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目的探讨饰胶蛋白聚糖(DCN)对大鼠系膜细胞(MsC)生长的抑制作用及其信号转导分子MAPKs和p21表达的影响。方法经脂质体介导将DCN基因转染体外培养的大鼠MsC,筛选和鉴定后,收集阳性细胞克隆的培养上清(DCN上清),将其加入正常MsC的培养液中,采用流式细胞仪检测细胞周期。用Western印迹法分别检测MAPKs,包括细胞外调节激酶(ERK)1/2、应激活化蛋白激酶(SAPK),氨基末端激酶(JNK)和p38和p21蛋白表达;用免疫荧光法观察p21在细胞中的表达。结果DCN上清明显抑制正常MsC的增殖,G2-M期细胞数明显减少,仅为对照组的35%(P<0.05);磷酸化ERK1/2及SAPK/JNK表达增强,分别为对照组的2.2倍、1.4倍及1.7倍、1.8倍;磷酸化p38无明显变化。DCN抗体呈浓度依赖性抑制磷酸化ERK1/2、SAPK/JNK的表达上调。DCN上清还可使细胞p21的表达明显增强,而DCN抗体也同样呈浓度依赖性地抑制其上调表达的作用,ERK1/2及SAPK/JNK通路抑制剂U0126和circumin均可抑制其上调表达的作用,分别为对照组的64%和61%;而p38通路抑制剂SB203580则对其无影响。结论DCN对肾MsC的生长有抑制作用,其机制可能经ERK1/2、SAPK/JNK和p21蛋白介导。
Objective To investigate the effect of decorin (DCN) on the growth of rat mesangial cells (MsC) and the expression of MAPKs and p21. Methods DCN gene was transfected into rat MsC cultured in vitro by lipofectamine. After screening and identification, the supernatant of DCN (DCN supernatant) was collected and added to the culture medium of normal MsC. Cytometry detects cell cycle. MAPKs were detected by Western blotting, including expression of extracellular regulated kinase (ERK) 1/2, SAPK, JNK and p38 and p21 protein. Immunocytochemistry was used to detect the expression of p21 in cells In the expression. Results The DCN supernatant significantly inhibited the proliferation of normal MsC cells, and the number of cells in G2-M phase was significantly decreased (35%, P <0.05). The phosphorylation of ERK1 / 2 and SAPK / JNK were increased in control group 2.2-fold, 1.4-fold and 1.7-fold, 1.8-fold; phosphorylated p38 no significant change. DCN antibody inhibited the phosphorylation of ERK1 / 2 in a concentration-dependent manner, and the expression of SAPK / JNK was up-regulated. DCN supernatants also significantly increased the expression of p21, while DCN antibody also inhibited its up-regulation in a concentration-dependent manner. Both ERK1 / 2 and SAPK / JNK pathway inhibitors U0126 and circumin could inhibit the up-regulated expression Which were respectively 64% and 61% of the control group, while the p38 pathway inhibitor SB203580 had no effect on it. Conclusions DCN can inhibit the growth of renal MsC and its mechanism may be mediated by ERK1 / 2, SAPK / JNK and p21 proteins.