We showed that decidualized stromal cells of luteal phase and pregnant human endometrium express tissue factor (TF), the primary initiator of hemostasis, thereby suggesting a mechanism by which perivascular decidual cells can mitigate the risk of hemorrhage during endovascular trophoblast invasion. Progestins enhanced TF mRNA and protein levels in monolayers of human endometrial stromal cells (HESCs), with estradiol (E2) + progestin, further enhancing TF levels despite a lack of response to E2 alone. This differential ovarian steroid response has been found for several decidualization markers. Further studies with cultured HESCs established that elevated TF levels are mediated by the progesterone receptor and are maintained for weeks in response to E2 plus progestin, thus simulating the chronic upregulation of TF levels observed in decidualized HESCs in vivo. Recent studies revealed that elevated TF expression during in vitro decidualization of HESCs involved both the EGFR and progesterone receptor. Thus, enhancement of TF mRNA and protein levels in the HESCs required co-incubation with a progestin (MPA) and an EGFR agonist such as EGF or TGF-alpha. In correspondence with co-elevation of EGFR and TF in decidualized HESCs in sections of luteal phase and pregnant endometrium, EGFR levels proved to be progestin-enhanced in the cultured HESCs. We established that progestin-enhanced TF expression in HESCs was trancriptionally regulated, then evaluated the relative roles of SP and EGR-1 sites on the TF promoter in regulating this expression. Transient transfections with a series of promoter constructs containing overlapping SP and EGR-1 sites and with constructs in which the EGR-1 and SP sites were systematically inactivated by site-directed mutagenesis established the dominance of SP sites in both basal and progestin-enhanced TF transcriptional activity. Additional experiments involving transient transfections with SPloverexpressing vectors and with a specific blocker of if Sp1 binding to its corresponding GC box specified the importance of the Sp1 transcription factor. These results were further validated by immunostaining, which revealed that the ratio of Sp1 to Sp3 increased during progestin-regulated decidualization of HESCs in vitro and in vivo. The absence of canonical estrogen and progesterone response elements from either the TF or Sp1 gene promoters suggests that the EGFR may help to mediate progestin-enhanced TF expression during decidualization of HESCs.