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Molecular and cellular characteristics of human and non-human primate multipotent stromal cells from the amnion and bone marrow during long term culture.

Pogozhykh O, Pogozhykh D, Neehus AL, Hoffmann A, Blasczyk R, Müller T - Stem Cell Res Ther (2015)

Bottom Line: Furthermore, human MSCs demonstrated the highest Sox2 levels vs. marmoset, whereas the marmoset exhibited significantly higher Lin28A values.Bisulfite sequencing of the Oct-4 promoter region displayed fewer methylations of CpG islands in the marmoset vs. human.Little is known about MSC characteristics from the preclinical animal model common marmoset vs. human during long term culture.

View Article: PubMed Central - PubMed

Affiliation: Institute for Transfusion Medicine, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany. pogozhykh.olena@mh-hannover.de.

ABSTRACT

Introduction: Multipotent stromal cells (MSCs) are among the key candidates in regenerative medicine. However variety of MSC sources and general heterogeneity lead to controversial data in functional characterization. Furthermore, despite intensive usage as preclinical animal model, little is known about MSCs of the common marmoset monkey.

Methods: MSCs derived from placental amnion and bone marrow samples from human and common marmoset were characterized in parallel over 12 passages to monitor similarities and significant differences (p ≤ 0.05, Student's t-test) in MSC markers and major histocompatibility complex (MHC) class I expression by immunohistochemistry, flow cytometry, real-time PCR, metabolic activity test, with special focus on pluripotency associated genes.

Results: Human and non-human primate MSCs were characterized for expression of MSC markers and capability of differentiation into mesenchymal lineages. MSCs could be cultured more than 100 days (26 passages), but metabolic activity was significantly enhanced in amnion vs. bone marrow MSCs. Interestingly, MHC class I expression is significantly reduced in amnion MSCs until passage 6 in human and marmoset, but not in bone marrow cells. For MSC markers, CD73 and CD105 levels remain unchanged in amnion MSCs and slightly decline in bone marrow at late passages; CD166 is significantly higher expressed in human MSCs, CD106 significantly lower vs. marmoset. All cultured MSCs showed pluripotency marker expression like Oct-4A at passage 3 significantly decreasing over time (passages 6-12) while Nanog expression was highest in human bone marrow MSCs. Furthermore, human MSCs demonstrated the highest Sox2 levels vs. marmoset, whereas the marmoset exhibited significantly higher Lin28A values. Bisulfite sequencing of the Oct-4 promoter region displayed fewer methylations of CpG islands in the marmoset vs. human.

Conclusions: Little is known about MSC characteristics from the preclinical animal model common marmoset vs. human during long term culture. Studied human and common marmoset samples share many similar features such as most MSC markers and reduced MHC class I expression in amnion cells vs. bone marrow. Furthermore, pluripotency markers indicate in both species a subpopulation of MSCs with true 'stemness', which could explain their high proliferation capacity, though possessing differences between human and marmoset in Lin28A and Sox2 expression.

No MeSH data available.


Related in: MedlinePlus

Evaluation of MHC class I expression by flow cytometry a and β2m by quantitative PCR b in MSCs. Amnion-derived cells from both species seem to regain immunogenicity triggered by MHC class I over time, whereas bone marrow MSCs already show full expression of MHC class I at passage 3. Interestingly, in marmoset expression of β2m is significantly lower than in human. *Significance by Student’s t test. β2m β2-microglobulin, MHC major histocompatibility complex
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Fig4: Evaluation of MHC class I expression by flow cytometry a and β2m by quantitative PCR b in MSCs. Amnion-derived cells from both species seem to regain immunogenicity triggered by MHC class I over time, whereas bone marrow MSCs already show full expression of MHC class I at passage 3. Interestingly, in marmoset expression of β2m is significantly lower than in human. *Significance by Student’s t test. β2m β2-microglobulin, MHC major histocompatibility complex

Mentions: Interestingly, the levels of MHC class I expression (in case of the human also called HLA) at the early passages were significantly lower in the amnion samples of both species in comparison with bone marrow. Further, we detected a significant increase in positive amnion cells of both species over time by FACS analysis (from 19.42 ± 0.43 at passage 3 to 79.47 ± 2.0 at passage 12 in human, and from 31.35 ± 0.3 at passage 3 to 87.21 ± 4.0 at passage 12 in marmoset; Fig. 4a), whereas roughly 80 % of bone marrow cells were positive for W6/32 antibody from the beginning at passage 3 and remained so until passage 12 (Fig. 4a) in both species. This was in part reflected by quantitative PCR results detecting expression of beta-2-microglobulin (β2m) (Fig. 4b). Here, interestingly, marmoset expressed overall significantly less β2m molecules independent of tissue origin and passage when compared with human (Fig. 4b).Fig. 4


Molecular and cellular characteristics of human and non-human primate multipotent stromal cells from the amnion and bone marrow during long term culture.

Pogozhykh O, Pogozhykh D, Neehus AL, Hoffmann A, Blasczyk R, Müller T - Stem Cell Res Ther (2015)

Evaluation of MHC class I expression by flow cytometry a and β2m by quantitative PCR b in MSCs. Amnion-derived cells from both species seem to regain immunogenicity triggered by MHC class I over time, whereas bone marrow MSCs already show full expression of MHC class I at passage 3. Interestingly, in marmoset expression of β2m is significantly lower than in human. *Significance by Student’s t test. β2m β2-microglobulin, MHC major histocompatibility complex
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4546288&req=5

Fig4: Evaluation of MHC class I expression by flow cytometry a and β2m by quantitative PCR b in MSCs. Amnion-derived cells from both species seem to regain immunogenicity triggered by MHC class I over time, whereas bone marrow MSCs already show full expression of MHC class I at passage 3. Interestingly, in marmoset expression of β2m is significantly lower than in human. *Significance by Student’s t test. β2m β2-microglobulin, MHC major histocompatibility complex
Mentions: Interestingly, the levels of MHC class I expression (in case of the human also called HLA) at the early passages were significantly lower in the amnion samples of both species in comparison with bone marrow. Further, we detected a significant increase in positive amnion cells of both species over time by FACS analysis (from 19.42 ± 0.43 at passage 3 to 79.47 ± 2.0 at passage 12 in human, and from 31.35 ± 0.3 at passage 3 to 87.21 ± 4.0 at passage 12 in marmoset; Fig. 4a), whereas roughly 80 % of bone marrow cells were positive for W6/32 antibody from the beginning at passage 3 and remained so until passage 12 (Fig. 4a) in both species. This was in part reflected by quantitative PCR results detecting expression of beta-2-microglobulin (β2m) (Fig. 4b). Here, interestingly, marmoset expressed overall significantly less β2m molecules independent of tissue origin and passage when compared with human (Fig. 4b).Fig. 4

Bottom Line: Furthermore, human MSCs demonstrated the highest Sox2 levels vs. marmoset, whereas the marmoset exhibited significantly higher Lin28A values.Bisulfite sequencing of the Oct-4 promoter region displayed fewer methylations of CpG islands in the marmoset vs. human.Little is known about MSC characteristics from the preclinical animal model common marmoset vs. human during long term culture.

View Article: PubMed Central - PubMed

Affiliation: Institute for Transfusion Medicine, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany. pogozhykh.olena@mh-hannover.de.

ABSTRACT

Introduction: Multipotent stromal cells (MSCs) are among the key candidates in regenerative medicine. However variety of MSC sources and general heterogeneity lead to controversial data in functional characterization. Furthermore, despite intensive usage as preclinical animal model, little is known about MSCs of the common marmoset monkey.

Methods: MSCs derived from placental amnion and bone marrow samples from human and common marmoset were characterized in parallel over 12 passages to monitor similarities and significant differences (p ≤ 0.05, Student's t-test) in MSC markers and major histocompatibility complex (MHC) class I expression by immunohistochemistry, flow cytometry, real-time PCR, metabolic activity test, with special focus on pluripotency associated genes.

Results: Human and non-human primate MSCs were characterized for expression of MSC markers and capability of differentiation into mesenchymal lineages. MSCs could be cultured more than 100 days (26 passages), but metabolic activity was significantly enhanced in amnion vs. bone marrow MSCs. Interestingly, MHC class I expression is significantly reduced in amnion MSCs until passage 6 in human and marmoset, but not in bone marrow cells. For MSC markers, CD73 and CD105 levels remain unchanged in amnion MSCs and slightly decline in bone marrow at late passages; CD166 is significantly higher expressed in human MSCs, CD106 significantly lower vs. marmoset. All cultured MSCs showed pluripotency marker expression like Oct-4A at passage 3 significantly decreasing over time (passages 6-12) while Nanog expression was highest in human bone marrow MSCs. Furthermore, human MSCs demonstrated the highest Sox2 levels vs. marmoset, whereas the marmoset exhibited significantly higher Lin28A values. Bisulfite sequencing of the Oct-4 promoter region displayed fewer methylations of CpG islands in the marmoset vs. human.

Conclusions: Little is known about MSC characteristics from the preclinical animal model common marmoset vs. human during long term culture. Studied human and common marmoset samples share many similar features such as most MSC markers and reduced MHC class I expression in amnion cells vs. bone marrow. Furthermore, pluripotency markers indicate in both species a subpopulation of MSCs with true 'stemness', which could explain their high proliferation capacity, though possessing differences between human and marmoset in Lin28A and Sox2 expression.

No MeSH data available.


Related in: MedlinePlus