TRANSCRIPTIONAL REGULATION OF THE TISSUE FACTOR GENE BY PROGESTINS INHUMAN ENDOMETRIAL STROMAL CELLS

Decidualization of estradiol (E-2)-primed human endometrial cells (HES Cs) by progesterone is associated with elevated levels of tissue facto r (TF), the primary initiator of hemostasis. Similarly, in cultured hu man HESCs, the synthetic progestin, medroxyprogesterone acetate (MPA), enhances TF protein and messenger ribonucleic acid (mRNA) levels. Alt hough ineffective alone, E-2 potentiates this progestin enhancement of TF expression by HESCs. The current study examines mechanisms underly ing MPA enhancement of TF mRNA expression in HESCs. In the presence of the transcription-blocking agent dichlororibofuranosylbenzimidazole, no significant differences were noted in the half-lives of TF mRNA iso lated from HESCs treated with E-2 alone or with E-2 plus MPA. This ind icates that MPA-enhanced TF mRNA levels do not reflect changes in the stability of the TF message. To test the effect of progestin on TF pro moter activity and to ascertain the mechanism of promoter regulation, primary or first passaged HESCs were transfected with TF promoter cons tructs spanning the regions -2106 to +121 (TFp(-2106)), -278 to +121 ( TFp(-278)), and -111 to +14 (TFp(-111)) bp upstream of the transcripti on start site. MPA was found to enhance TF transcription by 20-fold in HESCs transfected with TFp(-2106) after correcting for transfection e fficiencies with a beta-galactosidase reporter plasmid. Interestingly, levels of E-2-plus MPA-stimulated transcription were significantly in creased using TFp(-278) compared to TFp(-2106), suggesting that the re gion between -2106 and -278 bp may contain an inhibitory element. In a ddition, rates of MPA-stimulated transcription using TFp(-111) were si gnificantly reduced compared to values obtained using TFp(-2106) and w ere even further reduced compared to values obtained using TFp(-278). This suggests that regulatory elements in the -111 bp region of the TF promoter are necessary for progestin-mediated regulation of the TF ge ne in HESCs, but are not sufficient to account for maximal rates of TF gene transcription. Our results also demonstrated that induction of s teady state TF mRNA by MPA was abolished hi treating cells with E-2 pl us MPA in conjunction with the protein synthesis inhibitor cycloheximi de. In light of the absence of a complete progesterone or estrogen res ponse element in the published 5'-sequence of the TF promoter, our res ults suggest that progestin-enhanced transcription of TF mRNA in strom al cells may be mediated by an uncharacterized protein intermediate(s) .