2000 年度実績報告書

血管の細胞シグナル伝達網制御を多項目連続測定可能な光学システムの開発

研究課題

研究課題/領域番号	10557072
研究機関	九州大学
研究代表者	金出英夫九州大学, 大学院・医学研究院, 教授 (80038851)
研究分担者	平野勝也九州大学, 大学院・医学研究院, 講師 (80291516) 西村淳二九州大学, 大学院・医学研究院, 助教授 (90237727)
キーワード	細胞シグナル伝達 / 血管内皮細胞 / 血管平滑筋細胞 / fura-2 / BCECF / 光学線維束 / 血管緊張調節
研究概要	本研究によって、『細胞質Ca^<2+>濃度([Ca^<2+>]i)と[pH]i変化の同時連続測定システム』を開発した。本年度は、これを用いて次の研究を行った。【目的】平滑筋収縮は主に[Ca^<2+>]iにより調節される。一方、[pH]iはこの収縮性に影響を与えること、特にアルカリ化は収縮性を増強することが報告されている。[Ca^<2+>]iと[pH]iの同時測定によって、この機序の一端を解明した。【方法】ラット大動脈初代培養平滑筋細胞にCa^<2+>指示蛍光色素(Fura-2)とpH指示蛍光色素(BCECF)を二重負荷し、蛍光測光を行った。【結果】20mM NH_4Cl添加により、[pH]iは、静止期の7.24から7.78へと急速に上昇した。NH_4Cl投与中[pH]iは漸減した。NH_4Clを除くと[pH]iは急速に低下し、一旦、アシドーシス(6.73)に達し、徐々に刺激前値に回復した。NH_4Clによる[pH]i上昇にほぼ平行して[Ca^<2+>]iが上昇した。この[Ca^<2+>]i上昇は、100mM K^+脱分極による[Ca^<2+>]i上昇の50%に達した。NH_4Clを除くと[Ca^<2+>]iは刺激前値に復した。NH_4Clによる[Ca^<2+>]i上昇は、細胞外にCa^<2+>が無い場合には認められなかった。また、3mM NiCl_2を灌流液に添加しておくと、NH_4Clによる[Ca^<2+>]i上昇は消失した。10μMジルチアゼム処理により、100mM K^+脱分極による[Ca^<2+>]i上昇は対照の16%に抑止されたが、NH_4Clによる[Ca^<2+>]i上昇の抑制は見られなかった。一方、SKF96365処理は、100mM K^+脱分極による[Ca^<2+>]i上昇を50%に抑制し、NH_4Clによる[Ca^<2+>]i上昇は対照の20%に抑制した。【総括】アルカリ化は、細胞外からのCa^<2+>流入と[Ca^<2+>]I上昇を引き起こし、収縮を増強することが明かとなった。このCa^<2+>流入にはSKF96365感受性Ca^<2+>流入経路、おそらくは容量性Ca^<2+>流入が主要な経路であると考えられた。

研究成果
(20件)

すべてその他

すべて文献書誌 (20件)

[文献書誌] Ushio-Fukai M: "Changes in the cytosolic Ca^<2+> concentration and Ca^<2+>-sensitivity of the contractile apparatus during angiotensin II-induced desensitization in the rabbit femoral artery."Br J Pharmacol. 129. 425-436 (2000)
[文献書誌] Ushio-Fukai M: "The mechanism of the decrease in cytosolic Ca^<2+> concentrations induced by angiotensin II in the high K^+-depolarized rabbit femoral artery"Br J Pharmacol. 129. 437-447 (2000)
[文献書誌] Mizuno O: "Proteolysis and phosphorylation-mediated regulation of thrombin receptor activity in in situ endothelial cells."Eur J Pharmacol. 389. 13-23 (2000)
[文献書誌] Ihara E: "The mechanism of bradykinin-induced endothelium-dependent contraction and relaxation in the porcine interlobar renal artery."Br J Pharmacol. 129. 943-952 (2000)
[文献書誌] Mizuno O: "Stimulus-specific alteration of the relationship between cytosolic Ca^<2+> transients and nitric oxide production in endothelial cells ex vivo."Br J Pharmacol. 130. 1140-1146 (2000)
[文献書誌] Kawasaki J: "Dissociation between the Ca^<2+> signal and tube formation induced by vascular endothelial growth factor in bovine aortic endothelial cells."Eur J Pharmacol. 398. 19-29 (2000)
[文献書誌] Zeng Y: "Minimal requirement for nuclear localization of p27^<Kipl>, a cyclin-dependent kinase inhibitor."Biochem Biophys Res Commun. 274. 37-42 (2000)
[文献書誌] Shintani Y : "Mechanisms underlying the neurokinin A-induced contraction of the pregnant rat myometrium."Br J Pharmacol. 130. 1165-1173 (2000)
[文献書誌] Kuroiwa-Matsumoto M: "Mechanisms of the thapsigargin-induced Ca^<2+> entry in in situ endothelial cells of the porcine aortic valve and the endothelium-dependent relaxation in the porcine coronary artery."Br J Pharmacol. 131. 115-123 (2000)
[文献書誌] Hirano K : "Mitogen-induced up-regulation of non-smooth muscle isoform of α-tropomyosin in rat aortic smooth muscle cells."Eur J Pharmacol. 406. 209-218 (2000)
[文献書誌] Morisaki T: "A combination of cyclosporin-A (CsA) and interferon-gamma (INF-γ) induces apoptosis in human gastric carcinoma cells. : 3363-3374,200"Anticancer Research. 20. 3363-3373 (2000)
[文献書誌] Takeda K: "Peroxisome proliferator-activated receptor γ activators downregulate angiotensin II type 1 receptor in vascular smooth muscle cells."Circulation. 102. 1834-1839 (2000)
[文献書誌] Matoba T: "Hydorogen peroxide is an endothelium-derived hyperpolarizing factor in mice."J Clin Invest. 106. 1521-1530 (2000)
[文献書誌] Ieiri S: "Mechanism of vasorelaxation induced by a novel types of Ca^<2+> channel blocker SR33805 in the porcine coronary artery."Br J Pharmacol. 131. 1592-1596 (2000)
[文献書誌] Shintani Y: "Enhanced contractile response to thrombin in the pregnant rat myometrium.131 : 1619-1628, 2000"Br J Pharmacol. 131. 1619-1628 (2000)
[文献書誌] Derkach DN: "Thrombin causes endothelium-dependent biphasic regulation of vascular tone in the porcine renal interlobar artery."Br J Pharmacol. 131. 1635-1642 (2000)
[文献書誌] Toshima J: "Differential effects of progesterone and 17b-estradiol on the Ca^<2+> entry induced by thapsigargin and endothelin-1 in in situ endothelial cells."Biochem Biophys Acta. 1499. 109-121 (2000)
[文献書誌] Hirano K: "Cloning and functional expression of a degradation-resistant novel isoform of p27^<Kipl>.353 : 51-57, 2001"Biochem J. 353. 51-57 (2001)
[文献書誌] Setoguchi H: "Leukotriene C_4 enhances the contraction of porcine tracheal smooth muscle through the activation of Y-27632, a rho kinase inhibitor, sensitive pathway."Br J Pharmacol. 132. 111-118 (2001)
[文献書誌] Arimura T : "Identification, characterization and functional analysis of heart-specific myosin light chain phosphatase small subunit."J Biol Chem. (in press).

2000 年度 実績報告書

血管の細胞シグナル伝達網制御を多項目連続測定可能な光学システムの開発

研究代表者

金出 英夫 九州大学, 大学院・医学研究院, 教授 (80038851)

研究成果

[文献書誌] Ushio-Fukai M: "Changes in the cytosolic Ca^<2+> concentration and Ca^<2+>-sensitivity of the contractile apparatus during angiotensin II-induced desensitization in the rabbit femoral artery."Br J Pharmacol. 129. 425-436 (2000)

[文献書誌] Ushio-Fukai M: "The mechanism of the decrease in cytosolic Ca^<2+> concentrations induced by angiotensin II in the high K^+-depolarized rabbit femoral artery"Br J Pharmacol. 129. 437-447 (2000)

[文献書誌] Mizuno O: "Proteolysis and phosphorylation-mediated regulation of thrombin receptor activity in in situ endothelial cells."Eur J Pharmacol. 389. 13-23 (2000)

[文献書誌] Ihara E: "The mechanism of bradykinin-induced endothelium-dependent contraction and relaxation in the porcine interlobar renal artery."Br J Pharmacol. 129. 943-952 (2000)

[文献書誌] Mizuno O: "Stimulus-specific alteration of the relationship between cytosolic Ca^<2+> transients and nitric oxide production in endothelial cells ex vivo."Br J Pharmacol. 130. 1140-1146 (2000)

[文献書誌] Kawasaki J: "Dissociation between the Ca^<2+> signal and tube formation induced by vascular endothelial growth factor in bovine aortic endothelial cells."Eur J Pharmacol. 398. 19-29 (2000)

[文献書誌] Zeng Y: "Minimal requirement for nuclear localization of p27^<Kipl>, a cyclin-dependent kinase inhibitor."Biochem Biophys Res Commun. 274. 37-42 (2000)

[文献書誌] Shintani Y : "Mechanisms underlying the neurokinin A-induced contraction of the pregnant rat myometrium."Br J Pharmacol. 130. 1165-1173 (2000)

[文献書誌] Kuroiwa-Matsumoto M: "Mechanisms of the thapsigargin-induced Ca^<2+> entry in in situ endothelial cells of the porcine aortic valve and the endothelium-dependent relaxation in the porcine coronary artery."Br J Pharmacol. 131. 115-123 (2000)

[文献書誌] Hirano K : "Mitogen-induced up-regulation of non-smooth muscle isoform of α-tropomyosin in rat aortic smooth muscle cells."Eur J Pharmacol. 406. 209-218 (2000)

[文献書誌] Morisaki T: "A combination of cyclosporin-A (CsA) and interferon-gamma (INF-γ) induces apoptosis in human gastric carcinoma cells. : 3363-3374,200"Anticancer Research. 20. 3363-3373 (2000)

[文献書誌] Takeda K: "Peroxisome proliferator-activated receptor γ activators downregulate angiotensin II type 1 receptor in vascular smooth muscle cells."Circulation. 102. 1834-1839 (2000)

[文献書誌] Matoba T: "Hydorogen peroxide is an endothelium-derived hyperpolarizing factor in mice."J Clin Invest. 106. 1521-1530 (2000)

[文献書誌] Ieiri S: "Mechanism of vasorelaxation induced by a novel types of Ca^<2+> channel blocker SR33805 in the porcine coronary artery."Br J Pharmacol. 131. 1592-1596 (2000)

[文献書誌] Shintani Y: "Enhanced contractile response to thrombin in the pregnant rat myometrium.131 : 1619-1628, 2000"Br J Pharmacol. 131. 1619-1628 (2000)

[文献書誌] Derkach DN: "Thrombin causes endothelium-dependent biphasic regulation of vascular tone in the porcine renal interlobar artery."Br J Pharmacol. 131. 1635-1642 (2000)

[文献書誌] Toshima J: "Differential effects of progesterone and 17b-estradiol on the Ca^<2+> entry induced by thapsigargin and endothelin-1 in in situ endothelial cells."Biochem Biophys Acta. 1499. 109-121 (2000)

[文献書誌] Hirano K: "Cloning and functional expression of a degradation-resistant novel isoform of p27^<Kipl>.353 : 51-57, 2001"Biochem J. 353. 51-57 (2001)

[文献書誌] Setoguchi H: "Leukotriene C_4 enhances the contraction of porcine tracheal smooth muscle through the activation of Y-27632, a rho kinase inhibitor, sensitive pathway."Br J Pharmacol. 132. 111-118 (2001)

[文献書誌] Arimura T : "Identification, characterization and functional analysis of heart-specific myosin light chain phosphatase small subunit."J Biol Chem. (in press).

2000 年度実績報告書

金出英夫九州大学, 大学院・医学研究院, 教授 (80038851)