CHEMOKINE REGULATION IN MODELS OF ENDOMETRIOSIS

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Endometriosis is a common human gynecologic disorder associated with dysmenorrhea, pelvic pain and reduced fertility. Prevalence estimates range from 2-50%, although most scholars of endometriosis believe that it occurs in approximately 10% of reproductive aged American women. The annual United States health costs attributable to endometriosis exceed $1 billion. Recent studies suggest that endometriosis implants activate local peritoneal inflammatory responses that mediate the clinical symptoms. We hypothesize that the recruitment and subsequent accumulation of activated macrophages in the peritoneal cavity was an early and requisite step in the establishment of endometriosis implants and identified elevated concentrations of inflammatory and angiogenic cytokines (RANTES, IL-6, IL-8, VEGF) in peritoneal fluid of women with endometriosis. We propose to investigate the regulation of synthesis and secretion of one representative chemokine, RANTES, a potent chemoattractant for monocytes and T cells. We demonstrated that RANTES is localized in the stromal compartment of normal endometrium and endometriosis implants and that both mRNA and protein are expressed in endometrial stromal but not epithelial cells in vitro. In Specific Aim #1 we will use highly purified (>95%) primary stromal cell cultures to compare RANTES production in eutopic and ectopic cells from normal subjects, women with endometriosis and women with unexplained infertility. Our preliminary data indicate that RANTES protein secretion differs in the former two conditions. In Specific Aim #2, we will study the ability the natural ovarian steroid hormones (estradiol, progesterone), antagonists (tamoxifen, RU486, danazol) and other cytokines (TNF-alpha), IL-1alpha and beta, interferon-gamma) to modulate RANTES expression in vitro at the mRNA and protein levels. An expression vector containing 477 base pairs of the human RANTES gene promoter cloned upstream of a luciferase reporter will be used to map the transcriptional regulatory motifs in transiently transfected endometrial and endometriosis stromal cells. Specific Aim #3 will be executed in collaboration with Dr. Osteen using his in vivo model of human endometrium transplanted into the nude mouse peritoneal cavity. Hormones and cytokines that up- and down-regulate immunoreactive RANTES in vitro will be administered to mice bearing human endometrial implants to determine if these compounds regulate human RANTES in intact tissues in vivo. Cognate anti-hormones or cytokine neutralizing antibodies will be administered to confirm that the RANTES modulating effects are specific. It is likely that RANTES and other chemokines play early, requisite play early, requisite roles in the inflammatory process that accompanies this syndrome. These molecules should provide ideal targets for the future development of novel therapeutic antagonists for the medical treatment of endometriosis.