[Show abstract][Hide abstract]ABSTRACT: Recent studies have reported that induced pluripotent stem (iPS) cells from mice and humans can differentiate into primordial germ cells. However, whether iPS cells are capable of producing male germ cells is not known. The objective of this study was to investigate the differentiation potential of mouse iPS cells into spermatogonial stem cells and late-stage male germ cells. We used an approach that combines in vitro differentiation and in vivo transplantation. Embryoid bodies (EBs) were obtained from iPS cells using leukaemia inhibitor factor (LIF)-free medium. Quantitative PCR revealed a decrease in Oct4 expression and an increase in Stra8 and Vasa mRNA in the EBs derived from iPS cells. iPS cell-derived EBs were induced by retinoic acid to differentiate into spermatogonial stem cells (SSCs), as evidenced by their expression of VASA, as well as CDH1 and GFRα1, which are markers of SSCs. Furthermore, these germ cells derived from iPS cells were transplanted into recipient testes of mice that had been pre-treated with busulfan. Notably, iPS cell-derived SSCs were able to differentiate into male germ cells ranging from spermatogonia to round spermatids, as shown by VASA and SCP3 expression. This study demonstrates that iPS cells have the potential to differentiate into late-stage male germ cells. The derivation of male germ cells from iPS cells has potential applications in the treatment of male infertility and provides a model for uncovering the molecular mechanisms underlying male germ cell development.
[Show abstract][Hide abstract]ABSTRACT: To explore the expression profile of male germ cell-associated genes during the spontaneous differentiation of induced pluripotent stem cells (iPS) and assess the potency of their spontaneous differentiation into male germ cells in vitro.
Embryoid body (EB) formation was used to promote the spontaneous differentiation of iPS into male germ cells, and the expressions of germ cell-associated genes were detected by real-time PCR and PCR.
Real-time PCR and PCR revealed different expression levels of relevant genes at different times of iPS spontaneous differentiation into male germ cells. Each of the 9 genes analyzed exhibited one of the four temporal expression patterns: wavelike increase of Oct4, progressive decrease of Dppa3 and Stra8, wavelike decrease of Dazl, and decrease following initial increase of Tex14, Msy2, Scp1, Scp3 and Akap3.
Induced pluripotent stem cells express male germ cell-associated genes and male haploid genes during their spontaneous differentiation through EB formation, and have the potency of differentiating into male gametes.
Article · Jan 2011 · Zhonghua nan ke xue = National journal of andrology
[Show abstract][Hide abstract]ABSTRACT: To establish a stable recipient mouse model for stem cell transplantation into seminiferous tubules and improve the traditional techniques for transplantation.
Sixty male ICR mice were equally divided into Groups A, B, C and D, and injected with Busulfan at 15 mg/kg, 30 mg/kg, 40 mg/kg and 0 mg/kg, respectively. The survival rate was recorded every day, and the testis weight and spermatogenesis of testicular tubules were determined at 4, 8 and 12 weeks after the injection. We improved the stem cell transplantation technique and designed a new transplantation device, which connected the nozzle end, syringe and puncture needle by a three-way joint. The nozzle end was used for tentative injection, and the syringe for drawing and then injecting the cell suspension.
Only one mouse in Group C died after the injection. At 4 weeks after Busulfan treatment, the testis weight decreased apparently in Groups A, B and C, with significant differences from D (P < 0.05). The differences remained significant at 8 weeks (P < 0.05), except between Groups A and D (P > 0.05), but at 12 weeks none of the first three groups showed any significant difference from Group D (P > 0.05). At 4 and 8 weeks, the rate of hollow seminiferous tubules was < 50% in Group A and > 50% in Groups B and C, and almost returned to normal at 12 weeks, with no significant differences among the three groups (P > 0.05), but it remained unchanged in Group D. The improved transplantation device increased the success rate (> 90%), lowered the donor cell loss (< 50 microl cell suspension needed for both testes) and shortened the process ( < 10 min for one testis).
Intraperitoneal injection of Busulfan at 30 mg/kg is suitable for the establishment of the recipient mouse model of stem cell transplantation. The improved transplantation device and methods help promote the efficiency and success rate of the transplantation operation.
Article · Aug 2009 · Zhonghua nan ke xue = National journal of andrology