title: MCL-1 is modulated in Crohn's disease fibrosis by miR-29b via IL-6 and IL-8.
abstract: The miR-29 family is involved in fibrosis in multiple organs, including the intestine where miR-29b facilitates TGF-β-mediated up-regulation of collagen in mucosal fibroblasts from Crohn's disease (CD) patients. Myeloid cell leukemia-1 (MCL-1), a member of the B-cell CLL/Lymphoma 2 (BCL-2) apoptosis family, is involved in liver fibrosis and is targeted by miR-29b via its 3'-UTR in cultured cell lines. We investigate the role of MCL-1 and miR-29b in primary intestinal fibroblasts and tissue from stricturing CD patients. Transfection of CD intestinal fibroblasts with pre-miR-29b resulted in a significant increase in the mRNA expression of MCL-1 isoforms [MCL-1Long (L)/Extra Short (ES) and MCL-1Short (S)], although MCL-1S was expressed at significantly lower levels. Western blotting predominantly detected the anti-apoptotic MCL-1L isoform, and immunofluorescence showed that staining was localised in discrete nuclear foci. Transfection with pre-miR-29b or anti-miR-29b resulted in a significant increase or decrease, respectively, in MCL-1L foci. CD fibroblasts treated with IL-6 and IL-8, inflammatory cytokines upstream of MCL-1, increased the total mass of MCL-1L-positive foci. Furthermore, transfection of intestinal fibroblasts with pre-miR-29b resulted in an increase in mRNA and protein levels of IL-6 and IL-8. Finally, immunohistochemistry showed reduced MCL-1 protein expression in fibrotic CD samples compared to non-stricturing controls. Together, our findings suggest that induction of MCL-1 by IL-6/IL-8 may surmount any direct down-regulation by miR-29b via its 3'-UTR. We propose that an anti-fibrotic miR-29b/IL-6 IL-8/MCL-1L axis may influence intestinal fibrosis in CD. In the future, therapeutic modulation of this pathway might contribute to the management of fibrosis in CD.
authors.affiliation: Centre for Genomics and Child Health and National Centre for Bowel Research, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, 4 Newark St, Whitechapel, E1 2AT, London, UK.
authors.forename: Anke
authors.initials: A
authors.lastname: Nijhuis
authors.affiliation: Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, 4 Newark St, Whitechapel, E1 2AT, London, UK.
authors.forename: Renata
authors.initials: R
authors.lastname: Curciarello
authors.affiliation: Centre for Genomics and Child Health and National Centre for Bowel Research, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, 4 Newark St, Whitechapel, E1 2AT, London, UK.
authors.forename: Shameer
authors.initials: S
authors.lastname: Mehta
authors.affiliation: Department of Histopathology, The Royal London Hospital, London, UK.
authors.forename: Roger
authors.initials: R
authors.lastname: Feakins
authors.affiliation: Centre for Cell Biology and Cutaneous Research, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London, UK.
authors.forename: Cleo L
authors.initials: CL
authors.lastname: Bishop
authors.affiliation: Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, 4 Newark St, Whitechapel, E1 2AT, London, UK. james.lindsay@bartshealth.nhs.uk.
authors.forename: James O
authors.initials: JO
authors.lastname: Lindsay
authors.affiliation: Centre for Genomics and Child Health and National Centre for Bowel Research, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, 4 Newark St, Whitechapel, E1 2AT, London, UK. a.silver@qmul.ac.uk.
authors.forename: Andrew
authors.initials: A
authors.lastname: Silver
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D003424:Crohn Disease / Q000235:genetics* / Q000473:pathology*; D005347:Fibroblasts / Q000378:metabolism; D005355:Fibrosis; D006801:Humans; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism*; D016209:Interleukin-8 / Q000235:genetics / Q000378:metabolism*; D007422:Intestines / Q000473:pathology; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D064549:Myeloid Cell Leukemia Sequence 1 Protein / Q000235:genetics* / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D015854:Up-Regulation / Q000235:genetics
chemical_list: D020413:3' Untranslated Regions; D015850:Interleukin-6; D016209:Interleukin-8; C532045:MIRN29a microRNA, human; D035683:MicroRNAs; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D020033:Protein Isoforms; D012333:RNA, Messenger
keywords: Crohn’s disease; Fibrosis; MCL-1; miR-29b; microRNA