摘要
目的:探讨序列相似家族3成员C(FAM3C)在转化生长因子β1(TGF-β1)诱导人血管平滑肌细胞(VSMCs)表型转换中的作用及机制。方法:检测不同浓度(0、1、2、5及10μg/L)的TGF-β1作用24 h和10μg/L的TGF-β1作用不同时间(0、6、12、24及48 h)对原代人VSMCs中FAM3C表达的影响;检测TGF-β1(10μg/L)作用不同时间(0、6、12和24 h)对VSMCs表型转换的影响;敲减FAM3C、过表达FAM3C及加入蛋白激酶B(PKB/Akt)抑制剂Ⅷ(Akti-Ⅷ)后,Western blot检测骨桥蛋白(OPN)、波形蛋白(vimentin)、α-平滑肌肌动蛋白(α-SMA)、增殖细胞核抗原(PCNA)、FAM3C、Akt及磷酸化Akt(p-Akt)蛋白水平,CCK-8检测细胞活力,Transwell检测细胞迁移,免疫荧光检测vimentin及α-SMA蛋白表达。结果:随着TGF-β1作用浓度(0、1、2、5及10μg/L)和时间(0、6、12、24及48 h)的增加,VSMCs中FAM3C蛋白表达上调。TGF-β1(10μg/L)呈时间依赖性(0、6、12和24 h)上调OPN及vimentin表达,下调α-SMA表达。与si-NC组相比,敲减FAM3C后,OPN、vimentin、PCNA及p-Akt蛋白水平下调(P<0.05),α-SMA表达上调(P<0.05),细胞活力及迁移能力减弱(P<0.05);此外,敲减FAM3C减弱了TGF-β1对VSMCs的作用。而与LV-GFP组相比,过表达FAM3C后,OPN、vimentin、PCNA及p-Akt蛋白水平升高(P<0.05),α-SMA蛋白表达减少(P<0.05),细胞活力及迁移能力增强;过表达FAM3C促进了TGF-β1对VSMCs表型转换、活力及迁移的作用,而Akti-Ⅷ抑制了过表达FAM3C及与TGF-β1共作用对VSMCs表型的调节。结论:FAM3C通过Akt介导TGF-β1诱导的人VSMCs表型转换、活力增强及迁移,可能是干预内膜增生的潜在靶点。
AIM:To investigate the effect of family with sequence similarity 3 member C(FAM3C)on transforming growth factor-β1(TGF-β1)-induced phenotypic switch of human vascular smooth muscle cells(VSMCs),and to explore its mechanism.METHODS:Primary human VSMCs were treated with different concentrations(0,1,2,5 and10μg/L)of TGF-β1 for 24 h or 10μg/L TGF-β1 for different time(0,6,12,24 and 48 h),and then the expression of FAM3C in these cells was detected.The effect of TGF-β1(10μg/L)treatment for different time(0,6,12 and 24 h)on phenotypic switch of the VSMCs was detected.After knockdown or overexpression of FAM3C combined with protein kinase B(PKB/Akt)inhibitorⅧ(Akti-Ⅷ)treatment,the protein levels of osteopontin(OPN),vimentin,α-smooth muscle actin(α-SMA),proliferating cell nuclear antigen(PCNA),FAM3C,Akt and phosphorylated Akt(p-Akt)were detected by Western blot,cell viability and migration were detected by CCK-8 and Transwell assays,and the protein expression of vimentin andα-SMA was detected by immunofluorescence.RESULTS:Treatment with TGF-β1 up-regulated FAM3C protein expression in a concentration-and time-dependent manner.Treatment with TGF-β1(10μg/L)up-regulated the expression of OPN and vimentin,but down-regulated the expression ofα-SMA in a time-dependent manner.After knockdown of FAM3C,compared with si-NC group,the protein levels of OPN,vimentin,PCNA and p-Akt were down-regulated(P<0.05),the expression ofα-SMA was up-regulated(P<0.05),and the cell viability and migration were decreased(P<0.05).In addition,knockdown of FAM3C attenuated the effects of TGF-β1 on the VSMCs.After overexpression of FAM3C,compared with LV-GFP group,the protein levels of OPN,vimentin,PCNA and p-Akt increased in LV-FAM3C group(P<0.05),while the protein expression ofα-SMA decreased(P<0.05).Overexpression of FAM3C promoted the effects of TGF-β1 on phenotype switch,viability and migration of the VSMCs.However,the effects of FAM3C overexpression or combined with TGF-β1 on phenotypic switch of the VSMCs were attenuated by Akti-Ⅷ.CONCLUSION:FAM3C mediates TGF-β1-induced phenotypic switch,viability enhancement and migration of human VSMCs through Akt,and it may be a potential target for treatment of intimal hyperplasia.
作者
訾亚飞
王正力
訾亚婉
李雅馨
刘杨东
赵渝
ZI Ya-fei;WANG Zheng-li;ZI Ya-wan;LI Ya-xin;LIU Yang-dong;ZHAO Yu(Department of Vascular Surgery,First Affiliated Hospital,Chongqing Medical University,Chongqing 400016,China;Department of Pulmonary and Critical Care Medicine,First Affiliated Hospital,Chongqing Medical University,Chongqing 400016,China;Department of Vascular Surgery,South China Hospital,Shenzhen University,Shenzhen 518100,China)
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2022年第12期2130-2138,共9页
Chinese Journal of Pathophysiology
基金
重庆市卫生与计划生育委员会基金项目(No.2016ZDXM004)。
关键词
序列相似家族3成员C
转化生长因子Β1
血管平滑肌细胞
表型转换
Family with sequence similarity 3 member C
Transforming growth factor-β1
Vascular smooth muscle cells
Phenotypic switch
