title: Wnt Pathway Stabilizes MeCP2 Protein to Repress PPAR-γ in Activation of Hepatic Stellate Cells.
abstract: PPAR-γ is essential for differentiation of hepatic stellate cells (HSC), and its loss due to epigenetic repression by methyl-CpG binding protein 2 (MeCP2) causes HSC myofibroblastic activation mediated in part via Wnt pathway, the key cellular event in liver fibrosis. Decreased miR-132 was previously proposed to promote MeCP2 protein translation for Ppar-γ repression in activated HSC (aHSC). The present study aimed to test this notion and to better understand the mechanisms of MeCP2 upregulation in aHSC. MeCP2 protein is increased on day 3 to 7 as HSC become activated in primary culture on plastic, but this is accompanied by increased but not reduced miR-132 or miR-212 which is also expected to target MeCP2 due to its similar sequence with miR-132. The levels of these mRNAs are decreased 40~50% in aHSCs isolated from experimental cholestatic liver fibrosis but increased 6-8 fold in aHSC from hepatotoxic liver fibrosis in rats. Suppression of either or both of miR132 and miR212 with specific anti-miRNA oligonucleotides (anti-oligo), does not affect MeCP2 protein levels in aHSCs. The Wnt antagonist FJ9 which inhibits HSC activation, increases miR-132/miR-212, reduces MeCP2 and its enrichment at 5' Ppar-γ promoter, and restores Ppar-γ expression but the anti-oligo do not prevent Ppar-γ upregulation. The pan-NADPH oxidase (NOX) inhibitor diphenyleneiodonium (DPI) also reduces both MeCP2 and stabilized non-(S33/S37/Thr41)-phospho β-catenin and reverts aHSC to quiescent cells but do not affect miR-132/miR-212 levels. Wnt antagonism with FJ9 increases MeCP2 protein degradation in cultured HSC, and FJ9-mediated loss of MeCP2 is rescued by leupeptin but not by proteasome and lysozome inhibitors. In conclusion, canonical Wnt pathway increases MeCP2 protein due to protein stability which in turn represses Ppar-γ and activates HSC.
authors.affiliation: Southern California Research Center for ALPD and Cirrhosis, Department of Pathology, University of Southern California, Los Angeles, California, United States of America.
authors.forename: Soo-Mi
authors.initials: SM
authors.lastname: Kweon
authors.affiliation: Southern California Research Center for ALPD and Cirrhosis, Department of Pathology, University of Southern California, Los Angeles, California, United States of America.
authors.forename: Feng
authors.initials: F
authors.lastname: Chi
authors.affiliation: Southern California Research Center for ALPD and Cirrhosis, Department of Pathology, University of Southern California, Los Angeles, California, United States of America.
authors.forename: Reiichi
authors.initials: R
authors.lastname: Higashiyama
authors.affiliation: Southern California Research Center for ALPD and Cirrhosis, Department of Pathology, University of Southern California, Los Angeles, California, United States of America.
authors.forename: Keane
authors.initials: K
authors.lastname: Lai
authors.affiliation: Southern California Research Center for ALPD and Cirrhosis, Department of Pathology, University of Southern California, Los Angeles, California, United States of America.|Department of Veterans Affairs Greater Los Angeles Healthcare System, Los Angeles, California, United States of America.
authors.forename: Hidekazu
authors.initials: H
authors.lastname: Tsukamoto
mesh_terms: D000818:Animals; D054338:Cell Transdifferentiation / Q000235:genetics; D002478:Cells, Cultured; D063185:Epigenetic Repression; D055166:Hepatic Stellate Cells / Q000502:physiology*; D008103:Liver Cirrhosis / Q000235:genetics / Q000378:metabolism; D008297:Male; D051783:Methyl-CpG-Binding Protein 2 / Q000378:metabolism*; D035683:MicroRNAs / Q000502:physiology; D047495:PPAR gamma / Q000235:genetics* / Q000378:metabolism; D055550:Protein Stability; D051381:Rats; D017208:Rats, Wistar; D060449:Wnt Signaling Pathway / Q000235:genetics / Q000502:physiology*
chemical_list: C548821:MIRN132 microRNA, rat; C552427:MIRN212 microRNA, rat; C497394:Mecp2 protein, rat; D051783:Methyl-CpG-Binding Protein 2; D035683:MicroRNAs; D047495:PPAR gamma