Affiliation:
1. From the Leibniz Research Laboratories for Biotechnology and Artificial Organs (I.G., J.B., U.B., I.S.-R., A.H., U.M.) and Department of Functional and Applied Anatomy (A.S.), Hannover Medical School, Hannover; and Laboratory for Molecular Gastroenterology and Hepatology, Clinic for Internal Medicine, University of Kiel, Kiel (M.K.), Germany.
Abstract
Background—
Recent studies have suggested the differentiation of human endothelial progenitor cells (huEPCs) isolated from peripheral blood into cardiomyocytes. This study investigates whether, when cocultured, neonatal rat cardiomyocytes (NRCMs) can induce transdifferentiation of huEPCs into cardiomyocytes.
Methods and Results—
Coculture experiments with 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine (DiI)–labeled huEPCs and NRCMs have been performed. Cocultures have been analyzed by means of flow cytometry, 3D confocal laser microscopy, species-specific reverse transcriptase–polymerase chain reaction for the expression of human cardiac marker genes, and electron microscopy. Although fluorescence-activated cell sorting (FACS) analysis and conventional wide-field fluorescence microscopy suggested the existence of DiI
pos
cardiomyocytes in cocultures, no convincing evidence of cardiac differentiation of huEPCs has been obtained. Apparently, DiI
pos
cardiomyocytes were identified as necrotic NRCMs or NRCM-derived vesicles with high levels of autofluorescence or, alternatively, as NRCMs lying on top of or below labeled huEPCs or huEPC fragments. Accordingly, no expression of human Nkx2.5, GATA-4, or cardiac troponin I was detected.
Conclusions—
No convincing evidence of transdifferentiation of huEPCs into cardiomyocytes was obtained. Although we cannot exclude that recent contrary data may be due to slightly different culture protocols, our study has revealed that recently applied standard analysis tools including FACS and wide-field fluorescence microscopy are not sufficient to demonstrate transdifferentiation in coculture settings and can lead to misinterpretation of the data obtained solely with these methods.
Publisher
Ovid Technologies (Wolters Kluwer Health)
Subject
Physiology (medical),Cardiology and Cardiovascular Medicine
Cited by
87 articles.
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