In vitro generation of Sertoli-like and haploid spermatid-like cells from human umbilical cord perivascular cells

Autor: Clifford Librach, Tanya Barretto, Ekaterina Shlush, Shlomit Kenigsberg, Sonja A. Swanson, Andrée Gauthier-Fisher, Leila Maghen, Sergey I. Moskovtsev
Jazyk: angličtina
Předmět:
Male
0301 basic medicine
Cellular differentiation
Primary Cell Culture
Gene Expression
Medicine (miscellaneous)
Mice
SCID
Human mesenchymal stem cell
Biology
Biochemistry
Genetics and Molecular Biology (miscellaneous)
Mice
03 medical and health sciences
0302 clinical medicine
Paracrine Communication
Niche
medicine
Animals
Humans
Spermatogenesis
Cell potency
Sertoli Cells
Gene Expression Profiling
Research
Xenograft
Ex-vivo spermatogenesis
Cell Differentiation
Mesenchymal Stem Cells
Amniotic stem cells
Cell Biology
Fetal Blood
Human umbilical cord
Spermatids
Sertoli cell
Embryonic stem cell
Cell biology
030104 developmental biology
medicine.anatomical_structure
Paracrine
030220 oncology & carcinogenesis
Amniotic epithelial cells
Heterografts
Molecular Medicine
Perivascular cell
Stem cell
Spermatogonial stem cell
Germ cell
Biomarkers
Adult stem cell
Zdroj: Stem Cell Research & Therapy
ISSN: 1757-6512
DOI: 10.1186/s13287-017-0491-8
Popis: Background First trimester (FTM) and term human umbilical cord-derived perivascular cells (HUCPVCs), which are rich sources of mesenchymal stem cells (MSCs), can give rise to Sertoli cell (SC)-like as well as haploid germ cell (GC)-like cells in vitro using culture conditions that recapitulate the testicular niche. Gamete-like cells have been produced ex vivo using pluripotent stem cells as well as MSCs. However, the production of functional gametes from human stem cells has yet to be achieved. Methods Three independent lines of FTM and term HUCPVCs were cultured using a novel 5-week step-wise in vitro differentiation protocol recapitulating key physiological signals involved in testicular development. SC- and GC-associated phenotypical properties were assessed by real-time polymerase chain reaction (RT-PCR), quantitative PCR immunocytochemistry, flow cytometry, and fluorescence in-situ hybridization (FISH). Functional spermatogonial stem cell-like properties were assessed using a xenotranplantation assay. Results Within 3 weeks of differentiation, two morphologically distinct cell types emerged including large adherent cells and semi-attached round cells. Both early GC-associated markers (VASA, DAZL, GPR125, GFR1α) and SC-associated markers (FSHR, SOX9, AMH) were upregulated, and 5.7 ± 1.2% of these cells engrafted near the inner basal membrane in a xenograft assay. After 5 weeks in culture, 10–30% of the cells were haploid, had adopted a spermatid-like morphology, and expressed PRM1, Acrosin, and ODF2. Undifferentiated HUCPVCs secreted key factors known to regulate spermatogenesis (LIF, GDNF, BMP4, bFGF) and 10–20% of HUCPVCs co-expressed SSEA4, CD9, CD90, and CD49f. We hypothesize that the paracrine properties and cellular heterogeneity of HUCPVCs may explain their dual capacity to differentiate to both SC- and GC-like cells. Conclusions HUCPVCs recapitulate elements of the testicular niche including their ability to differentiate into cells with Sertoli-like and haploid spermatid-like properties in vitro. Our study supports the importance of generating a niche-like environment under ex vivo conditions aiming at creating mature GC, and highlights the plasticity of HUCPVCs. This could have future applications for the treatment of some cases of male infertility. Electronic supplementary material The online version of this article (doi:10.1186/s13287-017-0491-8) contains supplementary material, which is available to authorized users.
Databáze: OpenAIRE
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