氯化钴诱导低氧对3T3-L1脂肪细胞脂连蛋白表达的影响

Effects of hypoxia induced by CoCl_2 on expression of adiponectin in 3T3-L1 adipocytes

目的研究体外模拟低氧对小鼠3T3-L1脂肪细胞脂连蛋白(ADPN)表达的影响。方法体外培养小鼠3T3-L1前脂肪细胞,将其诱导分化成为成熟的脂肪细胞,油红O染色鉴定脂肪细胞分化程度和脂质积聚情况。氯化钴(CoCl2)化学模拟脂肪细胞低氧环境,采用实时PCR(Real-Time PCR)检测低氧诱导因子-1α(HIF-1α)和ADPN mRNA的表达,蛋白质印迹法(Western blotting)检测低氧诱导因子-1α(HIF-1α)的蛋白表达;酶联免疫吸附测定(ELISA)分析细胞培养上清液中ADPN蛋白的分泌水平。结果在CoCl2诱导3T3-L1脂肪细胞低氧的条件下,细胞内HIF-1αmRNA和蛋白的表达水平均显著上调;细胞内ADPN mRNA表达和细胞培养上清液中ADPN蛋白的分泌水平均显著下降;HIF-1αmRNA与ADPN mRNA的表达水平呈显著负相关(r=-0.854,P<0.001)。结论 CoCl2诱导低氧能够下调3T3-L1脂肪细胞ADPN的表达,该作用可能与脂肪细胞代谢功能障碍和胰岛素抵抗有关。

Objective To investigate the effects of hypoxia induced by CoC12 on the expression of adiponectin （ADPN） in mouse 3T3-L1 adipocytes. Methods 3T3-L1 preadipocytes were cultured and differentiated to matured adipocytes in vitro. Cell differentiation and lipid accumulation were determined by oil red O staining. CoC12 was used as a chemical hypoxia-inducible reagent to simulate hypoxic microenvironment. The expressions of hypoxia-inducible factor-lct （HIF-let） and ADPN mRNA were detected by Real-Time PCR. The protein expressions of HIF-lct were determined by Western blotting and the secretion levels of ADPN protein in cell culture supernatant were detected by ELISA. Results Under the hypoxia condition induced by COC12, the expression levels of HIF-la mRNA and protein in 3T3-L1 adipocytes were significantly increased, while the expressions of ADPN mRNA in 3T3-L1 adipocytes and the secretion levels of ADPN protein in cell culture supernatant were significantly decreased. The expression levels of HIF-lct mRNA and ADPN mRNA had significantly negative correlation （r = -0. 854, P 〈0. 001）. Conclusion The hypoxia induced by CoC12 can decrease the expression of ADPN in 3T3-L1 adipocytes. This effect may be relevant to the metabolic function impairment of adipose cells and insulin resistance.