Apocynin通过TGF-β1/Smad信号通路对脂多糖心肌纤维化小鼠的作用及其对ColⅠa1、ColⅢa1的影响

Effect of Apocynin via TGF-β1/Smad signaling pathway on lipopolysaccharide induced myocardial fibrosis in mice and its effect on ColⅠa1 and ColⅢa1

目的探讨Apocynin通过TGF-β1/Smad信号通路对脂多糖(LPS)心肌纤维化(MF)小鼠的作用及其对ColⅠa1、ColⅢa1的影响。方法将40只C57BL/6雄性小鼠随机分成正常对照组、Apocynin对照组、MF模型组和Apocynin干预组(n=10)。连续4周,每周一次腹腔注射LPS(10 mg/kg)的方法建立心肌纤维化模型。Apocynin干预组则于LPS(10 mg/kg)处理前0.5 h连续4周每周一次腹腔注射Apocynin(10 mg/kg)。各组小鼠于4周末颈椎脱臼处死后,心肌组织天狼星红染色检测心肌纤维化程度,采用免疫组织化学法检测各组小鼠心肌组织中TGF-β1,Smad2/3,p-Smad 2/3蛋白相对表达量;采用蛋白印记法检测小鼠心肌组织中ColⅠa1、ColⅢa1蛋白的表达量。结果 Apocynin对照组较正常对照组心肌组织胶原面积分数以及TGF-β1、Smad2/3、p-Smad2/3蛋白表达量和纤维化因子ColⅠa1、ColⅢa1表达量差异无统计学意义(P>0.05);MF模型组较正常对照组心肌组织胶原面积分数以及TGF-β1、Smad2/3、p-Smad2/3蛋白表达量和纤维化因子ColⅠa1、ColⅢa1表达量增加(P<0.01);Apocynin干预组较MF模型组心肌组织胶原面积分数以及TGF-β1、Smad2/3、p-Smad2/3蛋白表达量和纤维化因子ColⅠa1、ColⅢa1表达量降低(P<0.01)。结论 Apocynin通过TGF-β1/Smad信号通路介导了纤维化因子ColⅠa1、ColⅢa1蛋白的下调,从而减轻了LPS诱导的小鼠心肌纤维化。

Objective To investigate the effect of Apocynin on myocardial fibrosis(MF)mice and its effects on ColⅠa1 and ColⅢa1 through TGF-β1/Smad signaling pathway.Methods Forty C57BL/6 male mice were randomly divided into normal control group,Apocynin control group,MF model group and Apocynin intervention group(n=10).The myocardial fibrosis model was established by intraperitoneal injection of LPS(10 mg/kg)once a week for 4 weeks.Apocynin(10 mg/kg)was injected intraperitoneally once a week for 4 weeks in the Apocynin intervention group at 0.5 h before LPS(10 mg/kg)treatment.After each group of mice was sacrificed at the end of the 4th week after cervical dislocation,myocardial fibrosis was detected by Sirius Red staining.The expression of TGF-β1,Smad2/3,and p-Smad 2/3 in myocardial tissue of each group were detected by immunohistochemistry.The relative protein expression was detected by Western blotting.The expression of ColⅠa1 and ColⅢa1 protein in myocardial tissue was detected by Western blot.Results There was no significant difference in the collagen area fraction of TGF-β1,Smad2/3,p-Smad2/3 protein and the expression of fibrotic factors ColⅠa1 and ColⅢa1 between the Apocynin control group and the normal control group(P>0.05).Compared with the normal control group,the collagen area fraction and the expression levels of TGF-β1,Smad2/3,and p-Smad2/3 proteins and the expression of the fibrogenic factors ColⅠa1 and ColⅢa1 in the MF model group were significantly increased(P<0.01).Compared with the MF model group,the collagen area fraction and the expression of TGF-β1,Smad2/3,p-Smad2/3 protein and the expression of the fibrogenic factors ColⅠa1 and ColIIIa1 were significantly decreased in the Apocynin intervention group(P<0.01).Conclusion Apocynin mediated the down-regulation of the fibrogenic factors ColⅠa1 and ColⅢa1 through the TGF-β1/Smad signaling pathway,thereby reducing LPS-induced myocardial fibrosis in mice.