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目的研究姜黄素和氯沙坦对血管紧张素(Ang)Ⅱ诱导的内皮间质细胞转化(End MT)的作用及机制。方法培养人脐静脉内皮细胞(HUVECs)然后分为4组:对照组(不加刺激药物的正常培养组);AngⅡ组(加入刺激浓度为0.1μmol/L或1μmol/L的AngⅡ);姜黄素+AngⅡ组(先加入12.5μmol/L的姜黄素预孵1 h,再加入1μmol/L的AngⅡ);氯沙坦+AngⅡ组(先加入10μmol/L的氯沙坦孵育2 h,再加入1μmol/L的AngⅡ);刺激24 h后行划痕试验检测各组细胞迁移能力改变和CCK-8检测细胞活力;刺激5 d后,显微镜观察各组细胞形态变化和Western blot检测各组血管内皮钙黏蛋白(VE-cadherin)、α-平滑肌肌动蛋白(α-SMA)、转化生长因子β_1(TGF-β_1)蛋白表达量。结果 HUVECs经AngⅡ刺激后形态改变,长梭形样细胞明显增多;姜黄素和氯沙坦可维持内皮细胞铺路石样形态。AngⅡ呈浓度依赖性地降低HUVECs存活率(P<0.01);各组存活率均>50%(均P<0.05)。相比对照组,AngⅡ促进HUVECs的迁移(P<0.05);相比AngⅡ组,加入姜黄素和氯沙坦可抑制HUVECs的迁移(均P<0.05)。AngⅡ可诱导EndMT,AngⅡ刺激5 d后VE-cadherin表达减弱(P=0.003),而α-SMA和TGF-β_1的表达都明显增强(均P<0.01),姜黄素和氯沙坦使VE-cadherin表达明显增加,而α-SMA和TGF-β_1的表达均明显下调。结论姜黄素和氯沙坦通过下调TGF-β_1表达抑制AngⅡ诱导的EndMT。
Objective To investigate the effects and mechanisms of curcumin and losartan on endothelium-derived cells (End MT) induced by angiotensin (Ang Ⅱ). Methods Human umbilical vein endothelial cells (HUVECs) were cultured and divided into 4 groups: control group (normal group without stimulation); Ang Ⅱ group (AngⅡ with stimulation concentration of 0.1μmol / L or 1μmol / L); curcumin + AngⅡgroup (12.5μmol / L curcumin preincubated for 1 h, then added 1 μmol / L AngⅡ); losartan + Ang Ⅱ group (first added 10μmol / L losartan for 2 h, then added 1μmol / L AngⅡ). After 24 h of stimulation, the migration of cells in each group was detected by scratch test and the viability of CCK-8 cells was measured. After stimulated for 5 days, the morphological changes of cells in each group were observed by microscopy and the levels of vascular endothelial calcium VE-cadherin, α-SMA and TGF-β_1 were detected by immunohistochemistry. Results The morphology of HUVECs stimulated by Ang Ⅱ was changed, and spindle-like cells were significantly increased. Curcumin and losartan could maintain the morphology of endothelial cells. Ang Ⅱ reduced the survival rate of HUVECs in a concentration-dependent manner (P <0.01). The survival rates of all groups were all higher than 50% (all P <0.05). Compared with the control group, AngⅡ promoted the migration of HUVECs (P <0.05). Compared with AngⅡgroup, the addition of curcumin and losartan inhibited the migration of HUVECs (all P <0.05). AngⅡinduced EndMT. The expression of VE-cadherin decreased (P = 0.003) and the expression ofα-SMA and TGF-β_1 significantly increased after AngⅡ stimulation for5d (all P <0.01). Curcumin and losartan induced VE- cadherin expression was significantly increased, while the expression of α-SMA and TGF-β 1 were significantly down-regulated. Conclusion Curcumin and losartan can inhibit AngⅡ-induced EndMT by down-regulating the expression of TGF-β_1.