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目的观察PPAR-γ激活对血管紧张素Ⅱ诱导的体外培养条件下大鼠血管平滑肌(VSMCs)的细胞外基质(ECM)释放及结缔组织生长因子(CTGF)表达的影响。方法原代培养大鼠VSMCs,用不同浓度PPAR-γ激动剂rosiglitzone和/或抑制剂GW9662、BADGE预处理后,加入0.1μmol/L血管紧张素Ⅱ(AngⅡ)刺激24h。比色法检测各组细胞培养液中羟脯氨酸含量,实时荧光定量RT-PCR方法检测各组三型胶原(ColⅢ)、层粘连蛋白(FN)及CTGFmRNA表达,Western blotting检测各组CTGF、PPAR-γ蛋白表达情况,基于ELISA的DNA-binding检测各组PPAR-γ活性。结果 AngⅡ可增加VSMCs PPAR-γ表达,但明显抑制其活性(P<0.05,vs.Control),PPAR-γ激动剂rosiglitazone可显著增加PPAR-γ蛋白表达及活化(P<0.05,vs.AngⅡ)。rosiglitazone可降低细胞培养液中羟脯氨酸含量,下调VSMCs中ColⅢ、FN、CTGF mRNA表达,抑制CTGF蛋白表达,而其他PPAR-γ激动剂(pioglitazone、15-d-PGJ2)均有相似作用,PPAR-γ拮抗剂GW9662及BADGE可拮抗这一作用。结论在VSMCs中,PPAR-γ激活可抑制AngⅡ诱导的CTGF表达,同时抑制ECM沉积。提示PPAR-γ可能在血管纤维化中起重要作用。
Objective To observe the effect of PPAR-γ activation on the release of extracellular matrix (ECM) and the expression of connective tissue growth factor (CTGF) in rat vascular smooth muscle cells (VSMCs) induced by angiotensin Ⅱ. Methods Primary VSMCs were cultured in vitro and pretreated with various concentrations of PPAR-γ agonist rosiglitzone and / or inhibitors GW9662 and BADGE, and then stimulated with 0.1 μmol / L Ang Ⅱ for 24 h. The content of hydroxyproline in each group of cell culture medium was detected by colorimetric method. The expressions of Col Ⅲ, laminin (FN) and CTGF mRNA were detected by real-time fluorescent quantitative RT-PCR. The expressions of CTGF, PPAR-γ protein expression, ELISA-based DNA-binding assay PPAR-γ activity in each group. Results AngⅡ increased the PPAR-γ expression in VSMCs, but significantly inhibited the activity (P <0.05, vs.Control). The PPAR-γ agonist rosiglitazone significantly increased the PPAR-γ protein expression and activation (P <0.05, vs. AngⅡ) . Rosiglitazone reduced the content of hydroxyproline in cell culture medium, down-regulated the expression of ColⅢ, FN and CTGF mRNA in VSMCs and inhibited the expression of CTGF protein, while other PPAR-γ agonists (pioglitazone, 15-d-PGJ2) PPAR-γ antagonists GW9662 and BADGE can antagonize this effect. Conclusion In VSMCs, PPAR-γ activation can inhibit Ang Ⅱ-induced CTGF expression and inhibit ECM deposition. Tip PPAR-γ may play an important role in vascular fibrosis.