Dataset Information

BMP4 promotes SSEA-1(+) hUC-MSC differentiation into male germ-like cells in vitro.

ABSTRACT:

Objectives

Recent studies have demonstrated that primordial germ cells (PGC) can be differentiated from human umbilical cord mesenchymal stem cells (hUC-MSCs), and embryonic stem cells (ESCs) in vitro. Nevertheless, efficiencies were low and unstable. Here, whether hUC-MSCs can be induced to differentiate into germ-like cells with the aid of bone morphogenetic protein (BMP4) was investigated.

Materials and methods

Human umbilical cord mesenchymal stem cells were freshly isolated and cultured with BMP4. SSEA-1(+/-) cells were purified using magnetic-activated cell sorting (MACS) from the hUC-MSCs, and further induced with BMP4. Quantitative real-time PCR (qRT-PCR) and immunofluorescence analysis were used to determine PGC and germ-like cell-specific markers.

Results

Human umbilical cord mesenchymal stem cells differentiated into SSEA-1(+) spherical PGC-like cells efficiently with 12.5 ng/ml BMP4. qRT-PCR and immunofluorescence analysis demonstrated that SSEA-1(+) cells expressed higher levels of PGC-specific markers than SSEA-1(-) cells. Furthermore, SSEA-1(+) cells were induced with BMP4 to differentiate into STRA8, SCP3, DMRT1 and PLZF-positive male germ-like cells, and some sperm-like cells were obtained by 7-14 days after induction.

Conclusion

These results suggest that SSEA-1(+) hUC-MSCs can differentiate into male germ-like cells in the presence of BMP4. This study provides an efficient protocol to study germ-cell development using hUC-MSCs.

SUBMITTER: Li N

PROVIDER: S-EPMC6495181 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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BMP4 promotes SSEA-1(+) hUC-MSC differentiation into male germ-like cells in vitro.

Li N N Pan S S Zhu H H Mu H H Liu W W Hua J J

Cell proliferation 20140613 4

<h4>Objectives</h4>Recent studies have demonstrated that primordial germ cells (PGC) can be differentiated from human umbilical cord mesenchymal stem cells (hUC-MSCs), and embryonic stem cells (ESCs) in vitro. Nevertheless, efficiencies were low and unstable. Here, whether hUC-MSCs can be induced to differentiate into germ-like cells with the aid of bone morphogenetic protein (BMP4) was investigated.<h4>Materials and methods</h4>Human umbilical cord mesenchymal stem cells were freshly isolated a ...[more]

PMID: 24923741

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Project description:In vitro gamete derivation from stem cells has potential applications in animal reproduction as an alternative method for the dissemination of elite animal genetics, production of transgenic animals, and conservation of endangered species. Mesenchymal stem cells (MSCs) may be suitable candidates for in vitro gamete derivation considering their differentiative capacity and their potential for cell therapy. Due to its relevance in gametogenesis, it has been reported that retinoic acid (RA) and bone morphogenetic protein (BMP) 4 are able to upregulate the expression of specific markers associated to the early stages of germ cell (GCs) differentiation in bovine fetal MSCs (bfMSCs). In the present study, we used polycistronic vectors containing combinations of GC genes DAZL, STRA8, and BOULE followed by exposure to BMP4 or RA to induce GC differentiation of bovine fetal adipose tissue-derived MSC (AT-MSCs). Cells samples at Day 14 were analyzed according to the expression of pluripotent genes NANOG and OCT4 and GC genes DAZL, STRA8, BOULE, PIWI, c-KIT, and FRAGILIS using Q-PCR. Fetal and adult testis and AT-MSCs samples were also analyzed for the expression of DAZL, STRA8, and NANOG using immunofluorescence. Increased gene expression levels in the adult testis and cell-specific distribution of DAZL, STRA8, and NANOG in the fetal testis suggest that these markers are important components of the regulatory network that control the in vivo differentiation of bovine GCs. Overexpression of DAZL and STRA8 in bi-cistronic and DAZL, STRA8, and BOULE in tri-cistronic vectors resulted in the upregulation of OCT4, NANOG, and PIWIL2 in bovine fetal AT-MSCs. While BMP4 repressed NANOG expression, this treatment increased DAZL and c-KIT and activated FRAGILIS expression in bovine fetal AT-MSCs. Treatment with RA for 14 days increased the expression of DAZL and FRAGILIS and maintained the mRNA levels of STRA8 in bovine fetal AT-MSCs transfected with bi-cistronic and tri-cistronic vectors. Moreover, RA treatment repressed the expression of OCT4 and NANOG in these cells. Thus, overexpression of DAZL, STRA8, and BOULE induced the upregulation of the pluripotent markers and PIWIL2 in transfected bovine fetal AT-MSCs. The partial activation of GC gene expression by BMP4 and RA suggests that both factors possess common targets but induce different gene expression effects during GC differentiation in overexpressing bovine fetal AT-MSCs.

Dataset Information

BMP4 promotes SSEA-1(+) hUC-MSC differentiation into male germ-like cells in vitro.

Objectives

Materials and methods

Results

Conclusion

Publications

BMP4 promotes SSEA-1(+) hUC-MSC differentiation into male germ-like cells in vitro.

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