Survival, growth, and maturation of secondary follicles from prepubertal, young, and older adult rhesus monkeys during encapsulated three-dimensional culture: effects of gonadotropins and insulin

Abstract

A three-dimensional culture system supports the development of primate preantral follicles to the antral stage with appreciable steroid production. This study assessed i) whether in vitro developmental competence of follicles is age dependent, ii) the role of gonadotropins and insulin in supporting folliculogenesis, and iii) 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 μg/ml insulin, with or without recombinant human (rh) FSH (500 mIU/ml). No follicles survived in the 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 (NG;≤250 μm), slow-grow (SG; 250–500 μm), and fast-grow (FG;≥500 μm) according to their diameters. SG follicles cultured with 5 μg/ml insulin produced more ovarian steroids than those cultured with 0.05 μg/ml insulin by week 5. SG and FG follicles produced more AMH and VEGF than the NG, and levels peaked at weeks 2 and 5 respectively. After 100 ng/ml rh chorionic gonadotropin treatment for 34 h, more healthy oocytes were retrieved from young adults whose follicles were cultured with 5 μg/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.

Age-associated changes in bovine oocytes and granulosa cell complexes collected from early antral follicles

“…To date, a few pioneer studies have examined the effects of maternal age on pre antral follicle development. Xu et al [15] reported that pre antral follicles collected from young adult rhesus monkeys grew faster in vitro and secreted more steroids than those derived from older monkeys. In addition, Choi et al [16] reported that the overall number of follicles in the ovaries of aged mice decreased, and follicle formation of pre antral follicles cultured in vitro was lower in aged mice than in young mice, but the developmental ability to progress to the blastocyst stage was comparable between the two age groups.…”

“…Although no reports in cows, the effects of maternal aging on developmental ability of premature oocytes has been reported in other species. Xu et al [15] demonstrated that the growth rate of pre antral follicles derived from aged rhesus monkeys was slower than that of pre antral follicles derived from younger monkeys, when tested under in vitro conditions. In addition, in vitro culture of oocytes derived from early antral follicles (EAFs) of aged mice (63-67 weeks old) exhibited lower rates of pseudo-antrum formation Capsule N Itami conducted culture experiment and wrote this paper and R Kawahara-Miki conducted gene expression analysis and wrote this paper.…”

Anti-Müllerian hormone is produced heterogeneously in primate preantral follicles and is a potential biomarker for follicle growth and oocyte maturation in vitro

“…Open arrows, negative or modest staining of AMH in preantral follicles. Scale bar = 100 μm for a, b, e, and f; 200 μm for c and d. PM primordial follicle, ES early secondary follicle, LS late secondary follicle, SA small antral follicle, NC negative control Though follicle diameters did not differ at the beginning of culture, distinct cohorts of viable follicles were observed based on their growth rates and antrum formation [23]. One cohort (26.5 %) remained similar in size to the initial secondary follicles without significant change in diameters or antrum formation throughout 5 weeks of culture and was termed Bnongrowing^follicles.…”

Applicability of adult techniques for ovarian preservation to childhood cancer patients

“…The explanation for this might be threefold: dishomogeneous results in adults; pre-pubertal ovaries are made of primary and early secondary follicles, which may only partially be under FSH control [34]; the effect of such a hormonal treatment on cancer treatment outcome is currently unknown and could potentially be deleterious, thus precluding study protocol design.…”