Involvement of miRNA-29a in epigenetic regulation of transforming growth factor-β-induced epithelial-mesenchymal transition in hepatocellular carcinoma

摘要

Aim: Epithelial-mesenchymal transition (EMT) is a crucial process during cancer invasion and metastasis, which is accompanied by the suppressed expression of E-cadherin initiated by stimuli such as transforming growth factor (TGF)-β. Recent studies have shown that the epigenetic regulation of E-cadherin could be an alternate mechanism of EMT induction in hepatocellular carcinoma (HCC). miRNA-29a (miR-29a) is involved in the epigenetic regulation of genes by targeting DNA methyltransferases (DNMT), which methylate CpG islands to suppress the transcription of genes. We studied the involvement of miR-29a in TGF-β-induced EMT in HCC cells. Methods: We treated human HCC cell lines with TGF-β to induce EMT. To investigate DNA methylation in EMT, cells were treated with a methylation inhibitor, 5-Aza-2′-deoxycytidine (5-Aza) and methylation status of CpG islands in the E-cadherin promoter was examined using methylation-specific PCR. Precursor miR-29a (pre-miR-29a) was electroporated to force the expression of miR-29a in HCC cells in order to study the role of miR-29a in EMT. Results: TGF-β transformed HCC cells into a spindle-shaped morphology accompanied by a decrease of E-cadherin with the induction of methylation of its promoter. Pretreatment of the cells with 5-Aza blocked this suppression of E-cadherin, indicating the involvement of DNA methylation. TGF-β increased DNMT3B and DNMT1 and decreased miR-29a expression. The forced expression of miR-29a abrogated the suppression of E-cadherin induced by TGF-β. Conclusion: miR-29a could regulate TGF-β-induced EMT by affecting DNA methylation via the suppression of DNMT. These observations reveal the epigenetic regulation of genes by miRNA as a unique mechanism of EMT in HCC.