GENE EXPRESSION PROFILES IN HUMAN IMMORTALIZED ENDOMETRIOTIC EPITHELIAL AND STROMAL CELLS: CYCLOOXYGENASE-2 REGULATES CELL PROLIFERATION, MIGRATION AND INVASION

Endometriosis is a debilitating disease affects up to 20% of reproductive age women characterized by the presence of functional endometrial glandular epithelium and stroma out side the uterine cavity. The pathophysiology of endometriosis remains as enigma in reproductive medicine. Accumulating evidence indicates that steroids, growth factors, and cytokines, and prostaglandins may promote establishment and maintenance of endometriosis. Prostaglandin E2 (PGE2) promotes cell proliferation, migration, invasion, angiogenesis, antiapoptosis, pain and immunomodulation. Cyclooxygenase (COX-2) is the rate limiting enzyme governing the biosynthesis of PGE2. COX-2 is abundantly expressed in endometriotic lesions in human. Inhibition of COX-2 decreased the size and number of the endometriotic lesions in nude mice. In this study, we used immortalized human endometriotic epithelial cells (11-Z, 12-Z, 49-Z, and 108-Z) and stromal cells (22-B) as in vitro models to unravel the molecular and cellular aspects of establishment and maintenance of endometriosis in human. We investigated: (1) expression profiles of cassette of genes involved in steroid biosynthesis, cell cycle regulation, cell migration and invasion, angiogenesis, cytokine production, and prostaglandin biosynthesis, transport and signaling; and (2) role of COX-2 in endometriotic cell proliferation, migration and invasion. Expression of genes was studied by RT-PCR. Cell proliferation was measured by Coulter counter. Cell migration and invasion were studied using matrigel assay. Expression of COX-2 protein was studied by western blot. PGE2 levels were measured by ELISA. Endometriotic epithelial and stromal cells express variety of genes. Steroid acute regulatory protein, P450 aromatase, 17b hydroxydehydrogenase-1 (17bHSD-1), and estradiol receptor alpha and beta genes are highly expressed whereas 17bHSD-2 and progesterone receptor genes are barely detectable. Cyclins A2, D1, E1 and E2 genes are abundantly expressed. Cyclins A1 and D3 genes are moderately expressed. Cyclin D2 is poorly expressed. Cyclin dependent kinases 2, 4, and 6 and their inhibitors P21 and P27 genes are abundantly expressed. Matrix metalloproteinases MMP1, MMP2 and MMP9 genes are highly expressed whereas MMP3 and MMP7 are barely detectable. Tissue inhibitors of metalloproteinases TIMP1, TIMP2, TIMP3 and TIMP4 genes are expressed at various levels. Interleukin-1 beta gene is abundantly expressed. TNF alpha is abundantly expressed only in endometriotic stromal cells. Vascular endothelial growth factor and its receptors (VEGFR1, VEGFR2, and VEGFR3) and epidermal growth factor and its receptors (EGFR1, EGFR2 and EGFR3) are expressed at various levels. COX-1 and COX-2, prostaglandin E synthase 1, 2 and 3 are abundantly expressed whereas prostaglandin dehydrogenase is undetectable that results in increased PGE2 production. PGE2 receptors EP2 and EP4 are abundantly expressed while EP1 and EP3 are barely detectable. Inhibition of COX-2 with NS-398 significantly decreased the cell proliferation (55%), migration (65%) and invasion (75%) in both epithelial and stromal endometriotic cells. In summary, these endometriotic epithelial and stromal cells can be used as ideal model to study the cellcell communications and interactions in the establishment and maintenance of endometriosis in human. Targeting PGE2 biosynthetic and signaling pathways could emerge as a potential therapy for endometriosis in human. (poster)