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論文

論文
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 sphingosine 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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論文
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 receptors (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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論文
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-depen dent MLC 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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論文
Seok, Young Mi ; Choi, Young Whan ; Kim, Gyung-Duck ; Kim, Hye-Young ; Takuwa, Yoh ; Kim, In Kyeom
出版情報: Naunyn-Schmiedeberg's Archives of Pharmacology.  383  pp.45-56,  2011-01-01.  Springer Verlag (Germany)
URL: http://hdl.handle.net/2297/25789
概要: 金沢大学医薬保健研究域医学系<br />Gomisin A (GA) is an active ingredient of the fruits of Schisandra chinensis which has been wi dely used as a tonic in traditional Korean medicine. GA induces not only endothelium-dependent but also endothelium-independent relaxation in an isolated rat's thoracic aorta. This study was aimed to investigate the molecular mechanism by which GA induces endothelium-independent vasorelaxation. Rat aortic rings were denuded of endothelium, mounted in organ baths, and subjected to contraction or relaxation. We measured the amount of GTP RhoA as well as the phosphorylation level of 20 kDa myosin light chains (MLC20), myosin phosphatase-targeting subunit 1 (MYPT1) and protein kinase C-potentiated inhibitory protein for heterotrimeric myosin light-chain phosphatase of 17 kDa (CPI17). Pretreatment with GA dose-dependently inhibited the concentration-response curves in response to sodium fluoride (NaF) or thromboxane A2 agonist U46619, but not to phorbol 12, 13-dibutyrate (PDBu). GA decreased the activation of RhoA as well as the phosphorylation level of MLC20, MYPT1Thr855, and CPI17 induced by 8.0 mM NaF or 30 nM U46619. However, K+ channel blockers such as glibenclamide, apamin, or charybdotoxin did not affect the vascular relaxation induced by GA. Furthermore, GA did not affect the level of phosphorylation of CPI17 induced by PDBu. GA reduces vascular contraction through inhibition of RhoA/Rho-kinase pathway in endothelium-denuded rat aorta. © 2010 Springer-Verlag. 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 subtypes 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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論文
Seok, Young Mi ; Choi, Young Whan ; Kim, Gyung-Duck ; Kim, Hye-Young ; Takuwa, Yoh ; Kim, In Kyeom
出版情報: Naunyn-Schmiedeberg's Archives of Pharmacology.  383  pp.45-56,  2011-01-01.  Springer-Verlag
URL: http://hdl.handle.net/2297/26244
概要: 金沢大学医薬保健研究域医学系<br />Gomisin A (GA) is an active ingredient of the fruits of Schisandra chinensis which has been wi dely used as a tonic in traditional Korean medicine. GA induces not only endothelium-dependent but also endothelium-independent relaxation in an isolated rat's thoracic aorta. This study was aimed to investigate the molecular mechanism by which GA induces endothelium-independent vasorelaxation. Rat aortic rings were denuded of endothelium, mounted in organ baths, and subjected to contraction or relaxation. We measured the amount of GTP RhoA as well as the phosphorylation level of 20 kDa myosin light chains (MLC20), myosin phosphatase-targeting subunit 1 (MYPT1) and protein kinase C-potentiated inhibitory protein for heterotrimeric myosin light-chain phosphatase of 17 kDa (CPI17). Pretreatment with GA dose-dependently inhibited the concentration-response curves in response to sodium fluoride (NaF) or thromboxane A2 agonist U46619, but not to phorbol 12, 13-dibutyrate (PDBu). GA decreased the activation of RhoA as well as the phosphorylation level of MLC20, MYPT1Thr855, and CPI17 induced by 8.0 mM NaF or 30 nM U46619. However, K+ channel blockers such as glibenclamide, apamin, or charybdotoxin did not affect the vascular relaxation induced by GA. Furthermore, GA did not affect the level of phosphorylation of CPI17 induced by PDBu. GA reduces vascular contraction through inhibition of RhoA/Rho-kinase pathway in endothelium-denuded rat aorta. © 2010 Springer-Verlag. 続きを見る
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論文
Takabatake, Masayoshi ; Takuwa, Yoh ; Takuwa, Noriko ; Yasuno, Hironobu ; Matsumoto, Sayaka ; Shibutani, Makoto ; Mitsumori, Kunitoshi
出版情報: The journal of veterinary medical science.  70  pp.483-485,  2008-05-25.  日本獣医学会 = Japanese Society of Veterinary Science
URL: http://hdl.handle.net/2297/32519
概要: 32週給のS1P2遺伝子ヘテロ欠損雌性マウスに発生した腎複合上皮間質腫瘍(MESTK)について報告する.本病変は,尿細管様の管腔を形成する上皮成分およびそれらを取り囲む間質成分から成る二相性腫瘍であり,免疫染色において上皮成分はサイトケラチ ンに,間質成分はビメンチンおよび平滑筋マーカーにそれぞれ陽性であった.これらの結果から,ヒトWHO分類の診断基準に基づいて本症例はMESTKであると診断した. We report a case of mixed epithelial and stromal tumor of the kidney (MESTK) in a 32-week-old heterozygous sphingosine l-phosphate-2 (S1P2) receptor deficient female mouse. A white solid mass replacing the left kidney was observed at the left retroperitoneal wall. Histologically, the tumor mass consisted of dimorphic cellular components of epithelial and stromal cells. Epithelial cells formed various sized irregular-shaped tubular structures resembling renal tubules surrounded by stromal cells. Immunohistochemically, epithelial cells were positive for cytokeratin, while stromal cells showed positive immunoreactivity with a-smooth muscle actin as well as vimentin. Based on the morphological and immunohistochemical findings, this tumor was diagnosed as a MESTK. 続きを見る
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論文
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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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-Golg i network and 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
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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 biological 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