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Effects of vitamin A on in vitro maturation of pre-pubertal mouse spermatogonial stem cells.

ABSTRACT: Testicular tissue cryopreservation is the only potential option for fertility preservation in pre-pubertal boys exposed to gonadotoxic treatment. Completion of spermatogenesis after in vitro maturation is one of the future uses of harvested testicular tissue. The purpose of the current study was to evaluate the effects of vitamin A on in vitro maturation of fresh and frozen-thawed mouse pre-pubertal spermatogonial stem cells in an organ culture system. Pre-pubertal CD1 mouse fresh testes were cultured for 7 (D7), 9 (D9) and 11 (D11) days using an organ culture system. Basal medium was supplemented with different concentrations of retinol (Re) or retinoic acid (RA) alone or in combination. Seminiferous tubule morphology (tubule diameter, intra-tubular cell type), intra-tubular cell death and proliferation (PCNA antibody) and testosterone level were assessed at D7, D9 and D11. Pre-pubertal mouse testicular tissue were frozen after a soaking temperature performed at -7 °C, -8 °C or -9 °C and after thawing, were cultured for 9 days, using the culture medium preserving the best fresh tissue functionality. Retinoic acid at 10(-6)M and retinol at 3.3.10(-7)M, as well as retinol 10(-6)M are favourable for seminiferous tubule growth, maintenance of intra-tubular cell proliferation and germ cell differentiation of fresh pre-pubertal mouse spermatogonia. Structural and functional integrity of frozen-thawed testicular tissue appeared to be well-preserved after soaking temperature at -8 °C, after 9 days of organotypic culture using 10(-6)M retinol. RA and Re can control in vitro germ cell proliferation and differentiation. Re at a concentration of 10(-6)M maintains intra-tubular cell proliferation and the ability of spermatogonia to initiate spermatogenesis in fresh and frozen pre-pubertal mouse testicular tissue using a soaking temperature at -8 °C. Our data suggested a possible human application for in vitro maturation of cryopreserved pre-pubertal testicular tissue.

SUBMITTER: Travers A

PROVIDER: S-EPMC3857286 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Effects of vitamin A on in vitro maturation of pre-pubertal mouse spermatogonial stem cells.

Travers Albanne A Arkoun Brahim B Safsaf Athmane A Milazzo Jean-Pierre JP Absyte Anne A Bironneau Amandine A Perdrix Anne A Sibert Louis L Macé Bertrand B Cauliez Bruno B Rives Nathalie N

PloS one 20131209 12

Testicular tissue cryopreservation is the only potential option for fertility preservation in pre-pubertal boys exposed to gonadotoxic treatment. Completion of spermatogenesis after in vitro maturation is one of the future uses of harvested testicular tissue. The purpose of the current study was to evaluate the effects of vitamin A on in vitro maturation of fresh and frozen-thawed mouse pre-pubertal spermatogonial stem cells in an organ culture system. Pre-pubertal CD1 mouse fresh testes were cu ...[more]

PMID: 24349372

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Project description:Graphical abstract Highlights • Background: The risk of infertility after chemotherapy is unpredictable in children with cancer.• Our in vitro study demonstrated short-term damage of chemotherapy on immature mouse testis.• Cisplatin and doxorubicin induce acute spermatogonial stem cell loss that does not persist two months later.• High concentrations of cisplatin impair later phases of spermatogenesis in the long-term. Chemotherapy can affect testis development of young boys with cancer, reducing the chances of fatherhood in adulthood. Studies using experimental models are needed to determine the damage caused by individual chemotherapy drugs in order to predict the risk of infertility and direct patients towards appropriate fertility preservation options. Here, we investigated the individual role of two drugs, cisplatin and doxorubicin, using an in vitro culture model of prepubertal (postnatal day 5) mouse testis that supports induction and maintenance of full spermatogenesis. Twenty-four hour exposure with either drug at clinically-relevant doses (0.25, 0.5 or 0.75 ?g/mL for cisplatin, or 0.01, 0.03 or 0.05 ?g/mL for doxorubicin), induced an acute significant loss of spermatogonial stem cells (SSCs; PLZF+), proliferating SSCs (PLZF+BrdU+), total germ cells (MVH+), and spermatocytes (SCP3+) one week after chemotherapy exposure. By the time of the first (Week 4) and second (Week 8) waves of spermatogenesis, there was no longer any effect on SSC or proliferating SSC numbers in drug-exposed testis compared to untreated tissue: however, the populations of total germ cells and spermatocytes were still lower in the higher-dose cisplatin treated groups, along with a reduced frequency of round and elongated spermatids in both cisplatin- and doxorubicin-treated testis fragments. Overall, this study details a direct impairment of germ cell development following acute chemotherapy-induced damage during the prepubertal phase, most likely due to an effect on SSCs, using an in vitro culture system that successfully recapitulates key events of mouse spermatogenesis.

Dataset Information

Effects of vitamin A on in vitro maturation of pre-pubertal mouse spermatogonial stem cells.

Publications

Effects of vitamin A on in vitro maturation of pre-pubertal mouse spermatogonial stem cells.

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