PNPLA3 I148M Variant Activates Hepatic Stellate Cells via AMIGO2 Upregulation Using iPSC-Derived Model

A variant in the patatin-like phospholipase domain-containing protein 3 (PNPLA3) was reported to be related to metabolic-associated fatty liver disease. However, the mechanism by which this variant leads to liver fibrosis has not been unveiled yet. Using induced pluripotent stem cell (iPSC)-derived hepatic stellate cells (iHSC) containing a single nucleotide polymorphism (SNP) within PNPLA3, this study sought to clarify the mechanism through which this PNPLA3 SNP induces liver fibrosis.

Two types of iPSC (PNPLA3rs73840989(C):Wild, PNPLA3rs73840989(G):Variant) were differentiated into iHSC. iHSC were activated by TGF-β1.

TGF-β1 stimulation resulted in significantly higher secretion of the liver fibrosis markers αSMA and COL1A1 in Variant-iHSCs than in Wild-iHSCs. Variant-iHSC secreted significantly more PDGF and TGF-β and had significantly higher cell proliferative and migration ability. RNA sequencing showed that gene expression related to collagen-containing extracellular matrix and collagen metabolic processes was elevated in Variant-iHSC. We focused on AMIGO2 expression because Variant-iHSC expressed significantly more AMIGO2 than Wild-iHSC. AMIGO2 knockdown using siRNA in Variant-iHSC significantly reduced migration and proliferation ability. Additionally, significant changes were observed in Vimentin and E-cadherin, along with decreases in EMT activity and protein expression of αSMA and COL1A1. Staining for AMIGO2 in cirrhotic and normal liver samples showed that AMIGO2 protein expression was observed in PNPLA3-variant samples.

PNPLA3 SNP-induced liver fibrosis involves EMT of HSC via AMIGO2 expression.