Cytochrome Cyp2e1 is critical for joint effects of alcohol and lipids on hepatocellular steatosis and inflammation
Alcoholic steatohepatitis (ASH) and nonalcoholic steatohepatitis (NASH) are the most frequent conditions leading to elevated liver enzymes and liver cirrhosis, respectively, in the Western world. However, despite strong epidemiological evidence for combined effects on the progression of liver injury, the mutual interaction of the pathophysiological mechanisms is incompletely understood.
The aim of this study was to establish an in vitro model for joint effects of alcohol and lipids on hepatic steatosis and inflammation. Herein, we particularly wanted to assess the role of CYP2E1, which activates several hepatotoxins and contributes to alcoholic liver damage
Methods and Results: Initially, we established the dose range in which neither alcohol nor incubation with the free fatty acid (FFA) oleate affected viability or mitochondrial activity in primary human hepatocytes (PHH) and HepG2 hepatoma cells. Subsequently, we assessed the combined effect of alcohol (1 – 2%) and oleate (0.2 mM) on hepatocellular lipid accumulation in PHH. Under these conditions, alcohol significantly enhanced oleate induced cellular triglyceride content and FFA levels, while alcohol alone had only a minimal effect on hepatocellular lipid content. Analysis of heme oxygenase-1 (HMOX-1) expression and malondialdehyde levels revealed that the combination of alcohol and oleate caused significantly higher oxidative stress and lipid peroxidation than either of the two substances alone. The CYP2E1 inhibitor chlormethiazole and the antioxidant N-acetylcysteine blunted these combined effects of alcohol and oleate in PHH. In contrast to HepG2 C34 cells which do not express CYP2E1, HepG2 E47 cells which express CYP2E1 exhibited similar joint effects of alcohol and oleate as observed in PHH.
Conclusion: In summary, our new model allows the investigation of isolated or joint effects of alcohol and FFA on hepatocellular lipid metabolisms and inflammatory signaling. Our present findings indicate that CYP2E1 and CYP2E1-derived reactive oxygen species, respectively, play an important role in mediating synergistic effects of alcohol and lipids on hepatic steatosis and inflammation.