Cardiomyocyte protection by GATA-4 gene engineered mesenchymal stem cells is partially mediated by translocation of miR-221 in microvesicles

• Published in: PloS one Vol. 8; no. 8; p. e73304
• Main Authors: Yu, Bin, Gong, Min, Wang, Yigang, Millard, Ronald W, Pasha, Zeeshan, Yang, Yueting, Ashraf, Muhammad, Xu, Meifeng
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 28-08-2013; Public Library of Science (PLoS)
• Subjects: Angiogenesis; Animals; Apoptosis; Apoptosis Regulatory Proteins - genetics; Apoptosis Regulatory Proteins - metabolism; Biology; Bone marrow; Cardiomyocytes; Caspase; Caspase-3; CD63 antigen; CD9 antigen; Cell Hypoxia - genetics; Cell proliferation; Cloning; Coculture Techniques; Conditioning; Electron microscopy; GATA-4 gene; GATA4 Transcription Factor - genetics; GATA4 Transcription Factor - metabolism; Heart; Heart Ventricles - cytology; Heart Ventricles - metabolism; Hsp70 protein; Hypoxia; Ischemia; L-Lactate dehydrogenase; Laboratory animals; Lactate dehydrogenase; Lactic acid; Male; Medicine; Mesenchymal stem cells; Mesenchymal Stromal Cells - cytology; Mesenchymal Stromal Cells - metabolism; Mesenchyme; MicroRNAs; MicroRNAs - biosynthesis; MicroRNAs - genetics; miRNA; Myocytes, Cardiac - cytology; Myocytes, Cardiac - metabolism; Neonates; p53 Protein; Pathology; Penicillin; Rats; Rats, Sprague-Dawley; Rodents; Stem cell transplantation; Stem cells; Studies; Transfection; Translocation; Transplants & implants; Tumor Suppressor Protein p53 - genetics; Tumor Suppressor Protein p53 - metabolism; Ventricle; Viruses; United States > US; Ohio
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0073304

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).
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.
Our results demonstrate that cardioprotection by MSC(GATA-4) may be regulated in part by a transfer of anti-apoptotic miRs contained within MVs.