apelin-13下调caveolae可促进血管平滑肌细胞增殖

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(5 mmol/L,25 h)可明显增强apelin-13诱导的血管平滑肌细胞增殖。②apelin-13(0,1,2,4,8 µmol/L)刺激血管平滑肌细胞,caveolin-1的表达下调,在1 µmol/L时下调明显。③β-环糊精        (5 mmol/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。

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

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.