Fetal heart extract facilitates the differentiation of human umbilical cord blood-derived mesenchymal stem cells into heart muscle precursor cells (original) (raw)
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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...
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,
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.
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.
Stem Cells and Cloning: Advances and Applications, 2020
Background: The new therapeutic strategy of managing cardiac diseases is based on cell therapy; it highly suggests the use of multipotent mesenchymal stem/stromal cells (MSCs). MSCs widely used in researches are known to be isolated from bone marrow. However, this research seeks to use a human umbilical cord (HUC) as an alternative source of MSCs. Since HUC Wharton's jelly (WJ)-isolated MSCs originate as fetal tissue they are highly preferable for their potential advantages over other adult tissues. Methods: The researchers used enzymatic digestion to establish a primary HUC-WJisolated MSC line. Then, flow cytometry was used to characterize MSCs and hematopoietic stem cells (HSCs) markers' expression. In addition, the cardiac differentiation capacity of HUC-WJ-isolated MSCs in vitro was investigated by two protocols. Protocol-1 necessitates the dependence on merely 5-azacytidine (5-Aza), whereas in protocol-2, 5-Aza was supported by basic fibroblast growth factor (BFGF). The comparative study between the two protocols was applied by inspecting the ultrastructure of differentiated cells, measuring RT-PCR mRNA cardiac markers and the quantitative detection of cardiac proteins. Results: HUC-WJ isolated MSCs were expressed by CD90 +ve , CD105 +ve , CD106 +ve , CD45 −ve , and CD146 −ve. Remarkable TNNT1, NKX2.5, and Desmin mRNA expression and higher quantitative LDH and cTnI were detected by applying protocol-2. This same protocol-2 induced cardiac morphological features that were revealed by identifying cardiomyocyte-like cells and typical sarcomeres. Conclusion: HUC-WJ is proved to be an ethical and effective source of MSCs induced cardiac differentiation, whereas BFGF supports 5-Aza in MSCs-cardiomyocytes differentiation.
In vitro cardiomyogenic potential of human umbilical vein-derived mesenchymal stem cells
Biochemical and Biophysical Research Communications, 2006
Cardiomyocyte loss in the ischemically injured human heart often leads to irreversible defects in cardiac function. Recently, cellular cardiomyoplasty with mesenchymal stem cells, which are multipotent cells with the ability to differentiate into specialized cells under appropriate stimuli, has emerged as a new approach for repairing damaged myocardium. In the present study, the potential of human umbilical cord-derived mesenchymal stem cells to differentiate into cells with characteristics of cardiomyocyte was investigated. Mesenchymal stem cells were isolated from endothelial/subendothelial layers of the human umbilical cords using a method similar to that of human umbilical vein endothelial cell isolation. Isolated cells were characterized by transdifferentiation ability to adipocytes and osteoblasts, and also with flow cytometry analysis. After treatment with 5-azacytidine, the human umbilical cord-derived mesenchymal stem cells were morphologically transformed into cardiomyocyte-like cells and expressed cardiac differentiation markers. During the differentiation, cells were monitored by a phase contrast microscope and their morphological changes were demonstrated. Immunostaining of the differentiated cells for sarcomeric myosin (MF20), desmin, cardiac troponin I, and sarcomeric a-actinin was positive. RT-PCR analysis showed that these differentiated cells express cardiac-specific genes. Transmission electron microscopy revealed a cardiomyocyte-like ultrastructure and typical sarcomers. These observations confirm that human umbilical cord-derived mesenchymal stem cells can be chemically transformed into cardiomyocytes and can be considered as a source of cells for cellular cardiomyoplasty.
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
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.
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.