Irradiated Mouse Testes Efficiently Support Spermatogenesis Derived From Donor Germ Cells of Mice and Rats

Abstract

Testicular cell transplantation has been widely used to investigate the biology of spermatogonial stem cells, production of transgenic animals, and restoration of fertility in rodent models. One critical step in successful transplantation is the preparation of the recipient testes. Busulfan has been widely used, but irradiation has been often suggested as an alternative. There have only been limited reports of the use of irradiated animals as transplant recipients for studying differentiation of transplanted cells, and there has been no direct comparison of irradiation and busulfan as preparation methods. Mouse testes treated with local fractionated irradiation (1.5 + 12 Gy) were compared with busulfan-treated testes as recipients using mouse-to-mouse and rat-to-mouse germ cell transplantation. The fractionated irradiation schedule resulted in depletion of endogenous spermatogenesis similar to that produced by busulfan doses of 50-55 mg/kg. When immature mouse or rat testicular germ cells were transplanted into the irradiated testes, donor cells derived from either rat or mouse spermatogonial stem cells were able to form colonies of differentiated spermatogenic cells 10-13 weeks after transplantation with similar efficiencies as in busulfan-treated testes. Locally irradiated testes could be considered as an alternative to busulfan treatment for animal recipients of germ cell transplants that cannot endure the systemic toxicity of busulfan.

Dynamic expression pattern and subcellular localization of the Rhox10 homeobox transcription factor during early germ cell development

“…First, we examined the expression of Rhox10 in three germ cell-deficient mouse models: i) a mouse model of Klinefelter's syndrome (KS) that has a XXY karyotype and virtually lack all germ cells in the adult testis (Swerdloff et al 2011); ii) irradiated wild-type mice, which lose spermatogonial stem cells (SSCs) and thus contain only somatic cells 5 weeks after irradiation (Zhang et al 2006); and iii) juvenile spermatogonial depletion (jsd) mice, which lack all germ cells in adult testes except for undifferentiated type A spermatogonia (A s , A pr , and A al spermatogonia) (de Rooij et al 1999). Using qRT-PCR analysis, we found that testes from adult KS mice had nearly undetectable levels of Rhox10 mRNA ( Fig.…”

Donor Sertoli cells transplanted into irradiated rat testes stimulate partial recovery of endogenous spermatogenesis

“…To harvest single cells from the tubules, we removed the tunica and sequentially digested the tissue with enzymes at 35 8C in a shaking water bath as described in detail recently (Zhang et al 2006). Tubules were prepared by two digestions, Transplanted Sertoli cells stimulate spermatogenesis first with collagenase and then with collagenase and hyaluronidase in DMEM/F12 medium containing DNase I and fetal bovine serum for 20-30 min each.…”

“…Serial sections were prepared for immunohistochemistry, including antigen retrieval, inhibition of endogenous peroxidase, and blocking as described previously (Zhang et al 2006). Rabbit polyclonal (1:5000 dilution, Novus Biologicals, Littleton, CO, USA) or mouse monoclonal (1:1000 dilution, or Roche-Applied Science) anti-GFP was used to stain donor Sertoli and germ cells from the transgenic rats.…”

Cell‐specific ablation in the testis: what have we learned?

“…W/W v ) mice are unavailable or inappropriate (e.g. difficult to breed, immunocompatibility issues with donor cells, use of other species) (Zhang et al ., 2006). Even within mice, the removal of germ cells using busulfan is a strain‐dependent balance between the required toxicity to kill sufficient germ cells and the sensitivity of the animal to the wider toxicological effects of the drug, particularly on haematopoiesis (Meistrich et al ., 1978; Abuelhija et al ., 2013).…”

“…The adult Leydig cells are completely dependent on LH support for both development and adult function; in the absence of LH few adult Leydig cells develop and circulating testosterone levels are barely detectable (O'Shaughnessy et al ., 1998; Baker & O'Shaughnessy, 2001; Lei et al ., 2001; Zhang et al ., 2001). In contrast, the foetal Leydig cells in rodents appear to function largely independently of hormonal support (El Gehani et al ., 1998; O'Shaughnessy et al ., 1998; Baker et al ., 1999; Lei et al ., 2001; Zhang et al ., 2006) although in other species LH (or hCG in humans) is required for foetal androgen production (O'Shaughnessy & Fowler, 2011). …”