<p> In order to clarify human mucosal and systemic immunity against influenza viral infection, the serum titers of anti-influenza virus-specific nasal secretory IgA and serum IgG and their changes after subcutaneous vaccination were measured in a Japanese healthy adult population in the present study. We recruited 155 healthy adults in 2006, with an average age of 24.1 years (range: 19-60 years). The male-female ratio was 1: 1. Nasopharyngeal lavage fluid and serum specimens were obtained prior to vaccination and a month after subcutaneous vaccination with a trivalent influenza ether split hemagglutinin vaccine of the A (H1N1), A (H3N2) subtypes and type B influenza viruses. Nasopharyngeal lavage fluid specimens were obtained by the nasal spray and aspiration method. The anti-influenza virus-specific IgA titers in the nasopharyngeal lavage fluid and IgG titers in the serum against the A (H1N1), A (H3N2) subtype and type B influenza viruses were measured by ELISA. The anti-influenza virus-specific nasal lavage fluid IgA titers were represented as a ratio to the total IgA titers. About 70% of the subjects had nasal anti-viral IgA against the A (H1N1), A (H3N2) subtype and type B influenza viruses, and almost all had serum anti-viral IgG against the A (H1N1), A (H3N2) subtype and type B viruses. The serum antiviral IgG titers, but not the nasal antiviral IgA titers, were significantly elevated at 1 month after the subcutaneous vaccination. Moreover, the serum antiviral IgG titers were significantly elevated after vaccination only in subjects with low pre-vaccination IgG titers, and not in those with high pre-vaccination IgG titers. The nasal antiviral IgA titers in the subjects of our present study were significantly higher than those in patients with influenza infection reported from our previous study. The presence of nasal anti-influenza virus-specific IgA in about 70% of Japanese adults is considered as being suggestive of a history of influenza infection. The presence of anti-influenza virus-specific IgG in the serum in almost all Japanese adults could suggest a history of influenza infection or influenza vaccination. Currently available subcutaneous influenza vaccines induce systemic immunity, with the appearance of anti-viral IgG in the serum, in adults. However, subcutaneous vaccination does not appear to be capable of inducing mucosal immunity with the induction of antiviral secretory IgA in the nasopharynx. The present findings suggest that subcutaneous influenza vaccination can suppress the progression of influenza infection by inducing the appearance of antiviral IgG in serum, but not by inducing the appearance of antiviral IgA in the nasopharynx. The findings also suggest that subjects with low antiviral secretory IgA titers in the nasopharynx are at a higher risk of influenza infection.</p>
Junji Chida, Hideyuki Hara, Keiji Uchiyama, Etsuhisa Takahashi, Hironori Miyata, Hidetaka Kosako, Yukiko Tomioka, Toshihiro Ito, Hiroyuki Horiuchi, Haruo Matsuda, Hiroshi Kido and Suehiro Sakaguchi : Prion protein signaling induces M2 macrophage polarization and protects from lethal influenza infection in mice., PLoS Pathogens, Vol.16, No.8, e1008823, 2020.
(要約)
The cellular prion protein, PrPC, is a glycosylphosphatidylinositol anchored-membrane glycoprotein expressed most abundantly in neuronal and to a lesser extent in non-neuronal cells. Its conformational conversion into the amyloidogenic isoform in neurons is a key pathogenic event in prion diseases, including Creutzfeldt-Jakob disease in humans and scrapie and bovine spongiform encephalopathy in animals. However, the normal functions of PrPC remain largely unknown, particularly in non-neuronal cells. Here we show that stimulation of PrPC with anti-PrP monoclonal antibodies (mAbs) protected mice from lethal infection with influenza A viruses (IAVs), with abundant accumulation of anti-inflammatory M2 macrophages with activated Src family kinases (SFKs) in infected lungs. A SFK inhibitor dasatinib inhibited M2 macrophage accumulation in IAV-infected lungs after treatment with anti-PrP mAbs and abolished the anti-PrP mAb-induced protective activity against lethal influenza infection in mice. We also show that stimulation of PrPC with anti-PrP mAbs induced M2 polarization in peritoneal macrophages through SFK activation in vitro and in vivo. These results indicate that PrPC could activate SFK in macrophages and induce macrophage polarization to an anti-inflammatory M2 phenotype after stimulation with anti-PrP mAbs, thereby eliciting protective activity against lethal infection with IAVs in mice after treatment with anti-PrP mAbs. These results also highlight PrPC as a novel therapeutic target for IAV infection.
Makoto Irahara, Wakako Shinahara, Mayumi Sugimoto, Yukiko Ogawa, Keiji Shitsukawa, Kenji Kubota, Limin Yang, Yukihiro Ohya, Hirohisa Saito, Shoji Kagami, Kokichi Arisawa and Hiroshi Kido : Trajectories of class-switching-related egg and cow's milk allergen-specific immunoglobulin isotype formation and its modification by eczema with low- and high-affinity immunoglobulin E during early infancy, Immunity, Inflammation and Disease, Vol.7, No.2, 74-85, 2019.
(要約)
Allergen-specific immunoglobulin isotype formation associated with immunoglobulin class-switching during the lactation period is the immunological background for food allergy in infants. We analyzed the serial changes in the production of feeding type-related egg- and milk-specific immunoglobulin isotypes from birth to 6 months of age with or without eczema in 84 infants. Allergen-specific immunoglobulin G1 (IgG1), IgG2, IgG3, IgG4, IgA, and IgE levels of hen's egg and bovine milk were measured in cord blood and blood samples from infants at 2, 4, and 6 months of age by the densely carboxylated protein microarray. Formula and mixed feeding were associated with a rapid increase in cow's milk allergen-specific immunoglobulins and feeding type-related significant differences in casein-specific immunoglobulin levels were detected. Breast and mixed feeding were associated with slow but significant increase in ovalbumin-specific IgG1 and IgE levels, but not other immunoglobulins. We found two different immunoglobulin isotype formation at 6 months of age with low- or high-affinity IgE against ovalbumin. One isotype formation pattern had relatively high ovalbumin-specific IgG1 levels, detectable IgG2, and low-affinity IgE, while the other had low ovalbumin-specific IgG1 levels, undetectable IgG2, and high levels of high-affinity IgE. The incidence of eczema was significantly higher in the latter pattern (84.6%), compared with the remaining infants (42.2%). Feeding practice-related allergen sensitization and immunoglobulin isotype formation were identified during the lactation period. The development of eczema during the lactation period could potentially modify the immunoglobulin isotype formation with high levels of high-affinity IgE.
Takashi Kimoto, Hyejin Kim, Satoko Sakai, Etsuhisa Takahashi and Hiroshi Kido : Oral vaccination with influenza hemagglutinin combined with human pulmonary surfactant-mimicking synthetic adjuvant SF-10 induces efficient local and systemic immunity compared with nasal and subcutaneous vaccination and provides protective immunity in mice., Vaccine, Vol.37, No.4, 612-622, 2019.
(要約)
We reported previously that a synthetic mucosal adjuvant SF-10, which mimics human pulmonary surfactant, delivers antigen to mucosal dendritic cells in the nasal cavity and promotes induction of humoral and cellular immunity. The aim of the present study was to determine the effects of oral administration of antigen combined with SF-10 (antigen-SF-10) on systemic and local immunity. Oral administration of ovalbumin, a model antigen, combined with SF-10 enhanced ovalbumin uptake into intestinal antigen presenting MHC IICD11c cells and their CD11bCD103 and CD11bCD103 subtype dendritic cells, which are the major antigen presenting subsets of the intestinal tract, more efficiently compared to without SF-10. Oral vaccination with influenza hemagglutinin vaccine (HAv)-SF-10 induced HAv-specific IgA and IgG in the serum, and HAv-specific secretory IgA and IgG in bronchoalveolar lavage fluid, nasal washes, gastric extracts and fecal material; their levels were significantly higher than those induced by subcutaneous HAv or intranasal HAv and HAv-SF-10 vaccinations. Enzyme-linked immunospot assay showed high numbers of HAv-specific IgA and IgG antibody secreting cells in the gastrointestinal and respiratory mucosal lymphoid tissues after oral vaccination with HAv-SF-10, but no or very low induction following oral vaccination with HAv alone. Oral vaccination with HAv-SF-10 provided protective immunity against severe influenza A virus infection, which was significantly higher than that induced by HAv combined with cholera toxin. Oral vaccination with HAv-SF-10 was associated with unique cytokine production patterns in the spleen after HAv stimulation; including marked induction of HAv-responsive Th17 cytokines (e.g., IL-17A and IL-22), high induction of Th1 cytokines (e.g., IL-2 and IFN-γ) and moderate induction of Th2 cytokines (e.g., IL-4 and IL-5). These results indicate that oral vaccination with HAv-SF-10 induces more efficient systemic and local immunity than nasal or subcutaneous vaccination with characteristically high levels of secretory HAv-specific IgA in various mucosal organs and protective immunity.
Kiwako Yamamoto-Hanada, Tohru Kobayashi, Hywel C. Williams, Masashi Mikami, Mayako Saito-Abe, Kumiko Morita, Osamu Natsume, Miori Sato, Motoko Iwama, Yumiko Miyaji, Makiko Miyata, Shinichiro Inagaki, Fukuie Tatsuki, Narita Masami, Shoji F. Nakayama, Hiroshi Kido, Hirohisa Saito and Yukihiro Ohya : Early aggressive intervention for infantile atopic dermatitis to prevent development of food allergy: a multicenter, investigator-blinded, randomized, parallel group controlled trial (PACI Study) protocol for a randomized controlled trial., Clinical and Translational Allergy, Vol.8, 47, 2018.
(要約)
Atopic dermatitis is the first clinical manifestation of the atopic march, with the highest incidence in the first year of life. Those affected often go on to develop other allergic diseases including food allergy, asthma, and allergic rhinitis. Recent evidence suggests that sensitization to foods may occur through a defective skin barrier which is common in atopic dermatitis in early life. We hypothesize that therapeutic aggressive intervention to treat new onset atopic dermatitis may prevent the development of later allergen sensitization, and associated food allergy, asthma, and allergic rhinitis. This study is a multi-center, pragmatic, two-parallel group, assessor-blind, superiority, individually randomized controlled trial. Atopic dermatitis infants (N = 650) 7-13 weeks old who develop an itchy rash within the previous 28 days are randomly assigned to the aggressive treatment or the conventional treatment in a 1:1 ratio. The primary outcome is oral food challenge-proven IgE-mediated hen's egg allergy at the age of 28 weeks. This is a novel pragmatic RCT study to examine the efficacy of early aggressive treatment for atopic dermatitis to prevent later food allergy. If our hypothesis is correct, we hope that such a strategy might impact on disease prevention in countries where food allergy is common, and that our results might reduce the frequency and associated costs of all food allergies as well as hens egg food allergy. Long-term follow and other similar studies will help to determine whether such a strategy will reduce the burden of other allergic diseases such as asthma and allergic rhinitis. UMIN-CTR: UMIN000028043.
Jun Oda, Yukioka Tetsu, Kazunari Azuma, Takao Arai, Junji Chida and Hiroshi Kido : Endogenous genetic risk factor for serious heatstroke: the thermolabile phenotype of carnitine palmitoyltransferase II variant., Acute Medicine & Surgery, Vol.6, No.1, 25-29, 2018.
(要約)
In serious heatstroke, elevated body temperature (>40°C) is considered the main cause of illness. Mitochondrial carnitine palmitoyltransferase II (CPT II) plays an important role in adenosine triphosphate (ATP) generation from long-chain fatty acids, and its thermolabile phenotype of polymorphisms leads to ATP production loss under high fever. Whether by heatstroke or influenza, high fever suppresses mitochondrial ATP production in patients with the thermolabile phenotype of polymorphisms. We investigated the relation between polymorphism and severity of heatstroke with a body temperature of over 40°C. We analyzed blood chemistry test results, Japanese Association for Acute Medicine Disseminated Intravascular Coagulation (JAAM DIC), Acute Physiologic and Chronic Health Evaluation II, and Sequential Organ Failure Assessment (SOFA) scores, and polymorphisms in 24 consecutive patients with severe heatstroke at two university hospitals. Eleven patients carried thermolabile CPT II variants (rs2229291; c.1055T G [p.Phe352Cys]) (F352C), and the genotype frequency was greater in heatstroke patients than in healthy volunteers. There was no significant difference in body temperature or blood chemistry data at emergency room arrival between patients with and without the CPT II variants. However, hospital days were longer and initial antithrombin activity was significantly lower in the variant group, suggesting a possible link with early phase vascular endothelial cell dysfunction. The JAAM DIC diagnostic criteria and SOFA scores were also higher in the group. There were no differences in the serum albumin, serum creatine kinase, and fibrin degradation product levels, and platelet counts. In addition to known risks (e.g., environmental temperature and old age), the CPT II polymorphism [F352C] can be a predisposing genetic risk factor for serious heatstroke with organ disfunction, and lower antithrombin activity.
Junji Chida, Hideyuki Hara, Masashi Yano, Keiji Uchiyama, Rani Nandita Das, Etsuhisa Takahashi, Hironori Miyata, Yukiko Tomioka, Toshihiro Ito, Hiroshi Kido and Suehiro Sakaguchi : Prion protein protects mice from lethal infection with influenza A viruses., PLoS Pathogens, Vol.14, No.5, e1007049, 2018.
(要約)
The cellular prion protein, designated PrPC, is a membrane glycoprotein expressed abundantly in brains and to a lesser extent in other tissues. Conformational conversion of PrPC into the amyloidogenic isoform is a key pathogenic event in prion diseases. However, the physiological functions of PrPC remain largely unknown, particularly in non-neuronal tissues. Here, we show that PrPC is expressed in lung epithelial cells, including alveolar type 1 and 2 cells and bronchiolar Clara cells. Compared with wild-type (WT) mice, PrPC-null mice (Prnp0/0) were highly susceptible to influenza A viruses (IAVs), with higher mortality. Infected Prnp0/0 lungs were severely injured, with higher inflammation and higher apoptosis of epithelial cells, and contained higher reactive oxygen species (ROS) than control WT lungs. Treatment with a ROS scavenger or an inhibitor of xanthine oxidase (XO), a major ROS-generating enzyme in IAV-infected lungs, rescued Prnp0/0 mice from the lethal infection with IAV. Moreover, Prnp0/0 mice transgenic for PrP with a deletion of the Cu-binding octapeptide repeat (OR) region, Tg(PrPOR)/Prnp0/0 mice, were also highly susceptible to IAV infection. These results indicate that PrPC has a protective role against lethal infection with IAVs through the Cu-binding OR region by reducing ROS in infected lungs. Cu content and the activity of anti-oxidant enzyme Cu/Zn-dependent superoxide dismutase, SOD1, were lower in Prnp0/0 and Tg(PrPOR)/Prnp0/0 lungs than in WT lungs. It is thus conceivable that PrPC functions to maintain Cu content and regulate SOD1 through the OR region in lungs, thereby reducing ROS in IAV-infected lungs and eventually protecting them from lethal infection with IAVs. Our current results highlight the role of PrPC in protection against IAV infection, and suggest that PrPC might be a novel target molecule for anti-influenza therapeutics.
Hideki Hayashi, Yoshinao Kubo, Mai Izumida, Etsuhisa Takahashi, Hiroshi Kido, Ko Sato, Mutsuo Yamaya, Hidekazu Nishimura, Kou Nakayama and Toshifumi Matsuyama : Enterokinase enhances influenza A virus infection by activating trypsinogen in human cell lines., Frontiers in Cellular and Infection Microbiology, Vol.8, 91, 2018.
(要約)
Cleavage and activation of hemagglutinin (HA) by trypsin-like proteases in influenza A virus (IAV) are essential prerequisites for its successful infection and spread. In host cells, some transmembrane serine proteases such as TMPRSS2, TMPRSS4 and HAT, along with plasmin in the bloodstream, have been reported to cleave the HA precursor (HA) molecule into its active forms, HA and HA. Some trypsinogens can also enhance IAV proliferation in some cell types (e.g., rat cardiomyoblasts). However, the precise activation mechanism for this process is unclear, because the expression level of the physiological activator of the trypsinogens, the TMPRSS15 enterokinase, is expected to be very low in such cells, with the exception of duodenal cells. Here, we show that at least two variant enterokinases are expressed in various human cell lines, including A549 lung-derived cells. The exogenous expression of these enterokinases was able to enhance the proliferation of IAV in 293T human kidney cells, but the proliferation was reduced by knocking down the endogenous enterokinase in A549 cells. The enterokinase was able to enhance HA processing in the cells, which activated trypsinogen and in the IAV-infected cells also. Therefore, we conclude that enterokinase plays a role in IAV infection and proliferation by activating trypsinogen to process viral HA in human cell lines.
(キーワード)
Cell Line / Enzyme Activation / Hemagglutinin Glycoproteins, Influenza Virus / Host-Pathogen Interactions / Humans / Influenza A Virus, H1N1 Subtype / Influenza, Human / Protein Processing, Post-Translational / Serine Endopeptidases / トリプシン (trypsin)
Etsuhisa Takahashi, Irene L. Indalao, Takako Sawabuchi, Keiko Mizuno, Satoko Sakai, Takashi Kimoto, Hyejin Kim and Hiroshi Kido : Clarithromycin suppresses induction of monocyte chemoattractant protein-1 and matrix metalloproteinase-9 and inproves pathological changes in the lungs and heart of mice infected with influenza A virus., Comparative Immunology, Microbiology and Infectious Diseases, Vol.56, 6-13, 2018.
(要約)
The influenza A virus (IAV)-cytokine-trypsin/matrix metalloproteinase-9 (MMP-9) cycle is one of the important mechanisms of multiple organ failure in severe influenza. Clarithromycin, a macrolide antibiotic, has immune modulatory and anti-inflammatory effects. We analyzed the effects of clarithromycin on the induction of chemokines, cytokines, MMP-9, trypsin, vascular hyper-permeability and inflammatory aggravation in mice with IAV infection. IAV/Puerto Rico/8/34(H1N1) infection increased the levels of monocyte chemoattractant protein-1 (MCP-1) and cytokines in serum, and MMP-9 and trypsin in serum and/or the lungs and heart. Clarithromycin significantly suppressed the induction of serum MCP-1 and MMP-9 and vascular hyperpermeability in these organs in the early phase of infection, but did not suppress the induction of trypsin, IL-6 or IFN-γ. Histopathological examination showed that clarithromycin tended to reduce inflammatory cell accumulation in the lungs and heart. These results suggest that clarithromycin suppresses infection-related inflammation and reduces vascular hyperpermeability by suppressing the induction of MCP-1 and MMP-9.
Hyejin Kim, Takashi Kimoto, Satoko Sakai, Etsuhisa Takahashi and Hiroshi Kido : Adjuvanting influenza hemagglutinin vaccine with a human pulmonary surfactant-mimicking synthetic compound SF-10 induces local and systemic cell-mediated immunity in mice., PLoS ONE, Vol.13, No.1, e0191133, 2018.
(要約)
We reported previously that intranasal instillation of a synthetic human pulmonary surfactant with a carboxy vinyl polymer as a viscosity improver, named SF-10, shows potent adjuvanticity for humoral immunity in mice and cynomolgus monkeys. SF-10 effectively induces influenza hemagglutinin vaccine (HAv)-specific IgA in nasal and lung washes and IgG in sera with their neutralizing activities. Since CD8+ T cell-mediated protection is an important requirement for adaptive immunity, we investigated in this study the effects of SF-10 with antigen on local and systemic cell-mediated immunity. Nasal instillation of ovalbumin, a model antigen, combined with SF-10 efficiently delivered antigen to mucosal dendritic and epithelial cells and promoted cross-presentation in antigen presenting cells, yielding a high percentage of ovalbumin-specific cytotoxic T lymphocytes in the nasal mucosa, compared with ovalbumin alone. Nasal immunization of HAv-SF-10 also induced HAv-specific cytotoxic T lymphocytes and upregulated granzyme B expression in splenic CD8+ T cells with their high cytotoxicity against target cells pulsed with HA peptide. Furthermore, nasal vaccination of HAv-SF-10 significantly induced higher cytotoxic T lymphocytes-mediated cytotoxicity in the lungs and cervical lymph nodes in the early phase of influenza virus infection compared with HAv alone. Protective immunity induced by HAv-SF-10 against lethal influenza virus infection was partially and predominantly suppressed after depletion of CD8+ and CD4+ T cells (induced by intraperitoneal injection of the corresponding antibodies), respectively, suggesting that CD4+ T cells predominantly and CD8+ T cells partially contribute to the protective immunity in the advanced stage of influenza virus infection. These results suggest that SF-10 promotes effective antigen delivery to antigen presenting cells, activates CD8+ T cells via cross-presentation, and induces cell-mediated immune responses against antigen.
Masaya Takahashi, Kazuhiko Soejima, Shoichiro Taniuchi, Yasuko Hatano, Sohsaku Yamanouchi, Hideki Ishikawa, Makoto Irahara, Youhei Sasaki, Hiroshi Kido and Kazunari Kaneko : Oral immunotherapy combined with Omalizumab treatment induces early desensitization to cows milk in children with high-risk cows milk allergy: a randomized controlled trial., Scientific Reports, Vol.7, No.1, 17453, 2017.
(要約)
We evaluated the efficacy and safety of oral immunotherapy (OIT) combined with 24 weeks of omalizumab (OMB) at inducing desensitization in children with cow's milk allergy (CM) compared with an untreated group. The present study was a prospective randomized controlled trial. Sixteen patients (age, 6-14 years) with high IgE levels to CM were enrolled in the present study. Patients were randomized 1:1 to receive OMB-OIT group or untreated group. The primary outcome was the induction of desensitization at 8 weeks after OMB was discontinued in OMB-OIT treated group and at 32 weeks after study entry. None of the 6 children in the untreated group developed desensitization to CM while all of the 10 children in the OIT-OMB treated group achieved desensitization (P < 0.001). A significantly decreased wheal diameter in response to a skin prick test using CM was found in the OMB-OIT treated group (P < 0.05). These data suggest that OIT combined with OMB using microwave heated CM may help to induce desensitization for children with high-risk CM allergy. This prospective randomized controlled trial was intended for 50 participants but was prematurely discontinued due to overwhelming superiority of OMB combined with microwave heated OIT over CM avoidance.
Norio Kawamoto, Norio Kamemura, Hiroshi Kido and Toshiyuki Fukao : Detection of ovomucoid-specific low-affinity IgE in infants and its relationship to eczema., Pediatric Allergy and Immunology, Vol.28, No.4, 355-361, 2017.
(要約)
Allergen-specific low-affinity IgE was previously detected in cord blood by a highly sensitive densely carboxylated protein (DCP) chip, but not by ImmunoCAP. Here, we investigated the presence of low-affinity IgE during the early life of infants and observed its relationship with eczema. We conducted a birth cohort study, collecting sera at birth and 6 (6M) and 14 months (14M) of age (n=110). We monitored the ovomucoid (OM)- and egg white (EW)-specific-IgE (sIgE) by ImmunoCAP or DCP-chip and analyzed the antigen-affinity of sIgE by binding inhibition assays in the presence or absence of a mild chaotropic agent, diethyl amine (DEA). The low- and high-affinity OM-sIgEs and sensitization risk factors were analyzed by a multivariate logistic analysis. The OM-sIgE measured by DCP-chip significantly correlated with that measured by ImmunoCAP, but some samples assessed as OM-sIgE-positive by DCP-chip were considered OM-sIgE-negative by ImmunoCAP. Binding inhibition analysis after DEA treatment was performed for participants judged as OM-sIgE-positive by DCP-chip at 14M. The group assessed as negative for OM- and EW-sIgE by ImmunoCAP at 6M and 14M showed a larger binding inhibition curve shift after DEA treatment than did the group assessed as positive at these times, indicating the presence of low-affinity sIgE antibodies at 14M. The logistic regression analysis found that persistent eczema from 6M to 14M is a significant risk factor for developing high-affinity, but not low-affinity, sIgE. Human infant peripheral blood contains allergen-specific low-affinity sIgE. Persistent eczema is related to the development of high-affinity, but not low-affinity, IgE. This article is protected by copyright. All rights reserved.
Osamu Natsume, Shigenori Kabashima, Junko Nakazato, Kiwako Yamamoto-Hanada, Masami Narita, Mai Kondo, Mayako Saito, Ai Kishino, Tetsuya Takimoto, Eisuke Inoue, Julian Tang, Hiroshi Kido, Gary W. K. Wong, Kenji Matsumoto, Hirohisa Saito and Yukihiro Ohya : Two-step egg introduction for prevention of egg allergy in high-risk infants with eczema (PETIT): a randomised, double-blind, placebo-controlled trial, The Lancet, Vol.389, No.10066, 276-286, 2017.
(要約)
Evidence is accumulating that early consumption is more beneficial than is delayed introduction as a strategy for primary prevention of food allergy. However, allergic reactions caused by early introduction of such solid foods have been a problematic issue. We investigated whether or not early stepwise introduction of eggs to infants with eczema combined with optimal eczema treatment would prevent egg allergy at 1 year of age. In this randomised, double-blind, placebo-controlled trial, we enrolled infants 4-5 months of age with eczema from two centres in Japan. Exclusion criteria were being born before 37 weeks of gestational age, experience of ingestion of hen's eggs or egg products, history of immediate allergic reaction to hen's eggs, history of non-immediate allergic reaction to a particular type of food, and complications of any severe disease. Infants were randomly assigned (block size of four; stratified by institution and sex) to early introduction of egg or placebo (1:1). Participants in the egg group consumed orally 50 mg of heated egg powder per day from 6 months to 9 months of age and 250 mg per day thereafter until 12 months of age. We aggressively treated participants' eczema at entry and maintained control without exacerbations throughout the intervention period. Participants and physicians were masked to assignment, and allocation was concealed. The primary outcome was the proportion of participants with hen's egg allergy confirmed by open oral food challenges at 12 months of age, assessed blindly by standardised methods, in all randomly allocated participants who received the intervention. This trial is registered with the University Hospital Medical Information Network Clinical Trials Registry, number UMIN000008673. Between Sept 18, 2012, and Feb 13, 2015, we randomly allocated 147 participants (73 [50%] to the egg group and 74 [50%] to the placebo group). This trial was terminated on the basis of the results of the scheduled interim analysis of 100 participants, which showed a significant difference between the two groups (four [9%] of 47 participants had an egg allergy in the egg group vs 18 [38%] of 47 in the placebo group; risk ratio 0·222 [95% CI 0·081-0·607]; p=0·0012). In the primary analysis population, five (8%) of 60 participants had an egg allergy in the egg group compared with 23 (38%) of 61 in the placebo group (risk ratio 0·221 [0·090-0·543]; p=0·0001). The only difference in adverse events between groups was admissions to hospital (six [10%] of 60 in the egg group vs none in the placebo group; p=0·022). 19 acute events occurred in nine (15%) participants in the egg group versus 14 events in 11 (18%) participants in the placebo group after intake of the trial powder. Introduction of heated egg in a stepwise manner along with aggressive eczema treatment is a safe and efficacious way to prevent hen's egg allergy in high-risk infants. In this study, we developed a practical approach to overcome the second wave of the allergic epidemic caused by food allergy. Ministry of Health, Labour and Welfare, and National Centre for Child Health and Development, Japan.
I L. Indalao, T Sawabuchi, E Takahashi and Hiroshi Kido : IL-1β is a key cytokine which induces trypsin upregulation in influenza virus-cutokine-trypsin cycle., Archives of Virology, Vol.162, No.1, 201-211, 2016.
(要約)
Severe influenza is characterized by a cytokine storm, and the influenza virus-cytokine-trypsin cycle is one of the important mechanisms of viral multiplication and multiple organ failure. The aim of this study was to define the key cytokine(s) responsible for trypsin upregulation. Mice were infected with influenza virus strain A/Puerto Rico/8/34 (H1N1) or treated individually or with a combination of interleukin-1β, interleukin-6, and tumor necrosis factor α. The levels of these cytokines and trypsin in the lungs were monitored. The neutralizing effects of anti-IL-1β antibodies on cytokine and trypsin expression in human A549 cells and lung inflammation in the infected mice were examined. Infection induced interleukin-1β, interleukin-6, tumor necrosis factor α, and ectopic trypsin in mouse lungs in a dose- and time-dependent manner. Intraperitoneal administration of interleukin-1β combined with other cytokines tended to upregulate trypsin and cytokine expression in the lungs, but the combination without interleukin-1β did not induce trypsin. In contrast, incubation of A549 cells with interleukin-1β alone induced both cytokines and trypsin, and anti-interleukin-1β antibody treatment abrogated these effects. Administration of the antibody in the infected mice reduced lung inflammation area. These findings suggest that IL-1β plays a key role in trypsin upregulation and has a pathological role in multiple organ failure.
Shima Atsushi, Yasuno Tetsuhiko, Kohno Ryuichi, Yamada Kenji, Yamaguchi Seiji, Yamaguchi Miyoko, Hiroshi Kido and Fukuda Hideyoshi : First Japanese Case of Carnitine Palmitoyltransferase II Deficiency with the Homozygous Point Mutation S113L, Internal Medicine, Vol.55, No.18, 2659-2661, 2016.
Hiroshi Kido, Indalao L. Irene, Kim Hyejin, Kimoto Takashi, Sakai Satoko and Etsuhisa Takahashi : Energy metabolic disorder is a major risk factor in severe influenza virus infection: Proposals for new therapeutic options based on animal model experiments, Respiratory Investigation, Vol.54, No.5, 312-319, 2016.
(要約)
Severe influenza is characterized by cytokine storm and multiorgan failure. Influenza patients with underlying diseases show a rapid progression in disease severity. The major mechanism that underlies multiorgan failure during the progressive stage of infection, particularly in patients with underlying risk factors, is mitochondrial energy crisis. The relationship between the factors that determine infection severity, such as influenza virus, cytokines, cellular trypsin as a hemagglutinin processing protease for viral multiplication, accumulation of metabolic intermediates and ATP crisis in mitochondria, is termed the "influenza virus-cytokine-trypsin" cycle. This occurs during the initial stages of infection, and is interconnected with the "metabolic disorders-cytokine" cycle in the middle to late phase of infection. Experiments using animal models have highlighted the complex relationship between these two cycles. New treatment options have been proposed that target the ATP crisis and multiorgan failure during the late phase of infection, rather than antiviral treatments with neuraminidase inhibitors that work during the initial phase. These options are (i) restoration of glucose oxidation in mitochondria by diisopropylamine dichloroacetate, which inhibits infection-induced pyruvate dehydrogenase kinase 4 activity, and (ii) restoration of long-chain fatty acid oxidation in mitochondria by l-carnitine and bezafibrate, an agonist of peroxisome proliferation-activated receptors-β/δ, which transcriptionally upregulates carnitine palmitoyltransferase II. The latter is particularly effective in patients with influenza-associated encephalopathy who have thermolabile and short half-life compound variants of carnitine palmitoyltransferase II.
(キーワード)
Animals / Energy Metabolism / Humans / Influenza A virus / Influenza, Human / Metabolic Diseases / Mice / Risk Factors
Dai Mizuno, Takashi Kimoto, Satoko Sakai, Etsuhisa Takahashi, Hyejin Kim and Hiroshi Kido : Induction of systemic and mucosal immunity and maintenance of its memory against influenza A virus by nasal vaccination using a new mucosal adjuvant SF-10 derived from pulmonary surfactant in young cynomolgus monkeys, Vaccine, Vol.34, No.16, 1881-1888, 2016.
(要約)
Induction of systemic and mucosal immunity and maintenance of its memory was investigated in 12 young male cynomolgus monkeys after intranasal instillation of flu vaccine using a new mucosal adjuvant SF-10 derived from pulmonary surfactant constituents. Split-product of influenza virus A/California/7/2009(H1N1)pdm hemagglutinin vaccine (HAv) at 15μg with or without SF-10 and the adjuvant alone were instilled intranasally three times every 2 weeks. SF-10-adjuvanted HAv (SF-10-HAv) elicited significantly higher HAv-specific IgG and hemagglutinin inhibition (HI) titers in serum and HAv-specific secretory IgA and its neutralizing activities in nasal washes compared with HAv antigen and SF-10 alone. Significant cross-neutralizing activities of nasal washes after the third vaccination to several other H1N1 and H3N2 strains were observed. HI titers in serum and neutralizing activities in nasal washes reached peak levels at 6 weeks after initial vaccination, then gradually decreased after 10 weeks and returned to the baseline levels at 36 weeks. A single intranasal revaccination of SF-10-HAv at 36 weeks rapidly and significantly increased both immunity in serum and nasal washes compared with naïve monkeys. Revaccination by one or two doses achieved almost maximal immunity at 2 or 4 weeks after instillation. Statistically significant adverse effects (e.g., body weight loss, elevated body temperature, nasal discharge, change in peripheral blood leukocyte and platelet counts) were not observed for 2 weeks after vaccination of SF-10-HAv, HAv or SF-10 and also during the experimental period. These results in young monkey model suggest the potential of clinical use SF-10 for intranasal flu vaccine.
Mayumi Sugimoto, Norio Kamemura, Mizuho Nagao, Makoto Irahara, Shoji Kagami, Takao Fujisawa and Hiroshi Kido : Differential response in allergen-specific IgE, IgGs and IgA levels for predicting outcome of oral immunotherapy., Pediatric Allergy and Immunology, Vol.27, No.3, 276-282, 2016.
(要約)
The response to OIT was associated with significant increases in serum allergen-specific IgG1 levels after rush phase and high baseline IgA levels, compared with small changes in immunoglobulin response in low-responders. The characteristic IgG1 changes and IgA levels in the responders could be potentially useful biomarkers for the prediction of positive clinical response to OIT. This article is protected by copyright. All rights reserved.
Shoji Masaki, Arakai Yumie, Esumi Tomoyuki, Konami Shuntaro, Yamamoto Chihiro, Suzaki Yutaka, Etsuhisa Takahashi, Konishi Shiro, Hiroshi Kido and Kuzuhara Takashi : Bakuchiol is a phenolic isoprenoid with novel enantiomer-selective anti-influenza A virus activity involving Nrf2 activation, The Journal of Biological Chemistry, Vol.290, No.46, 28001-28017, 2015.
(要約)
Influenza represents a substantial threat to human health and requires novel therapeutic approaches. Bakuchiol is a phenolic isoprenoid compound present in Babchi (Psoralea corylifolia L.) seeds. We examined the anti-influenza viral activity of synthetic bakuchiol using Madin-Darby canine kidney cells. We found that the naturally occurring form, (+)-(S)-bakuchiol, and its enantiomer, (-)-(R)-bakuchiol, inhibited influenza A viral infection and growth and reduced the expression of viral mRNAs and proteins in these cells. Furthermore, these compounds markedly reduced the mRNA expression of the host cell influenza A virus-induced immune response genes, interferon-β and myxovirus-resistant protein 1. Interestingly, (+)-(S)-bakuchiol had greater efficacy than (-)-(R)-bakuchiol, indicating that chirality influenced anti-influenza virus activity. In vitro studies indicated that bakuchiol did not strongly inhibit the activities of influenza surface proteins or the M2 ion channel, expressed in Chinese hamster ovary cells. Analysis of luciferase reporter assay data unexpectedly indicated that bakuchiol may induce some host cell factor(s) that inhibited firefly and Renilla luciferases. Next generation sequencing and KeyMolnet analysis of influenza A virus-infected and non-infected cells exposed to bakuchiol revealed activation of transcriptional regulation by nuclear factor erythroid 2-related factor (Nrf), and an Nrf2 reporter assay showed that (+)-(S)-bakuchiol activated Nrf2. Additionally, (+)-(S)-bakuchiol up-regulated the mRNA levels of two Nrf2-induced genes, NAD(P)H quinone oxidoreductase 1 and glutathione S-transferase A3. These findings demonstrated that bakuchiol had enantiomer-selective anti-influenza viral activity involving a novel effect on the host cell oxidative stress response.
Hiroshi Kido : Influenza virus pathogenicity regulated by host cellular proteases, cytokines and metabolites, and its therapeutic options, Proceedings of the Japan Academy. Series B, Physical and biological sciences, Vol.91, No.8, 351-368, 2015.
(要約)
Influenza A virus (IAV) causes significant morbidity and mortality. The knowledge gained within the last decade on the pandemic IAV(H1N1)2009 improved our understanding not only of the viral pathogenicity but also the host cellular factors involved in the pathogenicity of multiorgan failure (MOF), such as cellular trypsin-type hemagglutinin (HA0) processing proteases for viral multiplication, cytokine storm, metabolic disorders and energy crisis. The HA processing proteases in the airway and organs for all IAV known to date have been identified. Recently, a new concept on the pathogenicity of MOF, the "influenza virus-cytokine-trypsin" cycle, has been proposed involving up-regulation of trypsin through pro-inflammatory cytokines, and potentiation of viral multiplication in various organs. Furthermore, the relationship between causative factors has been summarized as the "influenza virus-cytokine-trypsin" cycle interconnected with the "metabolic disorders-cytokine" cycle. These cycles provide new treatment concepts for ATP crisis and MOF. This review discusses IAV pathogenicity on cellular proteases, cytokines, metabolites and therapeutic options.
Yao Min, Cai Min, Yao Dengfu, Xu Xi, Yang Rongrong, Li Yuting, Zhang Yuanyuan, Hiroshi Kido and Yao Dengbing : Abbreviated half-lives and impaired fuel utilization in carnitine palmitoyltransferase II variant fibrobrasts, PLoS ONE, Vol.10, No.3, e0119936, 2015.
(要約)
Carnitine palmitoyltransferase II (CPT II) deficiency is one of the most common causes of fatty acid oxidation metabolism disorders. However, the molecular mechanism between CPT2 gene polymorphisms and metabolic stress has not been fully clarified. We previously reported that a number of patients show a thermal instable phenotype of compound hetero/homozygous variants of CPT II. To understand the mechanism of the metabolic disorder resulting from CPT II deficiency, the present study investigated CPT II variants in patient fibroblasts, [c.1102 G>A (p.V368I)] (heterozygous), [c.1102 G>A (p.V368I)] (homozygous), and [c.1055 T>G (p.F352C)] (heterozygous) + [c.1102 G>A (p.V368I)] (homozygous) compared with fibroblasts from healthy controls. CPT II variants exerted an effect of dominant negative on the homotetrameric proteins that showed thermal instability, reduced residual enzyme activities and a short half-life. Moreover, CPT II variant fibroblasts showed a significant decrease in fatty acid β-oxidation and adenosine triphosphate generation, combined with a reduced mitochondrial membrane potential, resulting in cellular apoptosis. Collectively, our data indicate that the CPT II deficiency induces an energy crisis of the fatty acid metabolic pathway. These findings may contribute to the elucidation of the genetic factors involved in metabolic disorder encephalopathy caused by the CPT II deficiency.
Maekawa Toshihiro, Kimoto Takashi, Dai Mizuno, Furukawa Yuichi, Ida Masayuki, Etsuhisa Takahashi, Izumo Takayuki, Ono Yoshiko, Shibata Hiroshi and Hiroshi Kido : Oral Administration of Lactobacillus pentosus Strain S-PT84 Enhances Anti-Influenza Virus-Specific IgG Production in Plasma after Limited Doses of Influenza Virus Vaccination in Mice, Journal of Vaccine & Immunotechnology, Vol.2, No.1, 2015.
25.
Norio Kamemura, Miwa Takashima, Hideaki Morita, Kenji Matsumoto, Hirohisa Saito and Hiroshi Kido : Measurement of allergen-specific secretory IgA in stool of neonates, infants and toddlers by protection against degradation of immunoglobulins and allergens., The Journal of Medical Investigation : JMI, Vol.62, No.3-4, 137-144, 2015.
(要約)
Allergen-specific SIgA levels in stool of neonates, infants and toddlers under 36 months of age could be analyzed using protease inhibitors, including PMSF and leupeptin.
Mineyoshi Hiyoshi, I.L Indalao, Mihiro Yano, Kazuhiko Yamane, Etsuhisa Takahashi and Hiroshi Kido : Influenza A virus infection of vascular endothelial cells induces GSK-3β-mediated β-catenin degradation in adherens junctions, with a resultant increase in membrane permeability., Archives of Virology, Vol.160, No.1, 225-234, 2014.
(要約)
Multiorgan failure with vascular hyperpermeability is the final outcome in the progression of seasonal influenza virus pneumonia and influenza-associated encephalopathy, and it is also common in infection with highly pathogenic avian influenza virus. However, the precise molecular mechanism by which influenza virus infection causes vascular endothelial cell hyperpermeability remains poorly defined. We investigated the mechanisms of hyperpermeability of human umbilical vein endothelial cells infected with influenza A virus (IAV)/Puerto Rico/8/34 (PR8) (H1N1). The levels of β-catenin, a key regulatory component of the vascular endothelial-cadherin cell adhesion complex, were markedly decreased during infection for 28 h, with increments of vascular hyperpermeability measured by transendothelial electrical resistance. Lactacystin (at 2 μM), a proteasome inhibitor, inhibited the decrease in β-catenin levels. Since the N-terminal phosphorylation of β-catenin by glycogen synthase kinase (GSK)-3β is the initiation step of proteasome-dependent degradation, we examined the effects of GSK-3β suppression by RNA interference in endothelial cells. IAV-infection-induced β-catenin degradation was significantly inhibited in GSK-3β-knockdown cells, and transfection of cells with recombinant β-catenin significantly suppressed IAV-induced hyperpermeability. These findings suggest that IAV infection induces GSK-3β-mediated β-catenin degradation in the adherens junctional complexes and induces vascular hyperpermeability. The in vitro findings of β-catenin degradation and activation of GSK-3β after IAV infection were confirmed in lungs of mice infected with IAV PR8 during the course of infection from day 0 to day 6. These results suggest that GSK-3β-mediated β-catenin degradation in adherens junctions is one of the key mechanisms of vascular hyperpermeability in severe influenza.
Pan Hai-Yan, Sun Hua-Mei, Xue Lu-jing, Pan Min, Wang Yi-Ping, Hiroshi Kido and Zhu Jian-Hua : Ectopic trypsin in the myocardium promotes dilated cardiomyopathy after influenza A virus infection, American Journal of Physiology, Heart and Circulatory Physiology, Vol.307, No.6, H922-H932, 2014.
(要約)
We have previously reported that ectopic trypsin in the myocardium triggers acute myocarditis after influenza A virus (IAV) infection. As myocarditis is a common precursor to dilated cardiomyopathy (DCM), the aim of the present study was to investigate the influence of trypsin on the progression of DCM after IAV infection. IAV-infected mice treated with saline or trypsin inhibitor were euthanized on days 0, 9, 20, 40 and 60 postinfection. Trypsin expression colocalized with myocardial inflammatory loci and IAV-induced myocarditis peaked on day 9 postinfection and alleviated by day 20 but persisted until day 60 postinfection, even though replication of IAV was not detected from day 20 postinfection. Similar time courses were observed for the activation of pro-matrix metalloproteinase (pro-MMP)-9 and expression of the proinflammatory cytokines IL-6, IL-1β, and TNF-α. Degradation of collagen type I, proliferation of ventricular interstitial collagen, and expression of collagen type I and III mRNA increased significantly during acute and chronic phases; collagen type III mRNA increased more significantly than collagen type I mRNA. Cardiac function progressively deteriorated with progressive left ventricular dilation. The trypsin inhibitor aprotinin suppressed pro-MMP-9 activation and cytokine release, alleviated myocardial inflammation, and restored collagen metabolism during acute and chronic phases of myocarditis. This effectively prevented ventricular dilation and improved cardiac function. These results suggest that ectopic trypsin in the myocardium promoted DCM through chronic activation of pro-MMP-9, persistent induction of cytokines, and mediation of collagen remodeling. Pharmacological inhibition of trypsin activity might be a promising approach for the prevention of viral cardiomyopathy.
(キーワード)
Animals / Cardiomyopathy, Dilated / Collagen Type I / Collagen Type III / Disease Models, Animal / Disease Progression / Enzyme Precursors / Hypertrophy, Left Ventricular / Inflammation Mediators / Influenza A Virus, H1N1 Subtype / Interleukin-1beta / Interleukin-6 / Male / Matrix Metalloproteinase 9 / Mice / Mice, Inbred BALB C / Myocarditis / Myocardium / Orthomyxoviridae Infections / RNA, Messenger / Time Factors / Trypsin / Trypsin Inhibitors / Tumor Necrosis Factor-alpha / Ventricular Dysfunction, Left / Ventricular Function, Left / Ventricular Remodeling / Virus Replication
Yamane Kazuhiro, Indalao L. Irene, Junji Chida, Yamamoto Yoshikazu, Hanawa Masaaki and Hiroshi Kido : Diisopropylamine dichloroacetate, a novel pyruvate dehydrogenase kinase 4 inhibitor, as a potential therapeutic agent for metabolic disorders and multiorgan failure in severe influenza, PLoS ONE, Vol.9, No.5, e98032, 2014.
(要約)
Severe influenza is characterized by cytokine storm and multiorgan failure with metabolic energy disorders and vascular hyperpermeability. In the regulation of energy homeostasis, the pyruvate dehydrogenase (PDH) complex plays an important role by catalyzing oxidative decarboxylation of pyruvate, linking glycolysis to the tricarboxylic acid cycle and fatty acid synthesis, and thus its activity is linked to energy homeostasis. The present study tested the effects of diisopropylamine dichloroacetate (DADA), a new PDH kinase 4 (PDK4) inhibitor, in mice with severe influenza. Infection of mice with influenza A PR/8/34(H1N1) virus resulted in marked down-regulation of PDH activity and ATP level, with selective up-regulation of PDK4 in the skeletal muscles, heart, liver and lungs. Oral administration of DADA at 12-h intervals for 14 days starting immediately after infection significantly restored PDH activity and ATP level in various organs, and ameliorated disorders of glucose and lipid metabolism in the blood, together with marked improvement of survival and suppression of cytokine storm, trypsin up-regulation and viral replication. These results indicate that through PDK4 inhibition, DADA effectively suppresses the host metabolic disorder-cytokine cycle, which is closely linked to the influenza virus-cytokine-trypsin cycle, resulting in prevention of multiorgan failure in severe influenza.
Endo Hiroshi, Mihiro Yano, Yuushi Okumura and Hiroshi Kido : Ibuprofen enhances the anticancer activity of cisplatin in lung cancer cells by inhibiting the heat shock protein 70, Cell Death & Disease, Vol.5, e1027, 2014.
(要約)
Hsp70 is often overexpressed in cancer cells, and the selective cellular survival advantage that it confers may contribute to the process of tumour formation. Thus, the pharmacological manipulation of Hsp70 levels in cancer cells may be an effective means of preventing the progression of tumours. We found that the downregulation of Hsp70 by ibuprofen in vitro enhances the antitumoural activity of cisplatin in lung cancer. Ibuprofen prominently suppressed the expression of Hsp70 in A549 cells derived from lung adenocarcinoma and sensitized them to cisplatin in association with an increase in the mitochondrial apoptotic cascade, whereas ibuprofen alone did not induce cell death. The cisplatin-dependent events occurring up- and downstream of mitochondrial disruption were accelerated by treatment with ibuprofen. The increase in cisplatin-induced apoptosis caused by the depletion of Hsp70 by RNA interference is evidence that the increased apoptosis by ibuprofen is mediated by its effect on Hsp70. Our observations indicate that the suppression of Hsp70 by ibuprofen mediates the sensitivity to cisplatin by enhancing apoptosis at several stages of the mitochondrial cascade. Ibuprofen, therefore, is a potential therapeutic agent that might allow lowering the doses of cisplatin and limiting the many challenge associated with its toxicity and development of drug resistance.
Kimoto Takashi, Dai Mizuno, Takei Tsunetomo, Kunimi Takuya, Ono Shinji, Sakai Satoko and Hiroshi Kido : Intranasal influenza vaccination using a new synthetic mucosal adjuvant SF-10: induction of potent local and systemic immunity with balanced Th1 and Th2 responses, Influenza and Other Respiratory Viruses, Vol.7, No.6, 1218-1226, 2013.
(要約)
We found previously that bovine pulmonary Surfacten® used in newborns with acute respiratory distress syndrome is a safe and efficacious antigen vehicle for intranasal vaccination. The objective of this study was to industrially produce a synthetic adjuvant mimicking Surfacten® for clinical use without risk of bovine spongiform encephalopathy. We selected three Surfacten lipids and surfactant protein (SP)-C as essential constituents for adjuvanticity. For replacement of the hydrophobic SP-C, we synthesized SP-related peptides and analyzed their adjuvanticity. We evaluated lyophilization to replace sonication for the binding of influenza virus hemagglutinin (HA) to the synthetic adjuvant. We also added a carboxy vinyl polymer (CVP) to the synthetic adjuvant and named the mixture as SF-10 adjuvant. HA combined with SF-10 was administered intranasally to mice, and induction of nasal-wash HA-specific secretory IgA (s-IgA) and serum IgG with Th1-/Th2-type cytokine responses in nasal cavity and virus challenge test were assessed. Intranasal immunization with HA-SF-10 induced significantly higher levels of anti-HA-specific nasal-wash s-IgA and serum IgG than those induced by HA-poly(I:C), a reported potent mucosal vaccine, and provided highly efficient protection against lethal doses of virus challenge in mice. Anti-HA-specific serum IgG levels induced by HA-SF-10 were almost equivalent to those induced by subcutaneous immunization of HA twice. Intranasal administration of HA-SF-10 induced balanced anti-HA-specific IgG1 and IgG2a in sera and IFN-γ- and IL-4-producing lymphocytes in nasal cavity without any induction of anti-HA IgE. The results suggest that HA-SF-10 is a promising nasal influenza vaccine and that SF-10 can be supplied in large quantities commercially.
W Shinahara, Etsuhisa Takahashi, T Sawabuchi, M Arai, N Hirotsu, Y Takasaki, S Shindo, K Shibao, T Yokoyama, K Nishikawa, M Mino, M Iwaya, Y Yamashita, S Suzuki, Dai Mizuno and Hiroshi Kido : Immunomodulator clarithromycin enhances mucosal and systemic immune responses and reduces re-infection rate in pediatric patients with influenza treated with antiviral neuraminidase inhibitors: a retrospective analysis., PLoS ONE, Vol.8, No.7, e70060, 2013.
(要約)
Treatment with antiviral neuraminidase inhibitors suppresses influenza viral replication and antigen production, resulting in marked attenuation of mucosal immunity and mild suppression of systemic immunity in mice. This study investigated the effects of immunomodulator clarithromycin (CAM) supplementation on mucosal and systemic immunity in pediatric patients with influenza treated with neuraminidase inhibitors. A retrospective, non-randomized case series study was conducted among five treatment groups of 195 children aged 5.9±3.3 years infected with influenza A in 2008/2009 season. The five treatment groups were oseltamivir (OSV), zanamivir (ZNV), OSV+CAM, ZNV+CAM and untreated groups. Anti-viral secretory IgA (S-IgA) levels in nasal washes and IgG levels in sera were measured. The re-infection rate was analyzed among the same five treatment groups in the 2009/2010 season. Treatment of influenza with OSV and ZNV for 5 days attenuated the induction of anti-viral S-IgA in nasal washes and anti-viral IgG in serum, compared with the untreated group. The combination of CAM plus OSV or ZNV boosted and restored the production of mucosal S-IgA and systemic IgG. The re-infection rates in the subsequent season were significantly higher in the OSV and ZNV groups than the untreated, while CAM+OSV and CAM+ZNV tended to reduce such rate. CAM restored the attenuated anti-viral mucosal and systemic immunity and reduced the re-infection rate in the subsequent year in pediatric patients with influenza treated with OSV and ZNV.
M Shoji, Etsuhisa Takahashi, D Hatakeyama, Y Iwai, Y Morita, R Shirayama, N Echigo, Hiroshi Kido, S Nakamura, T Mashino, T Okutani and T Kuzuhara : Anti-Influenza Activity of C60 Fullerene Derivatives., PLoS ONE, Vol.8, No.6, e66337, 2013.
(要約)
The H1N1 influenza A virus, which originated in swine, caused a global pandemic in 2009, and the highly pathogenic H5N1 avian influenza virus has also caused epidemics in Southeast Asia in recent years. Thus, the threat from influenza A remains a serious global health issue, and novel drugs that target these viruses are highly desirable. Influenza A RNA polymerase consists of the PA, PB1, and PB2 subunits, and the N-terminal domain of the PA subunit demonstrates endonuclease activity. Fullerene (C60) is a unique carbon molecule that forms a sphere. To identify potential new anti-influenza compounds, we screened 12 fullerene derivatives using an in vitro PA endonuclease inhibition assay. We identified 8 fullerene derivatives that inhibited the endonuclease activity of the PA N-terminal domain or full-length PA protein in vitro. We also performed in silico docking simulation analysis of the C60 fullerene and PA endonuclease, which suggested that fullerenes can bind to the active pocket of PA endonuclease. In a cell culture system, we found that several fullerene derivatives inhibit influenza A viral infection and the expression of influenza A nucleoprotein and nonstructural protein 1. These results indicate that fullerene derivatives are possible candidates for the development of novel anti-influenza drugs.
Masahiro Inoue, Kouichi Yasuda, Haruki Uemura, Natsumi Yasaka, Achim Schnaufer, Mihiro Yano, Hiroshi Kido, Daisuke Kohda, Hirofumi Doi, Toshihide Fukuma, Akihiko Tsuji and Nobuo Horikoshi : Trypanosoma brucei 14-3-3I and II proteins predominantly form a heterodimer structure that acts as a potent cell cycle regulator in vivo., The Journal of Biochemistry, Vol.153, No.5, 431-439, 2013.
(要約)
Hetero- and homodimerization of 14-3-3 proteins demonstrate distinctive functions in mammals and plants. Trypanosoma brucei 14-3-3I and II (Tb14-3-3I and II) play pivotal roles in motility, cytokinesis and the cell cycle; however, the significance and the mechanism of Tb14-3-3 dimerization are remained to be elucidated. We found that ectopically expressed epitope-tagged Tb14-3-3I and II proteins formed hetero- and homodimers with endogenous Tb14-3-3I and II proteins. However, we also found the ability to form hetero- or homodimers between Tb14-3-3I and II proteins was clearly affected by the sequence and location of the epitope tag used. We found a blue native polyacrylamide gel electrophoresis system followed by western blotting may distinguish monomer from dimer structure, and stable from unstable conformation of Tb14-3-3. Combined with co-immunoprecipitation results, we revealed that Tb14-3-3 proteins mainly existed as heterodimeric form. Furthermore, co-overexpression of Tb14-3-3I and II proteins in T. brucei induced aberrant numbers of organelles in cells, but overexpression of either isoform alone rarely produced such morphology. These results suggest that heterodimers play more significant roles than homodimers not only in the maintenance of steady-state levels of the 14-3-3 proteins but also in the regulation of cytokinesis.
Junji Chida, Rie Ono, Kazuhiko Yamane, Mineyoshi Hiyoshi, Masaji Nishimura, Mutsuo Onodera, Emiko Nakataki, Shichijo Koichi, Matsushita Masatomo and Hiroshi Kido : Blood lactate/ATP ratio, as an alarm index and real-time biomarker in critical illness, PLoS ONE, Vol.8, No.4, e60561, 2013.
(要約)
The acute physiology, age and chronic health evaluation (APACHE) II score and other related scores have been used for evaluation of illness severity in the intensive care unit (ICU), but there is still a need for real-time and sensitive prognostic biomarkers. Recently, alarmins from damaged tissues have been reported as alarm-signaling molecules. Although ATP is a member of the alarmins and its depletion in tissues closely correlates with multiple-organ failure, blood ATP level has not been evaluated in critical illness. To identify real-time prognostic biomarker of critical illness, we measured blood ATP levels and the lactate/ATP ratio (ATP-lactate energy risk score, A-LES) in critically ill patients. Blood samples were collected from 42 consecutive critically ill ICU patients and 155 healthy subjects. The prognostic values of blood ATP levels and A-LES were compared with APACHE II score. The mean ATP level (SD) in healthy subjects was 0.62 (0.19) mM with no significant age or gender differences. The median ATP level in severely ill patients at ICU admission was significantly low at 0.31 mM (interquartile range 0.25 to 0.44) than the level in moderately ill patient at 0.56 mM (0.38 to 0.70) (P<0.01). Assessment with ATP was further corrected by lactate and expressed as A-LES. The median A-LES was 2.7 (2.1 to 3.3) in patients with satisfactory outcome at discharge but was significantly higher in non-survivors at 38.9 (21.0 to 67.9) (P<0.01). Receiver operating characteristic analysis indicated that measurement of blood ATP and A-LES at ICU admission are as useful as APACHE II score for prediction of mortality. Blood ATP levels and A-LES are sensitive prognostic biomarkers of mortality at ICU admission. In addition, A-LES provided further real-time evaluation score of illness severity during ICU stay particularly for critically ill patients with APACHE II scores of ≥20.0.
(キーワード)
アデノシン三リン酸 (adenosine triphosphate) / Adolescent / Adult / Age Factors / Aged / Aged, 80 and over / Arteries / Biomarkers / Blood Chemical Analysis / 子ども (children) / Child, Preschool / Critical Illness / エネルギー代謝 (energy metabolism) / Female / Hemoglobins / Humans / 小児 (infant) / Infant, Newborn / Intensive Care Units / Lactic Acid / Male / Middle Aged / Prognosis / Reference Values / リスク (risk) / Time Factors / Veins / Young Adult
Etsuhisa Takahashi, Kosuke Kataoka, Irene L. Indalao, Keiko Konoha, Kazuyuki Fujii, Junji Chida, Dai Mizuno, Kohtaro Fujihashi and Hiroshi Kido : Oral clarithromycin enhances airway IgA immunity through induction of IgA class switching recombination and B-cell activating factor of the tumor necrosis factor family molecule on mucosal dendritic cells in mice infected with influenza A virus, Journal of Virology, Vol.86, No.20, 10924-1093, 2012.
(要約)
We previously reported that the macrolide antibiotic clarithromycin (CAM) enhanced the mucosal immune response in pediatric influenza, particularly in children treated with the antiviral neuraminidase inhibitor oseltamivir (OSV) with low production of mucosal antiviral secretory IgA (S-IgA). The aims of the present study were to confirm the effects of CAM on S-IgA immune responses, by using influenza A virus (IAV) H1N1-infected mice treated with or without OSV, and to determine the molecular mechanisms responsible for the induction of mucosal IgA class switching recombination in IAV-infected CAM-treated mice. The anti-IAV S-IgA responses and expression levels of IgA class switching recombination-associated molecules were examined in bronchus-lymphoid tissues and spleens of infected mice. We also assessed neutralization activities of S-IgA against IAV. Data show that CAM enhanced anti-IAV S-IgA induction in the airway of infected mice and restored the attenuated antiviral S-IgA levels in OSV-treated mice to the levels in the vehicle-treated mice. The expression levels of B-cell-activating factor of the tumor necrosis factor family (BAFF) molecule on mucosal dendritic cells as well as those of activation-induced cytidine deaminase and Iμ-Cα transcripts on B cells were enhanced by CAM, compared with the levels without CAM treatment, but CAM had no effect on the expression of the BAFF receptor on B cells. Enhancement by CAM of neutralization activities of airway S-IgA against IAV in vitro and reinfected mice was observed. This study identifies that CAM enhances S-IgA production and neutralizing activities through the induction of IgA class switching recombination and upregulation of BAFF molecules in mucosal dendritic cells in IAV-infected mice.
Ishii Ayumi, Kaminori Kanae, Mineyoshi Hiyoshi, Hiroshi Kido, Ohta Takeshi and Hiroaki Konishi : Inhibitory effect of SPE-39 due to tyrosine phosphorylation and ubiquitination on the function of Vps33B in the EGF-stimulated cells, FEBS Letters, Vol.586, No.16, 2245-2250, 2012.
(要約)
Although SPE-39 is a binding protein to Vps33B that is one of the subunit in the mammalian HOPS complex, the elements of SPE-39 function remain unknown. Here, we show that tyrosine phosphorylation of SPE-39 following EGF stimulation plays a role in the stability of SPE-39 itself. Ubiquitination of the C-terminal region of SPE-39 was also elevated in response to EGF stimulation, and this process was regulated by the phosphorylation of Tyr-11 in SPE-39. However, association of Vps33B with SPE-39 inhibited the elevation of ubiquitination of SPE-39 following EGF stimulation, which might be responsible for the stabilization of SPE-39. Furthermore, an opposing functional relationship between SPE-39 and Vps33B on the downregulation of the EGF receptor was observed in EGF-stimulated COS-7 cells.
(キーワード)
Animals / COS Cells / Carrier Proteins / Cercopithecus aethiops / DNA, Complementary / Epidermal Growth Factor / Gene Expression Regulation / Humans / Microscopy, Fluorescence / Phosphorylation / Protein Binding / Protein Interaction Mapping / Protein Structure, Tertiary / Time Factors / Tyrosine / Ubiquitin / Vesicular Transport Proteins
Cisa Fujimoto, Noriaki Takeda, A Matsunaga, A Sawada, T Tanaka, Takashi Kimoto, Wakako Shinahara, Takako Sawabuchi, Miyoko Yamaguchi, M Hayama, Hiroaki Yanagawa, Mihiro Yano and Hiroshi Kido : Induction and maintenance of anti-influenza antigen-specific nasal secretory IgA levels and serum IgG levels after influenza infection in adults., Influenza and Other Respiratory Viruses, Vol.6, No.6, 396-403, 2012.
(要約)
To determine the induction and changes in anti-influenza virus secretory IgA (s-IgA) levels in nasal washes and serum IgG levels in patients with influenza. The study recruited 16 patients with influenza aged 35.6 ± 9.6 years in 2007/2008 and 2008/2009 seasons. Nasal washes and serum were obtained throughout the first year. Anti-viral s-IgA levels and neutralization activities in nasal washes, and serum anti-viral IgG levels and hemagglutination inhibition (HI) titers were measured. Anti-viral(H1N1) s-IgA to total IgA ratio and neutralizing antibody titer were low in nasal washes of all patients, whereas serum levels of anti-viral IgG and HI titers varied widely at day 1.4 ± 1.0 postinfection. Both nasal s-IgA and serum IgG levels later increased significantly, reaching peak levels at day 9.6 ± 3.3 postinfection. The induced nasal s-IgA then returned toward the initial levels within 300 days, although the levels at day 143 ± 70 were 3.03-fold of the initial. Individual serum IgG levels also returned toward the initial levels within 300 days, although the mean levels remained high probably because of re-infection in a subgroup of patients. Although influenza A (H3N2) was a minor epidemic subtype in both flu seasons, a significant rise in nasal anti-viral (H3N2) s-IgA levels and a slightly increase in serum IgG levels were noted. Low levels of nasal anti-viral s-IgA and neutralizing antibody were noted compared with a wide range of serum anti-viral IgG and HI titers at the onset of infection. Elevated s-IgA and IgG returned toward the initial levels within 300 days of infection with minor exceptions.
(キーワード)
Adult / Antibodies, Viral / Female / Hemagglutination Inhibition Tests / Humans / Immunoglobulin A, Secretory / Immunoglobulin G / Influenza, Human / Male / Middle Aged / Nasal Mucosa / Neutralization Tests / Orthomyxoviridae / Retrospective Studies / Serum / Young Adult
Norio Kamemura, Hitomi Tada, Naoki Shimojo, Yoshinori Morita, Yoichi Kohno, Takao Ichioka, Koichi Suzuki, Kenji Kubota, Mineyoshi Hiyoshi and Hiroshi Kido : Intrauterine sensitization of allergen-specific IgE analyzed by a highly sensitive new allergen microarray., The Journal of Allergy and Clinical Immunology, Vol.130, No.1, 113-21.e2, 2012.
(要約)
Allergen-specific levels of IgE and IgA antibodies and their allergen profiles analyzed by the diamond-like-carbon allergen chip indicate that IgE antibodies in CB are of fetal origin. Food-allergen specific IgE antibodies were detected more often than inhalant-allergen specific IgE antibodies in CB, the reason of which remains unclarified.
Masaya Kubota, Junji Chida, Hideki Hoshino, Hiroshi Ozawa, Ayaka Koide, Hirohumi Kashii, Akiko Koyama, Yoko Mizuno, Ai Hoshino, Yamaguchi Miyoko, Dengbing Yao, Min Yao and Hiroshi Kido : Thermolabile CPT II variants and low blood ATP levels are closely related to severity of acute encephalopathy in Japanese children, Brain & Development, Vol.34, No.1, 20-27, 2012.
(要約)
Despite the decrease in Reye syndrome after the discontinuation of aspirin, acute encephalopathy (non-Reye syndrome type) has been continually reported in Japan. Recent studies suggested that the thermolabile phenotype of carnitine palmitoyltransferase II (CPT II) variation [F352C] was closely related to the pathomechanism of influenza-associated encephalopathy (IAE) in Japanese, causing mitochondrial ATP utilization failure during periods of high fever, resulting in brain edema. So, we analyzed CPT II polymorphism and peripheral blood ATP levels as a signal of "energy crisis" in 12 and 10 patients with acute encephalopathy, respectively. Out of the 12 patients with acute encephalopathy, six showed thermolabile CPT II variants [F352C], and of these six, two patients died in spite of intensive care. In contrast, the remaining six patients with no thermolabile CPT II variant [F352C] showed a relatively mild clinical course. Blood ATP levels of the 10 patients in the acute phase of encephalopathy were significantly lower than those during the convalescent phase and also those of patients with febrile seizure status. Our data suggest that the thermolabile F352C CPT II variant, found only in Japanese, might be one of the predisposing factors to trigger the pathomechanism of acute encephalopathy in the Japanese population, and that it is causally related to the severity of disease. The decreased blood ATP level seems to reflect systemic mitochondrial dysfunction including the blood brain barrier during the acute phase of encephalopathy.
Hiroshi Kido, Yuushi Okumura, Etsuhisa Takahashi, Haiyan Pan, Siye Wang, Dengbing Yao, Min Yao, Junji Chida and Mihiro Yano : Role of host cellular proteases in the pathogenesis of influenza and influenza-induced multiple organ failure, Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, Vol.1824, No.1, 186-194, 2012.
(要約)
Influenza A virus (IAV) is one of the most common infectious pathogens in humans. Since the IVA genome does not have the processing protease for the viral hemagglutinin (HA) envelope glycoprotein precursors, entry of this virus into cells and infectious organ tropism of IAV are primarily determined by host cellular trypsin-type HA processing proteases. Several secretion-type HA processing proteases for seasonal IAV in the airway, and ubiquitously expressed furin and pro-protein convertases for highly pathogenic avian influenza (HPAI) virus, have been reported. Recently, other HA-processing proteases for seasonal IAV and HPAI have been identified in the membrane fraction. These proteases proteolytically activate viral multiplication at the time of viral entry and budding. In addition to the role of host cellular proteases in IAV pathogenicity, IAV infection results in marked upregulation of cellular trypsins and matrix metalloproteinase-9 in various organs and cells, particularly endothelial cells, through induced pro-inflammatory cytokines. These host cellular factors interact with each other as the influenza virus-cytokine-protease cycle, which is the major mechanism that induces vascular hyperpermeability and multiorgan failure in severe influenza. This mini-review discusses the roles of cellular proteases in the pathogenesis of IAV and highlights the molecular mechanisms of upregulation of trypsins as effective targets for the control of IAV infection. This article is part of a Special Issue entitled: Proteolysis 50 years after the discovery of lysosome.
(キーワード)
Animals / Antigen Presentation / Birds / Capillary Permeability / Host-Pathogen Interactions / Humans / Immune System / Influenza A virus / Influenza in Birds / Influenza, Human / Models, Biological / Multiple Organ Failure / Peptide Hydrolases
Koichi Suzuki, Mineyoshi Hiyoshi, Hitomi Tada, Miwa Bando, Takao Ichioka, Norio Kamemura and Hiroshi Kido : Allergen diagnosis microarray with high-density immobilization capacity using diamond-like carbon-coated chips for profiling allergen-specific IgE and other immunoglobulins, Analytica Chimica Acta, Vol.706, No.2, 321-327, 2011.
(要約)
The diagnosis of antibody-mediated allergic disorders is based on clinical findings, skin prick tests and detection of allergen-specific IgE in serum. Here, we present a new microarray technique of high-density antigen immobilization using carboxylated arms on the surface of a diamond-like carbon (DLC)-coated chip. High immobilization capacity of antigen on DLC chip at (0.94-7.82)×10(9) molecules mm(-2) allowed the analysis of allergen-specific immunoglobulins against not only purified proteins but also natural allergen extracts with wide assay dynamic range. The higher sensitivity of the allergen-specific IgE detection on DLC chip was observed for comparison with the UniCAP system: the DLC chip allowed lowering the limit of dilution rate in UniCAP system to further dilution at 4-8-fold. High correlations (ϱ>0.9-0.85) of allergen-specific IgE values determined by the DLC chip and UniCAP were found in most of 20 different allergens tested. The DLC chip was useful to determine allergen-induced antibodies of IgA, IgG, IgG1, and IgG4 in sera, apart from IgE, as well as secretory IgA in saliva against the same series of allergens on the chip in a minimal amount (1-2 μL) of sample.
Min Yao, Dengbing Yao, Miyoko Yamaguchi, Junji Chida, Dengfu Yao and Hiroshi Kido : Bezafibrate upregulates carnitine palmitoyltransferase II expression and promotes mitochondrial energy crisis dissipation in fibroblasts of patients with influenza-associated encephalopathy, Molecular Genetics and Metabolism, Vol.104, No.3, 265-272, 2011.
(要約)
Influenza-associated encephalopathy (IAE) is characterized by persistently high fever, febrile convulsions, severe brain edema and high mortality. We reported previously that a large proportion of patients with disabling or fatal IAE exhibit a thermolabile phenotype of compound variants for [1055T>G/F352C] and [1102G>A/V368I] of carnitine palmitoyltransferase II (CPT II) and mitochondrial energy crisis during high fever. In the present study, we studied the effect of bezafibrate, a hypolipidemic pan-agonist of peroxisome proliferator-activated receptor (PPAR), on CPT II expression and mitochondrial energy metabolism in fibroblasts of IAE patients and wild type (WT) fibroblasts from a healthy volunteer at 37°C and 41°C. Although heat stress markedly upregulated CPT II, CPT IA and PPAR-δ mRNA expression levels, CPT II activity, β-oxidation and ATP levels in WT and IAE fibroblasts at 41°C were paradoxically downregulated probably due to the thermal instability of the corresponding enzymes. Bezafibrate significantly enhanced the expression levels of the above mRNAs and cellular functions of these enzymes in fibroblasts at 37°C. Bezafibrate-induced increase in CPT II activity also tended to restore the downregulated ATP levels, though moderately, and improved mitochondrial membrane potential even at 41°C to the levels at 37°C in fibroblasts of IAE patients. L-carnitine, a substrate of CPT II, boosted the effects of bezafibrate on cellular ATP levels in WT and IAE fibroblasts, even in severe IAE fibroblasts with thermolabile compound variations of F352C+V368I at 37°C and 41°C. The results suggest the potential usefulness of bezafibrate for the treatment of IAE.
(キーワード)
Adenosine Triphosphate / Base Sequence / Bezafibrate / Blotting, Western / Brain Diseases, Metabolic / Carnitine / Carnitine O-Palmitoyltransferase / DNA Primers / Energy Metabolism / Fibroblasts / Gene Expression Regulation / Genomics / Hot Temperature / Humans / Influenza, Human / Japan / Membrane Potential, Mitochondrial / Microscopy, Fluorescence / Mitochondria / Molecular Sequence Data / Peroxisome Proliferator-Activated Receptors / RNA, Messenger / Real-Time Polymerase Chain Reaction / Sequence Analysis, DNA / Time Factors
Dengbing Yao, Min Yao, Miyoko Yamaguchi, Junji Chida and Hiroshi Kido : Characterization of compound missense mutation and deletion of carnitine palmitoyltransferase II in a patient with adenovirus-associated encephalopathy, The Journal of Medical Investigation : JMI, Vol.58, No.3,4, 210-218, 2011.
(要約)
In mammals, carnitine palmitoyltransferase (CPT) system is a pivotal component of energy metabolism through mitochondrial fatty acid oxidation. The majority of patients with fatal or handicapped influenza-associated encephalopathy exhibit thermolabile compound homo/heterozygous mutations of CPT II. Compound CPT II mutations, [c.647A>G (p.Q216R)], [c.1102G>A (p.V368I)], [c.1939A>G (p.M647V)] and [c.745delG (p.G249EfsX16)], were found in a patient with adenovirus-associated encephalopathy and his family. The properties of these CPT II mutations were analyzed in COS-7 cells. CPT II mutations in the patient and his family were expressed in COS-7 cells and their molecular masses, enzyme activities, thermal instabilities and half-lives were analyzed. We identified two novel CPT II mutations in the patient, [c.647A>G (p.Q216R)] and [c.745delG (p.G249EfsX16)]. The CPT II Q216R mutation showed mild reduction of activity, thermal instability and short half-life but compound mutations with Q216R+V368I+M647V showed further enhancement of these disabilities, although mutations V368I and M647V had no such effects. CPT II mutation [c.745delG (p.G249EfsX16)] abolished enzyme activity and showed short half-life. The thermal instability and short half-life of the novel CPT II mutations, [c.647A>G (p.Q216R)] and [c.745delG (p.G249EfsX16)], could play important roles in energy crisis in the pathogenesis of virus-associated encephalopathy.
Saori Takeda, Ai Fujimoto, Emiko Yamauchi, Mineyoshi Hiyoshi, Hiroshi Kido, Takashi Watanabe, Kozo Kaibuchi, Takeshi Ohta and Hiroaki Konishi : Role of a tyrosine phosphorylation of SMG-9 in binding of SMG-9 to IQGAP and the NMD complex, Biochemical and Biophysical Research Communications, Vol.410, No.1, 29-33, 2011.
(要約)
SMG-9 is a component of the NMD complex, a heterotetramer that also includes SMG-1 and SMG-8 in the complex. SMG-9 was also originally identified as a tyrosine-phosphorylated protein but the role of the phosphorylation is not yet known. In this study, we determined that IQGAP protein, an actin cytoskeleton modifier acts as a binding partner with SMG-9 and this binding is regulated by phosphorylation of SMG-9 at Tyr-41. SMG-9 is co-localized with IQGAP1 as a part of the process of actin enrichment in non-stimulated cells, but not in the EGF-stimulated cells. Furthermore, an increase in the ability of SMG-9 to bind to SMG-8 occurs in response to EGF stimulation. These results suggest that tyrosine phosphorylation of SMG-9 may play a role in the formation of the NMD complex in the cells stimulated by the growth factor.
(キーワード)
Animals / COS Cells / Cercopithecus aethiops / HEK293 Cells / HeLa Cells / Humans / Phosphoproteins / Protein Binding / Serine / ras GTPase-Activating Proteins
Dai Mizuno, Tunetomo Takei, Akiho Fukuta, Wakako Shinahara, Etsuhisa Takahashi, Mihiro Yano and Hiroshi Kido : Surfactant protein C is an essential constituent for mucosal adjuvanticity of Surfacten, acting as an antigen delivery vehicle and inducing both local and systemic immunity, Vaccine, Vol.29, No.33, 5368-5378, 2011.
(要約)
We have reported that Surfacten(®) (St), a bovine pulmonary surfactant free of antigenic c-type lectins, is a useful mucosal adjuvant for nasal vaccination. To prepare ample supplies a synthetic adjuvant that mimics St, we analyzed essential constituents of St for mucosal adjuvanticity. Intranasal inoculation of influenza virus hemagglutinin (HA) vaccine combined with St free of surfactant protein (SP)-C resulted in failure of HA vaccine delivery to dendritic cells and loss of local and systemic immune responses. Naïve bovine SP-C, synthetic human or bovine SP-C peptide reconstituted with three major St lipids restored delivery activity and local and systemic immune responses to levels similar to those of St and provided almost complete protection against lethal doses of influenza virus challenge in mice. The delivery of fluoresceinated HA vaccine to cultured dendritic cells was significantly enhanced by co-administration of St or synthetic adjuvant, and moderately stimulated the expression of MHC class II and CD86. In addition, both St and synthetic adjuvant markedly sustained HA vaccine and achieved a wide antigen distribution in murine nasal cavity. These results suggest that synthetic mucosal adjuvant reconstituted with SP-C peptide and major St lipids is useful for ample supply of the potent mucosal adjuvant as an antigen delivery vehicle for intranasal vaccination.
Hiroshi Kido, Etsuhisa Takahashi, Kosuke Kataoka, Kazuyuki Fujii, Satoshi Suzuki, Kazuhiro Iwase and Chika Ito : Attenuation of respiratory immune responses by antiviral neuraminidase inhibitor treatment and boost of mucosal immunoglobulin A response by co-administration of immuno- modulator clarithromycin in paediatric influenza., Influenza and Other Respiratory Viruses, Vol.5, No.1, 240-243, 2011.
(キーワード)
Tamiflu, Infuenza virus, mucosal Immunology
49.
Etsuhisa Takahashi, Yuushi Okumura and Hiroshi Kido : Activation of the highly pathogenic avian influenza virus replication by membrane-bound proteases MSPL and TMPRSS13 and its inhibition by the protease inhibitors, Influenza and Other Respiratory Viruses, Vol.5, No.1, 276-279, 2011.
50.
Junji Chida, Siye Wang, Pan Hai-Yan, Le Quang Trong and Hiroshi Kido : Influenza virus-cytokine-protease cycles are principal mechanisms of multi-organ failure in severe influenza and therapeutic approaches, Influenza and Other Respiratory Viruses, Vol.5, No.1, 281-286, 2011.
51.
Pan Hai-Yan, Mihiro Yano and Hiroshi Kido : Effects of inhibitors of Toll-like receptors, protease-activated receptor-2 signalings and trypsin on influenza A virus replication and upregulation of cellular factors in cardiomyocytes, The Journal of Medical Investigation : JMI, Vol.58, No.1,2, 19-28, 2011.
(要約)
Severe influenza sometimes causes myocarditis. We recently found that influenza A virus (IAV) infection induces various cellular factors, such as proinflammatory cytokines IL-6, IL-1β and TNF-α, matrix metalloproteinases (MMPs) and ectopic trypsin in mice hearts and in H9c2 cardiomyocytes. The induction of these cellular factors in turn promotes viral replication, myocardial inflammation and cellular damage through their intracellular signal transductions in cooperation with the IAV-induced Toll-like receptors (TLRs) and proteinase-activated receptor-2 (PAR-2) signalings, although the precise nature of these interactions remain obscure. By using specific inhibitors of TLRs and PAR-2 signalings and trypsin inhibitor aprotinin, we analyzed the role of TLR signaling and PAR-2 signaling in the IAV-induced pathological changes in cardiomyocytes. Inhibitors of TLR7/8-Myeloid Differentiation factor 88-nuclear factor-κB signaling and aprotinin effectively suppressed IAV-induced upregulation of proinflammatory cytokines, MMPs, trypsinogen and viral replication. Inhibitor of TLR3-Toll/interleukin-1 receptor domain-containing adaptor inducing interferons-dependent signaling predominantly suppressed the upregulation of interferon-β, a key intracellular host immune response factor. In contrast to the suppressive effect of trypsin inhibitor aprotinin on IAV replication, PAR-2 inhibitor FSY-NH(2), induced marginal upregulation of trypsinogen and subsequent stimulation of IAV replication.
Hiroshi Kido and Kazumi Ishidoh : Nobuhiko Katunuma: an outstanding scientist in the field of proteolysis and warm-hearted 'Kendo Fighter' biochemist., The Journal of Biochemistry, Vol.148, No.5, 527-531, 2010.
(要約)
Professor Nobuhiko Katunuma is well known for his outstanding contribution to the understanding of proteolysis in general and cysteine proteinases and their inhibitors in mammals. In fact, he is a world pioneer in the field. In 1963, he started his highly successful scientific career as a Professor at the Institute for Enzyme Research, the University of Tokushima. During the initial 30 years of his career, he was interested in vitamin B6 metabolism and discovered the acceleration of turnover rates of pyridoxal enzyme in apoprotein formation. After this period, his interest expanded to lysosomal cystein proteinases and their endogenous inhibitors. After determining the crystal structure of human cathepsin B, he generated a series of chemically synthesized specific inhibitors of cathepsins. These inhibitors are currently used throughout the world and some of them have been applied therapeutically in various diseases. During his career and even at present, Professor Katunuma has been studying Biochemistry in Medicine and also practicing to become a 'Kendo sword fencing Fighter'.
(キーワード)
Biochemistry / Cysteine Proteinase Inhibitors / History, 20th Century / History, 21st Century / Japan / Peptide Hydrolases / Vitamin B 6
Hai-Yan Pan, Hirotsugu Yamada, Junji Chida, Siye Wang, Mihiro Yano, Min Yao, Jianhua Zhu and Hiroshi Kido : Up-regulation of ectopic trypsins in the myocardium by influenza A virus infection triggers acute myocarditis, Cardiovascular Research, Vol.89, No.3, 595-603, 2010.
(要約)
Influenza A virus (IAV) infection markedly up-regulates ectopic trypsins in various organs, viral envelope glycoprotein processing proteases, which are pre-requisites for virus entry and multiplication. We investigated the pathological roles of trypsin up-regulation in the progression of IAV-induced myocarditis, cytokine induction, and viral replication in the hearts, and also investigated the protective effects of trypsin inhibitor on cardiac dysfunction in vivo and selective knockdown of trypsin on IAV-induced cellular damage in cardiomyoblasts. The relationship of the expression among IAV RNA, trypsins, matrix metalloproteinase (MMP)-9, MMP-2, pro-inflammatory cytokines interleukin (IL)-6, IL-1β, and tumour necrosis factor-α was analysed in mice hearts and cardiomyoblasts after IAV infection. The severity of myocarditis was most noticeable during Day 6-9 post-infection, along with peak expression of viral RNA, trypsins, particularly trypsin , MMPs, and cytokines. Cardiac ATP levels were the lowest at Day 9. Up-regulated trypsins, viral protein, and tissue-injured loci in the myocardium were closely localized. Trypsin inhibitor aprotinin treatment in vivo and selective trypsin - and trypsin -knockdown, particularly the latter, in H9c2 cardiomyoblasts significantly suppressed viral replication, up-regulation of MMPs, and production of active MMP-9 and cytokines, resulting in marked protection against cellular damage, ATP depletion, and apoptosis. IAV infection-induced cardiac dysfunction monitored by echocardiography was improved significantly by aprotinin treatment. IAV-induced trypsins, particularly trypsin , in the myocardium trigger acute viral myocarditis through stimulation of IAV replication, proMMP-9 activation, and cytokine induction. These results suggest that up-regulation of trypsins is one of the key host pathological findings in IAV-induced myocarditis.
Siye Wang, Le Quang Trong, Kurihara Naoki, Junji Chida, Youssouf Cisse, Mihiro Yano and Hiroshi Kido : Influenza Virus Cytokine Protease Cycle in the Pathogenesis of Vascular Hyperpermeability in Severe Influenza, The Journal of Infectious Diseases, Vol.202, No.7, 991-1001, 2010.
(要約)
Severe influenza is characterized by cytokine storm and multiorgan failure with edema. The aim of this study was to define the impact of the cytokine storm on the pathogenesis of vascular hyperpermeability in severe influenza. Weanling mice were infected with influenza A WSN/33(H1N1) virus. The levels of proinflammatory cytokines, tumor necrosis factor (TNF) alpha, interleukin (IL) 6, IL-1beta, and trypsin were analyzed in the lung, brain, heart, and cultured human umbilical vein endothelial cells. The effects of transcriptional inhibitors on cytokine and trypsin expressions and viral replication were determined. Influenza A virus infection resulted in significant increases in TNF-alpha, IL-6, IL-1beta, viral hemagglutinin-processing protease trypsin levels, and viral replication with vascular hyperpermeability in lung and brain in the first 6 days of infection. Trypsin upregulation was suppressed by transcriptional inhibition of cytokines in vivo and by anti-cytokine antibodies in endothelial cells. Calcium mobilization and loss of tight junction constituent, zonula occludens-1, associated with cytokine- and trypsin-induced endothelial hyperpermeability were inhibited by a protease-activated receptor-2 antagonist and a trypsin inhibitor. The influenza virus-cytokine-protease cycle is one of the key mechanisms of vascular hyperpermeability in severe influenza.
Etsuhisa Takahashi, Kosuke Kataoka, Kazuyuki Fujii, Junji Chida, Dai Mizuno, Makoto Fukui, Hiro-O Ito, Kohtaro Fujihashi and Hiroshi Kido : Attenuation of inducible respiratory immune responses by oseltamivir treatment in mice infected with influenza A virus., Microbes and Infection, Vol.12, No.10, 778-783, 2010.
(要約)
The antiviral neuraminidase inhibitor oseltamivir (OSV) is widely used to suppress viral replication in the treatment of influenza. Here, we report that OSV administration significantly suppressed respiratory mucosal secretory IgA responses with respect to antigen (Ag)-specific antibody (Ab) production and also the induction of Ag-specific IgA Ab-forming cells, but not systemic IgG responses, in weanling mice as a model of pediatric influenza. Neutralizing activities of the airway fluids in oral OSV-treated mice were significantly less than those of sham-treated mice. Our findings suggest the risk of re-infection in patients showing a low mucosal response following OSV treatment.
(キーワード)
Animals / Antibodies, Viral / Antiviral Agents / Female / Immunity, Mucosal / Immunoglobulin A / Immunoglobulin G / Immunosuppression / Immunosuppressive Agents / Influenza A virus / Mice / Mice, Inbred BALB C / Orthomyxoviridae Infections / Oseltamivir
Yuushi Okumura, Etsuhisa Takahashi, Mihiro Yano, Ohuchi Masanobu, Daidoji Tomo, Nakaya Takaaki, Bőttcher Eva, Garten Woflgang, Klenk Hans-Dieter and Hiroshi Kido : Novel type II transmembrane serine proteases, MSPL and TMPRSS13, proteolytically activate membrane fusion activity of hemagglutinin of highly pathogenic avian influenza viruses and induce their multicycle replication., Journal of Virology, Vol.84, No.10, 5089-5096, 2010.
(要約)
Host cellular proteases induce influenza virus entry into cells by cleaving the viral surface envelope glycoprotein hemagglutinin (HA). However, details on the cellular proteases involved in this event are not fully available. We report here that ubiquitous type II transmembrane serine proteases, MSPL and its splice variant TMPRSS13, are novel candidates for proteases processing HA proteins of highly pathogenic avian influenza (HPAI) viruses, apart from the previously identified furin and proprotein convertases 5 and 6. HAs from all HPAI virus H5 and H7 strains have one of two cleavage site motifs, the R-X-K/R-R motif with R at position P4 and the K-K/R-K/T-R motif with K at position P4. In studies of synthetic 14-residue HPAI virus HA peptides with these cleavage site motifs, furin preferentially cleaved only HA peptides with the R-K-K-R motif in the presence of calcium and not peptides with the other motif, whereas MSPL and TMPRSS13 cleaved both types of HA peptides (those with the R/K-K-K-R motif) efficiently in the absence of calcium. Full-length recombinant HPAI virus HA with the K-K-K-R cleavage motif exhibited poor susceptibility to cleavage in the absence of MSPL or TMPRSS13 and the presence of furin in infected cells, but it was converted to mature HA subunits in transfected cells expressing MSPL or TMPRSS13, with membrane-fused giant-cell formation. This conversion and membrane fusion were suppressed by inhibitors of MSPL and TMPRSS13. Furthermore, infection with and multiplication of genetically modified live HPAI virus A/Crow/Kyoto/53/2004 (H5N1) with the K-K-K-R cleavage site motif were detected only in MSPL- and TMPRSS13-expressing cells.
Youssouf Cisse, Siye Wang, Isao Inoue and Hiroshi Kido : Rat model of influenza-associated encephalopathy (IAE): studies of electroencephalogram (EEG) in vivo, Neuroscience, Vol.165, No.4, 1127-1137, 2010.
(要約)
Influenza-associated encephalopathy (IAE) is characterized by severe neurological complications during high-grade fever with high morbidity and mortality in children. The major neurological complications during high-grade fever include convulsive seizures, loss of consciousness, neuropsychiatric behavior (hallucination, meaningless speech, disorientation, laughing alone); high voltage amplitude slow waves and the occurrence of theta oscillation are depicted on the electroencephalogram (EEG) in the IAE patients. At the early phase of the disease, the cytokines levels increase in severe cases. To understand the neuronal properties in the CNS leading to these neurological complications in IAE patients, we recorded EEG signals from the hippocampus and cortex of rats infected with influenza A/WSN/33 H1N1 virus (IAV) strain. Abnormal EEG activities were observed in all infected rats under anesthesia, including high voltage EEG burst amplitude and increased EEG spikes in the early phase (8 h-day 2) of infection, and these increases at the early phase were in parallel with a significant increase level of interleukin-6 (IL-6) in the serum. When the infected rats were heat-stressed by elevating the rat body core temperature to 39-41 degrees C, these abnormal EEG activities were enhanced, and the oscillation pattern shifted in most of rats from slow bursting waves (<1 Hz) to theta oscillation (3-6 Hz). These results indicate that the abnormal EEG activities in IAE patients could be well reproduced in anesthetized IAV infected rats under hyperthermia, hence this animal model will be useful for further understandings the mechanism of neuronal complications in IAE patient during high-grade fever.
Siye Wang, Le Quang Trong, Junji Chida, Youssouf Cisse, Mihiro Yano and Hiroshi Kido : Meckanisms of matrix metallo protease-9 wuregulation and tissue desfruction in rarious organs in influenza A virus infection, The Journal of Medical Investigation : JMI, Vol.57, No.1,2, 26-34, 2010.
(要約)
Severe influenza is characterized clinicopathologically by multiple organ failure, although the relationship amongst virus and host factors that influence this morbid outcome and the underlying mechanisms of action remain unclear. The present study identified marked upregulation of matrix metalloproteinase (MMP)-9 and pro-inflammatory cytokine tumor necrosis factor alpha (TNF-alpha) in various organs after intranasal infection of influenza A WSN virus. MMP-9 and TNF-alpha were upregulated in the lung, the site of initial infection, as well as in the brain and heart. The infection-induced MMP-9 upregulation was inhibited by anti-TNF-alpha antibodies and by anti-oxidative reagents pyrrolidine dithiocarbamate and N-acetyl-L-cysteine, which inhibit activation of nuclear factor kappa B (NF-kappaB), as well as by nordihydroguaiaretic acid, which inhibits activation of activator protein 1 (AP-1). In addition, MMP-9 upregulation via TNF-alpha was also suppressed by inhibitors of mitogen-activated protein kinases (MAPKs), such as extracellular signal-regulated kinase 1/2 and p38, and partly by a c-Jun N-terminal kinase inhibitor. These results indicated that the influenza-induced MMP-9 upregulation in various organs is mediated through MAPK-NF-kappaB- and/or AP-1-dependent mechanisms. Strategies that neutralize TNF-alpha as well as inhibitors of MAPK-NF-kappa B- and/or AP-1-dependent pathways may be useful for suppressing the MMP-9 effect and thus preventing multiple organ failure in severe influenza.
Yiu-Wing Kam, Yuushi Okumura, Hiroshi Kido, Lisa P. F. Ng, Roberto Bruzzone and Ralf Altmeyer : Cleavage of the SARS coronavirus spike glycoprotein by airway proteases enhances virus entry into human bronchial, PLoS ONE, Vol.4, No.11, e7870, 2009.
(要約)
Entry of enveloped viruses into host cells requires the activation of viral envelope glycoproteins through cleavage by either intracellular or extracellular proteases. In order to gain insight into the molecular basis of protease cleavage and its impact on the efficiency of viral entry, we investigated the susceptibility of a recombinant native full-length S-protein trimer (triSpike) of the severe acute respiratory syndrome coronavirus (SARS-CoV) to cleavage by various airway proteases. PURIFIED TRISPIKE PROTEINS WERE READILY CLEAVED IN VITRO BY THREE DIFFERENT AIRWAY PROTEASES: trypsin, plasmin and TMPRSS11a. High Performance Liquid Chromatography (HPLC) and amino acid sequencing analyses identified two arginine residues (R667 and R797) as potential protease cleavage site(s). The effect of protease-dependent enhancement of SARS-CoV infection was demonstrated with ACE2 expressing human bronchial epithelial cells 16HBE. Airway proteases regulate the infectivity of SARS-CoV in a fashion dependent on previous receptor binding. The role of arginine residues was further shown with mutant constructs (R667A, R797A or R797AR667A). Mutation of R667 or R797 did not affect the expression of S-protein but resulted in a differential efficacy of pseudotyping into SARS-CoVpp. The R667A SARS-CoVpp mutant exhibited a lack of virus entry enhancement following protease treatment. These results suggest that SARS S-protein is susceptible to airway protease cleavage and, furthermore, that protease mediated enhancement of virus entry depends on specific conformation of SARS S-protein upon ACE2 binding. These data have direct implications for the cell entry mechanism of SARS-CoV along the respiratory system and, furthermore expand the possibility of identifying potential therapeutic agents against SARS-CoV.
Sawabuchi Takako, Suzuki Satoshi, Isase Kazuhiro, Ito Chika, Dai Mizuno, Todari Hajime, Watanabe Isamu, Talukder R. Sadiqur, Junji Chida and Hiroshi Kido : Boost of mucosal secretory immunoglobulin A response by clarithromycin in paediatric influenza, Respirology, Vol.14, No.8, 1173-1179, 2009.
(要約)
The antiviral neuraminidase inhibitor oseltamivir (OSV) is used to treat influenza. The macrolide clarithromycin (CAM) is used to treat bacterial infections and has anti-inflammatory and immunomodulatory activities. This retrospective study investigated the immunomodulatory effects of CAM in children presenting with influenza A. The study recruited 40 children with acute influenza, and grouped them according to the treatment received: 5-day treatment with OSV (n = 14), CAM (n = 8), OSV + CAM (n = 12) and untreated (n = 6). The before and after treatment comparisons were made of the level of secretory IgA (sIgA) against influenza A virus (H3N2) and (H1N1), total sIgA, viral RNA copy numbers in nasopharyngeal aspirates and disease symptoms. Infection induced anti-viral mucosal sIgA in the nasopharyngeal aspirates of most patients of all treatment groups. Particularly prominent increases in the levels were found in the CAM and OSV + CAM groups. Low induction of anti-viral sIgA was observed in the OSV group, but the addition of CAM to OSV augmented sIgA production and restored local mucosal sIgA levels. The frequency of residual cough in the OSV + CAM group was significantly lower than in the other groups including the group treated with OSV. CAM boosted the nasopharyngeal mucosal immune response in children presenting with influenza A, even in those treated with OSV who had low production of mucosal anti-viral sIgA, and alleviated the symptoms of influenza.
Maki Nisino, Dai Mizuno, Kimoto Takashi, Shinahara Wakako, Fukuta Akiho, Tunetomo Takei, Kaori Sumida, Seiichiro Kitamura, Hiroshi Shiota and Hiroshi Kido : Influenza vaccine with Surfacten, a modified pulmonary surfactant, induces systemic and mucosal immune responses without side effects in minipigs, Vaccine, Vol.27, No.41, 5620-5627, 2009.
(要約)
Immune responses and side effects of intranasally administered flu vaccine with the commercial product Surfacten, a modified bovine pulmonary surfactant, were investigated in minipigs. The use of minipigs was based on the anatomical resemblance of nasal lymph nodes, the principal antigen uptake site of respiratory mucosal immunity, between pig and human. Intranasal instillation of HA vaccine adjuvanted with Surfacten elicited significantly higher serum hemagglutination inhibition titers than the antigen alone, with wide cross-neutralizing activities of secretory IgA in nasal washes. No significant induction of inflammatory cytokines or migration of inflammatory cells was observed at the site of immunization or serum after the first immunization. These data suggest the potential usefulness of Surfacten for mucosal vaccination.
Mineyoshi Hiyoshi, Hiroaki Konishi, Hirokazu Uemura, Hideki Matsuzaki, Hideo Tsukamoto, Ryusuke Sugimoto, Hideo Takeda, Satoru Dakeshita, Atsushi Kitayama, Hidenobu Takami, Fusakazu Sawachika, Hiroshi Kido and Kokichi Arisawa : D-dopachrome tautomerase is a candidate for key proteins to protect the rat liver damaged by carbon tetrachloride, Toxicology, Vol.255, No.1,2, 6-14, 2009.
(要約)
Carbon tetrachloride (CCl4) is known to induce liver damage. Animal experiments with CCl4 injections have revealed many findings, especially mechanisms of liver damage and liver regeneration. Recently, proteomic approaches have been introduced in various studies to evaluate the quantitative and qualitative changes in the comprehensive proteome level. The aim of this research is to elucidate the key protein for liver damage, liver protection and liver regeneration by using proteomic techniques. 50 % (v/v) CCl4 in corn oil was administered intraperitoneally to adult male rats at a dose of 4ml/kg body weight. Approximately 24h after the injection, the liver was removed and extracted proteins were analyzed with cleavable isotope coded affinity tag (cICAT) reagents, two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS). A twelvefold increase in D-dopachrome tautomerase (DDT) was indicated. This enzyme has been reported to be involved in the biosynthesis of melanin, an antioxidant. According to the histological analysis, melanin levels were increased in un-damaged hepatocytes of CCl4-treated rats. These results suggest that the increase in DDT is a response to liver damage, accelerates melanin biosynthesis and protects the liver from oxidative stress induced by CCl4.
(キーワード)
Animals / Blotting, Western / Body Weight / COS Cells / Carbon Tetrachloride Poisoning / Cercopithecus aethiops / Cloning, Molecular / DNA, Complementary / Drug-Induced Liver Injury / Genetic Vectors / Intramolecular Oxidoreductases / Liver / Male / Mass Spectrometry / Melanins / Organ Size / Rats / Rats, Wistar / Transfection
Hiroshi Kido, Yuushi Okumura, Etsuhisa Takahashi, HY Pan, Siye Wang, Junji Chida, Le Trong Quang and Mihiro Yano : Host envelope glycoprotein processing proteases are indispensable for entry into human cells by seasonal and highly pathogenic avian influenza viruses., Journal of Molecular and Genetic Medicine, Vol.3, No.1, 167-175, 2009.
(要約)
Influenza A virus (IAV) is one of the most common infectious pathogens in humans and causes considerable morbidity and mortality. The recent spread of highly-pathogenic avian IAV H5N1 viruses has reinforced the importance of pandemic preparedness. In the pathogenesis of IAV infection, cellular proteases play critical roles in the process of viral entry into cells that subsequently leads to tissue damage in the infected organs. Since there are no processing protease for the viral membrane fusion glycoprotein hemagglutinin precursor (HA(0)) in IAV, entry of the virus into cells is determined primarily by the host cellular HA(0) processing proteases that proteolytically activate membrane fusion activity. HA(0) of seasonal human IAV has the consensus cleavage site motif Q(E)-T/X-R and is selectively processed by at least seven different trypsin-type processing proteases identified to-date in animal model experiments using mouse-adapted IAV or gene expression system in MDCK cells. As is the case for the highly pathogenic avian influenza (HPAI) A virus, endoproteolytic processing of the HA(0) occurs through ubiquitous cellular processing proteases, which selectively recognize the multi-basic consensus cleavage site motifs, such as R-X-K/R-R, and K-X-K/R-R. The cleavage enzymes for the R-X-K/R-R motif, but not K-X-K/R-R motif, have been reported to be furin and pro-protein convertase (PC)5/6 in the trans-Golgi network. Here we report new members of type II transmembrane serine proteases of the cell membrane, mosaic serine protease large form (MSPL) and its splice variant TMPRSS13, which recognize and cleave both R-X-K/R-R and K-X-K/R-R motifs without calcium. Furthermore, IAV infection significantly up-regulates a latent ectopic pancreatic trypsin, one of the potent HA processing proteases, and pro-matrix metalloprotease-9, in various organs. These proteases may synergistically damage the blood-brain barrier in the brain and basement membrane of blood vessels in various organs, resulting in severe edema and multiple organ failure. In this review, we discuss these proteases as new drug target molecules for IAV treatment acting by inhibition of IAV multiplication and prevention of multiple organ failure, other than anti-viral agents, viral neuraminidase inhibitors.
(文献検索サイトへのリンク)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 19565019
Fujimoto Chisa, Hiroshi Kido, Sawabuchi Takako, Dai Mizuno, Hayama Masaki, Hiroaki Yanagawa and Noriaki Takeda : Evaluation of nasal IgA secretion in normal subjects by nasal spray and aspiration., Auris, Nasus, Larynx, Vol.36, No.3, 300-304, 2009.
(要約)
Nasal washing (NW) is a popular method for collecting human nasal lavage fluid. However, for NW the subject must be trained, and the method is unsuitable for field studies on untrained subjects. To overcome this problem, we have developed an easy and painless method, a nasal spray and aspiration (NSA) method. This method is different from NW in that the nasal cavity is misted over with saline, and the nasal lavage fluid is aspirated from the nostrils through a silicon tube. First, nasal lavage fluid was obtained twice by NSA with an interval of a week between lavages to evaluate intraindividual variability, and the IgA and protein levels in the nasal lavage fluid were measured by enzyme-linked immunosorbent assay and bicinchoninic acid assay, respectively. Next, the IgA value determined by NSA was compared with that by NW in another 12 normal subjects 2 days after NSA. In 10 normal subjects, mean volume of saline sprayed into the nose was 0.46+/-0.15 ml (mean+/-S.D.). Mean volume of aspirated nasal lavage fluid containing both sprayed saline and nasal secretion was 0.44+/-0.37 ml. The mean IgA level/mg protein in the nasal lavage fluid determined by NSA was 112+/-18 microg/mg protein at the first and 99+/-20 at the second times of measurement, being highly reproducible. The mean value by NSA was 114+/-19 microg/mg protein, being almost the same as that by NW of 99+/-27. These findings suggest that the IgA level/mg protein in nasal lavage fluid determined by NSA instead of NW might be useful for assessing the variability of nasal IgA secretion.
(キーワード)
Administration, Inhalation / Adult / Female / Humans / Immunoglobulin A / Indicators and Reagents / Male / Middle Aged / Nasal Lavage / Nasal Mucosa / Quinolines / Suction / Young Adult
Daisuke Hashimoto, Masaki Ohmuraya, Masahiko Hirota, Akitsugu Yamamoto, Koichi Suyama, Satoshi Ida, Yuushi Okumura, Etsuhisa Takahashi, Hiroshi Kido, Kimi Araki, Hideo Baba, Noboru Mizushima and Ken-ichi Yamamura : Involvement of autophagy in trypsinogen activation within the pancreatic acinar cells, The Journal of Cell Biology, Vol.18, No.7, 1065-1072, 2008.
(要約)
Autophagy is mostly a nonselective bulk degradation system within cells. Recent reports indicate that autophagy can act both as a protector and killer of the cell depending on the stage of the disease or the surrounding cellular environment (for review see Cuervo, A.M. 2004. Trends Cell Biol. 14:70-77). We found that cytoplasmic vacuoles induced in pancreatic acinar cells by experimental pancreatitis were autophagic in origin, as demonstrated by microtubule-associated protein 1 light chain 3 expression and electron microscopy experiments. To analyze the role of macroautophagy in acute pancreatitis, we produced conditional knockout mice lacking the autophagy-related 5 gene in acinar cells. Acute pancreatitis was not observed, except for very mild edema in a restricted area, in conditional knockout mice. Unexpectedly, trypsinogen activation was greatly reduced in the absence of autophagy. These results suggest that autophagy exerts devastating effects in pancreatic acinar cells by activation of trypsinogen to trypsin in the early stage of acute pancreatitis through delivering trypsinogen to the lysosome.
Shinichi Nakamuta, Hiroshi Endo, Youichiro Higashi, Aoi Kousaka, Aoi Kousaka, Mihiro Yano and Hiroshi Kido : HIV-1 gp120-mediated disruption of tight junction proteins by induction of proteasome-mediated degradation of zonula occludens-1 and -2 in human brain microvascular endothelial cells, Journal of NeuroVirology, Vol.14, No.3, 186-195, 2008.
(要約)
The infiltration of human immunodeficiency virus (HIV)-1, such as by HIV-infected leukocytes, across an injured blood-brain barrier (BBB) is a characteristic pathologic manifestation of HIV-1-associated dementia. HIV-1 gp120 has been implicated as a cause of breakdown of tight junctions between endothelial cells of the BBB, though the disrupting molecular mechanisms are unexplained. This study offers a new explanation for the increased BBB microvascular permeability, due to the degradation of tight junction proteins by the proteasome induced by gp120, and the negative regulation of this process by the scaffold protein, 14-3-3tau. gp120 reduced the amount of zonula occludens (ZO)-1 and ZO-2 in human brain microvascular endothelial cells (HBMECs). The treatment of HBMECs with the proteasome inhibitor, lactacystin, blocked the degradation of ZO-1 and ZO-2, suggesting that these proteins were targeted by gp120 for degradation by the proteasome. gp120 also specifically increased the expression of 14-3-3tau in HBMECs, and its down-regulation by RNAi facilitated the breakdown of tight junction proteins induced by gp120. Our results demonstrate the novel molecular mechanisms of the BBB breakdown by gp120.
Dengbing Yao, Hiroshi Mizuguchi, Miyoko Yamaguchi, Hiroshi Yamada, Junji Chida, Koji Shikata and Hiroshi Kido : Thermal instability of compound variants of carnitine palmitoyltransferase II and impaired mitochondrial fuel utilization in influenza-associated encephalopathy., Human Mutation, Vol.29, No.5, 718-727, 2008.
(要約)
Influenza-associated encephalopathy (IAE) is characterized by persistent high fever, febrile convulsions, severe brain edema, and high mortality in otherwise apparently healthy individuals. We have reported that a large proportion of patients suffering from disabling or fatal IAE, with transiently elevated serum acylcarnitine during high fever, exhibit a thermolabile phenotype of compound homo-/heterozygous variants of carnitine palmitoyltransferase II (CPT II, gene symbol CPT2). We characterized the enzymatic properties of five single and three compound CPT II variants in patients with IAE. The kinetic characteristics of WT and variant CPT IIs, expressed in COS-7 cells, indicated that the variants exert a dominant-negative effect on the homotetrameric protein of the enzyme. Among the variants, three compound variations found in patients with severe encephalopathy; [c.1055T>G (p.Phe352Cys); c.1102G>A (p.Val368Ile)], [c.1511C>T (p.Pro504Leu); c.1813G>C (p.Val605Leu)], and [c.1055T>G (p.Phe352Cys); c.1102G>A (p.Val368Ile); c.1813G>C (p.Val605Leu)], showed reduced activities, thermal instability, and short half-lives compared with the WT. Like other disease-causing mutant proteins, these variant proteins were poly-ubiquitinated and rapidly degraded by a lactacystin-sensitive proteasome pathway. COS-7 cells transfected with the compound variants had their fatty acid beta-oxidation decreased to 30-59% and intracellular ATP levels to 48-79%, and a marked reduction of mitochondrial membrane potential at 41 degrees C, compared with control cells transfected with WT at 37 degrees C. The unstable CPT II variants with decreased enzymatic activities may bring mitochondrial fuel utilization below the phenotypic threshold during high fever, and thus may play an important etiopathological role in the development of brain edema of IAE.
Arunasiri Iddamalgoda, Quang Trong Le, Kenichi Ito, Kiyotaka Tanaka, Hiroyuki Kojima and Hiroshi Kido : Mast cell tryptase and photoaging: possible involvement in the degradation of extra cellular matrix and basement membrane proteins, Archives of Dermatological Research, Vol.300, No.Suppl 1, S69-76, 2008.
(要約)
Mast cells are widely distributed in the connective tissue of the body, but are particularly prominent in tissues such as skin. An increased number of mast cells can be found in the dermis under inflammatory conditions and ultraviolet (UV) exposed skin. Previous investigations have identified matrix metalloproteinases (MMPs) as key enzymes in the degradation of extra cellular matrix (ECM). This study reports about the potential contribution of human mast cell tryptase as a new triggering enzyme in matrix degradation process. Recent studies suggest that mast cell-derived proteases can activate MMPs. We investigated both the degradation of cellular matrix components and activation of MMPs by human tryptase. Mast cells are increased in photoaged skin and the increase of mast cell tryptase in UV irradiated skin was confirmed. Human mast cell tryptase was purified from human tonsils by a series of standard chromatographic procedures. Degradation of collagen type I was achieved by incubation of human type I collagen with tryptase and the fragments were quantified by SDS-PAGE and staining with Coomassie Brilliant Blue 250-R (CBB). Treatment with tryptase resulted in the activation of proMMP-9 as revealed by gelatinolytic activity in type IV collagen zymography. When tryptase was incubated with human type IV collagen, gradual degradation of intact collagen was detected by Western blotting. Furthermore, type IV collagen degradation was observed in the basement membrane (BM) of a three-dimensional (3D) skin model. Degranulation of mast cells, which release tryptase, can activate MMPs and causes direct damage to ECM proteins. These findings strongly implicate that tryptase either alone or in conjunction with activation of MMPs, can participate in ECM damage and the possible destruction of BM leading to photoaging.
Hiroshi Kido and Yuushi Okumura : MSPL/TMPRSS13, Frontiers in Bioscience, Vol.13, 754-758, 2008.
(要約)
Two variant cDNAs, mosaic seine protease large-form (MSPL) and transmembrane protease serine 13 (TMPRSS13), have been identified from a human lung cDNA library by polymerase chain reaction. The deduced amino acid sequences of these proteins show a type II transmembrane protein structure with a unique and long cytoplasmic tail containing tandem repeat phosphorylation motifs of protein kinases, a transmembrane domain, and a trypsin-like serine protease domain at the extracellular C-terminal side. These proteins have an identical serine protease sequence except for the C-terminal ends, and the consensus protease domain exhibits 42, 39 and 43% sequence identity with those of plasma kallikrein, hepsin and transmembrane protease serine 2 (TMPRSS2), respectively. Although both genes are widely expressed in various tissues, they are predominantly expressed in human lung, placenta, pancreas and prostate. TMPRSS13 is expressed higher than MSPL in thymus, spleen and peripheral blood lymphocytes, particularly in CD8+ cells and CD19+ cells. Enzymatic properties of the recombinant soluble MSPL and TMPRSS13 show that these enzymes preferentially recognize the sites consisting of paired basic amino acid residues, and are strongly inhibited by aprotinin, benzamidine and Bowman-Birk trypsin inhibitor, but poorly inhibited by alpha 1-antitrypsin and leupeptin. These properties raise the possibility that MSPL and TMPRSS13 play roles in the proteolytic processing of prohormones, precursors of growth factors, and also play roles in the pathogenicity of many viruses and bacteria in vivo.
Masamichi Kuwajima, Hiroaki Fujihara, Hiroyoshi Sei, Asako Umehara, Masako Sei, Tomi T. Tsuda, Akiko Sukeno, Tatsuya Okamoto, Akiko Inubushi, Yoichi Ueta, Toshio Doi and Hiroshi Kido : Reduced carnitine level causes death from hypoglycemia: possible involvement of suppression of hypothalamic orexin expression during weaning period., Endocrine Journal, Vol.54, No.6, 911-925, 2007.
(要約)
The mechanism of onset of hypoglycemia in patients with carnitine deficiency has yet to be determined. Using mice with systemic carnitine deficiency (JVS mice), we examined this mechanism, focusing on the weaning period (days 14-28 postpartum). For normal mice, the survival rate was 100%, and no hypoglycemia was observed at all. Gastric lactose began to decrease on day 17, and cellulose increased sharply in amount thereafter. For JVS mice, the survival rate was 77% on day 14 and 28% on day 28. From day 21 on, hypoglycemia was noted. Gastric lactose had disappeared almost completely by day 17, and cellulose was almost undetectable from days 14 to 28. Expression of orexin mRNA in the hypothalamus did not differ between JVS and normal mice on day 14, but was suppressed in JVS mice on days 21 and 28. When JVS mice were fed a carnitine-rich diet, suppression of expression of orexin mRNA in hypothalamus was eliminated, and on day 28 lactose and cellulose were detected in the stomach without hypoglycemia. In conclusion, the suppression of the expression of orexin in the hypothalamus during the weaning period may be involved in the marked anorexia in JVS mice, which eventually leads to death from hypoglycemia.
J. Aoki, T. Yasuno, H. Sugie, Hiroshi Kido, I. Nishino, Y. Shigematsu, M. Kanazawa, M. Takayanagi, M. Kumai, K. Endo, H. Kaneoka, M. Yamaguchi, T. Fukuda and T. Yamamoto : A Japanese Adult form of CPT II Deficiency Associated with a Homozygous F383Y Mutation, Neurology, Vol.69, No.8, 804-806, 2007.
Masaki Hayama, Yuushi Okumura, Etsuhisa Takahashi, Aki Shimabukuro, Manabu Tamura, Noriaki Takeda, Takeshi Kubo and Hiroshi Kido : Identification and analysis of the promoter region of the type II transmembrane serine protease polyserase-1 and its transcript variants, Biological Chemistry, Vol.388, No.8, 853-858, 2007.
(要約)
Polyserase-1/TMPRSS9 and its alternative transcripts, serase-1B and serase-2B, are novel type II transmembrane serine proteases that may regulate physiological and pathological phenomena on the cell surface. To understand the mechanisms of gene expression and regulation of these transcripts, we cloned and characterized the 5' promoter region of the mouse polyserase-1 (mpolyserase-1) gene. Using 5'-rapid amplification of cDNA ends, we located the transcription initiation site 272 nucleotides upstream of the translation initiation site. Luciferase reporter gene analysis revealed that the region from +186 to +272 bp in the 5'-untranslated region (UTR), containing the GATA motif (AGATAA), glucocorticoid responsible element (TGTTCT), and E-box sequence (CAGGTG), is required for maximal promoter activity. Mutations introduced into the E-box sequence but not elsewhere in the promoter region caused a selective decrease in transcriptional activity. Furthermore, a DNA probe (+229 to +255 bp) containing the E-box sequence formed a single nuclear protein complex in a sequence-specific manner. These data suggest that the expression of mpolyserase-1 and its transcript variants is positively regulated by the E-box in its 5'-UTR, which might be responsible for the binding of basic helix-loop-helix transcription factors involved in the development of various organelles.
(キーワード)
5' Flanking Region / Animals / Base Sequence / Cell Line / Cloning, Molecular / E-Box Elements / Electrophoretic Mobility Shift Assay / Gene Expression Regulation, Enzymologic / Genes, Reporter / Mice / Molecular Sequence Data / Nuclear Proteins / Promoter Regions, Genetic / Protein Binding / RNA, Messenger / Serine Endopeptidases / Transcription Initiation Site
Hamuro Tsutomu, Hiroshi Kido, Asada Yujiro, Hatakeyama Kinta, Okumura Yuushi, Kunori Youichi, Kamimura Takashi, Iwanaga Sadaaki and Kamei Shintaro : Tissue factor pathway inhibitor is highly susceptible to chymase-mediated proteolysis, The FEBS Journal, Vol.274, No.12, 3065-3077, 2007.
(要約)
Tissue factor pathway inhibitor (TFPI) is a multivalent Kunitz-type protease inhibitor that primarily inhibits the extrinsic pathway of blood coagulation. It is synthesized by various cells and its expression level increases in inflammatory environments. Mast cells and neutrophils accumulate at sites of inflammation and vascular disease where they release proteinases as well as chemical mediators of these conditions. In this study, the interactions between TFPI and serine proteinases secreted from human mast cells and neutrophils were examined. TFPI inactivated human lung tryptase, and its inhibitory activity was stronger than that of antithrombin. In contrast, mast cell chymase rapidly cleaved TFPI even at an enzyme to substrate molar ratio of 1:500, resulting in markedly decreased TFPI anticoagulant and anti-(factor Xa) activities. N-terminal amino-acid sequencing and MS analyses of the proteolytic fragments revealed that chymase preferentially cleaved TFPI at Tyr159-Gly160, Phe181-Glu182, Leu89-Gln90, and Tyr268-Glu269, in that order, resulting in the separation of the three individual Kunitz domains. Neutrophil-derived proteinase 3 also cleaved TFPI, but the reaction was much slower than the chymase reaction. In contrast, alpha-chymotrypsin, which shows similar substrate specificities to those of chymase, resulted in a markedly lower level of TFPI degradation. These data indicate that TFPI is a novel and highly susceptible substrate of chymase. We propose that chymase-mediated proteolysis of TFPI may induce a thrombosis-prone state at inflammatory sites.
Mihiro Yano, Nakamuta Shinichi, Shiota Mayumi, Endo Hiroshi and Hiroshi Kido : Gatekeeper Role of 14-3-3τ Protein in HIV-1 gp120-Mediated Apotosis of Human Endothelial Cells by Inactivation of Bad, AIDS, Vol.21, No.8, 911-920, 2007.
(要約)
HIV-1-associated dementia (HAD) is a major neurological complication often observed in the advanced stages of AIDS. We have reported that 14-3-3 proteins in cerebrospinal fluid, reflecting neuronal cell destruction, is a real-time marker of HAD progression. This study was designed to examine the role of 14-3-3 proteins in HAD. An in-vitro human umbilical vein endothelial cells (HUVEC) model of gp120 protein-induced apoptosis to study the protective role of 14-3-3 in HIV-1 gp120/CXCR4-mediated cell death. The alpha-chemokine receptor-mediated cell death by HIV-1 envelope protein, gp120, the critical event that causes neuron loss and endothelial cell injury, was evaluated in HUVEC undergoing gp120-induced apoptosis through the CXCR4 receptor. We studied the effects of siRNA for each 14-3-3 isoform on the death of HUVEC treated with CXCR4-preferring gp120 (IIIB). Gp120 increased the expression of 14-3-3tau in HUVEC. The binding of Gp120 to CXCR4 induced apoptosis of HUVEC through decreased binding of 14-3-3tau to the pro-apoptotic molecule, Bad. Treatment of the cells with dsRNA against 14-3-3tau enhanced the gp120-mediated dephosphorylation of Bad and its association with Bcl-XL in mitochondria, accelerating the gp120-induced apoptosis, whereas suppression of Bad by RNAi rescued the cells from apoptosis triggered by gp120. The specific up-regulation of 14-3-3tau in HUVEC negatively regulated gp120/CXCR4-mediated cell death by protecting Bad dephosphorylation.
(キーワード)
14-3-3 Proteins / Apoptosis / Cells, Cultured / Cytochromes c / Cytosol / Down-Regulation / Endothelium, Vascular / HIV Envelope Protein gp120 / HIV Envelope Protein gp160 / Humans / Mitochondria / Phosphorylation / Recombinant Proteins / Reverse Transcriptase Polymerase Chain Reaction / Translocation, Genetic / Up-Regulation / bcl-Associated Death Protein
Hiroshi Kido, Yuushi Okumura, Hiroshi Yamada, Le Q. Trong and Mihiro Yano : Proteases Essential for Human Influenza Virus Entry into Cells and Their Inhibitors as Potential Therapeutic Agents, Current Pharmaceutical Design, Vol.13, No.4, 405-414, 2007.
(要約)
Influenza A virus (IAV) is one of the most common infectious pathogens in humans. Since IVA genome does not have the processing protease for the viral membrane fusion glycoprotein precursors, entry of this virus into cells is determined primarily by host cellular, trypsin-type, processing proteases that proteolytically activate the fusion glycoprotein precursors of IAV. At least five different processing proteases have been identified in the airways of animals and humans. These proteases determine the infectious organ tropism of IAV infection as well as the efficiency of viral multiplication in the airway, and sometimes in the brain. Proteases in the upper respiratory tract are suppressed by secretory leukoprotease inhibitor, and those in the lower respiratory tract are suppressed by pulmonary surfactant which, by adsorption, inhibits the interaction between the proteases and viral membrane proteins. Since protease activities predominate over those of endogenous inhibitory compounds under normal airway conditions, administration of protease inhibitors in the early-stage of infection significantly suppresses viral entry and viral multiplication. Several viral neuraminidase inhibitors are used clinically as anti-influenza virus agents, based on their inhibitory action on viral release from infected cells. Furthermore, protease inhibitors of viral entry could be potentially useful against influenza virus as well as neuraminidase inhibitor-resistant viruses. We also found that ambroxol, a mucolytic and anti-oxidant agent, up-regulates the levels of endogenous protease inhibitory compounds in the airway fluids in early-phase infection, and that clarithromycin, a macrolide antibiotic, increases IgA levels and mucosal immunity through augmentation of interleukin-12 levels in the airway. The combination of neuraminidase inhibitors and protease inhibitors, clarithromycin or ambroxol, could be potentially used as a potent anti-influenza therapy to minimize the emergence of drug-resistant mutant viruses.
Yuushi Okumura, M Hayama, Etsuhisa Takahashi, M Fujiuchi, A Shimabukuro, Mihiro Yano and Hiroshi Kido : Serase-1B, a new splice variant of polyserase-1/TMPRSS9, activates urokinase-type plasminogen activator and the proteolytic activation is negatively regulated by glycosaminoglycans., The Biochemical Journal, Vol.400, No.3, 551-561, 2006.
(要約)
Polyserase-1 (polyserine protease-1)/TMPRSS9 (transmembrane serine protease 9) is a type II transmembrane serine protease (TTSP) that possesses unique three tandem serine protease domains. However, the physiological function of each protease domain remains poorly understood. We discovered a new splice variant of polyserase-1, termed Serase-1B, which contains 34 extra amino acids consisting a SEA module (a domain found in sea urchin sperm protein, enterokinase and agrin) adjacent to the transmembrane domain and the first protease domain with a mucin-like box at the C-terminus. The tissue distribution of this enzyme by RT (reverse transcription)-PCR analysis revealed high expression in the liver, small intestine, pancreas, testis and peripheral blood CD14+ and CD8+ cells. To investigate the role of Serase-1B, a full-length form recombinant protein was produced. Interestingly, recombinant Serase-1B was partly secreted as a soluble inactive precursor and it was also activated by trypsin. This activated enzyme selectively cleaved synthetic peptides for trypsin and activated protein C, and it was inhibited by several natural serine protease inhibitors, such as aprotinin, alpha2-antiplasmin and plasminogen activator inhibitor 1. In addition, Serase-1B efficiently converted pro-uPA (urokinase-type plasminogen activator) into active uPA and this activation was strongly inhibited by these natural inhibitors. Furthermore, this activation was also negatively regulated by glycosaminoglycans. Our results indicate that Serase-1B is a novel member of TTSPs that might be involved in uPA/plasmin-mediated proteolysis and possibly implicated in biological events such as fibrinolysis and tumour progression.
(キーワード)
Alternative Splicing / Amino Acid Sequence / Animals / Base Sequence / Cell Line / Cloning, Molecular / Down-Regulation / Female / Gene Expression Regulation / Glycosaminoglycans / Humans / Male / Mice / Mice, Inbred C57BL / Molecular Sequence Data / Organ Specificity / Protein Isoforms / Serine Endopeptidases / Substrate Specificity / Urokinase-Type Plasminogen Activator
Recent progress in life science technology and the availability of much information on genes obtained by genome analysis has enabled us to analyze the changes of proteins on a large scale. Sets of proteins are called proteomes, and proteomics is the scientific field of proteome analysis including differential, post translational modification and interaction analyses. Various proteomic techniques, particularly two-dimensional gel electrophoresis (2-DE), mass spectrometry, protein chip methods, and surface plasmon resonance (SPR), are very useful for acquiring proteomes in cells, tissues and body fluid, and for analyzing interactions between a protein and other biofactors including proteins. A proteomic approach is also useful for determining biomarkers of diseases and key proteins involved in various stages of metabolism such as differentiation, cell cycle and apoptosis. Environmental pollutants including endocrine disruptors inhibit activities of various organs in wild animals and humans. Proteomic approaches could be very useful tools for elucidating the mechanisms of damage caused by environmental pollutants. In this review, we describe the application of a proteomic approach to the field of environmental medicine.
Dengfu Yao, Masamichi Kuwajima, Chen Ye, Mayumi Shiota, Yuushi Okumura, Hiroshi Yamada and Hiroshi Kido : Impaired long-chain fatty acid metabolism in mitochondria causes brain vascular invasion by a non-neurotropic epidemic influenza A virus in the newborn/suckling period: implications for influenza-associated encephalopathy, Molecular and Cellular Biochemistry, Vol.299, No.1-2, 85-92, 2006.
(要約)
The neuropathogenesis of influenza-associated encephalopathy in children and Reye's syndrome remains unclear. A surveillance effort conducted during 2000-2003 in South-West Japan reveals that almost all fatal and handicapped influenza-associated encephalopathy patients exhibit a disorder of mitochondrial beta-oxidation with elevated serum acylcarnitine ratios (C(16:0)+C(18:1))/C(2). Here we show invasion by a non-neurotropic epidemic influenza A H3N2 virus in cerebral capillaries with progressive brain edema after intranasal infection of mice having impaired mitochondrial beta-oxidation congenitally or posteriorly in the newborn/ suckling periods. Mice genetically lacking of carnitine transporter OCTN2, resulting in carnitine deficiency and impaired beta-oxidation, exhibited significant higher virus-genome numbers in the brain, accumulation of virus antigen exclusively in the cerebral capillaries and increased brain vascular permeability compared to in wild type mice. Mini-plasmin, which proteolytically potentiates influenza virus multiplication in vivo and destroys the blood-brain barrier, accumulated with virus antigen in the brain capillaries of OCTN2-deficient mice but only a little in wild-type mice. These results suggest that the impaired mitochondrial beta-oxidation changes the susceptibility to a non-neurotropic influenza A virus as to multiplication in the brain capillaries and to cause brain edema. These pathological findings in the brain of mice having impaired mitochondrial beta-oxidation after influenza virus infection may have implications for human influenza-associated encephalopathy.
Mihiro Yano, Nakamuta S., Wu X., Yuushi Okumura and Hiroshi Kido : A novel function of 14-3-3 protein: 14-3-3z is a heat shock-related molecular chaperone that dissolves thermal-aggregated proteins, Molecular Biology of the Cell, Vol.17, No.11, 4769-4779, 2006.
(要約)
The 14-3-3 proteins are highly conserved molecules that function as intracellular adaptors in a variety of biological processes, such as signal transduction, cell cycle control, and apoptosis. Here, we show that a 14-3-3 protein is a heat-shock protein (Hsp) that protects cells against physiological stress as its new cellular function. We have observed that, in Drosophila cells, the 14-3-3zeta is up-regulated under heat stress conditions, a process mediated by a heat shock transcription factor. As the biological action linked to heat stress, 14-3-3zeta interacted with apocytochrome c, a mitochondrial precursor protein of cytochrome c, in heat-treated cells, and the suppression of 14-3-3zeta expression by RNA interference resulted in the formation of significant amounts of aggregated apocytochrome c in the cytosol. The aggregated apocytochrome c was converted to a soluble form by the addition of 14-3-3zeta protein and ATP in vitro. 14-3-3zeta also resolubilized heat-aggregated citrate synthase and facilitated its reactivation in cooperation with Hsp70/Hsp40 in vitro. Our observations provide the first direct evidence that a 14-3-3 protein functions as a stress-induced molecular chaperone that dissolves and renaturalizes thermal-aggregated proteins.
木戸 博, Le Quang Trong, Yao Dengbing, 山田 博司, Wang Sie, 奥村 裕司 : インフルエンザウイルス感染の病態とプロテアーゼによる重症化機構, アレルギー·免疫, Vol.13, No.11, 1536-1544, 2006年.
84.
Hiroshi Yamada, Q.T. Le, A. Kousaka, Y. Higashi, M. Ttukane and Hiroshi Kido : Sendai virus infection up-regulates trypsin I and matrix metalloproteinase-9, triggering viral multiplication and matrix degrdation in rat lungs and lung L2 cells, Archives of Virology, Vol.151, No.12, 2529-2537, 2006.
(要約)
To elucidate the virus-host cell interaction, we analyzed quantitatively the expression of various cellular proteases and tumor necrosis factor-alpha (TNF-alpha) after Sendai virus infection in rat lungs and lung L2 cells. After infection, TNF-alpha mRNA levels increased rapidly to a peak on day one, and then trypsin I and matrix metalloproteinase (MMP)-9, but not MMP-2, were significantly up-regulated with a peak on day 2 in vivo. These up-regulations were confirmed in L2 cells. Up-regulation of proMMP-9 and its active convertase trypsin I seems to synergistically enhance virus multiplication and the destruction of lung matrix, resulting in the progression of pneumonia.
K. Kunimi, M. Maegawa, M. Kamada, S. Yamamoto, T. Yasui, T. Matsuzaki, A. Kuwahara, H. Furumoto, Y. Ohmoto, Hiroshi Kido and M. Irahara : Myeloid-related protein-8/14 is associated with proinflammatory cytokines in cervical mucus, Journal of Reproductive Immunology, Vol.71, No.1, 3-11, 2006.
(要約)
Myeloid-related protein-8 (MRP-8), MRP-14, and MRP-8/14 are found in a variety of inflammatory conditions and are involved in the host defense system. The objective of this study was to determine the concentrations of MRP-8, MRP-14, and MRP-8/14 in human cervical mucus and the associations between MRP-8/14 and proinflammatory cytokines. Samples of cervical mucus were obtained using a syringe from sexually active women (n=97) during the preovulatory phase. Samples from seven women were obtained using a swab placed in the cervical canal during the proliferative, preovulatory, and luteal phases. Concentrations of MRP-8, MRP-14, MRP-8/14, IL-1alpha, IL-6, IL-8, and granulocyte elastase were measured using an ELISA. The mean levels of MRP-8, MRP-14, and MRP-8/14 in cervical mucus were 1.87, 0.46, and 23.90microg/ml, respectively. The concentration of MRP-8/14 showed positive correlations with concentrations of IL-1alpha (p<0.0001), IL-8 (p<0.0001), and granulocyte elastase (p<0.0001). However, there were no significant differences in MRP-8/14 levels in the cervical mucus of each patient during the menstrual cycle. MRP-8/14 was mainly detected in human cervical mucus and showed a positive correlation with proinflammatory cytokines. The MRP-8/14 level in cervical mucus may be useful as a marker of inflammation of the uterine cervix.
木戸 博 : インフルエンザにおけるマクロライドの役割, Japan Medicine, No.1005, 18, 2006年.
87.
Trong Q. Le, M. Kawachi, Hiroshi Yamada, M. Shiota, Yuushi Okumura and Hiroshi Kido : Identification of Trypsin I as a candidate for influenza A virus and Sendai virus envelope glycoprotein processing protease in rat brain,, Biological Chemistry, Vol.387, No.4, 467-475, 2006.
(要約)
Extracellular cleavage of virus envelope fusion glycoprotein hemagglutinin (HA0) by host trypsin-like proteases is a prerequisite for the infectivity and pathogenicity of human influenza A viruses and Sendai virus. The common epidemic influenza A viruses are pneumotropic, but occasionally cause encephalopathy or encephalitis, although the HA0 processing enzyme in the brain has not been identified. In searching for the brain processing proteases, we identified a processing enzyme in rat brain that was inducible by infection with these viruses. The purified enzyme exhibited an apparent molecular mass of approximately 22 kDa on SDS-PAGE and the N-terminal amino acid sequence was consistent with that of rat pancreatic trypsin I. Its substrate specificities and inhibition profiles were the same as those of pancreatic trypsin I. In situ hybridization and immunohistochemical studies on trypsin I distribution revealed heavy deposits in the brain capillaries, particularly in the allocortex, as well as in clustered neuronal cells of the hippocampus. The purified enzyme efficiently processed the HA0 of human influenza A virus and the fusion glycoprotein precursor of Sendai virus. Our results suggest that trypsin I in the brain potentiates virus multiplication in the pathogenesis and progression of influenza-associated encephalopathy or encephalitis.
Y. Shiga, H. Wakabayashi, K. Miyazawa, Hiroshi Kido and Y. Itoyama : 14-3-3 Protein levels and isoform patterns in the cerebrospinal fluid of Creutzfeldt-Jakob diseasepatients in the progressive and terminal stages, Journal of Clinical Neuroscience, Vol.13, No.6, 661-665, 2006.
(要約)
To elucidate the diagnostic value and to establish the 14-3-3 isoform patterns in the cerebrospinal fluid (CSF) of Creutzfeldt-Jakob disease (CJD) patients, we analysed the 14-3-3 isoform patterns in the CSF of 11 CJD patients using the Western immunoassay technique. 14-3-3 protein was detected in the CSF of seven CJD patients in the progressive stage, but not in four patients in the terminal stages whose brains were severely atrophied. The amount of 14-3-3 protein measured semi-quantitatively in the CSF was correlated with that of neuron-specific enolase measured using an enzyme-linked immunosorbent assay in the same CSF. CJD patients showed five dominant 14-3-3 isoforms, gamma, epsilon, zeta, eta and beta, but 14-3-3 tau, which mainly originates from T lymphocytes, was not detected. 14-3-3 protein is released into the CSF as a consequence of the extensive and rapid destruction of the brain, and the presence of the five isoforms enhances the diagnostic value of 14-3-3 protein in the progressive stage.
(キーワード)
14-3-3 Proteins / Aged / Blotting, Western / Creutzfeldt-Jakob Syndrome / Disease Progression / Female / Humans / Male / Middle Aged / Phosphopyruvate Hydratase / Protein Isoforms
Dai Mizuno, Itsuka Kubo, Mikiko Ide-Kurihara, Tomoko Ichinomiya and Hiroshi Kido : Modified pulmonary surfactant is a potent adjuvant that stimulates the mucosal IgA production in response to the influenza virus antigen, The Journal of Immunology, Vol.176, No.2, 1122-1130, 2006.
(要約)
The intranasal administration of influenza hemagglutinin (HA) vaccine with Surfacten, a modified pulmonary surfactant free of antigenic c-type lectins, as a mucosal adjuvant induced the highest protective mucosal immunity in the airway. The intranasal immunization of mice with HA vaccine (0.2 microg)-Surfacten (0.2 microg) selectively induced the neutralizing anti-HA IgA, but not IgG, and conferred nearly maximal protection in the airway, without inducing a systemic response. In contrast, intranasal inoculation of vaccine with 0.2 microg of the potent mucosal adjuvant cholera toxin B* (CT-B*), prepared by adding 0.2% native CT to the B subunit of CT, induced both anti-HA IgA and IgG in the airway and in the serum. The intranasal administration of HA vaccine alone induced a limited amount of mucosal IgA against influenza virus. Although the s.c. administration of HA vaccine prominently induced serum IgG and IgA, Surfacten and CT-B* did not enhance their induction, and the concentrations of Abs leaking into the airways were insufficient to prevent viral multiplication. The intranasal administration of HA-Surfacten stimulated the expression of MHC class II, CD40, and CD86 molecules in the CD11c-positive cells isolated from the nasal mucosa, but not the expression of cells from the lungs or spleens. Lymphocytes isolated from the airway mucosa after intranasal HA-Surfacten immunization prominently induced TGF-beta1 which, compared with inoculation without Surfacten, promoted an Ag-specific mucosal IgA response. Surfacten alone, however, did not induce TGF-beta1. Our observations suggest that Surfacten, by mimicking the natural surfactant, is an effective mucosal adjuvant in the process of airway immunization.
A. Kurisaki, T. S. Hamazaki, K. Okabayashi, T. Iida, T. Nishine, R. Chonan, Hiroshi Kido, S. Tsunasawa, O. Nishimura, M. Asashima and H. Sugino : Chromatin-related proteins in pluripotent mouse embryonic stem cells are downregulated after removal of leukemia inhibitory factor, Biochemical and Biophysical Research Communications, Vol.335, No.3, 667-675, 2005.
(要約)
Embryonic stem (ES) cells have generated enormous interest due to their capacity to self-renew and the potential for growing many different cell types in vitro. Leukemia inhibitory factor (LIF), bone morphogenetic proteins, octamer-binding protein 3 or 4, and Nanog are important factors in the maintenance of pluripotency in mouse ES cells. However, the mechanisms by which these factors regulate the pluripotency remain poorly understood. To identify other proteins involved in this process, we did a proteomic analysis of mouse ES cells that were cultured in the presence or absence of LIF. More than 100 proteins were found to be involved specifically in either the differentiation process or the maintenance of undifferentiated state. Among these, chromatin-related proteins were identified as the major proteins in nuclear extracts of undifferentiated cells. Analysis with real-time RT-PCR revealed that enrichment of these proteins in pluripotent ES cells was regulated at the transcriptional levels. These results suggest that specific chromatin-related proteins may be involved in maintaining the unique properties of pluripotent ES cells.
Mihiro Yano, Y. Kanesaki, Y. Koumoto, M. Inoue and Hiroshi Kido : Chaperone Activities of the 26S and 20S Proteasome, Current Protein & Peptide Science, Vol.6, No.2, 197-203, 2005.
(要約)
The accumulation of misfolded or damaged proteins causes the failure of normal cell structure and functions necessary for growth and viability. To abort this adverse development, defective proteins must be rapidly repaired by molecular chaperones or destroyed by energy-dependent cytoplasmic proteases. A balance among these processes ultimately maintains cellular homeostasis. In eukaryotes, the 26S proteasome, a protease/chaperone complex, is a central component in the protein triage decision process. The 26S proteasome generally acts as a ubiquitination system, though it also selectively degrades structurally abnormal proteins in an ubiquitin-independent manner. In either case, all substrate proteins must undergo structural changes and stabilization necessary for their rapid degradation. It has, therefore, often been suggested that several chaperone functions are closely related to the stimulation of proteasomal degradation. This review summarizes recent discoveries pertaining to chaperone activities in the proteasomal degradation pathway, and to their regulation of protein breakdown mediated by the proteasome.
Y. Chen, H. Mizuguchi, D. Yao, M. Ide, Y. Kuroda, Y. Shigematsu and Hiroshi Kido : Thermolabile phenotype of carnitine palmitoyltransferase II variations as a predisposing factor for influenza-associated encephalopathy, FEBS Letters, Vol.579, No.10, 2040-2044, 2005.
(要約)
To assess the etiology of influenza-associated encephalopathy (IAE), a surveillance effort was conducted during 2000-2003 in South-West Japan. All fatal and handicapped patients except one (4/34 patients) exhibited a disorder of mitochondrial beta-oxidation evoked by the inactivated carnitine palmitoyltransferase II (CPT II) with transiently elevated serum acylcarnitine ratios (C(16:0) + C(18:1))/C(2) > 0.09 during high-grade fever. Analyses of genotypes and allele compositions of CPT II revealed a thermolabile phenotype of compound heterozygotes for [1055T > G/F352C] and [1102G > A/V368I], which shows a higher frequency in IAE patients than healthy volunteers (P < 0.025). The thermolabile phenotype of CPT II variations may be a principal genetic background of IAE in Japanese.
(キーワード)
Adolescent / Base Sequence / Carnitine O-Palmitoyltransferase / Child / Child, Preschool / DNA Primers / Encephalitis, Viral / Enzyme Stability / Female / Humans / Infant / Infant, Newborn / Influenza, Human / Male / Pedigree / Phenotype
M. Kita, Yuushi Okumura, S. D. Ohdachi, Y. Oba, M. Yoshikuni, Y. Nakamura, Hiroshi Kido and D. Uemura : Purification and characterization of brarinasin, a new tissue kallikurein-like protease from the short-tailed shrew Blarina brevicauda-comparative stadies with blarina toxin, Biological Chemistry, Vol.386, No.2, 177-182, 2005.
(要約)
A new tissue kallikrein-like protease, blarinasin, has been purified from the salivary glands of the short-tailed shrew Blarina brevicauda. Blarinasin is a 32-kDa N-glycosylated protease with isoelectric values ranging between 5.3 and 5.7, and an optimum pH of 8.5 for enzyme activity. The cloned blarinasin cDNA coded for a pre-pro-sequence and a mature peptide of 252 amino acids with a catalytic triad typical for serine proteases and 43.7-54.0% identity to other mammalian tissue kallikreins. Blarinasin preferentially hydrolysed Pro-Phe-Arg-4-methylcoumaryl-7-amide (MCA) and N-tert-butyloxycarbonyl-Val-Leu-Lys-MCA, and preferentially converted human high-molecular-weight kininogen (HK) to bradykinin. The activity of blarinasin was prominently inhibited by aprotinin (K(i) =3.4 nM). A similar kallikrein-like protease, the lethal venom blarina toxin, has previously been purified from the salivary glands of the shrew Blarina and shows 67.9% identity to blarinasin. However, blarinasin was not toxic in mice. Blarinasin is a very abundant kallikrein-like protease and represents 70-75% of kallikrein-like enzymes in the salivary gland of B. brevicauda.
Degfu Yao, Ye Chen, Masamichi Kuwajima, Mayumi Shiota and Hiroshi Kido : Accumulation of mini-plasmin in the cerebral capillaries causes vascular invasion of the murine brain by a pneumotropic influenza A virus: implications for influenza-associated encephalopathy, Biological Chemistry, Vol.385, No.6, 487-492, 2004.
(要約)
The infectivity and pathogenicity of influenza virus are primarily determined by host cellular trypsin-type processing proteases which cleave the viral membrane fusion glycoprotein hemagglutinin (HA). Therefore the distribution of the processing protease is a major determinant of the infectious organ tropism. The common epidemic human influenza A virus is pneumotropic and the HA processing proteases tryptase Clara, mini-plasmin, tryptase TC30 and ectopic anionic trypsin have all been isolated from mammalian airways. However, the pneumotropic influenza virus occasionally causes severe brain edema, particularly in children presenting with Reye's syndrome treated with aspirin, or in children with influenza-associated encephalopathy without antipyretic treatment. We have observed that, after influenza virus infection, the accumulation of mini-plasmin in the cerebral capillaries in mice with a congenital or acquired abnormality of mitochondrial beta-oxidation mimicking the pathological findings of Reye's syndrome, causes an invasion and multiplication of the pneumotropic influenza virus at these same locations. From these findings, we hypothesize that the accumulated mini-plasmin modifies the brain capillaries from a non-permissive to a permissive state, thereby allowing multiplication of pneumotropic influenza virus. In addition, mini-plasmin proteolytically destroys the blood-brain barrier. These pathologic findings, consistent with encephalopathy in mice with a systemic impairment of beta-oxidation, may have implications for human influenza encephalopathy.
Hiroshi Yamada, R. Chounan, Y. Higashi, N. Kurihara and Hiroshi Kido : Mitochondrial targeting sequence of the influenza A virus PB1-F2 protein and its function in mitochondria, FEBS Letters, Vol.578, No.3, 331-336, 2004.
(要約)
The influenza A virus PB1-F2 protein predominantly localizes in the mitochondria of virus-infected cells. A series of cDNAs encoding N- and C-terminal deletion mutants and site-directed mutagenesis of the basic residues of PB1-F2 appended to 3xFLAG revealed the domain from residues 46 to 75 to be both necessary and sufficient for mitochondrial targeting. In addition, the subdomain residues 63-75 and both Lys73 and Arg75 are minimally required for mitochondrial localization. Transfection of untagged- and 3xFLAG tagged-PB1-F2 into Vero, HeLa and MDCK cells changed the mitochondrial morphology from a filamentous to a dotted structure and suppressed the inner-membrane potential.
Hiroshi Kido, Yuushi Okumura, Hiroshi Yamada, Dai Mizuno, Youichirou Higashi and Mihiro Yano : Secretory leukoprotease inhibitor and pulmonary surfactant serve as principal defenses against influenza A virus infection in the airway and chemical agents up-regulating their levels may have therapeutic potential, Biological Chemistry, Vol.385, No.11, 1029-1034, 2004.
(要約)
Influenza A virus (IAV) is one of the most common infectious pathogens in humans. Entry of this virus into cells is primarily determined by host cellular trypsin-type processing proteases, which proteolytically activate viral membrane fusion glycoprotein precursors. Human IAV and murine parainfluenza virus type 1 Sendai virus are exclusively pneumotropic, and the infectious organ tropism of these viruses is determined by the susceptibility of the viral envelope glycoprotein to cleavage by proteases in the airway. Proteases in the upper respiratory tract are suppressed by secretory leukoprotease inhibitor, and those in the lower respiratory tract are suppressed by pulmonary surfactant, which by adsorption inhibits the interaction between the proteases and viral membrane proteins. Although the protease activities are predominant over the activities of inhibitory compounds under normal airway conditions, intranasal administration of inhibitors was able to significantly suppress multi-cycles of viral replication in the airway. In addition, we identified chemical agents that could act as defensive factors by up-regulating the levels of the natural inhibitors and immunoglobulin A (IgA) in airway fluids. One of these compounds, ambroxol, is a mucolytic and anti-oxidant agent that stimulates the release of secretory leukoprotease inhibitor and pulmonary surfactant in the early phase, and IgA in the late phase of infection at an optimal dose, i.e. a dose sufficient to inhibit virus proliferation and increase the survival rate of animals after treatment with a lethal dose of IAV. Another agent, clarithromycin, is a macrolide antibiotic that increases IgA levels through augmentation of interleukin-12 levels and mucosal immunization in the airway. In addition to the sialidase inhibitors, which prevent the release of IAV from infected cells, inhibitors of the processing proteases and chemical agents that augment mucosal immunity and/or levels of the relevant defensive compounds may also ultimately prove to be useful as new anti-influenza agents.
M. Kita, Y. Nakamura, Yuushi Okumura, S. D. Ohdachi, Y. Oba, M. Yoshikuni, Hiroshi Kido and D. Uemura : Blarina toxin, a mammalian lethal venom from the short-tailed shrew Blarina brevicauda: Isolation and characterization, Proceedings of the National Academy of Sciences of the United States of America, Vol.101, No.20, 7542-7547, 2004.
(要約)
Venomous mammals are rare, and their venoms have not been characterized. We have purified and characterized the blarina toxin (BLTX), a lethal mammalian venom with a tissue kallikrein-like activity from the submaxillary and sublingual glands of the short-tailed shrew Blarina brevicauda. Mice administered BLTX i.p. developed irregular respiration, paralysis, and convulsions before dying. Based on the amino acid sequence of purified protein, we cloned the BLTX cDNA. It consists of a prosequence and an active form of 253 aa with a typical catalytic triad of serine proteases, with a high identity with tissue kallikreins. BLTX is an N-linked microheterogeneous glycoprotein with a unique insertion of 10 residues, L(106)TFFYKTFLG(115). BLTX converted kininogens to kinins, which may be one of the toxic pathogens, and had dilatory effects on the blood vessel walls. The acute toxicity and proteolytic activity of BLTX were strongly inhibited by aprotinin, a kallikrein inhibitor, suggesting that its toxicity is due to a kallikrein-like activity of the venom.
木戸 博 : クラリスロマイシンに気道の粘膜免疫増強作用, Medical Tribune, Vol.37, No.42, 33, 2004年.
106.
Mihiro Yano, Y. Koumoto, Y. Kanesaki, Xueji Wu and Hiroshi Kido : 20S Proteasome prevents aggregation of heat-denatured proteins without PA700 regulatory subcomplex like a molecular chaperone, Biomacromolecules, Vol.5, No.4, 1465-1469, 2004.
M. Sakai, H. Miyake, S. Tashiro, Yuushi Okumura and Hiroshi Kido : Inhibitory effect of FK506 and Cyclosporine A on the growth and invation of human liver cancer cells, The Journal of Medical Investigation : JMI, Vol.51, No.1-2, 63-69, 2004.
(要約)
The prognosis of liver transplantation for liver cancer is determined by recurrence in the liver graft. In this study, the effects of immunosuppressors, FK506 and cyclosporine A (CsA) on the migration of liver cancer cells were investigated. The effects of FK506 at concentrations of 1-100 ng/mL and CsA at 1-1000 ng/mL on the growth of poorly and well differentiated human hepatocellular carcinoma cell lines, HLE and HuH-7, respectively, were examined. After treatment of these cells with FK506 and CsA, the growth of these cells, their cytotoxicities and invasion assay on the Matrigel basement membrane invasion chamber were evaluated. In addition, the effects of FK506 and CsA on the changes in the production of a soluble intercellular adhesion molecule-1 (sICAM-1) of these cells were measured. FK506 and CsA at concentrations of 1-10 ng/mL inhibited the growth of both HLE and HuH-7 and those immunosuppressors at concentrations over 100 ng/mL exhibited cytotoxicity on these cells. FK506 at concentration of 1 ng/mL significantly inhibited the invasion of poorly differentiated HLE, but not well differentiated HuH-7, after treatment for 2-5 days in culture (p<0.05), but FK 506 at 10 ng/mL showed less inhibitory efficient. CsA at concentrations of 1-10 ng/mL, however, did not inhibit or transiently inhibited the invasion of both cell lines. The production of ICAM-1 in HLE and HuH-7 was suppressed by FK506 at concentrations of 1-10 ng/mL after treatment for 3-5 days but the effect was not significant in the initial phase at days 1-2 and the last phase at days 5-6. FK506, but not CsA, at a clinical dose of 1 ng/mL significantly inhibited the invasion of the poorly-differentiated HLE, but not HuH-7 and also inhibited the production of sICAM-1 in HLE.
Xueji Wu, Mihiro Yano, H. Washida and Hiroshi Kido : The second metal-binding site of 70kDa heat-shock protein is essential for ADP-binding, ATP hydrolysis and ATP synthesis, The Biochemical Journal, Vol.378, No.Pt 3, 793-799, 2004.
(要約)
The chaperone activity of Hsp70 (70 kDa heat-shock protein) in protein folding and its conformational switch, including oligomeric and monomeric interconversion, are regulated by the hydrolysis of ATP and the ATP-ADP exchange cycle. The crystal structure of human ATPase domain shows two metal-binding sites, the first for ATP binding and a second, in close proximity to the first, whose function remains unknown [Sriram, Osipiuk, Freeman, Morimoto and Joachimiak (1997) Structure 5, 403-414]. In this study, we have characterized the second metal-binding motif by site-directed mutagenesis and the kinetics of ATP and ADP binding, and found that the second metal-binding site, comprising a loop co-ordinated by His-227, Glu-231 and Asp-232, participates both in ATP hydrolysis and ATP-synthetic activities, in co-operation with the first metal-binding site. The first metal-binding site, a catalytic centre, is essential for ATP binding and the second site for ADP binding in the reactions of ATP hydrolysis and ATP synthesis.
Ping Cui, Saimoon Sharmin, Yuushi Okumura, Hiroshi Yamada, Mihiro Yano, Dai Mizuno and Hiroshi Kido : Endothelin-1 peptides and IL-5 synergistically increase the expression of IL-13 in eosinophils., Biochemical and Biophysical Research Communications, Vol.315, No.4, 782-787, 2004.
(要約)
The 21-amino acid-length endothelin-1 (ET-1)(1-21) and its novel derivative, 31 amino acid-length ET-1(1-31), have proinflammatory properties and induce significant eosinophil migration mediated by an increase in the local levels of eotaxin and IL-5. We have analyzed by reverse transcription polymerase chain reaction and enzyme immunoassay the effects of ETs on the expression of IL-13 mRNA and protein in eosinophils with or without cell priming with IL-5. The expression of the ETA receptor (ETAR) and its membrane localization were detected in the eosinophils, whereas the ETB receptor was undetectable. ET peptides synergistically increased the expression of IL-13 in eosinophils after priming with IL-5, and the increase was blocked by the ETAR antagonist BQ123, though these peptides did not directly influence the expression. These results may explain the presence of eosinophilia in the airways' epithelium of patients suffering from asthma, along with an increase in immunoreactive ETs.
Natsuo Oka, Yuushi Okumura, Hiro-omi Kanayama, Hirofumi Izaki, Masumi Okamoto, Hiroshi Kido and Susumu Kagawa : Amiloride and Urinary Trypsin Inhibitor Inhibit Urothelial Cancer Invasion, European Urology, Vol.44, No.6, 737-741, 2003.
(要約)
To determine the contribution of urokinase-type plasminogen activator (uPA) and plasmin in the invasion of highly invasive urothelial cancer cells. We compared expression levels of mRNA and protease activity of uPA and plasmin formation in primary cultures of the noninvasive transitional cell carcinoma, UCT-1, and in the highly invasive type, UCT-2. By using in vitro cell invasion assay system, we evaluated the effects of amiloride and urinary trypsin inhibitor (UTI), which inhibit uPA and plasmin, respectively, on invasion by both cell lines. Expression levels of mRNA, protein, and activities of uPA were significantly higher (p<0.005) and resulted in more plasminogen activation in UCT-2 than in UCT-1. Amiloride and UTI significantly inhibited plasmin formation and the invasion of both cell lines (p<0.001). High expression levels of mRNA, activities of uPA and high plasmin formation significantly potentiated the invasiveness of urothelial cancer cells. Thus, inhibitors of uPA and plasmin, such as amiloride and UTI, respectively, could be useful therapeutic tools with which to treat urothelial cancer.
Eiko Murata, Saimoon Sharmin, Hiroshi Shiota, Mayumi Shiota, Mihiro Yano and Hiroshi Kido : The effect of topically applied secretory leukocyte protease inhibitor on the eosinophil response in the late phase of allergic conjunctivitis, Current Eye Research, Vol.26, No.5, 271-276, 2003.
(要約)
This study examined the effects of secretory leukocyte protease inhibitor (SLPI), a protease inhibitor in tears, in allergic conjunctivitis. Conjunctiva of male Hartley guinea pigs sensitized with ovalbumin were treated with SLPI or the vehicle 10 min before antigen challenge or simultaneously. The animals were sacrificed after antigen challenges of 0-24 h duration, and the inhibition of eosinophil conjunctival migration and degranulation by SLPI was analyzed histochemically. The effects of SLPI on mast cell chymase and tryptase were also examined. Treatment of sensitized guinea pigs with SLPI suppressed the conjunctival recruitment and degranulation of eosinophils after antigen challenge for 6 h, inhibiting the development of allergic conjunctivitis. The effects of SLPI were observed at concentrations > or =0.1 microM, with a peak at 5 microM. SLPI inhibited chymase in a dose-dependent manner, but had no effect on tryptase. The topical SLPI application may be therapeutic in allergic conjunctivitis.
Yuushi Okumura, M. Nishikawa, Ping Cui, M. Shiota, Y. Nakamura, M. Adachi, K. Kitamura, K. Tomita and Hiroshi Kido : Cloning and characterization of a novel transmembrane-type serine Protease from rat kidney, a new member among the sodium channel activators, Biological Chemistry, Vol.384, No.10-11, 1483-1495, 2003.
(要約)
We have cloned the gene of a new transmembrane-type serine protease from rat kidney, which activates sodium channels. The amino acid sequence deduced from a full-length cDNA revealed that transmembrane serine protease-1 (TMSP-1) is a member of the clan SA/family S1 of serine proteases, comprising a 30 amino acid prepropeptide, a mature form sequence of 274 amino acids starting with the Ile-Val-Gly-Gly-Gln motif, and a common catalytic triad of serine proteases. The hydrophobic amino acid sequence in the carboxy-terminus of this enzyme suggests that it is a glycosylphosphatidylinositol-anchored protein. As revealed by quantitative reverse transcription-polymerase chain reaction analysis, it is highly expressed in kidney, small intestine, and stomach, and moderately expressed in lung, thymus, spleen and skin. The recombinant protease had an optimal pH at 9.0, selectively cleaved synthetic peptide substrates of trypsin, and was inhibited by aprotinin, leupeptin and benzamidine. Immunohistochemical studies revealed that this protease is predominantly expressed in cells from collecting ducts of the renal medulla. We also demonstrate that a C-terminally truncated variant of TMSP-1 significantly activates the epithelial sodium channel, and that its mRNA levels are upregulated by aldosterone. These observations suggest that it is a new member of the trypsin-type transmembrane proteases, which regulate sodium balance by activating the epithelial sodium channel.
K. Hara, M. Shiota, Hiroshi Kido, K. Watanabe, K. Nagata and T. Toyoda : Inhibition of the protease activity of influenza virus RNA polymerase PA subunit by viral matrix protein, Microbiology and Immunology, Vol.47, No.7, 521-526, 2003.
(要約)
Influenza virus PA is a subunit of RNA-dependent RNA polymerase. We demonstrated that PA has a unique chymotrypsin-like serine protease activity with Ser624 as an active site. To obtain further insight into the role of the protease activity of PA in viral proliferation, we examined the interaction between PA and matrix protein (M1). Both M1 purified from virion and hexa-histidine-tagged M1 expressed in Escherichia coli bound to PA. Hexa-histidine-tagged M1 pulled down PA. The interaction of PA with M1 was sensitive to ionic strength, suggesting that the interaction is formed by electrostatic force. Using Suc-Leu-Leu-Val-Tyr-MCA, a specific substrate for PA protease, M1 was demonstrated to inhibit the amidolytic activity of PA, whereas M1 did not inhibit that of chymotrypsin or trypsin at all. These results suggest that M1 binds to and inhibits the amidolytic activity of PA.
Hiroshi Kido, Ping Cui, S. Sharmin, M. Shiota, Dengfu Yao, Chunxing Lin and Mihiro Yano : Human mast cell chymase and 31 amino acid endothelin-1. New inflammatory mediators in cardiovascular diseases, Cardiomyopathies and Heart Failure: Biomolecular, Infectious and Immune Mechanisms, 145-158, 2003.
118.
H. Kitamura, Ping Cui, S. Sharmin, Mihiro Yano and Hiroshi Kido : Binding of a new bioactive 31-amino-acid endothelin-1 to an endothelin ETB or ETB-like receptor in porcine lungs., European Journal of Pharmacology, Vol.465, No.1-2, 31-38, 2003.
(要約)
Endothelin-1-(1-31) is a new bioactive 31-amino-acid-length peptide generated from big endothelin-1 by chymase or other chymotrypsin-type proteases with various pathophysiologic functions. In this study, we have detected the specific and monophasic binding of [125I]endothelin-1-(1-31) in porcine lung membranes. Competition studies of [125I]endothelin-1-(1-31) binding by unlabeled endothelin-1-(1-31), endothelin-1, endothelin-3, and antagonists and agonists of endothelin ET(A) and ET(B) receptors suggest that the binding protein is an endothelin ET(B) or ET(B)-like receptor rather than an endothelin ET(A) receptor in porcine lungs. Kinetic studies showed that the affinity of endothelin-1-(1-31) to its receptor was approximately one order of magnitude lower than that of endothelin-1, and that the specific binding of endothelin-1-(1-31) was about 19% of endothelin-1 binding. The binding of [125I]endothelin-1-(1-31) was extremely slow, slower even than that of endothelin-1, and nearly irreversible. This unique quasi-irreversibility may explain the slow-onset and long-lasting biologic effects of this peptide in vivo.
Y. Nakamura, M. Inoue, Yuushi Okumura, M. Shiota, M. Nishikawa, S. Arase and Hiroshi Kido : Cloning, expression analysis, and tissue distribution of esp-1/testisin, a membrane-type serine protease from the rat., The Journal of Medical Investigation : JMI, Vol.50, No.1-2, 78-86, 2003.
Dengfu Yao, Masamichi Kuwajima and Hiroshi Kido : Pathologic mechanisms of influenza encephalitis with an abnormal expression of inflammatory cytokines and accumulation of mini-plasmin, The Journal of Medical Investigation : JMI, Vol.50, No.1-2, 1-8, 2003.
(要約)
The pathogenesis of influenza encephalopathy or encephalitis is poorly understood. This review summarizes our recent studies of the roles played by inflammatory cytokines, inducible nitric oxide synthase (iNOS), adhesion molecules and mini-plasmin in influenza encephalitis. After the intranasal infection of newborn mice with the non-neurotropic strain of influenza A virus (IAV) Aichi/2/68/H3N2, encephalitis and severe brain edema were observed within 3-5 days. IAV-RNA and abnormalities in the blood-brain barrier permeability were detected in association with an increase in the mRNA expressions of endothelin-1, iNOS, and tumor necrosis factor-alpha. Furthermore, the accumulation in the brain capillaries of mini-plasmin, which proteolytically induces the viral envelope fusion activity and allows the virus to enter the cells, changes the brain from non-susceptible to susceptible to non-neurotropic IAV multiplication. The accumulation of mini-plasmin was markedly greater in newborn mice with an impaired mitochondrial fatty acid metabolism. These inflammatory mediators and the accumulation of mini-plasmin in the brain may play an important role in the onset and progression of LAV encephalitis.
M. Sato, S. Yoshida, K. Iida, T. Tomosawa, Hiroshi Kido and M. Yamashita : A novel influenza A virus activating enzyme from porcine lung: Purification and characterization, Biological Chemistry, Vol.384, No.2, 219-227, 2003.
(要約)
Proteolytic activation of hemagglutinin, an envelope glycoprotein of the influenza virus, by host proteases is essential for infection and proliferation of the virus. However, there is no well-defined, inherent source of host proteases in man or swine, both of which are natural hosts for human influenza viruses. We have recently isolated a 32 kDa protein in a high salt extract from porcine lungs, which possess the hemagglutinin processing activity. In this study, we attempted to purify another hemagglutinin processing enzyme from porcine lung. The purified enzyme, named tryptase TC30, exhibited a molecular mass of about 30 kDa by SDS-PAGE and 28.5 kDa by gel filtration chromatography, suggesting that it is a monomer. Tryptase TC30 cleaved peptide substrates with Arg at the P1 position, and preferentially substrates with the Ser-Ile-Gin-Ser-Arg sequence corresponding to the HA cleavage site sequence of the A/PR/8/34 influenza virus. Among various inhibitors tested, trypsin-type serine protease inhibitors, such as aprotinin, antipain, benzamidine and leupeptin, efficiently inhibited the proteolytic activity of the enzyme. The N-terminal 40 amino acid sequence of tryptase TC30 exhibits more than 60% homology to mast cell tryptases from mice MCP-6 and human tryptase-alpha and -beta. These data indicate that tryptase TC30, the 30 kDa enzyme from porcine lung, is a novel hemagglutinin-cleaving enzyme.
M. Kihara, H. Kakegawa, Y. Matono, E. Murata, H. Tsuge, Hiroshi Kido and N. Kaunuma : Chondroitin sulfate proteoglycan Is a potent enhancer in the processing of procathepsin L, Biological Chemistry, Vol.383, No.12, 1925-1929, 2002.
(要約)
The acceleration effect of chondroitin-4-sulfate(CS-) proteoglycan on the processing of procathepsin L in vitro was investigated using enzyme purified from the culture medium of MLC cells. Procathepsin L was slightly processed even when it was incubated without CS-proteoglycan for 60 min in 50 mm acetate buffer, pH 5.5, and trace amounts of the 31 kDa mature form and 35-38 kDa intermediates of cathepsin L were formed. On the other hand, in the presence of CS-proteoglycan, procathepsin L was completely converted to the mature form within the same 60 minute time period. Moreover, Z-Phe-Arg-MCA hydrolyzing activity was increased significantly by the incubation with CS-proteoglycan, while no considerable increase in the activity was observed during the incubation without CS-proteoglycan. Since the specific cathepsin L inhibitor, CLIK-195, inhibited the processing of procathepsin L accelerated by CS-proteoglycan, the trace amount of cathepsin L activity may participate in the processing. These results suggest that CS-proteoglycan may play a role in accelerating the processing of procathepsin L as an endogenous enhancer in the extracellular environment in vivo.
(キーワード)
cysteine proteinase / MLC cell / procathepsin L / processing / proteoglycan / MAJOR EXCRETED PROTEIN / L IN-VITRO / CATHEPSIN-L / BONE-RESORPTION / IDENTIFICATION / MECHANISMS / ACTIVATION / SECRETION / SEQUENCE / FORM
T. Towatari, M. Ide, K. Ohba, Y. Chiba, M. Murakami, M. Shiota, M. Kawachi, Hiroshi Yamada and Hiroshi Kido : Identification of ectopic anionic trypsin I in rat lungs potentiating pneumotropic virus infectivity and increased enzyme level after virus infection, European Journal of Biochemistry, Vol.269, No.10, 2613-2621, 2002.
(要約)
Extracellular cleavage of virus envelope fusion glycoproteins by host cellular proteases is a prerequisite for the infectivity of mammalian and nonpathogenic avian influenza viruses, and Sendai virus. In search of such target processing proteases in the airway, we recently found a new candidate trypsin-like processing protease in rat lungs, which was induced by Sendai virus infection, and identified as ectopic rat anionic trypsin I. On SDS/PAGE under reducing and nonreducing conditions, the purified enzyme gave protein bands corresponding to 29 and 22 kDa, respectively, i.e. at the same positions as rat pancreatic anionic trypsin I. It exhibited an apparent molecular mass of 31 kDa on molecular sieve chromatography and its isoelectric point was pH 4.7. The amino-acid sequences of the N-terminus and proteolytic digest peptides of the purified enzyme were consistent with those of rat pancreatic anionic trypsin I. Its substrate specificities and inhibitor sensitivities were the same as those of the pancreatic enzyme. The purified enzyme efficiently processed the fusion glycoprotein precursor of Sendai virus and hemagglutinin of human influenza A virus, and potentiated the infectivity of Sendai virus in the same dose-dependent manner as the pancreatic one. Immunohistochemical studies revealed that this protease is located in the stromal cells in peri-bronchiolar regions. These results suggest that ectopic anionic trypsin I in rat lungs induced by virus infection may trigger virus spread in rat lungs.
Sharmin S., Shiota M., Murata E., Cui P., Kitamura H., Mihiro Yano and Hiroshi Kido : A novel bioactive 31-amino acid endothelin-1 peptide stimulates eosinophil recruitment, and increases the level of eotaxin and IL-5, Inflammation Research, Vol.51, No.4, 195-200, 2002.
(要約)
Investigation of the role of a novel inflammatory mediator 31-amino acid endothelin-1 [ET-1 (1-31)], a major ET derivative in granulocytes, in eosinophil recruitment after its subcutaneous administration to mice. Various ET-1 derivatives (100 pmol), with or without ET receptor antagonists (200 pmol), were administered subcutaneously to mice, and then the eosinophil migration into and chemokine levels in the injected loci were analyzed. ET-1 (1-31) and a 21-amino acid endothelin-1 (ET-1), but not big ET-1, induced eosinophil migration into the injected loci with a peak after administration for 12 h, and increased the levels of eotaxin and interleukin-5 with peaks at 6 and 24 h, respectively. These effects of ET-1(1-31) and ET-1 were significantly inhibited by an ETA receptor antagonist, BQ-123, but not by an ETB receptor antagonist, BQ-788. Novel bioactive ET-1 (1-31) induces local eosinophil migration, and increases in eotaxin and interleukin-5 through an ETA or ETA-like receptor.
(キーワード)
Animals / Cell Movement / Chemokine CCL11 / Chemokine CCL5 / Chemokines, CC / Cytokines / Endothelin-1 / Endothelins / Eosinophils / Indicators and Reagents / Injections, Subcutaneous / Interleukin-5 / Leukocyte Count / Male / Mice / Mice, Inbred BALB C / Mice, Inbred C3H / Oligopeptides / Peptide Fragments / Peptides, Cyclic / Piperidines / Receptor, Endothelin A / Receptor, Endothelin B / Receptors, Endothelin
Bing Yang, Deng Fu Yao, M. Ohuchi, M. Ide, Mihiro Yano, Yuushi Okumura and Hiroshi Kido : Ambroxol suppresses influenza-virus proliferation in the mouse airway by incresing antiviral factor levels, The European Respiratory Journal, Vol.19, No.5, 952-958, 2002.
(要約)
The protective effect of ambroxol, a mucolytic agent which has antioxidant properties and stimulates the release of pulmonary surfactant, against influenza-virus proliferation in the airway was investigated in mice. Ambroxol or the vehicle was administered intraperitoneally twice a day for 5-7 days to mice shortly after intranasal infection with a lethal dose of influenza A/Aichi/68 (H3N2) virus, and the survival rate, virus titre and levels of factors regulating virus proliferation in the airway fluid were analysed. Ambroxol significantly suppressed virus multiplication and improved the survival rate of mice. The effect of ambroxol reached a peak at 10 mg x kg(-1) x day(-1), higher doses being less effective. Ambroxol stimulated the release of suppressors of influenza-virus multiplication, such as pulmonary surfactant, mucus protease inhibitor, immunoglobulin (Ig)-A and IgG, although it stimulated the release of a trypsin-type protease that potentiates virus proliferation. In addition, ambroxol transiently suppressed release of the cytokines, tumour necrosis factor-alpha, interferon-gamma and interleukin-12, into airway fluid. Although ambroxol had several negative effects on the host defence system, overall it strikingly increased the concentrations of suppressors of influenza-virus multiplication in the airway.
Masahiko Maegawa, Masaharu Kamada, Satoshi Yamamoto, Shuji Yamano, Minoru Irahara, Hiroshi Kido and Toshihiro Aono : Involvement of carbohydrate molecules on zona pellucida in human fertilization, Journal of Reproductive Immunology, Vol.53, No.1-2, 79-89, 2002.
(要約)
Although the involvement of several receptors and ligand molecules in sperm-zona interaction in many species have been proposed, there has been a little analysis of the kinetics between these molecules during the interaction. In the present study, we applied the detection method using surface plasmon resonance (SPR) by a BIAcore apparatus for the analysis of the putative receptor-ligand interaction of sperm-egg binding. Mannose-BSA or [man](5)-[GlcNAc](2)-Asp was immobilized on the surface of a sensor chip. When concanavalin A (Con A) was delivered to each of two different sensor chips to evaluate their usefulness, the resonance signal after sample injection onto a [man](5)-[GlcNAc](2)-Asp-fixed chip decreased rapidly than the mannose-BSA-fixed chip. However, the amount of binding for Con A during the injection onto the [man](5)-[GlcNAc](2)-Asp-fixed chip was high. When acid sperm extracts (acid extracts) and fractions through a CM column, containing protease activity (protease fractions), were delivered to the mannose-BSA-fixed chip, the SPR signal during the injection was not obviously changed compared with that of the control. However, when sperm samples were delivered to the [man](5)-[GlcNAc](2)-Asp-fixed chip, the SPR response during the injection was enormous. These results suggest that the [man](5)-[GlcNAc](2)-Asp-fixed chip is more useful than the mannose-BSA-fixed chip for investigating the interactions with sperm extracts and that the sensitive method using SPR by a BIAcore apparatus is applicable for the analysis of the putative receptor-ligand interaction of sperm-egg binding.
Shuji Kondo, Shoji Kagami, Hiroshi Kido, Strutz Frank, Muller A. Gerhard and Yasuhiro Kuroda : Role of Mast Cell Tryptase in Renal Interstitial Fibrosis, Journal of the American Society of Nephrology, Vol.12, No.8, 1668-1676, 2001.
(要約)
Renal interstitial fibrosis is characterized by increased proliferation of fibroblasts and excessive accumulation of extracellular matrix. Mast cell tryptase has been implicated in the development of tissue fibrosis in skin and lungs. However, the significance of mast cell tryptase in human renal diseases has not been investigated. The potential role of mast cell-derived tryptase in the development of renal fibrosis was studied using immunohistochemical techniques and cultured human renal fibroblast cell lines. Semiquantitative immunostaining analysis of samples from 70 patients with several renal diseases, including IgA glomerulonephritis (GN) (n = 30), non-IgA GN (n = 5), membranous GN (n = 5), focal segmental glomerulosclerosis (n = 4), minor glomerular abnormalities (n = 5), lupus nephritis (n = 3), and acute or chronic tubulointerstitial nephritis (n = 18), revealed that the degree of renal interstitial fibrosis was well correlated with the number of infiltrating tryptase-positive mast cells (P < 0.01). Mast cells could not be detected in damaged glomeruli in any form of renal disease. [(3)H]Thymidine uptake experiments demonstrated that DNA synthesis by cultured renal fibroblasts was increased with the concentration of tryptase (0.5 to 5 nM) coincubated with heparin and was suppressed by coincubation with the protease inhibitors leupeptin and benzamidine hydrochloride. Tryptase alone also increased DNA synthesis by fibroblasts but exhibited less effectiveness, compared with the combination of tryptase and heparin. Conversely, heparin alone suppressed DNA synthesis by fibroblasts. Metabolic [(35)S]methionine-labeling experiments with cultured renal fibroblasts indicated that tryptase increased the synthesis of fibronectin and collagen type I, in a dose-dependent manner. These findings suggest that mast cell tryptase plays a role in the proliferation and extracellular matrix protein production of renal interstitial fibroblasts and thus contributes to the development of renal interstitial fibrosis.
Ping Cui, Kenji Tani, Hiroko Kitamura, Yuushi Okumura, Mihiro Yano, Daisuke Inui, Toshiaki Tamaki, Saburo Sone and Hiroshi Kido : A novel bioactive 31-amino acid endothelin-1 is a potent chemotactic peptide for human neutrophils and monocytes, Journal of Leukocyte Biology, Vol.70, No.2, 306-312, 2001.
(要約)
Endothelin (ET)-1(1-31) is a novel 31-amino acid-length peptide derived from big ET-1 by chymase or other chymotrypsin-type proteases and is a major ET derivative in human neutrophils. In this study, we revealed that ET-1(1-31), but not big ET, exhibited chemotactic activities toward human neutrophils and monocytes as an inflammatory mediator, although the effects were less potent than those of formyl-methionyl-leucyl-phenylalanine or interleukin-8. However, the chemotactic effects of ET-1(1-31) were much greater than those of the 21-amino acid ET-1, ET-1(1-21). Checkerboard analyses revealed that the effects are chemotactic rather than chemokinetic. The effects of ET-1(1-31) are not mediated by interleukin-8 or monocyte chemoattractant protein-1. The chemotactic effects and an increase in intracellular-free Ca(2)(+) caused by ET-1(1-31) were significantly inhibited by BQ123, an ET(A) receptor antagonist, but not by BQ788, an ET(B) receptor antagonist, suggesting that ET-1(1-31) mediates chemotaxis through an ET(A) or ET(A)-like receptor.
K. Hara, M. Shiota, Hiroshi Kido, Y. Ohtsu and T. Toyoda : Protease activity of influenza virus RNA polymerase PA subunit, International Congress, Vol.1219, 479-485, 2001.
139.
Wakabayashi H., Mihiro Yano, Tachikawa N., Oka S., Maeda M. and Hiroshi Kido : Increased concentrations of 14-3-3ϵ,γ and ζ isoforms in cerebrospinal fluid ofAIDS patients with neuronal destruction, Clinica Chimica Acta, Vol.312, No.1-2, 97-105, 2001.
(要約)
14-3-3 proteins are major evolutionarily conserved cytosolic proteins that regulate signal transduction, apoptosis and neurotransmitter synthesis. Five homologous 14-3-3 isoforms, beta, gamma, zeta, epsilon and eta, are reported in mammalian neurones. To elucidate the diagnostic value of 14-3-3 in cerebrospinal fluid (CSF), a highly specific antibody against each isoform and studies on the isoform patterns in patients with neuronal destruction are needed. In this study, we raised isoform-specific antibodies against 14-3-3 proteins and established a semiquantitative method of identification of each isoform by Western immunoblotting. We found that three isoforms, 14-3-3 epsilon, gamma and zeta, appeared in the CSF of HIV patients with AIDS dementia complex or cytomegalovirus encephalitis, but not in AIDS patients without neurological symptoms or the non-HIV patients examined. The isoform patterns in AIDS patients were different from those reported in Creutzfeldt-Jakob disease and herpes simplex encephalitis, suggesting that the isoform patterns may facilitate the differential diagnosis. A high frequency of 14-3-3 in CSF was observed in seriously ill AIDS patients, particularly those with CD4 levels of less than 20 mm(3). These findings suggested that 14-3-3 proteins were released from destroyed neural cells and are useful real-time markers of the rate and amount of neural cell destruction in these patients.
M. Murakami, T. Towatari, M. Ohuchi, M. Shiota, M. Akao, Yuushi Okumura, Marina A.A.Parry and Hiroshi Kido : Mini-plasmin found in the epithelial cells of bronchioles triggers infection by broad-spectrum influenza A viruses and Sendai virus, European Journal of Biochemistry, Vol.268, No.10, 2847-2855, 2001.
(要約)
Extracellular cleavage of virus envelope fusion glycoproteins by host cellular proteases is a prerequisite for the infectivity of mammalian and nonpathogenic avian influenza viruses, and Sendai virus. Here we report a protease present in the airway that, like tryptase Clara, can process influenza A virus haemagglutinin and Sendai virus envelope fusion glycoprotein. This protease was extracted from the membrane fraction of rat lungs, purified and then identified as a mini-plasmin. Mini-plasmin was distributed predominantly in the epithelial cells of the upward divisions of bronchioles and potentiated the replication of broad-spectrum influenza A viruses and Sendai virus, even that of the plasmin-insensitive influenza A virus strain. In comparison with plasmin, its increased hydrophobicity, leading to its higher local concentrations on membranes, and decreased molecular mass may enable mini-plasmin to gain ready access to the cleavage sites of various haemagglutinins and fusion glycoproteins after expression of these viral proteins on the cell surface. These findings suggest that mini-plasmin in the airway may play a pivotal role in the spread of viruses and their pathogenicity.
R.D. Kim, S. Sharmin, M. Inoue and Hiroshi Kido : Cloning and Expression of Novel Mosaic Serine Proteases with and without a Transmembrane Domain from Human Lung, Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression, Vol.1518, No.1-2, 204-209, 2001.
(要約)
Two cDNAs encoding novel mosaic proteins with a serine protease domain and potential regulatory modules, consisting of a protein kinase substrate and a low-density lipoprotein receptor, were cloned from a human lung cDNA library by PCR. One with a transmembrane domain (MSPL) and the other without one (MSPS) comprise 581 and 537 amino acids, respectively. Except for the C-terminal ends, the two isoforms had an identical serine protease domain exhibiting 42, 39 and 43% identity with those of plasma kallikrein, hepsin and transmembrane protease serine 2, respectively. Both genes were predominantly expressed in human lung, placenta, pancreas and prostate.
N. Okishima, M. Yoshizumi, K. Tsuchiya, Ping Cui, H. Kitamura, T. Tamaki and Hiroshi Kido : Determination of the Levels of Novel Bioactive 31-Amino-Acid Endothelins and Endothelins in Human Lungs, Life Sciences, Vol.68, No.18, 2073-2080, 2001.
(要約)
An effective method for determination of the levels of newly discovered 31-amino acid endothelins [ETs(1-31)] as well as big ETs and 21-amino acid ETs [ETs(1-21)], in human lungs has been developed. About 85% of ETs in human lung homogenates were recovered on acid extraction 8 times. Most of the published protocols for the determination of tissue ETs involve a reverse-phase minicolumn to separate proteins from peptides, after which the levels of ETs are directly determined by enzyme immunoassay. The levels determined, however, include fairly high amounts of non-bioactive ET metabolites in tissues and the data reported are diverse. We established an effective methods for the extraction and the separation of nine different muscle constricting ETs from their metabolites on a reverse-phase C18 column. Using this protocol, the levels of ETs in human lungs were determined by means of a sandwich-enzyme immunoassay specific for each ET derivative. The levels of ET-2(1-21) were the highest among those of ETs, and the levels of ETs(1-31) were in a similar range to those of big ETs but were lower than those of ETs(1-21). This method can be utilized to assess the pathophysiological roles of ETs(1-31) in various human organs.
K. Hara, M. Shiota, Hiroshi Kido, Y. Ohtsu, T. Kashiwagi, J. Iwahashi, N. Hamada, K. Mizoue, N. Tsumura, H. Kato and T. Toyoda : Influenza Virus RNA Polymerase PA Subunit is a Novel Serine Protease with Ser 624 in the Active Site, Genes to Cells, Vol.6, No.2, 87-97, 2001.
(要約)
Influenza virus RNA polymerase is a multifunctional enzyme that catalyses both transcription and replication of the RNA genome. The function of the influenza virus RNA polymerase PA subunit in viral replication is poorly understood, although the enzyme is known to be required for cRNA --> vRNA synthesis. The protease related activity of PA has been discussed ever since protease-inducing activity was demonstrated in transfection experiments. PA protein was highly purified from insect cells infected with the recombinant baculovirus carrying PA cDNA, and a novel chymotrypsin-type serine protease activity was identified with the synthetic peptide, Suc-LLVY-MCA, in the PA protein. [3H]DFP was crosslinked with PA and a mutational analysis revealed that serine624 was as an active site for the protease activity. These results constitute the demonstration of protease activity in PA subunit of the influenza virus RNA polymerase complexes.
Naoko Okishima, Masanori Yoshizumi, Koichiro Tsuchiya, Ping Cui, Hiroko Kitamura, Toshiaki Tamaki and Hiroshi Kido : Determination of the levels of novel 31-amino acid endothelins and endothelins in human lungs., Life Sciences, Vol.68, No.18, 2073-2080, 2001.
149.
Shuji Yoshikawa, Masaharu Kamada, Masahiko Maegawa, Satoshi Yamamoto, Minoru Irahara, Shuji Yamano, Toshihiro Aono, Hiroshi Kido and Samuel S Koide : Hormonal Control of mRNA Expression of Immunoglobulin Binding Factor in Uterine Cervix, Biochemical and Biophysical Research Communications, Vol.279, No.3, 898-903, 2000.
(要約)
Uterine cervical mucus contains an immunoglobulin binding factor (IgBF). It may play a role in preventing antibody production against sperm in the female reproductive tract. To elucidate the mechanism involved in the production of activated IgBF, we determined the effects of hormones on the expression of mRNAs of IgBF and of protein disulfide isomerase (PDI), activating enzyme, in uterine cervix by quantitative RT-PCR. The uterine cervices of female rats were excised at preovulatory, ovulatory, and postovulatory phases. The human uterine cervical adenocarcinoma cells (TCO-2) were cultured for 24 h in serum-free medium containing 17beta-estradiol or progesterone. Expression of IgBF and PDI mRNAs was significantly highest during the ovulatory phase. 17beta-estradiol stimulated the expression of both mRNAs in TCO-2; whereas progesterone was ineffective. In conclusion, estrogen regulates the production of IgBF by the endocervix and PDI in vivo, thereby increasing the level of activated IgBF in the female reproductive tract during the ovulatory phase, allowing sperm to enter the uterine cavity.
Y. Chen, M. Shiota, M. Ohuchi, T. Towatari, J. Tashiro, M. Murakami, Mihiro Yano, B. Yang and Hiroshi Kido : Mast Cell Tryptase from Pig Lungs Triggers Infection by Pneumotropic Sendai and Influenza A Viruses (Purification and Characterizatiion, European Journal of Biochemistry, Vol.267, No.11, 3189-3197, 2000.
(要約)
A novel trypsin-type serine proteinase, which processes the precursors of the envelope fusion glycoproteins of pneumotropic Sendai and human influenza A viruses, was purified to homogeneity from pig lungs. On SDS/PAGE, the purified enzyme gave a protein band corresponding to about 32 kDa, and has an apparent molecular mass of 120 kDa, as determined by gel permeation chromatography. Immunohistochemical staining with antibodies against this enzyme revealed that the enzyme is located in pig lung mast cells. The N-terminal 44-amino-acid sequence of the enzyme exhibits about 80% identity with those of mast cell tryptases from other species. Of the inhibitors tested, di-isopropyl fluorophosphate, antipain, leupeptin, benzamidine and a few proteinaceous inhibitors, such as mucus protease inhibitor and aprotinin, inhibited this enzyme activity. Heparin stabilized the enzyme, but high-ionic-strength conditions did not, unlike for human mast cell tryptase. The purified enzyme efficiently processed the fusion glycoprotein precursor of Sendai virus and slowly processed hemagglutinin of human influenza A virus, and triggered the infectivity of Sendai virus in a dose-dependent manner, although human mast cell tryptase beta and rat mast cell tryptase (rat MCP-7) from lungs did not process these fusion glycoproteins at all. These results suggest that mast cell tryptase in pig lungs is the possible trigger of the pneumotropic virus infections.
Mori H., Inoue M., Mihiro Yano, Wakabayashi H. and Hiroshi Kido : 14-3-3τAssociates with a Translational Control Factor FKBP12-Rapamycin-Associated Protein in T Cells after Stimulation by Pervanadate, FEBS Letters, Vol.467, No.1, 61-64, 2000.
(要約)
Proteins of the 14-3-3 family can associate with and/or modulate the activities of a variety of proteins, such as protooncogene and oncogene products, Cdc25 phosphatases and phosphatidylinositol 3-kinase, and thus are implicated in regulation of signaling pathways and the cell cycle. We report here that treatment of Jurkat T-cells with an inhibitor of protein tyrosine phosphatase, pervanadate, induces the association of 14-3-3tau with a translational control factor, FKBP12-rapamycin-associated protein (FRAP), with significant latter's autophosphorylation. Coimmunoprecipitation of various mutants of FRAP coexpressed with 14-3-3tau in COS-7 cells revealed that 14-3-3tau binds to the C-terminal side of FRAP at unknown site(s) different from the predicted binding motifs to date.
Kenji Tani, Fumitaka Ogushi, Hiroshi Kido, Tetsuya Kawano, Yuichi Kunori, Takashi Kamimura, Ping Cui and Saburo Sone : Chymase is a potent chemoattractant for human monocytes and neutrophils, Journal of Leukocyte Biology, Vol.67, No.4, 585-589, 2000.
(要約)
Chymase is a major chymotrypsin-like serine protease expressed in the secretory granules of mast cells in many mammalian species. In this study, we revealed the chemotactic activity of chymase for human mononuclear cells and neutrophils with a 48-well microchemotaxis chamber technique. Human chymase showed the potent chemotactic activity for monocytes and neutrophils dose-dependently in a concentration range from 0.1 to 10 microg/mL, corresponding to about 4-400 microM. The activity was as potent as that of N-formyl-methionyl-leucyl-phenylalanine. Chymase also stimulated cell migration of lymphocytes and purified T cells, but checkerboard analysis revealed that the effect was chemokinetic rather than chemotactic. Inhibition of chymase activities with chymase inhibitors, such as antileukoprotease and Bowman-Birk soybean trypsin inhibitor, significantly inhibited the chemotactic activity of chymase, suggesting that the proteolytic activity of chymase participates in the chemotactic activity. Our results suggest that mast cell chymase acts as a chemoattractant, and may play a role in the accumulation of inflammatory cells in development of the chronic inflammatory responses of allergic and nonallergic diseases.
Masanori Yoshizumi, Daisuke Inui, Kazuyoshi Kirima, Koichiro Tsuchiya, Hitoshi Houchi, Mami Azuma, Hiroaki Yasuoka, Hiroshi Kido and Toshiaki Tamaki : Comparison of the effects of endothelin-1,-2 and -3(1-31) on changes in [Ca2+]i in human coronary artery smooth muscle cells., The Japanese Journal of Pharmacology, Vol.81, 298-304, 1999.
162.
Hiroaki Yasuoka, Masanori Yoshizumi, Daisuke Inui, Naoko Okishima, Hitoshi Houchi, Kazuyoshi Kirima, Shuzo Oshita, Hiroshi Kido and Toshiaki Tamaki : Effect of endothelin-1 (1-31) on intracellular free calcium in cultured human mesangial cells., Life Sciences, Vol.65, No.22, PL267--272, 1999.
163.
H. Yasuoka, M. Yoshizumi, D. Inui, N. Okishima*, H. Houchi, K. Kirima, S. Oshita, Hiroshi Kido and T. Tamaki : Effect of Endothelin (1-31) on Intracellular Free Calcium in Cultured Human Mesangial Cells, Life Sci., Vol.65, No.22, 267-272, 1999.
164.
M. Inoue, M. Isobe, T. Itoyama and Hiroshi Kido : Structural analysis of esp-1 gene (PRSS 21),, Biochemical and Biophysical Research Communications, Vol.266, No.2, 564-568, 1999.
(要約)
The esp-1 gene is originally cloned from human eosinophils and encodes a membrane-type serine protease. This gene is ubiquitously expressed in various tissues but not kidney or muscle, and highly expressed in testis. Among granulocytes, this gene is expressed in eosinophils but not neutrophils, although both are derived from myeloid progenitors. In the present study, we have cloned the esp-1 genome using a BAC library, and determined exon-intron junctions: This gene spans approximately 4.6 kb, and consists of 6 exons and 5 introns. On radiation hybrid and FISH analyses, the esp-1 gene was mapped to 16p13.3. In addition, we have cloned a new splicing variant form of esp-1 from a HeLa cell cDNA library, which contains many esp-1 clones. Both RNase protection and primer extension analyses revealed the transcription initiation site of the esp-1 gene is located at nucleotide position -106, residue G. Dual-luciferase reporter analysis revealed a GC-rich region between nucleotide positions, -106 and -189 containing one AP-1/Sp-1 binding site is responsible for the minimum promoter activity in HeLa cells.
Mihiro Yano, Mori S. and Hiroshi Kido : Intrinsic Nucleoside Diphosphate Kinase-like Activity is a Novel Function of the 20S Proteasome, The Journal of Biological Chemistry, Vol.274, No.48, 34375-34382, 1999.
(要約)
The eukaryotic 20 S proteasome is the prototype of a new family of the N-terminal nucleophil hydrolases and is composed of numerous low molecular mass subunits arranged in a stack of four rings, each containing seven different alpha- or beta-subunits. Among the beta-type subunits in the yeast proteasome, three proteolytically active ones were identified, although the functions of the other beta- and alpha-type subunits remain to be clarified. We report here that the purified 20 S proteasome exhibits intrinsic nucleoside diphosphate (NDP) kinase-like activity. The proteasome exhibited a preference for ATP and dATP as phosphate donors, and a broad specificity for NDPs, other than GDP, as phosphate acceptors, unlike conventional NDP kinase, which catalyzes the transfer of gamma-phosphate between NDPs and nucleoside triphosphates. During the transfer of gamma-phosphate, the proteasome formed acid-labile phosphohistidine as autophosphorylated intermediates, and NDP-dependent dephosphorylation of the latter then occurred. These enzymatic properties are similar to those of the molecular chaperone, Hsp70, which also exhibits intrinsic NDP kinase-like activity, instead of ATPase activity. C5 among the beta-type subunits and C8 among the alpha-type subunits were autophosphorylated during the gamma-phosphate transfer reaction and were photoaffinity labeled with 8-azido-[alpha-(32)P]ATP, suggesting that the C5 and C8 subunits of the proteasome are responsible for the NDP kinase-like activity.
H. Takahashi, T. Iwata, Y. Kitagawa, R.H. Takahashi, Y. Sato, H. Wakabayashi, M. Takashima, Hiroshi Kido, Nagashima K., K. Kenney, C.J. Gibbs JR and T. Kurata : Increased levels of ϵ and γIsoform-Specific Increase of 14-3-3 Proteins in Cerebrospinal Fluid of Patients with Creutzfeldt-Jakob Disease, Clin. & Diag. Lab. Immunol., Vol.6, 983-985, 1999.
167.
T. Iwata, K. Sakai, M. Hori, S. Uchida, T. Towatri and Hiroshi Kido : Human Parathyroid Hormone (1-34) Transiently Increases the Excretion of Lysosomal Enzymes into Urine and the Size of Renal Lysosomes, The Journal of Biochemistry, Vol.126, No.3, 485-493, 1999.
(要約)
It has been reported that the urinary excretion of N-acetyl-beta-D-glucosaminidase (NAG), a lysosomal enzyme, transiently increases in human after treatment with human parathyroid hormone (hPTH)(1-34). We report here that hPTH(1-34) caused transient changes in the size and density of rat renal lysosomes following urinary excretion of NAG and other lysosomal enzymes tested. Percoll density gradient centrifugation revealed that hPTH(1-34) slightly but significantly increased the fraction of high density lysosomes (around 1.12 g/ml) 5-10 min after the treatment with hPTH(1-34), with a concomitant decrease in the fraction of intermediate density lysosomes (1.07-1.08 g/ml). On electron micrographs, some lysosomes in proximal tubules but not in distal tubules showed a change in morphology from circular to oval, and became enlarged and electron-dense 5-10 min after the treatment with hPTH(1-34). These responses to hPTH(1-34) were also reversible and transient. NAG excreted in urine after treatment with hPTH(1-34) had the molecular mass of a mature form in lysosomes and/or endosomes and was not a prepro-and/or pro-form of the enzyme. Thus, the changes in the density and size of renal lysosomes appear to be associated with the exocytosis of lysosomal enzymes by hPTH(1-34).
M. Yoshizumi, D. Inui, H. Houchi, H. Yasuoka, N. Okishima, Hiroshi Kido and T. Tamaki : Comparison of the Effects of Endothelin-1,-2 and -3 (1-31) on Changes in [Ca2+]i in Human Coronary Artery Smooth Muscle Cells, The Japanese Journal of Pharmacology, Vol.81, No.3, 298-304, 1999.
(要約)
We have previously found that human chymase selectively cleaves big endothelins (ETs) at the Tyr31-Gly32 bond to produce 31-amino-acid endothelins, ETs (1-31). In the present study, we investigated the effects of ETs (1-31) on changes in intracellular free Ca2+ ([Ca2+]i) in cultured human coronary artery smooth muscle cells (HCASMCs) using confocal laser microscopy. ETs (1-31) increased [Ca2+]i in a concentration-dependent manner. Phosphoramidon did not inhibit the increases in [Ca2+]i caused by ETs (1-31). The [Ca2+]i increases induced by ETs (1-31) were compared to those of ETs (1-21) and big ETs. ET-1 (1-21) was about 10-times more potent than big ET-1 or ET-1 (1-31), whereas big ET-2 was 10-times less potent than ET-2 (1-21) or ET-2 (1-31). ETs (1-31) may induce [Ca2+]i increase through ET(A)-type or ET(A)-type-like receptors. The 10(-12) M ET (1-31)-induced increases in [Ca2+]i were not affected by removal of extracellular Ca2+, but were inhibited by thapsigargin. These results suggested that ET-1, -2 and -3 (1-31) showed similar potencies in increasing [Ca2+]i and mechanisms of ET (1-31)-induced increases in [Ca2+]i may be similar among the three ETs (1-31).
T. Iwata, S. Uchida, M. Hori, K. Sakai, T. Towatari and Hiroshi Kido : Human Parathyroid Hormone (1-34) Increases Urinary Excretion of Lysosomal Enzymes in Rats, Life Sciences, Vol.65, No.17, 1725-1732, 1999.
(要約)
The kidney is the major target of parathyroid hormone (PTH), and PTH influences the urinary excretion of calcium, phosphate and hydrogen ions. It was previously reported that the urinary, excretion of N-acetyl-beta-D-glucosaminidase (NAG), a lysosomal enzyme, transiently increases after human PTH (hPTH) (1-34) infusion in normal subjects and idiopathic hypoparathyroidism patients, but not in pseudohypoparathyroidism type I patients. Here we report that intravenous infusion of hPTH(1-34) to rats transiently increased the urinary excretion of various lysosomal enzymes, such as beta-glucuronidase and acid phosphatase as well as NAG. However, it did not affect the urinary excretion of tubular brush border membrane enzymes, i.e. alkaline phosphatase, leucine aminopeptidase and gamma-glutamyl transpeptidase. Human PTH(1-34) dose-dependently increased the urinary excretion of NAG in rats with a peak at 30 min, which returned to a baseline within 60 min. The increase in the urinary NAG excretion caused by hPTH(1-34) positively correlated with the increase in the urinary cAMP excretion (r = 0.844, p < 0.01), and infusion of dibutyryl cAMP at a dose of 20 mg/kg similarly increased the urinary excretion of NAG. These results suggested that the increase in the urinary excretion of lysosomal enzymes caused by hPTH(1-34) may be a functional response to hPTH(1-34) occurring in the renal tubules via PTH signaling pathway.
D. Inui, M. Yoshizumi, N. Okishima, H. Houchi, K. Tsuchiya, Hiroshi Kido and T. Tamaki : Mechanism of Endothelin-1(1-31)-induced Calcium Signaling in Human Coronary Artery Smooth Muscle Cells., Am. J. Physiol., Vol.276, 1067-1072, 1999.
171.
M. Hara, A. Matsumori, K. Ono, Hiroshi Kido, M.-W. Hwang, T. Miyamoto, A. Iwasaki, M. Okada, K. Nakatani and S. Sasayama : Mast Cell Cause Apoptosis of Cardiomyocytes and Proliferation of Other Intramyocardial Cells In Vitro, Circulation, Vol.100, No.13, 1443-1449, 1999.
172.
Hiroshi Kido, M. Murakami, K. Oba, Y. Chen and T. Towatari : Cellular Proteinases Trigger the Infectivity of the Influenza A and Sendai Viruses, Molecules and Cells, Vol.9, No.3, 235-244, 1999.
(要約)
It has been proposed that the pathogenicity of the influenza and Sendai virus is primarily determined by host cellular proteases that activate viral infectivity. We isolated trypsin-type serine proteases from rat lungs, candidates for the processing proteases of viral envelope glycoproteins, such as tryptase Clara localized in the Clara cells of the bronchial epithelium and mini-plasmin. These enzymes specifically cleave the precursor of fusion glycoprotein HA of influenza virus at Arg325, and the F0 of Sendai virus at Arg116 in the consensus cleavage motif, Gln(Glu)-X-Arg, resulting in the induction of infectivity of these viruses. Proteolytic activation of viruses by these enzymes occurs extracellularly, probably on the surface and/or in the lumen of the respiratory tract. On the other hand, we isolated two compounds from human bronchial lavage, which inhibit the activity of tryptase Clara. One was a mucus protease inhibitor and the other was a pulmonary surfactant. These compounds inhibited multiple cycles of virus replication in vitro and in vivo, but did not themselves affect the hemagglutination and the infectivity of the virus. Administration of these compounds in the airway may be useful for preventing and treating infection with influenza virus and Sendai virus.
Hiroshi Kido, Y. Chen, M. Murakami, Y. Beppu and T. Towatari : Cellular Proteinases and Viral Infection: Influenza Virus, Sendai Virus and HIV-1, Proteases of Infectious Agents, 205-217, 1999.
Hiroshi Kido, Y. Beppu, Y. Imamura, Y. Chen, M. Murakami, K. Oba and T. Towatari : Human Mucus Protease Inhibitor and its Mutants are Novel Defensive Compounds against, Biopolymers, Vol.51, 79-86, 1999.
Okishima N., Hagiwara Y., Seito T., Mihiro Yano and Hiroshi Kido : Specific Sandwich-type Enzyme Immunoassays for Smooth Muscle Constricting Novel 31-Amino Acid Endothelins, Biochemical and Biophysical Research Communications, Vol.256, No.1, 1-5, 1999.
(要約)
We established highly sensitive and specific sandwich-enzyme immunoassays (EIAs) for three newly discovered bioactive 31-amino acid endothelins [ETs(1-31)], which can detect as little as 0.16 pg/well of ET-1(1-31), 0.39 pg/well of ET-2(1-31), and 0.16 pg/well of ET-3(1-31). The EIAs showed no crossreactivity with 21-amino acid endothelins [ETs(1-21)] or big ETs at the usual assay concentrations below 1-5 ng/ml. In reversed-phase HPLC, immunoreactive ETs(1-31) in the granulocytes of normal human subjects eluted at the exact positions of authentic ETs(1-31), except for the presence of one additional unknown immunoreactive ET-1(1-31). The results also indicate that ETs(1-31) exist in the granulocytes at levels higher than or similar to those of ETs(1-21). This study is the first to establish EIAs for novel bioactive ETs(1-31). These assays can be utilized to assess the pathophysiological roles of ETs(1-31).
Yuushi Okumura, Mihiro Yano, M. Murakami, T. Towatari and Hiroshi Kido : The Extracellular Processing of HIV-1 Envelope Glycoprotein gp160 by Human Plasmin., FEBS Lett.,, Vol.442, 39-42, 1999.
181.
M. Inoue, N. Kanbe, M. Kurosawa and Hiroshi Kido : Cloning and Tissue Distribution of a Novel Serine Protease esp-1 from Human Eosinophils, Biochemical and Biophysical Research Communications, Vol.252, No.2, 307-312, 1998.
(要約)
We have cloned a novel serine protease designated as esp-1 from human eosinophils. The amino acid sequence deduced from the cDNA showed that ESP-1 comprises a signal peptide of 18 amino acids, a propeptide of 23 amino acids, an active form sequence of 273 amino acids starting from an Ile-Val-Gly-Gly-Glu motif, the catalytic triad of serine proteases that has been characterized as the essential amino acid residues for the proteolytic activity, and a hydrophobic amino acid stretch in the carboxyl terminus, suggesting this enzyme is a novel membrane-type serine protease. The tissue distributions of esp-1 expression revealed that this protease is not only expressed in human eosinophils, but also widely expressed in mononuclear cells and various tissues other than skeletal muscle and kidney and is most abundant in testis and prostate, and moderately so in lung, spleen and pancreas.
(キーワード)
Amino Acid Sequence / Base Sequence / Cell Line / Cloning, Molecular / DNA Primers / DNA, Complementary / Eosinophils / Female / Gene Expression / Humans / Male / Molecular Sequence Data / Pregnancy / RNA, Messenger / Reverse Transcriptase Polymerase Chain Reaction / Serine Endopeptidases / Tissue Distribution / Transfection
M. Yoshizumi, S. Kim, S. Kagami, A. Hamaguchi, K. Tsuchiya, H. Houchi, H. Iwao, Hiroshi Kido and T. Tamaki : Effect of Endothelin-1(1-31) on Extracellular Signal-regulated Kinase and Proliferation of Human Coronary Artery Smooth Muscle Cells, British Journal of Pharmacology, Vol.125, No.5, 1019-1027, 1998.
(要約)
1. We have previously found that human chymase cleaves big endothelins (ETs) at the Tyr31-Gly32 bond and produces 31-amino acid ETs (1-31), without any further degradation products. In this study, we investigated the effect of synthetic ET-1 (1-31) on the proliferation of cultured human coronary artery smooth muscle cells (HCASMCs). 2. ET-1 (1-31) increased [3H]-thymidine incorporation and cell numbers to a similar extent as ET-1 at 100 nM. This ET-1 (1-31)-induced [3H]-thymidine uptake was not affected by phosphoramidon, an inhibitor of ET-converting enzyme. It was, however, inhibited by BQ123, an endothelin ET(A) receptor antagonist, but not by BQ788, an endothelin ET(B) receptor antagonist. 3. By using an in-gel kinase assay, we demonstrated that ET-1 (1-31) activated extracellular signal-regulated kinase 1/2 (ERK1/2) in a concentration-dependent manner (100 pM to 1 microM) in HCASMCs. ET-1 (1-31)-induced ERK1/2 activation was inhibited by BQ123, but not by BQ788 and phosphoramidon. Inhibition of protein kinase C (PKC) and ERK kinase also caused a reduction of ET-1 (1-31)-induced ERK1/2 activation, whereas tyrosine kinase inhibition had little effect. 4. Gel-mobility shift analysis revealed that the ERK1/2 activation was followed by an increase in transcription factor activator protein-1 DNA binding activity in HCASMCs. 5. Our results strongly suggest that ET-1 (1-31) itself stimulates HCASMC proliferation probably through endothelin ET(A) or ET(A)-like receptors. The underlining mechanism of cell growth by ET-1 (1-31) may be explained in part by PKC-dependent ERK1/2 activation. Since human chymase has been proposed to play a role in atherosclerosis, ET-1 (1-31) may be one of the mediators.
F. Kishi, K. Minami, N. Okishima, M. Murakami, S. Mori, Mihiro Yano, Y. Niwa, Y. Nakaya and Hiroshi Kido : Novel 31-Amino Acid-Length Endothelins Cause Constriction of Vascular Smooth Muscle, Biochemical and Biophysical Research Communications, Vol.248, No.2, 387-390, 1998.
(要約)
We have reported that human chymase specifically cleaves big endothelins (ETs) at the Try31-Gly32 bond, and produces novel trachea-constricting 31-amino-acid-length ETs, ETs(1-31). In this study, we investigated the effect of synthetic ETs(1-31) on the contractile activity toward porcine coronary arteries and rat aortae. Although ETs(1-31) exhibited less potent vasoconstrictile activity in these tissues than 21-amino-acid-length ETs(1-21), or a similar extent, ET-1(1-31) caused significantly slower-developing and longer-lasting contraction than ETs(1-21). The ETA receptor antagonist, BQ485, completely inhibited the activity of ET-1(1-31). The ETB receptor antagonist, BQ788, also inhibited the activity of ET-1(1-31) toward rat aortae more efficiently than that ET-1(1-21). Therefore, trachea-constricting peptides ETs(1-31) play roles as vasoconstrictors in a different manner from ETs(1-21).
T. Towatari, T. Miyamura, A. Kondo, I. Kato, M. Inoue, Mihiro Yano and Hiroshi Kido : The Structure of Asparagine-linked Oligosaccharide of Rat Liver Cathepsin L Reflect the Substrate Specificity of Lysosomal Alpha-Mannosidase, European Journal of Biochemistry, Vol.256, No.1, 163-169, 1998.
(要約)
We have studied the structures of asparagine-linked oligosaccharides of cathepsin L purified from rat liver in detail. The oligosaccharides released from rat liver cathepsin L on glycopeptidase-F treatment were tagged with 2-aminopyridine at their reducing ends. The pyridylamino (PA) derivatives were separated into seven fractions according to molecular size by normal-phase HPLC. The structure of each oligosaccharide thus isolated was analyzed by reversed-phase HPLC and characterized by ion-spray mass spectrometry and high-resolution proton nuclear magnetic resonance (1H-NMR) spectroscopy. Our results indicate that the asparagine-linked oligosaccharides of rat liver cathepsin L are of the oligomannose type, having two to six mannose residues. Among them, the five major ones are Manalpha1-6Manbeta1-4-GlcNAcbeta1-4GlcNAc, Manalpha1 -6Manalpha1-6Manbeta1-4GIcNAcbeta1-4GlcNAc, Manalpha1-6(Manalpha1-3)-Manalpha1-6Manbeta1- 4GlcNAcbeta1-4GlcNAc, Manalpha1-6(Manalpha1-3)Manalpha1-6(Manalpha1-3) Manbeta1-4Glc-NAcbeta1-4GlcNAc, and Manalpha1-6(Manalpha1-3)Manalpha1-6(Manalpha1-++ +2Manalpha1-3)Manbeta1-4GlcNAcbeta1-4Glc-NAc. Their structures are shown to be products of Man6GlcNAc2 hydrolysis with lysosomal alpha-mannosidase.
H. Mori, M. Kamada, M. Maegawa, S. Yamamoto, T. Aono, S. Futaki, Mihiro Yano, Hiroshi Kido and S.S. Koide : Enzymatic Activation of Immunoglobulin Binding Factor in Female Reproductive Tract, Biochemical and Biophysical Research Communications, Vol.246, No.2, 409-413, 1998.
(要約)
Human seminal plasma and cervical mucus contains an immunoglobulin binding factor (IgBF) which interacts with IgG as monomers under reducing condition. It may play a role in preventing antibody production against allogeneic sperms in the female reproductive tract. However, since IgBF is secreted as a homodimer that does not bind IgG, in vivo activation systems should be investigated. GSH reduces the inactive native dimer to the active monomer. Protein disulfide isomerase (PDI), a molecular chaperone, alters the configuration of dimers to active monomers. 20S proteasomes produced by activated T cells which cleave the dimers in the presence of GSH to active fragments. All these activating systems are widely distributed as cellular enzymes in vivo. Also PDI mRNAs are expressed in uterine cervix, endometrium and fallopian tube. Since these enzymes are produced upon stimulation by the immune system, we hypothesize that immunocompetent cells interact with allogeneic sperms, leading to the local production of these enzymes that will activate IgBF.
M. Abe, M. Kurosawa, O. Ishikawa, Y. Miyach and Hiroshi Kido : Mast Cell Tryptase Increases not Only the Proliferation of Human Dermal Fibroblast but Also Type I Collagen Production, Clin. & Exp. Allergy, Vol.28, 1509-1517, 1998.
M. Yoshizumi, D. Inui, N. Okishima, H. Houchi, K. Tsuchiya, H. Wakabayashi, Hiroshi Kido and T. Tamaki : Endothelin-1 (1-31), A Novel Vasoactive Peptide Increases [Ca2+ ]i in Human Coronary Artery Smooth Muscle Cells, European Journal of Pharmacology, Vol.348, No.2-3, 305-309, 1998.
(要約)
We have previously found that human chymase cleaves big endothelins at the Tyr31-Gly32 bond and produces 31-amino acid long endothelins-(1-31), without any further degradation products. In this study, we investigated the effect of synthetic endothelin-1-(1-31) on the intracellular free Ca2+ concentration ([Ca2+]i) in cultured human coronary artery smooth muscle cells. Endothelin-1-(1-31) increased [Ca2+]i in a concentration-dependent manner (10(-14) to 10(-10) M). This endothelin-1-(1-31)-induced [Ca2+]i increase was not affected by phosphoramidon (N-(alpha-Rhamnopyranosyloxyhydroxyphosphinyl)-L-Leucyl-L-Tryptoph an), an inhibitor of endothelin-converting enzyme. It was, however, inhibited by 10(-10) M BQ123 (Cyclo-(-D-Trp-D-Asp(ONa)-Pro-D-Val-Leu-)), an endothelin ET(A) receptor antagonist, but not by 10(-10) M BQ788 (N-cis-2,6-dimethylpiperidinocarbonyl-L-yMeLeu-D-Trp(COOM e)-D-Nle-ONa), an endothelin ET(B) receptor antagonist. These results suggest that endothelin-1-(1-31) by itself exhibits vasoactive properties probably through endothelin ET(A) receptors. Since human chymase has been reported to play a role in atherosclerosis, endothelin-1-(1-31) may be one of the candidate substances for its cause.
Hiroshi Kido, A. Nakano, N. Okishima, H. Wakabayashi, F. Kishi, Y. Nakaya, M. Yoshizumi and T. Tamaki : Human Chymase as the Enzyme Forming Novel Bioactive 31-Amino Acid Length Endothelins, Biological Chemistry, Vol.379, No.7, 885-891, 1998.
(要約)
We report the novel role of human chymase in the production of bioactive 31-amino acid length endothelins (ETs), which may play a role in allergies and vascular diseases. In the bronchi of asthmatic patients, the vascular tissue in atherosclerosis, and the heart muscle in cardiac hypertrophy, both ET-like immunoreactivity and the accumulation of mast cells significantly increase. Chymase from human mast cells selectively cleaves big ET-1, -2 and -3 at their Tyr31-Gly32 bonds, and produces novel bioactive 31-amino acid length ETs, ETs(1-31), without any further degradation products. However, chymases from other species, human cathepsin G, and porcine alpha-chymotrypsin, degrade big ETs. ETs(1-31) at concentrations between 10(-9) M and 10(-7) M exhibited various contractile potencies in rat tracheae and porcine coronary arteries in a dose-dependent manner. Furthermore, ET-1(1-31) at concentrations between 10(-14) M and 10(-10) M caused a significant increase in the intracellular free Ca2+ concentration. The contractile activity of ETs(1-31) may not be the consequence of conversion to the corresponding ETs(1-21) by phosphoramidon-sensitive ET converting enzyme(s) or other chymotrypsin-type proteases and metallo-endopeptidases, because the contractile activity was not significantly inhibited on treatment with inhibitors of these proteases prior to the addition of ET-1(1-31).
M. Hiromura, Mihiro Yano, H. Mori, M. Inoue and Hiroshi Kido : Intrinsic ADP-ATP Exchange Activity is a Novel function of the Molecular Chaperone, Hsp70, The Journal of Biological Chemistry, Vol.273, No.10, 5435-5438, 1998.
(要約)
Hsp70 is a multifunctional molecular chaperone whose interactions with protein substrates are regulated by ATP hydrolysis and ADP-ATP exchange. We show here that, in addition to ATPase activity, purified Hsp70 free from nucleoside-diphosphate (NDP) kinase exhibits intrinsic ADP-ATP exchange activity. The rate constants for ATP hydrolysis and ATP synthesis were in a similar range at the optimum pH of 7.5-8.5 in the presence of 5 mM ATP and 0.5 mM ADP. Hsp70 exhibited a considerably strict preference for ATP as a phosphate donor, and a biased substrate specificity, unlike NDP kinase; ADP, UDP, CDP > dTDP, dCDP > GDP, dGDP. During the reaction, Hsp70 formed an acid-labile autophosphorylated intermediate, and nucleoside diphosphate-dependent dephosphorylation of the latter then occurred. These properties of Hsp70 are not identical but similar to those of NDP kinase, but are not similar to those of adenylate kinase and ATP synthase.
Masanori Yoshizumi, Daisuke Inui, Naoko Okishima, Hitoshi Houchi, Koichiro Tsuchiya, Hideki Wakabayashi, Hiroshi Kido and Toshiaki Tamaki : Endothelin-1(1-31), a novel vasoactive peptide, increases [Ca2+]i in humen coronary artery smooth muscle cells., European Journal of Pharmacology, Vol.348, No.2-3, 305-309, 1998.
194.
Masanori Yoshizumi, Shokei Kim, Shoji Kagami, Akinori Hamaguchi, Koichiro Tsuchiya, Hitoshi Houchi, Hiroshi Iwao, Hiroshi Kido and Toshiaki Tamaki : Effect of endothelin-1(1-31)on extracellular signal-regulated kinase and proliferation of human coronary artery smooth muscle cells., British Journal of Pharmacology, Vol.125, No.5, 1019-1027, 1998.
195.
Hiroshi Kido, Ayako Nakano, Naoko Okishima, Hideki Wakabayashi, Fumiko Kishi, Yutaka Nakaya, Masanori Yoshizumi and Toshiaki Tamaki : Human chymase, an enzyme forming novel bioactive 31-amino acid length endothelins., The Journal of Biological Chemistry, Vol.379, 885-891, 1998.
196.
T. Brinkmann, J. Schafers, L. Gurtler, Hiroshi Kido, Y. Niwa, N. Katunuma and H. Tschesche : Inhibition of Tryptase TL2 from Human T4+lymphocytes and Inhibition of HIV-1 Replication in H9 Cells by Recombinant Aprotinin and Bikunin Homologues, Journal of Protein Chemistry, Vol.16, No.6, 651-660, 1997.
(要約)
The serine esterase TL2 from human T4+ lymphocytes is a binding component to HIV-1 glycoprotein gp120 and seems to play a role in the HIV-1 infection mechanism. Recombinant variants of the Kunitz-type serine proteinase inhibitor aprotinin were investigated for their ability to inhibit tryptase TL2 and the binding of gp120 to this enzyme. Furthermore, the viral replication of HIV-1 was investigated H9 cell cultures under the influence of recombinant aprotinin and bikunin variants. In contrast to native aprotinin, the recombinant variant [Arg15, Phe17, Glu52] aprotinin with a reactive-site sequence homologous to the V3 loop of HIV-1 gp120 showed a specific inhibition of tryptase TL2 (> 80%). However, the [Leu15, Phe17, Glu52] aprotinin variant with hydrophobic subsites was the most potent inhibitor of the binding of gp120 to tryptase TL2 (68%). Our results show that the enzyme activity of purified tryptase TL2 is inhibited not only by variants with basic amino acids, but also those with hydrophobic residues in the reactive-site region. Therefore, tryptase TL2 is not a typical trypsin-like or chymotrypsin-like protease. Investigations on inhibition of HIV-1 replication in H9 cell cultures showed that tryptase TL2 is involved in the mechanism of virus internalization into human lymphocytes. The [Leu15, Phe17, Glu52] aprotinin showed a significant retardation of syncytium formation over a period of 5 days in a 1 micro M concentration. Similar investigations were performed with recombinant variants of bikunin, the light chain of human inter-alpha-trypsin inhibitor. Only the single-headed variant [Arg94] delta 2 bikunin inhibited slightly the syncytium formation over a period of 2 days in a 2.2 micro M concentration. Wild-type bikunin and all full-length variants showed no effect, possibly due to steric hindrance by the second domain of the double-headed inhibitor.
Mihiro Yano, Mori S., Niwa Y., Inoue M. and Hiroshi Kido : Intrinsic Nucleoside Diphosphate Kinase-like Activity as a Novel Function of 14-3-3 Proteins, FEBS Letters, Vol.419, 244-248, 1997.
198.
Hiroshi Kido, Y. Beppu and Y. Immamura : Human Mucus Protease Inhibitor and its Mutants Prevent Infection by Influenza A and Sendai Viruses, Medical Aspects of Proteases and Protease Inhibitors, 118-127, 1997.
199.
A. Nakano, F. Kishi, K. Minami, H. Wakabayashi, Y. Nakaya and Hiroshi Kido : Selective Conversion of Big Endothelins to Tracheal Smooth Muscle-Constricting 31-Amino Acid Length Endothelins by Chymase from Human Mast Cells, The Journal of Immunology, Vol.159, No.4, 1987-1992, 1997.
(要約)
Chymase from human mast cells selectively cleaved big endothelins (ETs) at the Tyr31-Gly32 bond and produced novel trachea-constricting 31-amino acid-length endothelins, ETs(1-31), without any further degradation products. Chymases from other species, such as the enzymes from rat connective tissue and mucosal mast cells, and the other chymotrypsin-like proteases examined degraded big ETs. ETs(1-31) exhibited various contractile potencies as to the rat trachea in comparison with 21-amino acid-length endothelins, ETs(1-21), and big ETs: ET-1(1-21) > ET-1(1-31) > big ET-1; ET-2(1-31) > ET-2(1-21) > or = big ET-2; ET-3(1-21) > or = ET-3(1-31) > or = big ET-3. Among the ETs(1-31), ET-2(1-31) was the most potent constrictor, its potency being similar to that of ET-1(1-21) and stronger than that of ET-2(1-21). The contractile activity of ETs(1-31) may not be the consequence of conversion to the corresponding ETs(1-21) by phosphoramidon-sensitive ET-converting enzymes or other chymotrypsin-type proteases and metalloendopeptidases, because the contractile activity was not inhibited significantly on treatment with inhibitors of these proteases before the addition of ET-1(1-31). Inhibitors of chymotrypsin-type serine proteases, on the contrary, significantly enhanced the contractile activity exhibited by ET-1(1-31) and big ET-1, but not that by ET-1(1-21). These results suggest that protease(s) on the surface of the rat trachea tends to degrade ETs(1-31) and big ETs, and thereby reduces their contractile activity. Taken together, the results suggest that trachea-constricting ETs(1-31) generated by human chymase may play a role in the hyper-responsive airway in allergic inflammation.
T. Towatari, T. Miyamura, A. Kondo, I. Kato and Hiroshi Kido : Structure Studies on the Carbohydrate Moiety of RatLliver Cathepsin L, Proteolysis and Protein Turnover, 61-67, 1997.
201.
Hiroshi Kido, Y. Beppu, K. Sakai and T. Towatari : Protease Inhibitors and a Pulmonary Surfactant in Airway Fluid are Potential Defensive, Proteolysis and Protein Turnover, 545-554, 1997.
Hiroshi Kido, Y. Beppu, K. Sakai and T. Towatari : Molecular Basis of Proteolytic Activation of Sendai Virus Infection and the Defensive, Biol. Chem.,, Vol.378, 255-263, 1997.
Y. Beppu, Y. Imamura, M. Tashiro, T. Towatari, H. Ariga and Hiroshi Kido : Human Mucus Protease Inhibitor in Airway Fluids Is a Potential Defensive Compound against Infection with Influenza A and Sendai Viruses, The Journal of Biochemistry, Vol.121, No.2, 309-316, 1997.
(要約)
Tryptase Clara, a trypsin-like protease localized exclusively in and secreted from Clara cells to the bronchial epithelium of rat, proteolytically activates the infectivity of influenza A virus [H. Kido, Y. Yokogoshi, K. Sakai, M. Tashiro, Y. Kishino, A. Fukutomi, and N. Katunuma (1992) J. Biol. Chem. 267, 13573-13579]. We report here that human mucus protease inhibitor (MPI), a major inhibitor of granulocyte elastase in the lining fluids of the human respiratory tract, significantly inhibited proteolytic activation of the infectivity of influenza A and Sendai viruses by tryptase Clara in vitro and multi-cycles of mouse-adapted influenza A virus replication in rat lungs in vitro. Recombinant MPI and the C- but not the N-terminal domain of the MPI inhibited both the proteolytic activity of tryptase Clara and the activation of virus infection. The 50% inhibitory concentrations of recombinant MPI and the C-terminal domain for tryptase Clara with Sendai virus envelope glycoprotein as substrate were 7.4 and 61.6 nM, respectively. These results indicate that MPI is a defensive compound against virus infection. Since there is evidence suggesting that concentrations of MPI in respiratory fluids are insufficient for prevention of virus infection, administration of MPI in the airway may be useful for treatment of these virus infections.
Yuushi Okumura, H. Sato, M. Seiki and Hiroshi Kido : Proteolytic Activation of the Precursor of Membrane Type 1 Matrix Metalloproteinase by Human Plasmin, FEBS Letters, Vol.402, 181-184, 1997.
207.
M. Tashiro, Y. Beppu, K. Sakai and Hiroshi Kido : Inhibitory Effect of Pulmonary Surfactant on Sendai Virus Infection in Rat Lungs, Arch. Virol., Vol.141, 1571-1577, 1996.
208.
Hiroshi Kido, Y Niwa, Y Beppu and T Towatari : Cellular Proteases Involved in The Pathogenicity of Enveloped Animal Virus, Human Immunodeficiency Virus, Influenza Virus A and Sendai Virus, Advances in Enzyme Regulation, Vol.36, 325-347, 1996.
(要約)
In enveloped viruses, post-translational proteolytic activation is a critical step for the fusion activity and thus for the infectivity of the virus. In addition to the membrane receptors for the viruses, proteolytic activation is indispensable for effective virus spread in the infected host and it is a prime determinant for pathogenicity. Here we described the host cellular processing proteases, tryptase Clara and tryptase TL2, which proteolytically activate the infectivity of influenza A and Sendai viruses in the respiratory tract and HIV-1 in human CD4+ T cells, respectively. A novel trypsin-like protease, designated tryptase Clara, was purified from rat lung. The enzyme is localized in Clara cells of the bronchial epithelium and is secreted into the airway lumen. The enzyme specifically recognizes the consensus cleavage motif Gln(Glu)-X-Arg of influenza A and Sendai viruses and proteolytically activates the envelope fusion glycoproteins of the progeny viruses extracellularly in the airway lumen. Human mucus protease inhibitor and pulmonary surfactant in airway fluid inhibited the proteolytic activation of these viruses and also suppressed multiple cycles of viral replication in vitro. These results suggest that an imbalance between the amount of tryptase Clara and that of endogenous inhibitors in airway fluid is a prime determinant for pneumopathogenicity of the viruses. Therefore supplementing an endogenous inhibitor at therapeutic doses may protect against virus infection. In HIV-1 infection, binding of the gp120 envelope glycoprotein to the CD4 receptor is not sufficient in itself to allow virus entry, and an additional component(s) in the membrane is required for cell infection as a cofactor. We isolated a serine protease named tryptase TL2, in the membrane of CD4+ lymphocytes, which specifically binds to the V3 loop of HIV-1 gp120 as a cofactor. After binding, tryptase TL2 proteolytically processed gp120 into two protein species of 70 and 50 kDa and the cleavage was suppressed by a neutralizing antibody against the V3 loop. The amino acids that constitute the cleavage sites in the V3 loop of almost all HIV isolates are variable, but they are restricted to those which are susceptible to chymotryptic and/or tryptic enzyme. The multi-substrate specificity of tryptase TL2, which has tryptic and chymotryptic specificities, may correspond tot he variability of the V3 loop. The selective cleavage of the V3 loop by tryptase TL2 may lead to a conformational change of gp120, resulting in the dissociation of gp120 from gp41, exposing the fusogenic domain of the transmembrane protein gp41 following virus-host cell fusion.
Yuushi Okumura, A. Kudoh, M. Takashima, M. Inoue, K. Sakai and Hiroshi Kido : Purification and Characterization of a Novel Isoform of Mast Cell Tryptase from Rat Tongue, The Journal of Biochemistry, Vol.120, No.4, 856-864, 1996.
(要約)
Rat mast cell tryptase was purified to homogeneity from rat tongue by a series of standard chromatographic procedures. Since the enzyme gave band corresponding to molecular mass of 32-35 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis and exhibited a molecular mass of 135 kDa on gel filtration, it was presumed to be a noncovalently associated tetramer. The N-terminal amino acid sequence of 50 residues of the enzyme showed the highest degree of homology with the same region in mouse mast cell protease 7 (92%), and less homology to those of tryptases from man and dog, and peritoneal cells of rats and Mongolian gerbils. The inhibitor specificity of rat tongue tryptase was similar to that of rat peritoneal mast cell tryptase free from trypstatin: it was inhibited by alpha 1-antitrypsin, Kunitz-type soybean trypsin inhibitor and Bowman-Birk soybean trypsin inhibitor, but these inhibitors do not inhibit the tryptases from rat skin, human lung, and dog mast cells. Judging from these results, together with other enzymatic properties, the enzyme may be a novel isoform of tryptase in rat tongue. Analysis by differential staining with peroxidase-labeled lectins of the enzyme suggested that it has tri- and/or tetraantennary complex-type oligosaccharides containing a relatively high amount of sialic acid. The immunohistochemical distribution of this enzyme indicated that the reactive antigen was specific in connective tissue but not in mucosal mast cells.
Niwa Y., Mihiro Yano, Futaki S., Yuushi Okumura and Hiroshi Kido : T Cell Membrane Associated Serine Protease, Tryptase TL2, Binds Human Immunodeficiency Virus Type 1 gp120 and Cleaves The Third-Variable-Domain Loop of gp120. Neutralizing Antibodies of Human Immunodeficiency Virus Type I Inhibits Cleavage of gp120, Eur. J. Biochem., Vol.237, 64-70, 1996.
Hiroshi Kido, T. Towatari, Y. Niwa, Yuushi Okumura and Y. Beppu : Cellular Proteases Involved in the Pathogenicity of Human Immunodeficiency and Influenza Virus, Advances in Experimental Medicine and Biology, Vol.389, 233-240, 1996.
215.
Lihong Wang, Hideki Hayashi, Yasumasa Mitani, Kazuo Ishii, Tetsuo Ohnishi, Yasuharu Niwa, Hiroshi Kido and Yousuke Ebina : Cloning of a cDNA encoding a 190-kDa insulin receptor substrate-1-like protein of simian COS cells., Biochemical and Biophysical Research Communications, Vol.216, No.1, 321-328, 1995.
216.
L. Wang, H. Hayashi, Y. Mitani, K. Ishii, T. Ohnishi, Y. Niwa, Hiroshi Kido and Y. Ebina : Cloning of a cDNA Encoding a 190-kDa Insulin Receptor Substrate-1-like Protein of Simian CO5 Cells, Biochemical and Biophysical Research Communications, Vol.216, No.1, 321-328, 1995.
(要約)
Major insulin signals such as stimulation of glucose uptake and DNA synthesis and modification of hexose metabolism are mediated by the tyrosine-phosphorylated insulin receptor substrate-1 (IRS-1; pp180) in many species of cells. We cloned cDNA encoding a 190-kDa IRS-1-like protein (pp190) in simian COS cells and which is slightly larger than IRS-1 (pp180) of human, rat, and mouse cells. The deduced amino acid sequence of COS pp190 consisted of 1251 amino acids and was 96.4%, 87.9% and 88.7% identical to human, mouse and rat IRS-1. The COS pp190 bound to SH2 (src-homology 2) domains of p85, Grb2/Ash, and SH-PTP2, as did IRS-1. In IRS-1-knockout mice, insulin signals are thought to be mediated by IRS-2 (pp190), which is an alternative signaling molecule and is slightly larger than IRS-1. However, the COS pp190 may be a simian homologue of IRS-1, but not of IRS-2. The results of Southern blotting suggested the possibility that Chinese hamster ovary (CHO) cells have not only the IRS-1 gene but also a gene related to the COS pp190.
T. Ohshita and Hiroshi Kido : Simple Preparation of Rat Brain Lysosomes and Their Proteolytic Properties, Anal. Biochem., Vol.230, 41-47, 1995.
218.
Hiroshi Kido and N. Katunuma : Control of Tryptase and Chymase Activity by Protease Inhibitors, Mast Cell Proteases in Immunology and Biology, Vol.11, 237-255, 1995.
219.
Keiichi Kamoshita, Mayumi Shiota, Masafumi Sasaki, Yasuhiro Koga, Yuushi Okumura and Hiroshi Kido : Calcium Requirement and Inhibitor Spectrum for Intracellular HIV Type 1 gp160 Processing in Cultured HeLa Cells and CD4+ Lymphocytes: Similarity to Those of Viral Envelope Glycoprotein Maturase, The Journal of Biochemistry, Vol.117, No.6, 1244-1253, 1995.
N. Shimizu, M. Kobayashi, Hui-Yu Liu, Hiroshi Kido and H. Hoshino : Detection of Tryptase TL2 and CD26 Antigen in Brain-Derived Cells Non-Permissive to T-Cell-Line-Tropic Human Immunodeficiency Virus Type 1, FEBS Lett.,, Vol.358, 48-52, 1995.
222.
T. Yamaguchi, M. Fukase, T. Sugimoto, Hiroshi Kido and K. Chihara : Purification of Meprin from Human Kidney and its Role in Parathyroid Hormone Degradation, Biol. Chem. Hoppe-Seyler, Vol.375, 821-824, 1994.
223.
T. Brinkmann, J. Schafers, L. Gurtler, Hiroshi Kido, Y. Niwa, N. Katunuma and H. Tschesche : Inhibition of Tryptase TL2 from Human T4+ lymphocytes and In Vitro Inhibition of HIV-1 Replication in H9 Cells by Recombinant Aprotinin and Bikunin Variants, Journal AIDS-Forschung (AIFO),, Vol.9, 602, 1994.
224.
T. Ohshita and Hiroshi Kido : Involvement of a Cystatin-a-Sensitive Cysteine Proteinase in the Degradation of Native L-lactate Dehydrogenase and Serum Albumin by Rat Liver or Kidney Lysosomes, Eur. J. Biochem.,, Vol.225, 781-786, 1994.
225.
M. Inoue, T. Hoshino, T. Fukuma, Y. Niwa and Hiroshi Kido : Close Co-localization of CD4 and a Serine Esterase Tryptase TL2 on the Cell-surface of Human Monocytoid and CD4+ lymphoid Cells, Biochemical and Biophysical Research Communications, Vol.201, No.3, 1390-1395, 1994.
(要約)
Tryptase TL2, a serine esterase in the membrane of human monocytoid and CD4+ lymphoid cells, specifically binds to the V3 domain of HIV-1 gp120. Here we report that monoclonal antibodies against CD4 that recognize the epitope interacting with gp120 specifically blocked the immunostaining of cell-surface tryptase TL2, although the antibody does not cross-react with tryptase TL2. Down-regulation of cell-surface CD4 induced by HIV-1 Nef prevented this blocking effect. These data suggest that CD4 is closely co-localized with tryptase TL2 on the cell surface and that regulation of the expression of tryptase TL2 is not associated with that of CD4.
T. Yamaguchi, M. Fukase, Hiroshi Kido, T. Sugimoto, N. Katunuma and K. Chihara : Meprin is Predominantly Involved in Parathyroid Hormone Degradation by the Microvillar Membranes of Rat Kidney, Life Sciences, Vol.54, No.5, 381-386, 1994.
227.
Y. Matsunaga, Hiroshi Kido, K. Kawaji, K. Kamoshita, N. Katunuma and T. Ogura : Inhibitors of Chymotrypsin-like Proteases Inhibit Eosinophil Peroxidase Release from Activated Human Eosinophils, Arch. Biochem. Biophys., Vol.312, No.1, 67-74, 1994.
228.
S. Iwata, A.C. Schmidt, K. Titani, M. Suzuki, Hiroshi Kido, B. Gotoh, M. Hamaguchi and Y. Nagai : Assignment of Disulfide Bridges in the Fusion Glycoprotein of Sendai Virus, Journal of Virology, Vol.68, No.5, 3200-3206, 1994.
(要約)
The mature fusion (F) glycoprotein of the paramyxovirus family consists of two disulfide-linked subunits, the N-terminal F2 and the C-terminal F1 subunits, and contains 10 cysteine residues which are highly conserved at specific positions. The high level of conservation strongly suggests that they are indeed disulfide linked and play important roles in the folding and functioning of the molecule. However, it has not even been clarified which cysteine residues link the F2 and F1 subunits. This report describes our assignment of the disulfide bridges in purified Sendai virus F glycoprotein by fragmentation of the polypeptide and isolation of cystine-containing peptides and determination of their N-terminal sequences. The data demonstrate that all of the 10 cysteine residues participate in disulfide bridges and that Cys-70, the only cysteine in F2, and Cys-199, the most upstream cysteine in F1, form the interchain bond. Of the remaining eight cysteine residues clustered near the transmembrane domain of F1, the specific bridges identified are Cys-338 to Cys-347 and Cys-362 to Cys-370. Although no exact pairings between the subsequent four residues were defined, it seems likely that the most downstream, Cys-424, is linked to Cys-394, Cys-399, or Cys-401. Thus, we conclude that the cysteine-rich domain indeed contributes to the formation of a bunched structure containing at least two tandem cystine loops.
K. Sakai, T. Kohri, M. Tashiro, Y. Kishino and Hiroshi Kido : Sendai Virus Infection Changes the Subcellular Localization of Tryptase Clara in Rat Bronchiolar Epithelial Cells, Eur. Respir. J., Vol.7, 686-692, 1994.
231.
木戸 博, 丹羽 保晴, 井上 雅広 : HIV の外膜蛋白質 gp120 のV3領域と特異的に結合するトリプターゼ TL2, Medical Immunology, Vol.27, No.4, 351-359, 1994年.
232.
M. Takeda, S. Tanaka, Hiroshi Kido, S. Daikoku, M. Oka, K. Sakai and N. Katunuma : Chromaffin Cells Express Alzheimer Amyloid Precursor Protien in the Same Manner as Brain Cells, Neuroscience Letters, Vol.168, 57-60, 1994.
S Takada, Hiroshi Kido, A Fukutomi, T Mori and K Koike : Interaction of Hepatitis B Virus X Protein with a Serine Protease, Tryptase TL2, as an Inhibitor, Oncogene, Vol.9, No.2, 341-348, 1994.
(要約)
X protein of hepatitis B virus (HBV) transactivates transcription of various viral and cellular genes. It has been suggested that X protein plays a major role in hepatocarcinogenesis by HBV. The protein possesses amino acid sequence homology to the functionally essential domain of Kunitz-type serine protease inhibitors. This Kunitz domain-like sequence in X protein is indispensable for the transactivation function. To clarify whether X protein has a serine protease inhibitor activity, a search was made for serine proteases which interact with, but not degrade X protein. Tryptase TL2, one of serine proteases in hepatic cells, was found to directly interact with X protein without degradation. Moreover, the activities of tryptase TL2 and an analogous protease were substantially inhibited by X protein. These results suggest that transactivation function of X protein is exerted by modulation of the hepatic serine protease activity, giving rise to quantitative or qualitative change of cellular transcription factor(s) through protection from proteolytic degradation and/or suppression of processing.
Hiroshi Kido, K. Kamoshita and N. Katunuma : A Processing Protease for gp160 HIV-1 Envelope Glycoprotein Precursor in Human T4+ lymphocytes, Biological Functions of Proteases and Inhibitors, 199-212, 1993.
236.
Hiroshi Kido : Cellular processing proteases for the envelope glycoprotein precursors of human immunodeficiency virus-1 and of influenza and parainfluenza viruses, Proteolysis and Protein Turnover, 209-217, 1993.
237.
Y. Matsunaga, T. Saibara, Hiroshi Kido and N. Katunuma : Participation of cathepsin B in Processing of antigen Presentation to MHC-Class II, FEBS Letters, Vol.324, No.3, 325-330, 1993.
238.
Y. Sukenaga, Hiroshi Kido, A. Neki, M. Enomoto, K. Ishida, K. Takagi and N. Katunuma : Purification and Molecular Cloning of Chymase from Human Tonsils, FEBS Letters, Vol.323, No.1,2, 119-122, 1993.
M. Tashiro, M. Takeda, S. Tanaka, N. Nishimura, M. Takenaka and Hiroshi Kido : Antibody Against the Carboxyl Terminus of the F2 Subunit of Sendai Virus Fusion Glycoprotein Inhibits Proteolytic Activation, Virology, Vol.194, 882-885, 1993.
Hiroshi Kido, K. Sakai, Y. Kishino and M. Tashiro : Pulmonary Surfactant is A Potential Endogenous Inhibitor of Proteolytic Activation of Sendai Virus and Influenza A Virus, FEBS Lett.,, Vol.322, No.2, 115-119, 1993.
Hiroshi Kido, Keiichi Kamoshita, Akiko Fukutomi and Nobuhiko Katunuma : Processing Protease for gp160 Human Immunodeficiency Virus Type I Envelope Glycoprotein Precursor in Human T4+ lymphocytes: Purification and Characterization, The Journal of Biological Chemistry, Vol.268, No.18, 13406-13413, 1993.
(要約)
A processing protease for the human immunodeficiency virus type I (HIV-I) envelope glycoprotein gp160 precursor has been purified to homogeneity from the post-nuclear membrane fraction of a human T4+ lymphocyte clone. Most of the processing activity was found to be present in the fractions of endoplasmic reticulum and Golgi apparatus of the cells. The purified enzyme has a monomeric structure with a molecular mass of 26 +/- 3 kDa, as judged by gel-permeation liquid chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing and nonreducing conditions. The purified enzyme converted gp160 to gp120 and gp41, showing a pH optimum of 6.5-7.0. Direct amino acid sequencing of the amino terminus of the product gp41 revealed that the cleavage site of gp160 was between Arg511 and Ala512. The enzyme activity was inhibited by trypsin-type protease inhibitors, but was not affected by CaCl2, MgCl2 or chelating agents. The properties of the purified enzyme are clearly distinct from those of processing proteases reported previously. Although the significance of the enzyme in vivo is not currently certain, judging from its cleavage specificity and subcellular localization, this endopeptidase appears to be a processing enzyme for the human immunodeficiency virus type I gp160 precursor protein in human T cells.
(キーワード)
Amino Acid Sequence / CD4-Positive T-Lymphocytes / Cell Line / Cell Membrane / Chromatography, Liquid / Electrophoresis, Polyacrylamide Gel / Endopeptidases / Gene Products, env / HIV Envelope Protein gp160 / HIV-1 / Humans / Molecular Sequence Data / Protease Inhibitors / Protein Precursors / Protein Processing, Post-Translational / Substrate Specificity / Virus Replication
(文献検索サイトへのリンク)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 8099909
Hiroshi Kido, M. Takeda, H. Wakabayashi, S. Tanaka, N. Nishimura, M. Takenaka and M. Okada : Purification of a Trypsin-Type Protease from Human Umbilical Vein Endothelial Cells, That is Highly Sensitive to The Kunitz Inhibitor Domain Peptide of Alzheimer's Disease Amyloid Protein Precursor, Gerontology, Vol.39, 30-37, 1993.
Toru Yamaguchi, Hiroshi Kido, Riko Kitazawa, Sohei Kitazawa, Masaaki Fukase and Nobuhiko Katunuma : A Membrane-Bound Metallo-Endopeptidase from Rat Kidney: Its Immunological Characterization,, The Journal of Biochemistry, Vol.113, No.3, 299-303, 1993.
(要約)
The structure and location of a membrane-bound metallo-endopeptidase, previously purified from rat kidney [Yamaguchi et al. (1991) Eur. J. Biochem. 200, 563-571], were examined by immunochemical and immunohistochemical methods with a rabbit polyclonal antibody against the purified enzyme. On treatment with endoglycosidase F, the subunit of the purified enzyme (molecular mass = 88 kDa) was converted to a smaller form (78.5 kDa), indicating that the enzyme contained at least 11% N-linked carbohydrate. Treatment of kidney membranes with papain resulted in release of the enzyme, as shown by Western blotting analysis of the solubilized fraction. Immunoassays of rat tissues showed that only the kidney, and small and large intestine expressed significant amounts of the antigen. Moreover, immunohistochemical studies showed that the antigen was confined to the luminal surfaces of the proximal renal tubules and the intestinal villi. Thus, like another kidney membrane metallo-endopeptidase, meprin [Kounnas et al. (1991) J. Biol. Chem. 266, 17350-17357], the purified enzyme is shown to be a glycoprotein that is probably anchored in the plasma membrane, and located in the luminal surface of microvillar membranes of the kidney and intestine. These results indicate that our enzyme and meprin have clear structural and topological similarities.
H. Hayashi, Y. Nishioka, S. Kamohara, F. Kanai, K. Ishii, Y. Fukui, F. Shibasaki, T. Takenawa, Hiroshi Kido, N. Katunuma and Y. Ebina : The a-Type 85-kDa Subunit of Phosphatidylinositol 3-Kinase is Phosphorylated at Tyrosine 368, 580, and 607 by The Insulin Receptor, The Journal of Biological Chemistry, Vol.268, No.10, 7107-7117, 1993.
(要約)
We have shown previously that insulin stimulated the tyrosine phosphorylation of the alpha-type 85-kDa subunit (p85) of phosphatidylinositol (PI) 3-kinase in vitro and in vivo. In the present work, we identified the major tyrosine phosphorylation sites of the alpha-type p85 by the insulin receptor. [32P]Phosphopeptides obtained from lysylendopeptidase digestion of phosphorylated alpha-type p85 in intact cells after insulin treatment were analyzed using reverse-phase high performance liquid chromatography and thin layer electrophoresis. The tyrosine phosphorylation sites of alpha-type p85 in vivo were assigned to three major phosphopeptides, designated p1, p2, and p3. Highly purified insulin receptor also phosphorylated the purified p85 of PI 3-kinase from the bovine thymus at p1. The purified glutathione S-transferase (GST)-p85 (alpha-type) fusion protein and its truncated proteins from Escherichia coli were also phosphorylated by the purified insulin receptor at p1, p2, and p3 in vitro. Analysis of [32P]phosphopeptide of the truncated GST-p85 (alpha-type) fusion proteins and radiosequence analysis revealed that the p1, p2, and p3 phosphopeptides were phosphorylated at tyrosines 607, 580, and 368, respectively. In addition, phenylalanine substitutions at tyrosine 607 and 580 reduced the p1 and p2 phosphopeptides in vivo, respectively. We conclude that the alpha-type p85 of PI 3-kinase was phosphorylated at tyrosines 368, 580, and 607 by the insulin receptor in vivo.
K Sakai, Y Kawaguchi, Y Kishino and Hiroshi Kido : Electron Immunohistochemical Localization in Rat Bronchiolar Epithelial Cells of Tryptase Clara which Determines The Pneumotropism and Pathogenicity of Sendai Virus and Influenza Virus, The Journal of Histochemistry and Cytochemistry, Vol.41, No.1, 89-93, 1993.
(要約)
The intracellular localization in rat bronchiolar epithelial cells of a novel trypsin-like protease named tryptase Clara, a possible activator of inactive viral fusion glycoprotein of influenza A and wild-type Sendai virus in the respiratory tract, was examined by electron microscopy. In thin sections embedded in LR White, gold particles indicating immunoreactivity of tryptase Clara were detected specifically in secretory granules of Clara cells. No immunoreactivity was detected in bronchiolar ciliated cells, alveolar cells including epithelial Type I and II cells, or alveolar macrophages. Some granules enveloped in a thin membrane and labeled intensely with immunogold particles were seen protruding from peripheral and submembrane regions and a few were observed free in the airway lumen. Scanning electron microscopy also revealed smooth droplets along the main body of Clara cells. These data suggest that tryptase Clara is secreted into the bronchiolar lumen. These findings are the first to show the subcellular localization of tryptase Clara in rat bronchioles and suggest the site of proteolytic activation of the progeny of enveloped pneumotropic viruses, such as Sendai virus and influenza virus.
木戸 博 : エイズウイルス感染と細胞内プロテアーゼ, BIO medica, Vol.7, No.6, 19-25, 1992年.
256.
M. Tashiro, Y. Yokogoshi, K. Tobita, J.T. Seto, R. Rott and Hiroshi Kido : Tryptase Clara, An Activating Protease for Sendai Virus in Rat Lungs, is Involved in Pneumopathogenicity, Journal of Virology, Vol.66, No.12, 7211-7216, 1992.
(要約)
Tryptase Clara is an arginine-specific serine protease localized exclusively in and secreted from Clara cells of the bronchial epithelium of rats (H. Kido, Y. Yokogoshi, K. Sakai, M. Tashiro, Y. Kishino, A. Fukutomi, and N. Katunuma, J. Biol. Chem. 267:13573-13579, 1992). The purified protease was shown in vitro to behave similarly to trypsin, cleaving the precursor glycoprotein F of Sendai virus at residue Arg-116 and activating viral infectivity in a dose-dependent manner. Anti-tryptase Clara antibody inhibited viral activation by the protease in vitro in lung block cultures and in vivo in infected rats. When the enzyme-specific antibody was administered intranasally to rats that were also infected intranasally with Sendai virus, activation of progeny virus in the lungs was significantly inhibited. Thus, multiple cycles of viral replication were suppressed, resulting in a reduction in lung lesions and in the mortality rate. These findings indicate that tryptase Clara is an activating protease for Sendai virus in rat lungs and is therefore involved in pulmonary pathogenicity of the virus in rats.
木戸 博 : HIV (侵入に関与する細胞因子), Mebio, Vol.9, No.6, 32-40, 1992年.
258.
Hiroshi Kido, Yutaka Yokogoshi, Kentaro Sakai, Masato Tashiro, Yasuo Kishino, Aiko Fukutomi and Nobuhiko Katunuma : Isolation and Characterization of A Novel Trypsin-like Protease Found in Rat Bronchiolar Epithelial Clara Cells: A Possible Activator of The Viral Fusion Glycoprotein, The Journal of Biological Chemistry, Vol.267, No.19, 13573-13579, 1992.
(要約)
A novel trypsin-like protease associated with rat bronchiolar epithelial Clara cells, named Tryptase Clara, was purified to homogeneity from rat lung by a series of standard chromatographic procedures. The enzyme has apparent molecular masses of 180 +/- 16 kDa on gel filtration and 30 +/- 1.5 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. Its isoelectric point is pH 4.75. Studies with model peptide substrates showed that the enzyme preferentially recognizes a single arginine cleavage site, cleaving Boc-Gln-Ala-Arg-4-methylcoumaryl-7-amide most efficiently and having a pH optimum of 7.5 with this substrate. The enzyme is strongly inhibited by aprotinin, diisopropylfluorophosphate, antipain, leupeptin, and Kunitz-type soybean trypsin inhibitor, but inhibited only slightly by Bowman-Birk soybean trypsin inhibitor, benzamidine, and alpha 1-antitrypsin. Immunohistochemical studies indicated that the enzyme is located exclusively in the bronchiolar epithelial Clara cells and colocalized with surfactant. An immunoreactive protein with a molecular mass of 28.5 kDa was also detected in airway secretions by Western blotting analyses, suggesting that the 30-kDa protease in Clara cells is processed before or after its secretion. Proteolytic cleavage of the hemagglutinin of influenza virus is a prerequisite for the virus to become infectious. Tryptase Clara was shown to cleave the hemagglutinin and activate infectivity of influenza A virus in a dose-dependent way. These results suggest that the enzyme is a possible activator of inactive viral fusion glycoprotein in the respiratory tract and thus responsible for pneumopathogenicity of the virus.
Hiroshi Kido and N. Katunuma : Biomodulators of Glucocorticoid: Amplifiers and Suppressors of Glucocorticoid Action, Proceedings of The 1st International Congress on "Vitamins and Biofactors in Life Science", 279-282, 1992.
T. Yamaguchi, Hiroshi Kido and N. Katunuma : A Membrane-bound Metallo-endopeptidase from Rat Kidney:Characteristics of Its Hydrolysis of Peptide Hormones and Neuropeptides, Eur. J. Biochem., Vol.204, 547-552, 1992.
Hiroshi Kido, A. Fukutomi and N. Katunuma : A Novel Membrane-bound Serine Esterase in Human T4+ lymphocytes is A Binding Protein of Envelope Glycoprotein gp120 of HIV-1, Biomedica Biochimica Acta, Vol.50, No.4-6, 781-789, 1991.
(要約)
A novel membrane-bound serine esterase, named tryptase TL2, which is immunologically reactive with the antibody inhibiting induction of syncytia by human immunodeficiency virus-1 (HIV-1) (HATTORI, T., KOITO, A., TAKATSUKI, K., KIDO, H., and KATUNUMA, N., 1989, FEBS Lett., 248, 48-52), has been purified from a human T4+ lymphocyte clone. The enzyme has a molecular mass of 198 +/- 15 kDa, and is composed of two subunits of 32 kDa and four subunits of 28 kDa. The enzyme was strongly inhibited by the envelope glycoprotein gp120 of HIV-1, by synthetic peptides of V3 domains of gp120 s with the sequence GPGR in their center, which correspond to the principal neutralizing epitopes of the gp120s of various HIV-1 strains, by Kunitz-type inhibitors with the sequence GPCR in their active site, such as trypstatin, H130, and [Arg15, Glu52] aprotinin and by the microbial inhibitors leupeptin and antipain. This enzyme was specifically bound to the inhibitor V3 domain of gp120 of HIV-1, and this binding was blocked by the inhibitors of tryptase TL2, with a central motif GPCR or GPGR sequence in their center, but not by leupeptin and antipain without the motif. These findings suggest that tryptase TL2 is important in target site recognition and binding of HIV-1 in co-operation with CD4 receptor in the initial process of HIV-1 infection.
R. Sharma, Hiroshi Kido and N. Katunuma : H-7 Reduces The Nuclear Binding of [3H]Dexamethasone in Rat Liver Slice But Does Not Affect The Phosphorylation of Glucocorticoid Receptor, Biochem. Med. Metabol. Biol., Vol.46, 246-254, 1991.
T. Yamaguchi, Hiroshi Kido, M. Fukase, T. Fujita and N. Katunuma : A Membrane-bound Metallo-endopeptidase from Rat Kidney Hydrolyzing Parathyroid Hormone: Purification and Characterization, Eur. J. Biochem., Vol.200, No.2, 563-571, 1991.
Hiroshi Kido, A. Fukutomi and N. Katunuma : Tryptase TL2 in The Membrane of Human T4+ lymphocytes is A Novel Binding Protein of The V3 Domain of HIV-1 Envelope Glycoprotein gp120, FEBS Letters, Vol.286, No.1,2, 233-236, 1991.
271.
N. Katunuma and Hiroshi Kido : Recent Advances in Research on Tryptases and Endogenous Tryptase Inhibitors, Monogr. Allergy: Neutral Proteases of Mast Cells, Vol.27, 51-66, 1990.
Hiroshi Kido, A. Fukutomi, J. Schilling, Y. Wang, B. Cordell and N. Katunuma : Protease-specificity of Kunitz Inhibitor Domain of Alzheimer's Disease Amyloid Protein Precursor, Biochemical and Biophysical Research Communications, Vol.167, No.2, 716-721, 1990.
(要約)
The putative inhibitor domain of Alzheimer's disease amyloid protein precursor was purified from E. coli containing a synthetic gene encoding the Kunitz domain. The purified protein (A4 inhibitor) inhibited the activity of trypsin, forming a 1:1 molar complex with the enzyme. It also strongly inhibited plasmin (Ki = 7.5 x 10(-11) M) from human serum and tryptase (Ki = 2.2 x 10(-10) M) from rat mast cells (tryptase M). In addition, it inhibited rat pancreatic trypsin, alpha-chymotrypsin and kallikrein and human serum kallikrein, but did not inhibit rat chymase, pancreatic elastase, alpha-thrombin, urokinase, papain or cathepsin B.
Hiroshi Kido and N. Katunuma : Biological Functions of Chymase and Tryptase and their Inhibitors in Rat Mast Cells, Intracellular Proteolysis: Mechanisms and Regulations, 109-119, 1989.
N. Fukusen, Hiroshi Kido, Y. Kato, K. Ishidoh and N. Katunuma : Diacylglycerols Enhance the Anti-tumor Effect of Glucocorticoid on L5178Y Lymphoblasts in Vivo, Japanese Journal of Cancer Research, Vol.80, No.10, 963-967, 1988.
(要約)
1,2-Dioleoyl-rac-glycerol, a potent activator of protein kinase C, was found to enhance the growth-inhibitory effect of triamcinolone acetonide on L5178Y lymphoblasts in adrenalectomized, male DBA/2 mice. On the other hand, in mice without adrenalectomy, it markedly inhibited tumor growth without increasing the plasma level of corticosterone or adrenocorticotropic hormone or reducing the body weight. These results suggest that diacylglycerol enhances the action of endogenous glucocorticoid to a sufficient level to inhibit the growth of lymphoblasts. Of various diacylglycerols with different carbon chain lengths tested, 1,2-dioleoyl-rac-glycerol was the most potent growth inhibitor and was maximally effective at a dose of above 30 micrograms/100 g body weight. This finding suggests that diacylglycerols may be useful for enhancing the antitumor effect of a low dose of glucocorticoid or endogenous glucocorticoid on lymphoblasts without any significant side effect.
T. Hattori, A. Koito, K. Takatsuki, Hiroshi Kido and N. Katunuma : Involvement of Tryptase-related Cellular Protease(s) in Human Immunodeficiency Virus Type 1 Infection, FEBS Letters, Vol.248, No.1,, 48-52, 1988.
283.
N. Katunuma and Hiroshi Kido : Biological Functions of Serine Proteases in Mast Cells in Allergic Inflammation, Journal of Cellular Biochemistry, Vol.38, No.4, 291-301, 1988.
(要約)
Serine proteases in mast cell granules, such as chymase, atypical chymase, and tryptase, which are major proteins in the granules, may play important roles in the process of immunoglobulin E (IgE)-mediated degranulation and in pathobiological alterations in tissues. Indeed, inhibitors of chymase, substrate analogs, and antichymase F(ab')2, but not inhibitors of tryptase, markedly inhibited histamine release induced by IgE-receptor bridging but not that induced by Ca ionophore. In contrast, inhibitors of metalloprotease inhibited histamine release induced not only by IgE-receptor bridging but also by Ca ionophore. These results suggest that chymase and metalloprotease are involved at different steps in the process of degranulation. The extents of inhibition of histamine release were closely correlated with the amounts of the inhibitors of chymase accumulated in the granules. After degranulation, the released proteases may in part contribute to pathobiological alterations in allergic disorders through generations of C3a anaphylatoxin and thrombin by human and rat tryptase, respectively, and those of angiotensin II and a chemotactic factor of neutrophils by human and rat chymase, respectively. Moreover, chymase and atypical chymase from rat were shown to destroy type IV collagen, and human tryptase was found to hydrolyze various plasma proteins, such as fibrinogen and high-molecular-weight kininogen. The biological activities of tryptase and chymase from rat may be regulated by their dissociation from and association with trypstatin, an endogenous inhibitor of these proteases.
Hiroshi Kido, Y. Yokogoshi and N. Katunuma : Kunitz-type Protease Inhibitor Found in Rat Mast Cell: Purification, Properties and Amino Acid Sequence, The Journal of Biological Chemistry, Vol.263, No.34, 18104-18108, 1988.
(要約)
A low molecular weight serine protease inhibitor, named trypstatin, was purified from rat peritoneal mast cells. It is a single polypeptide with 61 amino acid residues and an Mr of 6610. Trypstatin markedly inhibits blood coagulation factor Xa (Ki = 1.2 x 10(-10) M) and tryptase (Ki = 3.6 x 10(-10) M) from rat mast cells, which have activities that convert prothrombin to thrombin. It also inhibits porcine pancreatic trypsin (Ki = 1.4 x 10(-8) M) and chymase (Ki = 2.4 x 10(-8) M) from rat mast cells, but not papain, alpha-thrombin, or porcine pancreatic elastase. Trypstatin forms a complex in a molar ratio of 1:1 with trypsin and one subunit of tryptase. The complete amino acid sequence of this inhibitor was determined and compared with those of Kunitz-type inhibitors. Trypstatin has a high degree of sequence homology with human and bovine inter-alpha-trypsin inhibitors, A4(751) Alzheimer's disease amyloid protein precursor, and basic pancreatic trypsin inhibitor. However, unlike other known Kunitz-type protease inhibitors, it inhibits factor Xa most strongly.
N. Katunuma, Y. Kato and Hiroshi Kido : Studies on Biomodulators of Glucocorticoid Action: Amplifiers and Suppressors of Glucocorticoid Action, Advances in Enzyme Regulation, Vol.27, 277-286, 1988.
Yasuhisa Kato, Hiroshi Kido, Naomi Fukusen and Nobuhiko Katunuma : Peptide Boronic Acids, Substrate Analogs, Inhibit Chymase, and Histamine Release from Rat Mast Cells, The Journal of Biochemistry, Vol.103, No.5, 5, 1988.
(要約)
Peptide boronic acids, such as methoxysuccinyl-Ala-Ala-Pro-(L)boro-Phe-OH, its pinacol ester, and t-butyloxycarbonyl-Phe-Pro-(L)boro-Phe-pinacol, inhibited the activity of chymase from connective tissue mast cells approximately 40- to 80-fold more than atypical chymase from mucosal mast cells, and did not inhibit trypsin. Only peptide boronic acids containing "L" forms of boronic acids were inhibitory. The Ki values of these peptide boronic acids for chymase were in the 60-170 nM concentration range, like those of the natural inhibitors tested, but all the natural inhibitors tested except Eglin C and chymostatin inhibited both chymase and trypsin. Thus these peptide boronic acids should be useful for selective inhibition of chymase with less inhibitory activity for atypical chymase and without inhibition of trypsin. These peptide boronic acids markedly inhibited histamine release induced by anti-rat immunoglobulin E, suggesting that chymase in connective tissue mast cells plays some role in the process of histamine release. These peptides are assumed to be therapeutically useful for treatment of allergic inflammations catalyzed by chymase.
Hiroshi Kido, N. Fukusen and N. Katunuma : Antibodies and Inhibitions of Chymase are Incorporated into Mast Cell Granules and Inhibit Histamine Release, Biol. Chem. Hoppe-Seyler, Vol.369, 95-100, 1988.
288.
Hiroshi Kido, N. Fukusen and N. Katunuma : Epidermal Growth Factor as a New Regulator of Induction of Tyrosine Aminotransferase and Tryptophan Oxygenase by Glucocorticoid, FEBS Letters, Vol.223, No.2, 223-226, 1987.
289.
Hiroshi Kido, N. Fukusen and N. Katunuma : Tumor-promoting Phorbol Ester Amplifies the Inductions of Tyrosine Aminotransferase and Ornithine Decarboxylase by Glucocorticoid, Biochemistry, Vol.26, No.8, 2349-2353, 1987.
290.
Hiroshi Kido, N. Fukusen and N. Katunuma : Inhibition by 1-(5-Isoguinolinesulfonyl)-2-Methyl-piperazine, an Inhibitor of Protein Kinase C, of Enzyme Induction by Glucocorticoid and of Nuclear Translocation of Glucocorticoid-Receptor Complexes, Bichem. Biophys. Res. Commun., Vol.144, No.1, 152-159, 1987.
291.
N. Katunuma and Hiroshi Kido : Studies on Biomodulators of Glucocorticoid Actions: The Nature and the Modes of Actions of Glucocorticoid Potency Amplifiers, Advances in Enzyme Regulation, Vol.26, 191-208, 1987.
(要約)
We have found many compounds that amplify the action of glucocorticoid without themselves having any glucocorticoid-like action and have proposed the concept of 'Glucocorticoid Action Biomodulators'. These biomodulators consist of 'Glucorticoid Sensitivity Amplifiers', which greatly amplify the action of glucocorticoid at doses of glucocorticoid that alone have minimal effects, and 'Glucocorticoid Potency Amplifiers', which markedly enhance the effect of glucocorticoid at doses that have maximal effects. Potent activators of protein kinase C, such as 1,2-racemic dioctanoylglycerol, 12-o-tetradecanoyl-phorbol-13-acetate, and epidermal growth factor (EGF), markedly enhanced the induction of tyrosine aminotransferase and ornithine decarboxylase by dexamethasone in adrenalectomized rats in vivo and in primary cultures of adult rat hepatocytes in vitro. They amplified enzyme induction by even a large amount of dexamethasone that had a maximal effect, but had no effect in the absence of glucocorticoid. These modes of amplification show that these compounds are 'Glucocorticoid Potency Amplifiers'. They amplified not only enzyme induction in liver but also growth inhibition by glucocorticoid of solid tumor L5178Y lymphoblasts. They specifically amplified the actions of glucocorticoids and did not amplify the actions of other steroids, such as 17-beta estradiol, glucagon and insulin. The induction of tyrosine aminotransferase by glucocorticoid and its amplification by EGF were both inhibited by 1-(5-iso-quinoline-sulfonyl)-2-methylpiperazine, an inhibitor of protein kinase C, and not by N-[2-(methylamino)-ethyl]-5-isoquinoline-sulfonamide, an inhibitor of cyclic nucleotide dependent protein kinases, suggesting that the induction and the amplification are mediated by protein kinase C.
N. Fukusen, Y. Kato, Hiroshi Kido and N. Katunuma : Kinetic Studies on the Inhibitions of Mast Cell Chymase by Natural Serine Proteae Inhibitors. Indications for Potential Biological Functions of these Inhibitors, Biochem. Med. Metab. Biol., Vol.38, 165-169, 1987.
Hiroshi Kido, N. Fukusen, K. Ishidoh and N. Katunuma : Diacylglycerol Amplified the Induction in Vivo of Tyrosine Aminotransferase and Ornithine Decarboxylase by Glucocorticoid, Biochemical and Biophysical Research Communications, Vol.138, No.1, 275-282, 1986.
(要約)
In adrenalectomized rats, diacylglycerol, a potent activator of protein kinase C, specifically enhanced the induction of tyrosine aminotransferase and ornithine decarboxylase by even maximally effective doses of dexamethasone phosphate, but itself had no effect on these enzyme inductions in the absence of glucocorticoid. The amplifications of enzyme induction by diacylglycerol was dose-dependent and the time courses of the amplified inductions were similar to those of the inductions by dexamethasone phosphate alone. Since diacylglycerol did not affect the induction of these enzymes by glucagon and insulin, its amplifying effect seemed to be specific for induction by glucocorticoids.
Hiroshi Kido, K. Izumi, H. Otsuka, N. Fukusen, Y. Kato and N. Katunuma : A Chymotrypsin-type Serine Protease in Rat Basophilic Leukemia Cells: Evidence for its Immunological Identity with Atypical Mast Cell Protease, The Journal of Immunology, Vol.136, No.3, 1061-1065, 1986.
(要約)
Activity of a chymotrypsin-type serine protease was found in a subline of rat basophilic leukemia (RBL-2H3) cells. The protease was immunologically cross-reactive with anti-atypical mast cell protease immunoglobulin (Ig) G, and its activity was inhibited in a dose-dependent manner by the antibody. The apparent m.w. of the protease that reacted with the antibody was 25,000, which was identical with that of atypical mast cell protease in rat mucosal mast cells. These results show that the chymotrypsin type serine protease in RBL-2H3 cells is immunologically identical with atypical mast cell protease, which was first purified from rat small intestine. Immunohistochemical studies showed that the protease was located not only in intracytoplasmic granules but also in organelles synthesizing protein, such as cisternae of the rough endoplasmic reticulum, perinuclear spaces, and the Golgi apparatus. However, no immunoreactivity was demonstrated in rat basophils. The activity of the protease increased in the exponential phase of growth of RBL-2H3 cells in which some activity was also detected in the medium, and it decreased in the late stationary phase.
(キーワード)
Animals / Basophils / Cell Line / Chymases / Chymotrypsin / Cross Reactions / Endopeptidases / Histocytochemistry / Immunoglobulin G / Leukemia / Male / Mast Cells / Rats / Rats, Inbred Strains / Serine Endopeptidases / Staining and Labeling
(文献検索サイトへのリンク)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 2416825
N. Katunuma, N. Fukusen and Hiroshi Kido : Biological Functions of Serine Proteases in the Granules of Rat Mast Cells, Advances in Enzyme Regulation, Vol.25, 241-255, 1986.
(要約)
The effects of specific low- and high-molecular weight inhibitors of chymase and tryptase and F(ab')2 of antichymase on histamine release from activated mast cells were examined. The release of histamine induced by anti-rat immunoglobulin E was markedly inhibited by F(ab')2 fragments of antichymase and the low-molecular weight inhibitor of chymase chymostatin, whereas release of histamine induced by calcium ionophore A23187 was inhibited only by chymostatin. Neither the inhibitor nor the antibody affected histamine release induced by compound 48/80. These results suggest that two main chymotrypsin-type proteases are involved in process of degranulation: one is chymase, which acts at a step before calcium entry, and the other is an unidentified protease, which acts at a step after calcium entry. These results are summarized in Figure 8. After degranulation, released chymase remains associated with the cell surface while released tryptase was found in the extracellular milieu. Tryptase converted bovine prothrombin to thrombin, as shown by increase in thrombin activity with a synthetic substrate, t-butyloxy-carbonyl-Val-Pro-Arg-4-methyl-coumaryl-7-amide. The apparent Km value toward prothrombin was relatively low (2.3 microM), suggesting that tryptase contributes to blood coagulation or the process of fibrosis in tissues. The proteolytic products of IgG1 produced by chymase had chemotactic activity for neutrophil leukocytes in vitro and in vivo. These findings indicate the possible functions of these proteases after degranulation.
Hiroshi Kido, N. Fukusen, N. Katunum, T. Morita and S. Iwanaga : Tryptase from Rat Mast Cells Converts Bovine Prothrombin to Thrombin, Biochemical and Biophysical Research Communications, Vol.132, No.2, 613-619, 1985.
(要約)
The effect of tryptase purified from rat peritoneal mast cells on bovine prothrombin was examined. Tryptase activated prothrombin, as evidenced by the increase in thrombin activity with a synthetic substrate, t-butyloxy-carbonyl-Val-Pro-Arg-4-methylcoumaryl-7-amide. The apparent Km value toward bovine prothrombin and the kcat value were 2.3 microM and 46.3 s-1, respectively. Studies on the time course of prothrombin activation by tryptase and by activated factor X (Xa), and analysis of the activation products on sodium dodecyl sulfate gel electrophoresis showed that the process of activation of prothrombin by tryptase was similar to that by Xa except that an intermediate of 67,000 daltons was formed.
Hiroshi Kido, N. Fukusen and N. Katunuma : Antibody and Inhibitor of Chymase Inhibit Histamine Release in Immunoglobulin E-Activated Mast Cells, Biochemistry International, Vol.10, No.6, 863-871, 1985.
(要約)
The low-molecular-weight inhibitor of chymase, chymostatin, and F(ab')2 fragments of anti-chymase markedly inhibited histamine release induced by anti-rat immunoglobulin E (IgE) but not that induced by compound 48/80. Inhibitors with molecular weights of more than 6,000, such as alpha 1-antichymotrypsin and aprotinin, and non-immunized F(ab')2 had no effect on histamine release. These results suggest that chymase in mast cell granules plays an essential role in the process of IgE-mediated degranulation. After degranulation, released chymase was associated with the cell surface while released tryptase was present in the extracellular milieu as a complex with a protein associated with tryptase (trypstatin).
Hiroshi Kido, N. Fukusen and N. Katunuma : Chymotrypsin-type and Trypsin-type Serine Protease in Rat Mast Cells: Properties and Functions, Arch. Biochem. Biophys, Vol.239, No.2, 436-443, 1985.
300.
N. Fukusen, Hiroshi Kido and N. Katunuma : Inhibition of Chymase Activity by Phosphoglycerides, Arch. Biochem. Biophys., Vol.237, No.1, 118-123, 1985.
301.
Hiroshi Kido, N. Fukusen and N. Katunuma : Inhibition of Chymase Activity by Long Ghain Fatty Acids, Arch. Biochem. Biophys., Vol.230, No.2, 610-615, 1984.
302.
Hiroshi Kido, N. Fukusen and N. Katunuma : A Simple Method for Purification of Chymase from Rat Tongue and Rat Peritoneal Cells, Anal. Biochem., Vol.137, 449-453, 1984.
303.
Hiroshi Kido : Purification and Structure of a New Nucleotide from Proteus Mirabilis that Amplifies Induction of Tyrosine Aminotransferase by Glucocorticoid, Biochemical and Biophysical Research Communications, Vol.86, No.2, 3258-3261, 1979.
304.
Hiroshi Kido : Amplification of Growth Inhibition by Glucocorticoid on L5178Y and L1210 Lymphoblasts in Vivo, Cancer Research, Vol.39, 3258-3261, 1979.
305.
Hiroshi Kido : Amplification of the Cytotoxic Action of Glucocorticoid in Cultured L5178Y Lymphoblasts, Cancer Research, Vol.38, 3100-3103, 1978.
306.
Hiroshi Kido : A New Factor from Enteric Bacteria of Rats Amplifying Induction of Liver Enzyme by Glucocorticoid, European Journal of Biochemistry, Vol.78, 541-546, 1977.
307.
Hiroshi Kido : Establishment of a Clonal Strain of Hepatoma Cells Derived fromMorris Hepatoma 8999, Gann : Japanese Journal of Cancer Research, Vol.68, No.5, 691-697, 1977.
学術論文(紀要・その他):
1.
Kenta Horimukai, Kumiko Morita, Masami Narita, Mai Kondo, Hiroshi Kitazawa, Makoto Nozaki, Yukiko Shigematsu, Kazue Yoshida, Hironori Niizeki, Ken-Ichiro Motomura, Haruhiko Sago, Tetsuya Takimoto, Eisuke Inoue, Norio Kamemura, Hiroshi Kido, Junzo Hisatsune, Motoyuki Sugai, Hiroyuki Murota, Ichiro Katayama, Takashi Sasaki, Masayuki Amagai, Hideaki Morita, Akio Matsuda, Kenji Matsumoto, Hirohisa Saito and Yukihiro Ohya : Application of moisturizer to neonates prevents development of atopic dermatitis., The Journal of Allergy and Clinical Immunology, Vol.134, No.4, 824-830.e6, 2014.
(要約)
Daily application of moisturizer during the first 32 weeks of life reduces the risk of AD/eczema in infants. Allergic sensitization during this time period is associated with the presence of eczematous skin but not with moisturizer use.
Junji Chida and Hiroshi Kido : Extraction and quantification of adenosine triphosphate in mammalian tissues and cells., Methods in Molecular Biology, Vol.1098, 21-32, 2014.
(要約)
Adenosine 5'-triphosphate (ATP) is the "energy currency" of organisms and plays central roles in bioenergetics, whereby its level is used to evaluate cell viability, proliferation, death, and energy transmission. In this chapter, we describe an improved and efficient method for extraction of ATP from tissues and cells using phenol-based reagents. The chaotropic extraction reagents reported so far co-precipitate ATP with insoluble proteins during extraction and with salts during neutralization. In comparison, the phenol-based reagents extract ATP well without the risks of co-precipitation. The extracted ATP can be quantified by the luciferase assay or high-performance liquid chromatography.
Hiroshi Kido, Dai Mizuno, Tunetomo Takei and Wakako Sihihara : New effective nasal immunization of influenza vaccine by a natural mucosal adjuvant from the lung and its synthetic compound., Options for the Control of Influenza VI, 616-618, 2008.
学術レター:
1.
Norio Kamemura, Norio Kawamoto, Ryosuke Nakamura, Reiko Teshima, Toshiyuki Fukao and Hiroshi Kido : Low-affinity allergen-specific IgE in cord blood and affinity maturation after birth., The Journal of Allergy and Clinical Immunology, Vol.133, No.3, 904-5.e6, 2013.
Hiroshi Kido : The role of secreted serine proteases of the host in influenza viral pathogenesis. Activation of viruses by host proteases, Activation of Viruses by Host Proteases, 135-151, 2018.
木戸 博, 千田 淳司, Min Yao, Wang Siye : インフルエンザ感染の重症化，多臓器不全の発症メカニズム, 日本臨牀, Vol.68, No.8, 1565-1573, 2010年8月.
(要約)
Severe influenza is characterized by cytokine storm and multi-organ failure with edema. We found that the "influenza virus-cytokine-trypsin/MMP-9 cycle" in the endothelial cells is one of the key mechanisms of vascular hyperpermeability, the major pathogen of multi-organ failure. Upregulated TNF-alpha, IL-6 and IL-beta induce ectopic pancreatic trypsin and pro-MMP-9 in the endothelial cells and in various organs. Trypsin mediates the viral hemagglutinin processing, which is crucial for viral entry and multicycles of replication. In addition, trypsin is the most potent pro-MMP-9 convertase to form active MMP-9 and both proteases synergistically destruct matrix around blood vessels. In addition upregulated trypsin triggers through its receptor, PAR-2, the modification of cellular functions, such as increase in calcium mobilization and mitochondrial membrane permeability, suppression of ATP generation and loss of tight junction constituent, zonula occludens-1. High risk patients who have impaired mitochondrial fuel utilization will easy get into energy crisis, resulting in vascular hyperpermeability in severe influenza.
Hiroshi Yamada, Higashi Youichiro, Kousaka Aoi, Enomoto Tadashi and Hiroshi Kido : Diclofenac augments intracellular influenza virus replication, Options for the Control of Influenza VI, Vol.1, No.1, 633-635, 2008.
65.
Hiroshi Kido, Dai Mizuno, Tsunetomo Takei, Maki Nishino and Wakako Shinahara : New effective nasal immunizations of influenza vaccine by a natural mucosal adjuvant from the lung and its synthetic compound, Options for the Control of Influenza VI, Vol.1, No.1, 616-618, 2008.
kawamoto Norio, Norio Kamemura, Hiroshi Kido and Fukao Toshiyuki : Detection of Ovomucoid-Specific Low-Affinity IgE in 14-Month-Old Infants and Its Relationship with Eczema, American Academy of Allergy Asthma and Immunology, ロサンゼルス (アメリカ合衆国), Mar. 2016.
2.
Ohya Yukihiro, Horimukai Kenta, Morita Kumiko, Narita Masami, Niizeki Hironori, Inoue Eisuke, Norio Kamemura, Hiroshi Kido and Saito Hirohisa : A randomized, controlled intervention trial of early emollient use in prevention of atopic dermatitis and allergic sensitization during infancy, Collegium Internatuonale Allergologicum, Petersberg, Germany, 91, Sep. 2014.
3.
Mayumi Sugimoto, Norio Kamemura, Suzuki Koichi, Kubota Kenji, Nagao Mizuho, Fujisawa Takao, Shoji Kagami and Hiroshi Kido : Analysis of Allergen-Specific Immunoglobulin in Rush Oral Immunotherapy for Severe Food Allergy by a Highly Sensitive Allergen Microarray, European Academy of Allergy and Clinical Immunology Congress 2014, Jun. 2014.
4.
Hiroshi Kido : Critical illness and energy metabolism-Blood lactate/ATP ratio as a real-time energy alarm index-, The 7th Asian Conference on Emergency Medicine, Oct. 2013.
5.
Naoki Shimojo, Norio Kamemura, Morita Yoshinori, Tada Hitomi, Kubota Kenji, Mineyoshi Hiyoshi, Suzuki Koichi, Ichioka Takao, Kohno Yoichi and Hiroshi Kido : Allergen sensitization develops in utero: analysis of allergen-specific antibodies by a highly-sensitive new allergen microarray, European Academy of Allergy and Clinical Immunology Congress 2012, Jun. 2012.
6.
Yuushi Okumura, Etsuhisa Takahashi and Hiroshi Kido : TYPE II TRANSMEMBRANE SERINE PROTEASES MSPL AND TMPRSS13 PROTEOLYTICALLY ACTIVATE MEMBRANE FUSION ACTIVITY OF HIGHLY PATHOGENIC AVIAN INFLUENZA VIRUS AND INDUCE THEIR MULTICYCLE REPLICATION, 7th General Meeting of the International Proteolisis Society, San Diego, Oct. 2011.
7.
Etsuhisa Takahashi, HY Pan, IL Indalao, Junji Chida and Hiroshi Kido : TRYPSIN KOCKDOWN AND TRYPSIN INHIBITOR ADMINISTRATION SUPPRESSED INFLUENZA VIRAL REPLICATION AND VIRUS-INDUCES MYOCARDITIS IN THE HEARTS AND CARDIOMYOBLAST CELLS, 7th General Meeting of the International Proteolisis, San Diego, Oct. 2011.
8.
Dai Mizuno, Kimoto Takashi, Tunetomo Takei, Kunimi Takuya and Hiroshi Kido : Synthetic antigen vehicle SF-10 adjuvant, mimicking human pulmonary surfactant, for effective intranasal flu vaccine, International Union of Microbiological Societies 2011, Sep. 2011.
9.
Dai Mizuno, Kimoto Takashi, Tunetomo Takei, Kunimi Takuya and Hiroshi Kido : Sy thetic antigen vehicle SF-10 adjuvant, mimicking human pulmonaru surfactant, for effective flu vaccine,, International Union of Microbiological Societies 2011, Sep. 2011.
10.
Etsuhisa Takahashi, Yuushi Okumura and Hiroshi Kido : Type II membrane-bound proteases, MSPL and TMPRSS13, cleave hemagglutinin of highly pathogenic avian influenza viruses and induce their multicycle replication, IUMS 2011, 札幌コンベンションセンター, Sep. 2011.
11.
Hiroshi Kido, Junji Chida, Siye Wang, Hai-Yan Pan and Min Yao : Influenza-Cytokine-Protease Cycle in the pathogenesis of vascular hyperpermeability in severe influenza and its possible treatments, BMB2010, Dec. 2010.
12.
Yuushi Okumura, Etsuhisa Takahashi, Oto Takahiro, Ohuchi Masanobu, Daidoji Tomo, Nakaya Takaaki, Bottcher Eva, Garten Wolfgan, Klenk Hans-Dieter and Hiroshi Kido : Novel type transmembrane serine proteases, MSPL and TMPRSS13,proteolytically activate membrane fusion activity of hemagglutinin of highly pathogenic avian influenza viruses and induce their multicycle replication, Cell symposia influenza, Washington D.C., USA, Dec. 2010.
13.
Hiroshi Kido, Etsuhisa Takahashi, Kosuke Kataoka, K. Fujii, S. Suzuki and C. Ito : Attenuation of Respiratory Immune Responses by Antiviral Neuraminidase Inhibitor Treatment and Boost of Mucosal Immunoglobulin A Response by Coadministration of Immunomodulator Clarithromycin in Pediatric Imfluenza, Options fot the Control of Influenza, Hong Kong SAR, China, Sep. 2010.
14.
Hiroshi Kido, Kataoka Kousuke, Takahashi Etsuhisa, Fujii Kazuyuki and Suzuki Satoshi : Attenuation of respiratory immune responses by antiviral neuraminidase inhibitor treatment and c0-administration of immuno-modulator clarithromycin in paediatric influenza, Options for the Control of Influenza VII, Hong Kong, Sep. 2010.
15.
Etsuhisa Takahashi, Yuushi Okumura and Hiroshi Kido : Novel type II transmembrane serine proteases, MSPL and TMPRSS13, proteolytically activate membrane fusion activity of hemagglutinin of highly pathogenic avian influenza viruses and induce their multicycle replication, Options for the Control of Influenza VII, China, Hong Kong, Sep. 2010.
16.
Takashi Kimono, Dai Mizuno, Tunetomo Takei, Takuya Kunimi, Shinji Ono, Wakako Shinahara and Hiroshi Kido : Synthetic antigen vehicle SF-10 adjuvant, mimicking human pulmonary surfactant, for effective intranasal flu vaccine., Options for the Control of Influenza VII, China, Hong Kong, Sep. 2010.
17.
Junji Chida, Siye Wang, Haiyan Pan, Min Yao, Dengbing Yao, Kazuhiko Yamane and Hiroshi Kido : Influenza virus-cytokine-protease cycle and mitochondrial ATP depletion are the principal mechanisms of multi-organ failure in severe influenza and therapeutic approaches, Options for the Control of Influenza VII, China, Hong Kong, Sep. 2010.
18.
Junji Chida, Siye Wang, Hai-Yan Pan, Min Yao, Dengbing Yao, Kazuhiko Yamane and Hiroshi Kido : Influenza virus-cytokine-protease cycle and mitochondrial ATP depletion are the principal mechanisms of multi-organ failure in severe influenza and therapeutic approaches., Oprions for the Control of Influenza, Hong Kong, Sep. 2010.
19.
Fujimoto Chisa, Hiroshi Kido, Yamaguchi Miyoko, Matsunaga Atsushi, Sawada Ayako, Tanaka Takeshi and Noriaki Takeda : Changes in Levels of Nasal IgA and Serum IgG Antibodies against Influenza Virus Antigen in Patients with Natural Influenza Infection, The 7th International Symposium on Tonsils and Mucosal Barriers of the Upper Airways, Asahikawa, Jul. 2010.
20.
Youssouf Cisse, Siye Wang, Isao Inoue and Hiroshi Kido : Hyperthermia induced abnormal neuronal responses in the hippocampus of rat brain after influenza A virus infection., Neuroelectronics Research Flanders, (Mini-Symposium), Leuven, Belgium, Oct. 2009.
21.
Yuushi Okumura and Hiroshi Kido : Role of the host cellular processing proteases in influenza virus infection., 6th General Meeting of the International Proteolysis Society, Queensland, Australia, Oct. 2009.
22.
Etsuhisa Takahashi, Yuushi Okumura and Hiroshi Kido : Type Transmembrane Serine Proteases, MSPL and TMPRSS13, Proteolytically Activate Membrane Fusion Activity of Highly Pathogenic Avian Influenza Virus Hemagglutinin., 6th General Meeting of the International Proteolysis Society, Queensland, Australia, Oct. 2009.
23.
Siye Wang, Le Quang Trong, Hiroshi Yamada and Hiroshi Kido : Influenza Virus-TNF-α-Protease Cycle for Progression of Influenza Virus Infection., 6th General Meeting of the International Proteolysis Society, Queensland, Australia, Oct. 2009.
24.
Etsuhisa Takahashi, Yuushi Okumura and Hiroshi Kido : Type II Transmembrane Serine Proteases, MSPL and TMPRSS13, Proteolytically Activate Membrane Fusion Activity of Highly Pathogenic Avian Influenza Virus Hemagglutinin., 6th General Meeting of the International Proteolysis Society, Queensland, Australia, Oct. 2009.
25.
Hiroshi Kido, Dai Mizuno, Tunetomo Takei, Shinahara Wakako, Fukuda Akiho and Kimoto Takashi : Immune responses to nasal vaccination of HA vaccine witha new natural mucosal adjuvant, pulmonary surfactantmedicine Surfacten and its synthetic compound in miceand mini-pigs, Second Circular and Provisional Conference Programme for IVW 2009The Third International Conference onInfluenza Vaccines for the World, France, Apr. 2009.
26.
Hiroshi Kido, Yuushi Okumura, Etsuhisa Takahashi and 潘 海燕 : Novel Proteolytic Activation Protease of Highly Pathogenic Avian Influenza Viruses which cover wide strains, even for non-susceptible strains by Furin and PC5/6., BirdFlu2008, Oxford, Sep. 2008.
27.
Junji Chida, Siye Wang, Tadashi Enomoto, Yuushi Ohnishi and Hiroshi Kido : Up-regulations of ectopic pancreatic trypsin mRNAs by influenza virus infection and pathological role of them in multi-organ failure after virus infection., 5th International Proteolysis Society, Patras, Oct. 2007.
Junpei Imajo, Masatake Akutagawa, Yohsuke Kinouchi, Fumio Shichijo and Hiroshi Kido : Effect of Carnitine Ingestion on EEG, Proceedings of 2006 World Congress on Medical Physics and Biomedical Engineering, Seoul, Korea, August 27-September 1, 2006, Vol.1, 1137-1140, Seoul, Aug. 2006.
29.
Mineyoshi Hiyoshi, Hirokazu Uemura, Hideo Takeda, Hiroshi Kido and Kokichi Arisawa : Quantitative proteomic analysis of testes in treated and untreated mice with bisphenol A by MALDI-TOF/TOF, 20th IUBMB International Congress of Biochemistry and Molecular Biology and 11th FAOBMB Congress, Kyoto, Jun. 2006.
30.
Dai Mizuno, Mikiko Ide-Kurihara, Tomoko Ichinomiya, Itsuka Kubo, Tunetomo Takei, Maki Nisino, Takayuki Usuki and Hiroshi Kido : Modified pulmonary surfactant is a novel antigen vehicle for influenza virus antigen to nasal-associated lymphoid tissues., 20th International Congress of Biochemistry and Molecular Biology, Kyoto, Jun. 2006.
31.
Satoshi Yamamoto, Anna Tani, Ayako Nakajima, Dan Kinoshita, Yuka Kasai, Masahiko Maegawa, Masaharu Kamada, Yuji Okumura, Hiroshi Kido and Minoru Irahara : Development of an Expression Vector Encoding Sperm Protein,Designated RSMP-B,as an Immunocontrraceptive Vaccine, IX International Congress of Reproductive Immunology, Vol.52, No.Supplement 1, 55, Kanagawa, Oct. 2004.
32.
Masahiko Maegawa, Satoshi Yamamoto, Ayako Nakajima, Dan Kinoshita, Masayo Kanayama, Minoru Irahara, Masaharu Kamada, Yuji Okumura and Hiroshi Kido : Development of an Expression Vector Encoding Sperm Protein,Designated rSMP-B,as an Immunocontrraceptive, The 4th Conference of the Pacific Rim Society for Fertility and Sterility, Okinawa, Mar. 2004.
Chisa Fujimoto, Hiroshi Kido and Noriaki Takeda : Induction and maintenance of anti-influenza antigenspecific nasal secretory IgA levels and serum IgG revuls after influenza infection in adults, 第41回日本免疫学会学術集会, Dec. 2012.
21.
木葉 敬子, 高橋 悦久, 片岡 宏介, Indalao Lorinda Irene, 木戸 博 : Airway mucosal IgA which reduced by oseltamivir is improved by combination with Clarithromycin in mice infected with influenza A virus, 第85回日本生化学会大会, 2012年12月.
22.
高橋 悦久, 奥村 裕司, Indalao L Irene, 木葉 敬子, 木戸 博 : Novel type II transmembrane serine proteases, MSPL/TMPRSS13 knockout mice attenuates multicycle replication of highly pathogenic avian influenza viruses, 第85回日本生化学会大会, 2012年12月.
Dai Mizuno, Tunetomo Takei, Takashi Kimono, Wakako Shinahara, Takuya Kunimi, Shinjji Ono and Hiroshi Kido : Effects of synthetic mucosal adjuvant SF10 on local and systemic antibody production by intranasal influenza vaccination., 第33回日本分子生物学会年会 第83回 日本生化学会大会合同大会,, Dec. 2010.
45.
Min Yao, Junji Chida, Hiyan Pan, Siye Wang and Hiroshi Kido : Effect of Bezafibrate on mitochondrial energy crisis in the fibroblast of severe influenza-associated encephalopathy patients, 第33回日本分子生物学会年会 第83回 日本生化学会大会合同大会,, Dec. 2010.
46.
Junji Chida, Min Yao, Dengbing Yao, Miyoko Yamaguchi and Hiroshi Kido : Pathogenesis of impaired systemic energy metabolism by severe influenza virus infection Analysis using mouse models of defect in mitochondrial -oxidation of long-chain fatty acids, 第33回日本分子生物学会年会 第83回 日本生化学会大会合同大会,, Dec. 2010.
47.
Hiroshi Kido, Junji Chida, Siye Wang, Hiyan Pan and Min Yao : Influenza-Cytokine-Protease Cycle in the pathogenesis of vascular hyper-permeability in severe influenza and its possible treatment, 第33回日本分子生物学会年会 第83回 日本生化学会大会合同大会,, Dec. 2010.
高橋 悦久, 奥村 裕司, 大内 正信, Klenk Hans-Dieter, 中屋 隆明, 大道寺 智, 木戸 博 : Novel type II transmembrane serine proteases, MSPL and TMPRSS13, proteolytically activate membrane fusion activity of hemagglutinin of highly pathogenic avian influenza viruses and induce their multicycle replication, 第83回日本生化学会大会, 2010年12月.
50.
Yao Min, 高橋 悦久, Pan Haiyan, 千田 淳司, 木戸 博 : A Type II Transmembrane Serine Protease Serase-1B, a New Splice Variant of Polyserase-1/TMPRSS9, Plays a Role in Adipogenesis., 第83回日本生化学会大会, 2010年12月.
51.
Pan Hai-Yan, 千田 淳司, 木戸 博 : Up-regulation of ectopic trypsin in myocardium triggers acute myocarditis in severe influenza, 生化学, 2010年12月.
52.
Yao Min, 千田 淳司, Pan Hai-Yan, Wang Siye, 木戸 博 : Effect of Bezafibrate on mitochondrial energy crisis in the fibroblast of severe influenza-associated encephalopathy patients, 生化学, 2010年12月.
53.
千田 淳司, Yao Min, Yao Dengbing, Miyoko Yamaguchi, 木戸 博 : Pathogenesis of impaired systemic energy metabolism by severe influenza virus infection Analysis using mouse models of defect in mitochondrial -oxidation of long-chain fatty acids-, 生化学, 2010年12月.
54.
Hiroshi Kido, Junji Chida, Siye Wang, Hai-Yan Pan and Min Yao : Influenza-Cytokine-Protease Cycle in the pathogenesis of vascular hyper-permeability in severe influenza and its possible treatment., Seikagaku, Dec. 2010.
Tidiane Seck Ahmed Cisse, Masatake Akutagawa, Takahiro Emoto, Yohsuke Kinouchi and Hiroshi Kido : The effect of Carnitine on human brain, Journal of Shikoku-Section Joint Convention of the Institutes of Electrical and Related Engineers, 205, Sep. 2010.
57.
千田 淳司, Wang Siye, Yao Dengbing, Yao Min, Miyoko Yamaguchi, 木戸 博 : インフルエンザ脳症の発症メカニズムの酵素学的な解析, --- ウイルス罹患後に高熱で誘発されるミトコンドリアの長鎖脂肪酸代謝障害 ---, 温熱生理研究会, 2010年9月.
Yuuji Onishi, Junji Chida, Youssouf Cisse and Hiroshi Kido : Molecular mechanism of the decrease in ATP levels by influenza virus infection., 第81回 日本生化学会大会, Dec. 2008.
Min Yao, Dengbing Yao, Miyoko Yamaguchi, Junji Chida and Hiroshi Kido : Effect of bezafibrate on mitochondrial energy crisis in the fibroblasts of severe influenza-associated encephalopathy patients., 第81回 日本生化学会大会, Dec. 2008.
水野 大, 武井 恒知, 西野 真紀, Wakako Shinahara, Takashi Kimoto, Akiho Fukuda, 木戸 博 : SP-C is the essential component for the adjuvanticity of Surfacten., 第30回日本分子生物学学会年会・第80回生化学会大会, 2007年12月.
87.
Tadashi Enomoto, Hiroshi Yamada, Junji Chida, Siye Wang and Hiroshi Kido : Mechanism of augmentation of symptomatic of influenza virus infection by antipyretic, diclophenac., 第30回 日本分子生物学会，第80回 日本生化学会大会, Dec. 2007.
Junji Chida, Tadashi Enomoto, Yuushi Ohnishi, Mariam Nasreen and Hiroshi Kido : Down-regulation of mitochondrial related-genes by influenza virus infection and pathological role of them in multi-organ failure after virus infection., 第30回 日本分子生物学会年会，第80回 日本生化学会大会, Dec. 2007.
奥村 裕司, 端山 昌樹, 高橋 悦久, 島袋 陽, 木戸 博 : Serase-1, a new splice variant of Polyserase-1/TMPRSS9, is an activator of pro-urokinase and is involved in urokinase/plasmin-mediated proteolysis., 第78回日本生化学会大会, 2005年10月.
奥村 裕司, 端山 昌樹, 高橋 悦久, 藤内 美恵子, 木戸 博 : Serase-1, a new member of the type II transmembrane serine protease, is highly expressed in ciliated epithelial cells., 第77回日本生化学会大会, 2004年10月.
奥村 裕司, 藤内 美恵子, 高橋 悦久, 木戸 博 : TMPRSS7, a new member of the type II transmembrane serine protease, has kallikrein-like characteristics., 第76回日本生化学会大会, 2003年10月.