THE PLASMINOGEN ACTIVATOR SYSTEM IS REGULATED BY PROSTAGLANDINS IN THE PRIMATE PERIOVULATORY FOLLICLE

Abstract

Successful ovulation requires prostaglandin (PG) production by periovulatory granulosa cells. Administration of a PG synthesis inhibitor prevented periovulatory events such as follicle rupture, while coadministration of PG synthesis inhibitor and PGE2 restored ovulation in primates. Follicle rupture requires proteolytic degradation of the follicle wall. The plasminogen activators (PA) tPA and uPA degrade extracellular matrix and likely contribute to follicle rupture. Plasminogen activator inhibitor type 1 (PAI-1) inactivates tPA and uPA. In several species, follicular PA activity increases in response to the ovulatory gonadotropin surge, with maximal PA activity measured just before ovulation. We hypothesized that 1) the ovulatory gonadotropin surge increases PA activity in the primate periovulatory follicle and 2) PGs mediate the ability of gonadotropin to increase ovarian PA activity, leading to follicle rupture in primates. Female cynomolgus macaques received gonadotropins to stimulate the development of multiple follicles. Granulosa cells, follicular fluid, and whole ovaries (n=3-4/time point) were obtained at 0, 12, 24 and 36 hours (h) after hCG administration; ovulation occurs about 40 hours after hCG. In some animals, hCG was coadministered with the PG synthesis inhibitor celecoxib to block follicular PG production during the periovulatory interval. mRNAs for tPA, uPA and PAI-1 were quantitated by real time RT-PCR in granulosa cells obtained throughout the periovulatory period. tPA and uPA mRNA levels tended to peak 12 h after hCG administration; celecoxib administration had no effect on tPA or uPA mRNA levels. PAI-1 mRNA was low 0-24 h after hCG but increased 17-fold above 0 h levels to peak 36 h after hCG administration; celecoxib treatment decreased PAI-1 mRNA to a level intermediate between levels measured at 0 h and 36 h hCG. Using immunofluorescence, granulosa cell tPA protein was low/non-detectable in ovaries obtained 0 h after hCG. Intense staining for tPA was present in ovaries obtained 12-36 h after hCG. In contrast, tPA protein was low/nondetectable in the ovaries obtained from animals treated for 36 h with hCG + celecoxib. uPA was not detected in monkey follicles by immunofluorescence. Similar to tPA, PAI-1 protein was non-detectable before hCG administration, was present 12-36 h after hCG administration, and was non-detectable in hCG + celecoxib-treated animals. Follicular PA activity as detected by in situ zymography was low before hCG administration and readily detectable 12, 24 and 36 h after hCG administration; PA activity was considerably attenuated by preincubation with an anti-tPA antibody. Animals treated for 36 h with hCG + celecoxib had low ovarian PA activity. In conclusion, tPA and PAI-1 expression by primate periovulatory follicles increased after exposure to an ovulatory dose of gonadotropin; hCG also increased ovarian PA activity. The PG synthesis inhibitor celecoxib reduced hCG-stimulated tPA and PAI-1 expression as well as PA activity. These data support the hypothesis that PGs mediate the ability of the ovulatory gonadotropin surge to increase PA activity in the granulosa cell layer, contributing to follicle rupture at ovulation. (platform)