title: Co-culture of spermatogonial stem cells with sertoli cells in the presence of testosterone and FSH improved differentiation via up-regulation of post meiotic genes.
abstract: Spermatogonial stem cells (SSCs) maintain spermatogenesis throughout life in the male. Maintenance of SSCs and induction of spermiogenesis in vitro may provide a therapeutic strategy to treat male infertility. This study investigated in vitro differentiation of mouse SSCs in presence or absence of Sertoli cells, hormones and vitamins. Spermatogonial populations were enriched from testes of 4-6 week old males by magnetic activated cell sorting and anti-Thy-1 antibody. Sertoli cells isolated from 6-8 week old testes were enriched using lectin-DSA-coated plates. Isolated SSCs were cultured in the presence of Leukemia inhibitory factor (LIF) for 7 days in gelatin-coated dishes, then dissociated and cultured for 7 days in media lacking LIF in the presence or absence of Sertoli cells, with or without FSH, testosterone and vitamins. After one week, the effects of Sertoli cells ± supplementary media on SSC differentiation was evaluated by microscopy and expression of meiotic and postmeiotic transcripts using RT-PCR. SSC colonies had limited development after LIF removal alone, exhibiting low expression of meiotic (Scp3, Th2b) but not postmeiotic transcript, and loss of Stra8 and Dazl expression. SSCs co-cultured with Sertoli cells, hormones and vitamins developed spermatid-like cells expressing postmeiotic markers (TP1, TP2, Prm1) at levels over 2-fold higher than Sertoli cells or hormone/vitamins alone. Our present SSC-Sertoli co-culture provides conditions that may allow efficient in vitro differentiation of SSCs for the treatment of male infertility.
authors: Department of Anatomical Sciences, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran.
authors: Bagher
authors: B
authors: Minaee Zanganeh
authors: Tayebeh
authors: T
authors: Rastegar
authors: Mehryar
authors: M
authors: Habibi Roudkenar
authors: Iraj
authors: I
authors: Ragerdi Kashani
authors: Fardin
authors: F
authors: Amidi
authors: Farid
authors: F
authors: Abolhasani
authors: Mohammad
authors: M
authors: Barbarestani
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D018920:Coculture Techniques; D005434:Flow Cytometry; D005640:Follicle Stimulating Hormone / Q000494:pharmacology*; D005786:Gene Expression Regulation; D018189:Immunomagnetic Separation; D053605:Leukemia Inhibitory Factor / Q000494:pharmacology; D008297:Male; D008540:Meiosis / Q000187:drug effects*; D051379:Mice; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012708:Sertoli Cells / Q000187:drug effects* / Q000378:metabolism; D013093:Spermatogonia / Q000187:drug effects* / Q000378:metabolism; D013234:Stem Cells / Q000187:drug effects* / Q000378:metabolism; D013739:Testosterone / Q000494:pharmacology*; D013997:Time Factors; D014815:Vitamins / Q000494:pharmacology
chemical_list: D053605:Leukemia Inhibitory Factor; D012333:RNA, Messenger; D014815:Vitamins; D013739:Testosterone; D005640:Follicle Stimulating Hormone