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In Vitro Cell Dev Biol Anim. 2016 Dec;52(10):1060-1071. Epub 2016 Aug 8.

Amniotic membrane mesenchymal stem cells can differentiate into germ cells in vitro.

Author information

1
Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran.
2
Department of Pharmacology and Toxicology, Faculty of Pharmacy and Pharmaceutical Sciences Research Center (PSRC), Tehran University of Medical Sciences (TUMS), Tehran, Iran.
3
Department of Pharmaceutics, Faculty of Pharmacy, Tehran University of Medical Sciences, Tehran, Iran.
4
Cellular and Molecular Research Center, Kurdistan University of Medical Sciences, Sanandaj, Iran.
5
Tashkhis Baft Aragene Company Ltd (TBA), Tehran, Iran.
6
Infertility and Reproductive Health Research Center, Health Research Institute, Babol University of Medical Sciences, P.O. Box: 47318-38711, Amirkola, Babol, Iran. ghasemzadeh78@gmail.com.

Abstract

This is the first report on differentiation of mouse amniotic membrane mesenchymal stem cells (AM-MSCs) into male germ cells (GCs). AM-MSCs have the multipotent differentiation capacity and can be differentiated into various cell types. In the present study, AM-MSCs were induced for differentiation into GCs. AM-MSCs were isolated from mouse embryonic membrane by enzymatic digestion. AM-MSCs were characterized with osteogenic and adipogenic differentiation test and flow cytometric analysis of some CD-markers. AM-MSCs were induced to differentiate into GCs using a creative two-step method. Passage-3 AM-MSCs were firstly treated with 25 ng/ml bone morphogenetic protein 4 (BMP4) for 5 d and in continuing with 1 μM retinoic acid (RA) for 12 d (total treatment time was 17 d). At the end of the treatment period, real-time reverse transcription (RT)-PCR was performed to evaluate the expression of GC-specific markers-Itgb1, Dazl, Stra8, Piwil2, Mvh, Oct4, and c-Kit- in the cells. Moreover, flow cytometry and immunofluorescence staining were performed to evaluate the expression of Mvh and Dazl at protein level. Real-time RT-PCR showed that most of the tested markers were upregulated in the treated AM-MSCs. Furthermore, flow cytometric and immunofluorescence analyses both revealed that a considerable part of the treated cells expressed GC-specific markers. The percentage of positive cells for Mvh and Dazl was about 23 and 46%, respectively. Our results indicated that a number of AM-MSCs successfully differentiated into the GCs. Finally, it seems that AM-MSCs would be a potential source of adult pluripotent stem cells for in vitro generation of GCs and cell-based therapies for treatment of infertility.

KEYWORDS:

Amniotic membrane mesenchymal stem cells; BMP4; Differentiation; Germ cells; Retinoic acid

PMID:
27503516
DOI:
10.1007/s11626-016-0073-6
[Indexed for MEDLINE]

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