CRAMP对小鼠心脏成纤维细胞激活、增殖和胶原分泌功能的影响  

Effects of CRAMP on mouse cardiac fibroblasts activation,proliferation and collagen secretion

作　　者：董伟[1] 罗强 高路[2] 肖莉丽[2] DONG Wei;LUO Qiang;GAO Lu;XIAO Lili(Department of Pediatrics,the First Affiliated Hospital,Zhengzhou University,Zhengzhou 450052;Department of Cardiovascular Medicine,the First Affiliated Hospital,Zhengzhou University,Zhengzhou 450052)

机构地区：[1]郑州大学第一附属医院儿科,郑州450052 [2]郑州大学第一附属医院心血管内科,郑州450052

出　　处：《郑州大学学报（医学版）》2022年第4期466-472,共7页Journal of Zhengzhou University(Medical Sciences)

基　　金：河南省科技厅科技攻关项目(202102310045);河南省高等学校重点科研项目(20B320039)。

摘　　要：目的:探讨CRAMP对小鼠心脏成纤维细胞激活、增殖和胶原分泌功能的影响。方法:分离培养小鼠心脏成纤维细胞。第一部分实验将细胞分为3组:对照组、转化生长因子β1(TGF-β1)组(10 mg/L 48 h)、CRAMP组(TGF-β1处理后给予CRAMP 0.5 mg/L 24 h),采用MTT法检测细胞增殖活性,采用免疫荧光染色检测α-SMA、Ⅲ型胶原和PCNA蛋白表达水平,采用RT-PCR检测Ⅲ型胶原和α-SMA mRNA表达水平,采用Western blot检测Smad信号通路蛋白表达水平。第二部分实验将细胞分为3组:TGF-β1组(10 mg/L 48 h)、SIS3组(TGF-β1处理后给予SIS310μmol/L 24 h)、CRAMP+SIS3组(TGF-β1处理后给予CRAMP 0.5 mg/L和SIS310μmol/L 24 h),采用免疫荧光染色和RT-PCR检测Ⅲ型胶原和α-SMA蛋白及mRNA表达水平。结果:TGF-β1组成纤维细胞增殖活性,PCNA、Ⅲ型胶原、α-SMA蛋白表达水平,Ⅲ型胶原、α-SMA mRNA表达水平,p-Smad2、p-Smad3和Smad4蛋白表达水平均高于对照组;而CRAMP组上述指标均低于TGF-β1组(P<0.05)。SIS3组、CRAMP+SIS3组细胞Ⅲ型胶原、α-SMA蛋白及mRNA表达水平均低于TGF-β1组(P<0.05)。结论:CRAMP可通过拮抗Smad3信号通路抑制小鼠心脏成纤维细胞的激活、增殖和胶原分泌。Aim:To explore the effects of CRAMP on activation,proliferation and collagen secretion of mouse cardiac fibroblasts.Methods:In experimentⅠ,mouse cardiac fibroblasts were isolated and cultured.The cells were divided into three groups,control group,transforming growth factorβ1(TGF-β1)group(10 mg/L 48 hours),and CRAMP group(TGF-β1 and CRAMP 0.5 mg/L 24 hours).Cell proliferation activity was detected using MTT method.CollagenⅢ,α-SMA and PCNA protein levels were detected using immunofluorescence staining.RT-PCR was used to detect the mRNA levels of collagenⅢandα-SMA.Western blot was used to detect the Smad signaling pathway proteins.In experimentⅡ,the cells were divided into three groups,TGF-β1 group(10 mg/L 48 hours),SIS3 group(TGF-β1 and SIS310μmol/L 24 hours)and CRAMP+SIS3 group(TGF-β1 and CRAMP 0.5 mg/L,SIS310μmol/L 24 hours).CollagenⅢandα-SMA protein and mRNA levels were detected using immunofluorescence staining and RT-PCR.Results:The proliferation activity,PCNA,collagenⅢandα-SMA protein levels,collagenⅢandα-SMA mRNA levels,p-Smad2,p-Smad3,Smad4 protein levels of the TGF-β1 group were higher than those of the control group,while the above indexes of the CRAMP group were lower than those of the TGF-β1 group(P<0.05).CollagenⅢandα-SMA protein and mRNA levels in SIS3 group and CRAMP+SIS3 group were lower than those of the TGF-β1 group(P<0.05).Conclusion:CRAMP could inhibit mouse cardiac fibroblast activation,proliferation and collagen secretion by antagonizing the Smad3 signaling pathway.

关 键 词：心肌纤维化 心脏成纤维细胞 CRAMP SMAD 小鼠 

分 类 号：R542.2[医药卫生—心血管疾病]