Identification of a metabolic, transcriptomic and molecular signature of PNPLA3-mediated acceleration of steatohepatitis

BACKGROUND & AIMS: The mechanisms by which the I148M mutant variant of the patatin-like phospholipase domain-containing 3 (PNPLA3(I148M)) drives development of nonalcoholic steatohepatitis (NASH) is not known. The aim of this study was to obtain insights on mechanisms underlying PNPLA3(I148M) induced acceleration of NASH. APPROACH & RESULTS: Hepatocyte-specific overexpression of empty vector (Luc), human wild-type PNPLA3 (PNPLA3(WT)), or PNPLA3(I148M) was achieved using adeno-associated virus (AAV)-8 in DIAMOND mice followed by chow diet or high fat Western diet with ad lib administration of sugar in drinking water (WDSW) for 8 weeks. Under WDSW, PNPLA3(I148M) overexpression accelerated steatohepatitis with increased steatosis, inflammation ballooning and fibrosis (p< 0.001 vs other groups for all). Silencing PNPLA3(I148M) after its initial overexpression abrogated these findings. PNPLA3(I148M) caused 22:6n3 docosahexanoic acid depletion and increased ceramides under WDSW in addition to increasing triglycerides and diglycerides especially enriched with unsaturated fatty acids. It also increased oxidative stress and ER-stress. Increased total ceramides was associated with STAT3 activation with downstream activation of multiple immune-inflammatory pathways at a transcriptomic level by network analyses. Silencing PNPLA3(I148M) reversed STAT3 activation. Conditioned media from HepG2 cells overexpressing PNPLA3(I148M) increased procollagen mRNA expression in LX2 cells; this was abrogated by hepatocyte STAT3 inhibition. CONCLUSIONS: Under WDSW, PNPLA3(I148M) overexpression promotes steatosis and NASH by metabolic reprogramming characterized by increased triglycerides and diglycerides, n3 PUFA depletion and increased ceramides with resultant STAT3 phosphorylation and downstream inflammatory pathway activation driving increased stellate cell fibrogenic activity.