红景天苷通过PDGFR/STAT3信号通路调控大鼠阴茎海绵体平滑肌细胞表型转化的实验研究

Salidroside regulates the phenotypic transformation of corpus cavernosum smooth muscle cells of rats through the PDGFR/STAT3 signaling pathway

目的:探索红景天苷(Salidroside)对大鼠阴茎海绵体平滑肌细胞(CCSMCs)表型转化的调控作用机制。方法:分离获取SD大鼠原代CCSMCs,在低氧环境中培养24 h并观察红景天苷(30μg/ml)的干预作用,Western印迹测HIF-1α、PDGF受体(PDGFR)及表型转化相关蛋白(α-SMA和Collagen I)的表达变化;CCSMCs培养体系中加入外源性PDGF-BB(20 ng/mL)处理24 h,同时观察红景天苷或PDGFR抑制剂Crenolanib(100 nmol/L)的干预作用,Western印迹检测PDGFR、磷酸化PDGFR、表型转化相关蛋白(α-SMA和Collagen I)、STAT3、磷酸化STAT3、STAT5及磷酸化STAT5蛋白的表达变化;在外源性PDGF-BB刺激CCSMCs的基础上,同时观察红景天苷、STAT3过表达、红景天苷+STAT3过表达的干预作用,Western印迹检测表型转化相关蛋白(α-SMA和Collagen I)、STAT3、磷酸化STAT3蛋白的表达变化。结果:CCSMCs在低氧环境中培养24 h后HIF-1α蛋白表达显著增高(P<0.05),提示成功构建CCSMCs低氧模型,同时CCSMCs中PDGFRα、PDGFRβ及Collagen I蛋白表达显著增高(P均<0.05),α-SMA蛋白表达显著下降(P<0.05),经红景天苷干预后,上述情况受到明显抑制(P均<0.05);CCSMCs受到外源性PDGF-BB刺激24 h后PDGFRα、PDGFRβ和STAT3磷酸化比例及Collagen I蛋白表达显著增高(P均<0.05),α-SMA蛋白表达显著下降(P<0.05),经Crenolanib或红景天苷干预后,上述情况受到明显抑制(P均<0.05),各组STAT5磷酸化比例差异无统计学意义(P>0.05);外源性PDGF-BB处理CCSMCs的同时运用红景天苷进行干预,若过表达STAT3可致CCSMCs中α-SMA蛋白表达显著下降(P<0.05),Collagen I蛋白表达显著增高(P<0.05)。结论:初步证实红景天苷可通过PDGFR/STAT3信号通路改善大鼠CCSMCs表型转化,尚需进一步验证探讨。

Objective:To explore the mechanism of Salidroside regulating the phenotypic transformation of cavernous smooth muscle cells(CCSMCs)in rats.Methods:Primary CCSMCs were isolated from male SD rats,cultured in hypoxic environment for 24 hours,and treated with Salidroside at 30μg/mL.Then the expressions of HIF-1α,platelet-derived growth factor receptor(PDGFR)and phenotypic transformation-related proteinsα-SMA and Collagen I were detected by Western blot.The culture system of the CCSMCs was treated with exogenous PDGF-BB at 20 ng/mL for 24 hours,and the effects of Salidroside or PDGFR inhibitor Crenolanib(100 nmol/L)were observed.The expressions of PDGFR,phosphorylated PDGFR,phenotypic transformation-related proteinsα-SMA and Collagen I,STAT3,phosphorylated STAT3,STAT5 and phosphorylated STAT5 were determined by Western blot.The intervention effects of Salidroside and/or the overexpression of STAT3 were observed after stimulation of the CCSMCs by exogenous PDGF-BB,followed by detection of the expressions of phenotypic transformation-related proteinsα-SMA and Collagen I,STAT3 and phosphorylated STAT3 proteins.Results:The expression of HIF-1αin the CCSMCs was significantly upregulated after cultured in hypoxic environment for 24 hours(P<0.05),suggesting the successful construction of the hypoxia model of CCSMCs.Meanwhile,the expressions of PDGFRα,PDGFRβand Collagen I were remarkably increased(all P<0.05),and that ofα-SMA markedly decreased(P<0.05)in the CCSMCs.However,the expressions of the all the proteins above were significantly inhibited by Salidroside intervention(all P<0.05).After stimulated by exogenous PDGF-BB for 24 hours,the phosphorylation ratios of PDGFRα,PDGFRβand STAT3 and the expression of Collagen I were significantly elevated(all P<0.05),that ofα-SMA remarkably reduced(P<0.05),and all were inhibited by intervention with crenolanib or Salidroside(all P<0.05).No statistically significant difference was observed in the STAT5 phosphorylation ratio between different groups(P>0.05).Overexpression of STAT3 in the CCSMCs treated with exogenous PDGF-BB and Salidroside significantly decreased the expression ofα-SMA(P<0.05)and increased that of Collagen I(P<0.05).Conclusion:Salidroside could improve the phenotypic transformation of CCSMCs in male rats through the PDGFR/STAT3 signaling pathway,which needs further exploration and verification.