ILK-siRNA抑制衰老大鼠系膜细胞纤维连接蛋白积聚的研究

目的观察整合素连接激酶（integrin linked kinase,ILK）小干扰RNA（small interference,siRNA）对衰老大鼠系膜细胞细胞外基质纤维连接蛋白（fibronectin,FN）的影响,探讨ILK在衰老肾脏肾小球硬化中的作用.方法分离培养3月龄和24月龄雄性Wistar大鼠原代肾小球系膜细胞（mesangial cells,MCs）,脂质体转染抑制ILK表达的siRNA或无关siRNA（con siRNA）.利用RT-PCR、Western blot和免疫荧光结合激光共聚焦显微镜技术检测ILK和FN的表达情况.结果ILK-siRNA转染可抑制3月龄和24月龄大鼠系膜细胞ILK和FN的表达水平（P＜0.05）;对24月龄大鼠系膜细胞的抑制程度更显著（P＜0.05）,几乎使ILK和FN下降到正常青年大鼠系膜细胞的水平.结论通过RNAi技术抑制ILK的表达,可减轻衰老大鼠系膜细胞FN的积聚,为肾脏器官衰老的防治提供新的干预靶点.

ILK-siRNA inhibites the accumulation of fibronectin in mesangial cells of aged rats

Objective To observe the effect of small interference RNA(siRNA) of intergrin linked kinase(ILK) on the accumulation of fibronectin(FN) in the mesangial cells of aged rats,then to explore the role of ILK in the process of the age-related glomerular sclerosis.Methods Primary mesangial cells(MCs) of 3-month-old and 24-month-old Wistar rats were isolated and cultured.Lipofectamine was applied to transfect ILK-siRNA,which could inhibit ILK expression,and ILK-con siRNA to the above cells.The expressions of ILK and FN were detected by real-time PCR,western-blot,and immunofluorescence combined with confocal laser scanning microscopy.Results The expressions of ILK and FN were inhibited by ILK siRNA transfection both in 3-month-old and 24-month-old MCs(P<0.05),especially in 24-month-old MCs(P<0.05),almost decreased to the normal levels of 3-month-old MCs.Conclusions FN in senescent MCs could be decreased by inhibiting ILK through RNA interference,which provide a new intervene target for preventing renal senescence.