Survival, growth, and maturation of secondary follicles from prepubertal, young and older adult, rhesus monkeys during encapsulated three-dimensional (3D) culture: effects of gonadotropins and insulin.
A three-dimensional culture system supports development of primate preantral follicles to the antral stage with appreciable steroid production. The present study assessed: (1) whether in vitro developmental competence of follicles is age-dependent, (2) the role of gonadotropins and insulin in supporting folliculogenesis, and (3) anti-Müllerian hormone (AMH) and vascular endothelial growth factor (VEGF) production by growing follicles. Ovaries were obtained from prepubertal, young and older adult, rhesus macaques. Secondary follicles were encapsulated into alginate beads and cultured individually for 40 days in media containing 0.05 or 5 microg/ml insulin, with or without recombinant human (rh) follicle stimulating hormone (FSH, 500 mIU/ml). No follicles survived in culture without rhFSH. In the presence of rhFSH, survival was lower for follicles from older animals, whereas growth, i.e., follicle diameter, was less by day 40 for follicles from prepubertal animals. The surviving follicles were categorized as no-grow (</= 250 microm), slow-grow (250-500 microm), and fast-grow (>/= 500 microm) according to their diameters. Slow-grow follicles cultured with 5 microg/ml insulin produced more ovarian steroids compared with those with 0.05 microg/ml insulin by week 5. Slow- and fast-grow follicles produced more AMH and VEGF than the no-grows, and levels peaked at week 2 and 5, respectively. After 100 ng/ml rh chorionic gonadotropin treatment for 34 hrs, more healthy oocytes were retrieved from young adults whose follicles were cultured with 5 microg/ml insulin. This culture system offers an opportunity to characterize the endocrine and paracrine function of primate follicles that influence follicle growth and oocyte maturation.
Xu J, Bernuci MP, Lawson MS, Yeoman RR, Fisher TE, Zelinski MB, Stouffer RL. Reproduction, Aug. Epub 2010. PMID 20729335