H_2O_2诱导小鼠肌原细胞系C2C12 UII/UT系统的高表达

H_2O_2 Induced the Up-regulation of UII/UT System in Mouse Myogenic Cell Line C2C12

目的:观察外源给予H2O2对C2C12细胞中UII/UT系统表达的影响。方法:培养小鼠肌原细胞系C2C12细胞株,应用胎盘蓝染色和测定细胞培养液中LDH的含量检测H2O2对细胞损伤的影响,应用放射免疫的方法检测细胞培养液和裂解液中UII的含量,应用RT-PCR法测定C2C12中UT mRNA的表达,应用Western Blot检测不同浓度H2O2对肌原细胞系C2C12中UT蛋白表达的影响。结果:外源给予不同浓度的H2O2,C2C12细胞裂解液中UII的含量各组间无明显差异,但增加了细胞孵育液中UII的含量,10-4 M和10-3 M时分别增加了83.1%(p<0.01)和94.5%(P<0.01)。低浓度H2O2(0.5×10-6 M,10-6 M,10-5 M,10-4 M)刺激明显增加了UT m RNA的表达,而高浓度的H2O2(10-3M)刺激反而使UT mRNA的表达降低。高浓度的H2O2(10-6 M,10-5M,10-4 M,10-3M)刺激使UT蛋白表达分别增加了200.1%、255.6%、111.1%和100.1%(均p<0.05)。结论:H2O2作为T2DM发病因素之一的活性氧族,可能是T2DM时骨骼肌组织中UII/UT系统表达增加的一个因素。

Objective： To observe the effect of H2O2 on the expression of UII/UT system in C2C12. Methods： Cell injury was determined by Placenta blue staining and LDH kit, the content of UII in cell lysate and cell incubation fluid were assayed by RIA, mRNA levels were assayed by RT-PCR, Protein levels were assayed by Western Blot. Results： UII content in cell incubation fluid was increased after H2O2 of different concentration was administrated, the UII content was increased by 83.1 %（P〈0.01） and 94.5 %（P〈0.01） when the concentration of H2O2 was elevated to 10-4M and 10-3M. The simulation of different concentration of H2O2（0.5×10-6M, 10-6M, 10-5M and 10-4M） significantly increased the expression of UT mRNA. However, The expression of UT mRNA was decreased significantly when the concentration of H2O2 was elevated to 10-3M. The UT protein expression was increased by 200.1%, 200.1%, 255.6% and200.1% when the concentration of H2O2 was elevated to 10-6M, 10-5M, 10-4M and 10-3M（P〈0.05）. Conclusions： H2O2 might be a factor involved in up-regulating the expression of UII/UT system for the skeletal muscle of Type 2 diabetes mellitus.