Identification and Modulation of a PPARG-regulated Anti-fibrotic MicroRNA Network in Liver Fibrosis and Hepatocellular Carcinoma

Abstract

The molecular processes underlying chronic liver disease and its progression from fibrosis to cirrhosis and hepatocellular carcinoma (HCC) have been extensively studied in the past, but reliable treatment options remain limited. An important feature of many diseases is the dysregulation of microRNAs (miRNAs). While the functions of miRNAs during hepatic fibrosis and HCC have been described for various individual miRNAs, few studies focus on miRNAs as integral parts of signaling networks. Therefore, this study aims to identify a signaling network of functionally connected miRNAs and target genes that contributes to fibrosis and HCC. For this, the SRF VP16iHep mouse model of fibrotic HCC was analyzed by global miRNA and transcriptome profiling in combination with bioinformatic tools. The thus identified anti fibrotic miRNA network (AF miRNA network) is comprised of 8 miRNAs and 54 extracellular matrix (ECM ) related target genes that influence the fibrotic tumor microenvironment. The AF miRNA network is not only dysregulated in murine and human HCC but also in murine liver fibrosis independent of HCC. Mechanistically, it was shown that the AF miRNA network is dysregulated in activated hepatic stellate cells (HSCs) during fibrogenesis, and that the AF miRNAs are collectively and directly regulated by the transcription factor peroxisome proliferator activated receptor gamma (PPARG). This study further shows that the AF miRNA network is not only conserved between human and mouse, but also to the rat. This expands the available model organisms in which to study the AF miRNA network with implications for human patients. Expression patterns of the AF miRNA network were then compared in several rat and mouse models of liver fibrosis, revealing that AF miRNA dysregulation is highly context specific and varies considerably between the analyzed fibrosis models. Additionally, it was investigated whether modulation of the PPARG regulated AF miRNA network inhibits liver fibrosis progression. Indeed, the PPARG agonist pioglitazone, while ineffective in vivo, enhanced AF miRNA expression in vitro in immortalized HSC cell lines. Since the thus increased AF miRNA expression inhibited the expression of their fibrotic target genes, it can be concluded that PPARG mediated AF miRNA modulation is a possible approach to reduce the fibrotic activity of HSCs during fibrogenesis. Overall, this study demonstrates how functionally connected miRNAs collectively affect complex biological processes, in this case the microenvironment of fibrosis and HCC. The characterization of the AF miRNA network in different liver fibrosis models as well as the investigation of how the AF miRNA network can be modulated in vitro and in vivo present important steps towards understanding the relevance of the AF miRNAs as potential therapeutic targets or predictors of disease outcome.