ERK1/2 MAPK通路在TIMP-1抑制高糖诱导的大鼠肾小球系膜细胞凋亡中的作用

目的 探讨细胞外信号调节激酶(ERK1/2)丝裂原激活蛋白激酶(MAPK)通路在基质金属蛋白酶1组织抑制剂(TIMP-1)抑制高糖诱导的大鼠肾小球系膜细胞凋亡中的作用。 方法 将人正义、反义TIMP-1用脂质体法转染大鼠肾小球系膜细胞，并将细胞分为正常对照组、未转染组、空载体组、正义转染组和反义转染组。各组细胞分别用高糖(25 mmol/L)和(或)MEK(ERK1/2通路中的MAPK激酶)特异性抑制剂PD98059(50 μmol/L)刺激24 h。应用透射电镜观察细胞内部结构变化。Western 印迹观察磷酸化(p)的ERK1/2、P90rsk、Bad及总的P90rsk、Bad、Bcl-2蛋白的表达。 结果 加入PD98059后，各组细胞内可见凋亡小体形成、细胞皱缩、核固缩、染色体边集及膜泡样突变等现象，与未转染组比较，以反义转染组最明显，凋亡细胞数目最多；正义转染组细胞形态改变不十分明显，凋亡细胞数目相对较少。此外，加入PD98059后，ERK1/2、P90rsk、Bcl-2及Bad的磷酸化明显减少(P < 0.05)，ERK1/2和P90rsk的磷酸化以正义转染组最明显(P < 0.01)；总P90rsk、总Bad表达量在各组之间无显著差异。 结论 ERK1/2 通路在TIMP-1抑制高糖诱导的大鼠肾小球系膜细胞凋亡过程中起重要作用。

The role of ERK1/2 MAPK pathway in the inhibition of TIMP-1 on apoptosis in rat mesangial cells induced by high glucose

Objective To investigate the mechanism of ERK1/2 signal pathway in the inhibition of TIMP-1 on apoptosis of rat glomerular mesangial cells induced by high glucose. Methods The human TIMP-1 sense， antisense plasmid were transferred into rat mesangial cells through liposome. The cells were divided into control group， non-transfected group，veotor group，sense group and antisense group. After cultured in high glucose (HG 25 mmol/L) with the presence of PD98059(50 µmol/L)for 24 hours， cell structure was observed by electron microcope. Western blot was performed to test the expression of phospho-ERK1/2， phospho-P90rsk， phospho-Bad， Bad， P90rsk and Bcl-2. Results Electron microcope showed that PD98059 caused a significant increase of apoptosis bodies in every group of rat mesangial cells. In contrast with the control group， the expression levels of phospho-ERK1/2， phospho-P90rsk， phospho-Bad and Bcl-2 were inhibited by antisense TIMP-1 and PD98059(P < 0.05). Among these groups，the expression levels of total Bad and total P90rsk protein were not different. Conclusion TIMP-1 exerts its anti-apoptoic effect， at least in part， by activation of ERK1/2 pathway.