Ethanol-Induced Expression of ET-1 and ET-BR in Liver Sinusoidal Endothelial Cells and Human Endothelial Cells Involves Hypoxia-Inducible Factor-1α and MicroRNA-199

Chronic alcohol consumption leads to inflammation and cirrhosis of the liver. In this study, we observed that liver sinusoidal endothelial cells (LSEC) derived from ethanol-fed rats showed several fold increases in the mRNA expression of endothelin-1 (ET-1), hypoxia-inducible factor-1alpha (HIF-1alpha), and inflammatory cytochemokines compared with control rat LSEC. We also observed the same results in acute ethanol-treated LSEC from control rats and human dermal microvascular endothelial cells. Ethanol-mediated ET-1 expression involved NADPH oxidase and HIF-1alpha activation. Furthermore, ethanol increased the expression of the ET-1 cognate receptor ET-BR in Kupffer cells and THP-1 monocytic cells, which also involved HIF-1alpha activation. Promoter analysis and chromatin immunoprecipitation showed that hypoxia response element sites in the proximal promoter of ET-1 and ET-BR were required for the binding of HIF-1alpha to up-regulate their expression. We showed that microRNAs, miR-199 among several microRNAs, attenuated HIF-1alpha and ET-1 expression, while anti-miR-199 reversed the effects, suggesting that ethanol-induced miR-199 down-regulation may contribute to augmented HIF-1alpha and ET-1 expression. Our studies, for the first time to our knowledge, show that ethanol-mediated ET-1 and ET-BR expression involve HIF-1alpha, independent of hypoxia. Additionally, ethanol-induced ET-1 expression in rat LSEC is regulated by miR-199, while in human endothelial cells, ET-1 expression is regulated by miR-199 and miR-155, indicating that these microRNAs may function as novel negative regulators to control ET-1 transcription and, thus, homeostatic levels of ET-1 to maintain microcirculatory tone.