Endometriosis Alters the Peripheral and Endometrial Expression of Regulatory T Cells in a Non-Human Primate.

Endometriosis is a gynecological condition that is characterized by extreme abdominal pain and also decreased fertility. Regulatory T cells (Tregs) have immunosuppressive activity critical for embryonic implantation and likewise the acceptance of tissue engraftment. We hypothesize that endometriosis decreases the peripheral and endometrial Treg profile whereas ectopic lesions have increased Treg localization. This study characterized Treg expression in the peripheral circulation, uterine endometrium and ectopic lesions in an induced model of endometriosis in the non-human primate (Papio Anubis). Peripheral blood and endometrium were obtained at different timepoints during the menstrual cycle prior to induction of disease. The timepoints were defined as follows: mense (day (d) 1-4 post mense (PM)), late proliferative (d9-12PM), mid secretory (d9-11 post ovulation (PO)) and late secretory (d12-16 PO). Animals were randomly assigned to control (n=5) or diseased (n=16) treatment groups. Endometriosis was induced by intraperitoneal injection of autologous menstrual tissue for 2 consecutive months on day 1 or 2 of mense. Peripheral blood and endometrial tissue were collected at d9-11PO at 1, 3, 6, 9, 12 and 15 months post-induction of disease. Lymphocytes were isolated using Ficoll Paque density separation of blood and then labeled with CD4/CD25/FOXP3 specific antibodies to identify 3 populations of Tregs by FACS analysis. The proportions of Treg populations of total CD4 positive cells were analyzed using non-parametric statistical analysis to determine experimental significance. Endometrium was obtained by endometriectomy at the time of laparotomy for each surgical timepoint and processed for RNA or IHC. Ectopic lesions were excised during necropsy at the 15 mth timepoint and processed for RNA of IHC. FOXP3 (marker of Tregs) and CXCL12 (chemoattractant for Tregs) RNA was analyzed by qRT-PCR and relative fold differences were determined by non-parametric statistical analysis for each gene. Endometrium and ectopic lesions were analyzed for Treg localization by IHC using a FOXP3 antibody. In control animals the proportion of peripheral natural Tregs (nTregs) was reduced during the mid and late secretory stage of the menstrual cycle. In addition, endometrial CXCL12 gene expression was highest during the proliferative phase. Induction of disease decreased peripheral Treg expression at early timepoints and remained low throughout the timecourse. FOXP3 gene expression and Treg localization were also decreased in eutopic endometrium, whereas these parameters were increased in ectopic lesions. We speculate that the decrease in Treg localization in the endometrium may be due to the decrease in endometrial CXCL12 gene expression (p<0.05) early in disease. CXCL12 is important for recruiting Tregs to the endometrium from the peripheral circulation. Endometriosis decreases the peripheral population of Tregs thereby reducing the ability of these cells to be recruited to the endometrium. However ectopic lesions have enhanced expression of Tregs for suppression of immune clearance. The reduction in the number of the peripheral Tregs may be a causative factor of endometriosis associated infertility while the increase in ectopic Treg expression may aid in lesion development. These data indicate that endometriosis disrupts Treg recruitment in both eutopic and ectopic endometrium. (Supported by SCCPRR U54HD20093 and U54HD40093). (platform)