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目的探讨LYRM1基因沉默对成熟脂肪细胞线粒体形态及相关基因的影响。方法以3T3-L1前体脂肪细胞为工具细胞,建立LYRM1基因沉默细胞株,以转染空载质粒的3T3-L1脂肪细胞作为阴性对照,将前体脂肪细胞诱导分化为成熟脂肪细胞,采用油红O染色观察脂肪细胞分化状态,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)1/2 mRNA、线粒体动态相关基因(Drp)1 mRNA、线粒体分裂基因(Fis)1 mRNA的表达水平。结果 1.油红O染色诱导分化第8天的脂肪细胞,发现85%以上脂肪细胞已分化为成熟脂肪细胞,细胞形态大而圆,胞质含大量被油红O染成亮红色的脂滴,比较发现2组细胞无论脂滴的大小、数量均无显著差异。2.透射电镜观察发现2组细胞的线粒体形态基本正常,线粒体无明显肿胀、皱缩,线粒体嵴清晰可见,2组细胞间线粒体数目无明显差异。3.LYRM1基因沉默成熟脂肪细胞的Mfn2 mRNA表达水平显著高于对照组成熟脂肪细胞。4.LYRM1基因沉默成熟脂肪细胞的Mfn1 mRNA、Drp1 mRNA和Fis1 mRNA表达水平与对照组成熟脂肪细胞的表达水平无明显差异。结论 LYRM1基因沉默能够引起成熟脂肪细胞Mfn2 mRNA表达水平升高,对线粒体形态结构并无显著影响,提示LYRM1基因沉默可部分影响成熟脂肪细胞线粒体形态相关基因。
Objective To investigate the effects of LYRM1 gene silencing on mitochondrial morphology and related genes in mature adipocytes. Methods 3T3-L1 preadipocytes were used as tool cells to establish LYRM1 gene silencing cell lines. The 3T3-L1 adipocytes transfected with empty vector were used as negative control. The precursor adipocytes were induced to differentiate into mature adipocytes by using oil Red O staining was used to observe the differentiation status of adipocytes. Transmission electron microscopy was used to observe the mitochondrial morphology of mature adipocytes. Mfn 1/2 mRNA and the expression of Drp 1 mRNA in mature adipocytes were detected by real-time fluorescence quantitative PCR. , Mitochondrial fission gene (Fis) 1 mRNA expression levels. Results Oil red O staining induced differentiation of adipocytes on day 8 and found that more than 85% of adipocytes had differentiated into mature adipocytes with large and round cells. The cytoplasm contained a large number of lipid droplets which were dyed bright red with oil red O , No significant difference in the number of lipid droplets was found between the two groups of cells. Transmission electron microscopy showed that the morphology of mitochondria in the two groups of cells was normal, mitochondria had no obvious swelling and shrinkage, and the mitochondrial crest was clearly visible. There was no significant difference in the number of mitochondria between the two groups. The expression level of Mfn2 mRNA in mature adipocytes with LYRM1 gene silencing was significantly higher than that in mature adipocytes in control group. The expression of Mfn1 mRNA, Drp1 mRNA and Fis1 mRNA in mature adipocytes silenced by LYRM1 gene was not significantly different from that of mature adipocytes in control group. Conclusion Silencing of LYRM1 can induce Mfn2 mRNA expression in mature adipocytes, which has no significant effect on mitochondrial morphology and structure, suggesting that LYRM1 gene silencing may partially affect mitochondrial morphology-related genes in mature adipocytes.