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目的研究肝脏脂肪酸氧化关键分子SIRT3编码基因变异与非酒精性脂肪性肝病(nonalcoholic fatty liver disease,NAFLD)发生的关系,并在小鼠体内研究人源SIRT3保护性等位型基因过表达对脂肪肝相关表型的影响。方法建立非酒精性脂肪肝的人群研究队列,提取基因组DNA对编码区基因变异rs11246020进行基因分型,并与疾病发病与否做关联分析;建立小鼠NAFLD模型,在小鼠肝脏过表达人源SIRT3基因,8周后检测小鼠肝脏相关生化指标及肝脏HE染色、油红O染色情况,并提取RNA和蛋白采用Real-time PCR和Western blot检测Pparα、Acox1、cpt1a、cpt1b和SIRT3的转录或表达水平。结果与对照组相比,SIRT3基因编码区错意突变rs11246020的GA+AA基因型在NAFLD组分布较低,而GG基因型分布较高,差异具有统计学意义(P<0.05);在小鼠肝脏过表达人源SIRT3的A等位型基因后,肝脏脂肪蓄积降低,肝细胞空泡化程度降低,肝脏和血清游离脂肪酸以及肝脏甘油三酯、总胆固醇水平显著降低(P<0.05),肝脏脂肪酸代谢相关基因Pparα和Acox1基因转录水平降低(P<0.05)。同时,小鼠体质量上升被有效控制(P<0.05)。结论 SIRT3基因变异rs11246020与NAFLD风险相关,小鼠体内过表达SIRT3可显著改善脂肪酸代谢和肝脏脂肪蓄积。
Objective To study the relationship between the gene mutation of SIRT3, a key fatty acid oxidation molecule, and the occurrence of nonalcoholic fatty liver disease (NAFLD) in mice and to study the effect of overexpression of protective SIRT3 alleles on fatty liver Effect of related phenotypes. Methods A cohort of non-alcoholic fatty liver patients was established. Genomic DNA was extracted to genotype the gene mutation rs11246020 in the coding region and correlated with the onset of disease. The mouse NAFLD model was established to overexpress the human SIRT3 gene was detected after 8 weeks. Liver biochemical markers and liver HE staining and oil red O staining were detected in 8 weeks and RNA and protein were extracted. Real-time PCR and Western blot were used to detect the transcription of Pparα, Acox1, cpt1a, cpt1b and SIRT3 The expression level. Results Compared with the control group, the GA + AA genotype of rs11246020 in the missense mutation region of SIRT3 gene was lower in NAFLD group and higher in GG genotype. The difference was statistically significant (P <0.05) Liver overexpression of human SIRT3 A allele decreased liver fat accumulation, hepatocytes vacuolization decreased, liver and serum free fatty acids and liver triglyceride, total cholesterol levels were significantly lower (P <0.05), the liver The transcriptional level of Pparα and Acox1 in fatty acid metabolism decreased (P <0.05). At the same time, the increase of body weight in mice was effectively controlled (P <0.05). Conclusion SIRT3 gene mutation rs11246020 is associated with the risk of NAFLD. Overexpression of SIRT3 in mice can significantly improve fatty acid metabolism and liver fat accumulation.