Role of antral follicle development and cumulus cells on in vitro fertilization of mouse oocytes.

Abstract

The role exerted by antral follicle development and by companion granulosa cells on the ability of mouse oocytes at metaphase II to fuse with spermatozoa (as estimated by the kinetics of sperm-egg fusion), and to support normal fertilization (estimated by the formation of a male pronucleus) was investigated. After 17 h of culture in vitro, nuclear maturation occurred in 82% of oocytes derived from early antral follicles devoid of granulosa cells (denuded oocytes) and in 95% of oocytes derived from preovulatory follicles and cultured cumulus-intact or cumulus-free. Among the matured oocytes, 95% of cumulus-intact oocytes were arrested at metaphase II, while 61% of denuded and 50% of cumulus-free oocytes were arrested at metaphase I. The competence of denuded and cumulus-free oocytes to override the metaphase I stage was not affected by the addition of preantral granulosa, early antral granulosa or cumulus cells to the maturation medium. The kinetics of sperm-oocyte fusion were comparable in cumulus-intact and cumulus-free oocytes, and were more rapid than those of the early antral oocytes for insemination periods up to 60 min. However, when denuded oocytes were matured in medium containing early antral granulosa or cumulus cells, the kinetics of sperm-egg fusion was greatly accelerated to values equivalent to those of preovulatory oocytes. The ability of denuded oocytes to fuse with spermatozoa was unaffected by the addition of preantral granulosa cells to the maturation medium. Sixty minutes after insemination, the fertilization rates were 39% for denuded oocytes, 50% for cumulus-free oocytes and 73% for cumulus-intact oocytes. The fertilization rate of denuded and cumulus-free oocytes was significantly improved by the addition to the maturation medium of cumulus cells, but not by the addition of preantral or early antral granulosa cells. Taken together, the present results show that in mice, the maturation of both oolemma and ooplasm, required for successful fertilization, is acquired together with the development of antral follicles and is positively affected by cumulus cells during meiotic progression.