Yan X M, Yuan S H, Xu X, Ai H Y, Liang H
Department of Endocrinology, the Third Affiliated Hospital, Sun Yat-sen University, Guangdong Provincial Key Laboratory of Diabetology, Guangzhou 510630, China.
Zhonghua Yi Xue Za Zhi. 2018 Sep 11;98(34):2754-2759. doi: 10.3760/cma.j.issn.0376-2491.2018.34.015.
To explore the influence of patatin-like phospholipase domain containing-3 (PNPLA3) wild type 148I/I and mutant type 148M/M on HepG2 cell proliferation and the relative mechanisms. HepG2 cell line stably overexpressing PNPLA3 148I/I, 148M/M and negative control (NC) were set up. Cell counting kit-8 (CCK8) assay was used to measure cell viability. Edu assay was used to determine the ability of cell proliferation. Western blot was used to detect the protein levels in the phosphatidylinositol 3-kinases (PI3K) pathway. Enzyme-linked immunosorbent assay (ELISA) was used to detect proliferation-related PNPLA3 metabolites[arachidonic acid (AA) and lysophosphatidic acid (LPA)]. Quantitative real-time PCR was used to detect the expression level of prostaglandin G/H synthase 2 (PTGS2) and proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) associated with PNPLA3. The cell viability of overexpression of PNPLA3 148M/M group was about 1/3 times higher than that of overexpression of PNPLA3 148I/I group, and the difference was statistically significant[(98.02±1.29)% vs (71.51±2.89)%, <0.001]. There was no significant difference between overexpression of PNPLA3 148M/M group and negative control group[(98.02±1.29)% vs (100±2.61)%, =0.181]. The proliferative activity of overexpression of PNPLA3 148M/M group was about 1/3 times higher than that of overexpression of PNPLA3 148I/I group, and the difference was statistically significant(46.46±1.83 vs 35.96±2.65, =0.001). There was no significant difference between overexpression of PNPLA3 148M/M group and negative control group(46.46±1.83 vs 46.64±7.33, =0.965). The PGC1α mRNA expression, total PI3K, PAKT, P-mammalian target of rapamycin (P-mTOR) and PGC1α protein expression levels in the overexpression of PNPLA3 148M/M group were higher than those in the overexpression of PNPLA3 148I/I group, but there were no significant differences in AA and LPA levels, as well as PTGS2 mRNA expression levels. PNPLA3 148M/M cell proliferation was stronger than PNPLA3 148I/I.
探讨含帕他丁样磷脂酶结构域蛋白3（PNPLA3）野生型148I/I和突变型148M/M对HepG2细胞增殖的影响及相关机制。构建稳定过表达PNPLA3 148I/I、148M/M的HepG2细胞系及阴性对照（NC）。采用细胞计数试剂盒-8（CCK8）法检测细胞活力。采用Edu法测定细胞增殖能力。采用蛋白质免疫印迹法检测磷脂酰肌醇3激酶（PI3K）通路中的蛋白水平。采用酶联免疫吸附测定（ELISA）法检测与PNPLA3相关的增殖相关代谢产物[花生四烯酸（AA）和溶血磷脂酸（LPA）]。采用定量实时聚合酶链反应检测与PNPLA3相关的前列腺素G/H合酶2（PTGS2）和增殖激活受体γ共激活因子1α（PGC1α）的表达水平。PNPLA3 148M/M过表达组的细胞活力比PNPLA3 148I/I过表达组高约1/3倍，差异有统计学意义[（98.02±1.29）% vs（71.51±2.89）%，<0.001]。PNPLA3 148M/M过表达组与阴性对照组之间无显著差异[（98.02±1.29）% vs（100±2.61）%，=0.181]。PNPLA3 148M/M过表达组的增殖活性比PNPLA3 148I/I过表达组高约1/3倍，差异有统计学意义（46.46±1.83 vs 35.96±2.65，=0.001）。PNPLA3 148M/M过表达组与阴性对照组之间无显著差异（46.46±1.83 vs 46.64±7.33，=0.965）。PNPLA3 148M/M过表达组的PGC1α mRNA表达、总PI3K、PAKT、磷酸化哺乳动物雷帕霉素靶蛋白（P-mTOR）和PGC1α蛋白表达水平均高于PNPLA3 148I/I过表达组，但AA和LPA水平以及PTGS2 mRNA表达水平无显著差异。PNPLA3 148M/M细胞增殖强于PNPLA3 148I/I。
