Division of Gastroenterology and Hepatology, University of Michigan Health System, Ann Arbor, MI 48109, USA.
Department of Computational Medicine and Bioinformatics, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
Int J Mol Sci. 2021 Sep 9;22(18):9758. doi: 10.3390/ijms22189758.
Non-alcoholic fatty liver disease (NAFLD) is caused by excess lipid accumulation in hepatocytes. Genome-wide association studies have identified a strong association of NAFLD with non-synonymous E167K amino acid mutation in the transmembrane 6 superfamily member 2 (TM6SF2) protein. The E167K mutation reduces TM6SF2 stability, and its carriers display increased hepatic lipids and lower serum triglycerides. However, the effects of TM6SF2 on hepatic lipid metabolism are not completely understood. We overexpressed wild-type or E167K variant of or knocked down expression in lipid-treated Huh-7 cells and used untargeted lipidomic analysis, RNAseq transcriptome analysis, and fluorescent imaging to determine changes in hepatic lipid metabolism. Both knockdown and E167K overexpression increased hepatic lipid accumulation, while wild-type overexpression decreased acylglyceride levels. We also observed lipid chain remodeling for acylglycerides by knockdown, leading to a relative increase in species with shorter, more saturated side chains. RNA-sequencing revealed differential expression of several lipid metabolizing genes, including genes belonging to AKR1 family and lipases, primarily in cells with knockdown. Taken together, our data show that overexpression of gene or its loss-of-function changes hepatic lipid species composition and expression of lipid metabolizing genes. Additionally, our data further confirms a loss-of-function effect for the E167K variant.
非酒精性脂肪性肝病 (NAFLD) 是由肝细胞内脂质蓄积过多引起的。全基因组关联研究发现，NAFLD 与跨膜 6 超家族成员 2（TM6SF2）蛋白中 E167K 氨基酸的非同义突变有很强的相关性。E167K 突变降低了 TM6SF2 的稳定性，其携带者表现出肝内脂质增加和血清甘油三酯降低。然而，TM6SF2 对肝内脂质代谢的影响尚不完全清楚。我们在脂质处理的 Huh-7 细胞中转染野生型或 E167K 突变型或敲低 表达，并进行非靶向脂质组学分析、RNAseq 转录组分析和荧光成像，以确定肝内脂质代谢的变化。 敲低和 E167K 过表达均增加肝内脂质蓄积，而野生型过表达则降低酰基甘油水平。我们还观察到 敲低导致酰基甘油的脂质链重塑，导致短链、更饱和的侧链相对增加。RNA-seq 揭示了几种脂质代谢基因的差异表达，包括 AKR1 家族和脂肪酶的基因，主要在 敲低的细胞中。综上所述，我们的数据表明， 基因的过表达或功能丧失会改变肝内脂质种类组成和脂质代谢基因的表达。此外，我们的数据进一步证实了 E167K 变体的功能丧失效应。
