益心泰含药血浆对血管紧张素Ⅱ诱导的心肌成纤维细胞增殖、凋亡的影响及机制研究  

Mechanism of Yixintai Medicated Plasma on Proliferation and Apoptosis of Cardiac Fibroblasts Induced by AngiotensionⅡ

作　　者：杨金伟[1] 郭志华[1] 李雅[1] 张彤瑜 邓奕辉[1] YANG Jinwei;GUO Zhihua;LI Ya;ZHANG Tongyu;DENG Yihui(Hunan University of Chinese Medicine,Changsha 410208 Hunan,China)

机构地区：[1]湖南中医药大学,湖南长沙410208

出　　处：《中药新药与临床药理》2021年第9期1245-1253,共9页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：湖南省中医药科研计划项目(2020002,201516);湖南省“国内一流建设学科”中医学开放基金项目(2018ZYX44);湖南省研究生科研创新项目(CX2018B471)。

摘　　要：目的探讨益心泰含药血浆对血管紧张素Ⅱ(AngⅡ)诱导的心肌成纤维细胞(CFs)增殖、凋亡的影响及相关机制。方法将分化完全的CFs随机分为空白血浆组、模型组、益心泰含药血浆组,每组6个复孔。除空白血浆组外,其余各组以AngⅡ(10-7 mol·L^(-1))诱导构建细胞增殖及纤维化模型。空白血浆组及模型组以15%空白血浆培养,益心泰含药血浆组以15%益心泰含药血浆培养,干预24 h。采用CCK-8法检测CFs细胞增殖情况;ELISA法测定细胞中Ⅰ型胶原(CollagenⅠ)、Ⅲ型胶原(CollagenⅢ)及转化生长因子(TGF-β1)含量;流式细胞术检测细胞凋亡及细胞周期;Western Blot法检测细胞中TGF-β1、B细胞白血病/淋巴瘤-2(Bcl-2)及Bcl-2相关X蛋白(Bax)的蛋白表达水平;RT-qPCR法检测CFs的增殖细胞核抗原(PCNA)、α-平滑肌肌动蛋白(α-SMA)、CollagenⅠ/ⅢmRNA表达水平。结果15%益心泰含药血浆对CFs无明显的促细胞增殖或抑制作用。与空白血浆组比较,模型组细胞G0/G1期的细胞百分率明显降低(P<0.01),S期和G2/M期的细胞百分率明显升高(P<0.01);细胞凋亡率有所下降,但差异无统计学意义(P>0.05);Ⅰ/Ⅲ型胶原含量及TGF-β1水平均显著升高(P<0.01);TGF-β1、Bcl-2蛋白表达明显上调(P<0.01),Bax蛋白表达明显下调(P<0.01);PCNA、α-SMA、CollagenⅠ/ⅢmRNA表达水平均明显升高(P<0.01)。与模型组比较,益心泰含药血浆组细胞的G0/G1期细胞百分率明显升高(P<0.01),S期和G2/M期的细胞百分率明显降低(P<0.01),细胞凋亡率明显升高(P<0.01);Ⅰ、Ⅲ型胶原含量及TGF-β1水平明显降低(P<0.01);TGF-β1、Bcl-2蛋白表达明显下调(P<0.01),Bax蛋白表达明显上调(P<0.01);PCNA、α-SMA、CollagenⅠ/ⅢmRNA表达水平均明显降低(P<0.01)。结论益心泰含药血浆能够抑制CFs细胞增殖分化及纤维化,可能与其调控Bax/Bcl-2平衡及下调纤维化相关因子表达有关。Objective To investigate the mechanism of Yixintai medicated plasma on AngiotensionⅡ(AngⅡ)-induced proliferation and apoptosis of Cardiac fibroblasts(CFs).Methods CF cells were pretreatment with AngⅡ(10-7 mol·L^(-1))to establish an experimental model for proliferation,which were randomly divided into 3 groups:blank plasma group,model group and Yixintai medicated plasma group,6 replicates in each group.The blank plasma group and model group were cultured with 15%blank plasma,and the Yixintai medicated plasma groups was cultured with 15%drug containing plasma,for 24 h respectively.The proliferation of CFs were detected by CCK-8 essay.ELISA was used to detect levels of Collagen typeⅠ(CollagenⅠ),Collagen typeⅢ(CollagenⅢ)and transforming growth factor-β1(TGF-β1).The cell cycle and apoptosis were determined using flow cytometry.The expression of B-cell lymphoma-2(Bcl-2),Bcl-2-associated X(Bax)and TGF-β1 were detected by Western Blot.RT-qPCR was used to examine the gene transcript levels of proliferating cell nuclear antigen(PCNA),α-smooth muscle actin(α-SMA),CollagenⅠand CollagenⅢ.Results 15%Yixintai medicated plasma showed no inhibition of cells.Compared with the blank plasma group,the results of flow cytometry demonstrated that AngⅡinduced G0/G1 phase block and increased S and G2/M phase,which delayed cell cycle progression in model group(P<0.01);the apoptosis rate decreased with no statistically significance(P>0.05);the contents of CollagenⅠ,CollagenⅢand TGF-β1 were significantly increased(P<0.01);the expression of Bcl-2 and TGF-β1 increased obviously,while the expression of Bax decreased(P<0.01);the mRNA expression levls of PCNA,α-SMA,CollagenⅠand CollagenⅢwere significantly up regulated in model group(P<0.01).Compared with model group,Yixintai medicated plasma could apparently arrested cells in G0/G1 and decreased those in S and G2/M phase(P<0.01);the apoptosis rate and CollagenⅠ,CollagenⅢ,TGF-β1 level reduced(P<0.01).Yixintai medicated plasma can significantly restrain th

关 键 词：益心泰含药血浆 心肌成纤维细胞 血管紧张素Ⅱ 细胞增殖 细胞凋亡 TGF-Β1 BAX/BCL-2 

分 类 号：R285.5[医药卫生—中药学]