腺苷及腺苷A1受体在小鼠肾小球系膜细胞收缩中的作用

The role of adenosine and A1AR in the contraction of mice glomerular mesangial cells

目的:观察腺苷A1受体(A1AR)在小鼠肾小球系膜细胞的表达,探讨腺苷和A1AR是否参与肾小球系膜细胞的收缩。方法:采用分样筛法提取原代小鼠肾小球系膜细胞,CCK8试剂盒检测细胞增殖,免疫荧光染色和Western blot观察A1AR在系膜细胞中的表达,每2min拍照动态观察血管紧张素Ⅱ(AngⅡ)刺激30min前后细胞面积的变化,对比腺苷、A1AR激动剂、A1AR抑制剂对细胞收缩的影响。结果:腺苷对系膜细胞的增殖无明显影响,A1AR主要表达于小鼠肾小球系膜细胞的胞浆中。腺苷和A1AR激动剂刺激48h后,系膜细胞在接触AngⅡ30min后的收缩反应显著增强,面积变化值分别达到8947±3658pixels和9813±1640pixels,显著大于正常对照组(4268±2131pixels,P<0.05);而使用A1AR抑制剂刺激48h后,系膜细胞的收缩反应有减弱趋势。结论:小鼠系膜细胞存在A1AR表达,腺苷及A1AR是调节系膜细胞收缩的重要机制。

Objective:To investigate the expression of adenosine A1 receptor(A1 AR)in mice glomerular mesangial cells,and the role of adenosine and A1 AR in mesangial contraction.Methods:Sample sieve method was used in the separation of mice primary mesangial cells.CCK-8 kit was applied to evaluate the proliferation of cells.A1 AR was identified by immunofluorescent staining and western blot in mice mesangial cells.The morphological changes every two minutes during the 30 minutes period after angiotensinⅡstimulation were observed.The effects of adenosine,A1 AR agonists and A1 AR inhibitors on the contraction of mesangial cells were analyzed by comparation of the cell area changes before and 30 minutes after angiotensinⅡstimulation.Results:Different concentrations of adenosine such as 0.1μM,1μM,10μM and 100μM showed no effects on the proliferation of mesangial cells.A1 AR expressed mainly in the cytoplasm of mice mesangial cells.After stimulation of adenosine or A1 AR agonist for 48 hours,the cell area decreased reacting to angiotensinⅡby8947±3658 pixels and 9813±1640 pixels,respectively;significantly stronger than that of control which was4268±2131 pixels(P<0.05),while stimulation of A1 AR inhibitor for 48 hours alleviated the contraction.Conclusion:A1 AR expressed mainly in the cytoplasm of mice mesangial cells,adenosine and A1 AR played an important role in mesangial contraction.