A two-step serum-free culture system supports development of human oocytes from primordial follicles in the presence of activin.

BACKGROUND The objective of this study was to determine whether follicles grown within human ovarian cortical strip culture for 6 days in serum-free medium could be isolated at the secondary stage of pre-antral development and grown in vitro to the late pre-antral/early antral stage during a 4 day culture period. METHODS Ovarian cortical biopsies were obtained from six women aged 26-40 years, with informed consent, during elective Caesarean section. Small tissue slices of ovarian cortex, with underlying stromal tissue removed, were cultured in serum-free medium for 6 days and at the end of this period pre-antral (secondary) follicles were dissected from the strips. Seventy-four intact pre-antral follicles ranging in size (66-132 microm) (mean size 100 microm +/- 3.4) were selected for further culture. Follicles were placed individually within V-shaped microwell culture plates in serum-free medium in the presence (n = 38) or absence (n = 36) of 100 ng/ml of human recombinant activin A. RESULTS Pre-antral follicles grown for 4 days in the presence of activin A grew to a larger size (mean diameter 143 microm +/- 7.4) than those grown in control medium (mean diameter 111 microm +/- 8) (P < 0.005). Ninety percent of follicles cultured in the presence of activin A increased in size during the first 2 days of culture compared with only 36% of follicles in control medium (P > 0.005). Of the follicles surviving the entire culture period, 30% of those cultured in the presence of activin A showed normal morphology with intact oocytes and antral formation. None of the follicles grown in control medium developed antral cavities and >90% of those follicles collected at the end of the culture period showed signs of oocyte degeneration. CONCLUSIONS The results reported here demonstrate that under certain conditions, it is possible to achieve accelerated oocyte/follicle development from human primordial/primary follicles. This provides the first encouraging step towards achieving full in vitro growth of human oocytes.