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目的:研究低氧诱导因子HIF-1α在颏舌肌成肌分化和线粒体生物发生中的作用,探索可能的调控机制。方法:构建HIF-1α敲降质粒,慢病毒载体转染小鼠颏舌肌成肌细胞,设阴性对照组,用含2%马血清的分化培养基在低氧(1%O2)环境下诱导成肌细胞分化,western blot检测成肌细胞分化2、4、6 d,PGC-1β、AMPKα1、p AMPKα1的表达水平,免疫细胞化学染色观察低氧分化4 d时AMPK通道激活剂与阻滞剂对PGC-1β的表达影响,western blot定量分析。结果:1 HIF-1α敲降组的PGC-1β表达量高于对照组(P<0.05),且分化4 d时表达最高(P<0.05);2AMPKα1总蛋白表达量无明显差异,p AMPKα1的表达在HIF-1α敲降组远高于对照组(P<0.05)。3AMPK通路促进剂AICAR增加PGC-1β的表达(P<0.05),抑制剂Compound C明显抑制PGC-1β的表达(P<0.05)。结论:HIF-1α下调了低氧环境中PGC-1β的表达,PGC-1β的表达受AMPK通路的正向调节作用。
Objective: To investigate the role of hypoxia-inducible factor HIF-1α in genioglossal myogenic differentiation and mitochondrial biogenesis in genioglossus and to explore possible regulatory mechanisms. METHODS: HIF-1α knockdown plasmid was constructed. The lentiviral vector was transfected into genioglossus muscle of genioglossus. Negative control group was induced by hypoxia (1% O2) with 2% horse serum differentiation medium The expression of PGC-1β, AMPKα1, p AMPKα1 in myoblasts at 2, 4 and 6 days after differentiation was detected by western blot. The expressions of AMPKα1 and AMPKα1 were detected by immunocytochemical staining at 4 days after hypoxia On the expression of PGC-1β, western blot quantitative analysis. Results: 1 The expression level of PGC-1β in knockdown HIF-1α group was higher than that in control group (P <0.05), and the expression level of PGC-1β was the highest at 4 d (P <0.05); 2 There was no significant difference in the expression of total AMPKα1 protein The expression of HIF-1α knockdown group was much higher than that of the control group (P <0.05). AICAR, a 3AMPK pathway enhancer, increased the expression of PGC-1β (P <0.05). Inhibitor Compound C significantly inhibited the expression of PGC-1β (P <0.05). Conclusion: HIF-1α down-regulated the expression of PGC-1β in hypoxic environment. The expression of PGC-1β was positively regulated by AMPK pathway.