The Potentiality of Human Umbilical Cord Isolated Mesenchymal Stem/Stromal Cells for Cardiomyocyte Generation (original) (raw)
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Journal of Tissue Engineering and Regenerative Medicine, 2008
and therapeutics, due to their ability to undergo proliferation and differentiation. Complications associated with bone marrow-derived MSCs has prompted researchers to explore alternative sources of MSCs. The human umbilical cord is one such source; it is easily available and its collection is non-invasive. The sources of MSCs are non-controversial and thus they are not subjected to ethical constraints, as in the case of embryonic stem cells. MSCs are multipotent stem cells and has the ability to differentiate into various cell types of the mesodermal lineage. The aim of this study was to establish a reproducible method for the isolation of MSCs from human umbilical cord, as the few methods published till date gave inconsistent results and had a mixed population of contaminating endothelial cells. In our isolation strategy, we isolated a pure population of MSCs from Wharton's jelly of the human umbilical cord, which is very rich in collagen, and we used a high concentration of collagenase enzyme in the isolation of MSCs. Extensive phenotypic characterization analysis of these cells, using flow cytometry and antibody staining methods, have shown that we were able to isolate a pure population of the mesenchymal lineage cells that is devoid of haematopoietic and endothelial cell contaminants. When these MSCs were subjected to cardiomyocyte differentiation, we observed a change in the morphological characteristics, which was accompanied by the formation of myotube structures and spontaneous beating after 21 days.
2000
IW, Heyndrickx GR. Pretreatment of adult bone marrow mesenchymal stem cells with cardiomyogenic growth factors and repair of the chronically infarcted myocardium. The in vivo cardiac differentiation and functional effects of unmodified adult bone marrow mesenchymal stem cells (MSCs) after myocardial infarction (MI) is controversial. We postulated that ex vivo pretreatment of autologous MSCs using cardiomyogenic growth factors will lead to cardiomyogenic specification and will result in superior biological and functional effects on cardiac regeneration of chronically infarcted myocardium. We used a chronic dog MI model generated by ligation of the coronary artery (n ϭ 30). Autologous dog bone marrow MSCs were isolated, culture expanded, and specified into a cardiac lineage by adding growth factors, including basic FGF, IGF-1, and bone morphogenetic protein-2. Dogs underwent cell injection Ͼ8 wk after the infarction and were randomized into two groups. Group A dogs (n ϭ 20) received MSCs specified with growth factors (147 Ϯ 96 ϫ 10 6 ), and group B (n ϭ 10) received unmodified MSCs (168 Ϯ 24 ϫ 10 6 ). After the growth factor treatment, MSCs stained positive for the early muscle and cardiac markers desmin, antimyocyte enhancer factor-2, and Nkx2-5. In group A dogs, prespecified MSCs colocalized with troponin I and cardiac myosin. At 12 wk, group A dogs showed a significantly larger increase in regional wall thickening of the infarcted territory (from 22 Ϯ 8 to 32 Ϯ 6% in group A; P Ͻ 0.05 vs. baseline and group B, and from 19 Ϯ 7 to 21 Ϯ 7% in group B, respectively) and a decrease in the wall motion score index (from 1.60 Ϯ 0.05 to 1.35 Ϯ 0.03 in group A; P Ͻ 0.05 vs. baseline and group B, and from 1.58 Ϯ 0.07 vs. 1.56 Ϯ 0.08 in group B, respectively). The biological ex vivo cardiomyogenic specification of adult MSCs before their transplantation is feasible and appears to improve their in vivo cardiac differentiation as well as the functional recovery in a dog model of the chronically infarcted myocardium. cardiac repair; myogenesis; chronic myocardial infarction; heart failure SEVERAL CELL TYPES HAVE BEEN utilized in the regeneration of damaged heart tissue (2, 4, 9, 11, 13, 16 -18, 23-25). After transplantation to the damaged heart, skeletal myoblasts dif-Address for reprint requests and other correspondence: J. Bartunek,
Stem Cells, 2007
We tested the cardiomyogenic potential of the human umbilical cord blood-derived mesenchymal stem cells (UCBM-SCs). Both the number and function of stem cells may be depressed in senile patients with severe coronary risk factors. Therefore, stem cells obtained from such patients may not function well. For this reason, UCBMSCs are potentially a new cell source for stem cell-based therapy, since such cells can be obtained from younger populations and are being routinely utilized for clinical patients. The human UCBM-SCs (5 ؋ 10 3 per cm 2 ) were cocultured with fetal murine cardiomyocytes ([CM] 1 ؋ 10 5 per cm 2 ). On day 5 of cocultivation, approximately half of the green fluorescent protein (GFP)-labeled UCBMSCs contracted rhythmically and synchronously, suggesting the presence of electrical communication between the UCBMSCs. The fractional shortening of the contracted UCBMSCs was 6.5% ؎ 0.7% (n ؍ 20). The
Cardiomyocyte differentiation of perinatally‑derived mesenchymal stem cells
Molecular medicine reports, 2013
Coronary heart disease is major cause of mortality worldwide and several risk factors have been shown to play a role in its pathogenesis, including smoking, obesity, hypertension and hypercholesterolemia. A number of therapeutic methods have been developed to improve the quality of patients' lives, including stem cell therapy using mesenchymal stem cells (MSCs). Perinatal sources, including the placenta (PL) and umbilical cord (UC), are rich sources of MSCs and have been identified as a potential source of cells for therapeutic use. Their role in cardiogenic differentiation is also of contemporary medical interest. The present study demonstrated the induced differentiation of MSCs obtained from the UC, PL and Wharton's jelly (WJ) into cardiomyocytes, using 10 µM 5‑azacytidine. The characteristics of the MSCs from each source were studied and their morphology was compared. An immunofluorescence analysis for the cardiac‑specific markers, GATA4 and Troponin T (TnT), was perform...
Vox Sanguinis, 2008
Background and Objectives Mesenchymal stem/progenitor cells (MSCs) are multipotent progenitors that differentiate into such lineages as bone, fat, cartilage and stromal cells that support haemopoiesis. Bone marrow MSCs can also contribute to cardiac repair, although the mechanism for this is unclear. Here, we examine the potential of MSCs from different sources to generate cardiomyocytes in vitro , as a means for predicting their therapeutic potential after myocardial infarction. Materials and Methods Mesenchymal stem/progenitor cells were isolated from the perivascular tissue and Wharton's jelly of the umbilical cord and from cord blood. Their immunophenotype and differentiation potential to generate osteoblasts, chondrocytes, adipocytes and cardiomyoxcytes in vitro was compared with those of bone marrow MSCs. Results Mesenchymal stem/progenitor cells isolated from umbilical cord and cord blood were phenotypically similar to bone marrow MSCs, the exception being in the expression of CD106, which was absent on umbilical cord MSCs, and CD146 that was highly expressed in cord blood MSCs. They have variable abilities to give rise to osteoblasts, chondrocytes and adipocytes, with bone marrow MSCs being the most robust. While a small proportion (~0•07%) of bone marrow MSCs could generate cardiomyocyte-like cells in vitro, those from umbilical cord and cord blood did not express cardiac markers either spontaneously or after treatment with 5-azacytidine. Conclusion Although MSCs may be useful for such clinical applications as bone or cartilage repair, the results presented here indicate that such cells do not generate cardiomyocytes frequently enough for cardiac repair. Their efficacy in heart repair is likely to be due to paracrine mechanisms.
Cardiac Differentiation of HumanWhartos Jelly Stem Cells – Experimental Comparison of Protocols
Cardiomyoplasty represents a promising approach for the repair of the injured heart, but is hampered by the availability of appropriate cells. Mesenchymal stem cells derived from the human umbilical cord tissue (UCMSC) can be obtained in large amounts without medical intervention, exhibit self renewal and immunological naivity as well as multipotency. In the present study, different published protocols of cardiac differentiation designed for different stem cell types were compared to differentiate UCMSC into cardiomyocyte-like cells (cUCMSC). Cardiac differentiation of UCMSC was driven by cell treatment with 5-azacytidine, oxytocin as well as by forming of "embryoid bodies". The morphological and immunocytochemical analysis of cUCMSC with an extensive panel of cardiac markers showed that oxytocin is a more potent inducer of cardiac differentiation than 5-azacytidine and the forming of "embryoid bodies". cUCMSC reveal a cardiomyocyte-like structure and the expression of cardiomyocyte associated proteins. The easy accessibility and the ability of UCMSC to differentiate into cells with characteristics of cardiomyocytes render UCMSC an attractive candidate for cell based therapies and cardiac tissue engineering.
Differentiation, 2010
Bone marrow mesenchymal stromal cells (BM-MSCs) with regenerative potential have been identified in heart. Whether these cells become new cardiac lineage cells by phenomena of transdifferentiation or fusion is also being investigated. Although, these mechanisms give cardiomyocytes, it has to be considered that MSCs transplantation could carry out ossification and calcification processes. An alternative might be the use of myocytes; however, the problem is the arrythmia. For those reasons, is that we investigated how to obtain cardiomyocyte-like cells from human MSCs (hMSCs). The aim of the present work was to evaluate a nuclear reprogramming of the hMSCs by a neonatal rat cardiomyocytes extract (EX) using Streptolysin O (SLO) treatment. hMSCs treated with 57.5 ng/ml SLO presented balllike, stick-like and myotube-like morphology. In the absence of cardiomyogenic stimuli, hMSCs expressed markers of cardiac phenotype-like sarcomeric a-actinin, connexin-43 and GATA-4. However, when hMSCs were treated with SLO +EX or 10 mM of 5-azacytidine (5-AZA), the expression of these markers were significantly increased and furthermore, expressed SERCA-2, cardiac Troponin I, b-MyHC, desmin, MLC-2a and MLC-2v thus showing the phenotype of mature cardiomyocytes. PCR analysis showed that cardiomyocyte-related genes, such as b1-adrenergic receptor (b1-AR), MLC-2a and cardiac Troponin T, were expressed after SLO + EX treatment like with 5-AZA. We concluded that the extract of neonatal rat cardiomyocytes could promote a nuclear modification of hMSCs to cardiomyogenic-like cells differentiation. Since the 5-AZA treatment appears to be genotoxic and taking into account the obtained results, the nuclear reprogramming by cell extract may be an approach leading to the identification of soluble factors that drives the reprogramming.
International Journal of Stem Cells
Background and Objectives: Most studies in cardiac regeneration have explored bone marrow mesenchymal stem cells (BM-MSC) with variable therapeutic effects. Amniotic fluid MSC (AF-MSC) having extended self-renewal and multipotent properties may be superior to bone marrow MSC (BM-MSC). However, a comparison of their cardiomyogenic potency has not been studied yet. Methods: The 5-azacytidine (5-aza) treated AF-MSC and BM-MSC were evaluated for the expression of GATA-4, Nkx2.5 and ISL-1 transcripts and proteins by quantitative RT-PCR and Western blotting, respectively as well as for the expression of cardiomyogenic differentiation markers cardiac troponin-T (cTNT), beta myosin heavy chain (βMHC) and alpha sarcomeric actinin (ASA) by immunocytochemistry. Results: The AF-MSC as compared to BM-MSC had significantly higher expression of GATA-4 (183.06±29.85 vs. 9.80±0.05; p＜0.01), Nkx2.5 (8.3±1.4 vs. 1.82±0.32; p＜0.05), and ISL-1 (39.59±4.05 vs. 4.36±0.39; p＜0.01) genes as well as GATA-4 (2.01±0.5 vs. 0.6±0.1; p＜0.05), NKx2.5 (1.9±0.14 vs. 0.8±0.2; p＜0.01) and ISL-1 (1.7±0.3 vs. 0.9±0.1; p＜0.05) proteins. The AF-MSC also had significantly elevated expression of cTNT (5.0×10 4 ±0.6×10 4 vs. 3.5 ×10 4 ±0.8×10 4 ; p＜0.01), β-MHC (15.7×10 4 ±0.9×10 4 vs. 8.2×10 4 ±0.6×10 4 ; p＜0.01) and ASA (18.6×10 4 ±4.9×10 4 vs. 13.1×10 4 ±3.0×10 4 ; p＜0.05) than BM-MSC. Conclusions: Our data suggest that AF-MSC have greater cardiomyogenic potency than BM-MSC, and thus may be a better source of MSC for therapeutic applications in cardiac regenerative medicine.
Cytotechnology, 2014
Human umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) are a promising stem cell source with the potential to modulate the immune system as well as the capacity to differentiate into osteoblasts, chondrocytes, and adipocytes. In previous publications, UCB-MSCs have been successfully differentiated into cardiomyocytes. This study aimed to improve the efficacy of differentiation of UCB-MSCs into cardiomyocytes by combining 5-azacytidine (Aza) with mouse fetal heart extract (HE) in the induction medium. UCB-MSCs were isolated from umbilical cord blood according to a published protocol. Murine fetal hearts were used to produce fetal HE using a rapid freeze-thaw procedure. MSCs at the 3rd to 5th passage were differentiated into cardiomyocytes in two kinds of induction medium: complete culture medium plus Aza (Aza group) and complete culture medium plus Aza and fetal HE (Aza + HE group). The results showed that the cells in both kinds of induction medium exhibited the phenot...
Basic Research in Cardiology, 2010
The ability of human umbilical cord blood-derived mesenchymal stem cells (UCBMSCs) to transdifferentiate towards cardiomyocytes remains unclear. The aim of this study was to direct UCBMSCs to the cardiac lineage by exposure to: (1) 5-azacytidine (AZ) or dimethyl sulfoxide (DMSO); (2) a combination of growth factors involved in early cardiomyogenesis (BMP-2 + bFGF + IGF-1); (3) the Wnt signaling activators lithium chloride (LiCl) and phorbol-12-myristate-13-acetate (PMA); and (4) direct contact with neonatal rat cardiomyocytes. Expression of cardiomyocyte-specific proteins and β-catenin were assessed by quantitative RT-PCR, immunofluorescence and Western blot. Cocultures of human UCBMSCs with neonatal rat cardiomyocytes were also analyzed for the presence of calcium oscillations and changes in electrical potential using Fura Red and di-4-ANEPPS confocal imaging, respectively. Induction of cardiac-specific proteins was not detected in 5-AZ- or DMSO-treated cells. Following DMSO addition, β-catenin cytoplasmic expression increased, but did not translocate into cell nuclei to promote cardiac gene activation. Likewise, neither co-stimulation with BMP-2 + bFGF + IGF-1, nor exposure to LiCl and PMA resulted in the acquisition of a cardiac phenotype by UCBMSCs. Direct contact with neonatal rat cardiomyocytes promoted neither the expression of cardiomyocyte-specific proteins, nor the presence of calcium rhythmic oscillations and potential-dependent fluorescence emission in UCBMSCs. The cardiomyogenic stimuli investigated in this study failed to transdifferentiate human UCBMSCs. Alternative strategies or regulatory factors and signaling pathways may be better suited to recruit UCBMSCs into cardiac cell lineage.