Bone morphogenetic protein 15 induces differentiation of mesenchymal stem cell derived from human follicular fluid to oocyte like cell
- Published
- Accepted
- Subject Areas
- Cell Biology, Molecular Biology, Anatomy and Physiology, Gynecology and Obstetrics, Histology
- Keywords
- Follicular fluid, Mesenchymal stem cells, Oocyte like cell, Bone morphogenetic protein15
- Copyright
- © 2019 Taheri Moghadam et al.
- Licence
- This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ Preprints) and either DOI or URL of the article must be cited.
- Cite this article
- 2019. Bone morphogenetic protein 15 induces differentiation of mesenchymal stem cell derived from human follicular fluid to oocyte like cell. PeerJ Preprints 7:e28006v1 https://doi.org/10.7287/peerj.preprints.28006v1
Abstract
Background. To study the effect of Bone morphogenetic protein 15 on differentiation potential of mesenchymal stem cell derived from human follicular fluid to oocyte like cell. Methods. Human FF derived cells were collected from 78 women in assisted fertilization program, and cultured in differentiation medium containing human recombinant BMP15 for 21 days. Mesenchymal stem cells and OLCs were characterized by real-time PCR and immunocytochemistry (ICC) staining. Results. MSCs expressed germ line stem cell markers, such as OCT4 and NANOG. After 15 days, OLCs formed and expressed zona pellucida markers (ZP2, ZP3), and reached 20 – 30 µm in diameters. Ten days after induction with BMP15, round cells remarkably developed, and the maximum size of OLCs reached 115 µm. Finally, a decrease ranging from 0.04 to 4.5 in the expression of pluripotency and oocyte specific markers was observed in the cells cultured in BMP15 supplemented medium. Our work demonstrates, FF derived MSCs have an innate potency to differentiate into OLCs, and BMP15 is effective in stimulating the differentiation of these cells, which may give an in vitro model to examine human germ cell development.
Author Comment
This is a submission to PeerJ for review.
Supplemental Information
DNA synthesis by BrdU incorporation
Graph pad prism 6 data analysis method
Cover letter
All authors have approved the manuscript and agree with its submission to PeerJ.
ELISA for estradiol assays
Determination of estradiol production in medium within process of differentiation, by graph pad prism 6 software.
Graph analysis
cytoplasmic immunostaining for vimentin analysis by graphpad prism6.
RT-PCR raw data
CT number of amplification plot for ZP3
RT-PCR
CT number of amplification plot for ZP2 and ZP3 weak 1.
RT-PCR- raw data
CT number of treated group.ZP3 and Nanog
RT-PCR raw data
CT numbers of ZP2 control group.
RT-PCR raw data
CT numbers of OCT4 and Nanog after treatment with induction medium.
Sampling the ovarian follicular fluids
Hypo-osmotic lysis technique was used to enrich the follicular cells and elimination of red blood cells from FF.
Adipogenic differentiation of human FF cells. Raw dat
Oil Red O staining
Osteoinduction prestaining
osteogenic differentiation of human FF cells. Von Kossa staining of follicular fluid derived cells with differentiation medium during three weeks
Adipogenic media .Raw data
before staining procedure
Cell morphology raw data
Primary cell culture of FF planted on plate .
cell morphology.Raw data
fibroblast-like shape.
Cell morphology after treatment
morphology of FF derived cells culture in medium supplemented with BMP15
cell morphology
figurevshow different size of germ cell and OLCs developed spontaneously from human FF derived cells.
DAPI stainig.raw data
Pluripotent activities of human FF-derived cells
FITC raw data
Pluripotent activities of human FF-derived cells
DAPI for OCT4.Raw data
pluripotency markers of OCT4
cytoplasmic immunostaining for vimentin
Raw data.
cytoplasmic immunostaining for vimentin.Raw data
vimentin immunostaining of FF derived adherent cells
vimentin ICC of FF derived adherent cells
DAPI and FITC staining.
DAPI OLC
after treatment with BMP15, using germ cell marker immunolocalization.
OLC for FITC
after treatment with BMP15 using germ cell marker immunolocalization
OLC for DAPI
after treatment with BMP15 using germ cell marker immunolocalization
Control negative
Control negative for using germ cell marker immunolocalization. data not shown in main document.
DAPI for negative control
the nuclei were counterstained and observed by 4′,6-Diamidino-2-phenylindole (DAPI)
