The Protein Kinase A Pathway-Regulated Transcriptome of Endometrial Stromal Fibroblasts Reveals Compromised Differentiation and Persistent Proliferative Potential in Endometriosis

Abstract

Intrinsic abnormalities in transplanted eutopic endometrium are believed to contribute to the pathogenesis of pelvic endometriosis. Herein we investigated transcriptomic differences in human endometrial stromal fibroblasts (hESFs) from women with (hESF_endo) vs. without (hESF_nonendo) endometriosis, in response to activation of the protein kinase A (PKA) pathway with 8-bromoadenosine-cAMP (8-Br-cAMP). hESF_nonendo (n = 4) and hESF_endo (n = 4) were isolated from eutopic endometrium and treated ± 0.5 mm 8-Br-cAMP for 96 h. Purified total RNA was subjected to microarray analysis using the whole-genome Gene 1.0 ST Affymetrix platform. A total of 691 genes were regulated in cAMP-treated hESF_nonendovs. 158 genes in hESF_endo, suggesting a blunted response to cAMP/PKA pathway activation in women with disease. Real-time PCR and ELISA validated the decreased expression of decidualization markers in hESF_endo compared with hESF_nonendo. In the absence of disease, 8-Br-cAMP down-regulated progression through the cell cycle via a decrease in cyclin D1, cyclin-dependent kinase 6, and cell division cycle 2 and an increase in cyclin-dependent kinase inhibitor 1A. However, cell cycle components in hESF_endo were not responsive to 8-Br-cAMP, resulting in persistence of a proliferative phenotype. hESF_endo treated with 8-Br-cAMP exhibited altered expression of immune response, extracellular matrix, cytoskeleton, and apoptosis genes. Changes in phosphodiesterase expression and activity were not different among experimental groups. These data support that eutopic hESF_endo with increased proliferative potential can seed the pelvic cavity via retrograde menstruation and promote establishment, survival, and proliferation of endometriosis lesions, independent of hydrolysis of cAMP and likely due to an inherent abnormality in the PKA pathway.