糜酶介导心脏成纤维细胞增殖的信息交流机制

Information exchange mechanism of proliferation of cardiac fibroblasts induced by chymase

目的探讨糜酶介导大鼠心脏成纤维细胞(CFs)增殖的细胞内信息交流机制。方法采用胰酶消化法分离、培养新生大鼠的CFs,通过四氮唑盐(MTT)比色法(A490值)测定细胞数目,免疫印迹法(Western blot)检测转化生长因子-β1(TGF-β1)、p-Smad2/3和过氧化物酶体增殖物激活受体α(PPARα)的蛋白表达水平。结果糜酶促使CFs数目增加并呈剂量依赖性,15、30和60μg/L组的A490值分别为0.263±0.033、0.348±0.031和0.387±0.026,均较对照组(0.201±0.019)显著增多(P<0.01)。15、30和60μg/L糜酶组的TGF-β1蛋白表达水平分别为0.968±0.069,1.782±0.058和2.656±0.085,p-Smad2/3蛋白表达水平分别为0.506±0.019,0.629±0.038和0.883±0.031,均明显高于对照组(0.333±0.023和0.287±0.016,P<0.05或<0.01)。糜酶可减低CFs的PPARα蛋白表达水平,15、30和60μg/L糜酶组的PPARα蛋白表达分别为0.430±0.025、0.289±0.026和0.165±0.013,均较对照组(0.740±0.041)显著减少(P<0.05或<0.01)。结论糜酶能促进大鼠CFS增殖,其机制与TGF-β1和p-Smad2/3蛋白表达增加、PPARα蛋白表达减少有关,提示TGF-β1/Smad和PPARα信号通路存在信息交流。

Objective To investigate the mechanism of intracellular information exchange in chymase-mediated proliferation of cardiac fibroblasts(CFs)in rats.Methods The CFs of neonatal rats were isolated and cultured by trypsinization.The number of cells was determined by MTT assay(A490 value),and protein expression levels of transforming growth factor1(TGF1),p Smad2/3 and peroxisome proliferator-activated receptorα(PPARα)were detected by Western blot.Results The chymase increased the number of CFs in a dose-dependent manner.The A490 values in 15,30,and 60μg/L groups were 0.263±0.033,0.348±0.031,0.387±0.026,respectively,which were significantly higher than those(0.201±0.019)in control group(P<0.01).The expression levels of TGF-β1 protein in 15,30,and 60μg/L chymase groups were 0.968±0.069,1.78±0.058,and 2.656±0.085,respectively,whereas those of p Smad2/3 protein were 0.506±0.019,0.629±0.038,and 0.883±0.031,respectively,which were significantly higher than those in control group(0.333±0.023 and 0.287±0.016;P<0.05 or<0.01).The chymase reduced the expression levels of PPARαprotein in CFs,and the PPARαprotein expression levels in 15,30,and 60μg/L chymase groups were 0.430±0.025,0.289±0.026,and 0.165±0.013,respectively,which were significantly lower than those in control group(0.740±0.041;P<0.05 or<0.01).Conclusion The chymase can promote the proliferation of CFS in rats,which is associated with up-regulation of TGF-β1 and p Smad2/3 protein expression as well as down-regulation of PPARαprotein expression,which suggests that there exists the information exchange in TGF-β1/Smad and PPARαsignaling pathways.Chymase can promote the proliferation of rat CFs,which is associated with the upregulation of TGF-β1 and p-Smad2/3 protein,the downregulation of PPARαprotein.It indicated that there was a cross-talk between TGF-β1/Smad and PPARαsignal passway.