When gonadal sex reversal occurs in mammalian species, the resultant XX males and XY females become infertile or subfertile, suggesting critical roles of sex chromosomes in germ cell differentiation. The objective of our study is to clarify the mechanism of infertility in the B6.YTIR (XY) sex-reversed female mouse, which can be attributed to a failure in the second meiotic division in its mature oocytes. In the present study, we examined the orientation of MII-spindles and the time course of cell cycle progression after oocyte activation.
Materials and methods
XX and XY females at 29-31 dpp were injected with 5IU eCG and sacrificed 45-47 hours later. GV-stage oocytes surrounded by cumulus cells (CCs) were collected from large follicles and cultured for 19 hours. At the end of culture, the oocytes were denuded of CCs and the MII-oocytes that had extruded the first polar body were subjected to following experiments: 1. After oocytes were fixed and stained with anti-α-tubulin-FITC and DAPI, the orientation of spindles was analyzed by confocal microscopy; 2. After oocytes were activated with 10 mM SrCl2, their cell cycle progression and polar body extrusion was monitored by live-image video for 5 hours, followed by Hoechst staining for visualizing pronuclei.
No difference was found in the orientation of MIIspindles between the oocytes from XY females and those from XX females. Only 37.4% (n=99) of MII-oocytes from XY females extruded the second polar body within 4 hours post activation while 69.7% (n=89) of oocytes from XX females did so. The time required for the degradation of the first-polarbody was significantly longer while those for both the extrusion of the second polar body and formation of pronuclei were shorter in the oocytes from XY females, compared to those from XX females.
A lack of coordination between sister chromatid segregation and the second polar body extrusion in the MII oocytes after activation is the major defect responsible for infertility of the B6.YTIR female mouse.