Abstract 9485: Translocation of miRNA from Mesenchymal Stem Cells Protects Cardiomyocytes against Ischemic Injury in vitro
microRNAs (miR) play an important role in cell survival. In this study, we investigated whether mesenchymal stem cells (MSC) mediated cardioprotection was associated with the regulation of miR as well as their targeted proteins.
Methods: MSC were harvested from rat bone marrow. Cardiomyocytes (CM) were obtained from neonatal rat ventricles. In vitro cardioprotection by MSC was assayed by lactate dehydrogenase (LDH) release, Annexin-V staining, and DNA fragmentation following cells exposure to hypoxia for 48 hours. The expression of miR in MSC and CM was assessed by quantitative real-time PCR. miR targeted proteins were evaluated using a luciferase reporter assay and western blotting. To further investigate the role of miR, pre-miR precursor molecules were introduced into MSC using Lipofectamine 2000.
Results: miR-221 was expressed 12-fold more in MSC than in CM (p<0.05). The p53-upregulated modulator of apoptosis (PUMA), one of miR-221 targeted proteins, was lower in MSC than in CM (Fig. A). When co-cultured with CM in a dual-chamber system, MSC markedly reduced LDH release and the number of annoxin-V positive CM, and increased CM survival in hypoxic culture. Transfection of miR-221 into MSC significantly increased MSC mediated cardioprotection compared to MSC transfected with negative control miR. In CM co-cultures with MSC, miR-221 transferred from MSC to CM was confirmed by using fluorescence in situ hybridization (FISH) and immunostaining. Moreover, less PUMA protein was found in CM co-cultured with MSC than in CM cultured alone (Fig. B).
Conclusions: Cardioprotective effects of MSC are putatively associated with intercellular translocation of specific miR and regulation of their targeted proteins such as PUMA in CM.
- © 2012 by American Heart Association, Inc.