Isolation and characterization of Oct-4+/HLA-G+ mesenchymal stem cells from human umbilical cord matrix: differentiation potential and detection of new markers.
ABSTRACT The presence of multipotent cells in several adult and embryo-related tissues opened new paths for their use in regenerative medicine. Extraembryonic tissues such as umbilical cord are considered a promising source of stem cells, potentially useful in therapy. The characterization of cells from the umbilical cord matrix (Wharton's Jelly) and amniotic membrane revealed the presence of a population of mesenchymal-like cells, sharing a set of core-markers expressed by "mesenchymal stem cells". Several reports enlightened the differentiation capabilities of these cells, even if at times the lack of an extensive characterization of surface markers and immune co-stimulators expression revealed hidden pitfalls when in vivo transplantation was performed. The present work describes a novel isolation protocol for obtaining mesenchymal stem cells from the umbilical cord matrix. These cells are clonogenic, retain long telomeres, can undergo several population doublings in vitro, and can be differentiated in mature mesenchymal tissues as bone and adipose. We describe for the first time that these cells, besides expressing all of the core-markers for mesenchymal stem cells, feature also the expression, at both protein and mRNA level, of tolerogenic molecules and markers of all the three main lineages, potentially important for both their differentiative potential as well as immunological features.
Article: Isolation and characterization of Wharton's jelly-derived multipotent mesenchymal stromal cells obtained from bovine umbilical cord and maintained in a defined serum-free three-dimensional system.[show abstract] [hide abstract]
ABSTRACT: The possibility for isolating bovine mesenchymal multipotent cells (MSCs) from fetal adnexa is an interesting prospect because of the potential for these cells to be used for biotechnological applications. Bone marrow and adipose tissue are the most common sources of MSCs derived from adult animals. However, little knowledge exists about the characteristics of these progenitors cells in the bovine species. Traditionally most cell cultures are developed in two dimensional (2D) environments. In mammalian tissue, cells connect not only to each other, but also support structures called the extracellular matrix (ECM). The three-dimensional (3D) cultures may play a potential role in cell biotechnology, especially in tissue therapy. In this study, bovine-derived umbilical cord Wharton's jelly (UC-WJ) cells were isolated, characterized and maintained under 3D-free serum condition as an alternative of stem cell source for future cell banking. Bovine-derived UC-WJ cells, collected individually from 5 different umbilical cords sources, were successfully cultured under serum-free conditions and were capable to support 60 consecutive passages using commercial Stemline® mesenchymal stem cells expansion medium. Moreover, the UC-WJ cells were differentiated into osteocytes, chondrocytes, adipocytes and neural-like cells and cultured separately. Additionally, the genes that are considered important embryonic, POU5F1 and ITSN1, and mesenchymal cell markers, CD105+, CD29+, CD73+ and CD90+ in MSCs were also expressed in five bovine-derived UC-WJ cultures. Morphology of proliferating cells typically appeared fibroblast-like spindle shape presenting the same viability and number. These characteristics were not affected during passages. There were 60 chromosomes at the metaphase, with acrocentric morphology and intense telomerase activity. Moreover, the proliferative capacity of T cells in response to a mitogen stimulus was suppressed when bovine-derived UC-WJ cells was included in the culture which demonstrated the immunossupression profile typically observed among isolated mesenchymal cells from other species. After classified the UC-WJ cells as mesenchymal stromal phenotype the in vitro 3D cultures was performed using the AlgiMatrix® protocol. Based on the size of spheroids (283,07 μm ± 43,10 μm) we found that three weeks of culture was the best period to growth the UC-WJ cells on 3D dimension. The initial cell density was measured and the best value was 1.5 × 106 cells/well. We described for the first time the isolation and characterization of UC-WJ cells in a serum-free condition and maintenance of primitive mesenchymal phenotype. The culture was stable under 60 consecutive passages with no genetic abnormalities and proliferating ratios. Taken together all results, it was possible to demonstrate an easy way to isolate and culture of bovine-derived UC-WJ cells under 2D and 3D serum-free condition, from fetal adnexa with a great potential in cell therapy and biotechnology.BMC Biotechnology 05/2012; 12:18. · 2.35 Impact Factor
Article: Isolation and characterization of CD276+/HLA-E+ human sub-endocardial mesenchymal stem cells from chronic heart failure patients: analysis of differentiative potential and immunomodulatory markers expression.[show abstract] [hide abstract]
ABSTRACT: Mesenchymal stem cells (MSC) are virtually present in all postnatal organs as well as in perinatal tissues. MSC can be differentiated towards severalmature cytotypes and interestingly hold potentially relevant immunomodulatory features. Myocardial infarction results in severe tissue damage, cardiomyocytes loss, and eventually heart failure. Cellular cardiomyoplasty represents a promising approach for myocardial repair. Clinical trials using MSC are underway for a number of heart diseases, even if their outcomes are hampered by low long-term improvements and the possible presence of complications related to cellular therapy administration. Therefore elucidating the presence and role of MSC which reside in the post-infarct human heart should provide essential alternatives for therapy. In the present paper we show a novel method to reproducibly isolate and culture MSC from the sub-endocardial zone of human left ventricle from patients undergoing heart transplant for post-infarct chronic heart failure. By using both immunocytochemistry and RT-PCR, we demonstrated that these cells do express key MSC markers, do express heart-specific transcription factors in their undifferentiated state, while lacking strictly cardiomyocyte-specific proteins. Moreover, these cells do express immunomodulatory molecules which should disclose their further potential in immune modulation processes in the post-infarct microenvironment. Standard MSC trilineage differentiation experiments were also performed. The present paper adds new data on the basic biological features of heart-resident MSC which populate the organ following myocardial infarction. The use of heart-derived MSC to promote in-organ repair or as a cellular source for cardiomyoplasty is a fascinating and challenging task, which deserves further research efforts.Stem cells and development 09/2012; · 4.15 Impact Factor
Stem cells and development 05/2012; 21(15):2900-3. · 4.15 Impact Factor