Regulation of prostaglandin F2α receptor mRNA in human granulosa-luteal cells by human chorionic gonadotrophin and prostaglandin F2α

This study examined the effects of prostaglandin F-2 alpha (PGF(2 alpha)) and human chorionic gonadotropin (hCG) on the levels of PGF(2 alpha)-receptor (PGF(2 alpha)-R) mRNA and steroidogenesis, in the human granulosa luteal cell (hGLC). Human GLCs collected from patients undergoing in vitro fertilization, were cultured for 24 h, after which cells were exposed to culture media containing either vehicle, hCG (1 IU/mL), or hCG plus PGF(2 alpha) (10-(11)-10(-6) M)I for a further 24 h. Following the treatment period, media were collected and stored (-20 degrees C) until assayed for progesterone and 1 7 beta-estradiol (estradiol). Immediately following the treatment period, cells were extracted for total RNA. Transcripts for PGF(2 alpha)-R were detected by PCR with two different sets of oligonucleotide primers based on the published human and rat PGF(2 alpha)-R sequences. PCR products were confirmed to be those of PGF(2 alpha)-R by size and by Southern blot hybridization with an internal oligo nucleotide probe. All experiments were performed a minimum of three times, on cells from a minimum of three separate patients. Prostaglandin F-2 alpha-R mRNA was significantly downregulated, whereas progesterone and estradiol production were significantly stimulated by hCG. Conversely, both low (10(-11) M) and high concentrations (10(-6) M) of PGF(2 alpha) restored PGF(2 alpha)-R m RNA levels to those of the controls, whereas steroidogenesis was significantly inhibited by these conditions. At a concentration of 10(-9) M PGF(2 alpha)-R mRNA was barely detectable. Progesterone and estradiol production were inversely related to PGF(2 alpha)-R levels, since hCG-stimulated progesterone and estradiol production were completely restored in the presence of 10(-9) M PGF(2 alpha). Messenger RNA levels for the housekeeping gene p-actin were unaltered by the above treatments. In conclusion, in the human granulosa luteal cell, PGF(2)alpha-R mRNA levels are inversely related to hCG-stimulated steroidogenesis (which was biphasic in nature). Moreover, in the presence of hCG, PGF(2 alpha) downregulates its receptor mRNA, thus providing a potential form of negative feedback on its own actions, which may be important in rescuing the corpus luteum from PGF(2 alpha)-mediated luteolysis should pregnancy occur.