Open AccessCardiomyocyte Protection by GATA-4 Gene Engineered Mesenchymal Stem Cells Is Partially Mediated by Translocation of miR-221 in Microvesicles
Author(s) -
Bin Yu,
Min Gong,
Yigang Wang,
Ronald W. Millard,
Zeeshan Pasha,
Yueting Yang,
Muhammad Ashraf,
Meifeng Xu
Publication year - 2013
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0073304
Subject(s) - cardioprotection , mesenchymal stem cell , microbiology and biotechnology , microvesicles , biology , stem cell , microrna , transfection , cell culture , medicine , genetics , gene , ischemia , cardiology
microRNAs (miRs), a novel class of small non-coding RNAs, are involved in cell proliferation, differentiation, development, and death. In this study, we found that miR-221 translocation by microvesicles (MVs) plays an important role in cardioprotection mediated by GATA-4 overexpressed mesenchymal stem cells (MSC). Methods and Results Adult rat bone marrow MSC and neonatal rat ventricle cardiomyocytes (CM) were harvested as primary cultures. MSC were transduced with GATA-4 (MSC GATA-4 ) using the murine stem cell virus (pMSCV) retroviral expression system. Empty vector transfection was used as a control (MSC Null ). The expression of miRs was assessed by real-time PCR and localized using in situ hybridization (ISH). MVs collected from MSC cultures were characterized by expression of CD9, CD63, and HSP70, and photographed with electron microscopy. Cardioprotection during hypoxia afforded by conditioned medium (CdM) from MSC cultures was evaluated by lactate dehydrogenase (LDH) release, MTS uptake by CM, and caspase 3/7 activity. Expression of miR-221/222 was significantly higher in MSC than in CM and miR-221 was upregulated in MSC GATA-4 . MSC overexpression of miR-221 significantly enhanced cardioprotection by reducing the expression of p53 upregulated modulator of apoptosis (PUMA). Moreover, expression of PUMA was significantly decreased in CM co-cultured with MSC. MVs derived from MSC expressed high levels of miR-221, and were internalized quickly by CM as documented in images obtained from a Time-Lapse Imaging System. Conclusions Our results demonstrate that cardioprotection by MSC GATA-4 may be regulated in part by a transfer of anti-apoptotic miRs contained within MVs.