红景天苷对低氧环境下大鼠阴茎海绵体平滑肌细胞缝隙连接蛋白43表达的影响

Effect of salidroside on the expression of connexin43 in the corpus cavernosum smooth muscle cells of hypoxic rats

目的:探讨红景天苷对低氧环境下SD大鼠阴茎海绵体平滑肌细胞(CCSMC)缝隙连接蛋白43(Cx43)表达的影响。方法:体外培养SD大鼠CCSMC,免疫组化法鉴定细胞;设常氧对照组(21%O_2)、低氧组(1%O_2)、低氧+红景天苷8μg/ml组、低氧+红景天苷16μg/ml组、低氧+红景天苷32μg/ml组和低氧+红景天苷64μg/ml组,分别培养48 h;Western印迹测定Cx43相对表达量。结果:体外培养CCSMC生长良好,抗α-平滑肌肌动蛋白(α-SMA)和抗desmin抗体免疫组化染色阳性,CCSMC占比90%以上。MTT法结果显示,红景天苷浓度≤64μg/ml时,对CCSMC无明显毒性;而128μg/ml和256μg/ml红景天苷对CCSMC有一定的抑制作用。未加红景天苷干预时,与常氧对照组相比,低氧组Cx43总蛋白和磷酸化蛋白表达均显著升高(P<0.01和P<0.05),磷酸化比例未见明显变化(P>0.05);与低氧不加药组相比,不同浓度红景天苷干预后Cx43表达量仍显著升高,但Cx43蛋白磷酸化比例显著降低,差异均有显著性(P均<0.05)。结论:低氧促进大鼠CCSMC Cx43表达升高,红景天苷浓度≤64μg/ml无法逆转上述缺氧改变,但降低了Cx43磷酸化比例。推测红景天苷能通过降低Cx43磷酸化比例抑制缺氧引起的缝隙连接通道形成,保护平滑肌细胞功能。

Objective： To investigate the effect of salidroside on the expression of the connexin43（Cx43） protein in the corpus cavernosum smooth muscle cells（ CCSMCs） of hypoxic SD rats. Methods： CCSMCs were cultured in vitro and identified by immunofluorescence staining. The cells were divided into six groups： normal control（ 21% O2）,hypoxia（1% O2）,hypoxia ＋ salidroside（HS） 8 μg/ml,HS 16 μg/ml,HS 32 μg/ml,and HS 64 μg/ml,and cultured for 48 hours. Then the relative expression of Cx43 in different groups was detected by Western blot. Results： The in vitro cultured CCSMCs grew well and 90% of the cells showed positivity for α-SMA and desmin on immunohistochemistry. Salidroside ≤64 μg / ml produced no obvious toxicity on the CCSMCs. The expressions of Cx43 and phosphorylated proteins were dramatically increased in the hypoxia group as compared with the normal control（ P 0. 01 and P 0. 05）. The HS groups all showed significantly higher expression of Cx43 than the hypoxia group（ P 0. 01）,but the phosphorylation rate of the Cx43 proteins was remarkably decreased（ P 0. 01）. Conclusion： Hypoxia increases the expression of Cx43 in the CCSMCs of SD rats. Salidroside ≤64 μg / ml cannot reverse the hypoxia-induced change but can reduce the dephosphorylation of Cx43 in CCSMCs. It is deduced that salidroside can protect CCSMCs by decreasing the phosphorylation of Cx43 and suppressing hypoxia-induced formation of the gap junction channel.