五味子乙素通过调控细胞周期蛋白抑制大鼠心肌成纤维细胞增殖研究

Schisandrin B Inhibits Cardiac Fibroblast Proliferation via Regulating Cell Cycle Protein

目的观察五味子乙素(Schisandrin B,SchB)对血管紧张素Ⅱ(AngⅡ)诱导的新生大鼠心肌成纤维细胞(CFb)增殖、细胞周期、细胞周期蛋白(cyclin)A和E、细胞周期抑制蛋白P21^(cip1)和P27^(kip1)蛋白的表达水平及Ⅰ型胶原(TypeⅠcollagen)和Ⅲ型胶原(TypeⅢcollagen)蛋白表达水平的影响.方法将CFb分为对照组、模型组、SchB低剂量组(SchB-L,1μmol/L)、SchB中剂量组(SchB-M,10μmol/L)和SchB高剂量组(SchB-H,30μmol/L)共5组.采用胰酶消化、差速贴壁法培养CFb,四氮唑盐(MTT)比色法检测CFb增殖情况,流式细胞术检测细胞周期变化,酶联免疫法(ELISA)法检测CFb中Ⅰ和Ⅲ型胶原蛋白表达水平,免疫印迹(Wb)法检测细胞周期蛋白A和E、P21^(cip1)和P27^(kip1)蛋白的表达水平.结果与AngⅡ组比较,SchB干预组能显著降低CFb增殖率(P<0.05或P<0.001),降低CFb中collagenⅠ和collagenⅢ蛋白表达水平(P<0.05或P<0.01),升高CFb中G_(0)/G_(1)期细胞百分率,并降低S期细胞百分率(P<0.05或P<0.01),明显降低CFb中cyclinA和cyclinE蛋白表达水平(P<0.01),增加CFb中P21^(cip1)和P27^(kip1)蛋白表达水平(P<0.01).结论SchB抑制AngⅡ诱导的CFb增殖和胶原蛋白沉积,其作用机制可能与调控细胞周期蛋白表达水平有关.

Objective To observe the effects of Schisandrin B(SchB)on neonatal rat cardiac fibroblasts(CFb)proliferation,cell cycle,cyclin A,E,P21^(cip1),P27^(kip1) protein expression and Type I,Ⅲcollagen protein expression induced by angiotensinⅡ(AngⅡ).Method The experimental CFb was divided into five groups:control group,model group,low dose of SchB group(SchB-L,1μmol/L),medium dose of SchB group(SchB-M,10μmol/L)and high dose of SchB(SchB-H,30μmol/L).CFb was cultured by trypsin digestion and differential adhesion method,MTT colorimetry was used to detect cell proliferation,flow cytometry method was used to detect cell cycle,enzyme linked immunosorbent assay(ELISA)was used to detect TypeⅠandⅢcollagen protein expression levels,the protein expression of cyclinsA,E,P21^(cip1) and P27^(kip1) was detected by Western blotting method.Results Compared with AngⅡgroup,the proliferation inhibiting rate of CFb was significantly decreased(P<0.05 or P<0.01),the protein expression levels of TypeⅠandⅢcollagen were decreased(P<0.05 or P<0.1),the percentage of CFb in G_(0)/G_(1) phase was increased,and the percentage of CFb in S phase was decreased(P<0.05 or P<0.01),the protein expression level of cyclinA and cyclinE were significantly decreased(P<0.01)and the expression levels of P21^(cip1) and P27^(kip1) protein were significantly increased(P<0.01).Conclusion SchB can inhibit the proli-feration and collagen deposition of CFb induced by AngⅡ,and its mechanism is related to regulating cyclin protein expression possibly.