糜酶介导心脏成纤维细胞增殖的信息交流机制
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摘要
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,and60μg/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 were0. 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 PPARα protein in CFs,and the PPARα protein 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 PPARα protein expression,which suggests that there exists the information exchange in TGF-β1/Smad and PPARα signaling 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 PPARα protein. It indicated that there was a cross-talk between TGF-β1/Smad and PPARα signal passway.
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,and60μg/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 were0. 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 PPARα protein in CFs,and the PPARα protein 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 PPARα protein expression,which suggests that there exists the information exchange in TGF-β1/Smad and PPARα signaling 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 PPARα protein. It indicated that there was a cross-talk between TGF-β1/Smad and PPARα signal passway.
引文
1 Li J,Jubair S,Levick SP,et al.The artocrine role of tryptase in pressure overload-induced mast cell activation,chymase release and cardiac fibrosis.IJC Metab Endocr,2016,10:16-23.
2 Dong X,Zhang C,Ma S,et al.High concentrations of mast cell chymase facilitate the transduction of the transforming growth factor-β1/Smads signaling pathway in skin fibroblasts.Exp Ther Med,2015,9:955-960.
3 刘展晨,王靓,施慧,等.TGF-β1/Smad信号通路在心室重构中的调控机制研究进展.广西中医药大学学报,2016,19:87-90.
4 黄华,李宇声,金鑫,等.钩藤碱对自发性高血压大鼠心室重构过程中TGF-β1/Smad通路的影响.中国病理生理杂志,2015,31:1365-1370.
5 Drosatos K,Pollak NM,Pol CJ,et al.Cardiac myocyte KLF5 Regulates PPARa expression and cardiac function.Circ Res,2016,118:241-253.
6 Ajith TA,Jayakumar TG.Peroxisome proliferator-activated receptors in cardiac energy metabolism and cardiovascular disease.Clin Exp Pharmacol Physiol,2016,43:649-658.
7 张永,肖颖彬,郝嘉,等.过氧化物酶体增殖物激活受体α在心血管疾病中的研究进展.实用医学杂志,2012,28:1211-1213.
8 赵晓燕,赵连友,张兴凯,等.糜酶介导自发性高血压大鼠心肌纤维化的作用机制.西北国防医学杂志,2010,31:255-257.
9 Takai S,Jin D.Improvement of cardiovascular remodelling by chymase inhibitor.Clin Exp Pharmacol Physiol,2016,43:387-393.
10 王先梅,杨丽霞,宋武战,等.心脏糜酶活性改变在心肌肥厚患者心肌纤维化中的意义.中国病理生理杂志,2007,23:2327-2331.
11 宋晶,郭家娟,李相军,等.不同浓度的糜酶对自发性高血压大鼠心肌肥厚细胞的构建.中国老年杂志,2015,35:5751-5752.
12 Luo K.Signaling cross talk between TGF-β/Smad and other signaling pathways.Cold Spring Harb Perspect Biol,2017,9:32-38.
13 Wang J,Wang G,Sun GW.Role of PPARαin down-regulating AGEinduced TGF-βand MMP-9 expressions in chondrocytes.Genet Mol Res,2016,15:116-120.
14 Wu Y,Wang BX,Guo YY,et al.The effect of relgulation of PPAR-αon cardiac hypertrophy and the relationship between the effect of PPAR-αwith PI3K/Akt/m TOR pathway.Zhongguo Ying Yong Sheng Li Xue Za Zhi,2015,31:284-288.
15 Shimojo N,Jesmin NS,Sakai I S,et al.Fish oil constituent eicosapentaenoic acid inhibits endothelin-induced cardiomyocyte hypertrophy via PPAR-α.Life Sci,2014,118:173-178.
16 Lam VH,Zhang L,Huqi A,et al.Activating PPARαprevents postischemic contractile dysfunction in hypertrophied neonatal hearts.Circ Res,2015,117:41-51.
2 Dong X,Zhang C,Ma S,et al.High concentrations of mast cell chymase facilitate the transduction of the transforming growth factor-β1/Smads signaling pathway in skin fibroblasts.Exp Ther Med,2015,9:955-960.
3 刘展晨,王靓,施慧,等.TGF-β1/Smad信号通路在心室重构中的调控机制研究进展.广西中医药大学学报,2016,19:87-90.
4 黄华,李宇声,金鑫,等.钩藤碱对自发性高血压大鼠心室重构过程中TGF-β1/Smad通路的影响.中国病理生理杂志,2015,31:1365-1370.
5 Drosatos K,Pollak NM,Pol CJ,et al.Cardiac myocyte KLF5 Regulates PPARa expression and cardiac function.Circ Res,2016,118:241-253.
6 Ajith TA,Jayakumar TG.Peroxisome proliferator-activated receptors in cardiac energy metabolism and cardiovascular disease.Clin Exp Pharmacol Physiol,2016,43:649-658.
7 张永,肖颖彬,郝嘉,等.过氧化物酶体增殖物激活受体α在心血管疾病中的研究进展.实用医学杂志,2012,28:1211-1213.
8 赵晓燕,赵连友,张兴凯,等.糜酶介导自发性高血压大鼠心肌纤维化的作用机制.西北国防医学杂志,2010,31:255-257.
9 Takai S,Jin D.Improvement of cardiovascular remodelling by chymase inhibitor.Clin Exp Pharmacol Physiol,2016,43:387-393.
10 王先梅,杨丽霞,宋武战,等.心脏糜酶活性改变在心肌肥厚患者心肌纤维化中的意义.中国病理生理杂志,2007,23:2327-2331.
11 宋晶,郭家娟,李相军,等.不同浓度的糜酶对自发性高血压大鼠心肌肥厚细胞的构建.中国老年杂志,2015,35:5751-5752.
12 Luo K.Signaling cross talk between TGF-β/Smad and other signaling pathways.Cold Spring Harb Perspect Biol,2017,9:32-38.
13 Wang J,Wang G,Sun GW.Role of PPARαin down-regulating AGEinduced TGF-βand MMP-9 expressions in chondrocytes.Genet Mol Res,2016,15:116-120.
14 Wu Y,Wang BX,Guo YY,et al.The effect of relgulation of PPAR-αon cardiac hypertrophy and the relationship between the effect of PPAR-αwith PI3K/Akt/m TOR pathway.Zhongguo Ying Yong Sheng Li Xue Za Zhi,2015,31:284-288.
15 Shimojo N,Jesmin NS,Sakai I S,et al.Fish oil constituent eicosapentaenoic acid inhibits endothelin-induced cardiomyocyte hypertrophy via PPAR-α.Life Sci,2014,118:173-178.
16 Lam VH,Zhang L,Huqi A,et al.Activating PPARαprevents postischemic contractile dysfunction in hypertrophied neonatal hearts.Circ Res,2015,117:41-51.