Linze XIA, Fumiya Kano, Noboru Hashimoto, Cheng DING, Yang XU, Hideharu HIBI, Tomonori Iwasaki, Eiji Tanaka and Akihito Yamamoto : Conditioned Medium from Stem Cells of Human Exfoliated Deciduous Teeth Partially Alters the Expression of Inflammation-associated Molecules of Mouse Condylar Chondrocytes via Secreted Frizzled-related Protein 1, Journal of Oral Health and Biosciences, Vol.35, No.2, 52-60, 2023.
Fumiya Kano, Noboru Hashimoto, Yao Liu, Linze Xia, Takaaki Nishihara, Wakana Oki, Keita Kawarabayashi, Noriko Mizusawa, Keiko Aota, Takayoshi Sakai, Masayuki Azuma, Hideharu Hibi, Tomonori Iwasaki, Tsutomu Iwamoto, Nobuyasu Horimai and Akihito Yamamoto : Therapeutic benefits of factors derived from stem cells from human exfoliated deciduous teeth for radiation-induced mouse xerostomia, Scientific Reports, Vol.13, No.1, 2706-2719, 2023.
(Summary)
Radiation therapy for head and neck cancers is frequently associated with adverse effects on the surrounding normal tissue. Irreversible damage to radiation-sensitive acinar cells in the salivary gland (SG) causes severe radiation-induced xerostomia (RIX). Currently, there are no effective drugs for treating RIX. We investigated the efficacy of treatment with conditioned medium derived from stem cells from human exfoliated deciduous teeth (SHED-CM) in a mouse RIX model. Intravenous administration of SHED-CM, but not fibroblast-CM (Fibro-CM), prevented radiation-induced cutaneous ulcer formation (p < 0.0001) and maintained SG function (p < 0.0001). SHED-CM treatment enhanced the expression of multiple antioxidant genes in mouse RIX and human acinar cells and strongly suppressed radiation-induced oxidative stress. The therapeutic effects of SHED-CM were abolished by the superoxide dismutase inhibitor diethyldithiocarbamate (p < 0.0001). Notably, quantitative liquid chromatography-tandem mass spectrometry shotgun proteomics of SHED-CM and Fibro-CM identified eight proteins activating the endogenous antioxidant system, which were more abundant in SHED-CM than in Fibro-CM (p < 0.0001). Neutralizing antibodies against those activators reduced antioxidant activity of SHED-CM (anti-PDGF-D; p = 0.0001, anti-HGF; p = 0.003). Our results suggest that SHED-CM may provide substantial therapeutic benefits for RIX primarily through the activation of multiple antioxidant enzyme genes in the target tissue.
Anrizandy Narwidina, Aya Miyazaki, Kokoro Iwata, Rika Kurogoushi, Asuna Sugimoto, Yasusei Kudo, Keita Kawarabayashi, Yoshihito Yamakawa, Yuki Akazawa, Takamasa Kitamura, Hiroshi Nakagawa, Kimiko Ueda Yamaguchi, Tomokazu Hasegawa, Keigo Yoshizaki, Satoshi Fukumoto, Akihito Yamamoto, Naozumi Ishimaru, Tomonori Iwasaki and Tsutomu Iwamoto : Iroquois homeobox 3 regulates odontoblast proliferation and differentiation mediated by Wnt5a expression., Biochemical and Biophysical Research Communications, Vol.650, 47-54, 2023.
(Summary)
Iroquois homeobox (Irx) genes are TALE-class homeobox genes that are evolutionarily conserved across species and have multiple critical cellular functions in fundamental tissue development processes. Previous studies have shown that Irxs genes are expressed during tooth development. However, the precise roles of genes in teeth remain unclear. Here, we demonstrated for the first time that Irx3 is an essential molecule for the proliferation and differentiation of odontoblasts. Using cDNA synthesized from postnatal day 1 (P1) tooth germs, we examined the expression of all Irx genes (Irx1-Irx6) by RT-PCR and found that all genes except Irx4 were expressed in the tooth tissue. Irx1-Irx3 a were expressed in the dental epithelial cell line M3H1 cells, while Irx3 and Irx5 were expressed in the dental mesenchymal cell line mDP cells. Only Irx3 was expressed in both undifferentiated cell lines. Immunostaining also revealed the presence of IRX3 in the dental epithelial cells and mesenchymal condensation. Inhibition of endogenous Irx3 by siRNA blocks the proliferation and differentiation of mDP cells. Wnt3a, Wnt5a, and Bmp4 are factors involved in odontoblast differentiation and were highly expressed in mDP cells by quantitative PCR analysis. Interestingly, the expression of Wnt5a (but not Wnt3a or Bmp4) was suppressed by Irx3 siRNA. These results suggest that Irx3 plays an essential role in part through the regulation of Wnt5a expression during odontoblast proliferation and differentiation.
YAO LIU, Fumiya Kano, Noboru Hashimoto, LINZE XIA, Qiao Zhou, Xingmei Feng, Hideharu Hibi, Aya Miyazaki, Tsutomu Iwamoto, Yoshizo Matsuka, Zhijun Zhang, Eiji Tanaka and Akihito Yamamoto : Conditioned Medium From the Stem Cells of Human Exfoliated Deciduous Teeth Ameliorates Neuropathic Pain in a Partial Sciatic Nerve Ligation Model., Frontiers in Pharmacology, Vol.13, 745020, 2022.
(Summary)
In neuropathic pain (NP), injury or diseases of the somatosensory system often result in highly debilitating chronic pain. Currently, there is no effective drug for the complete and definitive treatment of NP. We investigated the therapeutic potential of conditioned medium (CM) derived from stem cells from human exfoliated deciduous teeth (SHED-CM) against NP using a mouse partial sciatic nerve ligation (PSL) model. Abnormal pain sensation, such as tactile allodynia and hyperalgesia, can be caused by PSL. In the behavioral test, intravenous administration of SHED-CM greatly improved the PSL-induced hypersensitivity. We found that treatment with SHED-CM resulted in the recruitment of M2 macrophages in the injured sciatic nerve and ipsilateral L4/L5 dorsal root ganglion and suppressed microglial activation in the spinal cord. Notably, specific depletion of the anti-inflammatory M2 macrophages by mannosylated-Clodrosome markedly reduced the antinociceptive effect of SHED-CM. Intravenous administration of CM from M2 induced by SHED-CM (M2-CM) ameliorated the PSL-induced hypersensitivity. We found that M2-CM directly suppressed the expression of nociceptive receptors as well as proinflammatory mediators in Schwann cells. Taken together, our data suggest that SHED-CM ameliorates NP through the induction of the analgesic anti-inflammatory M2 macrophages. Thus, SHED-CM may be a novel therapeutic candidate for NP.
Tomoyuki Ueda, Taisei Ito, Masatoshi Inden, Hisaka Kurita, Akihito Yamamoto and Isao Hozumi : Stem Cells From Human Exfoliated Deciduous Teeth-Conditioned Medium (SHED-CM) is a Promising Treatment for Amyotrophic Lateral Sclerosis, Frontiers in Pharmacology, Vol.3, No.13, 2022.
(Summary)
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder, characterized by the loss of upper and lower motor neurons, for which an effective treatment has yet to be developed. Previous reports have shown that excessive oxidative stress, related to mitochondrial dysfunction and the accumulation of misfolding protein, contributes to ALS pathology. In terms of treatment, it remains necessary to identify effective medicines for multiple therapeutic targets and have additive effects against several disorders. In this study, we investigated stem cells from human exfoliated deciduous teeth (SHED), which release many factors, such as neurotrophic factors and cytokines, and are applied to treat neurological diseases. Specifically, we examined whether SHED-conditioned medium (CM), i.e., the serum-free culture supernatant of SHED, reduced mutant SOD1-induced intracellular aggregates and neurotoxicity. We found that SHED-CM significantly suppressed the mutant SOD1-induced intracellular aggregates and neurotoxicity. The neuroprotective effects of SHED-CM are partly related to heat shock protein and the activation of insulin-like growth factor-1 receptor. SHED-CM also had a protective effect on induced pluripotent stem cell-derived motor neurons. Moreover, SHED-CM was effective against not only familial ALS but also sporadic ALS. Overall, these results suggest that SHED-CM could be a promising treatment for slowing the progression of ALS.
Kokoro Iwata, Keita Kawarabayashi, Keigo Yoshizaki, Tian Tian, Kan Saito, Asuna Sugimoto, Rika Kurogoushi, Aya Yamada, Akihito Yamamoto, Yasusei Kudo, Naozumi Ishimaru, Satoshi Fukumoto and Tsutomu Iwamoto : von Willebrand factor D and EGF domains regulate ameloblast differentiation and enamel formation., Journal of Cellular Physiology, Vol.237, No.3, 1964-1979, 2021.
(Summary)
Cell- and tissue-specific extracellular matrix (ECM) composition plays an important role in organ development, including teeth, by regulating cell behaviors, such as cell proliferation and differentiation. Here, we demonstrate for the first time that von Willebrand factor D and epidermal growth factor (EGF) domains (Vwde), a previously uncharacterized ECM protein, is specifically expressed in teeth and regulates cell proliferation and differentiation in inner enamel epithelial cells (IEEs) and enamel formation. We identified the Vwde as a novel ECM protein through bioinformatics using the NCBI expressed sequence tag database for mice. Vwde complementary DNA encodes 1773 amino acids containing a signal peptide, a von Willebrand factor type D domain, and tandem calcium-binding EGF-like domains. Real-time polymerase chain reaction demonstrated that Vwde is highly expressed in tooth tissue but not in other tissues including the brain, lung, heart, liver, kidney, and bone. In situ hybridization revealed that the IEEs expressed Vwde messenger RNA in developing teeth. Immunostaining showed that VWDE was localized at the proximal and the distal ends of the pericellular regions of the IEEs. Vwde was induced during the differentiation of mouse dental epithelium-derived M3H1 cells. Vwde-transfected M3H1 cells secreted VWDE protein into the culture medium and inhibited cell proliferation, whereas ameloblastic differentiation was promoted. Furthermore, Vwde increased the phosphorylation of extracellular signal-regulated kinase 1/2 and protein kinase B and strongly induced the expression of the intercellular junction protein, N-cadherin (Ncad). Interestingly, the suppression of endogenous Vwde inhibited the expression of Ncad. Finally, we created Vwde-knockout mice using the CRISPR-Cas9 system. Vwde-null mice showed low mineral density, rough surface, and cracks in the enamel, indicating the enamel hypoplasia phenotype. Our findings suggest that Vwde assembling the matrix underneath the IEEs is essential for Ncad expression and enamel formation.
Arief Waskitho, Yumiko Yamamoto, S Raman, Fumiya Kano, Huijiao Yan, R Raju, S Afroz, Tsuyoshi Morita, Daisuke Ikutame, Kazuo Okura, Masamitsu Ohshima, Akihito Yamamoto, Otto Baba and Yoshizo Matsuka : Peripherally Administered Botulinum Toxin Type A Localizes Bilaterally in Trigeminal Ganglia of Animal Model, Toxins, Vol.13, No.10, 704, 2021.
Hisanori Muto, Takanori Ito, Taku Tanaka, Shinya Yokoyama, Kenta Yamamoto, Norihiro Imai, Yoji Ishizu, Keiko Maeda, Takashi Honda, Tetsuya Ishikawa, Asuka Kato, Taichi Ohshiro, Fumiya Kano, Akihito Yamamoto, Kiyoshi Sakai, Hideharu Hibi, Masatoshi Ishigami and Mitsuhiro Fujishiro : Conditioned medium from stem cells derived from human exfoliated deciduous teeth ameliorates NASH via the GutLiver axis, Scientific Reports, Vol.18778 (2021), No.11, 2021.
(Summary)
Non-alcoholic steatohepatitis (NASH) occurrence has been increasing and is becoming a major cause of liver cirrhosis and liver cancer. However, effective treatments for NASH are still lacking. We examined the benefits of serum-free conditioned medium from stem cells derived from human exfoliated deciduous teeth (SHED-CM) on a murine non-alcoholic steatohepatitis (NASH) model induced by a combination of Western diet (WD) and repeated administration of low doses of carbon tetrachloride intraperitoneally, focusing on the gut-liver axis. We showed that repeated intravenous administration of SHED-CM significantly ameliorated histological liver fibrosis and inflammation in a murine NASH model. SHED-CM inhibited parenchymal cell apoptosis and reduced the activation of inflammatory macrophages. Gene expression of pro-inflammatory and pro-fibrotic mediators (such as Tnf-α, Tgf-β, and Ccl-2) in the liver was reduced in mice treated with SHED-CM. Furthermore, SHED-CM protected intestinal tight junctions and maintained intestinal barrier function, while suppressing gene expression of the receptor for endotoxin, Toll-like receptor 4, in the liver. SHED-CM promoted the recovery of Caco-2 monolayer dysfunction induced by IFN-γ and TNF-α in vitro. Our findings suggest that SHED-CM may inhibit NASH fibrosis via the gut-liver axis, in addition to its protective effect on hepatocytes and the induction of macrophages with unique anti-inflammatory phenotypes.
Tsendsuren Khurel-Ochir, Takashi Izawa, Akihiko Iwasa, Fumiya Kano, Akihito Yamamoto and Eiji Tanaka : The immunoregulatory role of p21 in the development of the temporomandibular joint-osteoarthritis., Clinical and Experimental Dental Research, Vol.7, No.3, 313-322, 2021.
(Summary)
Our findings indicate that p21 in chondrocytes contributes to regulate matrix synthesis via the control ofm aggrecan and MMP-13 expression under mechanical stress. Thus, p21 might regulate the pathogenesis of osteoarthritis in the TMJ.
Naoko Ogasawara, Fumiya Kano, Noboru Hashimoto, Hiroki Mori, YAO LIU, LINZE XIA, Takuma Sakamaki, Hideharu Hibi, Tsutomu Iwamoto, Eiji Tanaka and Akihito Yamamoto : Factors Secreted from Dental Pulp Stem Cells Show Multifaceted Benefits for Treating Experimental Temporomandibular Joint Osteoarthritis, Osteoarthritis and Cartilage, Vol.28, No.6, 831-841, 2020.
(Summary)
Temporomandibular joint osteoarthritis (TMJOA) is a degenerative disease characterized by progressive cartilage degeneration, abnormal bone remodeling, and chronic pain. In this study, we aimed to investigate effective therapies to reverse or suppress TMJOA progression. To this end, we performed intravenous administration of serum free conditioned media from human exfoliated deciduous teeth stem cells (SHED-CM) into a mechanical-stress induced murine TMJOA model. SHED-CM administration markedly suppressed temporal muscle inflammation, and improved bone integrity and surface smoothness of the destroyed condylar cartilage. Moreover, SHED-CM treatment decreased the number of IL-1β, iNOS, and MMP-13 expressing chondrocytes, whereas it specifically increased PCNA-positive cells in the multipotent polymorphic cell layer. Notably, the numbers of TUNEL-positive apoptotic chondrocytes in the SHED-CM treated condyles were significantly lower than in those treated with DMEM, whereas the proteoglycan positive area was restored to a level similar to that of the sham treated group, demonstrating that SHED-CM treatment regenerated the mechanical-stress injured condylar cartilage and subchondral bone. Secretome analysis revealed that SHED-CM contained multiple therapeutic factors that act in osteochondral regeneration. Our data demonstrated that SHED-CM treatment promoted the regeneration and repair of mechanical-stress induced mouse TMJOA. Our observations suggest that SHED-CM has potential to be a potent tissue-regenerating therapeutic agent for patients with severe TMJOA.
Yuma Kitase, Yoshiaki Sato, Kazuto Ueda, Toshihiko Suzuki, Alkisti Mikrogeorgiou, Yuichiro Sugiyama, Kohki Matsubara, Yuka Tsukagoshi Okabe, Shinobu Shimizu, Hitoshi Hirata, Hiroshi Yukawa, Yoshinobu Baba, Masahiro Tsuji, Yoshiyuki Takahashi, Akihito Yamamoto and Masahiro Hayakawa : A Novel Treatment with Stem Cells from Human Exfoliated Deciduous Teeth for Hypoxic-Ischemic Encephalopathy in Neonatal Rats, Stem Cells and Development, Vol.29, No.2, 63-74, 2020.
(Summary)
Recently, cell therapy has been developed as a novel treatment for perinatal hypoxic-ischemic encephalopathy (HIE), which is an important cause of neurological disorder and death, and stem cells from human exfoliated deciduous teeth (SHED) express early markers for mesenchymal and neuroectodermal stem cells. We investigated the treatment effect of SHED for HIE in neonatal rats. Seven-day-old rats underwent ligation of the left carotid artery and were exposed to 8% hypoxic treatment. SHED (1 × 10 cells) were injected via the right external jugular vein 24 h after the insult. The effect of intravenous administration of SHED cells was evaluated neurologically and pathophysiologically. In the evaluation of engraftment using quantum dots 655, only a few SHED were detected in the injured cortex. In the immunohistological evaluation 24 h after injection, the numbers of positive cells of active caspase-3 and anti-4 hydroxynonenal antiserum were lower in the SHED group than in the vehicle group. The number of Iba-1 cells in the cortex was higher in the SHED group. However, the proportion of M1 microglia (Iba-1/ED-1) was significantly decreased, whereas M2 microglia (Iba-1/CD206) tended to increase in the SHED group. In the behavioral tests performed 5 months after hypoxic treatment, compared to the vehicle group, the SHED group showed significant elongation of the endurance time in the rotarod treadmill test, significantly ameliorated proportion of using the impaired hand in the cylinder test, significantly lower ratio of right/left front paw area in gait analysis, and significantly higher avoidance rate in the active avoidance test. In the in vitro experiment with cultured neurons exposed to oxygen-glucose deprivation, we confirmed the neuroprotective effect of the condition medium of SHED. These results suggested that intravenous administration of SHED exerted a treatment effect both histologically and functionally, possibly via a paracrine effect.
These results suggested that SHED-CM might have a therapeutic effect on diabetic polyneuropathy through promoting neurite outgrowth, and the increase in capillaries might contribute to the improvement of neural function.
Noboru Hashimoto, Shizuka Ito, Akiko Tsuchida, H Robiul Bhuiyan, Tetsuya Okajima, Akihito Yamamoto, Keiko Furukawa, Yuhsuke Ohmi and Koichi Furukawa : The ceramide moiety of disialoganglioside (GD3) is essential for GD3 recognition by the sialic acid-binding lectin SIGLEC7 on the cell surface., The Journal of Biological Chemistry, Vol.294, No.28, 10833-10845, 2019.
(Summary)
) gene, involved in phytoceramide synthesis, disclosing that DES2 inhibition confers SIGLEC7 binding. Furthermore, knocking out fatty acid 2-hydroxylase also resulted in the emergence of SIGLEC7 binding to the cell surface. To analyze the effects of binding between SIGLEC7 and various GD3 species on natural killer function, we investigated cytotoxicity of peripheral blood mononuclear cells from healthy donors toward GD3S-transfected DLD-1 (DLD-1-GD3S) cells and DLD-1-GD3S cells with modified ceramides. We found that cytotoxicity is suppressed in DLD-1-GD3S cells with dehydroxylated GD3s. These results indicate that the ceramide structures in glycosphingolipids affect SIGLEC7 binding and distribution on the cell surface and influence cell sensitivity to killing by SIGLEC7-expressing effector cells.
Jun Ishikawa, Fumiya Kano, Yuji Ando, Hideharu Hibi and Akihito Yamamoto : Monocyte chemoattractant protein-1 and secreted ectodomain of sialic acid-binding Ig-like lectin-9 enhance bone regeneration by inducing M2 macrophages, Journal of Oral and Maxillofacial Surgery, Medicine, and Pathology, Vol.31, No.3, 169-174, 2019.
Takanori Ito, Masatoshi Ishigami, Yoshihiro Matsushita, Marina Hirata, Kohki Matsubara, Tetsuya Ishikawa, Hideharu Hibi, Minoru Ueda, Yoshiki Hirooka, Hidemi Goto and Akihito Yamamoto : Secreted Ectodomain of SIGLEC-9 and MCP-1 Synergistically Improve Acute Liver Failure in Rats by Altering Macrophage Polarity., Scientific Reports, Vol.7, 2017.
(Summary)
Effective treatments for acute liver failure (ALF) are still lacking. We recently reported that a single intravenous administration of serum-free conditioned medium from stem cells derived from human exfoliated deciduous teeth (SHED-CM) into the D-galactosamine (D-Gal)-induced rat ALF model improves the liver injury. However, the specific factors in SHED-CM that are responsible for resolving ALF remain unclear. Here we found that depleting SHED-CM of two anti-inflammatory M2 macrophage inducers-monocyte chemoattractant protein-1 (MCP-1) and the secreted ectodomain of sialic acid-binding Ig-like lectin-9 (sSiglec-9)-abolished its ability to resolve rat ALF. Furthermore, treatment with MCP-1/sSiglec-9 alone dramatically improved the survival of ALF rats. This treatment induced anti-inflammatory M2, suppressed hepatocyte apoptosis, and promoted hepatocyte proliferation. Treatment with an M2-depletion reagent (mannosylated clodronate liposomes) suppressed the recovery. In addition, MCP-1 and sSiglec-9 synergistically promoted the M2 differentiation of bone marrow-derived macrophages via CCR2, accompanied by the production of multiple liver-regenerating factors. The conditioned medium from MCP-1/sSiglec-9-activated M2 macrophages, but not from interleukin-4-induced ones, suppressed the D-Gal- and LPS-induced apoptosis of primary hepatocytes and promoted their proliferation in vitro. The unique combination of MCP-1/sSiglec-9 ameliorates rat ALF by inhibiting hepatocellular apoptosis and promoting liver regeneration through the induction of anti-inflammatory/tissue-repairing M2 macrophages.
Hirohiko Okamura, Kaya Yoshida, Hiroyuki Morimoto, Jumpei Teramachi, Kazuhiko Ochiai, Tatsuji Haneji and Akihito Yamamoto : Role of Protein Phosphatase 2A in Osteoblast Differentiation and Function., Journal of Clinical Medicine, Vol.6, No.3, 2017.
(Summary)
The reversible phosphorylation of proteins plays hugely important roles in a variety of cellular processes, such as differentiation, proliferation, and apoptosis. These processes are strictly controlled by protein kinases (phosphorylation) and phosphatases (de-phosphorylation). Here we provide a brief history of the study of protein phosphorylation, including a summary of different types of protein kinases and phosphatases. One of the most physiologically important serine/threonine phosphatases is PP2A. This review provides a description of the phenotypes of various PP2A transgenic mice and further focuses on the known functions of PP2A in bone formation, including its role in osteoblast differentiation and function. A reduction in PP2A promotes bone formation and osteoblast differentiation through the regulation of bone-related transcription factors such as Osterix. Interestingly, downregulation of PP2A also stimulates adipocyte differentiation from undifferentiated mesenchymal cells under the appropriate adipogenic differentiation conditions. In osteoblasts, PP2A is also involved in the ability to control osteoclastogenesis as well as in the proliferation and metastasis of osteosarcoma cells. Thus, PP2A is considered to be a comprehensive factor in controlling the differentiation and function of cells derived from mesenchymal cells such as osteoblasts and adipocytes.
(Keyword)
osteoblast / protein dephosphorylation / protein phosphatase
Fumiya Kano, Kohki Matsubara, Minoru Ueda, Hideharu Hibi and Akihito Yamamoto : Secreted Ectodomain of Sialic Acid-Binding Ig-Like Lectin-9 and Monocyte Chemoattractant Protein-1 Synergistically Regenerate Transected Rat Peripheral Nerves by Altering Macrophage Polarity, Stem Cells, Vol.35, No.3, 641-653, 2017.
(Summary)
Peripheral nerves (PNs) exhibit remarkable self-repairing reparative activity after a simple crush or cut injury. However, the neuronal transection involving a nerve gap overwhelms their repairing activity and causes persistent paralysis. Here, we show that an implantation of the serum-free conditioned medium from stem cells from human exfoliated deciduous teeth (SHED-CM) immersed in a collagen sponge into the nerve gap formed by rat facial nerves transection restored the neurological function. In contrast, SHED-CM specifically depleted of a set of anti-inflammatory M2 macrophage inducers, monocyte chemoattractant protein-1 (MCP-1) and the secreted ectodomain of sialic acid-binding Ig-like lectin-9 (sSiglec-9) lost the ability to restore neurological function in this model. Notably, the combination of MCP-1 and sSiglec-9 induced the polarization of M2 macrophages in vitro, resulting in the expression of multiple trophic factors that enhanced proliferation, migration, and differentiation of Schwann cells, blood vessel formation, and nerve fiber extension. Furthermore, the implantation of a collagen graft containing MCP-1/sSiglec-9 into the nerve gap induced anti-inflammatory M2 macrophage polarization, generated a Schwann-cell bridge instead of fibrotic scar, induced axonal regrowth, and restored nerve function. The specific elimination of M2 macrophages by Mannosylated-Clodrosome suppressed the MCP-1/sSiglec-9-mediated neurological recovery. Taken together, our data suggest that MCP-1/sSiglec-9 regenerates PNs by inducing tissue-repairing M2 macrophages and may provide therapeutic benefits for severe peripheral nerve injuries. Stem Cells 2017;35:641-653.
This study demonstrated that a single intravenous administration of stem cells from human exfoliated deciduous teeth (SHEDs) or of the serum-free conditioned medium (CM) derived from SHEDs markedly improved mouse liver fibrosis (LF). SHED-CM suppressed chronic inflammation, eliminated activated hepatic stellate cells by inducing their apoptosis, protected hepatocytes from undergoing apoptosis, and induced differentiation of tissue-repairing macrophages expressing high levels of the profibrinolytic factor matrix metalloproteinase 13. Furthermore, hepatocyte growth factor played a central role in the SHED-CM-mediated resolution of LF. This is the first report demonstrating the multifaceted therapeutic benefits of secreted factors derived from SHEDs for LF.
Takuya Matsumoto, Nobunori Takahashi, Toshihisa Kojima, Yutaka Yoshioka, Jun Ishikawa, Koichi Furukawa, Kenji Ono, Makoto Sawada, Naoki Ishiguro and Akihito Yamamoto : Soluble Siglec-9 suppresses arthritis in a collagen-induced arthritis mouse model and inhibits M1 activation of RAW264.7 macrophages., Arthritis Research & Therapy, Vol.18, No.1, 2016.
(Summary)
In this study, we have demonstrated the therapeutic effects of sSiglec-9 in a murine CIA model. The mechanism underlying these effects involves the suppression of M1 proinflammatory macrophages by inhibiting the NF-kB pathway. sSiglec-9 may provide a novel therapeutic option for patients with rheumatoid arthritis refractory to currently available drugs.
Chiaki Shimojima, Hideyuki Takeuchi, Shijie Jin, Bijay Parajuli, Hisashi Hattori, Akio Suzumura, Hideharu Hibi, Minoru Ueda and Akihito Yamamoto : Conditioned Medium from the Stem Cells of Human Exfoliated Deciduous Teeth Ameliorates Experimental Autoimmune Encephalomyelitis., The Journal of Immunology, Vol.196, No.10, 4164-4171, 2016.
(Summary)
Multiple sclerosis (MS) is a major neuroinflammatory demyelinating disease of the CNS. Current MS treatments, including immunomodulators and immunosuppressants, do not result in complete remission. Stem cells from human exfoliated deciduous teeth (SHEDs) are mesenchymal stem cells derived from dental pulp. Both SHED and SHED-conditioned medium (SHED-CM) exhibit immunomodulatory and regenerative activities and have the potential to treat various diseases. In this study, we investigated the efficacy of SHED-CM in treating experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. EAE mice treated with a single injection of SHED-CM exhibited significantly improved disease scores, reduced demyelination and axonal injury, and reduced inflammatory cell infiltration and proinflammatory cytokine expression in the spinal cord, which was associated with a shift in the microglia/macrophage phenotype from M1 to M2. SHED-CM also inhibited the proliferation of myelin oligodendrocyte glycoprotein-specific CD4(+) T cells, as well as their production of proinflammatory cytokines in vitro. Treatment of EAE mice with the secreted ectodomain of sialic acid-binding Ig-like lectin-9, a major component of SHED-CM, recapitulated the effects of SHED-CM treatment. Our data suggest that SHED-CM and secreted ectodomain of sialic acid-binding Ig-like lectin-9 may be novel therapeutic treatments for autoimmune diseases, such as MS.
Masahito Fujio, Zhe Xing, Niyaz Sharabi, Ying Xue, Akihito Yamamoto, Hideharu Hibi, Minoru Ueda, Inge Fristad and Kamal Mustafa : Conditioned media from hypoxic-cultured human dental pulp cells promotes bone healing during distraction osteogenesis., Journal of Tissue Engineering and Regenerative Medicine, 2015.
Jun Ishikawa, Nobunori Takahashi, Takuya Matsumoto, Yutaka Yoshioka, Noriyuki Yamamoto, Masaya Nishikawa, Hideharu Hibi, Naoki Ishigro, Minoru Ueda, Koichi Furukawa and Akihito Yamamoto : Factors secreted from dental pulp stem cells show multifaceted benefits for treating experimental rheumatoid arthritis., Bone, Vol.83, 210-219, 2015.
(Summary)
Rheumatoid arthritis (RA) is an autoimmune disease characterized by synovial hyperplasia and chronic inflammation, which lead to the progressive destruction of cartilage and bone in the joints. Numerous studies have reported that administrations of various types of MSCs improve arthritis symptoms in animal models, by paracrine mechanisms. However, the therapeutic effects of the secreted factors alone, without the cell graft, have been uncertain. Here, we show that a single intravenous administration of serum-free conditioned medium (CM) from human deciduous dental pulp stem cells (SHED-CM) into anti-collagen type II antibody-induced arthritis (CAIA), a mouse model of rheumatoid arthritis (RA), markedly improved the arthritis symptoms and joint destruction. The therapeutic efficacy of SHED-CM was associated with an induction of anti-inflammatory M2 macrophages in the CAIA joints and the abrogation of RANKL expression. SHED-CM specifically depleted of an M2 macrophage inducer, the secreted ectodomain of sialic acid-binding Ig-like lectin-9 (ED-Siglec-9), exhibited a reduced ability to induce M2-related gene expression and attenuate CAIA. SHED-CM also inhibited the RANKL-induced osteoclastogenesis in vitro. Collectively, our findings suggest that SHED-CM provides multifaceted therapeutic effects for treating CAIA, including the ED-Siglec-9-dependent induction of M2 macrophage polarization and inhibition of osteoclastogenesis. Thus, SHED-CM may represent a novel anti-inflammatory and reparative therapy for RA.
Satoshi Yamaguchi, Rei Shibata, Noriyuki Yamamoto, Masaya Nishikawa, Hideharu Hibi, Tohru Tanigawa, Minoru Ueda, Toyoaki Murohara and Akihito Yamamoto : Dental pulp-derived stem cell conditioned medium reduces cardiac injury following ischemia-reperfusion., Scientific Reports, Vol.5, 16295, 2015.
(Summary)
Stem cells from human exfoliated deciduous teeth (SHEDs) can regenerate various tissues. We investigated the impact of SHED-conditioned medium (SHED-CM) on myocardial injury in a mouse model of ischemia-reperfusion (I/R). Wild-type (WT) mice were subjected to myocardial ischemia followed by reperfusion. SHED-CM was intravenously injected at 5 min after reperfusion. Administration of SHED-CM reduced myocardial infarct size as well as decreased apoptosis and inflammatory cytokine levels, such as TNF-, IL-6, and IL-, in the myocardium following I/R. In cultured cardiac myocytes, SHED-CM significantly suppressed apoptosis under hypoxia/serum-deprivation and reduced LPS-induced expression of pro-inflammatory genes. Furthermore, anti-apoptotic action of SHED-CM was stronger than bone marrow-derived stem cell (BMSC)-CM or adipose-derived stem cell (ADSC)-CM in cardiac myocytes. SHED-CM contains a higher concentration of hepatocyte growth factor (HGF) than BMSC-CM and ADSC-CM, and neutralization of HGF attenuated the inhibitory actions of SHED-CM on apoptosis in cardiac myocytes. Finally, WT mice were intravenously treated with an HGF-depleted SHED-CM, followed by myocardial I/R. HGF depletion significantly attenuated the inhibitory actions of SHED-CM on myocardial infarct size and apoptosis after I/R. SHED-CM protects the heart from acute ischemic injury because it suppresses inflammation and apoptosis. SHED-CM could be a useful treatment option for acute myocardial infarction.
Yuka Hattori, Hangsoo Kim, Naotake Tsuboi, Akihito Yamamoto, Shinichi Akiyama, Yiqin Shi, Takayuki Katsuno, Tomoki Kosugi, Minoru Ueda, Seiichi Matsuo and Shoichi Maruyama : Therapeutic Potential of Stem Cells from Human Exfoliated Deciduous Teeth in Models of Acute Kidney Injury., PLoS ONE, Vol.10, No.10, e0140121, 2015.
(Summary)
SHED attenuated the levels of inflammatory cytokines and improved kidney function in AKI induced by IRI. SHED secreted factors reduced MCP-1 and increased HGF expression, which promoted wound healing. These results suggest that SHED might provide a novel stem cell resource, which can be applied for the treatment of ischemic kidney injury.
Takako Izumoto-Akita, Shin Tsunekawa, Akihito Yamamoto, Eita Uenishi, Kota Ishikawa, Hidetada Ogata, Atsushi Iida, Makoto Ikeniwa, Kaori Hosokawa, Yasuhiro Niwa, Ryuya Maekawa, Yuichiro Yamauchi, Yusuke Seino, Yoji Hamada, Hideharu Hibi, Hiroshi Arima, Minoru Ueda and Yutaka Oiso : Secreted factors from dental pulp stem cells improve glucose intolerance in streptozotocin-induced diabetic mice by increasing pancreatic -cell function., BMJ Open Diabetes Research & Care, Vol.3, No.1, e000128, 2015.
(Summary)
These findings suggest that SHED-CM provides direct protection and encourages the propagation of -cells, and has potential as a novel strategy for treatment of diabetes.
Tsuneyuki Mita, Yoko Furukawa-Hibi, Hideyuki Takeuchi, Hisashi Hattori, Kiyofumi Yamada, Hideharu Hibi, Minoru Ueda and Akihito Yamamoto : Conditioned medium from the stem cells of human dental pulp improves cognitive function in a mouse model of Alzheimer's disease., Behavioural Brain Research, Vol.293, 189-197, 2015.
(Summary)
Alzheimer's disease (AD) is a progressive, neurodegenerative disease characterized by a decline in cognitive abilities and the appearance of -amyloid plaques in the brain. Although the pathogenic mechanisms associated with AD are not fully understood, activated microglia releasing various neurotoxic factors, including pro-inflammatory cytokines and oxidative stress mediators, appear to play major roles. Here, we investigated the therapeutic benefits of a serum-free conditioned medium (CM) derived from the stem cells of human exfoliated deciduous teeth (SHEDs) in a mouse model of AD. The intranasal administration of SHEDs in these mice resulted in substantially improved cognitive function. SHED-CM contained factors involved in multiple neuroregenerative mechanisms, such as neuroprotection, axonal elongation, neurotransmission, the suppression of inflammation, and microglial regulation. Notably, SHED-CM attenuated the pro-inflammatory responses induced by -amyloid plaques, and generated an anti-inflammatory/tissue-regenerating environment, which was accompanied by the induction of anti-inflammatory M2-like microglia. Our data suggest that SHED-CM may provide significant therapeutic benefits for AD.
Hirotaka Wakayama, Naozumi Hashimoto, Yoshihiro Matsushita, Kohki Matsubara, Noriyuki Yamamoto, Yoshinori Hasegawa, Minoru Ueda and Akihito Yamamoto : Factors secreted from dental pulp stem cells show multifaceted benefits for treating acute lung injury in mice., Cytotherapy, Vol.17, No.8, 1119-1129, 2015.
(Summary)
Our results suggest that SHED-secreted factors provide multifaceted therapeutic effects, including a strong M2-inducing activity, for treating BLM-induced ALI. This work may open new avenues for research on stem cell-based ARDS therapies.
Hiromi Fujii, Kohki Matsubara, Kiyoshi Sakai, Mikako Ito, Kinji Ohno, Minoru Ueda and Akihito Yamamoto : Dopaminergic differentiation of stem cells from human deciduous teeth and their therapeutic benefits for Parkinsonian rats., Brain Research, Vol.1613, 59-72, 2015.
(Summary)
Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by the loss of nigrostriatal dopaminergic (DAergic) neurons and the depletion of striatal dopamine. Here we show that DAergic-neuron-like cells could be efficiently induced from stem cells derived from human exfoliated deciduous teeth (SHEDs), and that these induced cells had therapeutic benefits in a 6-OHDA-induced Parkinsonian rat model. In our protocol, EGF and bFGF signaling activated the SHED's expression of proneural genes, Ngn2 and Mash1, and subsequent treatment with brain-derived neurotrophic factor (BDNF) promoted their maturation into DAergic neuron-like SHEDs (dSHEDs). A hypoxic DAergic differentiation protocol improved cell viability and enhanced the expression of multiple neurotrophic factors, including BDNF, GDNF, NT-3, and HGF. Engrafted dSHEDs survived in the striatum of Parkinsonian rats, improved the DA level more efficiently than engrafted undifferentiated SHEDs, and promoted the recovery from neurological deficits. Our findings further suggested that paracrine effects of dSHEDs contributed to neuroprotection against 6-OHDA-induced neurodegeneration and to nigrostriatal tract restoration. In addition, we found that the conditioned medium derived from dSHEDs protected primary neurons against 6-OHDA toxicity and accelerated neurite outgrowth in vitro. Thus, our data suggest that stem cells derived from dental pulp may have therapeutic benefits for PD.
Kohki Matsubara, Yoshihiro Matsushita, Kiyoshi Sakai, Fumiya Kano, Megumi Kondo, Mariko Noda, Noboru Hashimoto, Shiro Imagama, Naoki Ishiguro, Akio Suzumura, Minoru Ueda, Koichi Furukawa and Akihito Yamamoto : Secreted ectodomain of sialic acid-binding Ig-like lectin-9 and monocyte chemoattractant protein-1 promote recovery after rat spinal cord injury by altering macrophage polarity., The Journal of Neuroscience, Vol.35, No.6, 2452-2464, 2015.
(Summary)
Engrafted mesenchymal stem cells from human deciduous dental pulp (SHEDs) support recovery from neural insults via paracrine mechanisms that are poorly understood. Here we show that the conditioned serum-free medium (CM) from SHEDs, administered intrathecally into rat injured spinal cord during the acute postinjury period, caused remarkable functional recovery. The ability of SHED-CM to induce recovery was associated with an immunoregulatory activity that induced anti-inflammatory M2-like macrophages. Secretome analysis of the SHED-CM revealed a previously unrecognized set of inducers for anti-inflammatory M2-like macrophages: monocyte chemoattractant protein-1 (MCP-1) and the secreted ectodomain of sialic acid-binding Ig-like lectin-9 (ED-Siglec-9). Depleting MCP-1 and ED-Siglec-9 from the SHED-CM prominently reduced its ability to induce M2-like macrophages and to promote functional recovery after spinal cord injury (SCI). The combination of MCP-1 and ED-Siglec-9 synergistically promoted the M2-like differentiation of bone marrow-derived macrophages in vitro, and this effect was abolished by a selective antagonist for CC chemokine receptor 2 (CCR2) or by the genetic knock-out of CCR2. Furthermore, MCP-1 and ED-Siglec-9 administration into the injured spinal cord induced M2-like macrophages and led to a marked recovery of hindlimb locomotor function after SCI. The inhibition of this M2 induction through the inactivation of CCR2 function abolished the therapeutic effects of both SHED-CM and MCP-1/ED-Siglec-9. Macrophages activated by MCP-1 and ED-Siglec-9 extended neurite and suppressed apoptosis of primary cerebellar granule neurons against the neurotoxic effects of chondroitin sulfate proteoglycans. Our data suggest that the unique combination of MCP-1 and ED-Siglec-9 repairs the SCI through anti-inflammatory M2-like macrophage induction.
Fumiya Kano, Kohki Matsubara and Akihito Yamamoto : The development of new therapy facial nerve injury using the m2 macrophage induction factors, Inflammation Research, Vol.64, No.Sup2, S229-S230, 2015.
35.
Sun Yang, Finne-Wistrand Anna, Waag Thilo, Xing Zhe, Yassin Mohamed, Akihito Yamamoto, Mustafa Kamal, Steinmüller-Nethl Doris, Krueger Anke and Albertsson Ann-Christine : Reinforced Degradable Biocomposite by Homogenously Distributed Functionalized Nanodiamond Particles, Macromolecular Materials and Engineering, Vol.300, No.4, 436-447, 2015.
Yuji Ando, Kohki Matsubara, Jun Ishikawa, Masahito Fujio, Ryutaro Shohara, Hideharu Hibi, Minoru Ueda and Akihito Yamamoto : Stem cell-conditioned medium accelerates distraction osteogenesis through multiple regenerative mechanisms., Bone, Vol.61, 82-90, 2014.
(Summary)
Distraction osteogenesis (DO) successfully induces large-scale skeletal tissue regeneration, but it involves an undesirably long treatment period. A high-speed DO mouse model (H-DO) with a distraction speed twice that of a control DO model failed to generate new bone callus in the distraction gap. Here we demonstrate that the local administration of serum-free conditioned medium from human mesenchymal stem cells (MSC-CM) accelerated callus formation in the mouse H-DO model. Secretomic analysis identified factors contained in MSC-CM that recruit murine bone marrow stromal cells (mBMSCs) and endothelial cells/endothelial progenitor cells (EC/EPCs), inhibit inflammation and apoptosis, and promote osteoblast differentiation, angiogenesis, and cell proliferation. Functional assays identified MCP-1/-3 and IL-3/-6 as essential factors in recruiting mBMSCs and EC/EPCs. IL-3/-6 also enhanced the osteogenic differentiation of mBMSCs. MSC-CM that had been depleted of MCP-1/-3 failed to recruit mBMSCs, and consequently failed to promote callus formation. Taken together, our data suggest that MSCs produce a broad repertoire of trophic factors with tissue-regenerative activities that accelerate healing in the DO process.
Akihito Yamamoto, Kohki Matsubara, Fumiya Kano and Kiyoshi Sakai : Analysis of the neuroregenerative activities of mesenchymal stem cells in functional recovery after rat spinal cord injury., Methods in Molecular Biology, Vol.1213, 321-328, 2014.
(Summary)
Spinal cord injury (SCI) involves concurrent, interacting pathological processes, and requires a multifaceted therapeutic strategy. Stem cell-based transplantation holds great promise as such an approach. We have reported that stem cells derived from human dental pulp have remarkable neuroregenerative activity, and that when transplanted into animal models of SCI, these cells promote functional recovery by inhibiting massive SCI-induced apoptosis, preserving neural fibers and myelin, regenerating transected axons, and replacing damaged cells by differentiating into oligodendrocytes. Here, we introduce some details of our experimental procedures, which may serve as a guide for designing experiments to evaluate the therapeutic benefits of various types of stem cells.
Akihito Yamamoto, Kiyoshi Sakai, Kohki Matsubara, Fumiya Kano and Minoru Ueda : Multifaceted neuro-regenerative activities of human dental pulp stem cells for functional recovery after spinal cord injury., Neuroscience Research, Vol.78, 16-20, 2013.
(Summary)
Spinal cord injury (SCI) often leads to persistent functional deficits due to the loss of neurons and glia and to limited axonal regeneration after such injury. Recently, three independent groups have reported marked recovery of hindlimb locomotor function after the transplantation of human adult dental pulp stem cells (DPSCs) and stem cells from human exfoliated deciduous teeth (SHEDs) into rats or mice with acute, sub-acute or chronic SCI. This review summarizes the primary characteristics of human dental pulp stem cells and their therapeutic benefits for treating SCI. Experimental data from multiple preclinical studies suggest that pulp stem cells may promote functional recovery after SCI through multifaceted neuro-regenerative activities.
Mari Yamagata, Akihito Yamamoto, Eisuke Kako, Naoko Kaneko, Kohki Matsubara, Kiyoshi Sakai, Kazunobu Sawamoto and Minoru Ueda : Human dental pulp-derived stem cells protect against hypoxic-ischemic brain injury in neonatal mice., Stroke, Vol.44, No.2, 551-554, 2012.
(Summary)
SHED transplantation into the HI-injured brain resulted in remarkable neurological and pathophysiological recovery. Our findings indicate that paracrine factors derived from SHED support a neuroprotective microenvironment in the HI brain. SHED graft and SHED-conditioned medium may provide a novel neuroprotective therapy for HI.
Ryutaro Shohara, Akihito Yamamoto, Sachiko Takikawa, Akira Iwase, Hideharu Hibi, Fumitaka Kikkawa and Minoru Ueda : Mesenchymal stromal cells of human umbilical cord Wharton's jelly accelerate wound healing by paracrine mechanisms., Cytotherapy, Vol.14, No.10, 1171-1181, 2012.
(Summary)
Our results show that HUCPVC promotes wound healing via multifaceted paracrine mechanisms. Together with their ability to differentiate into the osteogenic linage, HUCPVC may provide significant therapeutic benefits for treating wounds in neonatal patients.
Kiyoshi Sakai, Akihito Yamamoto, Kohki Matsubara, Shoko Nakamura, Mami Naruse, Mari Yamagata, Kazuma Sakamoto, Ryoji Tauchi, Norimitsu Wakao, Shiro Imagama, Hideharu Hibi, Kenji Kadomatsu, Naoki Ishiguro and Minoru Ueda : Human dental pulp-derived stem cells promote locomotor recovery after complete transection of the rat spinal cord by multiple neuro-regenerative mechanisms., The Journal of Clinical Investigation, Vol.122, No.1, 80-90, 2011.
(Summary)
Spinal cord injury (SCI) often leads to persistent functional deficits due to loss of neurons and glia and to limited axonal regeneration after injury. Here we report that transplantation of human dental pulp stem cells into the completely transected adult rat spinal cord resulted in marked recovery of hind limb locomotor functions. Transplantation of human bone marrow stromal cells or skin-derived fibroblasts led to substantially less recovery of locomotor function. The human dental pulp stem cells exhibited three major neuroregenerative activities. First, they inhibited the SCI-induced apoptosis of neurons, astrocytes, and oligodendrocytes, which improved the preservation of neuronal filaments and myelin sheaths. Second, they promoted the regeneration of transected axons by directly inhibiting multiple axon growth inhibitors, including chondroitin sulfate proteoglycan and myelin-associated glycoprotein, via paracrine mechanisms. Last, they replaced lost cells by differentiating into mature oligodendrocytes under the extreme conditions of SCI. Our data demonstrate that tooth-derived stem cells may provide therapeutic benefits for treating SCI through both cell-autonomous and paracrine neuroregenerative activities.
Masahito Fujio, Akihito Yamamoto, Yuji Ando, Ryutaro Shohara, Kazuhiko Kinoshita, Tadashi Kaneko, Hideharu Hibi and Minoru Ueda : Stromal cell-derived factor-1 enhances distraction osteogenesis-mediated skeletal tissue regeneration through the recruitment of endothelial precursors., Bone, Vol.49, No.4, 693-700, 2011.
(Summary)
Distraction osteogenesis (DO) is a unique therapy that induces skeletal tissue regeneration without stem/progenitor cell transplantation. Although the self-regeneration property of DO provides many clinical benefits, the long treatment period required is a major drawback. A high-speed DO mouse model (H-DO), in which the distraction was done two times faster than in control DO (C-DO) mice, failed to generate new bone callus in the DO gap. We found that this was caused by the unsuccessful recruitment of bone marrow endothelial cells (BM-ECs)/endothelial progenitor cells (EPCs) into the gap. We then tested the ability of a local application of stromal cell-derived factor-1 (SDF-1), a major chemo-attractant for BM-ECs/EPCs, to accelerate the bone regeneration in H-DO. Our data showed that, in H-DO, SDF-1 induced callus formation in the gap through the recruitment of BM-ECs/EPCs, the maturation of neo-blood vessels, and increased blood flow. These results indicate that the active recruitment of endogenous BM-ECs/EPCs may provide a substantial clinical benefit for shortening the treatment period of DO.
Yingsong Zhu, Akiko Tsuchida, Akihito Yamamoto, Keiko Furukawa, Orie Tajima, Noriyo Tokuda, Shinichi Aizawa, Takeshi Urano, Kenji Kadomatsu and Koichi Furukawa : Expression and roles of a xenopus head-forming gene homologue in human cancer cell lines., Nagoya Journal of Medical Science, Vol.70, No.3-4, 73-82, 2008.
(Summary)
Molecular mechanisms for both morphogenesis and carcinogenesis have frequently overlapped, and similar signaling pathways are often involved in these processes. Yamamoto et al. identified a novel protein that induces head formation in Xenopus (Yamamoto et al. Cell, 120, 223-225, 2005). This new protein, named Shisa, plays unique roles in head formation by suppressing the maturation processes of receptors for Wnt and FGF at the endoplasmic reticulum. Here, we have identified a human homologue of the shisa gene (hu-shisa-2), and analyzed its expression in various human cancer cell lines by real-time reverse transcription polymerase chain reaction. High levels of mRNA expression were observed in some neuroectoderm-derived human cancer cell lines and small cell lung cancer cell lines. Intracellular localization of hu-Shisa-2 protein was also analyzed, indicating that it is present in the endoplasmic reticulum. Over-expression of hu-Shisa-2 resulted in increased cell growth and invasion, suggesting that hu-Shisa-2 is involved in the evolution and/or progression of human cancers.
Kenryo Furushima, Akihito Yamamoto, Takashi Nagano, Mikihito Shibata, Hitoshi Miyachi, Takaya Abe, Naoko Ohshima, Hiroshi Kiyonari and Shinichi Aizawa : Mouse homologues of Shisa antagonistic to Wnt and Fgf signalings., Developmental Biology, Vol.306, No.2, 480-492, 2007.
(Summary)
In an effort to identify Otx2 targets in mouse anterior neuroectoderm we identified a gene, mShisa, which is homologous to xShisa1 that we previously reported as a head inducer in Xenopus. mShisa encodes an antagonist against both Wnt and Fgf signalings; it inhibits these signalings cell-autonomously as xShisa1 does. The mShisa expression is lost or greatly reduced in Otx2 mutant visceral endoderm, anterior mesendoderm and anterior neuroectoderm. However, mShisa mutants exhibited no defects in head development. Shisa is composed of five subfamilies, but normal head development in mShisa mutants is unlikely to be explained in terms of the compensation of mShisa deficiency by its paralogues or by known Wnt antagonists in anterior visceral endoderm and/or anterior mesendoderm.
Takashi Nagano, Shoko Takehara, Maiko Takahashi, Shinichi Aizawa and Akihito Yamamoto : Shisa2 promotes the maturation of somitic precursors and transition to the segmental fate in Xenopus embryos., Development, Vol.133, No.23, 4643-4654, 2006.
(Summary)
In vertebrate somitogenesis, FGF and Wnt signals constitute a morphogenetic gradient that controls the maturation of the presomitic mesoderm (PSM) as well as the transition to segmental units. It remains unclear, however, whether there is a regulatory mechanism that promotes the transition by a direct regulation of FGF and Wnt signaling in the PSM. Here we show that Shisa2, a member of a novel Shisa gene family, plays an essential role in segmental patterning during Xenopus somitogenesis. Shisa2 encodes an endoplasmic reticulum (ER) protein that cell-autonomously inhibits FGF and Wnt signaling by preventing the maturation and the cell-surface expression of their receptors. Shisa2 is expressed in the PSM and its knockdown caused a reduction in somite number by the delayed maturation of PSM and anterior shift of the transition; however, the phase of the segmental clock remained intact. These phenotypes were abolished by the inhibition of both FGF and Wnt signals, but by neither alone. We therefore propose that the individual inhibition of both types of signaling by the regulation of receptor maturation in the ER plays an essential role in the establishment of proper segmental patterning.
Akihito Yamamoto, Takashi Nagano, Shoko Takehara, Masahiko Hibi and Shinichi Aizawa : Shisa promotes head formation through the inhibition of receptor protein maturation for the caudalizing factors, Wnt and FGF., Cell, Vol.120, No.2, 223-235, 2005.
(Summary)
Head formation requires simultaneous inhibition of multiple caudalizing signals during early vertebrate embryogenesis. We identified a novel antagonist against Wnt and FGF signaling for head formation, Shisa, which functions cell autonomously in the endoplasmic reticulum (ER). Shisa is specifically expressed in the prospective head ectoderm and the Spemann organizer of Xenopus gastrulae. Overexpression of Shisa inhibited both Wnt and FGF signaling in Xenopus embryos and in a cell line. Loss of Shisa function sensitized the neuroectoderm to Wnt signaling and suppressed head formation during gastrulation. Shisa physically interacted with immature forms of the Wnt receptor Frizzled and the FGF receptor within the ER and inhibited their posttranslational maturation and trafficking to the cell surface. Taken together, these findings indicate that Shisa is a novel molecule that controls head formation by regulating the establishment of the receptors for caudalizing factors.
Akihito Yamamoto, C Kemp, D Bachiller, D Geissert and M Robertis E De : Mouse paraxial protocadherin is expressed in trunk mesoderm and is not essential for mouse development., Genesis : the journal of genetics and development, Vol.27, No.2, 49-57, 2000.
(Summary)
Paraxial protocadherin (PAPC) is a cell adhesion molecule that marks cells undergoing convergence-extension cell movements in Xenopus and zebrafish gastrulating embryos. Here a mouse homologue (mpapc) was identified and characterized. During early- to mid-gastrulation, mpapc is expressed in the primitive streak as the trunk mesoderm undergoes morphogenetic cell movements. At head-fold stage mpapc expression becomes localized to paraxial regions in which somites are formed in the segmental plate. At later stages, mpapc displays a complex expression pattern in cerebral cortex, olfactory bulb, inferior colliculus, and in longitudinal stripes in hindbrain. To analyze the effect of the loss of PAPC function during mouse development, a null allele of the mouse papc gene was generated. Homozygous animals show no defects in their skeleton and are viable and fertile.
(Keyword)
Amino Acid Sequence / Animals / Cadherins / Embryonic and Fetal Development / Gene Expression Regulation, Developmental / Mesoderm / Mice / Molecular Sequence Data
(Link to Search Site for Scientific Articles)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 10890978
A Sawada, A Fritz, J Y Jiang, Akihito Yamamoto, K Yamasu, A Kuroiwa, Y Saga and H Takeda : Zebrafish Mesp family genes, mesp-a and mesp-b are segmentally expressed in the presomitic mesoderm, and Mesp-b confers the anterior identity to the developing somites., Development, Vol.127, No.8, 1691-1702, 2000.
(Summary)
Segmentation of a vertebrate embryo begins with the subdivision of the paraxial mesoderm into somites through a not-well-understood process. Recent studies provided evidence that the Notch-Delta and the FGFR (fibroblast growth factor receptor) signalling pathways are required for segmentation. In addition, the Mesp family of bHLH transcription factors have been implicated in establishing a segmental prepattern in the presomitic mesoderm. In this study, we have characterized zebrafish mesp-a and mesp-b genes that are closely related to Mesp family genes in other vertebrates. During gastrulation, only mesp-a is expressed in the paraxial mesoderm at the blastoderm margin. During the segmentation period, both genes are segmentally expressed in one to three stripes in the anterior parts of somite primordia. In fused somites (fss) embryos, in which all early somite boundary formation is blocked, initial mesp-a expression at the gastrula stage remains intact, but the expression of mesp-a and mesp-b is not detected during the segmentation period. This suggests that these genes are downstream targets of fss at the segmentation stage. Comparison with her1 expression (Müller, M., von Weizsäcker, E. and Campos-Ortega, J. A. (1996) Development 122, 2071-2078) suggests that, like her1, mesp genes are not expressed in primordia of the first several somites. Furthermore, we found that zebrafish her1 expression oscillates in the presomitic mesoderm. The her1 stripe, which first appears in the tailbud region, moves in a caudal to rostral direction, and it finally overlaps the most rostral mesp stripe. Thus, in the trunk region, both her1 and mesp transcripts are detected in every somite primordium posterior to the forming somites. Ectopic expression of Mesp-b in embryos causes a loss of the posterior identity within the somite primordium, leading to a segmentation defect. These embryos show a reduction in expression of the posterior genes, myoD and notch5, with uniform expression of the anterior genes, FGFR1, papc and notch6. These observations suggest that zebrafish mesp genes are involved in anteroposterior specification within the presumptive somites, by regulating the essential signalling pathways mediated by Notch-Delta and FGFR.
H S Kim, Akihito Yamamoto, T Bouwmeester, E Agius and M E Robertis : The role of paraxial protocadherin in selective adhesion and cell movements of the mesoderm during Xenopus gastrulation., Development, Vol.125, No.23, 4681-4690, 1998.
(Summary)
Paraxial Protocadherin (PAPC) encodes a transmembrane protein expressed initially in Spemann's organizer and then in paraxial mesoderm. Together with another member of the protocadherin family, Axial Protocadherin (AXPC), it subdivides gastrulating mesoderm into paraxial and axial domains. PAPC has potent homotypic cell adhesion activity in cell dissociation and reaggregation assays. Gain- and loss-of-function microinjection studies indicate that PAPC plays an important role in the convergence and extension movements that drive Xenopus gastrulation. Thus, PAPC is not only an adhesion molecule but also a component of the machinery that drives gastrulation movements in Xenopus. PAPC may provide a link between regulatory genes in Spemann's organizer and the execution of cell behaviors during morphogenesis.
K Takamiya, Akihito Yamamoto, K Furukawa, J Zhao, S Fukumoto, S Yamashiro, M Okada, M Haraguchi, M Shin, M Kishikawa, H Shiku, S Aizawa and K Furukawa : Complex gangliosides are essential in spermatogenesis of mice: possible roles in the transport of testosterone., Proceedings of the National Academy of Sciences of the United States of America, Vol.95, No.21, 12147-12152, 1998.
(Summary)
Mice, homozygous for disrupted ganglioside GM2/GD2 synthase (EC 2.4. 1.94) gene and lacking all complex gangliosides, do not display any major neurologic abnormalities. Further examination of these mutant mice, however, revealed that the males were sterile and aspermatogenic. In the seminiferous tubules of the mutant mice, a number of multinuclear giant cells and vacuolated Sertoli cells were observed. The levels of testosterone in the serum of these mice were very low, although testosterone production equaled that produced in wild-type mice. Testosterone was found to be accumulated in interstitial Leydig cells, and intratesticularly injected testosterone was poorly drained in seminiferous fluid in the mutant mice. These results suggested that complex gangliosides are essential in the transport of testosterone to the seminiferous tubules and bloodstream from Leydig cells. Our results provide insights into roles of gangliosides in vivo.
S Fukumoto, Akihito Yamamoto, T Hasegawa, K Abe, K Takamiya, M Okada, J Z Min, K Furukawa, H Miyazaki, Y Tsuji, G Goto, M Suzuki, H Shiku and K Furukawa : Genetic remodeling of gangliosides resulted in the enhanced reactions to the foreign substances in skin., Glycobiology, Vol.7, No.8, 1111-1120, 1997.
(Summary)
Several lines of transgenic mice with gangliosides GM2/GD2 synthase gene were established, and the expression levels of the transgene in brain, liver, spleen and thymus were analyzed by comparing with those in their litter mates. Among four tissues, brain and skin showed markedly high expression levels of the transgene in Northern blotting. Particularly, transgenic mice skin showed about 10-fold higher expression of GM2/GD2 synthase gene than the wild type mice skin. Therefore, alterations in the morphology, glycolipid components, and responses to the exogenous stimulations in the transgenic mice skin were examined. Gangliosides in the transgenic skin were dramatically converted from GM3 to GM1, whereas no morphological changes were observed. However, when skin flap test was performed with insertion of nylon membranes under the skin flaps, much stronger inflammatory reactions consisting of edema, marked thickness, and cell infiltration were observed in the transgenic mice compared with the wild type. Similar enhanced inflammatory reaction was also observed in the skin injected by silicon gel, and in the peritoneal reaction to the injected casein. Main cell population in these inflammatory reactions consisted of neutrophils, suggesting an increased sensitivity of neutrophils to chemotactic factors in the transgenic mice.
M Robertis E De, S Kim, L Leyns, S Piccolo, D Bachiller, E Agius, A J Belo, Akihito Yamamoto, A Hainski-Brousseau, B Brizuela, O Wessely, B Lu and T Bouwmeester : Patterning by genes expressed in Spemann's organizer., Cold Spring Harbor Symposia on Quantitative Biology, Vol.62, 169-175, 1997.
K Takamiya, Akihito Yamamoto, K Furukawa, S Yamashiro, M Shin, M Okada, S Fukumoto, M Haraguchi, N Takeda, K Fujimura, M Sakae, M Kishikawa, H Shiku, K Furukawa and S Aizawa : Mice with disrupted GM2/GD2 synthase gene lack complex gangliosides but exhibit only subtle defects in their nervous system., Proceedings of the National Academy of Sciences of the United States of America, Vol.93, No.20, 10662-10667, 1996.
(Summary)
Gangliosides, sialic acid-containing glycosphingolipids, are abundant in the vertebrate (mammalian) nervous system. Their composition is spatially and developmentally regulated, and gangliosides have been widely believed to lay essential roles in establishment of the nervous system, especially in neuritogenesis and synaptogenesis. However, this has never been tested directly. Here we report the generation of mice with a disrupted beta 1,4-N-acetylgalactosaminyltransferase (GM2/GD2 synthase; EC 2.4.1.92) gene. The mice lacked all complex gangliosides. Nevertheless, they did not show any major histological defects in their nervous systems or in gross behavior. Just a slight reduction in the neural conduction velocity from the tibial nerve to the somatosensory cortex, but not to the lumbar spine, was detected. These findings suggest that complex gangliosides are required in neuronal functions but not in the morphogenesis and organogenesis of the brain. The higher levels of GM3 and GD3 expressed in the brains of these mutant mice may be able to compensate for the lack of complex gangliosides.
Akihito Yamamoto, S Yamashiro, S Fukumoto, M Haraguchi, M Atsuta, H Shiku and K Furukawa : Site restricted and neuron dominant expression of alpha 2,8sialyltransferase gene in the adult mouse brain and retina., Glycoconjugate Journal, Vol.13, No.3, 471-480, 1996.
(Summary)
Gene expression of the alpha 2,8sialyltransferase (alpha 2,8S-T) responsible for GD3 synthesis in the adult mouse brain and retina was analysed by reverse transcription-polymerase chain reaction/Southern blotting (RT-PCR/Southern) and in situ hybridization. Among various portions of the brain, high levels of 9.5 kb mRNA were observed in the retina and midbrain. Results of RT-PCR/Southern did not necessarily correlate with the enzyme activities in the individual sites. In situ hybridization analysis revealed that this gene was characteristically expressed in the inner segment of photoreceptor cells, some nuclei in the midbrain, cranial nerve nuclei in the pons-medulla, Purkinje cells in the cerebellum, pyramidal cells of the hippocampus and granular cells of the dentate gyrus. In the retina, the alpha 2,8S-T gene was broadly expressed over the layers during development, and retained high expression levels in the photoreceptor cells of adult mice consistent with high expression of GD3. Destruction of neurons in the hippocampus and dentate gyrus by injection of kainic acid and colchicine respectively resulted in the disappearance of the hybridization signal, suggesting that the alpha 2,8S-T gene was mainly expressed by neurons.
(Keyword)
aging / Animals / Astrocytes / Base Sequence / Blotting, Southern / brain / Colchicine / DNA Primers / Embryonic and Fetal Development / Female / Gangliosides / Gene Expression Regulation, Developmental / Gene Expression Regulation, Enzymologic / In Situ Hybridization / Kainic Acid / Male / Mesencephalon / Mice / Mice, Inbred BALB C / Neurons / Organ Specificity / Polymerase Chain Reaction / RNA, Messenger / retina / Sialyltransferases
(Link to Search Site for Scientific Articles)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 8781978
Akihito Yamamoto, M Haraguchi, S Yamashiro, S Fukumoto, K Furukawa, K Takamiya, M Atsuta, H Shiku and K Furukawa : Heterogeneity in the expression pattern of two ganglioside synthase genes during mouse brain development., Journal of Neurochemistry, Vol.66, No.1, 26-34, 1996.
(Summary)
Gangliosides are synthesized by sequential catalytic reaction of multiple glycosyltransferases. GM2/GD2 synthase and GD3 synthase are key enzymes for ganglioside synthesis, because their relative activities regulate the main profiles of ganglioside expression. Mouse GD3 synthase (EC 2.4.99.8) cDNA was cloned by eukaryotic expression cloning, and its mRNA expression as well as that of GM2/GD2 synthase gene during the development of the mouse CNS was analyzed by using northern blotting, reverse transcription-polymerase chain reaction, and in situ hybridization. When brain tissue was analyzed as a whole mass, a typical pattern corresponding to the reported findings obtained by biochemical analyses was observed, i.e., high expression of GD3 synthase gene in the early stage and gradual increase of GM2/GD2 synthase gene expression in the late stage of the development. However, the results of in situ hybridization of these two genes revealed that the expression kinetics of these two genes were heterogeneous among various sites in the brain under development. These findings suggest that various expression patterns of the two genes reflect differences in the course of the development of individual sites, and also different ganglioside components are required in individual portions of the brain for development and maintenance of the function.
Akihito Yamamoto, S Yamashiro, K Takamiya, M Atsuta, H Shiku and K Furukawa : Diverse expression of beta 1,4-N-acetylgalactosaminyltransferase gene in the adult mouse brain., Journal of Neurochemistry, Vol.65, No.6, 2417-2424, 1995.
(Summary)
Among various tissues of mouse, beta 1,4-N-acetylgalactosaminyltransferase (GM2/GD2 synthase) gene is expressed predominantly in the brain. Further analysis of the gene expression in the mouse CNS was performed by northern blotting and by enzyme assays using extracts from various parts of the CNS. In situ hybridization was also done to investigate the distribution of cells generating GM2/GD2 synthase. In northern blots, diverse levels of the gene expression were observed, depending on the regions examined. By in situ hybridization, pyramidal cells in the hippocampus, granular cells in dentate gyrus and cerebral cortex, Purkinje cells in cerebellum, and mitral cells in the olfactory bulb expressed high levels of the mRNA; these results corresponded to the results obtained by northern blot. Enzyme levels in these sites were accordingly high. However, enzyme levels in certain areas with low mRNA intensities, such as thalamus and pons medulla, were higher than expected from the results of northern blotting. The significance of the high gene expression in certain areas for brain function and the reason for the discrepancy between mRNA level and enzyme activity in some regions are discussed.
(Keyword)
Animals / Blotting, Northern / Brain / Central Nervous System / Gangliosides / Gene Expression / In Situ Hybridization / Mice / Mice, Inbred BALB C / Mice, Inbred C57BL / N-Acetylgalactosaminyltransferases / RNA, Messenger / Tissue Distribution
(Link to Search Site for Scientific Articles)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 7595535
S Yamashiro, M Haraguchi, K Furukawa, K Takamiya, Akihito Yamamoto, Y Nagata, O K Lloyd, H Shiku and K Furukawa : Substrate specificity of beta 1,4-N-acetylgalactosaminyltransferase in vitro and in cDNA-transfected cells. GM2/GD2 synthase efficiently generates asialo-GM2 in certain cells., The Journal of Biological Chemistry, Vol.270, No.11, 6149-6155, 1995.
(Summary)
The substrate specificity of beta 1,4-N-acetylgalactosaminyltransferase has been analyzed using a fusion enzyme which consisted of the catalytic domain of the enzyme and the IgG binding domain of protein A, and also by extracts from cDNA transfectants. Both enzyme sources were capable of producing not only GM2 and GD2, but also asialo-GM2, GalNAc-sialylparagloboside, and Gal-NAc-GD1a from appropriate acceptors, although the efficiencies were at most 1-3% of those of GM2/GD2. The biological significance of these low specificities was studied with transient and stable transfectant cells. From the results of transient expression of the cDNA, asialo-GM2 expression appeared to inversely correlate with GM2 synthase levels in those lines. Consequently, GM2 seemed to be preferentially synthesized when both GM3 and lactosylceramide are available, and asialo-GM2 is synthesized in the absence of GM3 synthesis. However, the results of double immunostaining of CHO transfectants with anti-GM2 and anti-asialo-GM2 antibodies indicated that another factor may be involved in asialo-GM2 synthesis. From the in vitro assay using mixed acceptors, it was concluded that the presence of certain levels of GM2 might enhance the asialo-GM2 synthesis. These results suggest that even acceptors showing low efficiencies in vitro might be used in certain cells depending on the availability of precursors, expression levels of other gangliosides, as well as the kinetic properties of the enzyme, and the compartmentation of the glycosylation machineries in the cells.
(Keyword)
Animals / CHO Cells / Carbohydrate Conformation / Carbohydrate Sequence / Cricetinae / Gangliosides / Gene Expression / Glycolipids / Humans / Kinetics / L Cells (Cell Line) / Melanoma, Experimental / Mice / Molecular Sequence Data / N-Acetylgalactosaminyltransferases / Rats / Recombinant Fusion Proteins / Staphylococcal Protein A / Substrate Specificity / Time Factors / Transfection
(Link to Search Site for Scientific Articles)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 7890749
K Takamiya, Akihito Yamamoto, S Yamashiro, K Furukawa, M Haraguchi, M Okada, T Ikeda, H Shiku and K Furukawa : T cell receptor-mediated stimulation of mouse thymocytes induces up-regulation of the GM2/GD2 synthase gene., FEBS Letters, Vol.358, No.1, 79-83, 1995.
(Summary)
cDNA clones of the mouse GM2/GD2 synthase (EC 2.4.1.92) gene were isolated, and their analyses revealed that the protein has a type II transmembrane structure with 533 amino acids, which was very similar to the human homolog except for the mRNA size. The mRNA level in thymocytes dramatically increased after treatment with anti-CD3 monoclonal antibody, whereas it was not elevated when treated with prostaglandin E2. In situ hybridization showed an elevation of mRNA levels in medullar thymocytes, suggesting that T cell receptor-mediated signaling induces up-regulation of the GM2/GD2 synthase gene in mature thymocytes.
M Haraguchi, S Yamashiro, Akihito Yamamoto, K Furukawa, K Takamiya, O K Lloyd, H Shiku and K Furukawa : Isolation of GD3 synthase gene by expression cloning of GM3 alpha-2,8-sialyltransferase cDNA using anti-GD2 monoclonal antibody., Proceedings of the National Academy of Sciences of the United States of America, Vol.91, No.22, 10455-10459, 1994.
(Summary)
For the isolation of ganglioside GD3 synthase (EC 2.4.99.8) cDNA, we developed an expression cloning approach that used an anti-GD2 monoclonal antibody for selection. A host recipient cell line that we have named KF3027-Hyg5 was also utilized. This cell line expresses high levels of GM2 as well as GM3 but no GD3 or GD2 and was constructed from mouse B16 melanoma cells transfected with the polyoma large tumor antigen gene (KF3027) and the previously cloned beta-1,4-N-acetylgalactosaminyltransferase (EC 2.4.1.92) cDNA. Four rounds of transfection, monoclonal antibody 3F8 panning, and Hirt extraction resulted in the isolation of two cDNA clones, transfection of which directed the expression of GD3 in KF3027 and B16 melanoma cells and GD3 and GD2 in KF3027-Hyg5 cells. The cDNA contained a 1650-bp insert and a single open reading frame. The deduced amino acid predicted a type II membrane topology consisting of cytoplasmic (14 aa), transmembrane (18 aa), and catalytic (309 aa) domains. The sequence also predicted the presence of a sialyl motif similar to that found in the other sialyltransferases cloned so far. As expected, mRNA of this gene (2.6 kb) was strongly expressed in human melanoma lines.
K Yukawa, T Yasui, Akihito Yamamoto, H Shiku, T Kishimoto and H Kikutani : Epoc-1: a POU-domain gene expressed in murine epidermal basal cells and thymic stromal cells., Gene, Vol.133, No.2, 163-169, 1993.
(Summary)
POU-domain transcription factors are known as developmental regulators which control organ development and cell phenotypes. In order to clarify the roles of POU-domain transcription factors in cell differentiation, we cloned a novel POU family gene, Epoc-1, from a murine thymus cDNA library. The amino acid (aa) sequence of the POU-specific domain of Epoc-1 is almost identical to those of Oct-1 and Oct-2. However, within the POU-homeodomain, 13 out of 60 aa differ between Epoc-1 and Oct-2. Recombinant Epoc-1 products were found to bind specifically to the octamer sequence. Epoc-1 was found to be expressed in skin, thymus, stomach and testis. In situ hybridization experiments and RNase protection assays indicated that Epoc-1 is expressed in the epidermal basal cells of the skin, which contain stem cells unipotent for keratinocyte differentiation and in thymic stromal elements. These results suggest that Epoc-1 might be one of the developmental regulators which controls epidermal development and thymic organogenesis.
R Suto, H Udono, Akihito Yamamoto, H Shiku and E Nakayama : Effect of accessory cells on stimulation of murine T-cell leukemia with antibodies to the CD3/T cell antigen receptor complex., Gann : Japanese Journal of Cancer Research, Vol.84, No.4, 438-444, 1993.
(Summary)
Stimulation of EL4 and RL male 1 leukemia cells in vitro with immobilized anti-CD3 epsilon monoclonal antibody (mAb) (145-2C11) or anti-TCR beta mAb (H57-597) in the absence of accessory cells induced interleukin-2 (IL-2) production, and caused growth inhibition. The growth inhibition was, however, transient and the tumors started to grow again within 5 days in immobilizing plates treated with antibodies at concentrations of 2.5-100 micrograms/ml. Addition of mitomycin C-treated accessory cells to the culture inhibited IL-2 production and resulted in augmented and persistent growth inhibition. No recovery of tumor growth was observed. Furthermore, DNA from EL4 and RL male 1 leukemia cells stimulated with anti-CD3/TCR mAbs was fragmented even in the absence of accessory cells, but fragmentation was much greater in the presence of accessory cells. Marginal and high expression of the bcl-2 gene were observed in EL4 and RL male 1, respectively, indicating that apoptosis of these leukemias mediated by signalling through the CD3/TCR complex has no direct relationship with expression of the bcl-2 gene.
Akihito Yamamoto, M Atsuta and K Hamatani : Restricted expression of recombination activating gene (RAG-1) in mouse lymphoid tissues., Cell Biochemistry and Function, Vol.10, No.2, 71-77, 1992.
(Summary)
In an attempt to determine the distribution of recombinase activity in the mouse thymus, spleen and lymph nodes, we used the in situ hybridization method to examine the expression of the recombination activating genes RAG-1 and RAG-2. Expression of RAG-1 was found in most cortical thymocytes but not in the majority of medullary thymocytes. Although hybridization signals of RAG-2 were not as intense as those of RAG-1, the localization of RAG-2 transcripts was similar to that of RAG-1. In the spleen, expression of RAG-1 was found only in limited cells near the sinus, and the majority of the cells within the follicle were negative for RAG-1 transcript. In nude mice, RAG-1-expressing cells were detected in the same regions, which suggests that in situ hybridization signals of RAG-1 in the spleen are due to the cells of B cell origin. In the lymph nodes, expression of RAG-1 was found only in the medullary region. Expression of RAG-2 transcript in the spleen and the lymph nodes, if any, was too faint to determine the specific localization. These results suggest that most of the cortical thymocytes and some cells in the spleen are capable of rearranging T cell receptor genes and immunoglobulin genes, respectively, but the possibility of some other explanation could not be ruled out in RAG-1 expressing cells of the spleen and the lymph nodes.
Eiji Tanaka, YAO LIU, LINZE XIA, Naoko Ogasawara, Takuma Sakamaki, Fumiya Kano, Noboru Hashimoto, Xingmei Feng and Akihito Yamamoto : Effectiveness of low-intensity pulsed ultrasound on osteoarthritis of the temporomandibular joint: A review., Annals of Biomedical Engineering, Vol.48, No.8, 2158-2170, Jun. 2020.
(Summary)
Loading is indispensable for the growth, development, and maintenance of joint tissues, including mandibular condylar cartilage, but excessive loading or reduced host adaptive capacity can considerably damage the temporomandibular joint (TMJ), leading to temporomandibular joint osteoarthritis (TMJ-OA). TMJ-OA, associated with other pathological conditions and aging processes, is a highly degenerative disease affecting the articular cartilage. Many treatment modalities for TMJ-OA have been developed. Traditional clinical treatment includes mainly nonsurgical options, such as occlusal splints. However, non-invasive therapy does not achieve joint tissue repair and regeneration. Growing evidence suggests that low-intensity pulsed ultrasound (LIPUS) accelerates bone fracture healing and regeneration, as well as having extraordinary effects in terms of soft tissue repair and regeneration. The latter have received much attention, and various studies have been performed to evaluate the potential role of LIPUS in tissue regeneration including that applied to articular cartilage. The present article provides an overview of the status of LIPUS stimulation used to prevent the onset and progression of TMJ-OA and enhance the tissue regeneration of mandibular condylar cartilage. The etiology and management of TMJ-OA are explained briefly, animal models of TMJ-OA are described, and the effectiveness of LIPUS on cell metabolism and tissue regeneration in the TMJ is discussed.
Akihito Yamamoto, S Kiyoshi, M Kohki, Fumiya Kano and U Minoru : Multifaceted neuro-regenerative activities of human dental pulp stem cells for functional recovery after spinal cord injury., Neuroscience Research, Vol.78, 16-20, 2014.
Journal of Clinical and Experimental Medicine, Vol.239, No.8, 847-850, 2011.
5.
Akihito Yamamoto : Molecular basis for specification of the vertebrate head field, Interface Oral Health Science, 27-32, 2009.
6.
Akihito Yamamoto : Regulation of Wnt and FGF Signaling within the Endoplasmic Reticulum for Head Formation, Journal of Oral Biosciences, Vol.48, No.1, 18-21, 2006.
Fumiya Kano, M Kohki, U Minoru, H Hideharu and Akihito Yamamoto : Secreted Ectodomain of Sialic Acid-Binding Ig-Like Lectin-9 and Monocyte Chemoattractant Protein-1 Synergistically Regenerate Transected Rat Peripheral Nerves by Altering Macrophage Polarity., 13th World Congress on Inflammation., London, Jul. 2017.
2.
Jumpei Teramachi, Masahiro Hiasa, Oda Asuka, Hirofumi Tenshin, Ryota Amachi, Takeshi Harada, Shingen Nakamura, Hirokazu Miki, Itsuro Endo, Akihito Yamamoto, Tatsuji Haneji, Toshio Matsumoto and Masahiro Abe : Therapeutic impact of TAK1 inhibition on myeloma tumor progression and bone destruction, Cancer and Bone Society Conference 2017, Indianapolis, May 2017.
3.
Fumiya Kano, M Kohki, U Minoru, H Hideharu and Akihito Yamamoto : SECRETED ECTODOMAIN OF SIALIC ACID-BINDING IMMUNOGLOBULIN-LIKE LECTIN-9 AND MONOCYTE CHEMOATTRACTANT PROTEIN-1 DERIVED FROM DENTAL PULP STEM CELLS SYNERGISTICALLY REGENERATE TRANSECTED RAT PERIPHERAL NERVES BY ALTERING MACROPHAGE POLARITY., The 23th International Conference on Oral and Maxillofacial Surgery., Hong Kong, Mar. 2017.
4.
Fumiya Kano, M Kohki and Akihito Yamamoto : THE DEVELOPMENT OF NEW THERAPY FACIAL NERVE INJURY USING THE M2 MACROPHAGE INDUCTION FACTORS., The 22nd International Conference on Oral and Maxillofacial Surgery., Melbourne, Oct. 2015.
5.
Fumiya Kano, M Kohki and Akihito Yamamoto : THE DEVELOPMENT OF NEW THERAPY FACIAL NERVE INJURY USING THE M2 MACROPHAGE INDUCTION FACTORS., 12th World Congress on Inflammation, Boston, Aug. 2015.
6.
Fumiya Kano, M Kohki, Akihito Yamamoto and U Minoru : THE DEVELOPMENT OF NEW THERAPY FACIAL NERVE INJURY USING THE M2 MACROPHAGE INDUCTION FACTORS., AAOMS 96th Annual Meeting, Hawaii, Sep. 2014.
Proceeding of Domestic Conference:
1.
Wakana Oki, Fumiya Kano, Takaaki Nishihara, Noboru Hashimoto and Akihito Yamamoto : Therapeutic benefits of factors derived from stem cells from human exfoliated deciduous teeth for radiation-induced mouse xerostomia, 第64回歯科基礎医学会学術大会, Sep. 2022.
(Keyword)
歯髄幹細胞 / 放射線性口腔乾燥 / 培養上清
2.
Yoshiaki Sayama, Risa Morioka, Fumiya Kano, Noboru Hashimoto and Akihito Yamamoto : Development of a novel treatment for rheumatoid arthritis using soluble sialic acid binding lectin, 第76回NPO法人日本口腔科学会学術集会, Apr. 2022.
(Keyword)
Siglec-9 / rheumatoid arthritis
3.
Wakana Oki, Fumiya Kano, Takaaki Nishihara, Noboru Hashimoto and Akihito Yamamoto : Therapeutic benefits of factors derived from stem cells from human exfoliated deciduous teeth for radiation-induced mouse xerostomia, 第76回NPO法人日本口腔科学会学術集会, Apr. 2022.
(Keyword)
Dental pulp stem cell / Conditioned medium / xerostomia
4.
Risa Morioka, Yoshiaki Sayama, Fumiya Kano, Noboru Hashimoto and Akihito Yamamoto : 分泌型シアル酸認識レクチン-9とケモカインMCP-1 を用いた関節リウマチの新規治療法の開発, 四国歯学会第59回例会, Mar. 2022.
(Keyword)
Siglec-9 / MCP-1 / 関節リウマチ
5.
Waskitho Arief, Yumiko Yamamoto, Raman Swarna Lakshmi, Fumiya Kano, Huijiao Yan, Kazuo Okura, Daisuke Ikutame, Masamitsu Ohshima, Otto Baba, Akihito Yamamoto and Yoshizo Matsuka : Bilateral botulinum toxin type A effect on orofacial neuropathic pain of animal model, 徳島大学脳科学クラスターミニリトリート, Feb. 2022.
6.
LINZE XIA, Fumiya Kano, Noboru Hashimoto and Akihito Yamamoto : Conditioned Medium from the Stem Cells of Human Exfoliated Deciduous Teeth Ameliorates Experimental Temporomandibular Joint Osteoarthritis by Inducing M2 Phenotype of Macrophages, 蔵本免疫懇話会, Nov. 2021.
(Keyword)
Conditioned Medium / Stem Cells of Human Exfoliated Deciduous Teet / TMJ0A / M2macrophage
Fumiya Kano, YAO LIU, Noboru Hashimoto, Yoshizo Matsuka, Eiji Tanaka and Akihito Yamamoto : Conditioned Medium From the Stem Cells of Human Exfoliated Deciduous Teeth Ameliorates Neuropathic Pain in a Partial Sciatic Nerve Ligation Model, 第63回歯科基礎医学会学術大会, Sep. 2021.
(Keyword)
歯髄幹細胞 / 培養上清 / 神経障害性疼痛
9.
Takaaki Nishihara, Wakana Oki, Fumiya Kano, Noboru Hashimoto, Keita Kawarabayashi and Akihito Yamamoto : Therapeutic benefits of factors derived from stem cells from human exfoliated deciduous teeth for radiation-induced mouse xerostomia, 四国歯学会第57回例会, Mar. 2021.
(Keyword)
Dental pulp stem cell / Conditioned medium / xerostomia
10.
LINZE XIA, Fumiya Kano, Noboru Hashimoto, Naoko Ogasawara, YAO LIU, Eiji Tanaka and Akihito Yamamoto : :Analysis of the Therapeutic Mechanisms of SHED-CM for Experimentally Induced Temporomandibular Joint Osteoarthritis in Mice., Shikoku Dental Research, Mar. 2021.
11.
Fumiya Kano, Naoko Ogasawara, Noboru Hashimoto, Hiroki Mori, Aya Miyazaki, Tsutomu Iwamoto, Eiji Tanaka and Akihito Yamamoto : Factors secreted from dental pulp stem cells show multifaceted benefits for treating experimental temporomandibular joint osteoarthritis, 第62回歯科基礎医学会学術大会, Sep. 2020.
(Keyword)
TMJ-OA / SHED / SHED-CM
12.
Noboru Hashimoto, Fumiya Kano, Keita Kawarabayashi, YAO LIU, LINZE XIA and Akihito Yamamoto : ヒト乳歯歯髄幹細胞由来培養液における皮膚炎モデルマウス治療効果因子のプロテオーム解析, 第41回日本炎症・再生医学会, Jul. 2020.
13.
Fumiya Kano, YAO LIU, Noboru Hashimoto, Yoshizo Matsuka, Eiji Tanaka and Akihito Yamamoto : 歯髄幹細胞由来無血清培養上清は坐骨神経結紮モデルマウスの神経障害性疼痛を改善する, 日本炎症・再生医学会, Jul. 2020.
14.
Noboru Hashimoto, Fumiya Kano, Keita Kawarabayashi, YAO LIU and Akihito Yamamoto : ヒト乳歯歯髄幹細胞由来培養液における皮膚炎モデルマウス治療効果因子のプロテオーム解析, 第19回日本再生医療学会総会, May 2020.
15.
Fumiya Kano, Naoko Ogasawara, Noboru Hashimoto, Hiroki Mori, Aya Miyazaki, Tsutomu Iwamoto, Eiji Tanaka and Akihito Yamamoto : 歯髄幹細胞由来無血清培養上清を用いた変形性顎関節症の治療法開発, 第19回日本再生医療学会, 20, Mar. 2020.
Fumiya Kano, 西須 大徳, 神尾 尚伸, 鶴田 剛士, 坂口 晃平, 日比 英晴 and Akihito Yamamoto : A case of referred pain related to the antagonistic tooth by dental pulpitis progressed after crown restoration, 第24回日本口腔顔面痛学会学術大会, Sep. 2019.
23.
Fumiya Kano, 西須 大徳, 神尾 尚伸, 鶴田 剛士, 坂口 晃平, 日比 英晴 and Akihito Yamamoto : A case of referred pain related to the antagonistic tooth by dental pulpitis progressed after crown restoration, 第29 回日本口腔内科学会, Sep. 2019.
Fumiya Kano, Noboru Hashimoto and Akihito Yamamoto : Therapeutic benefits of factors derived from stem cells from human exfoliated deciduous teeth for radiation-induced mouse xerostomia, 四国免疫フォーラム, Jun. 2023.
Akihito Yamamoto : COMPOSITION FOR TREATMENT OF DAMEGED PART, US 16/043,395 (2018/7/24) (Jul. 2018), US 2018/0325946(2018/11/15) (Nov. 2018), US 2021/00000000 (Jul. 2021).
3.
Akihito Yamamoto : 組織修復活性組成物及びその利用, 15/938,839 (Mar. 2018), US 10,507,230 (Dec. 2019), US 10,507,230 (Dec. 2019).
Akihito Yamamoto : COMPOSITION HAVING TISSUE-REPAIRING ACTIVITY, AND USE THEREFOR, EP 13864639.3 (2013/12/24) (Dec. 2013), EP 2937088 (2015/10/28) (Oct. 2015), EP2937088A1 (Mar. 2018).
Elucidation of intracellular signaling mechanisms involved in cell fate determination through regulation of the extracellular environment (Project/Area Number: 23H03112 )
Analysis of joint regeneration mechanism by secretory sialic acid recognition lectin (Project/Area Number: 21H03055 )
Analysis of the mechanisms of organ formation by cell-cell and cell-extracellular communications (Project/Area Number: 20H03898 )
Novel therapeutic strategies for amyotrophic lateral sclerosis using human deciduous dental pulp stem cells and their active ingredients (Project/Area Number: 17K19666 )
Cell fate determination of mesenchymal stem cells by extracellular mechanical stress (Project/Area Number: 17H04414 )
Development of a novel treatment method for xerostomia by dental pulp stem cell-derived humoral factor (Project/Area Number: 17H04404 )
The co-transplantation of Islets with Dental Pulp Stem Cells Improved Survival and Function of Transplanted Beta-Cells (Project/Area Number: 16K09742 )
Bone regeneration with cell-derived factors in response to severe culture environment (Project/Area Number: 16H05540 )
Evaluation of the therapeutic benefits of anti-inflammatory M2 macrophage inducer for distraction osteogenesis (Project/Area Number: 15K15736 )
Effect of Stem Cells derived from Human Exfoliated Decidious Teeth in animal model of acute liver failure-correlation between inflammation and regeneration in liver (Project/Area Number: 15K08996 )
Development of vascular network construction method in vivo for enhancing tissue regeneration (Project/Area Number: 26670863 )
Development of regenerative therapy against severe peripheral nerve injury with a novel anti-inflammatory M2 macrophages inducer derived from dental pulp stem cells (Project/Area Number: 26293427 )
Development of the method for bone regeneration mimicing distraction osteogenesis for extensive bone defects without distraction devices (Project/Area Number: 25293408 )
Cell destruction and reloading treatment for bone graft in oral cancer surgery (Project/Area Number: 23659942 )
Analysis of molecular forces for stem/progenitor cell homing in bone regeneration (Project/Area Number: 23659915 )
Development of new regenerative therapy for central nerve system using the conditioned media derived from human dental pulp stem cells (Project/Area Number: 23659914 )
Bone regeneration following vascular network formation (Project/Area Number: 22390384 )
Stem cells from human-exfoliated deciduous teeth can regenerate central nervous system (Project/Area Number: 22390372 )
Analysis of benefits of dental pulp stem cells from permanent and milk teeth for the treatment of the intractable diseases and of their banking system. (Project/Area Number: 21390524 )