Electrophysiological properties of mouse bone marrow c-kit+ cells co-cultured onto neonatal cardiac myocytes.
OBJECTIVE
Controversy about hematopoietic stem cells reprogramming into cardiac myocytes is currently supported by positive and negative findings. In fact, some reports have shown the ability of stem cells from the bone marrow (BM) to differentiate into cardiac myocytes and to contribute to myocardium repair, while others have reported the opposite.


METHODS
C-kit(+) cells from mouse bone marrow were co-cultured onto neonatal cardiac myocytes. Hematopoietic stem cell-derived cells were analyzed by investigating the expression of cardiac markers and ion channels and by single-cell electrophysiological recordings.


RESULTS
Groups of undifferentiated c-kit(+) cells displayed only outward currents. Co-cultured c-kit(+) stem cells on neonatal cardiac myocytes expressed cardiac markers and Na(+) and Ca(2+) voltage-gated ion channels. However, Na(+) and Ca(2+) currents were not detected by electrophysiological patch-clamp recordings even if caffeine and cyclopiazonic acid treatment showed the presence of intracellular calcium stores. This suggests that these channels, although expressed, were not functional and thus do not allow the coupling between excitation and contraction that is typical of cardiac myocytes. Nevertheless, co-cultured cells had a more hyperpolarized resting membrane potential and, at least in a subset of cells, displayed voltage-gated inward rectifier currents and outward currents. Co-cultured c-kit(+)-derived cells were not connected to surrounding cardiac myocytes through gap junctions. To induce a more pronounced differentiation, co-cultured cells were treated with BMP-4 and TGF-beta, two factors that were shown to trigger a cardiac myocyte differentiation pathway in embryonic stem (ES) cells. Even under these conditions, c-kit(+) cells did not differentiate into functionally active cardiac myocytes. However, TGF-beta/BMP-4-treated cells were hyperpolarized and showed and increased inward rectifier current density.


CONCLUSIONS
Our study shows that mouse BM hematopoietic stem cells exhibit a limited plasticity to transdifferentiate into cardiac myocytes in culture.
Lagostena
Laura
L
Laboratorio di Biologia Vascolare e Terapia Genica, Centro Cardiologico Monzino, Istituto di Ricovero e Cura a Carattere Scientifico, Milan, Italy.
Avitabile
Daniele
D
De Falco
Elena
E
Orlandi
Alessia
A
Grassi
Francesca
F
Iachininoto
Maria Grazia
MG
Ragone
Gianluca
G
Fucile
Sergio
S
Pompilio
Giulio
G
Eusebi
Fabrizio
F
Pesce
Maurizio
M
Capogrossi
Maurizio C
MC
D000818:Animals; D000831:Animals, Newborn; D015415:Biomarkers / Q000032:analysis; D001854:Bone Marrow Cells / Q000502:physiology*; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000378:metabolism; D002454:Cell Differentiation; D018920:Coculture Techniques; D004594:Electrophysiology; D005455:Fluorescent Antibody Technique; D006412:Hematopoietic Stem Cells / Q000378:metabolism; D006801:Humans; D007473:Ion Channels / Q000378:metabolism*; D008564:Membrane Potentials; D051379:Mice; D018613:Microscopy, Confocal; D032383:Myocytes, Cardiac / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015222:Sodium Channels / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000494:pharmacology
C525809:BMP4 protein, human; D015415:Biomarkers; C525810:Bmp4 protein, mouse; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D020746:Calcium Channels, L-Type; D007473:Ion Channels; D015222:Sodium Channels; D016212:Transforming Growth Factor beta; D019009:Proto-Oncogene Proteins c-kit