Cardiomyocytes can be generated from marrow stromal cells in vitro
Shinji Makino, … , Akihiro Umezawa, Satoshi Ogawa
Shinji Makino, … , Akihiro Umezawa, Satoshi Ogawa
Published March 1, 1999
Citation Information: J Clin Invest. 1999;103(5):697-705. https://doi.org/10.1172/JCI5298.
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Article

Cardiomyocytes can be generated from marrow stromal cells in vitro

Abstract

We have isolated a cardiomyogenic cell line (CMG) from murine bone marrow stromal cells. Stromal cells were immortalized, treated with 5-azacytidine, and spontaneously beating cells were repeatedly screened. The cells showed a fibroblast-like morphology, but the morphology changed after 5-azacytidine treatment in ∼30% of the cells; they connected with adjoining cells after one week, formed myotube-like structures, began spontaneously beating after two weeks, and beat synchronously after three weeks. They expressed atrial natriuretic peptide and brain natriuretic peptide and were stained with anti-myosin, anti-desmin, and anti-actinin antibodies. Electron microscopy revealed a cardiomyocyte-like ultrastructure, including typical sarcomeres, a centrally positioned nucleus, and atrial granules. These cells had several types of action potentials, such as sinus node–like and ventricular cell–like action potentials. All cells had a long action potential duration or plateau, a relatively shallow resting membrane potential, and a pacemaker-like late diastolic slow depolarization. Analysis of the isoform of contractile protein genes, such as myosin heavy chain, myosin light chain, and α-actin, indicated that their muscle phenotype was similar to that of fetal ventricular cardiomyocytes. These cells expressed Nkx2.5/Csx, GATA4, TEF-1, and MEF-2C mRNA before 5-azacytidine treatment and expressed MEF-2A and MEF-2D after treatment. This new cell line provides a powerful model for the study of cardiomyocyte differentiation.

Authors

Shinji Makino, Keiichi Fukuda, Shunichirou Miyoshi, Fusako Konishi, Hiroaki Kodama, Jing Pan, Motoaki Sano, Toshiyuki Takahashi, Shingo Hori, Hitoshi Abe, Jun-ichi Hata, Akihiro Umezawa, Satoshi Ogawa

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Figure 6

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Representative tracing of the action potential of CMG myotubes. Action p...
Representative tracing of the action potential of CMG myotubes. Action potential recordings using a conventional microelectrode were obtained from the spontaneously beating cells at day 28 after 5-azacytidine treatment. We categorized these action potentials into two groups: a sinus node–like action potential (a) or a ventricular cardiomyocyte–like action potential (b). These action potentials have a relatively shallow resting membrane potential with late diastolic slow depolarization: a pacemaker-like potential. The ventricular cardiomyocyte–like action potential had peak notch-plateau characteristics, with an initial tall overshoot in phase 0, a repolarizing notch in phase 1, and a second depolarizing plateau in phase 2, whereas the sinus node–like action potential had none of these features. Beating cycle length, action potential amplitude, action potential duration, and most diastolic potential are given in Table 2.