1030-LBP: Hepatitis C virus induced changes in microRNA-107 and microRNA-449a modulate the inflammatory chemokine CCL2 by targeting IL6 receptor complex in patients with hepatic fibrosis

Abstract

Aim Hepatitis C virus (HCV) mediated liver diseases are one of the major health issues in United States and worldwide. This study is to identify the role of HCV induced changes in microRNAs (miRNA) that controls various cell surface receptors and gene regulatory complexes involved in patients with HCV mediated inflammation and fibrosis. Methods Computational sequence analyses and reporter assays using the CCL2 promoter region were carried out to analyze transcription factors. Gene expression analyses were done using q-PCR. Gene overexpression and knockdowns in human hepatocytes were carried out using cDNA plasmids and siRNAs. Chromatin (ChIP) and protein (CoIP) immunoprecipitations were done to establish interactions between transcription factors and DNA. Results We report here that down-regulation of miRNA-107 and miRNA-449a following HCV infection in patients modulate expression of the CCL2, an inflammatory chemokine up-regulated in patients with chronic liver diseases, by targeting components of the IL6R complex. Computational analysis for DNA bound transcription factors in the CCL2 promoter identified adjacent binding sites for CCAAT/CEBP α , spleen focus forming virus, proviral integration oncogene (SPI1/PU.1), and STAT3. We demonstrate that CEBP α , PU.1 and STAT3 interacted with each other physically to co-operatively bind to the promoter and activate CCL2 expression. Analysis of IL6R and JAK1 expression in HCV patients showed significant up regulation when there is impaired miRNA-107 and miRNA-449a expression along with up regulation of PU.1 and STAT3 but not CEBP α . miRNA-449a and miRNA-107 target IL6R and JAK1 respectively, inhibit IL6 signaling and impair STAT3 activation. Conclusions Our results demonstrate a novel gene regulatory mechanism wherein HCV induced changes in miRNAs (miRNA-449a and miRNA-107) regulate CCL2 expression by activation of the IL6 mediated signaling cascade in HCV patients which we propose will result in HCV mediated induction of inflammatory responses and fibrosis.