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目的:研究全反式维甲酸(all-trans retinoic acid,ATRA)对转化生长因子β(transforming growth factor-β,TGF-β)刺激系膜细胞环氧化酶2(COX-2)及Smad3,Smad7蛋白表达的影响,探讨ATRA在TGF-β/Smad信号通路中对系膜细胞表达COX-2的影响作用。方法:取培养第4代的大鼠肾小球系膜细胞分组:(1)对照组;(2)TGF-β刺激组(5、10ng/ml);(3)TGF-β+NS398[COX-2抑制剂组(10μmol/LNS398+10ng/mlTGF-β)];(4)TGF-β+Staurosporine(Smad抑制剂)组(5nmol/ml Staurosporine+10ng/mlTGF-β);(5)药物组:ATRA(10-3、10-6、10-9mol/L)组,在不同的作用时间(4h,12h,24h),WesternBlot印迹法测定各组COX-2、Smad3、Smad7蛋白的表达变化。结果:(1)在外源性TGF-β刺激下,COX-2、Smad3、Smad7蛋白的表达明显增加,各组与对照组比较,差异有统计学意义(P<0.05)。(2)加入COX-2抑制剂NS-398和Smad抑制剂Staurosporine后,COX-2、Smad3、Smad7表达均减少,与TGF-β组比较,差异有统计学意义(P<0.05)。(3)ATRA各浓度组和TGF-β组比较COX-2、Smad3、Smad7蛋白的表达明显减少(P<0.05),呈时间和浓度依赖性。(4)COX-2蛋白的表达与Smad3、Smad7蛋白的表达均呈显著正相关(r=0.978,r=0.942,P<0.05)。结论:ATRA可能通过TGF-β/Smad信号通路抑制系膜细胞COX-2的表达,显示其具有良好的抗纤维化和抗细胞增殖作用。
Objective: To investigate the effect of all-trans retinoic acid (ATRA) on the expression of cyclooxygenase-2 (COX-2) and Smad3 in mesangial cells induced by transforming growth factor- Smad7 protein expression, explore the ATRA TGF-β / Smad signaling pathway mesangial cells expression of COX-2 impact. Methods: The rat glomerular mesangial cells were divided into 4 groups: (1) control group, (2) TGF-β stimulation group (5 and 10 ng / ml), (3) TGF-β + NS398 [COX -2 inhibitor group (10μmol / L NS398 + 10ng / ml TGF-β)]; (4) TGF-β + Staurosporine group (5nmol / ml Staurosporine + 10ng / : ATRA (10-3, 10-6, 10-9mol / L). The expressions of COX-2, Smad3 and Smad7 in each group were measured by western blotting at different time points (4h, 12h, 24h) Results: (1) The expression of COX-2, Smad3 and Smad7 were significantly increased in exogenous TGF-β stimulation group, the difference was statistically significant between each group and the control group (P <0.05). (2) The expression of COX-2, Smad3 and Smad7 decreased after adding COX-2 inhibitor NS-398 and Smad inhibitor Staurosporine, which had statistical significance compared with TGF-β group (P <0.05). (3) The expressions of COX-2, Smad3 and Smad7 in ATRA groups were significantly decreased compared with those in TGF-β group (P <0.05) in a time and concentration-dependent manner. (4) The expression of COX-2 protein was positively correlated with the expression of Smad3 and Smad7 (r = 0.978, r = 0.942, P <0.05). CONCLUSION: ATRA may inhibit COX-2 expression in mesangial cells through TGF-β / Smad signaling pathway, indicating that it has good anti-fibrosis and anti-cell proliferation effects.