LYRM1基因过表达对大鼠骨骼肌细胞线粒体功能的影响

Effect of Over-Expression of LYR Motif Containing 1 Gene on Mitochondrial Function in Skeletal Muscle Cells of Rats

目的评价LYRM1基因过表达对成熟骨骼肌细胞线粒体功能的影响。方法体外培养大鼠骨骼肌细胞株(L6),分别将pcDNA3.1及LYRM1-pcDNA3.1转染L6,通过G418筛选稳定表达株,应用Western blot技术验证其转染情况。诱导L6分化成熟后电镜观察线粒体形态;经Mitotracker red染色,应用流式细胞仪检测线粒体膜电位变化。结果 Western blot技术证实LYRM1表达质粒转染成功;LYRM1-pcDNA3.1组细胞线粒体凝集、形态扭曲变形及线粒体空泡化、线粒体嵴排列不规则,甚至断裂、溶解、消失;流式细胞仪检测结果显示LYRM1-pcDNA3.1组荧光值为3.7667±0.1007,pcDNA3.1组荧光值为8.7633±0.2286,二组比较差异有统计学意义(P<0.05)。结论 LYRM1基因在骨骼肌细胞中过表达能明显改变线粒体形态,降低骨骼肌细胞线粒体的膜电位,提示LYRM1基因在骨骼肌细胞中过表达影响了线粒体的功能。

Objective To explore the effect of LYR motif containing 1（LYRM1）gene over-expression on mitochondrial function in skeletal muscle cells of rats.Methods Rat myoblasts（L6）were transfected with either an empty expression vector（pcDNA3.1Myc/His B）or a LYRM1 expression vector.After screening culture by G418,the expression of LYRM1 protein was identified by Western blot.Ultra-structural analysis of mitochondrial rat skeletal muscle cells were visualized by transmission electron microscopy after differentiation.Simultaneously,the mitochondrial membrane potential was determined according to fluorescence intensity of mitotracker red.Results The LYRM1 protein was transfected successfully.Mitochondria morphology of LYRM1-pcDNA3.1 L6 was altered obviously.LYRM1-forced cells displayed condensed mitochondria with twisted and condensed cristae.By flow cytometry,the relative fluorescence intensity in LYRM1-pcDNA3.1 group was 3.766 7±0.100 7,while in pcDNA3.1 group was 8.763 3±0.228 6,the difference between them was significant（P0.05）.Conclusions In skeletal muscle cells with LYRM1 over-expression,mitochondria morphology was changed manifestly.Meanwhile,mitochondrial membrane potential of skeletal muscle cells was downregulated.It is implicated that mitochondrial function was influenced in rat skeletal muscle cells with LYRM1 over-expression.