Novel in vitro models of alcohol-mediated hepatic injury

Only a fraction of drinkers develops significant hepatic inflammation, and even less progresses to significant hepatic fibrosis. There is evidence for combined effects of alcohol and nonalcoholic fatty liver disease on the progression of liver injury. Still, the mechanisms of this combined pathophysiological effect are not well understood.The aim of this study was to establish in vitro models to study the direct effects of alcohol on hepatocytes and indirect hepatocyte mediated effects on hepatic stellate cells (HSC).

Methods and Results: Hepatoma cells and primary human hepatocytes (PHH) were incubated with different alcohol concentrations (0.1–1%) for 48h, which did not affect cell viability or mitochondrial activity but led to dose-dependent hepatocellular lipid accumulation as assessed by analysis of triglyceride content and free fatty acid (FFA) levels. Subsequently, human HSC were incubated with conditioned media (CM) from alcohol-stimulated hepatocytes, which significantly induced proliferation, and pro-inflammatory (IL–8, RANTES) and pro-fibrogenic (TGF-beta) gene expression compared to HSC incubated with CM from control PHHs. At higher concentrations (>5%) alcohol dose-dependently induced cytotoxicity and apoptosis in PHHs. However, the toxic alcohol effects in PHHs, in which cellular steatosis had been induced by pre-incubation with FFAs, occurred already at lower concentrations, which also was associated with higher ROS-formation and pro-inflammatory and pro-fibrogenic gene expression.

Conclusion: Our in vitro data indicate the release of soluble factors as a potential mechanism for development and progression of fibrogenesis in alcoholic liver disease, the identification of these factors may lead to novel prognostic markers. Further, we provide a novel model to study the interactive effects of alcoholic- and non-alcoholic fatty liver injury, which may be used to identify and test novel targets for anti-fibrotic therapies.