低氧对C2C12细胞分化不同阶段自噬和肌细胞生成素表达的影响

目的:研究低氧对C2C12细胞分化不同阶段自噬相关蛋白Beclin1、微管相关蛋白轻链3(microtubule-associated protein light chain 3,LC3B)和肌细胞生成素(myogenin)表达的影响。方法:C2C12细胞诱导分化1、2、3、4 d,在常氧和低氧状态下观察细胞分化情况。免疫细胞化学染色法检测自噬相关蛋白LC3B和Beclin1的表达。Western blot检测Beclin1、LC3B和myogenin的蛋白表达水平,采用SPSS 22.0软件对数据进行统计学分析。结果:1.与常氧组比较,低氧组分化形成的肌管数量较少,形态细小;2.免疫细胞化学染色结果显示,分化1、2、3、4 d低氧组细胞LC3B和Beclin1荧光强度均低于常氧组;3.Western blot结果显示分化前中期低氧组Beclin1、LC3B和myogenin蛋白表达水平显著低于常氧组(P<0.05)。随着分化时间的增加,低氧状态下LC3B的表达水平进一步降低,而myogenin表达水平回升。结论:低氧抑制C2C12细胞分化;低氧能降低细胞分化前中期Beclin1、LC3B和myogenin蛋白表达水平。

Effects of Hypoxia on Autophagy and Myogenin Expression in Different Stages of Differentiation of C2C12 Cells

Objective:To study the effects of hypoxia on the expression of Beclin1,LC3B and myogenin in different stages of differentiation of C2C12 cells.Methods:C2C12 cells were induced to differentiate for 1,2,3 and 4 days.The differentiation of C2C12 cells was observed under normoxia and hypoxia.Immunocytochemistry was used to detect the expression of autophagy-related proteins LC3B and Beclin1.Western blotting was used to detect the protein expression levels of Beclin1,LC3B and myogenin.SPSS22.0 software package was used to analyze the data.Results:The number of myotubes in hypoxia group was less than that in normoxia group,and the morphology of cells was smaller.Immunocytochemistry showed that the fluorescence intensity of LC3B and Beclin1 of hypoxia group at 1,2,3 and 4 days of differentiation was lower than that in normoxia group.Western blotting showed that the expression levels of Beclin1,LC3B and myogenin in the early and middle stages of differentiation in hypoxic group were significantly lower than the normoxia group(P<0.05).With the increase of differentiation time,the expression level of LC3B decreased further,while the expression level of myogenin increased.Conclusions:Hypoxia inhibits the differentiation of C2C12 cells;hypoxia can reduce the expression levels of Beclin1,LC3B and myogenin in the early and middle stages of cell differentiation.