Endometrial inflammasome activation accompanies menstruation and may have implications for systemic inflammatory events of the menstrual cycle

STUDY QUESTION Does NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3) inflammasome activation within decidualized endometrial stromal cells accompany menstruation and is this reflected systemically? SUMMARY ANSWER Components of the NLRP3 inflammasome immunolocalize to decidualized endometrial stromal cells immediately prior to menstruation, and are activated in an in vitro model of menstruation, as evidenced by downstream interleukin (IL)-1beta and IL-18 release, this being reflected systemically in vivo. WHAT IS KNOWN ALREADY Menstruation is a highly inflammatory event associated with activation of NFκB (nuclear factor kappa-light-chain-enhancer of activated B cells), local release of chemokines and cytokines and inflammatory leukocyte influx. Systemically, chemokines and cytokines fluctuate across the menstrual cycle. STUDY DESIGN, SIZE, DURATION This study examined the NLRP3 inflammasome and activation of downstream IL-1beta and IL-18 in endometrial tissues from women of known fertility (≥1 previous parous pregnancy) across the menstrual cycle (n ≥ 8 per cycle phase), serum from women during the proliferative, secretory and menstrual phases (≥9 per cycle phase) of the cycle and menstrual fluid collected on Day 2 of menses (n = 18). Endometrial stromal cells isolated from endometrial tissue biopsies (n = 10 in total) were used for an in vitro model of pre-menstrual hormone withdrawal. PARTICIPANTS/MATERIALS, SETTING, METHODS Expression and localization of components of the NLRP3 inflammasome (NLRP3 & apoptosis-associated speck–caspase recruit domain [ASC]) in endometrial tissues was performed by immunohistochemistry. Unbiased digital quantification of immunohistochemical staining allowed determination of different patterns of expression across the menstrual cycle. Serum from women across the menstrual cycle was examined for IL-1beta and IL-18 concentrations by ELISA. An in vitro model of hormone withdrawal from estrogen/progestin decidualized endometrial stromal cells was used to more carefully examine activation of the NLRP3 inflammasome. Endometrial stromal cells isolated from endometrial tissue biopsies (n = 10) were treated with estrogen/medroxyprogesterone acetate for 12 days to induce decidualization (assessed by release of prolactin) followed by withdrawal of steroid hormone support. Activation of NLRP3, & ASC in these cells was examined on Days 0–3 after hormone withdrawal by Western immunoblotting. Release of IL-1beta and IL-18 examined during decidualization and across the same time course of hormone withdrawal by ELISA. Specific involvement of NLRP3 inflammasome activation in IL-1beta and IL-18 release after hormone withdrawal was investigated via application of the NLRP3 inflammasome inhibitor MCC950 at the time of hormone withdrawal. MAIN RESULTS AND THE ROLE OF CHANCE Critical components of the NLRP3 inflammasome (NLRP3, ASC) were increased in menstrual phase endometrial tissues versus early secretory phase tissues (P LARGE SCALE DATA N/A. LIMITATIONS, REASONS FOR CAUTION This study uses descriptive and semi-quantitative measures of NLRP3 inflammasome activation within endometrial tissues. Further, the in vitro model of pre-menstrual hormone withdrawal may not accurately recapitulate the in vivo environment as only one cell type is present and medroxyprogesterone acetate replaced natural progesterone due to its longer stability. WIDER IMPLICATIONS OF THE FINDINGS We provide novel evidence that the NLRP3 inflammasome is activated within decidualized endometrial stromal cells immediately prior to menses and that local activation of the inflammasome within the endometrium appears to be reflected systemically in by activation of downstream IL-1beta and IL-18. Given the prevalence of menstrual disorders associated with inflammation including dysmenorrhoea and aspects of pre-menstrual syndrome, the inflammasome could be a novel target for ameliorating such burdens. STUDY FUNDING/COMPETING INTEREST(S) The authors have no competing interests. J.E. was supported by a Fielding Foundation fellowship, NHMRC project grants (#1139489 and #1141946) and The Hudson Institute of Medical Research. L.A.S. was supported by The Hudson Institute of Medical Research and J.H. by an Australian Government Research Training Program Scholarship. We acknowledge the Victorian Government’s Operating Infrastructure funding to the Hudson Institute. TRIAL REGISTRATION NUMBER N/A