miR-15a/15b Cluster Modulates Survival of Mesenchymal Stem Cells to Improve Its Therapeutic Efficacy of Myocardial Infarction

Background-The poor viability of transplanted mesenchymal stem cells (MSCs) hampers their therapeutic efficacy for ischemic heart disease. MicroRNAs are involved in regulation of MSC survival and function. The present study was designed to investigate the molecular effects of miR-15a/15b on MSC survival, focusing on the role of vascular endothelial growth factor receptor 2. Methods and Results-We first harvested donor luc(Luciferase)-MSCs (5x 10(5)) isolated from the luciferase transgenic mice with FVB background. Luc-MSCs were transfected with miR-15a/15b mimics or inhibitors and cultured under oxygen glucose deprivation condition for 12 hours to mimics the harsh microenvironment in infarcted heart; they were subjected to MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide Thiazolyl Blue Tetrazolium Bromide) assay, bioluminescence imaging, quantitative reverse transcription-polymerase chain reaction, transferase-mediated deoxyuridine triphosphate-digoxigenin nick-end labeling assay, and flow cytometry. Furthermore, the levels of vascular endothelial growth factor receptor 2, protein kinase B, p(Phosphorylate)-protein kinase B, Bcl-2, Bax, and caspase-3 proteins were available by Western blotting assay. In vivo, acute myocardial infarction was induced in 24 mice by coronary ligation, with subsequent receipt of Luc-MSCs, Luc-MSCs+miR-15a/15b inhibitors, or PBS treatment. The therapeutic procedure and treatment effects were tracked and assessed using bioluminescence imaging and echocardiographic measurement. Next, ex vivo imaging and immunohistochemistry were conducted to verify the distribution of MSCs. We demonstrated that miR-15a/15b targeted vascular endothelial growth factor receptor 2 to modulate MSC survival, possibly via phosphatidylinositol 3-kinase/protein kinase B signaling pathway, which was proved by bioluminescence imaging, immunohistochemistry analysis, and echocardiographic measurement. Conclusions-Luc-MSCs could be followed dynamically in vitro and in vivo by bioluminescence imaging, and the role of miR-15a/b could be inferred from the loss of signals from luc-MSCs. This finding may have practical clinical implications in miR-15a/15b- modified MSC transplantation in treating myocardial infarction.