Abstract
BACKGROUND: We have shown that type I collagen matrix treatment at 3h post-myocardial infarction (MI) leads to less inflammation and improved cardiac function, but the underlying mechanisms remain to be better characterized. MicroRNAs are emerging as key players in the regulation of the post-MI environment. We previously identified that matrix-treated hearts had down-regulated expression of miR-92a, a miRNA with inflammatory effects that is normally upregulated after MI. The goal of this study was to elucidate a possible role of miR-92a in the anti-inflammatory/pro-wound healing effect of matrix treatment post-MI. METHODS/RESULTS: C57BL/6J mice underwent LAD ligation to induce MI. Hearts were removed at 6h, 1d, 2d, 3d, and 7d post-MI and RNA was extracted from infarct and peri-infarct tissue. PCR analysis revealed that miR-92a was initially upregulated at 6h, gradually decreased from 1d to 3d, and then increased again at 7d. The first peak in miR-92a expression coincides with initiation of the inflammatory phase, so we assessed the macrophage response in matrix-treated mice. Three hours post-MI, a separate group of mice received intramyocardial injections of PBS or collagen matrix. Tissues were harvested at 2 days, and miR-92a was confirmed by PCR to be down-regulated in the matrixvs. PBS-treated hearts (p<0.0005). Matrix-treated hearts also had a reduced number of M1 inflammatory macrophages at 2 days (p<0.05). Bone marrow-derived macrophages (BMDMs) were used in vitro to further evaluate matrix regulation of miR-92a-mediated pathways in macrophages. Integrins a5 (ITGa5) and aV (ITGaV), involved in cell-matrix interactions, as well as inflammatory regulators MKK4, CD69, S1PR1 and RAP1b were identified as putative miR-92a targets. BMDMs cultured on the collagen matrix had significantly reduced expression of miR-92a compared to those cultured on TCPS (p<0.001), while the miR-92a target genes integrins a5 and aV were significantly up-regulated. Forced over-expression of miR-92a in matrixcultured BMDMs caused a decrease in the expression of MKK4 (p1⁄40.01), CD69 (p<0.001), ITGa5 (p1⁄40.001), ITGaV (p1⁄40.02), and S1PR1 (p1⁄40.01). Finally, the matrix also enhanced the differentiation of M2 wound healing macrophages as indicated by increased expression of IL-10. CONCLUSION: We report that the beneficial effects of matrix treatment post-MI may be mediated, at least in part, through its ability to regulate miR-92a and pro-wound healing mechanisms in macrophages. These results present the matrix as a novel non-pharmacological approach to locally regulate miRNAs in vivo for reducing inflammation and protecting the myocardium post-MI.
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