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Nur77 Inhibits Isoproterenol‐Induced Cardiac Fibrosis through Upregulation of Relaxin‐3
Author(s) -
Guo ZhiFu,
Zhang Guanxin,
You Xiaohua,
Yi Bing,
Cheng Fang,
Sun Jianxin
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.765.11
Subject(s) - relaxin , nerve growth factor ib , cardiac fibrosis , fibrosis , downregulation and upregulation , nuclear receptor , receptor , biology , endocrinology , medicine , microbiology and biotechnology , transcription factor , gene , genetics
Objective The orphan nuclear receptor Nur77 plays critical roles in cardiovascular diseases and its expression is markedly induced in the heart after excessive β‐adrenergic receptor (β‐AR) activation and ischemia/reperfusion injury. However, the functional significance of Nur77 in cardiac fibrosis remains unclear. The objective of this study is to determine whether Nur77 is involved in the development of cardiac fibrosis, and if so, to determine the molecular mechanism (s) involved. Methods and Results By using a mouse model of isoproterenol (ISO)‐induced cardiac fibrosis, we show that the ISO‐induced cardiac fibrosis was markedly exacerbated in Nur77 knockout mice, hence highlighting a critical importance of Nur77 in cardiac remodeling. To determine the molecular mechanism involved, we investigated whether Nur77 affects the cardiac expression of relaxin, which has been shown to be a key anti‐fibrotic molecule in the heart under various pathophysiological circumstances. By using qPCR, we found that both relaxin‐1 and relaxin‐3 mRNA are predominantly expressed in mouse cardiomyocytes, as compared with that in cardiac fibroblasts. Importantly, we demonstrate that overexpressing Nur77 robustly increases the expression of relaxin‐3, but not relaxin‐1 in cardiomyocytes, suggesting the specificity of Nur77 regulation on relaxin‐3 in the heart. Indeed, sequential analysis identified two NGFI‐B response elements (NBRE) (5′‐AAAGGTCA‐3′) in the promoter region of relaxin‐3. Site‐directed mutagenesis and promoter functional analysis suggest that these two NBRE motifs are mainly responsible for the Nur77‐induced relaxin‐3 expression in cardiomyocytes. Furthermore, we show that Nur77 specifically binds to the promoter region of relaxin‐3, as demonstrated by electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assays, under both basal and ISO‐stimulated conditions. Moreover, ISO stimulation markedly increases the expression of relaxin‐3 in wild‐type (WT) mice, but not in Nur77 knockout (KO) mice, further indicating the important role of Nur77 in regulating relaxin‐3 expression in the heart. Continuous infusion of relaxin‐3 through the mini‐osmotic pump implanted subcutaneously for 14 days significantly attenuated cardiac fibrosis of both WT and Nur77 KO mice, as determined by Masson's trichrome (MTC) staining and hydroxyproline assay. Furthermore, pharmacological activation of Nur77 by cytosporone B (CsnB) potently inhibited ISO‐induced cardiac fibrosis in wild‐type mice, but not in Nur77 KO mice. Conclusion Together, these results demonstrate for the first time that Nur77 is a novel regulator for cardiac fibrosis through upregulating of relaxin‐3, and that targeted activation of Nur77 in the heart may represent a potentially novel therapeutic strategy for preventing cardiac fibrosis. Support or Funding Information This research was supported by the US National Institutes of Health (HL103869)