Tadashi Nakanishi, Kanako Takahashi, Kazumi Ozaki, Hideaki Nakae and Takashi Matsuo : Proceedings of the International Conference on Dentin/Pulp Complex 2001, --- An Immunohistologic Study on the Localization of Selected Bacteria and Chemokines in Human Deep-Carious Teeth ---, Quintessence Publishing CO.,Ltd., Tokyo, Jun. 2002.
学術論文(審査論文):
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Katsuhiro Mieda, Tadashi Nakanishi, Hitomi Kuramoto, Yoshitaka Hosokawa, Ikuko Hosokawa, Daisuke Takegawa and Keiichi Hosaka : Sudachitin reduces inflammatory mediator expression in toll-like receptor 2 ligand-stimulated human dental pulp cells, Cell Biochemistry and Biophysics, 83, 2, 2431-2439, 2025.
(要約)
Sudachitin, which is a polymethoxy flavonoid derived from the peer of Citrus sudachi, has several biological properties. However, the effect of sudachitin on human dental pulp cells (HDPCs) remains unclear. The aim of this study was to investigate whether sudachitin could decrease the expression of inflammatory mediators such as cytokines and prostaglandin in HDPCs stimulated with Pam3CSK4, a ligand for toll-like receptor (TLR) 2. HDPCs were pre-incubated with different concentrations of sudachitin (6.25, 12.5, 25, or 50 μM) and stimulated with Pam3CSK4 (100 ng/mL). The quantification of inflammatory cytokines (interleukin (IL)-6, IL-8, and C-X-C motif chemokine ligand (CXCL) 10) and prostaglandin E (PGE) were performed by enzyme-linked immunosorbent assay (ELISA). The expression of cyclooxygenase (COX)-2, a key enzyme for PGE formation, was analyzed by western blot. Moreover, the activations of cell signal pathways were examined by western blot analysis. Sudachitin suppressed IL-6, IL-8, CXCL10, and PGE production and COX-2 protein expression in Pam3CSK4-stimulated HDPCs. In addition, we revealed that nuclear factor-kappa B (NF-κB) and protein kinase B (Akt) pathways in the Pam3CSK4-stimulated HDPCs were inhibited by sudachitin treatment. These findings suggest that sudachitin can reduce inflammatory mediator production in HDPCs stimulated with TLR2 ligand by inhibiting NF-κB and Akt activations.
Tadashi Nakanishi, Katsuhiro Mieda, Hitomi Kuramoto and Daisuke Takegawa : Effect of interleukin-17A on inflammatory mediator production in interleukin-1β-stimulated human dental pulp fibroblasts, European Journal of Oral Sciences, 132, 5, e13019, 2024.
(要約)
In response to pro-inflammatory cytokines such as interleukin (IL)-1β, dental pulp fibroblasts produce various inflammatory mediators, including IL-6, IL-8, CC chemokine ligand 20 (CCL20), and CXC chemokine ligand 10 (CXCL10), leading to the progression of pulpitis. IL-17/IL-17A (IL-17A) is a pro-inflammatory cytokine secreted by T helper (Th) 17 cells following their recruitment to inflamed sites; however, the roles of IL-17A during pulpitis remain unclear. The purpose of this study was to investigate the effect of IL-17A on IL-6, IL-8, CCL20 and CXCL10 production by human dental pulp fibroblasts (HDPFs) in vitro. IL-17A at a concentration of 100 ng/ml induced the production of 10 times more IL-8 and 4 times more CXCL10, but not IL-6 and CCL20, compared to controls. Co-stimulation of HDPFs with IL-17A and IL-1β synergistically enhanced the production of IL-6, CCL20, IL-8 and CXCL10. IL-1β increased expression of IL-17 receptor/IL-17RA (IL-17R) on HDPFs. Moreover, the cell signal pathways of p38 mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) were more potently activated by simultaneous stimulation with IL-17A and IL-1β. These findings suggest that IL-17A participates in the progression of dental pulp inflammation through the enhanced production of inflammatory mediators in HDPFs.
Hitomi Kuramoto, Tadashi Nakanishi, Daisuke Takegawa, MIEDA Katsuhiro and Keiichi Hosaka : Caffeic Acid Phenethyl Ester Induces Vascular Endothelial Growth Factor Production and Inhibits CXCL10 Production in Human Dental Pulp Cells, Current Issues in Molecular Biology, 44, 11, 5691-5699, 2022.
(要約)
The survival rate of root non-vital teeth is lower than that of vital teeth. Therefore, to preserve the dental pulp is very important. The vascular endothelial growth factor (VEGF) is the most potent angiogenic factor involved in the vitality of dental pulp including reparative dentin formation. Caffeic acid phenethyl ester (CAPE) is a physiologically active substance of propolis and has some bioactivities such as anti-inflammatory effects. However, there are no reports on the effects of CAPE on dental pulp inflammation. In this study, we investigated the effects of CAPE on VEGF and inflammatory cytokine production in human dental pulp cells (HDPCs) to apply CAPE to an ideal dental pulp protective agent. We found that CAPE induced VEGF production from HDPCs. Moreover, CAPE induced the phosphorylation of p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinases (ERK), and stress-activated protein kinase/c-Jun N-terminal kinase (SAP/JNK) in HDPCs. Furthermore, CAPE inhibited C-X-C motif chemokine ligand 10 (CXCL10) production in Pam3CSK4- and tumor necrosis factor-alpha (TNF-α)-stimulated HDPCs. In conclusion, these results suggest that CAPE might be useful as a novel biological material for vital pulp therapy by exerting the effects of VEGF production and anti-inflammatory activities.
Hitomi Kuramoto, Kouji Hirao, Hiromichi Yumoto, Yuki Hosokawa, Tadashi Nakanishi, Daisuke Takegawa, Ayako Washio, Chiaki Kitamura and Takashi Matsuo : Caffeic acid phenethyl ester (CAPE) induces VEGF expression and production in rat odontoblastic cells, BioMed Research International, Article ID 5390720, 2019.
(要約)
Caffeic acid phenethyl ester (CAPE), the main component of propolis, has various biological activities including anti-inflammatory effect and wound healing promotion. Odontoblasts located in the outermost layer of dental pulp play crucial roles such as production of growth factors and formation of hard tissue termed reparative dentin in host defense against dental caries. In this study, we investigated the effects of CAPE on the upregulation of vascular endothelial growth factor (VEGF) and calcification activities of odontoblasts, leading to development of novel therapy for dental pulp inflammation caused by dental caries. CAPE significantly induced mRNA expression and production of VEGF in rat clonal odontoblast-like KN-3 cells cultured in normal medium or osteogenic induction medium. CAPE treatment enhanced nuclear factor-kappa B (NF-B) transcription factor activation, and furthermore, the specific inhibitor of NF-B significantly reduced VEGF production. The expression of VEGF receptor- (VEGFR-) 2, not VEGFR-1, was up regulated in KN-3 cells treated with CAPE. In addition, VEGF significantly increased mineralization activity in KN-3 cells. These findings suggest that CAPE might be useful as a novel biological material for the dental pulp conservative therapy.
Yuki Hosokawa, Kouji Hirao, Hiromichi Yumoto, Ayako Washio, Tadashi Nakanishi, Daisuke Takegawa, Chiaki Kitamura and Takashi Matsuo : Functional roles of NOD1 in odontoblasts on dental pulp innate immunity, BioMed Research International, 2016, 9325436, 2016.
(要約)
Caries-related pathogens are first recognized by odontoblasts and induce inflammatory events that develop to pulpitis. Generally, initial sensing of microbial pathogens is mediated by pattern recognition receptors, such as Toll-like receptor and nucleotide-binding oligomerization domain (NOD); however, little is known about NODs in odontoblasts. In this study, the levels of NODs expressed in rat odontoblastic cell line, KN-3, were assessed by flow cytometry and the levels of chemokines in NOD-specific ligand-stimulated KN-3 cells were analyzed by real-time PCR and ELISA. The signal transduction pathway activated with NOD-specific ligand was assessed by blocking assay with specific inhibitors and reporter assay. In KN-3 cells, the expression level of NOD1 was stronger than that of NOD2 and the production of chemokines, such as CINC-1, CINC-2, CCL20, and MCP-1, was upregulated by stimulation with NOD1-specific ligand, but not with NOD2-specific ligand. CINC-2 and CCL20 production by stimulation with NOD1-specific ligand was reduced by p38 MAPK and AP-1 signaling inhibitors. Furthermore, the reporter assay demonstrated AP-1 activation in NOD1-specific ligand-stimulated KN-3 cells. These findings indicated that NOD1 expressed in odontoblasts functions to upregulate the chemokines expression via p38-AP-1 signaling pathway and suggested that NOD1 may play important roles in the initiation and progression of pulpitis.
Tadashi Nakanishi, Kayo Mukai, Yoshitaka Hosokawa, Daisuke Takegawa and Takashi Matsuo : Catechins inhibit vascular endothelial growth factor production and cyclooxygenase-2 expression in human dental pulp cells, International Endodontic Journal, 48, 3, 277-282, 2015.
(要約)
To investigate the effect of catechins on vascular endothelial growth factor (VEGF) production and cyclooxygenase-2 (COX-2) expression in human dental pulp cells (HDPC) stimulated with bacteria-derived factors or pro-inflammatory cytokines. Morphologically fibroblastic cells established from explant cultures of healthy human dental pulp tissues were used as HDPC. HDPC pre-treated with catechins, epigallocatechin-3-gallate (EGCG) or epicatechin gallate (ECG), were exposed to lipopolysaccharide (LPS), peptidoglycan (PG), interlukin-1β (IL-1β) or tumour necrosis factor-α (TNF-α). VEGF production was examined by enzyme-linked immunosorbent assay, and COX-2 expression was assessed by immunoblot. EGCG and ECG significantly reduced LPS- or PG-mediated VEGF production in the HDPC in a dose-dependent manner. EGCG also prevented IL-1β-mediated VEGF production. Although TNF-α did not enhance VEGF production in the dental pulp cells, treatment of 20 μg mL(-1) of EGCG decreased the level of VEGF. In addition, the catechins attenuated COX-2 expression induced by LPS and IL-1β. The up-regulated VEGF and COX-2 expressions in the HDPC stimulated with these bacteria-derived factors or IL-1β were diminished by the treatment of EGCG and ECG. These findings suggest that the catechins may be beneficial as an anti-inflammatory tool of the treatment for pulpal inflammation.
Daisuke Takegawa, Tadashi Nakanishi, Kouji Hirao, Hiromichi Yumoto, Kanako Takahashi and Takashi Matsuo : Modulatory roles of interferon-γ through indoleamine 2, 3-dioxygenase induction in innate immune response of dental pulp cells., Journal of Endodontics, 40, 9, 1382-1387, 2014.
(要約)
Marked infiltration of inflammatory cells such as activated T cells producing interferon-γ (IFN-γ) is observed in severe pulpitis. However, the roles of IFN-γ in the innate immune response of dental pulp have not been reported. Indoleamine 2, 3-dioxygenase (IDO) is a regulator of immune responses, and the IDO expression is induced by IFN-γ in many cells whose expression in dental pulp is unknown. The purpose of this study was to determine the role of IFN-γ in the immune response through microbial pattern recognition receptors (PRRs) such as Toll-like receptors or nucleotide-binding oligomerization domain-like receptors on the production of proinflammatory cytokines such as CXCL10 and interleukin (IL)-6 and the expression of IDO in cultured human dental pulp cells (HDPCs). HDPCs were established from explant cultures of healthy pulp tissues. CXCL10 and IL-6 production was determined using enzyme-linked immunosorbent assay. Confirmation of IDO localization in dental pulp tissues was examined using immunohistochemistry. IDO expression in HDPCs was analyzed by immunoblot. IFN-γ significantly up-regulated CXCL10 and IL-6 production in the HDPCs stimulated with ligands for PRRs in a concentration-dependent manner. The expression of IDO was detected in inflamed pulp tissue. In addition, IFN-γ in combination with the PRR ligands enhanced IDO expression in HDPCs compared with IFN-γ alone. Moreover, CXCL10 production in IFN-γ-stimulated HDPCs was inhibited by an IDO inhibitor. This study showed the synergistic effects by IFN-γ on cytokine production and IDO expression in HDPCs, suggesting that IFN-γ may modulate the innate immune response of dental pulp.
Tadashi Nakanishi, Daisuke Takegawa, Kouji Hirao, Kanako Takahashi, Hiromichi Yumoto and Takashi Matsuo : Roles of dental pulp fibroblasts in the recognition of bacterium-related factors and subsequent development of pulpitis, Japanese Dental Science Review, 47, 2, 161-166, 2011.
Yoshitaka Hosokawa, Ikuko Hosokawa, Satoru Shindo, Kazumi Ozaki, Tadashi Nakanishi, Hideaki Nakae and Takashi Matsuo : Black tea polyphenol inhibits CXCL10 production in oncostatin M-stimulated human gingival fibroblasts., International Immunopharmacology, 11, 6, 670-674, 2011.
(要約)
CXC chemokine ligand 10 (CXCL10) plays an important role in the infiltration of Th1 cells and thus in the exacerbation of periodontal disease. Theaflavin-3,3'-digallate (TFDG), polyphenol in black tea, has some beneficial effects but the effect of TFDG on CXCL10 production from human gingival fibroblasts (HGFs) is uncertain. In this study, we investigated the mechanisms by which TFDG may inhibit oncostatin M (OSM)-induced CXCL10 production in human gingival fibroblasts. TFDG prevented OSM-mediated CXCL10 production by HGFs in a dose dependent manner. TFDG significantly inhibited OSM-induced phosphorylation of c-Jun N terminal kinase (JNK), protein kinase B (Akt) (Ser473) that are related to CXCL10 production from OSM-stimulated HGFs. In addition, TFDG suppressed OSM receptor (OSMR) β expression on HGFs. These data provide a novel mechanism where the black tea flavonoid, theaflavin, could provide direct benefits in periodontal disease.
Gilson C. N. Franco, Mikihito Kajiya, Tadashi Nakanishi, Kouji Ohta, Pedro L. Rosalen, Francisco C. Groppo, Cory W. O. Ernst, Janie L. Boyesen, John D. Bartlett, Philip Stashenko, Martin A. Taubman and Toshihisa Kawai : Inhibition of matrix metalloproteinase-9 activity by doxycycline ameliorates RANK ligand-induced osteoclast differentiation in vitro and in vivo., Experimental Cell Research, 317, 10, 1454-1464, 2011.
(要約)
Tetracycline antibiotics, including doxycycli (DOX), have been used to treat bone resorptive diseases, partially because of their activity to suppress osteoclastogenesis induced by receptor activator of nuclear factor kappa B ligand (RANKL). However, their precise inhibitory mechanism remains unclear. Therefore, the present study examined the effect of Dox on osteoclastogenesis signaling induced by RANKL, both in vitro and in vivo. Although Dox inhibited RANKL-induced osteoclastogenesis and down-modulated the mRNA expression of functional osteoclast markers, including tartrate-resistant acid phosphatase (TRAP) and cathepsin K, Dox neither affected RANKL-induced MAPKs phosphorylation nor NFATc1 gene expression in RAW264.7 murine monocytic cells. Gelatin zymography and Western blot analyses showed that Dox down-regulated the enzyme activity of RANKL-induced MMP-9, but without affecting its protein expression. Furthermore, MMP-9 enzyme inhibitor also attenuated both RANKL-induced osteoclastogenesis and up-regulation of TRAP and cathepsin K mRNA expression, indicating that MMP-9 enzyme action is engaged in the promotion of RANKL-induced osteoclastogenesis. Finally, Dox treatment abrogated RANKL-induced osteoclastogenesis and TRAP activity in mouse calvaria along with the suppression of MMP9 enzyme activity, again without affecting the expression of MMP9 protein. These findings suggested that Dox inhibits RANKL-induced osteoclastogenesis by its inhibitory effect on MMP-9 enzyme activity independent of the MAPK-NFATc1 signaling cascade.
(キーワード)
Acid Phosphatase / Animals / Anti-Bacterial Agents / Blotting, Western / Bone Resorption / Bone and Bones / Cathepsin K / Cell Differentiation / Cells, Cultured / Doxycycline / Isoenzymes / Male / Matrix Metalloproteinase 9 / Mice / Mice, Inbred BALB C / NFATC Transcription Factors / Osteoclasts / Phosphorylation / RANK Ligand / RNA, Messenger / Reverse Transcriptase Polymerase Chain Reaction / Skull
Yoshitaka Hosokawa, Ikuko Hosokawa, Kazumi Ozaki, Tadashi Nakanishi, Hideaki Nakae and Takashi Matsuo : Tea polyphenols inhibit IL-6 production in tumor necrosis factor superfamily 14-stimulated human gingival fibroblasts., Molecular Nutrition & Food Research, 54 Suppl 2, S151-8, 2010.
(要約)
IL-6 is well recognized to be a potent bone resorptive agent and thus in the development of periodontal disease. Epigallocatechin gallate (EGCG) and epicatechin gallate (ECG), the major catechins in green tea, and theaflavin-3,3'-digallate (TFDG), polyphenol in black tea, have multiple beneficial effects, but the effects of catechins and theaflavins on IL-6 production in human gingival fibroblasts (HGFs) are not known. In this study, we investigated the mechanisms by which EGCG, ECG, and TFDG inhibit tumor necrosis factor superfamily 14 (TNFSF14)-induced IL-6 production in HGFs. We detected TNFSF14 mRNA expression in human diseased periodontal tissues. TNFSF14 increased IL-6 production in HGFs in a concentration-dependent manner. EGCG, ECG, and TFDG prevented TNFSF14-mediated IL-6 production in HGFs. EGCG, ECG, and TFDG prevented TNFSF14-induced extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and nuclear factor-kappaB activation in HGFs. Inhibitors of ERK, JNK, and nuclear factor-kappaB decreased TNFSF14-induced IL-6 production. In addition, EGCG, ECG, and TFDG attenuated TNFSF14 receptor expression on HGFs. These data provide a novel mechanism through which the green tea and black tea polyphenols could be used to provide direct benefits in periodontal disease.
Yoshitaka Hosokawa, Ikuko Hosokawa, Kazumi Ozaki, Tadashi Nakanishi, Hideaki Nakae and Takashi Matsuo : Catechins inhibit CXCL10 production from oncostatin M-stimulated human gingival fibroblasts, The Journal of Nutritional Biochemistry, 21, 7, 659-664, 2010.
(要約)
CXC chemokine ligand 10 (CXCL10) plays a pivotal role in the recruitment of Th1 cells and, thus, in the development of periodontal disease. Epigallocatechin gallate (EGCG) and epicatechin gallate (ECG), the major catechins derived from green tea, have multiple beneficial effects, but the effects of catechins on CXCL10 production from human gingival fibroblasts (HGFs) is not known. In this study, we investigated the mechanisms by which EGCG and ECG inhibit oncostatin M (OSM)-induced CXCL10 production in HGFs. HGFs constitutively expressed glycoprotein 130 and OSM receptor beta (OSMR beta), which are OSM receptors. OSM increased CXCL10 production in a concentration-dependent manner. EGCG and ECG prevented OSM-mediated CXCL10 production by HGFs. Inhibitors of p38 mitogen-activated protein kinase, c-Jun N-terminal kinase (JNK), phosphatidylinositol-3-OH kinase and signal transducer and activator of transcription (STAT)3 decreased OSM-induced CXCL10 production. EGCG significantly prevented OSM-induced phosphorylation of JNK, Akt (Ser473) and STAT3 (Tyr705 and Ser727). ECG prevented phosphorylation of JNK and Akt (Ser473). In addition, EGCG and ECG attenuated OSMR beta expression on HGFs. These data provide a novel mechanism through which the green tea flavonoids, catechins, can provide direct benefits in periodontal disease.
Tadashi Nakanishi, Kayo Mukai, Hiromichi Yumoto, Kouji Hirao, Yoshitaka Hosokawa and Takashi Matsuo : Anti-inflammatory effect of catechin on cultured dental pulp cells affected by bacteria-derived factors, European Journal of Oral Sciences, 118, 2, 145-150, 2010.
(要約)
Catechins (bioactive polyphenols in green tea) are known to exhibit potent anti-inflammatory properties. However, the anti-inflammatory effects of catechins on inflamed dental pulp tissue are not known. In this study, we investigated the effect of epigallocatechin-3-gallate (EGCG) and epicatechin gallate (ECG), the major components of green tea catechins, on the expression of pro-inflammatory cytokines and adhesion molecules in human dental pulp cells stimulated with bacteria-derived factors such as lipopolysaccharide (LPS) and peptidoglycan (PG). The expression of interleukin (IL)-6 and of IL-8 was examined using the reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assays. The expression of intercellular adhesion molecule-1 (ICAM-1) and of vascular cell adhesion molecule-1 (VCAM-1) on dental pulp cells was analyzed using flow cytometry. The presence of EGCG and ECG significantly reduced, in a concentration-dependent manner, the expression of IL-6 and IL-8 in dental pulp cells exposed to LPS or PG. Increased expression of ICAM-1 and VCAM-1 on the dental pulp cells in response to bacterial components was also decreased by treatment with EGCG and ECG. These findings suggest that green tea catechins may prevent the exacerbation of pulpitis.
Yoshitaka Hosokawa, Ikuko Hosokawa, Kazumi Ozaki, Tadashi Nakanishi, Hideaki Nakae and Takashi Matsuo : Catechins Inhibit CCL20 Production in IL-17A-stimulated Human Gingival Fibroblasts, Cellular Physiology and Biochemistry, 24, 5-6, 391-396, 2009.
(要約)
CC chemokine ligand 20 (CCL20) plays a pivotal role in the recruitment of Th17 cells and thus in the development of periodontal disease. Epigallocatechin gallate (EGCG) and epicatechin gallate (ECG), the major catechins in green tea, have multiple beneficial effects, but the effects of catechins on CCL20 production in human gingival fibroblasts (HGFs) are not known. In this study, we investigated the mechanisms by which EGCG and ECG inhibit interleukin (IL)-17A-induced CCL20 production in human gingival fibroblasts. IL-17A increased CCL20 production in HGFs in a concentration-dependent manner. EGCG and ECG prevented IL-17A-mediated CCL20 production in HGFs. Inhibitors of p38 mitogen-activated protein kinase (MAPK) or extracellular signal-regulated kinase (ERK) decreased IL-17A-induced CCL20 production. EGCG and ECG prevented IL-17A-induced phosphorylation of p38 MAPK and ERK in HGFs. In addition, EGCG and ECG attenuated IL-17 receptor expression on HGFs. These data provide a novel mechanism through which the green tea flavonoids catechins could be used to provide direct benefits in periodontal disease.
Kouji Hirao, Hiromichi Yumoto, Kanako Takahashi, Kayo Mukai, Tadashi Nakanishi and Takashi Matsuo : Roles of TLR2, TLR4, NOD2, and NOD1 in pulp fibroblasts., Journal of Dental Research, 88, 8, 762-767, 2009.
(要約)
Pulp fibroblasts express various pro-inflammatory mediators leading to marked infiltration of inflammatory cells in the progression of pulpitis. We hypothesized that pulp fibroblasts play roles in the recognition of invaded caries-related bacteria and the subsequent innate immune responses. We found clear expressions of TLR2, NOD1, and NOD2 and a faint expression of TLR4 in human dental pulp fibroblasts (HDPF) by RT-PCR and flow cytometry. We also observed that various pro-inflammatory mediators, including cytokines, chemokines, adhesion molecules, prostaglandin E(2) and its key enzyme COX-2, not iNOS or caspase-1, were markedly up-regulated by stimulation with these TLR and NOD agonists. More over, the NOD2 agonist acted synergistically with the TLR2, not the TLR4, agonist to stimulate the production of pro-inflammatory mediators in HDPF. These findings indicate that TLR2, TLR4, NOD2, and NOD1 in HDPF are functional receptors, and NOD2 is a modulator of signals transmitted through TLR2 in pulpal immune responses, leading to progressive pulpitis.
Kanako Takahashi, Tadashi Nakanishi, Hiromichi Yumoto, Tomohiko Adachi and Takashi Matsuo : CCL20 production is induced in human dental pulp upon stimulation by Streptococcus mutans and pro-inflammatory cytokines, Oral Microbiology and Immunology, 23, 4, 320-327, 2008.
(要約)
Pulpitis is characterized by the marked infiltration of inflammatory cells in response to an invasion of caries-related bacteria. It is well known that chemokines regulate the trafficking of lymphocytes, and CC chemokine ligand 20 (CCL20) has been recently shown to play a crucial role in the recruitment of memory T cells and immature dendritic cells into inflammatory lesions. We previously reported that CCL20 was mainly expressed in microvascular endothelial cells and macrophages that accumulated in inflamed pulp tissues and that its specific receptor, CCR6, was expressed on infiltrated lymphocytes. However, the mechanism of CCL20 expression remains unclear. In this study, we investigated the expression of CCL20 in monocytes/macrophages, endothelial cells, and pulpal fibroblasts after stimulation with Streptococcus mutans, a representative of caries-related bacteria, or proinflammatory cytokines. CCL20 messenger RNA was detected by reverse transcription-polymerase chain reaction in inflamed pulp, but not in clinically normal pulp. By enzyme-linked immunosorbent assay, S. mutans induced a human monocytic cell line, differentiated macrophage-like THP-1 cells, and human umbilical vein endothelial cells (HUVEC) to produce an increased amount of CCL20. Lipoteichoic acid from S. mutans also elicited CCL20 production by HUVEC. Moreover, CCL20 production from pulpal fibroblasts was increased by stimulation with inetrleukin-1beta and tumor necrosis factor-alpha. Our results indicate that CCL20 expression is induced by stimulation with caries-related bacteria that have invaded deeply into the dentinal tubules as well as by proinflammatory cytokines in the inflamed pulpal lesions. It may be involved in the progression of pulpitis via accumulation of inflammatory cells.
Tomohiko Adachi, Tadashi Nakanishi, Hiromichi Yumoto, Kouji Hirao, Kanako Takahashi, Kayo Mukai, Hideaki Nakae and Takashi Matsuo : Caries-related bacteria and cytokines induce CXCL10 in dental pulp., Journal of Dental Research, 86, 12, 1217-1222, 2007.
(要約)
Marked infiltration of inflammatory cells, such as activated T-cells, is observed in the progression of pulpitis; however, little is known about the mechanism of their recruitment into pulpal lesions. It has been recently demonstrated that CXC chemokine ligand 10 (CXCL10) chemoattracts CXC chemokine receptor 3 (CXCR3)-positive activated T-cells. We therefore examined whether CXCL10 is involved in the pathogenesis of pulpitis. CXCL10 mRNA expression levels in clinically inflamed dental pulp were higher than those in healthy dental pulp. Immunostaining results revealed that CXCL10 was detected in macrophages, endothelial cells, and fibroblasts in inflamed dental pulp, and that CXCR3 expression was observed mainly on T-cells. Moreover, cultured dental pulp fibroblasts produced CXCL10 after stimulation with live caries-related bacteria, peptidoglycans, and pro-inflammatory cytokines. In contrast, heat-killed bacteria did not induce CXCL10 secretion. These findings suggest that CXCL10-CXCR3 may play an important role in the pulpal immune response to caries-related bacterial invasion. Abbreviations: CXCL10, CXC chemokine ligand 10; CXCR3, CXC chemokine receptor 3; IFN, interferon; FBS, fetal bovine serum; LTA, lipoteichoic acid; PGN, peptidoglycan; IL, interleukin; TNF, tumor necrosis factor; PBS, phosphate-buffered saline; ELISA, enzyme-linked immunosorbent assay; CCL, C-C chemokine ligand; TLR, Toll-like receptor; NOD, nucleotide oligomerization domain; HDPF, human dental pulp fibroblasts.
Cory W. Ernst, Jang Eun Lee, Tadashi Nakanishi, Nadeem Y. Karimbux, Taia M. Rezende, Philip Stashenko, Makoto Seki, Martin A Taubman and Toshihisa Kawai : Diminished forkhead box P3/CD25 double-positive T regulatory cells are associated with the increased nuclear factor-kB ligand (RANKL(+)) T cells in bone resorption lesion of periodontal disease, Clinical and Experimental Immunology, 148, 2, 271-280, 2007.
(要約)
Periodontal disease involves multi-bacterial infections accompanied by inflammatory bone resorption lesions. The abundant T and B lymphocyte infiltrates are the major sources of the osteoclast differentiation factor, receptor activator for nuclear factor-kappaB ligand (RANKL) which, in turn, contributes to the development of bone resorption in periodontal disease. In the present study, we found that the concentrations of RANKL and regulatory T cell (T(reg))-associated cytokine, interleukin (IL)-10, in the periodontal tissue homogenates were correlated negatively, whereas RANKL and proinflammatory cytokine, IL-1beta, showed positive correlation. Also, according to the fluorescent-immunohistochemistry, the frequency of forkhead box P3 (FoxP3)/CD25 double-positive cells was diminished strikingly in the bone resorption lesion of periodontal disease compared to healthy gingival tissue, while CD25 or FoxP3 single positive cells were still observed in lesions where abundant RANKL+ lymphocytes were present. Very importantly, few or no expressions of FoxP3 by the RANKL+ lymphocytes were observed in the diseased periodontal tissues. Finally, IL-10 suppressed both soluble RANKL (sRANKL) and membrane RANKL (mRANKL) expression by peripheral blood mononuclear cells (PBMC) activated in vitro in a bacterial antigen-specific manner. Taken together, these results suggested that FoxP3/CD25 double-positive T(reg) cells may play a role in the down-regulation of RANKL expression by activated lymphocytes in periodontal diseased tissues. This leads to the conclusion that the phenomenon of diminished CD25+FoxP3+ T(reg) cells appears to be associated with the increased RANKL+ T cells in the bone resorption lesion of periodontal disease.
Yoshitaka Hosokawa, Tadashi Nakanishi, Daisuke Yamaguchi, Hideaki Nakae and Takashi Matsuo : Expression of fractalkine (CX3CL1) and its receptor, CX3CR1, in periodontal diseased tissue, Clinical and Experimental Immunology, 139, 3, 506-512, 2005.
(要約)
The regulatory role of chemokines and chemokine receptors on specific leucocyte recruitment into periodontal diseased tissue is poorly characterized. We observed that leucocytes infiltrating inflamed gingival tissue expressed marked levels of CX3CR1. In periodontal diseased tissue, the expression of fractalkine and CX3CR1 mRNA was detected by reverse transcription-polymerase chain reaction (RT-PCR) and further, fractalkine was distributed mainly on endothelial cells, as shown by immunohistochemistry. Moreover, we can detect CX3CR1-expressing cells infiltrated in periodontal diseased tissue by immunohistochemical staining. Furthermore, fractalkine production by human umbilical vein endothelial cells (HUVEC) was up-regulated by pathogen-associated molecular patterns (PAMPs), including Porphyromonas gingivalis lipopolysaccharide (LPS). Thus, these findings suggested that CX3CR1 and the corresponding chemokine, fractalkine may have an important regulatory role on specific leucocyte migration into inflamed periodontal tissue.
Tadashi Nakanishi, Kanako Takahashi, Yoshitaka Hosokawa, Tomohiko Adachi, Hideaki Nakae and Takashi Matsuo : Expression of macrophage inflammatory protein 3alpha in human inflamed dental pulp tissue, Journal of Endodontics, 31, 2, 84-87, 2005.
(要約)
Severe pulpitis resulting from dental caries is characterized by marked inflammatory infiltrate such as lymphocytes. Little is known about the recruitment of these cells into the dental pulp lesions of carious teeth. Macrophage inflammatory protein-3alpha (MIP-3alpha), a CC chemokine attracts CC chemokine receptor 6 (CCR6)-expressing T cells. We examined the distribution of MIP-3alpha-positive and/or CCR6-positive cells in human inflamed and normal dental pulp by immunohistochemistry. MIP-3alpha was observed in all inflamed pulp sections, and was mostly distributed in macrophages that had accumulated in the area adjacent to carious lesions. Furthermore, CCR6 expression was also observed in the infiltrating lymphocytes. In contrast, MIP-3alpha and CCR6 were rarely detected in normal pulp. These findings suggest that MIP-3alpha plays a role in the advancement of pulpal inflammation via the recruitment of CCR6-expressing lymphocytes.
Takashi Matsuo, Toshiyuki Shirakami, Kazumi Ozaki, Tadashi Nakanishi, Hiromichi Yumoto and Shigeyuki Ebisu : An immunohistological study of the localization of bacteria invading root pulpal walls of teeth with periapical lesions, Journal of Endodontics, 29, 3, 194-200, 2003.
(要約)
We immunohistologically examined the prevalence and localization of bacteria invading dentinal tubules of the roots of teeth with infected canals. Forty extracted teeth with apical lesions were selected and divided into two groups: a group of untreated teeth and a group of canal-enlarged teeth. The bacteria in the specimens were detected by Brown-Brenn stain and the labeled-streptavidin-biotin method with specific antisera for 16-bacteria. Seventy percent of the examined teeth showed bacteria invading the dentinal tubules of the roots. Fusobacterium nucleatum, Eubacterium alactolyticum, E. nodatum, Lactobacillus casei, and Peptostreptococcus micros were abundant. Even in the canal-enlarged group, invasion of bacteria was observed in 65% of teeth. This study revealed the actual condition of bacteria in infected root dentin and suggested that the canal-enlargement procedure could not completely remove all the bacteria in the infected dentinal tubules of the root.
Yoshitaka Hosokawa, Tadashi Nakanishi, Daisuke Yamaguchi, Kanako Takahashi, Hiromichi Yumoto, Kazumi Ozaki and Takashi Matsuo : Macrophage Inflammatory Protein 3α-CC chemokine receptor 6 interactions play an important role in CD4+ T-cell accumulation in periodontal diseased tissue., Clinical and Experimental Immunology, 128, 3, 548-554, 2002.
(要約)
The regulatory role of chemokines and chemokine receptors on specific lymphocyte recruitment into periodontal diseased tissue is poorly characterized. We observed that lymphocytes infiltrating inflamed gingival tissue expressed marked levels of CCR6. In periodontal diseased tissue, the expression of MIP-3alpha mRNA was detected by RT-PCR and further, MIP-3alpha was distributed in the basal layer of gingival epithelial cells, microvascular endothelial cells and the areas of inflammatory cells as shown by immunohistochemistry. Moreover, CCR6-expressing cells infiltrated into periodontal diseased tissue, and the proportion of CCR6-positive CD4+ T cells was significantly elevated in periodontal diseased tissue compared with peripheral blood in the same patients. Furthermore, gingival lymphocytes isolated from patients showed migration toward MIP-3alpha in an in vitro chemotaxis assay in which migration was abrogated by specific antibody to CCR6. Thus, these findings suggested that CCR6 and the corresponding chemokine, MIP-3alpha may have an important regulatory role in specific lymphocyte migration into inflamed periodontal tissue.
Tadashi Nakanishi, Hirotoshi Shimizu, Yoshitaka Hosokawa and Takashi Matsuo : An immunohistological study on cyclooxygenase-2 in human dental pulp., Journal of Endodontics, 27, 6, 385-388, 2001.
(要約)
Characteristics of cyclooxygenase-2 (COX-2) expressing cells in human dental pulp were immunohistologically studied. Extirpated pulpal tissues from extracted teeth were examined to elucidate the localization and distribution of COX-2. Pulpal tissues were examined by the labeled streptavidin biotin method using specific mouse monoclonal antibodies for COX-2. Cell types of the COX-2 expressing cells were also investigated by the double stain technique using both monoclonal antibodies for CD68/macrophage and anti-COX-2. COX-2 expressing cells could be found in all of the inflamed pulps, and these cells were mostly distributed close to the area of accumulation of inflammatory cells. COX-2 was mainly expressed in fibroblasts rather than macrophages. In contrast, COX-2 expressing cells were scarcely found in the normal pulps. These findings indicate that pulpal fibroblasts, as well as macrophages, may participate in the production of prostaglandin through COX-2 expression in pulpal inflammation, and might be involved in the pathogenesis of irreversible pulpitis.
Takashi Matsuo, Tadashi Nakanishi, Hirotoshi Shimizu and Shigeyuki Ebisu : A clinical study of direct pulp capping applied to carious-exposed pulps, Journal of Endodontics, 22, 10, 551-556, 1996.
Takashi Matsuo, Tadashi Nakanishi and Shigeyuki Ebisu : Immunoglobulins in periapical exudates of infected root canals: correlations with the clinical findings of the involved teeth, Endodontics & dental traumatology, 11, 2, 95-99, 1995.
(要約)
This study quantified the concentrations of IgG and IgA in the periapical exudates obtained from 69 single-rooted teeth using enzyme-linked immunosorbent assays, and examined their correlation with clinical findings of the involved teeth. Changes in the levels of these immunoglobulins during root canal treatment were also investigated. The average levels of IgG (85.61 mg/ml) was significantly higher than that of IgA (0.65 mg/ml) (P < 0.001). The exudates from the canals with large lesions (> or = 1 cm in diameter on radiographs) showed significantly higher IgG and IgA levels than small lesions (< 1 cm in diameter) (P < 0.01). The tendency for these to be a decrease in the levels of IgG and IgA was observed following the root canal treatment. These observations suggest that IgG and IgA are involved in the immunopathogenesis of periapical lesions.
(キーワード)
Bacterial Infections / Enzyme-Linked Immunosorbent Assay / Exudates and Transudates / Humans / Immunoglobulin A / Immunoglobulin G / Periapical Periodontitis / Pulpitis / Radiography / Root Canal Therapy / Suppuration / Time Factors / Toothache
Tadashi Nakanishi, Takashi Matsuo and Shigeyuki Ebisu : Quantitative analysis of immunoglobulins and inflammatory factors in human pulpal blood from exposed pulps, Journal of Endodontics, 21, 3, 131-136, 1995.
(要約)
The levels of immunoglobulins (IgG, IgA, and IgM) and inflammatory factors (elastase, prostaglandin E2, interleukin-1 alpha, interleukin-1 beta, interleukin-6, and tumor necrosis factor-alpha) in the blood of human dental pulp were quantified, using enzyme-linked immunosorbent assays. The pulpal blood samples were obtained with nylon fiber pellets from normal and inflamed dental pulp at pulp sites exposed on pulpectomy. Statistically significant differences between normal and inflamed pulp were found in the levels of IgG (p < 0.01), IgA (p < 0.05), IgM (p < 0.01), elastase (p < 0.05), and prostaglandin E2 (p < 0.01). These findings indicate that these factors play an important role in the pathogenesis of pulpal disease, and the sampling methods used in this study are useful for examination of pulpal inflammation.
松尾 敬志, 中西 正, 湯本 浩通, 平尾 功治, 篠原 千尋, Toshihiko Tominaga : 感染根管治療の考え方, Journal of Oral Health and Biosciences, 28, 2, 87-92, 2016年.
(要約)
It is a well-established fact that bacterial infection of the dental pulp ultimately results in the formation of apical periodontitis. And apical periodontitis may develop periapical leisons consisting granulomas and cysts. In this article, we show the pathologic condition of infected root canal systems and periapical lesions immunohistologically. We also describe the possible causes of refractory apical periodontitis that conventional root canal therapy could not work effectively. Finally,we explain our newly developing treatment for refractory apical periodontitis named electro-magnetic apical treatment, and illustrate clinical efficacy of this treatment.
Hiromichi Yumoto, Kouji Hirao, Yuki Hosokawa, Hitomi Kuramoto, Daisuke Takegawa, Tadashi Nakanishi and Takashi Matsuo : The Roles of Odontoblasts in Dental Pulp Innate Immunity, Japanese Dental Science Review, 54, 3, 105-117, Aug. 2018.
(要約)
Odontoblasts located in the outermost layer of dental pulp form a natural barrier between mineralized tissues, dentin, and soft tissues, dental pulp, of the vital tooth, and they first recognize caries-related pathogens and sense external irritations. Therefore, odontoblasts possess a specialized innate immune system to fight oral pathogens invading into dentin. Generally, the rapid initial sensing of microbial pathogens, especially pathogen-associated molecular patterns (PAMPs) shared by microorganisms, are mediated by pattern recognition receptors (PRRs), such as Toll-like receptor and the nucleotide-binding oligomerization domain (NOD). The innate immune responses in odontoblasts initiated by sensing oral pathogens provide host protective events, such as inflammatory reactions, to produce a variety of pro-inflammatory mediators, including chemokines and cytokines. These attract various inflammatory cells and cause antibacterial reactions, such as the production of defensins, to kill microorganisms in the proximal region of the odontoblast layer. This review focuses on innate immunity, especially cellular and molecular mechanisms regarding the sensing of PAMPs from oral pathogens by PRRs, in odontoblasts and provides information for future studies for the development of novel therapeutic strategies, including diagnosis and treatment, to prevent exceeding dental pulp inflammation and preserve the dental pulp tissues.
Tadashi Nakanishi : Dental Pulp Responses to Dental Caries, Dentisphere 4; International Scientific Meeting 5th ASEAN Plus Tokushima Joint International Conference, Nov. 2019.
2.
Yuki Hosokawa, Hiromichi Yumoto, Kouji Hirao, Hitomi Kuramoto, Tadashi Nakanishi, Daisuke Takegawa and Takashi Matsuo : Anti-inflammatory effects of polyphenols on rat odontoblastic cells, 95th General Session & Exhibition of the International Association for Dental Research (San Francisco), Mar. 2017.
3.
Daisuke Takegawa, Tadashi Nakanishi, Kouji Hirao, Hiromichi Yumoto, Yuki Hosokawa and Takashi Matsuo : Interferon-gamma modulates the innate immune response in odontoblast-like cells, 95th General session and exhibition of the IADR, Mar. 2017.
4.
Tadashi Nakanishi, Daisuke Takegawa, Kouji Hirao, Hiromichi Yumoto, Yuki Hosokawa and Takashi Matsuo : Effect of interleukin-17A on CCL20 production from odontoblast-like cells, 95th General session and exhibition of the IADR, Mar. 2017.
5.
Tadashi Nakanishi, Daisuke Takegawa, Hiromichi Yumoto, Kouji Hirao and Takashi Matsuo : Interleukin-17 Regulates Inflammatory Mediator Expression in IL-1beta-stimulated Dental Pulp Cells, 93th General Session & Exhibition of the International Association for Dental Research, Boston (USA), Mar. 2015.
6.
Tadashi Nakanishi, Daisuke Takegawa, Kouji Hirao, Hiromichi Yumoto, Kanako Takahashi and Takashi Matsuo : Prostaglandin F2a Regulates Cytokine Expression in Dental Pulp Cells, 91th General Session & Exhibition of the IADR, Seattle, USA, Mar. 2013.
7.
Daisuke Takegawa, Tadashi Nakanishi, Kouji Hirao, Hiromichi Yumoto, Kanako Takahashi and Takashi Matsuo : Interferon-g modulates the innate immune response of dental pulp cells, 91th General Session & Exhibition of the IADR, Seattle, USA, Mar. 2013.
8.
Tadashi Nakanishi, Kouji Hirao, Takegawa Daisuke, Hiromichi Yumoto, Kanako Takahashi and Takashi Matsuo : Catechins suppress cyclooxygenase-2 expression in human dental pulp cells, 88th General Session and Exhibition of the IADR, Jul. 2010.
9.
Takegawa Daisuke, Tadashi Nakanishi, Kouji Hirao, Hiromichi Yumoto, Kanako Takahashi and Takashi Matsuo : Interferon-gamma enhances toll-like receptor ligand-induced cytokine production in pulpal cells, 88th General Session and Exhibition of the IADR, Jul. 2010.
10.
Kayo Mukai, Tadashi Nakanishi, Hiromichi Yumoto, Kanako Takahashi, Kouji Hirao and Takashi Matsuo : Effects of Catechins on Cytokine Production in Human Odontoblast-like Cells, 86th General Session and Exhibition of the International Association for Dental Research, Jul. 2008.
11.
Kouji Hirao, Hiromichi Yumoto, Tadashi Nakanishi, Kanako Takahashi, Kayo Mukai, Miho Yoneda and Takashi Matsuo : Functional Analysis of Pattern Recognition Receptor Expression in Pulpal Cells, 86th General Session and Exhibition of the International Association for Dental Research, Jul. 2008.
12.
Kayo Mukai, Tadashi Nakanishi, Yoshitaka Hosokawa, Kanako Takahashi and Takashi Matsuo : Inhibitory effects of catechins on inflammatory reactions in pulp fibroblasts, 84th General Session & Exhibition of the IADR, Brisbane, Jun. 2006.
13.
Tadashi Nakanishi, Jean Eastcott, Martin A Taubman and Toshihisa Kawai : Characteristics of regulatory dendritic cells induced by gingival epithelial cells, 35th Annual Meeting & Exhibition of the AADR, Orlando, Mar. 2006.
14.
Tadashi Nakanishi, Jean Eastcott, Martin A Taubman and Toshihisa Kawai : Induction of regulatory dendritic cells by gingival epithelial cells, 83rd General session of IADR (International Association for Dental Research), Baltimore, Mar. 2005.
15.
Tadashi Nakanishi, Tomohiko Adachi, Kanako Takahashi and Takashi Matsuo : Expression of interferon-gamma -inducible protein 10/CXCL10 in inflamed dental pulp, 82th General Session of the IADR(INternational Association of Dental Research), Honolulu, Mar. 2004.
16.
Kanako Takahashi, Tadashi Nakanishi, Tomohiko Adachi, Natsuko Ida, Hideaki Nakae and Takashi Matsuo : Effects of Streptococcus mutans on MIP-3 α production by macrophages, 82th General Session of the IADR(INternational Association of Dental Research), Honolulu, Mar. 2004.
17.
Daisuke Yamaguchi, Tadashi Nakanishi, Kanako Takahashi, Hideaki Nakae and Takashi Matsuo : Expression of macrophage inflammatory protein-3alpha/CCL20 in human gingival epithelial cells, 81th General Session of the IADR(INternational Association of Dental Research), エルサレム, Jun. 2003.
18.
Yoshitaka Hosokawa, Tadashi Nakanishi, Daisuke Yamaguchi, Hiromichi Yumoto and Takashi Matsuo : LARC-CCR6 Interactions play an important role in CD4+ T cell accumulation in periodontal diseased tissue, 80th General Session and Exhibition of the International Association for Dental Research, San Diego(USA), Mar. 2002.
19.
Tadashi Nakanishi, Kanako Takahashi, Yoshitaka Hosokawa, Daisuke Yamaguchi, Chihiro Shinohara, Hiromichi Yumoto and Takashi Matsuo : Expression of macrophage inflammatory protein-3 α in human pulpitis, 80th General SEssion ofthe IADR(International Association of Dental Research), San Diego(USA), Mar. 2002.
20.
Tadashi Nakanishi, Kanako Takahashi, Kazumi Ozaki, Hideaki Nakae and Takashi Matsuo : An immunohistological study on the localization of selected bacteria and chemokines in human deep-carious teeth, The International Conference on Dentin/Pulp complexs, Chiba, Jul. 2001.
21.
Hiromichi Yumoto, Daisuke Yamaguchi, Yoshitaka Hosokawa, Chihiro Shinohara, Tadashi Nakanishi, Hirotoshi Shimizu, Masayoshi Yamada, Hideaki Nakae and Takashi Matsuo : Expression of IL-18 and IL-18 receptor in human periodontally diseased tissue, Modern Periodontology New Directions in 21st Century-, Kanagawa(Japan), Jun. 2001.