Role of SUMOylation in Alcohol‐induced Liver Fibrosis

Abstract

Background & AimAlcohol‐induced liver fibrosis is characterized by excessive deposition of extracellular matrix (ECM) components, especially collagen in response to chronic hepatocellular damage. If alcohol consumption is continued, fibrosis may progress to cirrhosis which is a major cause of morbidity and mortality. Reactive oxygen species (ROS) generated in hepatocytes by alcohol metabolism plays an important role in producing liver damage and initiating hepatic fibrogenesis. ROS disrupts lipids, proteins and DNA inducing necrosis and apoptosis of hepatocytes and amplifies the inflammatory response via Kupffer cells that directly induces hepatic stellate cells (HSCs) activation, resulting in the initiation of fibrosis. SUMOylation is a post‐translational modification that plays an important role in a wide range of cellular processes. SUMOylation is oxidative stress inducible and involves the covalent attachment of a member of the SUMO (small ubiquitin‐like modifier) to the target protein. We previously demonstrated that the level of Ubiquitin‐conjugation enzyme 9 (Ubc9), a sole E2‐conjugating enzyme essential for SUMOylation, is induced in the livers of intragastric ethanol‐infusion (EI) treated mice and lipopolysaccharide‐induced inflammation promotes Ubc9 phosphorylation and its activity in Kupffer cells. Our aim was to examine whether the dysregulated SUMOylation could regulate the ethanol‐induced liver fibrosis and elucidate the molecular mechanism(s).MethodsStudies were done using in vivo binge ethanol‐fed mice, primary mouse and human HSCs and LX‐2 cells. Oxidative stress markers were measured using commercial kits.ResultsWe found that ethanol treatment for 48 hours (25mM) in LX‐2 and HSCs cells increased ROS production, Ubc9, α‐SMA and COL I mRNA levels, well known as activated HSCs markers, by 1.5‐fold (H2O2), 1.4‐fold (Ubc9), 1.3‐fold (α‐SMA) and 1.5‐fold (COL I), respectively. RNAi‐mediated knockdown of Ubc9 to 75% of control blocked ethanol‐induced ROS production and the increase in α‐SMA and COL I mRNA levels. Interestingly, we found that silencing SUMO‐1 in activated LX‐2 apoptosis was induced by 50%, while quiescent LX‐2 cells in matrigel were not affected. This finding confirmed that sumoylation is playing a key role on stellate cells activation. In addition, isolated mouse stellate cells from binge ethanol‐fed mice overexpressed high level of Ubc9 mRNA and protein levels compared with control mice.ConclusionThis data is strongly suggesting that ethanol is inducing HSCs activation in human and mouse liver involving the oxidative stress‐induced sumoylation machinery. Identifying the molecular mechanisms will require knowing which proteins are affected and what the functional outcomes of a change in sumoylation status is on those proteins, which will pave the way for future exploring.Support or Funding InformationSupported by NIH grant 5 K01 AA022372‐04, maria lauda tomasi