Stiffness-driven Human Hepatocytes Dysfunction via Metabolic-Redox Cross Talk in Liver Fibrogenesis

Liver stiffness (LS) is currently the best clinical predictor of liver fibrosis. However, the molecular mechanisms that account for the stiffness predilection to hepatocytes dysfunction have been underexplored. The overall goal of this study is to introduce a new paradigm that LS is a driver of hepatic dysfunction during fibrosis. Using a multidisciplinary approach, we investigated the role of LS in altering redox-bioenergetic network driving hepatocytes dysfunction. Primary human hepatocytes (PHH) were cultured on an innovative biomimetic platform “BEASTS (Bio-Engineered Adhesive Siloxane substrate with Tunable Stiffness)” that recreates physiologic (2 kPa) and pathologic stiffness (8, 15, 25, 55 kPa). We demonstrated that stiffness impedes urea, albumin production, and expression of drug transporter gene and epithelial cell phenotype markers. NMR metabolomics demonstrated that stiffness regulates metabolic pathways in PHHs including glycolysis, glutamate metabolism, TCA cycle, GSH metabolism, and mitochondrial function. Also, PHHs on fibrotic stiffness inhibits ATP production and maximal respiration and increases glycolysis and glycolytic capacity which parallels metabolic changes observed in fibrosis patients. PHHs cultured on fibrotic stiffness increased ROS; and decreased reduced glutathione levels culminating in apoptosis. Similar metabolic changes were observed in hepatocytes isolated from TAA-induced liver fibrosis in vivo model. These data demonstrates the plausible role of LS in driving hepatocytes dysfunction via redox-metabolic crosstalk during fibrosis.