Chronic ethanol feeding alters miRNA expression dynamics during liver regeneration.

BACKGROUND: Adaptation to chronic ethanol (EtOH) treatment of rats results in a changed functional state of the liver and greatly inhibits its regenerative ability, which may contribute to the progression of alcoholic liver disease. METHODS: In this study, we investigated the effect of chronic EtOH intake on hepatic microRNA (miRNA) expression in male Sprague-Dawley rats during the initial 24 hours of liver regeneration following 70% partial hepatectomy (PHx) using miRNA microarrays. miRNA expression during adaptation to EtOH was investigated using RT-qPCR. Nuclear factor kappa B (NFκB) binding at target miRNA promoters was investigated with chromatin immunoprecipitation. RESULTS: Unsupervised clustering of miRNA expression profiles suggested that miRNA expression was more affected by chronic EtOH feeding than by the acute challenge of liver regeneration after PHx. Several miRNAs that were significantly altered by chronic EtOH feeding, including miR-34a, miR-103, miR-107, and miR-122 have been reported to play a role in regulating hepatic metabolism and the onset of these miRNA changes occurred gradually during the time course of EtOH feeding. Chronic EtOH feeding also altered the dynamic miRNA profile during liver regeneration. Promoter analysis predicted a role for NFκB in the immediate-early miRNA response to PHx. NFκB binding at target miRNA promoters in the chronic EtOH-fed group was significantly altered and these changes directly correlated with the observed expression dynamics of the target miRNA. CONCLUSIONS: Chronic EtOH consumption alters the hepatic miRNA expression profile such that the response of the metabolism-associated miRNAs occurs during long-term adaptation to EtOH rather than as an acute transient response to EtOH metabolism. Additionally, the dynamic miRNA program during liver regeneration in response to PHx is altered in the chronically EtOH-fed liver and these differences reflect, in part, differences in miRNA expression between the EtOH-adapted and control livers at the baseline state prior to PHx

A novel comparative pattern analysis approach identifies chronic alcohol mediated dysregulation of transcriptomic dynamics during liver regeneration.

BACKGROUND: Liver regeneration is inhibited by chronic ethanol consumption and this impaired repair response may contribute to the risk for alcoholic liver disease. We developed and applied a novel data analysis approach to assess the effect of chronic ethanol intake in the mechanisms responsible for liver regeneration. We performed a time series transcriptomic profiling study of the regeneration response after 2/3(rd) partial hepatectomy (PHx) in ethanol-fed and isocaloric control rats. RESULTS: We developed a novel data analysis approach focusing on comparative pattern counts (COMPACT) to exhaustively identify the dominant and subtle differential expression patterns. Approximately 6500 genes were differentially regulated in Ethanol or Control groups within 24 h after PHx. Adaptation to chronic ethanol intake significantly altered the immediate early gene expression patterns and nearly completely abrogated the cell cycle induction in hepatocytes post PHx. The patterns highlighted by COMPACT analysis contained several non-parenchymal cell specific markers indicating their aberrant transcriptional response as a novel mechanism through which chronic ethanol intake deregulates the integrated liver tissue response. CONCLUSIONS: Our novel comparative pattern analysis revealed new insights into ethanol-mediated molecular changes in non-parenchymal liver cells as a possible contribution to the defective liver regeneration phenotype. The results revealed for the first time an ethanol-induced shift of hepatic stellate cells from a pro-regenerative phenotype to that of an anti-regenerative state after PHx. Our results can form the basis for novel interventions targeting the non-parenchymal cells in normalizing the dysfunctional repair response process in alcoholic liver disease. Our approach is illustrated online at http://compact.jefferson.edu

Adaptation to ethanol dysregulates hepatic miRNA expression and alters the dynamic response of miRNAs during liver regeneration

MicroRNAs (miRNAs) are short, non-coding RNAs which post-transcriptionally regulate their target messenger RNAs by translational inhibition and/or mRNA destabilization and degradation. miRNAs have been implicated in the regulation of cell cycle, metabolism, development, cancer, and responses to toxins. Adaptation to chronic ethanol treatment of rats results in a changed functional state of the liver and greatly inhibits its regenerative ability, which may contribute to the progression of alcoholic liver disease. Chronic ethanol consumption alters the hepatic miRNA expression profile such that, for instance, the response of several metabolism-associated miRNAs occurs during long-term adaptation to ethanol. Additionally, the dynamic miRNA program during liver regeneration in response to partial hepatectomy is altered in the chronically ethanol-fed rat and these differences reflect, in part, differences in miRNA expression between the ethanol-adapted and control livers at the baseline state prior to partial hepatectomy. miR-21 is pro-proliferative in many contexts, most notably cancer. We find miR-21 expression to increase during liver regeneration following partial hepatectomy. However, the induction of miR-21 is more robust in the livers from regeneration-inhibited ethanol-fed animals compared to the controls. Global gene expression analyses of predicted miR-21 targets suggest that miR-21 induction has a greater effect in the remnant of the ethanol-fed rat than in that of the control rat. Additionally, our analysis suggests that miR-21 may have a widespread regulatory role and a range of effects that are not predominantly associated with the regulation of cell proliferation during inhibited regeneration in the liver of the ethanol-fed rat. Inhibition of miR-21 in vivo does not affect DNA synthesis or expression of cell cycle markers through S phase following partial hepatectomy. However, miR-21 inhibition may affect later stages of the cell cycle. Our data also suggest that few predicted targets of miR-21 are affected by miR-21 inhibition, indicating that the actions of miR-21 are context dependent