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Differentiation of human umbilical cord matrix mesenchymal stem cells into neural-like progenitor cells and maturation into an oligodendroglial-like lineage.

Leite C, Silva NT, Mendes S, Ribeiro A, de Faria JP, Lourenço T, dos Santos F, Andrade PZ, Cardoso CM, Vieira M, Paiva A, da Silva CL, Cabral JM, Relvas JB, Grãos M - PLoS ONE (2014)

Bottom Line: In the present work MSCs were isolated from the umbilical cord matrix (Wharton's jelly) of human umbilical cord samples.The cells were additionally subjected to an oligodendroglial-oriented step-wise differentiation protocol in order to test their neural- and oligodendroglial-like differentiation capacity.The results confirmed the neural-like plasticity of MSCs, and suggested that the cells presented an oligodendroglial-like phenotype throughout the differentiation protocol, in several aspects sharing characteristics common to those of bona-fide oligodendrocyte precursor cells and differentiated oligodendrocytes.

View Article: PubMed Central - PubMed

Affiliation: Biocant - Technology Transfer Association, Biocant Park, Cantanhede, Portugal.

ABSTRACT
Mesenchymal stem cells (MSCs) are viewed as safe, readily available and promising adult stem cells, which are currently used in several clinical trials. Additionally, their soluble-factor secretion and multi-lineage differentiation capacities place MSCs in the forefront of stem cell types with expected near-future clinical applications. In the present work MSCs were isolated from the umbilical cord matrix (Wharton's jelly) of human umbilical cord samples. The cells were thoroughly characterized and confirmed as bona-fide MSCs, presenting in vitro low generation time, high proliferative and colony-forming unit-fibroblast (CFU-F) capacity, typical MSC immunophenotype and osteogenic, chondrogenic and adipogenic differentiation capacity. The cells were additionally subjected to an oligodendroglial-oriented step-wise differentiation protocol in order to test their neural- and oligodendroglial-like differentiation capacity. The results confirmed the neural-like plasticity of MSCs, and suggested that the cells presented an oligodendroglial-like phenotype throughout the differentiation protocol, in several aspects sharing characteristics common to those of bona-fide oligodendrocyte precursor cells and differentiated oligodendrocytes.

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Cell morphology and expression of nestin in hUCM-MSC and during the first steps of differentiation.Phase contrast images of undifferentiated hUCM-MSCs (A), neuroectodermal-like induced MSCs – niMSCs (B) and NSC-like cells after 3 (C) and 18 days (D) in culture in NSC induction medium. Immunofluorescence microscopy images for the neural precursor marker nestin (in green) in undifferentiated MSCs (E) and NSC-like cells (F). Counterstaining of nuclei was performed with DAPI (in blue). In (A-D) the scale bar represents 200 µm and in (E-F) represents 50 µm. Images are representative of at least 3 independent experiments.
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pone-0111059-g003: Cell morphology and expression of nestin in hUCM-MSC and during the first steps of differentiation.Phase contrast images of undifferentiated hUCM-MSCs (A), neuroectodermal-like induced MSCs – niMSCs (B) and NSC-like cells after 3 (C) and 18 days (D) in culture in NSC induction medium. Immunofluorescence microscopy images for the neural precursor marker nestin (in green) in undifferentiated MSCs (E) and NSC-like cells (F). Counterstaining of nuclei was performed with DAPI (in blue). In (A-D) the scale bar represents 200 µm and in (E-F) represents 50 µm. Images are representative of at least 3 independent experiments.

Mentions: After assuring the fidelity of the UCM-MSCs obtained, we focused on the differentiation of these cells into oligodendrocyte-like cells using a stepwise protocol, based on the literature for neural- and oligodendroglial-like induction of distinct types of stem cells, including embryonic stem cells [24], [25], [39] and MSCs [7], [9], [22]. MSCs were subjected to distinct soluble factors, culture surfaces and coating conditions during the differentiation process (Figure S3). Briefly, undifferentiated MSCs (Figures 3A and 4A-C) were treated with epidermal growth factor (EGF) for 3 days for generation of neuroectodermal-induced MSCs (niMSCs) (Figure 3B). Next, niMSCs were cultured on non-treated polystyrene in the presence of EGF and basic fibroblast growth factor (bFGF) for induction into neural stem-like (NSC-like) cells (Figures 3C, D, F and 4D-F). The latter were then cultured on tissue culture polystyrene using different coating conditions (Figure S3) for the generation of oligodendrocyte precursor-like (OPC-like) cells (Figure 4G-O). Finally, these progenitor cells were seeded on poly-D-lysine (PDL) and laminin-2/merosin (MN) double-coated surfaces for the generation of oligodendrocyte-like (OL-like) cells (Figures 5 and 6).


Differentiation of human umbilical cord matrix mesenchymal stem cells into neural-like progenitor cells and maturation into an oligodendroglial-like lineage.

Leite C, Silva NT, Mendes S, Ribeiro A, de Faria JP, Lourenço T, dos Santos F, Andrade PZ, Cardoso CM, Vieira M, Paiva A, da Silva CL, Cabral JM, Relvas JB, Grãos M - PLoS ONE (2014)

Cell morphology and expression of nestin in hUCM-MSC and during the first steps of differentiation.Phase contrast images of undifferentiated hUCM-MSCs (A), neuroectodermal-like induced MSCs – niMSCs (B) and NSC-like cells after 3 (C) and 18 days (D) in culture in NSC induction medium. Immunofluorescence microscopy images for the neural precursor marker nestin (in green) in undifferentiated MSCs (E) and NSC-like cells (F). Counterstaining of nuclei was performed with DAPI (in blue). In (A-D) the scale bar represents 200 µm and in (E-F) represents 50 µm. Images are representative of at least 3 independent experiments.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4214693&req=5

pone-0111059-g003: Cell morphology and expression of nestin in hUCM-MSC and during the first steps of differentiation.Phase contrast images of undifferentiated hUCM-MSCs (A), neuroectodermal-like induced MSCs – niMSCs (B) and NSC-like cells after 3 (C) and 18 days (D) in culture in NSC induction medium. Immunofluorescence microscopy images for the neural precursor marker nestin (in green) in undifferentiated MSCs (E) and NSC-like cells (F). Counterstaining of nuclei was performed with DAPI (in blue). In (A-D) the scale bar represents 200 µm and in (E-F) represents 50 µm. Images are representative of at least 3 independent experiments.
Mentions: After assuring the fidelity of the UCM-MSCs obtained, we focused on the differentiation of these cells into oligodendrocyte-like cells using a stepwise protocol, based on the literature for neural- and oligodendroglial-like induction of distinct types of stem cells, including embryonic stem cells [24], [25], [39] and MSCs [7], [9], [22]. MSCs were subjected to distinct soluble factors, culture surfaces and coating conditions during the differentiation process (Figure S3). Briefly, undifferentiated MSCs (Figures 3A and 4A-C) were treated with epidermal growth factor (EGF) for 3 days for generation of neuroectodermal-induced MSCs (niMSCs) (Figure 3B). Next, niMSCs were cultured on non-treated polystyrene in the presence of EGF and basic fibroblast growth factor (bFGF) for induction into neural stem-like (NSC-like) cells (Figures 3C, D, F and 4D-F). The latter were then cultured on tissue culture polystyrene using different coating conditions (Figure S3) for the generation of oligodendrocyte precursor-like (OPC-like) cells (Figure 4G-O). Finally, these progenitor cells were seeded on poly-D-lysine (PDL) and laminin-2/merosin (MN) double-coated surfaces for the generation of oligodendrocyte-like (OL-like) cells (Figures 5 and 6).

Bottom Line: In the present work MSCs were isolated from the umbilical cord matrix (Wharton's jelly) of human umbilical cord samples.The cells were additionally subjected to an oligodendroglial-oriented step-wise differentiation protocol in order to test their neural- and oligodendroglial-like differentiation capacity.The results confirmed the neural-like plasticity of MSCs, and suggested that the cells presented an oligodendroglial-like phenotype throughout the differentiation protocol, in several aspects sharing characteristics common to those of bona-fide oligodendrocyte precursor cells and differentiated oligodendrocytes.

View Article: PubMed Central - PubMed

Affiliation: Biocant - Technology Transfer Association, Biocant Park, Cantanhede, Portugal.

ABSTRACT
Mesenchymal stem cells (MSCs) are viewed as safe, readily available and promising adult stem cells, which are currently used in several clinical trials. Additionally, their soluble-factor secretion and multi-lineage differentiation capacities place MSCs in the forefront of stem cell types with expected near-future clinical applications. In the present work MSCs were isolated from the umbilical cord matrix (Wharton's jelly) of human umbilical cord samples. The cells were thoroughly characterized and confirmed as bona-fide MSCs, presenting in vitro low generation time, high proliferative and colony-forming unit-fibroblast (CFU-F) capacity, typical MSC immunophenotype and osteogenic, chondrogenic and adipogenic differentiation capacity. The cells were additionally subjected to an oligodendroglial-oriented step-wise differentiation protocol in order to test their neural- and oligodendroglial-like differentiation capacity. The results confirmed the neural-like plasticity of MSCs, and suggested that the cells presented an oligodendroglial-like phenotype throughout the differentiation protocol, in several aspects sharing characteristics common to those of bona-fide oligodendrocyte precursor cells and differentiated oligodendrocytes.

Show MeSH
Related in: MedlinePlus