Uterine natural killer cell progenitor populations predict successful implantation in women with endometriosis-associated infertility.

PROBLEM: Uterine natural killer (uNK) cells play a critical role early in gestation. As we previously identified altered uNK cell development in endometriosis-associated infertility, we herein sought to characterize natural killer (NK) cell profiles in endometriosis that may predict embryo implantation. METHOD OF STUDY: Study participants had a surgical diagnosis of endometriosis-associated infertility. Endometrial tissue and peripheral blood were obtained from 58 women. Thirty-three patients underwent artificial reproductive technology (IVF, ICSI, or IUI) within a mean of 9.5 months of surgery. NK and hematopoietic progenitor cells from endometrium and blood were analyzed by flow cytometry. Successful implantation was defined as a positive pregnancy test. RESULTS: In successful implantation, populations of endometrial CD34+ hematopoietic stem cells were higher (3.97% vs 0.69%; P < .0004), and coexpression of NK cell marker CD56 was increased (81.1% vs 60.9%; P < .034) compared with patients who had failed implantation. In contrast, levels of blood NK progenitors were similar in both groups. CONCLUSION: Our study revealed that uterine NK progenitor cell populations are markedly different in patients with endometriosis who proceed to successful or failed embryo implantation and may define a novel predictor of implantation success. Our findings also highlight the fundamental differences inherent in NK cell repertoires between blood and uterine compartments.

Steroids Regulate CXCL4 in the Human Endometrium During Menstruation to Enable Efficient Endometrial Repair.

Context: Repair of the endometrial surface at menstruation must be efficient to minimize blood loss and optimize reproductive function. The mechanism and regulation of endometrial repair remain undefined. Objective: To determine the presence/regulation of CXCL4 in the human endometrium as a putative repair factor at menses. Patients/Setting: Endometrial tissue was collected throughout the menstrual cycle from healthy women attending the gynecology department. Menstrual blood loss was objectively measured in a subset, and heavy menstrual bleeding (HMB) was defined as >80 mL per cycle. Monocytes were isolated from peripheral blood. Design: CXCL4 messenger RNA (mRNA) and protein were identified by quantitative reverse transcription polymerase chain reaction and immunohistochemistry. The function/regulation of endometrial CXCL4 was explored by in vitro cell culture. Results: CXCL4 mRNA concentrations were significantly increased during menstruation. Intense staining for CXCL4 was detected in late secretory and menstrual tissue, localized to stromal, epithelial and endothelial cells. Colocalization identified positive staining in CD68+ macrophages. Treatment of human endometrial stromal and endothelial cells (hESCs and HEECs, respectively) with steroids revealed differential regulation of CXCL4. Progesterone withdrawal resulted in significant increases in CXCL4 mRNA and protein in hESCs, whereas cortisol significantly increased CXCL4 in HEECs. In women with HMB, CXCL4 was reduced in endothelial cells during the menstrual phase compared with women with normal menstrual bleeding. Cortisol-exposed macrophages displayed increased chemotaxis toward CXCL4 compared with macrophages incubated with estrogen or progesterone. Conclusions: These data implicate CXCL4 in endometrial repair after menses. Reduced cortisol at the time of menses may contribute to delayed endometrial repair and HMB, in part by mechanisms involving aberrant expression of CXCL4.

Increased uNK Progenitor Cells in Women With Endometriosis and Infertility are Associated With Low Levels of Endometrial Stem Cell Factor.

PROBLEM: Uterine natural killer (uNK) cells play a significant role in successful human pregnancy. Having previously demonstrated uNK cell progenitors in human endometrium, we hypothesized that abnormal uNK cell maturation contributes to infertility in women with endometriosis. We aimed to characterize uNK cells at different developmental stages in women with and without endometriosis and to investigate possible mechanisms to explain any differences. METHOD OF STUDY: We characterized uNK cell development in women with and without endometriosis using flow cytometry, protein array and in vitro experiments. RESULTS: We found increased proportions of uNK cells at developmental stages 1 and 2 in endometrium from women with endometriosis (n = 36; mean = 21.2%) when compared with healthy fertile women (n = 9; mean = 7.0%). Protein array analysis revealed significantly lower levels of stem cell factor (SCF) in the eutopic endometrium of women with endometriosis when compared to healthy women. Addition of SCF to endometrial progenitor cells in vitro restored uNK cell maturation. CONCLUSION: We have shown that women with endometriosis have low levels of endometrial SCF, which we hypothesize contributes to abnormal maturation of local uNK cell populations. This defect may also compromise embryo implantation and hence contribute to endometriosis-associated infertility. SCF replacement may be a new therapeutic approach.

Cortisol regulates the paracrine action of macrophages by inducing vasoactive gene expression in endometrial cells.

The human endometrium undergoes inflammation and tissue repair during menstruation. We hypothesized that the local availability of bioactive glucocorticoids plays an important role in immune cell-vascular cell interactions in endometrium during tissue repair at menstruation, acting either directly or indirectly via tissue resident macrophages. We sought to determine whether endometrial macrophages are direct targets for glucocorticoids; whether cortisol-treated macrophages have a paracrine effect on angiogenic gene expression by endometrial endothelial cells; and whether endometrial macrophages express angiogenic factors. Human endometrium (n = 41) was collected with ethical approval and subject consent. Donor peripheral blood monocyte-derived macrophages were treated with estradiol, progesterone, or cortisol. The effect of peripheral blood monocyte-derived macrophage secretory products on the expression of angiogenic RNAs by endothelial cells was examined. Immunofluorescence was used to examine localization in macrophages and other endometrial cell types across the menstrual cycle. Endometrial macrophages express the glucocorticoid receptor. In vitro culture with supernatants from cortisol-treated peripheral blood monocyte-derived macrophages resulted in altered endometrial endothelial cell expression of the angiogenic genes, CXCL2, CXCL8, CTGF, and VEGFC These data highlight the importance of local cortisol in regulating paracrine actions of macrophages in the endometrium. CXCL2 and CXCL8 were detected in endometrial macrophages in situ. The expression of these factors was highest in the endometrium during the menstrual phase, consistent with these factors having a role in endometrial repair. Our data have indicated that activation of macrophages with glucocorticoids might have paracrine effects by increasing angiogenic factor expression by endometrial endothelial cells. This might reflect possible roles for macrophages in endometrial repair of the vascular bed after menstruation.

Natural Killer Cells: Key Players in Endometriosis.

Endometriosis affects more than 10% of women, causing significant pain and morbidity. It is also a significant cause of infertility. The aetiology of the disease remains an enigma, and the mechanisms responsible for the associated infertility are unclear. A role for immune cells in endometriosis has been postulated, with attention directed towards natural killer (NK) cells and macrophages. NK cells kill tumours and infected cells but also have roles in tissue remodelling in several organs including the uterus and are key to successful pregnancy. Here, we explore evidence (from peer-reviewed published articles) of phenotypic and functional abnormalities in NK cell subpopulations of women with endometriosis. It is clear that peripheral blood NK cells and peritoneal NK cells have reduced cytotoxic function in women with endometriosis. Uterine NK cells have a vital role in infertility, but very little research has been carried out in this area. We propose that abnormal u NK cell activity may contribute to the pathogenesis of endometriosis and its associated infertility and that future research should focus on this complex area.

The importance of the macrophage within the human endometrium.

The human endometrium is exposed to cyclical fluctuations of ovarian-derived sex steroids resulting in proliferation, differentiation (decidualization), and menstruation. An influx of leukocytes (up to 15% macrophages) occurs during the latter stages of the menstrual cycle, including menses. We believe the endometrial macrophage is likely to play an important role during the menstrual cycle, especially in the context of tissue degradation (menstruation), which requires regulated repair, regeneration, and phagocytic clearance of endometrial tissue debris to re-establish tissue integrity in preparation for fertility. The phenotype and regulation of the macrophage within the endometrium during the menstrual cycle and interactions with other cell types that constitute the endometrium are currently unknown and are important areas of study. Understanding the many roles of the endometrial macrophage is crucial to our body of knowledge concerning functionality of the endometrium as well as to our understanding of disorders of the menstrual cycle, which have major impacts on the health and well-being of women.