ALTERED ENDOMETRIAL GENE EXPRESSION IN A BABOON MODEL OF ENDOMETRIOSIS

Endometriosis is one of the most common causes of chronic pelvic pain and infertility. However, the causative factors associated with reduced fecundity have not been clearly elucidated. Due to the limitations associated with controlled studies in women with endometriosis, we have developed an induced endometriosis model in the baboon by inoculating the peritoneal cavity with menstrual endometrium on two consecutive menstrual cycles. To determine if the presence of endometriotic lesions influenced the gene expression in the eutopic endometrium, endometrial tissues were harvested at 1,3,6,9,12 and 15 months (m) from the same cohort of animals during the window of uterine receptivity. Microarray expression profiling for elucidation of molecular players participating in the maturation of the endometrium during the window of implantation was done on an Affymetrix platform. Independent hierarchical clustering analysis with all samples revealed two major dendogram branches comprised of a) 1 and 3m and b) 6m, 15m, spontaneous endometriotic samples, together with controls. A Heatmap of gene expression revealed dysregulation of many genes 1 and 3m after induction of disease, with lower levels of differential gene expression in 6m, 15m and spontaneous endometriotic samples. These data further suggest that the eutopic endometrium undergoes sequential changes that are initially characterized by a dominance of estrogen (E2) regulated genes, such as FOS, CYR61 and EMMPRIN followed by the development of progesterone (P) resistance within the endometrium and a decrease in P regulated genes such as HOXA10 and Calcitonin. Overall, our studies imply that the genes showing transient dysregulation during the initial phases of the disease are E2 regulated genes, while P regulated genes are permanently down regulated, as the disease progresses. These changes in gene expression were correlated with aberrant endometrial morphology and epigenetic changes. Increased levels of DNMT1 mRNA and protein were observed in endometria from baboons with disease compared to controls. Bisulfite sequencing revealed significantly increased levels of methylation in the F1 region within the promoter of the HOXA10 gene. MSRF identified 14 hypo- and two hypermethylated DNA fragments in baboons with endometriosis. Hypomethylation and subsequent increased levels of genes involved in cell growth and differentiation (i.e.,WNT2B) may dysregulate differentiation of the secretory endometrium of baboons with disease Furthermore, these changes in gene expression and morphology were associated with an altered steroid receptor distribution. Both estrogen receptors (alpha and beta) were significantly decreased in the stromal cells while PR levels were reduced in the glandular epithelial cells during the transition phase. Also, although PR was present in the stromal cells, this receptor was less responsive to ligand stimulation. The chaperone immunophilin FKBP52 is critical for P action and the effects of FKBP52 are primarily controlled by PRA. The absence of FKBP52 also skews the uterus towards an exaggerated E2 response during the window of receptivity. In our endometriosis model the decrease in both PR and FKBP52 is associated with the development of P resistance further validating our hypothesis that endometriosis associated infertility is a result of diminished P action in the eutopic endometrium. Using a well established non-human primate model, we have demonstrated that the presence of endometriotic lesions results in an altered eutopic endometrial environment. In many ways these changes replicate those reported for women with endometriosis. By sequential analysis of the eutopic endometrium in the same animals during the progression of the disease, we have also shown that early in the disease process there is a transitory dominance of an estrogenic phenotype, but as the disease progresses a more permanent P4 resistant phenotype results. [NIH HD 40093]