Genomics of human fatty liver disease reveal mechanistically linked lipid droplet–associated gene regulations in bland steatosis and nonalcoholic steatohepatitis

Abstract

Nonalcoholic fatty liver disease (NAFLD) is a common disorder hallmarked by excessive lipid deposits. Based on our recent research on lipid droplet (LD) formation in hepatocytes, we investigated LD-associated gene regulations in NAFLD of different grades, that is, steatosis vs steatohepatitis by comparing liver biopsies from healthy controls (N=13) and NAFLD patients (N=102). On average, more than 700 differentially expressed genes (DEGs) were identified of which 146 are mechanistically linked to LD formation. We identified 51 LD-associated DEGs frequently regulated in patient samples (range ≥5 to ≤102) with the liver-receptor homolog-1(NR5A2), that is, a key regulator of cholesterol metabolism being commonly repressed among 100 patients examined. With bland steatosis, notable regulations involved hypoxia-inducible lipid droplet-associated-protein and diacylglycerol-O-acyltransferase-2 renowned for their role in LD-growth. Conversely, nonalcoholic steatohepatitis-associated DEGs coded for epidermal growth factor receptor and TLR4 signaling with decreased expression of the GTPase Rab5 and the lipid phosphohydrolase PPAP2B thus highlighting adaptive responses to inflammation, LDL-mediated endocytosis and lipogenesis, respectively. Studies with steatotic primary human hepatocyte cultures demonstrated induction of LD-associated PLIN2, CIDEC, DNAAF1, whereas repressed expression of CPT1A, ANGPTL4, and PKLR informed on burdened mitochondrial metabolism. Equally, repressed expression of the B-lymphocyte chemoattractant CXCL13 and STAT4 as well as induced FGF21 evidenced amelioration of steatosis-related inflammation. In-vitro/in-vivo patient sample comparisons confirmed C-reactive protein, SOCS3, NR5A2, and SOD2 as commonly regulated. Lastly, STRING network analysis highlighted potential "druggable" targets with PLIN2, CIDEC, and hypoxia-inducible lipid droplet-associated-protein being confirmed by immunofluorescence microscopy. In conclusion, steatosis and steatohepatitis specific gene regulations informed on the pathogenesis of NAFLD to broaden the perspective of targeted therapies.