BackgroundThe small molecule 6-bromoindirubin-30-oxime (BIO), a glycogen synthase kinase 3 (GSK3) inhibitor, is a pharmacological agent known to maintain self-renewal in human and mouse embryonic stem cells (ESCs). However, the precise role of GSK3 in immortalized pancreatic mesenchymal stem cells (iPMSCs) growth and survival is not completely understood at present.ResultsTo determine whether this molecule is involved in controlling the proliferation of iPMSCs, we examined the effect of BIO on iPMSCs. We found that the inactivation of GSK3 by BIO can robustly stimulate iPMSCs proliferation and mass formation as shown by QRT-PCR, western blotting, 5-Bromo-2-deoxyuridine (BrdU) immunostaining assay and tunel assay. However, we did not find the related roles of BIO on β cell differentiation by immunostaining, QRT-PCR assay, glucose-stimulated insulin release and C-peptide content analysis.ConclusionsThese results suggest that BIO plays a key role in the regulation of cell mass proliferation and maintenance of the undifferentiated state of iPMSCs.

Recent studies have demonstrated that germ-like cells could be differentiated from human umbilical cord mesenchymal stem cells (hUC-MSCs) in vitro. Whether the sexuality of hUC-MSCs affects the formation efficiency of germ-like cells derived from hUC-MSCs is still unclear. To clearly test the formation efficiency of oocyte-like cells from male and female hUC-MSCs, obtained hUC-MSCs were induced by 20% follicular fluid (FF) according to the method that has been proved by our previous studies. Results showed that hUC-MSCs differentiated into oocyte-like structures and expressed germ cell makers. It was noted that the presence of advanced oocyte-like cells in male hUC-MSCs (m-hUC-MSCs) was similar as that in female hUC-MSCs (f-hUC-MSCs); however, the expression of germ cell's specific markers in m-hUC-MSCs was delayed compared with that in f-hUC-MSCs. In addition, immunofluorescence analysis demonstrated that germ cell-specific markers, Oct4, Vasa, Dazl, ZP2, ZP3 and Stra8, were expressed on the 14th day after induction in both f-hUC-MSCs and m-hUC-MSCs. However, the size of oocyte-like cells from f-hUC-MSCs was larger than that in m-hUC-MSCs. The level of secreted oestradiol was significantly higher in f-hUC-MSCs than m-hUC-MSCs. We sought to determine whether critical germ cell's transcription factor-Figlα will promote the development of oocyte-like cells. Some germ cell-specific markers were increased when exogenous Figlα was transfected into hUC-MSCs. This process implied that germ-like cells might be produced by over-expression of exogenous germ cell-specific gene, and this process was similar as that in production of germ cells in induced pluripotent stem cells (iPSCs). Finally, to verify the feasibility that hUC-MSCs differentiate into germ cells, hUC-MSCs were transplanted into seminiferous tubules and kidney capsule of mouse, respectively, and we found the transplanted cells differentiated into germ-like cells in recipient's seminiferous tubules and kidney capsule. This study will provide a simple model to study mammalian germ cell specification using hUC-MSCs in vitro.

Umbilical cord (UC) has been suggested as a new source of mesenchymal stem cells (MSCs). In this report, we isolated MSCs from the fetal UC of goat and investigated their multipotency of differentiation into germ cells in vitro, in the presence of 0-20 % bovine follicular fluid (FF). The phenotypes, capacity of proliferation and expression of MSC markers were served as the indexes of multipotency of the isolated UC-MSCs, those were ascertained by growth curves, RT-PCR and immunofluorescent staining, respectively. Our results showed that the UC-MSCs shared a similar immunophenotype to those cells reported in mouse and human bone marrow MSCs, as well as some characteristics seen in embryonic stem cells (ESCs). In addition, our data also demonstrated that a dose-dependent function of FF on the states of differentiation of goat UC-MSCs. From 2 to 20 % of the FF can promote the proliferation of goat UC-MSC, especially the 5 % concentration of follicular fluid promote proliferation was significantly higher than 2 %. In contrast, higher concentration of follicular fluid (>10 %) induced goat UC-MSCs differentiation into oocyte-like cells. These findings provide an efficient model to study the mechanism on cell proliferation and germ cell differentiation in livestock using FF.

This study was designed to investigate the effect of platelet-derived growth factor (PDGF) on the proliferation of human umbilical cord mesenchymal stem cells (UC-MSCs) and further explore the mechanism of PDGF in promoting the proliferation of UC-MSCs. The human UC-MSCs were treated with different concentrations of PDGF, and the effects were evaluated by counting the cell number, the cell viability, the expression of PDGF receptors analyzed by RT-PCR, and the detection of the gene expression of cell proliferation, cell cycle and pluripotency, and Brdu assay by immunofluorescent staining and Quantitative real-time (QRT-PCR). The results showed that PDGF could promote the proliferation of UC-MSCs in vitro in a dose-dependent way, and 10 to 50 ng/ml PDGF had a significant proliferation effect on UC-MSCs; the most obvious concentration was 50 ng/ml. Significant inhibition on the proliferation of UC-MSCs was observed when the concentration of PDGF was higher than 100 ng/ml, and all cells died when the concentration reached 200 ng/ml PDGF. The PDGF-treated cells had stronger proliferation and antiapoptotic capacity than the control group by Brdu staining. The expression of the proliferation-related genes C-MYC, PCNA and TERT and cell cycle-related genes cyclin A, cyclin 1 and CDK2 were up-regulated in PDGF medium compared with control. However, pluripotent gene OCT4 was not significantly different between cells cultured in PDGF and cells analyzed by immunofluorescence and QRT-PCR. The PDGF could promote the proliferation of human UC-MSCs in vitro.

These results may provide a new resource for the study of oogenesis and therapy for ovarian sterility.