Abstract
ObjectiveThe purpose of this study was to assess the effect of collagen composition on engraftment of progenitor cells within infarcted myocardium.BackgroundWe previously reported that intramyocardial penetration of stem/progenitor cells in epicardial patches was enhanced when collagen was reduced in hearts overexpressing adenylyl cyclase-6 (AC6). In this study we hypothesized an alternative strategy wherein overexpression of microRNA-29b (miR-29b), inhibiting mRNAs that encode cardiac fibroblast proteins involved in fibrosis, would similarly facilitate progenitor cell migration into infarcted rat myocardium.Methods In vitro: A tri-cell patch (Tri-P) consisting of cardiac sodium-calcium exchanger-1 (NCX1) positive iPSC (iPSCNCX1+), endothelial cells (EC), and mouse embryonic fibroblasts (MEF) was created, co-cultured, and seeded on isolated peritoneum. The expression of fibrosis-related genes was analyzed in cardiac fibroblasts (CFb) by qPCR and Western blot. In vivo: Nude rat hearts were administered mimic miRNA-29b (miR-29b), miRNA-29b inhibitor (Anti-29b), or negative mimic (Ctrl) before creation of an ischemically induced regional myocardial infarction (MI). The Tri-P was placed over the infarcted region 7 days later. Angiomyogenesis was analyzed by micro-CT imaging and immunofluorescent staining. Echocardiography was performed weekly.ResultsThe number of green fluorescent protein positive (GFP+) cells, capillary density, and heart function were significantly increased in hearts overexpressing miR-29b as compared with Ctrl and Anti-29b groups. Conversely, down-regulation of miR-29b with anti-29b in vitro and in vivo induced interstitial fibrosis and cardiac remodeling.ConclusionOverexpression of miR-29b significantly reduced scar formation after MI and facilitated iPSCNCX1+ penetration from the cell patch into the infarcted area, resulting in restoration of heart function after MI.
Highlights
In response to myocardial infarction (MI), cardiac remodeling is activated by structural rearrangement and involves cardiomyocyte hypertrophy, cell death, myofibroblast differentiation, collagen types I and III deposition, fibrosis, and scar formation
To analyze the role of miR-29b on cardiac fibrosis, the total mRNA of rat neonatal cardiomyocytes and cardiac fibroblasts (CFb) were isolated respectively, and qPCR revealed that miR-29b was expressed significantly more in CFb than in Neonatal cardiomyocytes (neoCM) (p,0.05, Fig. 1A)
To further study the molecular signaling pathway involved in fibrosis, CFb were transfected with negative mimic (CFbCtrl), miR-29b mimic (CFbmiR-29b), or miR-29b inhibitor (CFbAnti-29b)
Summary
In response to MI, cardiac remodeling is activated by structural rearrangement and involves cardiomyocyte hypertrophy, cell death, myofibroblast differentiation, collagen types I and III deposition, fibrosis, and scar formation. Fibrosis and activation of cardiac fibroblasts, the major non-myocyte component (90%) of heart tissue [1], are primary events in ventricular remodeling, rather than a secondary response to cardiac myocyte injury [2]. We previously reported that intramyocardial penetration of stem/progenitor cells in epicardial patches was enhanced when collagen was reduced in hearts overexpressing adenylyl cyclase-6 (AC6). In this study we hypothesized an alternative strategy wherein overexpression of microRNA-29b (miR-29b), inhibiting mRNAs that encode cardiac fibroblast proteins involved in fibrosis, would facilitate progenitor cell migration into infarcted rat myocardium
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