Progesterone secretion by granulosa cells from different sized follicles of human ovaries after short term incubation

Studies on the ability of granulosa to secrete progesterone after hormonal treatment have not addressed the possibility of variations in response associated with the stage of follicular development. Accordingly, granulosa were isolated from follicular fluid obtained from apparently healthy ovaries of patients undergoing tubal bypass. Follicle size was estimated by laparoscopic and ultrasonic measurements, and granulosa cells were prepared by centrifugation and treatment with EGTA and hypertonic sucrose solution to minimize aggregation. Cell viability was determined and 10(6) cells/ml medium 199D were incubated for 24 h without or with ovine LH (1 microgram/ml) or 17 beta-estradiol (5 micrograms/ml). Progesterone secretion by granulosa increased from 6 to 30 ng/24 h/10(6) cells and correlated with follicle sizes of 5--25 mm. Compared to untreated cells obtained from the same follicle, granulosa from follicles ranging from 5--15 mm secreted 2.5 times more progesterone when treated with LH. This effect of LH diminished with further increases in follicle size, with LH having no effect on cells from follicles larger than 20 mm. Compared to untreated cells from the same follicles, 17 beta-estradiol caused a progressive inhibition of progesterone secretion by granulosa that correlated with follicle sizes between 5--18 mm, with almost 60% inhibition occurring in cells from 18-mm follicles. Thereafter, the inhibitory effect of 17 beta-estradiol diminished, and no effect was found in granulosa from follicles larger than 22 mm. These results indicate that LH and 17 beta-estradiol have maximal in vitro effects on progesterone secretion by granulosa from follicles with diameters of 15--18 mm. The change in response to LH and 17 beta-estradiol observed in samples from larger follicles may be due to the fact that these cells already have maximal binding of LH and 17 beta-estradiol to their receptors. Alternatively, it is possible that cells from larger follicles are desensitized to LH and 17 beta-estradiol.