Epigenetic regulation of microRNA-370 by interleukin-6 in malignant human cholangiocytes

Interleukin-6 (IL-6) is overexpressed and contributes to tumor cell growth in cholangiocarcinoma. Enforced IL-6 production can alter the expression of specific microRNAs (miRNAs) involved in tumor growth, and moreover can modulate expression of methylation-dependent genes. Thus, we assessed the methylation-dependent regulation of miRNA expression in human malignant cholangiocytes stably transfected to overexpress IL-6. The expression of the methyltransferases DNA methyltransferase enzyme-1 and HASJ4442 was increased by IL-6 overexpression, but was decreased by the methylation inhibitor 5-aza-2'-deoxycytidine (5-aza-CdR). Expression profiling identified seven miRNAs that were significantly downregulated by IL-6 overexpression (<0.4-fold) and upregulated (>2-fold) by 5-aza-CdR. One of these, miR-370, is embedded in a CpG island. Although 5-aza-CdR increased miR-370 expression by 2.1-fold in malignant cells, the expression in nonmalignant cells was unchanged. The oncogene mitogen-activated protein kinase kinase kinase 8 (MAP3K8) was identified as a target of miR-370, and its expression was decreased by 5-aza-CdR in cholangiocarcinoma cells. Overexpression of IL-6 reduced miR-370 expression and reinstated MAP3K8 expression in vitro as well as in tumor cell xenografts in vivo. Thus, IL-6 may contribute to tumor growth by modulation of expression of selected miRNAs, such as miR-370. These studies define a mechanism by which inflammation-associated cytokines can epigenetically modulate gene expression and directly contribute to tumor biology.