apelin-13下调caveolae可促进血管平滑肌细胞增殖

apelin-13 promotes proliferation of vascular smooth muscle cells by downregulating caveolae expression

背景:结合课题组以往研究成果,提出caveolae可能参与apelin-13促血管平滑肌细胞增殖的假设。目的:实验观察细胞膜特殊凹陷结构caveolae参与G蛋白偶联受体APJ的内源性配体apelin-13促进大鼠血管平滑肌细胞增殖的作用。方法:采用组织贴块法培养大鼠胸主动脉血管平滑肌细胞,用MTT方法观察血管平滑肌细胞增殖,Western Blotting方法观察信号蛋白表达,免疫共沉淀技术检测信号分子复合物的形成。结果与结论:①caveolae结构破坏剂β-环糊精(5mmol/L,25h)可明显增强apelin-13诱导的血管平滑肌细胞增殖。②apelin-13(0,1,2,4,8μmol/L)刺激血管平滑肌细胞,caveolin-1的表达下调,在1μmol/L时下调明显。③β-环糊精(5mmol/L)破坏caveolae后,可使apelin-13下调caveolin-1表达的作用增强。④对照组(体积分数为0.1%胎牛血清孵育)及处理组(apelin-13刺激)caveolin-1与PI3K及ERK1/2均有复合物形成,在apelin-13刺激的情况caveolin-1-PI3K复合物减少、caveolin-1-ERK1/2复合物减少,即apelin-13可能促进caveolin-1与PI3K及ERK1/2解离。结果提示细胞膜特殊凹陷结构caveolae参与apelin-13促血管平滑肌细胞增殖作用。

BACKGROUND：Previous results have shown that caveolae is likely to participate in apelin-13 promotion of vascular smooth muscle cell proliferation.OBJECTIVE：APJ is a G-protein coupled receptor,and its ligand is apelin peptide.Previously,we reported PI3K and ERK1/2 signal pathway mediated proliferation of vascular smooth muscle cells（VSMCs） induced by apelin-13.This study is to observe caveolae involved in rat VSMCs proliferation induced by apelin-13 and to determine whether caveolin-1 protein binds to signal molecules PI3K,ERK1/2 to mediate VSMC proliferation induced by apelin-13.METHODS：VSMCs were prepared from male Sprague-Dawley rat thoracic aorta by the primary-explant method.The effect of β-cyclodextrin on cell proliferation induced by apelin-13 was measured by MTT assay.VSMCs were incubated with apelin-13 and β-cyclodextrin（β-CD） ,caveolin-1 expression was detected by western blot assay.Formation of high molecular weight polyprotein component including caveolin-1 and PI3K /ERK1/2 was detected by immunoprecipitation.RESULTS AND CONCLUSION：β-CD（5 mmol/L,25 hours） significantly enhanced VSMCs proliferation induced by apelin-13（P〈0.05） .Treating VSMCs with apelin-13（0,1,2,4,8 μmol/L） downregulated apelin-13-induced expression of caveolin-1 and the effect was distinct at 1μmol/L（P〈0.05） .Pretreatment of the cells with 5 mmol/L β-CD could strength the downregulation of apelin-13-induced caveolin-1 expression（P〈0.05） .Apelin-13 may induce dissociation of PI3K（ERK1/2） with caveolin-1 in VSMCs compared with the control group.These findings suggest that caveolae is involved in apelin-13-induced VSMCs proliferation.