丝裂原活化蛋白激酶类和蛋白激酶C在转化生长因子β1诱导肾小管细胞结缔组织生长因子表达中的作用

目的了解转化生长因子β1（TGF-β1）诱导肾小管细胞结缔组织生长因子（CTGF）表达的机制，特别是蛋白激酶C（PKC）和丝裂原活化蛋白激酶（MAPK）在CTGF基因表达中的作用及其对Smad磷酸化的影响。方法分别应用PKC抑制剂G06850以及MAPK的3个组成成分ERK、JNK和p38MAPK的抑制剂PD98059、U0126、SP600125和SB203580阻断相应通路，观察其对TGF．131诱导的CTGF表达以及Smad2／Smad3磷酸化的影响。结果TGF-β1（5μg／L）以时间依赖方式诱导HK-2细胞中Smad2／Smad3的磷酸化，从基础值0．87±0．09上升至2h时高峰2．350±0.11。PKC抑制剂G06850（5μmol／L）和ERK抑制剂PD98059（10μmol／L）、U0126（10μmol／L）可部分抑制TGF-β1诱导的CTGF表达，而p38MAPK抑制剂SB203580（20μmol／L）和JNK抑制剂SP600125（10μmol／L）对TGF-β1诱导的CTGF的表达无影响。PKC抑制剂G06850（5μmol／L）可减少TGF-β1诱导的Smad2／Smad3磷酸化，而ERK抑制剂PD98059（10μmol／L）和U0126（10μmol／L）对Smad2／Smad3的磷酸化没有影响。结论在肾小管上皮细胞中，TGF-β1诱导CTGF的表达需要PKC和Ras／MEK／ERK的参与。PKC以Smad依赖的方式参与肾小管上皮细胞中TGF-β1诱导的CTGF的表达，而Ras／MEK／ERK对CTGF表达的调节不依赖于Smads。

Role of MAPK and PKC pathway in transforming growth factor β1-induced connective tissue growth factor expression

Objective To investigate the signal transduction events that regulate CTGF synthesis and secretion in response to TGF-β1, especially the role of protein kinase C (PKC) and mitogen-activated protein kinase (MAPK) in TGF-β1-induced CTGF expression. Methods PKC was inhibited with Go6850 and ERK, JNK, p38 MAPK was blocked by PD98059, U0126, SP600125 and SB203580 respectively, and their effects on CTGF expression and Smad2/Smad3 phosphorylation wereanalyzed. Results TGF-β1 （5 μg/L） induced Smad2/Smad3 phosphorylation in a time-dependent manner from baseline 0.87±0.09 to 2.35±0.11 for 2 hours. The PKC inhibitor Go6850（5 μmol/L） and the ERK inhibitor PD98059（10 μmol/L） and U0126 （10 μmol/L）partially attenuated TGF-β1-stimulated expression of CTGF, while the p38 MAPK inhibitor SB203580（20 μmol/L） or the JNK inhibitor SP600125（10 μmol/L） had no such effect. In addition, phosphorylation of Smad2/Smad3 was not blocked by U0126（10 μmol/L） or PD98059 （10 μmol/L）, but by Go6850. Conclusions Ras/MEK/ERK and PKC activity are required for the TGF-β1 induction of CTGF in tubular epithelial cells. Furthermore, PKC activity is involved in the TGF-β1-induced CTGF expression in a smad-dependent manner, while Ras/MEK/ERK modulates CTGF expression independently on Smads.