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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 connexin 43 (Cx43) in the corpus cavernosum smooth muscle cells (CCSMCs) of SD rats under hypoxia. Methods: The CCSMCs of SD rats were cultured in vitro and the cells were identified by immunohistochemistry. The cells in normoxia control group (21% O2), hypoxia group (1% O2), hypoxia + salidroside 8μg / ml group + Salidroside 16μg / ml group, hypoxia + salidroside 32μg / ml group and hypoxia + salidroside 64μg / ml group, were cultured for 48 h; Cx43 relative expression was measured by Western blot. Results: CCSMC grew well in vitro. Immunohistochemical staining of α-smooth muscle actin (α-SMA) and anti-desmin antibody was positive, accounting for more than 90% of CCSMC. The result of MTT showed that salidroside had no obvious toxicity to CCSMC when its concentration was less than 64μg / ml, while salidroside at 128μg / ml and 256μg / ml had some inhibitory effect on CCSMC. Compared with normoxia control group, the expressions of Cx43 protein and phosphorylated protein in hypoxia group were significantly increased (P <0.01 and P <0.05), and no significant changes in phosphorylation ratio (P <0.05). Compared with hypoxia group, the expression of Cx43 in salidroside at different concentrations was still significantly increased, but the phosphorylation of Cx43 protein was significantly decreased (all P <0.05 ). CONCLUSION: Hypoxia can promote the expression of Cx43 in CCSMC. The concentration of salidroside ≤64μg / ml can not reverse the hypoxia, but reduce the proportion of Cx43 phosphorylation. It is speculated that salidroside can protect the function of smooth muscle cells by inhibiting the formation of gap junctional channels induced by hypoxia by decreasing the phosphorylation of Cx43.