Up-regulation of Cyclooxygenase-2 Expression and Prostaglandin Synthesis in Endometrial Stromal Cells by Malignant Endometrial Epithelial Cells A PARACRINE EFFECT MEDIATED BY PROSTAGLANDIN E2 AND NUCLEAR FACTOR-κB

Abstract We investigated the regulation of prostaglandin production in normal endometrial stromal cells (ESC) by malignant endometrial epithelial cells. We found that cyclooxygenase (COX)-2 mRNA and protein levels and prostaglandin (PG)E2production in ESC were significantly increased by Ishikawa malignant endometrial epithelial cell conditioned medium (MECM). By using transient transfection assays, we found that the −360/−218-bp region of the COX-2 promoter gene was critical for MECM induction of promoter activity. This MECM-responsive region contained a variant nuclear factor (NF)-κB site at −222 to −213 that, when mutated, completely abolished COX-2 promoter activation by MECM. Employing electrophoretic mobility shift assays, we further demonstrated that binding of NF-κB p65 to this NF-κB-binding site is, in part, responsible for the COX-2 promoter activation by MECM. To investigate further the potential effects of MECM onCOX-2 mRNA stability, ESC were treated with MECM in the absence or presence of actinomycin D, a general transcription inhibitor. We found that MECM significantly increased COX-2mRNA stability. Intriguingly, we found that PGE2 was one of the major factors in MECM, which was responsible for up-regulating COX-2 expression in ESC. ECC-1 and HEC-1A malignant endometrial epithelial cell lines also produced significantly increased quantities of PGE2. In conclusion, malignant endometrial epithelial cells secrete PGE2 that induces COX-2 expression in normal endometrial stromal cells in a paracrine fashion through activation of transcription and stabilization of COX-2mRNA.