Expression of glucocorticoid and androgen receptors in bone marrow-derived hematopoietic and nonhematopoietic murine endometrial cells.

OBJECTIVE: To determine whether bone marrow (BM)-derived cells engrafting the murine endometrium express the glucocorticoid receptor (GR) and androgen receptor (AR). Recent data demonstrate that BM is a long-term source of multiple hematopoietic and nonhematopoietic endometrial cell types. Important roles for glucocorticoids and androgens in regulating endometrial functions, including decidualization and early embryo attachment/invasion, have very recently emerged. Whether endometrial cells of BM origin express glucocorticoid or ARs has not been previously studied. DESIGN: Animal study. SETTING: Basic science laboratory. ANIMAL(S): Wild-type C57BL/6J male mice expressing enhanced green fluorescent protein (GFP) and syngeneic wild-type C57BL/6J female mice aged 6-9 weeks. INTERVENTION(S): Murine bone marrow transplant. MAIN OUTCOME MEASURE(S): Bone marrow cells were harvested from adult wild-type C57BL/6 mice and subjected to flow cytometry to identify the percentage of hematopoietic and nonhematopoietic cells expressing GR or AR. Uterine tissue sections from lethally irradiated syngeneic adult female C57BL/6 mice that had been recipients of BM transplants from adult male transgenic donor mice ubiquitously expressing GFP were studied. Immunohistochemistry was performed in the uterine tissue sections of the recipient mice at 5, 9, and 12 months after transplant using specific anti-GR, anti-AR, anti-GFP, anti-CD45 (pan leukocyte marker), and anti-F4/80 (murine macrophage marker) primary antibodies. Confocal laser microscopy was used to localize and quantitate BM-derived (GFP+) cell types in the endometrial stromal and epithelial compartments and determine whether BM-derived cell types in the murine endometrium express GR or AR. RESULT(S): Hematopoietic cells comprised 93.6%-96.6% of all cells in the BM, of which 98.1% ± 0.2% expressed GR and 92.2% ± 4.4% expressed AR. Nonhematopoietic cells comprised 0.4%-1.3% of BM, of which 52.8% ± 5.9% expressed GR and 48.9% ± 3.4% expressed AR. After BM transplant, the proportion of cells originating from BM in the endometrial stromal compartment increased over time, reaching 13.5% ± 2.3% at 12 months after transplant. In the epithelial compartments, <1% of the cells were of BM origin at 12 months after transplant. Most (60%-72%) GR+ and/or AR+ BM-derived cells in the stroma were hematopoietic (CD45+) cells, of which 37%-51% were macrophages. Nonetheless, 28%-33% of GR+ cells, and 28%-40% of AR+ BM-derived cells, were nonhematopoietic (CD45-) stromal cells of BM origin. CONCLUSION(S): A substantial number of BM-derived cells express GR and AR, suggesting a role for these cells in both glucocorticoid-regulated and androgen-regulated endometrial functions, such as proliferation and/or decidualization.

The role of mesenchymal-epithelial transition in endometrial function.

BACKGROUND: The human uterine endometrium undergoes significant remodeling and regeneration on a rapid and repeated basis, after parturition, menstruation, and in some cases, injury. The ability of the adult endometrium to undergo cyclic regeneration and differentiation/decidualization is essential for successful human reproduction. Multiple key physiologic functions of the endometrium require the cells of this tissue to transition between mesenchymal and epithelial phenotypes, processes known as mesenchymal-epithelial transition (MET) and epithelial-mesenchymal transition (EMT). Although MET/EMT processes have been widely characterized in embryonic development and in the context of malignancy, mounting evidence demonstrates the importance of MET/EMT in allowing the endometrium the phenotypic and functional flexibility necessary for successful decidualization, regeneration/re-epithelialization and embryo implantation. OBJECTIVE AND RATIONALE: The objective of this review is to provide a comprehensive summary of the observations concerning MET and EMT and their regulation in physiologic uterine functions, specifically in the context of endometrial regeneration, decidualization and embryo implantation. SEARCH METHODS: Using variations of the search terms 'mesenchymal-epithelial transition', 'mesenchymal-epithelial transformation', 'epithelial-mesenchymal transition', 'epithelial-mesenchymal transformation', 'uterus', 'endometrial regeneration', 'endometrial decidualization', 'embryo implantation', a search of the published literature between 1970 and 2018 was conducted using the PubMed database. In addition, we searched the reference lists of all publications included in this review for additional relevant original studies. OUTCOMES: Multiple studies demonstrate that endometrial stromal cells contribute to the regeneration of both the stromal and epithelial cell compartments of the uterus, implicating a role for MET in mechanisms responsible for endometrial regeneration and re-epithelialization. During decidualization, endometrial stromal cells undergo morphologic and functional changes consistent with MET in order to accommodate embryo implantation. Under the influence of estradiol, progesterone and multiple other factors, endometrial stromal fibroblasts acquire epithelioid characteristics, such as expanded cytoplasm and rough endoplasmic reticulum required for greater secretory capacity, rounded nuclei, increased expression of junctional proteins which allow for increased cell-cell communication, and a reorganized actin cytoskeleton. During embryo implantation, in response to both maternal and embryonic-derived signals, the maternal luminal epithelium as well as the decidualized stromal cells acquire the mesenchymal characteristics of increased migration/motility, thus undergoing EMT in order to accommodate the invading trophoblast. WIDER IMPLICATIONS: Overall, the findings support important roles for MET/EMT in multiple endometrial functions required for successful reproduction. The endometrium may be considered a unique wound healing model, given its ability to repeatedly undergo repair without scarring or loss of function. Future studies to elucidate how MET/EMT mechanisms may contribute to scar-free endometrial repair will have considerable potential to advance studies of wound healing mechanisms in other tissues.

Association of Cervical Microglandular Hyperplasia With Exogenous Progestin Exposure.

OBJECTIVES: Although microglandular hyperplasia is a benign endocervical lesion, it may raise concern of malignancy in an inexperienced pathologist, because clinicians may not be familiar with it on a report. It has been reported to occur in association with progestational stimulation, but this has not been well studied. The aim of the study was to evaluate this potential association. MATERIALS AND METHODS: This was an observational case control study of patients seen from January 01, 1991 to November 01, 2014 at the University Hospital, Newark, New Jersey. Cases of microglandular hyperplasia and controls were identified from the files of the Department of Pathology. Controls were cases of endocervical curettages for whom no microglandular hyperplasia was detected. Medical records were reviewed for evidence of exogenous progestin exposure within the previous 6 months. RESULTS: Eighty-nine cases of microglandular hyperplasia and 97 controls were identified. Of the cases of microglandular hyperplasia, 26 (29.2%) of 89 cases had exposure to progestational medication, significantly greater than that of controls (10/97, 10.3%; p = .0014). CONCLUSIONS: Our data show that a significantly higher percentage of women with microglandular hyperplasia had progestin exposure than women who do not exhibit this benign lesion. Clinicians and pathologists should be aware of this association, as well as the benign nature of the lesion.

Myometrial artery calcifications and aging.

OBJECTIVE: This study aims to determine whether myometrial artery calcifications increase with age and whether uterine sections are an appropriate model for studying vascular aging. METHODS: An observational study of 172 women (aged 45 y or older) who underwent hysterectomy for benign indications at the University Hospital (Newark, NJ) between July 1, 2009 and June 1, 2012 was performed. Women with a history of malignancy, undocumented last menstrual period, or unavailable uterine tissue slides were excluded. H&E-stained uterine sections were evaluated for myometrial artery calcifications (defined as the presence of acellular densely basophilic material within the media of vessels) by a single pathologist in a blinded manner. RESULTS: Between July 1, 2009 and June 1, 2012, hysterectomies were performed on 441 women, 172 of whom met inclusion criteria. Seventeen women (9.9%) had myometrial artery calcifications detectable on H&E-stained tissue sections. None of 84 women aged 45 to 49 years, 2 of 51 women (3.9%) aged 50 to 59 years (aged 56 and 58 y), 10 of 27 women (37%) aged 60 to 69 years, and 5 of 10 women (50%) aged 70 to 81 years had myometrial artery calcifications. The prevalence of myometrial artery calcifications significantly increased with advancing age (P = 0.022). CONCLUSIONS: Myometrial artery calcifications increase with advancing age. Histological sections of uterine tissue from hysterectomy specimens seem to be a useful model for evaluating vascular aging markers.

Effect of single embryo transfer on the risk of preterm birth associated with in vitro fertilization.

PURPOSE: To determine whether elective single embryo transfer (eSET) reduces the risk of preterm delivery associated with in vitro fertilization (IVF). METHODS: This is an observational study of 3125 eSET cycles performed from 2008 to 2009 and reported to the Society for Assisted Reproductive Technology (SART) database. Preterm delivery rates were compared to the overall preterm delivery rate among all patients undergoing IVF over the same time period. RESULTS: The 3125 eSET cycles resulted in 1507 live births (live birth rate 48.2 %) Among these deliveries were 27 twins (1.8 %) and one set of triplets (0.07 %). The overall preterm delivery rate (20-37 weeks gestation) following eSET was 17.6 % (269/1527). This is significantly greater than the preterm birth rate for all patients undergoing IVF over the same time period (12 %, P < 0.001). CONCLUSIONS: Elective single embryo transfer does not reduce the risk of preterm delivery associated with in vitro fertilization (IVF).

Cervical conization for cervical intraepithelial neoplasia (CIN) 2 and 3 in HIV-positive women: a case-control study.

OBJECTIVE: The purposes of the study were to investigate the outcomes of cervical conization for cervical intraepithelial neoplasia (CIN) 2 and 3 in HIV-positive women and age-matched HIV-negative controls and to determine whether positive margin, positive endocervical curettage, CD4 count, or viral load was associated with the persistence of CIN 2,3 or residual CIN 2,3 on the specimen from repeat excision procedure or hysterectomy. MATERIALS AND METHODS: HIV-positive women and HIV-negative controls with CIN 2,3 on cervical conization were enrolled in the study. Patients who underwent repeat conization or hysterectomy were identified, and the specimens were evaluated for residual CIN 2,3. CD4 count and viral load within 8 weeks of procedure were analyzed. RESULTS: A total of 44 patients and 44 age-matched controls were identified. Persistent CIN 2,3 was diagnosed in 28 HIV-positive (63.6%) and 14 HIV-negative patients (31.8%; odds ratio [OR] = 4.7, 95% confidence interval [CI] = 1.9-11.5, p < .001). In HIV-positive women, a positive margin was associated with a higher persistence rate after cervical conization (OR = 5.3, 95% CI = 1.17-24.14, p = .03). In HIV-negative patients, positive endocervical curettage was associated with a higher persistence rate after conization (OR = 12, 95% CI = 2.24-64.23, p = .004). Of HIV-positive women, 75% had residual CIN 2,3 on the specimen from repeat procedure compared to 45.2% of controls (OR = 3.6, 95% CI = 1.3-10.6, p = .018). CD4 count or viral load was not associated with the rate of residual disease or persistence rate after cervical conization, but the lowest OR that the sample size allowed to assess with 90% power was 5.02. CONCLUSIONS: HIV-positive women have a higher rate of residual disease and higher persistence rate after conization for CIN 2,3 than age-matched HIV-negative controls.

Experimental evidence for bone marrow as a source of nonhematopoietic endometrial stromal and epithelial compartment cells in a murine model.

Human endometrium has the remarkable ability to regenerate all cellular compartments with every menstrual cycle; the cellular source remains unknown. The objective of the present study was to determine whether the bone marrow (BM) is a source of multiple endometrial cell types using a murine BM transplant model. BM cells were harvested from transgenic donor mice that ubiquitously express green fluorescent protein (GFP) and were injected into lethally irradiated, syngeneic female recipient mice. Recipients with successful hematopoietic reconstitution were sacrificed at 3, 5, 9, and 12 mo posttransplant, after which hysterectomy was performed. Numbers of GFP-positive, CD45-positive, and CD45-negative cells in the endometrial stromal and epithelial compartments were determined. In the stromal compartment, BM-derived cells (BMDCs) were detectable as early as 3 mo posttransplant, and the BM remained a long-term contributor of nonhematopoietic endometrial cells. Nonhematopoietic endometrial cells comprised 47.3%-72.2% of total BMDCs in the stromal compartment at 12 mo posttransplant. In contrast, BMDCs were not detected in the glandular or luminal epithelial compartments until 12 mo posttransplant. These data demonstrate that the BM is a significant source of nonhematopoietic endometrial stromal compartment cells and contributes to a much lesser extent to the epithelial compartments. That BM is a source of nonhematopoietic cells in the endometrial stromal and epithelial compartments provides a potential mechanism by which monthly regeneration of the endometrium may occur.

Reproductive tract actions of relaxin in models of human pregnancy.

Elucidating the role(s) of relaxin in women has been greatly hampered by its species specificity. Suitable experimental models of relaxin action in women are limited. We established a non-human primate model of early human pregnancy to study the effects of relaxin in vivo and used three well characterized in vitro models of human endometrial function for study of mechanisms involved. Results from these studies clearly demonstrate that relaxin is an import ant factor in human endometrium which causes accommodation to and maintenance of early pregnancy and uses multiple physiological and biochemical mediators.

Endometrial stem cells and reproduction.

Abnormal endometrial function remains a significant cause of implantation failure, recurrent pregnancy loss, and other pathologies responsible for female infertility. The development of novel therapies to treat infertility due to endometrial dysfunction requires an understanding of the latest advancements in endometrial cell biology, such as the role of endometrial stem cells. The remarkable regenerative capacity of the human endometrium is absolutely essential for successful reproduction and likely requires a population of stem cells in the endometrium. The purpose of this review is to provide an introduction to some of the newest concepts in endometrial stem cell biology.

Collagen expression in the pregnant human cervix is decreased with labor.

OBJECTIVE: The current study tested the hypothesis that collagen content in the pregnant cervix decreases with labor, using morphologically preserved specimens, avoiding limitations of earlier studies. Collagen abundance remote from pregnancy was also evaluated. MATERIALS AND METHODS: Histologic sections of postpartum cervix obtained from 22 cases of total hysterectomy performed immediately after delivery: 13 cases performed after delivery with no labor and 9 cases in which labor had ensued before delivery. Cervices from 10 nonpregnant uteri served as additional controls. Sections were stained, and quantitative histomorphometric assessment of relative collagen abundance was performed using computer-assisted image analysis. Data were assessed for differences using rank sum tests. Relationships between cervical collagen abundance and age, parity, ethnicity, or mode of delivery were also assessed. RESULTS: Quantitative assessment of collagen abundance in trichrome-stained cervical sections revealed significantly decreased cervical collagen expression in sections from pregnant uteri. Mean percent collagen was 73.5% ± 3.5% (±SEM) in cervices from nonpregnant uteri (n = 10) and 21.5% ± 2.2% in cervices from pregnant uteri (n = 22, p < .0001). Cervical collagen content was significantly lower (p = .04) in cervices from cases in which labor had ensued before delivery (mean percent collagen = 16.1% ± 3.4%, n = 9) than in those in which delivery occurred with no labor (25.3% ± 2.3%, n = 13). No relationships between collagen expression and age, parity, ethnicity, or mode of delivery were observed. CONCLUSIONS: Collagen expression seems to be reduced in the postpartum cervix, particularly after labor has ensued.

Is the diagnosis of atypical glandular cells of undetermined significance associated with greater risk at an inner city hospital?

OBJECTIVE: To test the hypothesis that the high-risk patients at an inner city hospital with atypical glandular cells of undetermined significance (AGC) on their Pap smears have a higher rate of underlying significant pathology than that reported in published data. STUDY DESIGN: This was an Institutional Review Board-approved retrospective review of all AGC Pap smears performed at University Hospital, Newark, New Jersey, between January 1, 2001, and July 30, 2008. We defined significant pathology as cervical intraepithelial neoplasia 2 (CIN 2) or greater, endocervical adenocarcinoma in situ or greater, or simple hyperplasia or greater of the endometrium. RESULTS: Medical records of 126 patients were reviewed. Forty did not meet inclusion criteria; 86 patients were included in the analysis. Thirty of the 86 (34.9%) patients were found to have significant pathology. CONCLUSION: Patients with AGC Pap results at our inner city hospital have a high risk for underlying significant pathology.

Body mass index of patients with endometrial hyperplasia: comparison to patients with proliferative endometrium and abnormal bleeding.

OBJECTIVE: Endometrial hyperplasia is a known risk factor for the development of endometrial cancer, particularly atypical hyperplasia, with a subsequent risk of up to 30%. Of the known risk factors for endometrial hyperplasia, obesity is the most preventable, but there is a paucity of data addressing the association. We tested the hypothesis that patients with endometrial hyperplasia have a higher body mass index (BMI) than patients with abnormal bleeding who are found to have proliferative endometrium. STUDY DESIGN: This was an Institutional Review Board-approved retrospective study using University Hospital Department of Pathology records. All patients who had endometrial sampling performed between January 1, 2001, and July 30, 2008, were included. The experimental group consisted of patients with endometrial hyperplasia including simple, complex and atypical hyperplasia. The control group consisted of patients who underwent endometrial sampling for abnormal bleeding during the same time period and were diagnosed with proliferative endometrium. BMI was calculated based on documented height and weight within 30 days of endometrial sampling. RESULTS: Forty-two patients with hyperplasia and 103 patients with proliferative endometrium met inclusion criteria, including documented height and weight and nonexposure to hormones. The median BMI in the hyperplasia group was 38 kg/m2 (95% CI 34.8-42.4) and 30 kg/m2 (95% CI 29.9-33.3) in the proliferative group (p < 0.0001). CONCLUSION: These data suggest that higher BMI is associated with endometrial hyperplasia as compared to women with lower BMIs and abnormal bleeding.

Endometrial expression of relaxin and relaxin receptor in endometriosis.

Our studies demonstrate significantly lower expression of relaxin and its receptor in ectopic endometriotic tissues than their expression in eutopic endometrium and in endometrium from normal controls. These data suggest that in normal and eutopic endometrium, relaxin may exert a protective effect against endometriosis.

Circulating relaxin and cervical length in midpregnancy are independently associated with spontaneous preterm birth.

OBJECTIVE: We sought to examine associations among serum relaxin levels, cervical length, and spontaneous preterm birth (sPTB). STUDY DESIGN: We conducted a case-control study within a cohort of 1080 singleton pregnant women. In all, 38 women (3.5%) delivered spontaneously preterm (< 37 completed weeks of gestation). Relaxin was measured in serum in gestational weeks 12 and 19, cervical length only in week 19. Associations to sPTB were evaluated by logistic regression analysis and receiver operating characteristic curves. RESULTS: Cervical length and relaxin in week 19, but not week 12 were associated to sPTB. There were no correlations between cervical length and relaxin level in week 12 (controls: Spearman rho: -0.07, P = .45; cases: Spearman rho: -0.07, P = .72) or week 19 (controls: Spearman rho: -0.03, P = .77; cases: Spearman rho: 0.04, P = .84). CONCLUSION: There were no correlation between relaxin concentrations and the cervical length, indicating that relaxin is probably not the cause of preterm shortening of the cervix.

Relaxin receptor LGR7 (RXFP1) is regulated by estrogen.

Estrogen regulates LGR7 (RXFP1) mRNA expression in an in vitro model of human term pregnancy cervix that utilizes lower uterine segment fibroblasts. LGR7 mRNA levels were increased by estradiol to mean levels of 152%+/- 5.9% above those in untreated control cells. Therefore, estradiol may amplify relaxin's actions in the cervix.

Relaxin in human pregnancy.

Relaxin has beneficial effects upon the endometrium which are responsible for establishment of pregnancy. We have demonstrated that relaxin stimulates endometrial decidualization, the structural and biochemical changes in endometrial parenchymal cells, and the accompanying angiogenesis, modulation of matrix metalloproteinase activity, and increased concentrations in local immune cells which are required for implantation. Our recent data also demonstrate that either too much or too little relaxin can be detrimental. Elevated circulating maternal relaxin concentrations (hyperrelaxinemia) are associated with premature birth. This is likely due to the effects of relaxin at the level of the cervix, via upsetting the balance in the maintenance of cervical connective tissue architecture. In addition, the absence of circulating relaxin during pregnancy in women may have negative consequences upon glucose metabolism.

Relaxin in endometriosis.

Endometriosis is an important contributing factor to chronic pelvic pain and infertility. Matrix metalloproteinases (MMPs) and vascular endothelial growth factor (VEGF) have each been implicated in the establishment of endometriotic lesions. Since relaxin regulates the expression of MMPs and VEGF in the endometrium, we tested the hypothesis that relaxin plays a role in endometriosis by comparing the expression of relaxin mRNA and its LGR7 (RXFP1) receptor mRNA in normal human endometrium to those in samples from patients with endometriosis.

Expression of LGR7 in the primate corpus luteum implicates the corpus luteum as a relaxin target organ.

In women, the corpus luteum is the source of circulating relaxin. No previous studies have addressed whether the corpus luteum is also a relaxin target organ. We determined relaxin receptor LGR7 mRNA expression in human term pregnancy corpora lutea and nonhuman primate corpora lutea obtained during the menstrual cycle. Real-time reverse transcription-PCR demonstrated the expression of LGR7 mRNA in both human and rhesus monkey corpora lutea. Rhesus monkey corpora lutea were obtained from naturally cycling animals following documented luteinizing hormone (LH) surges at early, mid-, mid-late, and late luteal phases. Luteal expression of LGR7 mRNA did not show temporal variation. Since the primate corpus luteum is LH dependent, we assessed LGR7 mRNA expression in corpora lutea from rhesus monkeys treated with a gonadotropin-releasing hormone (GnRH) antagonist, which significantly suppressed pituitary LH levels. GnRH antagonist treatment, which also inhibits both progesterone and relaxin production, resulted in a fivefold increase in luteal LGR7 mRNA expression. These data suggest that luteal LGR7 mRNA expression may be regulated by relaxin and/or LH and that the primate corpus luteum is a target organ for relaxin.

Inflammation in reproductive disorders.

Inflammatory disorders account for a significant percentage of gynecologic disease, particularly in reproductive age women. Inflammation is a basic method by which we respond to infection, irritation, or injury. Inflammation is now recognized as a type of nonspecific immune response, either acute or chronic. In gynecology, inflammation leads to anatomic disorders primarily as a result of infectious disease; however inflammation can affect ovulation and hormone production as well as be associated with endometriosis. Similarly, immune cell trafficking is an important component of cyclic endometrial development in each menstrual cycle. These immune cells are required for endometrial function, producing a vast array of inflammatory cytokines. Inflammation alters endometrial receptivity, however it may also play a role in tissue repair and remodeling. Finally, inflammation affects the trophoblast and trophoblast-endometrial interaction. Some components of the immune response are required for optimal fertility and normal tissue remodeling. A better understanding of the necessary role of inflammation in reproduction will allow more rational and targeted treatment of inflammatory disorders in reproductive medicine.

Longitudinal changes in hypothalamic and ovarian function in perimenopausal women with anovulatory cycles: relationship with vasomotor symptoms.

OBJECTIVE: To determine whether the transition to menopausal status is unidirectional and predictable with aging. DESIGN: Longitudinal evaluation of the menstrual cycle hormone patterns and experience of vasomotor symptoms in an anovulatory, perimenopausal cohort, during cycles that occurred 1 and 2 years after an anovulatory cycle. SETTING: Academic center. PATIENT(S): One hundred fifty-nine of 840 women in the Daily Hormone Study, a substudy of the Study of Women's Health across the Nation (SWAN), had anovulatory cycles. Their menstrual cycle patterns were previously described. This report describes their cycle patterns and vasomotor symptoms in the subsequent 2 years. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Daily urinary hormone levels of FSH, LH, and estrogen and P metabolites and reports of daily occurrence of vasomotor symptoms. RESULT(S): While a tendency to develop cycles having a loss of negative feedback of estrogen on LH secretion was seen before menopause, there is no clear progression of cycle patterns in anovulatory women. Anovulation did not predict menopause within 2 years. Vasomotor symptoms occur before menopause, as experienced by 73% of the women. Vasomotor symptoms were not related to cycle pattern. CONCLUSION(S): Any cycle pattern may be related to vasomotor symptoms. The best predictor of vasomotor symptoms is a prior history of vasomotor symptoms.