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Zhao, Juanjuan ; Okamoto, Yasuo ; Asano, Yuya ; Ishimaru, Kazuhiro ; Aki, Sho ; Yoshioka, Kazuaki ; Takuwa, Noriko ; Wada, Takashi ; Inagaki, Yutaka ; Takahashi, Chiaki ; Nishiuchi, Takumi ; Takuwa, Yoh ; 安藝, 翔 ; 吉岡, 和晃 ; 多久和, 典子 ; 和田, 隆志 ; 髙橋, 智聡 ; 西内, 巧 ; 多久和, 陽
出版情報: PLoS ONE.  13  pp.e0197604-,  2018-05-21.  Public Library of Science
URL: http://hdl.handle.net/2297/00053881
概要: 金沢大学医薬保健研究域医学系<br />Idiopathic pulmonary fibrosis is a devastating disease with poor prognosis. The pathogenic role of t he lysophospholipid mediator sphingosine-1-phosphate and its receptor S1PR2 in lung fibrosis is unknown. We show here that genetic deletion of S1pr2 strikingly attenuated lung fibrosis induced by repeated injections of bleomycin in mice. We observed by using S1pr2 LacZ/+ mice that S1PR2 was expressed in alveolar macrophages, vascular endothelial cells and alveolar epithelial cells in the lung and that S1PR2-expressing cells accumulated in the fibrotic legions. Bone marrow chimera experiments suggested that S1PR2 in bone marrow–derived cells contributes to the development of lung fibrosis. Depletion of macrophages greatly attenuated lung fibrosis. Bleomycin administration stimulated the mRNA expression of the profibrotic cytokines IL-13 and IL-4 and the M2 markers including arginase 1, Fizz1/Retnla, Ccl17 and Ccl24 in cells collected from broncho-alveolar lavage fluids (BALF), and S1pr2 deletion markedly diminished the stimulated expression of these genes. BALF cells from bleomycin–administered wild-type mice showed a marked increase in phosphorylation of STAT6, a transcription factor which is activated downstream of IL-13, compared with saline–administered wild-type mice. Interestingly, in bleomycin–adminis-tered S1pr2 -/- mice, STAT6 phosphorylation in BALF cells was substantially diminished compared with wild-type mice. Finally, pharmacological S1PR2 blockade in S1pr2 +/+ mice alleviated bleomycin–induced lung fibrosis. Thus, S1PR2 facilitates lung fibrosis through the mechanisms involving augmentation of IL-13 expression and its signaling in BALF cells, and represents a novel target for treating lung fibrosis. © 2018 Zhao et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 続きを見る
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Pham, Hoa Q. ; Yoshioka, Kazuaki ; Mohri, Hiromi ; Nakata, Hiroki ; Aki, Sho ; Ishimaru, Kazuhiro ; Takuwa, Noriko ; Takuwa, Yoh ; 吉岡, 和晃 ; 仲田, 浩規 ; 安藝, 翔 ; 多久和, 典子 ; 多久和, 陽
出版情報: Genes Cells.  23  pp.670-687,  2018-08.  John Wiley & Sons
URL: http://hdl.handle.net/2297/00053882
概要: 金沢大学医薬保健研究域医学系<br />Phosphatidylinositol 3-phosphate (PI(3)P) is the predominant phosphoinositide species in early endos omes and autophagosomes, in which PI(3)P dictates traffic of these organelles. Phosphoinositide levels are tightly regulated by lipid-kinases and -phosphatases; however, a phosphatase that converts PI(3)P back to phosphatidylinositol in the endosomal and autophagosomal compartments is not fully understood. We investigated the subcellular distribution and functions of myotubularin-related protein-4 (MTMR4), which is distinct among other MTMRs in that it possesses a PI(3)P-binding FYVE domain, in lung alveolar epithelium-derived A549 cells. MTMR4 was localized mainly in late endosomes and autophagosomes. MTMR4 knockdown markedly suppressed the motility, fusion, and fission of PI(3)P-enriched structures, resulting in decreases in late endosomes, autophagosomes, and lysosomes, and enlargement of PI(3)P-enriched early and late endosomes. In amino acid- and serum-starved cells, MTMR4 knockdown decreased both autophagosomes and autolysosomes and markedly increased PI(3)P-containing autophagosomes and late endosomes, suggesting that the fusion with lysosomes of autophagosomes and late endosomes might be impaired. Notably, MTMR4 knockdown inhibited the nuclear translocation of starvation stress responsive transcription factor-EB (TFEB) with reduced expression of lysosome-related genes in starved cells. These findings indicate that MTMR4 is essential for the integrity of endocytic and autophagic pathways. © 2018 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.<br />Embargo Period 12 months 続きを見る
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Aung, Khin Thuzar ; Yoshioka, Kazuaki ; Aki, Sho ; Ishimaru, Kazuhiro ; Takuwa, Noriko ; Takuwa, Yoh ; 吉岡, 和晃 ; 安藝, 翔 ; 多久和, 典子 ; 多久和, 陽
出版情報: Journal of Physiological Sciences.  69  pp.263-280,  2019-03-01.  Physiological Society of Japan 日本生理学会 / Springer Nature
URL: http://hdl.handle.net/2297/00053883
概要: 金沢大学医薬保健研究域医学系<br />Pinocytosis is an important fundamental cellular process that is used by the cell to transport fluid and solutes. Phosphoinositide 3-kinases (PI3Ks) regulate a diverse array of dynamic membrane events. However, it is not well-understood which PI3K isoforms are involved in specific mechanisms of pinocytosis. We performed knockdown studies of endogenous PI3K isoforms and clathrin heavy chain (CHC) mediated by small interfering RNA (siRNA). The results demonstrated that the class II PI3K PI3K-C2α and PI3K-C2β, but not the class I or III PI3K, were required for pinocytosis, based on an evaluation of fluorescein-5-isothiocyanate (FITC)–dextran uptake in endothelial cells. Pinocytosis was partially dependent on both clathrin and dynamin, and both PI3K-C2α and PI3K-C2β were required for clathrin-mediated—but not clathrin-non-mediated—FITC-dextran uptake at the step leading up to its delivery to early endosomes. Both PI3K-C2α and PI3K-C2β were co-localized with clathrin-coated pits and vesicles. However, PI3K-C2β, but not PI3K-C2α, was highly co-localized with actin filament-associated clathrin-coated structures and required for actin filament formation at the clathrin-coated structures. These results indicate that PI3K-C2α and PI3K-C2β play differential, indispensable roles in clathrin-mediated pinocytosis. © 2018, The Physiological Society of Japan and Springer Japan KK, part of Springer Nature.<br />Embargo Period 12 months 続きを見る
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Sarker, Md Azadul Kabir ; Aki, Sho ; Yoshioka, Kazuaki ; Kuno, Kouji ; Okamoto, Yasuo ; Ishimaru, Kazuhiro ; Takuwa, Noriko ; Takuwa, Yoh ; 安藝, 翔 ; 吉岡, 和晃 ; 多久和, 典子 ; 多久和, 陽
出版情報: Endocrinology.  160  pp.235-248,  2019-01-01.  Endocrine Society / Oxford University Press
URL: http://hdl.handle.net/2297/00053884
概要: 金沢大学医薬保健研究域医学系<br />Class II phosphoinositide 3-kinases (PI3Ks), PI3K-C2α and PI3K-C2β, are highly homologous and distin ct from class I and class III PI3Ks in catalytic products and domain structures. In contrast to class I and class III PI3Ks, physiological roles of PI3K-C2α and PI3K-C2β are not fully understood. Because we previously demonstrated that PI3K-C2α is involved in vascular smooth muscle contraction, we studied the phenotypes of smooth muscle-specific knockout (KO) mice of PI3K-C2α and PI3K-C2β. The pup numbers born from single PI3K-C2α-KO and single PI3K-C2β-KO mothers were similar to those of control mothers, but those from double KO (DKO) mothers were smaller compared with control mice. However, the number of intrauterine fetuses in pregnant DKO mothers was similar to that in control mice. Both spontaneous and oxytocin-induced contraction of isolated uterine smooth muscle (USM) strips was diminished in DKO mice but not in either of the single KO mice, compared with control mice. Furthermore, contraction of USM of DKO mice was less sensitive to a Rho kinase inhibitor. Mechanistically, the extent of oxytocin-induced myosin light chain phosphorylation was greatly reduced in USM from DKO mice compared with control mice. The oxytocin-induced rise in the intracellular Ca2+ concentration in USM was similar in DKO and control mice. However, Rho activation in the intracellular compartment was substantially attenuated in DKO mice compared with control mice, as evaluated by fluorescence resonance energy transfer imaging technique. These data indicate that both PI3K-C2α and PI3K-C2β are required for normal USM contraction and parturition mainly through their involvement in Rho activation.<br />Embargo Period 12 months 続きを見る
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Qia, Xun ; Okamoto, Yasuo ; Murakawa, Tomomi ; Wang, Fei ; Oyama, Osamu ; Ohkawa, Ryunosuke ; Yoshioka, Kazuaki ; Du, Wa ; Sugimoto, Naotoshi ; Yatomi, Yutaka ; Takuwa, Noriko ; Takuwa, Yoh
出版情報: European Journal of Pharmacology.  634  pp.121-131,  2010-05-01.  Elsevier BV
URL: http://hdl.handle.net/2297/23922
概要: 金沢大学医薬保健研究域医学系<br />Therapeutic angiogenesis is a promising strategy for treating ischemia. The lysophospholipid mediato r sphingosine-1-phosphate (S1P) acts on vascular endothelial cells to stimulate migration and tube formation, and plays the critical role in developmental angiogenesis. We developed poly(lactic-co-glycolic-acid) (PLGA)-based S1P-containing microparticles (PLGA-S1P), which are biodegradable and continuously release S1P, and studied the effects of PLGA-S1P on neovascularization in murine ischemic hindlimbs. Intramuscular injections of PLGA-S1P stimulated blood flow in C57BL/6 mice dose-dependently, with repeated administrations at a 3-day interval, rather than a single bolus or 6-day interval, over 28. days conferring the optimal stimulating effect. In Balb/c mice that exhibit limb necrosis and dysfunction due to retarded blood flow recovery, injections of PLGA-S1P stimulated blood flow with alleviation of limb necrosis and dysfunction. PLGA-S1P alone did not induce edema in ischemic limbs, and rather blocked vascular endothelial growth factor-induced edema. PLGA-S1P not only increased the microvessel densities in ischemic muscle, but promoted coverage of vessels with smooth muscle cells and pericytes, thus stabilizing vessels. PLGA-S1P stimulated Akt and ERK with increased phosphorylation of endothelial nitric oxide synthase in ischemic muscle. The effects of the nitric oxide synthase inhibitor, Nω-nitro-l-arginine methylester, showed that PLGA-S1P-induced blood flow stimulation was partially dependent on nitric oxide. Injections of PLGA-S1P also increased the expression of angiogenic factors and the recruitment of CD45-, CD11b- and Gr-1-positive myeloid cells, which are implicated in post-ischemic angiogenesis, into ischemic muscle. These results indicate that PLGA-based, sustained local delivery of S1P is a potentially useful therapeutic modality for stimulating post-ischemic angiogenesis. © 2010 Elsevier B.V. 続きを見る
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Takuwa, Yoh ; Okamoto, Yasuo ; Yoshioka, Kazuaki ; Takuwa, Noriko
出版情報: BBA - Molecular and Cell Biology of Lipids.  781  pp.483-488,  2008-09-01.  Elsevier
URL: http://hdl.handle.net/2297/11734
概要: 金沢大学医薬保健研究域医学系<br />The plasma lysophospholipid mediator sphingosine-1-phosphate (S1P) is produced exclusively by sphing osine kinase (SPHK) 1 and SPHK2 in vivo, and plays diverse biological and pathophysiological roles by acting largely through three members of the G protein-coupled S1P receptors, S1P1, S1P2 and S1P3. S1P1 expressed on endothelial cells mediates embryonic vascular maturation and maintains vascular integrity by contributing to eNOS activation, inhibiting vascular permeability and inducing endothelial cell chemotaxis via Gi-coupled mechanisms. By contrast, S1P2, is expressed in high levels on vascular smooth muscle cells (VSMCs) and certain types of tumor cells, inhibiting Rac and cell migration via a G12/13-and Rho-dependent mechanism. In rat neointimal VSMCs, S1P1 is upregulated to mediate local production of platelet-derived growth factor, which is a key player in vascular remodeling. S1P3 expressed on endothelial cells also mediates chemotaxis toward S1P and vasorelaxation via NO production in certain vascular bed, playing protective roles for vascular integrity. S1P3 expressed on VSMCs and cardiac sinoatrial node cells mediates vasopressor and negative chronotropic effect, respectively. In addition, S1P3, together with S1P2 and SPHK1, is suggested to play a protective role against acute myocardial ischemia. However, our recent work indicates that overexpressed SPHK1 is involved in cardiomyocyte degeneration and fibrosis in vivo, in part through S1P activation of the S1P3 signaling. We also demonstrated that exogenously administered S1P accelerates neovascularization and blood flow recovery in ischemic limbs, suggesting its usefulness for angiogenic therapy. These results provide evidence for S1P receptor subtype-specific pharmacological intervention as a novel therapeutic approach to cardiovascular diseases and cancer. © 2008 Elsevier B.V. All rights reserved. 続きを見る
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Du, Wa ; Takuwa, Noriko ; Yoshioka, Kazuaki ; Okamoto, Yasuo ; Gonda, Koichi ; Sugihara, Kazushi ; Fukamizu, Akiyoshi ; Asano, Masahide ; Takuwa, Yoh
出版情報: Cancer Research.  70  pp.772-781,  2010-01-15.  American Association for Cancer Research
URL: http://hdl.handle.net/2297/21765
概要: 金沢大学医薬保健研究域医学系<br />Sphingosine-1-phosphate (S1P) has been implicated in tumor angiogenesis by acting through the Gi-cou pled chemotactic receptor S1P1. Here, we report that the distinct receptor S1P2 is responsible for mediating the G12/13/Rho-dependent inhibitory effects of S1P on Akt, Rac, and cell migration, thereby negatively regulating tumor angiogenesis and tumor growth. By using S1P2LacZ/+ mice, we found that S1P2 was expressed in both tumor and normal blood vessels in many organs, in both endothelial cells (EC) and vascular smooth muscle cells, as well as in tumor-associated, CD11b-positive bone marrow-derived cells (BMDC). Lewis lung carcinoma or B16 melanoma cells implanted in S1P2-deficient (S1P2-/-) mice displayed accelerated tumor growth and angiogenesis with enhanced association of vascular smooth muscle cells and pericytes. S1P2-/- ECs exhibited enhanced Rac activity, Akt phosphorylation, cell migration, proliferation, and tube formation in vitro. Coinjection of S1P2-/- ECs and tumor cells into wild-type mice also produced a relative enhancement of tumor growth and angiogenesis in vivo. S1P2-/- mice were also more efficient at recruiting CD11b-positive BMDCs into tumors compared with wild-type siblings. Bone marrow chimera experiments revealed that S1P2 acted in BMDCs to promote tumor growth and angiogenesis. Our results indicate that, in contrast to endothelial S1P1, which stimulates tumor angiogenesis, S1P 2 on ECs and BMDCs mediates a potent inhibition of tumor angiogenesis, suggesting a novel therapeutic tactic for anticancer treatment. ©2010 AACR. 続きを見る
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Wang, Fei ; Okamoto, Yasuo ; Inoki, Isao ; Yoshioka, Kazuaki ; Du, Wa ; Qi, Xun ; Takuwa, Noriko ; Gonda, Koichi ; Yamamoto, Yasuhiko ; Ohkawa, Ryunosuke ; Nishiuchi, Takumi ; Sugimoto, Naotoshi ; Yatomi, Yutaka ; Mitsumori, Kunitoshi ; Asano, Masahide ; Kinoshita, Makoto ; Takuwa, Yoh
出版情報: The journal of clinical investigation.  120  pp.3979-3995,  2010-11-01.  American Society for Clinical Investigation
URL: http://hdl.handle.net/2297/25352
概要: 金沢大学医薬保健研究域医学系<br />Sphingosine-1-phosphate (S1P) is a biologically active sphingolipid that has pleiotropic effects in a variety of cell types including ECs, SMCs, and macrophages, all of which are central to the development of atherosclerosis. It may therefore exert stimulatory and inhibitory effects on atherosclerosis. Here, we investigated the role of the S1P receptor S1PR2 in atherosclerosis by analyzing S1pr2–/– mice with an Apoe–/– background. S1PR2 was expressed in macrophages, ECs, and SMCs in atherosclerotic aortas. In S1pr2–/–Apoe–/– mice fed a high-cholesterol diet for 4 months, the area of the atherosclerotic plaque was markedly decreased, with reduced macrophage density, increased SMC density, increased eNOS phosphorylation, and downregulation of proinflammatory cytokines compared with S1pr2+/+Apoe–/– mice. Bone marrow chimera experiments indicated a major role for macrophage S1PR2 in atherogenesis. S1pr2–/–Apoe–/– macrophages showed diminished Rho/Rho kinase/NF-κB (ROCK/NF-κB) activity. Consequently, they also displayed reduced cytokine expression, reduced oxidized LDL uptake, and stimulated cholesterol efflux associated with decreased scavenger receptor expression and increased cholesterol efflux transporter expression. S1pr2–/–Apoe–/– ECs also showed reduced ROCK and NF-κB activities, with decreased MCP-1 expression and elevated eNOS phosphorylation. Pharmacologic S1PR2 blockade in S1pr2+/+Apoe–/– mice diminished the atherosclerotic plaque area in aortas and modified LDL accumulation in macrophages. We conclude therefore that S1PR2 plays a critical role in atherogenesis and may serve as a novel therapeutic target for atherosclerosis. 続きを見る
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Tanahashi, Hiroshi ; Yoshioka, Kazuaki
出版情報: Neuroscience Letters.  439  pp.293-297,  2008-07-18.  Elsevier
URL: http://hdl.handle.net/2297/10014
概要: 金沢大学医薬保健研究域医学系<br />In a previous study, we reported that Alzheimer's disease-associated presenilin-2 interacts with a L IM-domain protein, namely, DRAL/FHL2/SLIM3. In this study, we investigated whether DRAL modifies the metabolism of the amyloid precursor protein (APP). We used small interfering RNA (siRNA) to knockdown DRAL in COS7 and HEK293 cells that stably overexpress APP695. We found that the knockdown was accompanied by a decrease in the amount of secreted α-secretase-cleaved APP and the membrane-bound C-terminal fragment C83 and an increase in the amount of secreted β-amyloid peptide (Aβ) from the cells. We also found that in addition to a disintegrin and metalloprotease (ADAM)-17, DRAL binds to ADAM-10. Thus, DRAL may be involved in the processing of APP through the α-secretase pathway. © 2008 Elsevier Ireland Ltd. All rights reserved. 続きを見る
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Takuwa, Yoh ; Okamoto, Yasuo ; Yoshioka, Kazuaki ; Takuwa, Noriko
出版情報: Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids.  1781  pp.483-488,  2008-09-01.  Elsevier
URL: http://hdl.handle.net/2297/11867
概要: 金沢大学医薬保健研究域医学系<br />The plasma lysophospholipid mediator sphingosine-1-phosphate (S1P) is produced exclusively by sphing osine kinase (SPHK) 1 and SPHK2 in vivo, and plays diverse biological and pathophysiological roles by acting largely through three members of the G protein-coupled S1P receptors, S1P1, S1P2 and S1P3. S1P1 expressed on endothelial cells mediates embryonic vascular maturation and maintains vascular integrity by contributing to eNOS activation, inhibiting vascular permeability and inducing endothelial cell chemotaxis via Gi-coupled mechanisms. By contrast, S1P2, is expressed in high levels on vascular smooth muscle cells (VSMCs) and certain types of tumor cells, inhibiting Rac and cell migration via a G12/13-and Rho-dependent mechanism. In rat neointimal VSMCs, S1P1 is upregulated to mediate local production of platelet-derived growth factor, which is a key player in vascular remodeling. S1P3 expressed on endothelial cells also mediates chemotaxis toward S1P and vasorelaxation via NO production in certain vascular bed, playing protective roles for vascular integrity. S1P3 expressed on VSMCs and cardiac sinoatrial node cells mediates vasopressor and negative chronotropic effect, respectively. In addition, S1P3, together with S1P2 and SPHK1, is suggested to play a protective role against acute myocardial ischemia. However, our recent work indicates that overexpressed SPHK1 is involved in cardiomyocyte degeneration and fibrosis in vivo, in part through S1P activation of the S1P3 signaling. We also demonstrated that exogenously administered S1P accelerates neovascularization and blood flow recovery in ischemic limbs, suggesting its usefulness for angiogenic therapy. These results provide evidence for S1P receptor subtype-specific pharmacological intervention as a novel therapeutic approach to cardiovascular diseases and cancer. © 2008 Elsevier B.V. All rights reserved. 続きを見る
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Takashima, Shinichiro ; Sugimoto, Naotoshi ; Takuwa, Noriko ; Okamoto, Yasuo ; Yoshioka, Kazuaki ; Takamura, Masayuki ; Takata, Shigeo ; Kaneko, Shuichi ; Takuwa, Yoh
出版情報: Cardiovascular Research.  79  pp.689-697,  2008-09-01.  Elsevier
URL: http://hdl.handle.net/2297/12043
概要: 金沢大学医薬保健研究域医学系<br />Aims: The lysophospholipid mediator sphingosine-1-phosphate (S1P) activates G protein-coupled recept ors (GPCRs) to induce potent inhibition of platelet-derived growth factor (PDGF)-induced Rac activation and, thereby, chemotaxis in rat vascular smooth muscle cells (VSMCs). We explored the heterotrimeric G protein and the downstream mechanism that mediated S1P inhibition of Rac and cell migration in VSMCs. Methods and results: S1P inhibition of PDGF-induced cell migration and Rac activation in VSMCs was abolished by the selective S1P2 receptor antagonist JTE-013. The C-terminal peptides of Gα subunits (Gα-CTs) act as specific inhibitors of respective G protein-GPCR coupling. Adenovirus-mediated expression of Gα12-CT, Gα13-CT, and Gα q-CT, but not that of Gαs-CT or LacZ or pertussis toxin treatment, abrogated S1P inhibition of PDGF-induced Rac activation and migration, indicating that both G12/13 and Gq classes are necessary for the S1P inhibition. The expression of Gαq-CT as well as Gα12-CT and Gα13-CT also abolished S1P-induced Rho stimulation. C3 toxin, but not a Rho kinase inhibitor or a dominant negative form of Rho kinase, abolished S1P inhibition of PDGF-induced Rac activation and cell migration. The angiotensin II receptor AT1, which robustly couples to Gq, did not mediate either Rho activation or inhibition of PDGF-induced Rac activation or migration, suggesting that activation of Gq alone was not sufficient for Rho activation and resultant Rac inhibition. However, the AT1 receptor fused to Gα12 was able to induce not only Rho stimulation but also inhibition of PDGF-induced Rac activation and migration. Phospholipase C inhibition did not affect S1P-induced Rho activation, and protein kinase C activation by a phorbol ester did not mimic S1P action, suggesting that S1P inhibition of migration or Rac was not dependent on the phospholipase C pathway. Conclusion: These observations together suggest that S1P2 mediates inhibition of Rac and migration through the coordinated action of G 12/13 and Gq for Rho activation in VSMCs. © The Author 2008.. 続きを見る
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Seok, Young Mi ; Azam, Mohammed Ali ; Okamoto, Yasuo ; Sato, Atsushi ; Yoshioka, Kazuaki ; Maeda, Masataka ; Kim, In Kyeom ; Takuwa, Yoh
出版情報: Hypertension.  56  pp.934-941,  2010-11-01.  American Heart Association
URL: http://hdl.handle.net/2297/25788
概要: 金沢大学医薬保健研究域医学系<br />Rho-mediated inhibition of myosin light chain (MLC) phosphatase (MLCP), together with Ca-dependent M LC kinase activation, constitutes the major signaling mechanisms for vascular smooth muscle contraction. We recently unveiled the involvement of Ca-induced, phosphoinositide 3-kinase (PI3K) class IIα isoform (PI3K-C2α)-dependent Rho activation and resultant Rho kinase-dependent MLCP suppression in membrane depolarization- and receptor agonist-induced contraction. It is unknown whether Ca- and PI3K-C2α- dependent regulation of MLCP is altered in vascular smooth muscle of hypertensive animals and is involved in hypertension. Therefore, we studied the role of the Ca-PI3K-C2α-Rho-MLCP pathway in spontaneously hypertensive rats (SHRs). PI3K-C2α was readily detected in various vascular beds of Wistar-Kyoto rats and activated by high KCl. High KCl also stimulated vascular Rho activity and phosphorylation of the MLCP regulatory subunit MYPT1 at Thr in a PI3K inhibitor wortmannin-sensitive manner. In mesenteric and other vessels of SHRs at the hypertensive but not the prehypertensive stage, the activity of PI3K-C2α but not class I PI3K p110α was elevated with concomitant rises of Rho activity and Thr-phosphorylation of MYPT1, as compared with normotensive controls. Infusion of the Ca channel antagonist nicardipine reduced blood pressure with suppression of vascular activity of PI3K-C2α-Rho and phosphorylation of MYPT1 in hypertensive SHRs. Infusion of wortmannin lowered blood pressure with inhibition of PI3K-C2α-Rho activities and MYPT1 phosphorylation in hypertensive SHRs. These observations suggest that an increased activity of the Ca-PI3K-C2α-Rho signaling pathway with resultant augmented MLCP suppression contributes to hypertension in SHRs. The Ca- and PI3K-C2α-dependent Rho stimulation in vascular smooth muscle may be a novel, promising target for treating hypertension. © 2010 American Heart Association, Inc. 続きを見る
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Takuwa, Noriko ; Ohkura, Sei-Ichiro ; Takashima, Shin-ichiro ; Ohtani, Keisuke ; Okamoto, Yasuo ; Tanaka, Tamotsu ; Hirano, Kaoru ; Usui, Soichiro ; Wang, Fei ; Du, Wa ; Yoshioka, Kazuaki ; Banno, Yoshiko ; Sasaki, Motoko ; Ichi, Ikuyo ; Okamura, Miwa ; Sugimoto, Naotoshi ; Mizugishi, Kiyomi ; Nakanuma, Yasuni ; Ishii, Isao ; Takamura, Masayuki ; Kaneko, Shuichi ; Kojo, Shosuke ; Satouchi, Kiyoshi ; Mitumori, Kunitoshi ; Chun, Jerold ; Takuwa, Yoh
出版情報: Cardiovascular Research.  85  pp.484-493,  2010-02-01.  Oxford University Press (OUP)
URL: http://hdl.handle.net/2297/21766
概要: 金沢大学医薬保健研究域医学系<br />Aims Sphingosine kinase 1 (SPHK1), its product sphingosine-1-phosphate (S1P), and S1P receptor subty pes have been suggested to play protective roles for cardiomyocytes in animal models of ischaemic preconditioning and cardiac ischaemia/reperfusion injury. To get more insight into roles for SPHK1 in vivo, we have generated SPHK1-transgenic (TG) mice and analysed the cardiac phenotype.Methods and results SPHK1-TG mice overexpressed SPHK1 in diverse tissues, with a nearly 20-fold increase in enzymatic activity. The TG mice grew normally with normal blood chemistry, cell counts, heart rate, and blood pressure. Unexpectedly, TG mice with high but not low expression levels of SPHK1 developed progressive myocardial degeneration and fibrosis, with upregulation of embryonic genes, elevated RhoA and Rac1 activity, stimulation of Smad3 phosphorylation, and increased levels of oxidative stress markers. Treatment of juvenile TG mice with pitavastatin, an established inhibitor of the Rho family G proteins, or deletion of S1P3, a major myocardial S1P receptor subtype that couples to Rho GTPases and transactivates Smad signalling, both inhibited cardiac fibrosis with concomitant inhibition of SPHK1-dependent Smad-3 phosphorylation. In addition, the anti-oxidant N-2-mercaptopropyonylglycine, which reduces reactive oxygen species (ROS), also inhibited cardiac fibrosis. In in vivo ischaemia/reperfusion injury, the size of myocardial infarct was 30 decreased in SPHK1-TG mice compared with wild-type mice.Conclusion These results suggest that chronic activation of SPHK1-S1P signalling results in both pathological cardiac remodelling through ROS mediated by S1P3 and favourable cardioprotective effects. 続きを見る
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Takuwaa, Yoh ; Okamoto, Yasuo ; Yoshioka, Kazuaki ; Takuwa, Noriko
出版情報: BioFactors.  38  pp.329-337,  2012-09-01.  International Union of Biochemistry and Molecular Biology, Inc. / Wiley-Blackwell
URL: http://hdl.handle.net/2297/31980
概要: Sphingosine-1-phosphate (S1P), which acts as both the extracellular and intracellular messenger, exerts pleiotropic biol ogical activities including regulation of formation of the vasculature, vascular barrier integrity, and lymphocyte trafficking. Many of these S1P actions are mediated by five members of the G protein-coupled S1P receptors (S1P 1-S1P 5) with overlapping but distinct coupling to heterotrimeric G proteins. The biological activities of S1P are based largely on the cellular actions of S1P on migration, adhesion, and proliferation. Notably, S1P often exhibits receptor subtype-specific, bimodal effects in these cellular actions. For example, S1P 1 mediates cell migration toward S1P, that is, chemotaxis, via G i/Rac pathway whereas S1P 2 mediates inhibition of migration toward a chemoattractant, that is, chemorepulsion, via G 12/13/Rho pathway, which induces Rac inhibition. In addition, S1P 1 mediates stimulation of cell proliferation through the G i-mediated signaling pathways including phosphatidylinositol 3-kinase (PI3K)/Akt and ERK whereas S1P 2 mediates inhibition of cell proliferation through mechanisms involving G 12/13/Rho/Rho kinase/PTEN-dependent Akt inhibition. These differential effects of S1P receptor subtypes on migration and proliferation lead to bimodal regulation of various biological responses. An observed biological response is likely determined by an integrated outcome of the counteracting signals input by S1P receptor subtypes. More recent studies identified the new intracellular targets of S1P including the inflammatory signaling molecule TRAF2 and histone deacetylases HDAC1 and HDAC2. These interactions of S1P regulate NF-κB activity and gene expression, respectively. Development of S1P receptor agonists and antagonists with improved receptor subtype-selectivity, inhibitors, or modulators of sphingolipid-metabolizing enzymes, and their optimal drug delivery system provide novel therapeutic tactics. © 2012 International Union of Biochemistry and Molecular Biology, Inc.<br />発行後1年より全文公開. 続きを見る
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Takuwa, Yoh ; Ikeda, Hitoshi ; Okamoto, Yasuo ; Takuwa, Noriko ; Yoshioka, Kazuaki
出版情報: Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids.  1831  pp.185-192,  2013-01-01.  Elsevier
URL: http://hdl.handle.net/2297/32827
概要: Fibrosis is a pathological process characterized by massive deposition of extracellular matrix (ECM) such as type I/III collagens and fibronectin that are secreted by an expanded pool of myofibroblasts, which are phenotypically altered fibroblasts with more contractile, proliferative, migratory and secretory activities. Fibrosis occurs in various organs including the lung, heart, liver and kidney, resulting in loss of normal tissue architecture and functions. Myofibroblasts could originate from multiple sources including tissue-resident fibroblasts, epithelial and endothelial cells through mechanisms of epithelial/endothelial-mesenchymal transition (EMT/EndMT), and bone marrow-derived circulating progenitors called fibrocytes. Emerging evidence in recent years shows that sphingosine-1-phosphate (S1P) acts on several types of target cells and is engaged in pro-fibrotic inflammatory process and fibrogenic process through multiple mechanisms, which include vascular permeability change, leukocyte infiltration, and migration, proliferation and myofibroblast differentiation of fibroblasts. Many of these S1P actions are receptor subtype-specific. In these actions, S1P has multiple cross-talks with other cytokines, particularly transforming growth factor-β (TGFβ), which plays a major role in fibrosis. The cross-talks include the regulation of S1P production through altered expression and activity of sphingosine kinases in fibrotic lesions, altered expression of S1P receptors, and S1P receptor-mediated transactivation of TGFβ signaling pathway. These cross-talks may give rise to a feed-forward, amplifying loop between S1P and TGFβ, and possibly with other cytokines in stimulating fibrogenesis. Another lysophospholipid mediator lysophosphatidic acid has also been recently implicated in fibrosis. The lysophospholipid signaling pathways represent novel, promising therapeutic targets for treating refractory fibrotic diseases. This article is part of a Special Issue entitled Advances in Lysophospholipid Research. © 2012 Elsevier B.V. 続きを見る
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論文
Yoshioka, Kazuaki ; Yoshida, Kotaro ; Cui, Hong ; Wakayama, Tomohiko ; Takuwa, Noriko ; Okamoto, Yasuo ; Du, Wa ; Qi, Xun ; Asanuma, Ken ; Sugihara, Kazushi ; Aki, Sho ; Miyazawa, Hidekazu ; Biswas, Kuntal ; Nagakura, Chisa ; Ueno, Masaya ; Iseki, Shoichi ; Schwartz, Robert J. ; Okamoto, Hiroshi ; Sasaki, Takehiko ; Matsui, Osamu ; Asano, Masahide ; Adams, Ralf H. ; Takakura, Nobuyuki ; Takuwa, Yoh
出版情報: Nature Medicine.  18  pp.1560-1569,  2012-10-01.  Nature Publishing Group
URL: http://hdl.handle.net/2297/32825
概要: The class II α-isoform of phosphatidylinositol 3-kinase (PI3K-C2α) is localized in endosomes, the trans-Golgi network an d clathrin-coated vesicles; however, its functional role is not well understood. Global or endothelial-cell-specific deficiency of PI3K-C2α resulted in embryonic lethality caused by defects in sprouting angiogenesis and vascular maturation. PI3K-C2α knockdown in endothelial cells resulted in a decrease in the number of PI3-phosphate-enriched endosomes, impaired endosomal trafficking, defective delivery of VE-cadherin to endothelial cell junctions and defective junction assembly. PI3K-C2α knockdown also impaired endothelial cell signaling, including vascular endothelial growth factor receptor internalization and endosomal RhoA activation. Together, the effects of PI3K-C2α knockdown led to defective endothelial cell migration, proliferation, tube formation and barrier integrity. Endothelial PI3K-C2α deficiency in vivo suppressed postischemic and tumor angiogenesis and diminished vascular barrier function with a greatly augmented susceptibility to anaphylaxis and a higher incidence of dissecting aortic aneurysm formation in response to angiotensin II infusion. Thus, PI3K-C2α has a crucial role in vascular formation and barrier integrity and represents a new therapeutic target for vascular disease.<br />In Press / 2013-03-18公開予定. 続きを見る
17.

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論文
Takuwa, Yoh ; Okamoto, Yasuo ; Yoshioka, Kazuaki ; Takuwa, Noriko
出版情報: BioFactors.  38  pp.329-337,  2012-09-01.  International Union of Biochemistry and Molecular Biology, Inc / Wiley-Blackwell
URL: http://hdl.handle.net/2297/32828
概要: Sphingosine-1-phosphate (S1P), which acts as both the extracellular and intracellular messenger, exerts pleiotropic biol ogical activities including regulation of formation of the vasculature, vascular barrier integrity, and lymphocyte trafficking. Many of these S1P actions are mediated by five members of the G protein-coupled S1P receptors (S1P 1-S1P 5) with overlapping but distinct coupling to heterotrimeric G proteins. The biological activities of S1P are based largely on the cellular actions of S1P on migration, adhesion, and proliferation. Notably, S1P often exhibits receptor subtype-specific, bimodal effects in these cellular actions. For example, S1P 1 mediates cell migration toward S1P, that is, chemotaxis, via G i/Rac pathway whereas S1P 2 mediates inhibition of migration toward a chemoattractant, that is, chemorepulsion, via G 12/13/Rho pathway, which induces Rac inhibition. In addition, S1P 1 mediates stimulation of cell proliferation through the G i-mediated signaling pathways including phosphatidylinositol 3-kinase (PI3K)/Akt and ERK whereas S1P 2 mediates inhibition of cell proliferation through mechanisms involving G 12/13/Rho/Rho kinase/PTEN-dependent Akt inhibition. These differential effects of S1P receptor subtypes on migration and proliferation lead to bimodal regulation of various biological responses. An observed biological response is likely determined by an integrated outcome of the counteracting signals input by S1P receptor subtypes. More recent studies identified the new intracellular targets of S1P including the inflammatory signaling molecule TRAF2 and histone deacetylases HDAC1 and HDAC2. These interactions of S1P regulate NF-κB activity and gene expression, respectively. Development of S1P receptor agonists and antagonists with improved receptor subtype-selectivity, inhibitors, or modulators of sphingolipid-metabolizing enzymes, and their optimal drug delivery system provide novel therapeutic tactics. © 2012 International Union of Biochemistry and Molecular Biology, Inc. 続きを見る
18.

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論文
Cui, Hong ; Okamoto, Yasuo ; Yoshioka, Kazuaki ; Du, Wa ; Takuwa, Noriko ; Zhang, Wei ; Asano, Masahide ; Shibamoto, Toshishige ; Takuwa, Yoh
出版情報: Journal of Allergy and Clinical Immunology.  132  pp.1205-1214,  2013-11-01.  Elsevier
URL: http://hdl.handle.net/2297/35640
概要: Background: Sphingosine-1-phosphate receptor 2 (S1P2) is expressed in vascular endothelial cells (ECs). However, the rol e of S1P2 in vascular barrier integrity and anaphylaxis is not well understood. Endothelial nitric oxide synthase (eNOS) generates nitric oxide to mediate vascular leakage, compromising survival in patients with anaphylaxis. We recently observed that endothelial S1P2 inhibits Akt, an activating kinase of eNOS. Objective: We tested the hypothesis that endothelial S1P2 might suppress eNOS, exerting a protective effect against endothelial barrier disruption and anaphylaxis. Methods: Mice deficient in S1P2 and eNOS underwent antigen challenge or platelet-activating factor (PAF) injection. Analyses were performed to examine vascular permeability and the underlying mechanisms. Results: S1pr2 deletion augmented vascular leakage and lethality after either antigen challenge or PAF injection. PAF injection induced activation of Akt and eNOS in the aortas and lungs of S1pr2-null mice, which were augmented compared with values seen in wild-type mice. Consistently, PAF-induced increase in cyclic guanosine monophosphate levels in the aorta was enhanced in S1pr-null mice. Genetic Nos3 deletion or pharmacologic eNOS blockade protected S1pr2-null mice from aggravation of barrier disruption after antigen challenge and PAF injection. ECs isolated from S1pr2-null mice exhibited greater stimulation of Akt and eNOS, with enhanced nitric oxide production in response to sphingosine-1-phosphate or PAF, compared with that seen in wild-type ECs. Moreover, S1pr2-deficient ECs showed more severe disassembly of adherens junctions with augmented S-nitrosylation of β-catenin in response to PAF, which was restored by pharmacologic eNOS blockade. Conclusion: S1P2 diminishes harmful robust eNOS stimulation and thereby attenuates vascular barrier disruption, suggesting potential usefulness of S1P2 agonists as novel therapeutic agents for anaphylaxis. © 2013 American Academy of Allergy, Asthma & Immunology. 続きを見る
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論文
Aki, Sho ; Yoshioka, Kazuaki ; Okamoto, Yasuo ; Takuwa, Noriko ; Takuwa, Yoh
出版情報: Journal of Biological Chemistry.  290  pp.6086-6105,  2015-03-06.  American Society for Biochemistry and Molecular Biology
URL: http://hdl.handle.net/2297/41360
概要: We have recently demonstrated that the PI3K class II-α isoform (PI3K-C2α), which generates phosphatidylinositol 3-phosph ate and phosphatidylinositol 3,4-bisphosphates, plays crucial roles in angiogenesis, by analyzing PI3K-C2α knock-out mice. The PI3K-C2α actions are mediated at least in part through its participation in the internalization of VEGF receptor-2 and sphingosine-1-phosphate receptor S1P1 and thereby their signaling on endosomes. TGFβ, which is also an essential angiogenic factor, signals via the serine/threonine kinase receptor complex to induce phosphorylation of Smad2 and Smad3 (Smad2/3). SARA (Smad anchor for receptor activation) protein, which is localized in early endosomes through its FYVE domain, is required for Smad2/3 signaling. In the present study, we showed that PI3K-C2α knockdown nearly completely abolished TGFβ1-induced phosphorylation and nuclear translocation of Smad2/3 in vascular endothelial cells (ECs). PI3K-C2α was necessary for TGFβ-induced increase in phosphatidylinositol 3,4-bisphosphates in the plasma membrane and TGFβ receptor internalization into the SARA-containing early endosomes, but not for phosphatidylinositol 3-phosphate enrichment or localization of SARA in the early endosomes. PI3K-C2α was also required for TGFβ receptor-mediated formation of SARA-Smad2/3 complex. Inhibition of dynamin, which is required for the clathrin-dependent receptor endocytosis, suppressed both TGFβ receptor internalization and Smad2/3 phosphorylation. TGFβ1 stimulated Smad-dependent VEGF-A expression, VEGF receptor-mediated EC migration, and capillary-like tube formation, which were all abolished by either PI3K-C2α knockdown or a dynamin inhibitor. Finally, TGFβ1-induced microvessel formation in Matrigel plugs was greatly attenuated in EC-specific PI3K-C2α-deleted mice. These observations indicate that PI3K-C2α plays the pivotal role in TGFβ receptor endocytosis and thereby Smad2/3 signaling, participating in angiogenic actions of TGFβ. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc. 続きを見る
20.

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論文
Cui, Hong ; Okamoto, Yasuo ; Yoshioka, Kazuaki ; Du, Wa ; Takuwa, Noriko ; Zhang, Wei ; Asano, Masahide ; Shibamoto, Toshishige ; Takuwa, Yoh
出版情報: Journal of Allergy and Clinical Immunology.  132  pp.1205-1214,  2013-11-01.  Elsevier
URL: http://hdl.handle.net/2297/36259
概要: Background Sphingosine-1-phosphate receptor 2 (S1P2) is expressed in vascular endothelial cells (ECs). However, the role of S1P 2 in vascular barrier integrity and anaphylaxis is not well understood. Endothelial nitric oxide synthase (eNOS) generates nitric oxide to mediate vascular leakage, compromising survival in patients with anaphylaxis. We recently observed that endothelial S1P2 inhibits Akt, an activating kinase of eNOS. Objective We tested the hypothesis that endothelial S1P 2 might suppress eNOS, exerting a protective effect against endothelial barrier disruption and anaphylaxis. Methods Mice deficient in S1P2 and eNOS underwent antigen challenge or platelet-activating factor (PAF) injection. Analyses were performed to examine vascular permeability and the underlying mechanisms. Results S1pr2 deletion augmented vascular leakage and lethality after either antigen challenge or PAF injection. PAF injection induced activation of Akt and eNOS in the aortas and lungs of S1pr2-null mice, which were augmented compared with values seen in wild-type mice. Consistently, PAF-induced increase in cyclic guanosine monophosphate levels in the aorta was enhanced in S1pr-null mice. Genetic Nos3 deletion or pharmacologic eNOS blockade protected S1pr2-null mice from aggravation of barrier disruption after antigen challenge and PAF injection. ECs isolated from S1pr2-null mice exhibited greater stimulation of Akt and eNOS, with enhanced nitric oxide production in response to sphingosine-1-phosphate or PAF, compared with that seen in wild-type ECs. Moreover, S1pr2-deficient ECs showed more severe disassembly of adherens junctions with augmented S-nitrosylation of β-catenin in response to PAF, which was restored by pharmacologic eNOS blockade. Conclusion S1P2 diminishes harmful robust eNOS stimulation and thereby attenuates vascular barrier disruption, suggesting potential usefulness of S1P2 agonists as novel therapeutic agents for anaphylaxis. © 2013 American Academy of Allergy, Asthma & Immunology. 続きを見る
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論文
吉岡, 和晃 ; Yoshioka, Kazuaki
出版情報: 令和1(2019)年度 科学研究費補助金 基盤研究(C) 研究成果報告書 = 2019 Fiscal Year Final Research Report.  2017-04-01 - 2020-03-31  pp.7p.-,  2020-05-18. 
URL: http://hdl.handle.net/2297/00057963
概要: 金沢大学医薬保健研究域医学系<br />細胞膜に存在するホスファチジルイノシトール(PI)は脂質リン酸化酵素PI3キナーゼにより種々の3’-ホスホイノシチド(3’-PI)に変換され、細胞内小胞輸送やシグナル伝達に関与する。Myotubula rin-related protein (MTMR)ファミリーはPI-3リン酸(PI(3)P)をPIへと変換する3’-PIホスファターゼである。本研究課題では、MTMR4が肺組織において II型肺胞上皮細胞(AECII)に高発現し、エンドソーム~リソソーム(エンドリソソーム)、オートファゴソームといった細胞内クリアランスを担う小器官の機能調節に重要な役割を果たすことを明らかにした。<br />Phosphatidylinositol 3-phosphate (PI(3)P) is the predominant phosphoinositide species in early endosomes and autophagosomes, in which PI(3)P dictates traffic of these organelles. Phosphoinositide levels are tightly regulated by lipid-kinases and-phosphatases; however, a phosphatase that converts PI(3)P back to phosphatidylinositol in the endosomal and autophagosomal compartments is not fully understood. We investigated the subcellular distribution and functions of myotubularin-related protein-4 (MTMR4), which is distinct among other MTMRs in that it possesses a PI(3)P-binding FYVE domain, in lung alveolar epithelial cells. MTMR4 knockdown markedly suppressed the motility, fusion, and fission of PI(3)P-enriched structures, resulting in decreases in late endosomes, autophagosomes, and lysosomes. This project has unveiled that MTMR4 is essential for the integrity of endocytic and autophagic pathways.<br />研究課題/領域番号:17K08532, 研究期間(年度):2017-04-01 - 2020-03-31<br />出典:「膜動現象を制御するイノシトールリン脂質脱リン酸化酵素の肺発達・成熟における役割」研究成果報告書 課題番号17K08532(KAKEN:科学研究費助成事業データベース(国立情報学研究所))(https://kaken.nii.ac.jp/report/KAKENHI-PROJECT-17K08532/17K08532seika/)を加工して作成 続きを見る
22.

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論文
吉岡, 和晃 ; Yoshioka, Kazuaki
出版情報: 平成28(2016)年度 科学研究費補助金 基盤研究(C) 研究成果報告書 = 2016 Fiscal Year Final Research Report.  2014-04-01 - 2017-03-31  pp.5p.-,  2017-05-26. 
URL: http://hdl.handle.net/2297/00050990
概要: 金沢大学医薬保健研究域医学系<br />当研究室では、クラスII型PI3K-C2α KO マウスは胎生期の血管形成不全により胎生致死となり、C2αが胎生期血管形成及び生後の血管恒常性維持に必要であることを見いだしていた。本研究では、内皮にお けるPI(3)Pレベル調節機構を解析するために、PI(3)P脂質脱リン酸化酵素(MTM)種を同定した。更にMTM-KOマウスを作出して表現型を解析した結果、ホモKOは出生直後24時間以内に全例死亡することを見出した。以上の結果から、内皮における脂質リン酸化酵素C2aと脱リン酸化酵素MTMは強調して働き、膜小胞上でPI(3)Pレベルを調節することによって、内皮の恒常性を維持していることが示唆された。<br />Phosphatidylinositol 3-kinases (PI3Ks) are the family of lipid kinases responsible for the generation of 3’-phosphoinositides. We have recently reported that class II a-isoform PI3K (C2a) plays a crucial role in developmental and pathological angiogenesis, and is indispensable for the maintenance of the VE-cadherin-mediated EC barrier function and receptor endocytosis. These defects result in impaired vascular homeostasis under the pathophysiological conditions. In this study, we found that among myotubularin-related protein (MTMR) family members, the expression of MTMRx is relatively abundant in ECs. We found that, in HUVECs, MTMRx was localized mainly in the endosomes. Next we analyzed the phenotype of MTMRx global KO mice. MTMRx mice showed normally development but die perinatally. We conclude that C2a regulates vascular homeostasis through controlling PI(3)P+-membrane trafficking, cooperating with MTMRx in ECs.<br />研究課題/領域番号:26460292, 研究期間(年度):2014-04-01 - 2017-03-31 続きを見る
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論文
吉岡, 和晃 ; Yoshioka, Kazuaki
出版情報: 平成25(2013)年度 科学研究費補助金 基盤研究(C) 研究成果報告書 = 2013 Fiscal Year Final Research Report.  2011-2013  pp.6p.-,  2014-05-20.  金沢大学医薬保健研究域医学系
URL: http://hdl.handle.net/2297/00050991
概要: 金沢大学医薬保健研究域医学系<br />本研究において申請者らは、発生期・生後の生理的および病的(虚血、腫瘍)血管新生ならびに血管恒常性維持にクラスII型PI3K-C2αが重要な役割を果たすことを明らかにした。内皮細胞においてC2αをノック ダウンすると、エンドソーム輸送の障害、VE-カドヘリンの配送異常が引き起こされた。C2αヘテロ欠損マウスでは、アンジオテンシンII投与に対する解離性大動脈瘤形成の発症率上昇が見られた。以上のことから、C2αは血管内皮細胞において、小胞輸送の制御を介した物質輸送及びエンドソーム上でのシグナル伝達に必須であり、これらの作用により血管形成と血管の健全性維持に重要な役割を担うことが明らかになった。<br />Although class I PI3Ks and class III Vps34 are well-characterized, the physiological roles of PI3K class IIa (C2a) remain largely unknown. Global C2a-null mice and EC-specific C2a conditional KO mice showed embryonic lethality due to defects in sprouting angiogenesis and vascular maturation. In cultured ECs, siRNA-mediated knockdown of C2a resulted in impaired endosomal trafficking. C2a knockdown also impaired cell signaling including VEGF receptor-2 internalization and RhoA activation on endosomes, but not Akt and ERK. Consequently, endosomal delivery of VE-cadherin to EC junctions was disturbed, leading to defects in VE-cadherin transport and assembly and barrier integrity. C2a haplo-insufficient mice exhibited defective postnatal angiogenesis and vascular barrier integrity with a higher incidence of dissecting aortic aneurysm formation on angiotensin-II infusion. Thus, C2a plays a crucial role in vascular formation and barrier integrity.<br />研究課題/領域番号:23590257, 研究期間(年度):2011-2013 続きを見る
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論文
吉岡, 和晃 ; Yoshioka, Kazuaki
出版情報: 平成22(2010)年度 科学研究費補助金 基盤研究(C) 研究成果報告書 = 2010 Fiscal Year Final Research Report.  2008-2010  pp.6p.-,  2011-05-19.  金沢大学医薬保健研究域医学系
URL: http://hdl.handle.net/2297/00050992
概要: 本研究では、"クラスII型PI3キナーゼC2α"の機能、特に血管恒常性における役割を明らかにするために、C2α遺伝子KOマウスを用いてC2αの生理機能と血管障害における役割を解析することを目的として実施した。KOマウスの表現型解析から、胎生 期血管形成プロセスの異常は血管新生、特に血管成熟過程に起因する可能性が高いと考えられた。また、C2αヘテロKOマウス血管障害モデルでは、解離性大動脈瘤が高頻度に観察された。C2αはこれまで全く生理機能が知られてなく、クラスI型PI3キナーゼとは異なる基質特異性、細胞内分布を示すことから、血管恒常性維持に必須の新たな機能的役割を持つ細胞内制御因子でありことが明らかとなった。<br />Phosphatidylinositol-3-OH kinase (PI3K) family, which comprises three classes, regulates a diverse array of cellular processes through the generation of 3-phosphoinositides. Although class I PI3Ks including p110α, p110β, p110δ and p110γ isoforms were well characterized among three classes, the in vivo physiological functions of class II PI3Ks, which comprise three members PI3K-C2α (C2α), C2β and C2γ, remain largely unknown. We found that Pik3c2a gene-global disruption (C2α KO) and conditional loss of C2α in endothelial cells (ECs) in mice, but not in vascular smooth muscles (VSMCs) and cardiomyocytes, caused embryonic lethality due to impairment of developmental angiogenesis characterized by incomplete EC sprouting and mural-cell (VSMCs and pericytes) coverage. Finally, C2α-haploinsufficient mice are alive, but exhibit vascular hyperpermeability and a higher incidence of dissecting aortic aneurysms with rupture on angiotensin-II (AngII) infusion. These results provide the first evidence that C2α plays a novel essential role in endothelial physiology, particularly angiogenesis and barrier integrity, through regulating endosomal trafficking and underscore broader roles for PI3K family members in vascular physiology.<br />研究課題/領域番号:20590207, 研究期間(年度):2008-2010 続きを見る
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論文
吉岡, 和晃 ; Yoshioka, Kazuaki
出版情報: 平成17(2005)年度 科学研究費補助金 若手研究(B) 研究概要 = 2005 Research Project Summary.  2004 – 2005  pp.2p.-,  2016-04-21. 
URL: http://hdl.handle.net/2297/00061063
概要: 金沢大学医薬保健研究域医学系<br />血管平滑筋のトーヌス調節において、ノルアドレナリンなどの生理活性物質が果たす役割もさることながら、血管平滑筋に備わる"自己調節機構"は極めて重要である。しかしながら、その制御機構に関する知見は乏しく、 詳細な解析が必要である。本研究では、血管平滑筋トーヌスの調節機構のうち、特にRho-Rhoキナーゼからなるカルシウム感受性調節に的を絞って、伸展刺激によるその制御分子機構を明らかにすることを目的とした。本研究によって、血管内圧に由来する伸展力が、従来考えられていたCa^<2+>-MLCK系以外にRho-Rhoキナーゼから成るCa^<2+>に対する感受性調節系により血管平滑筋トーヌスを制御している可能性が示された。このRho-Rhoキナーゼ及びミオシン・ホスファターゼ調節系において注目すべき事は、新規脂質キナーゼが大きく関与していることが薬理学的な解析から明らかになった点である。これを受けて、現在この脂質キナーゼの遺伝子欠損マウスの作製に取りかかっている。既に組換えES細胞を樹立後、凝集法によりキメラマウス及びヘテロ変異体マウスの作製に成功している。今後、ホモ変異体マウスの表現型の解析を始め、ホモ変異体マウス由来の摘出血管及び培養血管平滑筋細胞を用いた血管トーヌス制御機構を解析することが可能である。これにより、伸展力の作用機序の全貌が明らかにすることによって、将来的には血管トーヌス自己調節機構を標的とした新しい薬剤の開発も期待できる。これらの薬物は、脳循環改善薬、高血圧治療薬、抗血管れん縮薬として期待される。<br />研究課題/領域番号:16790133, 研究期間(年度):2004 – 2005<br />出典:「伸展力を介した血管平滑筋トーヌスのカルシウム感受性制御」研究成果報告書 課題番号16790133(KAKEN:科学研究費助成事業データベース(国立情報学研究所))(https://kaken.nii.ac.jp/ja/grant/KAKENHI-PROJECT-16790133/)を加工して作成 続きを見る