Effect of different genotypes of PNPLA3 I148M on hepatocyte proliferation

Abstract

Objective: 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. Methods: 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. Results: 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)%, P<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)%, P=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, P=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, P=0.965). The PGC1α mRNA expression, total PI3K, PThr-308AKT, PSer2448-mammalian target of rapamycin (PSer2448-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. Conclusion: PNPLA3 148M/M cell proliferation was stronger than PNPLA3 148I/I.