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目的探讨NYGGF4(又称PID1)基因沉默对脂肪细胞线粒体代谢的影响。方法以3T3-L1前体脂肪细胞为研究对象,体外培养稳定转染NYGGF4基因沉默载体(pGPU6/GFP/Neo-shRNA,NYGGF4-RNAi)的3T3-L1前体脂肪细胞,以转染空载体(pG-PU6/GFP/Neo-shRNA,NC shRNA)的3T3-L1前体脂肪细胞作为对照,经1-甲基-3异丁基黄嘌呤加地塞米松、胰岛素方案诱导分化为成熟脂肪细胞,采用实时定量RCR技术检测成熟脂肪细胞中线粒体代谢酶指标:己糖激酶、乙酰辅酶A羧化酶柠檬酸合成酶、肉毒碱棕榈酰转移酶1、细胞色素C基因表达水平。结果 NYGGF4表达沉默与NYGGF4基因过表达相比,可以部分逆转3T3-L1脂肪细胞中己糖激酶、乙酰辅酶A羧化酶、柠檬酸合成酶、肉毒碱棕榈酰转移酶1、细胞色素C基因表达水平,差异均有统计学意义。结论NYGGF4基因过表达,下调3T3-L1脂肪细胞中定位于线粒体的代谢关键酶,提示NYGGF4基因的动态平衡对维持3T3-L1脂肪细胞的线粒体代谢具有重要作用。
Objective To investigate the effect of NYGGF4 (also known as PID1) gene silencing on the mitochondrial metabolism of adipocytes. Methods 3T3-L1 preadipocytes were cultured in vitro and transfected into 3T3-L1 preadipocytes stably transfected with NYGPA4 gene silencing vector (pGPU6 / GFP / Neo-shRNA, NYGGF4-RNAi) pG-PU6 / GFP / Neo-shRNA, NC shRNA) was used as a control to induce differentiation into mature adipocytes by using 1-methyl-3-isobutylxanthine plus dexamethasone and insulin. Real-time quantitative RCR was used to detect mitochondrial metabolic enzymes in mature adipocytes: hexokinase, acetyl-CoA carboxylase citrate synthase, carnitine palmitoyl transferase 1 and cytochrome C gene expression. Results Compared with the NYGGF4 gene overexpression, NYGGF4 silencing could partially reverse the effects of hexokinase, acetyl-CoA carboxylase, citrate synthase, carnitine palmitoyltransferase 1 and cytochrome C gene in 3T3-L1 adipocytes The expression levels and differences were statistically significant. Conclusion Overexpression of NYGGF4 down-regulates mitochondrial metabolism-critical enzymes in 3T3-L1 adipocytes, suggesting that homeostasis of NYGGF4 may play an important role in maintaining mitochondrial metabolism in 3T3-L1 adipocytes.