Transcriptional regulation of miR-224 upregulated in human HCCs by NFκB inflammatory pathways

Giuseppe Montalto, Rossana De Iaco, Paolo D'Onorio De Meo, Melchiorre Cervello, Massimo Levrero, Paolo D'Onorio De Meo, Francesca Guerrieri, Valeria Schinzari, Cecilia Scisciani, Giovanni Raimondo, Natalia Pediconi, Stefania Vossio, Teresa Pollicino

Research output: Contribution to journalArticlepeer-review

114 Citations (Scopus)


Background & Aims: miR-224 is up-regulated in human HCCs as compared to both paired peri-tumoral cirrhotic tissues and cirrhotic livers without HCC. Here, we have cloned the miR-224 regulatory region and characterized its transcriptional regulation by the NFκB-dependent inflammatory pathways. Methods: Mature miRNA expression was evaluated by a 2 step stem-loop real-time RT-PCR. The recruitment of polymerase II and transcription factors on the pre-miR-224 promoter has been assessed by ChIPSeq and ChIP. Results: We found miR-224 levels strongly up-regulated in both peri-tumoral cirrhotic livers and HCC samples as compared to normal livers. In silico analysis of the putative miR-224 promoter revealed multiple NFκB sites. We showed that LTα and TNFα activate transcription from the miR-224 promoter and of endogenous miR-224 expression in HCC cell lines, whereas the expression of miR-224 target API5 was reduced. Exogenously expressed p65/RelA activates the miR-224 promoter and a dominant negative form of IκBα (IκBSR) represses it. ChIP analysis showed that p65/NFκB is recruited on the miR-224 promoter and that its binding sharply increases after exposure to LPS, TNFα, and LTα. Altogether these findings link the inflammatory signals to NFκB-mediated activation of miR-224 expression. An antago-miR specific for miR-224 blocked LPS and LTα stimulated HCC cells migration and invasion. Conversely, the IKK inhibitor BMS-345541 blocks pre-miR-224-induced cellular migration and invasion. Conclusions: Our results identify p65/NFκB as a direct transcriptional regulator of miR-224 expression and link miR-224 up-regulation with the activation of the LPS, LTα, and TNFα inflammatory pathways and cell migration/invasion in HCC. © 2011 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.
Original languageEnglish
Pages (from-to)855-861
Number of pages7
JournalJournal of Hepatology
Publication statusPublished - 2012

All Science Journal Classification (ASJC) codes

  • Hepatology


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