Abstract 11639: Cytoglobin: A Novel Epicardial Regulator of Epithelial to Mesenchymal Transition (EMT) Within the Ischemic Heart
Cytoglobin (Cygb), a nuclear stress-responsive hemoprotein, is abundantly expressed in adult epicardial cells (EPDCs). Data from our laboratory indicate Cygb is a repressor of p53 transcriptional activity within myocytes. We hypothesize that Cygb serves an important role in maintaining EPDC multipotency and EMT following ischemia-reperfusion (IR) injury. To test our hypothesis, we generated mice with epicardial-specific deletion of Cygb (Cygb-/-). These mice are viable with normal cardiac function [ECHO: FS(%) 64±3 vs 66±2 p=NS; Cygb-/- vs control (Ctrl), n=6]. Cygb-/- EPDCs have increased transcript levels of pro-fibrotic genes (MMP2, Mucin, Keratins) while the expression of cardiogenic and endothelial markers are down regulated. After IR injury, Cygb-/- mice develop worse maladaptive cardiac remodeling with enhanced fibrosis [%fibrosis: 14.4±2 vs 3.7±0.8 p<0.05; FS(%): 23±8 vs 54±5 p<0.05; Cygb-/- vs Ctrl, n=3-6]. Utilizing EPDCs isolated from IR injured hearts and immortalized epicardial cells, we show that cells lacking Cygb fail to undergo EMT resulting in impaired epicardial cell migration (total migrated cells: 24±2.7x103 vs 37±4x103 p<0.05; Cygb-/- vs Ctrl, n=3). ChIP-on-ChIP data reveal the presence of Cygb on the promoter of several key EMT genes including β-catenin and Stat3. Immunohistochemistry and qRT PCR data demonstrate down regulation of β-catenin and up regulation of Stat3 expression in Cygb-/- EPDCs at baseline and after IR injury. Bioinformatics analyses reveal the presence of two putative p53 binding sites in the β-catenin and Stat3 promoter. A ChIP assay verified the co-occupancy of Cygb and p53 at the identified sites within the β-catenin and Stat3 promoter. qRT PCR data reveal a reciprocal expression pattern between Cygb levels and p53 target genes (p21, Gadd45A, Plk3), indicating a functional Cygb-p53 axis in epicardial cells. A standard luciferase reporter assay indicates p53 inhibits β-catenin transcription. Collectively, our data demonstrate that Cygb modulates epicardial EMT by directly regulating β-catenin and Stat3 expression in a p53 dependent manner. Loss of Cygb impairs EMT and activates a pro-fibrotic gene signature culminating in excessive fibrosis and maladaptive cardiac remodeling after IR injury.
- © 2013 by American Heart Association, Inc.