In vitro cardiomyocyte differentiation of umbilical cord blood cells: crucial role for c-kit(+) cells.
OBJECTIVE
Although bone marrow c-kit(+) progenitor cells support myocardial regeneration, the cardiomyocyte differentiation potential of umbilical cord blood (UCB) c-kit(+) cells is unknown.


METHODS
UCB mononuclear cells (MNCs) and c-kit(+) cells purified by use of immunomagnetic beads were used. Cardiomyocyte differentiation was induced with (i) α-minimum essential medium (MEM) with cyclosporine A, (ii) α-MEM with bone morphogenic protein 4 (BMP-4) and transforming growth factor-β (TGF-β) or (iii) MEM with dexamethasone. The expression of cardiac markers (GATA4, GATA6, β-myosin heavy chain, α-sarcomeric actin and cardiac Troponin T) was investigated, and whole-cell current and voltage-clamp recordings were performed.


RESULTS
Although c-kit(+) cells revealed an immature gene profile, with high expression of CD34, CD133, aldehyde dehydrogenase-A1 and c-myc RNAs, purified c-kit(+) cells did not succeed in differentiating into cardiomyocyte-like cells in culture. In contrast, MNCs (either in α-MEM plus cyclosporine A or in α-MEM plus BMP-4 and TGF-β) produced large, adherent cells expressing several cardiac genes and exhibiting an excitable phenotype. Cardiomyocyte-like cell formation was prevented by removing the c-kit(+) cell fraction from MNCs. Furthermore, after co-culturing carboxyfluorescein diacetate succynimidyl ester (CFSE)-tracked c-kit(+) cells together with c-kit(-) cells, we found that cardiac Troponin T--expressing cells were also CFSE(+).


CONCLUSIONS
We show that UCB contains progenitors endowed with differentiation potential into cardiomyocytes: these cells reside in the c-kit(+) fraction and require the presence of abundant accessory cells to accomplish the differentiation. These preliminary observations provide the basis for consider the storage of autologous UCB in patients with prenatal diagnosis of congenital heart diseases potentially amenable by myocardial regenerative approaches.
Institute of Hematology, Catholic University of the Sacred Heart, Rome, Italy.
Maria Grazia
MG
Iachininoto
Institute of Hematology, Catholic University of the Sacred Heart, Rome, Italy.
Sara
S
Capodimonti
Institute of Human Physiology, Catholic University of the Sacred Heart, Rome, Italy.
Maria Vittoria
MV
Podda
Institute of Hematology, Catholic University of the Sacred Heart, Rome, Italy.
Caterina Giovanna
CG
Valentini
Institute of Hematology, Catholic University of the Sacred Heart, Rome, Italy.
Maria
M
Bianchi
Department of Cardiovascular Sciences, Catholic University of the Sacred Heart, Rome, Italy.
Antonio Maria
AM
Leone
Institute of Hematology, Catholic University of the Sacred Heart, Rome, Italy. Electronic address: lteofili@rm.unicatt.it.
Luciana
L
Teofili
Institute of Hematology, Catholic University of the Sacred Heart, Rome, Italy.
Giuseppe
G
Leone
D015415:Biomarkers / Q000032:analysis; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D018920:Coculture Techniques; D005312:Fetal Blood / Q000166:cytology* / Q000187:drug effects; D005786:Gene Expression Regulation; D006801:Humans; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000378:metabolism; D009930:Organic Chemicals / Q000494:pharmacology; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism*; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000378:metabolism / Q000494:pharmacology; D024723:Ventricular Myosins / Q000378:metabolism
D015415:Biomarkers; D009930:Organic Chemicals; D016212:Transforming Growth Factor beta; C420642:alpha minimal essential medium; D019009:Proto-Oncogene Proteins c-kit; D024723:Ventricular Myosins
c-kit; cardiomyocytes; umbilical cord blood