NYGGF4基因过表达对成熟脂肪细胞线粒体形态及动力学的影响

Influence of Over-Expression of NYGGF4 Gene on Mitochondrial Morphology and Dynamics in Adipocytes

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 explore the effect of NYGGF4 gene over-expression on mitochondrial morphology and dynamics in 3T3-L1 adipocytes.Methods 3T3-L1 preadipocytes transfected with either an empty expression vector or a NYGGF4 expression vector were induced to differentiation.The ultrastructure of the mitochondria in adipocytes was visualized using transmission electron microscope after diffe-rentiation.The levels of mitofusin（Mfn）1 mRNA,Mfn2 mRNA and dynamin related protein 1（Drp1） mRNA expression were detected by real-time quantitative PCR.The levels of Mfn1,Mfn2 protein and Drp1 protein expression were detected by Western blot.Results 1.The mitochondria in NYGGF4-over expressing adipocytes were smaller in size with abnormal morphology.Signs of mitochondrial damage ranged from swelling and reduced density to virtually hollow mitochondria.2.The levels of Mfn1 mRNA and Mfn1 protein were dramatically increased in NYGGF4-over expressing adipocytes compared with controls.3.There was no significant difference in the levels of Mfn2 and Drp1 both in mRNA and protein expression between the 2 groups.Conclusions In adipocytes with NYGGF4 gene over-expression,mitochondrial morphology is changed manifestly.Meanwhile,the expressions of Mfn1 mRNA and protein are up-regulated.It can implicate that mitochondrial morphology and dynamics are influenced in adipocytes with NYGGF4 gene over-expression.