P0454 : An additional heme oxygenase-1 knockout increases maturation of dendritic cells and liver inflammation in Mdr2 knockout mice

Background and Aims: DNA methylation (5mC) is an epigenetic mark that is an established regulator of gene transcription with an important role in liver fibrosis. Currently, there is only very basic knowledge available as to how DNA methylation controls the phenotype of hepatic stellate cells (HSC), the key cell type responsible for onset and progression of liver fibrosis. Evidence has shown that presence of higher levels of DNA methylation around gene promoters correlate with low or no transcriptional activity. However, recently discovered 5-hydroxymethylcytosine (5hmC), generated by oxidation of 5mC through action of TET enzymes, is actually involved in gene activation and its patterns are often altered in human diseases. In this study we investigated the role of 5mC and 5hmC in liver fibrosis. Methods: Levels of 5mC and 5hmC were assessed by slot blot in a range of animal liver fibrosis models and diseased human livers. Expression levels of TET enzymes and DNA methyltransferases (DNMTs) were measured by qRT-PCR and WB. Reduced representation bisulfide sequencing (RRBS) method was used to examine 5mC and 5hmC patterns in quiescent and in vivo activated rat HSC. Results: Expression of TET enzymes and therefore global levels of 5hmC, are both downregulated in activated rat HSC and in diseased human livers. Moreover, DNMTs expression is changed in human and rat samples suggesting changed levels of 5mC. Using RRBS, we examined the exact genomic positions of changed methylation patterns in quiescent and in vivo activated rat HSC. Conclusions: We found significant differences in TET and DNMT protein expression between quiescent and activated rat HSC as well as in human samples, which in turn cause alteration in the HSC DNA methylome and hydroxymethylome. Changes in DNA methylation during HSC activation may bring new insights into the molecular events underpinning fibrogenesis and may provide biomarkers for disease progression as well as potential new drug targets.