论文部分内容阅读
目的探讨NYGGF4基因过表达对成熟3T3-L1脂肪细胞线粒体形态及动力学的影响。方法以3T3-L1前体脂肪细胞为载体,建立NYGGF4基因稳定过表达细胞株,以空载质粒转染的细胞为对照,将前体脂肪细胞诱导分化为成熟脂肪细胞。采用透射电镜观察成熟脂肪细胞的线粒体形态,采用荧光定量PCR技术检测成熟脂肪细胞中线粒体融合基因(Mfn)1 mRNA、Mfn2 mRNA、线粒体动态相关基因(Drp)1 mRNA的表达水平。采用Western blot方法检测Mfn1蛋白、Mfn2蛋白、Drp1蛋白的表达水平。结果 1.电镜观察发现NYGGF4基因过表达的成熟脂肪细胞线粒体体积变小、数量明显减少,线粒体嵴断裂、减少、消失,部分线粒体肿胀,甚至呈空泡状;对照组成熟脂肪细胞的线粒体形态基本正常,线粒体嵴清晰可见,线粒体无明显肿胀、皱缩。2.NYGGF4基因过表达成熟脂肪细胞Mfn1 mRNA及Mfn1蛋白表达水平均显著高于对照组。3.NYGGF4基因过表达成熟脂肪细胞的Mfn2 mRNA、Drp1mRNA及相应蛋白表达水平与对照组比较差异均无统计学意义。结论在3T3-L1成熟脂肪细胞中,NYGGF4基因过表达可导致线粒体形态发生变化、数量减少,同时上调Mfn1 mRNA和Mfn1蛋白表达水平,提示NYGGF4基因在成熟脂肪细胞中过表达能影响细胞线粒体形态及动力学。
Objective To investigate the effect of NYGGF4 overexpression on mitochondria morphology and kinetics of mature 3T3-L1 adipocytes. Methods 3T3-L1 preadipocytes were used as a vector to establish stable over-expressing NYGGF4 cell lines. The transfected cells were used as control to differentiate preadipocytes into mature adipocytes. The morphology of mitochondria of mature adipocytes was observed by transmission electron microscopy. The expression of Mfn1 mRNA, Mfn2 mRNA and the expression of Drp-1 mRNA in mature adipocytes were detected by real-time fluorescence quantitative PCR. Western blot was used to detect the expression of Mfn1, Mfn2 and Drp1. The results of electron microscopy showed that the mitochondria of mature adipocytes overexpressing NYGGF4 became smaller and smaller in number, mitochondrial cristae were broken, reduced and disappeared, some mitochondria swollen and even vacuolized. The mitochondrial morphology of mature adipocytes in control group Normal, mitochondrial cristae clearly visible, mitochondria no obvious swelling, shrinkage. The expression of Mfn1 mRNA and Mfn1 in overexpressed mature adipocytes ofNYGGF4were significantly higher than that of the control group. Mfn2 mRNA, Drp1mRNA and corresponding protein expression level of NGF gene over-expressing mature adipocytes were not significantly different from the control group. Conclusion Overexpression of NYGGF4 in 3T3-L1 mature adipocytes results in a change in mitochondrial morphology and a decrease in the number of mitochondria, as well as up-regulation of Mfn1 mRNA and Mfn1 protein expression, suggesting that overexpression of NYGGF4 in mature adipocytes may affect mitochondrial morphology and dynamics.