In vitro primary follicle culture: Investigation into factors involved in primary follicle development

The ovary is comprised of individual functional units, follicles, composed of an oocytic germ cell and surrounding somatic granulosa and theca cells. Folliculogenesis is a complicated and highly regulated maturation process culminating in the production of a meiotically competent oocyte for fertilization. The inability to produce oocytes capable of fertilization is a growing concern among reproductive aged women. Reproducible in vitro follicle culture systems enhance our understanding of follicle development and assist in developing alternative methods for women who are in need of fertility preservation due to reproductive diseases or cancer therapies causing sterility. Current in vitro follicle cultures are not capable of sustaining primary follicle development. Combining tissue engineering principles with ovarian biology led to the development of a novel in vitro follicle culture system. An alginate hydrogel scaffold provides the three-dimensional mechanical and architectural support maintaining the follicular unit throughout growth and maturation leading to survival of the primary follicle in culture. The alginate scaffold was applied to the in vitro culture of primary follicles to determine the effects of various growth factors. We have determined that follicle stimulating hormone (FSH) promotes primary follicle development in vitro by supporting granulosa cell proliferation, production of the ovarian steroids, estradiol and progesterone, and theca cell activity. However, in the in vivo environment FSH acts as a granulosa cell maintenance factor rather than a promoter of primary follicle development. Previous to these findings, FSH was known to be required during later stages of follicle development and it was thought that FSH did not perform any function during primary follicle development. This new discovery of a role for FSH in primary follicle development is an important first step in examining its etiology and other factors involved at this stage of folliculogenesis.; Localization of the extracellular matrix (ECM) throughout folliculogenesis is important in providing the most comprehensive in vivo-like environment for in vitro folliculogenesis. Type I collagen, type IV collagen, fibronectin, and laminin distribution was determined. Laminin, found within the basement membrane of primary follicles, addition to the alginate hydrogel led to the promotion of primary follicle growth indicating a role for the ECM during follicle development. The in vitro culture of primary follicles is an important new system that can be applied to examining factors involved in this early stage of folliculogenesis.