Different Dosages of Progesterone in Luteal Phase Support Reflect Varying Endometrial microRNA Expression in Frozen Embryo Transfer Cycles.

Despite serum progesterone being a widely accepted method for luteal phase support during embryo transfer cycles, debates persist regarding the optimal strategy for guiding clinical decisions on progesterone dosages to maximize reproductive outcomes. This retrospective study explored the utility of microRNA (miRNA) biomarkers in guiding personalized progesterone dosage adjustments for frozen embryo transfer (FET) cycles in 22 in vitro fertilization (IVF) patients undergoing hormone replacement therapy. Utilizing MIRA, an miRNA-based endometrial receptivity test, we analyzed patients' miRNA expression profiles before and after progesterone dosage adjustments to determine suitable dosages and assess endometrial status. Despite patients receiving identical progesterone dosages, variations in miRNA profiles were observed in the initial cycle, and all patients presented a displaced window of implantation. Following dosage adjustments based on their miRNA profiles, 91% of patients successfully transitioned their endometrium towards the receptive stages. However, two patients continued to exhibit persistent displaced receptivity despite the adjustments. Given the evident variation in endometrial status and serum progesterone levels among individuals, analyzing miRNA expression profiles may address the challenge of inter-personal variation in serum progesterone levels, to deliver more personalized dosage adjustments and facilitate personalized luteal phase support in IVF.

Competent blastocyst and receptivity endometrium improved clinical pregnancy in fresh embryo transfer cycles: a retrospective cohort study.

BACKGROUND: Embryo quality is usually regarded as a key predictor of successful implantation and clinical pregnancy potential. The identification of embryos that have the capacity to implant and result in a healthy pregnancy is a crucial part of in vitro fertilization (IVF). Usually, morphologically high-quality embryos are chosen for embryo transfer in IVF treatment. The aim of this study was to assess the association between the available blastocyst formation rate and the clinical pregnancy outcome following the first fresh embryo transfer cycle and provide systematic individual treatment to adjust endometrial receptivity for the next transfer cycle. METHODS: This retrospective, single-center study included 512 fresh embryo transfers conducted between 11/2019 and 08/2021, which consisted of 385 cleavage-stage (Day 3) and 127 blastocyst-stage (Day 5) embryo transfers. The two groups were divided into a clinical pregnancy group and a nonclinical pregnancy group for comparison. The association between the available blastocyst formation rate and the clinical pregnancy rate in the Day 3 and Day 5 transfer groups were considered. RESULTS: In the Day 3 group, there were 275 clinical pregnancies, and the clinical pregnancy rate was 71.43%. Although the two pronuclei (2PN) oocyte rate and available embryo rate at Day 3 were significantly higher in the clinical pregnancy group than the nonclinical pregnancy group (P < 0.05), the blastocyst formation rate and the available blastocyst formation rate were not significantly different between the clinical pregnancy group and the nonclinical pregnancy group (P > 0.05). In the Day 5 group, there were 81 clinical pregnancies, and the clinical pregnancy rate was 63.78%. No baseline characteristics showed any obvious differences between the clinical pregnancy group and nonclinical pregnancy group (P > 0.05). The blastocyst formation rate in the nonclinical pregnancy group was higher than that in the clinical pregnancy group, but the difference was not statistically significant (81.06% vs. 77.03%, P = 0.083). Interestingly, the available blastocyst formation rate and the Day 5 available blastocyst formation rate were significantly higher in the nonclinical pregnancy group than the clinical pregnancy group (66.19% vs. 60.79%, P = 0.014; 54.58% vs. 46.98%, P = 0.007). CONCLUSIONS: In fresh cycles, the available blastocyst formation rate was not associated with the clinical pregnancy outcome for Day 3 embryo transfers, and the available blastocyst formation rate was not positively correlated with the clinical pregnancy outcome for Day 5 embryo transfers.

Macrophage polarization in adenomyosis: A review.

Adenomyosis (AM) is a common gynecological disorder characterized by the presence of endometrial glands and stroma within the uterine myometrium. It is associated with abnormal uterine bleeding (AUB), dysmenorrhea, and infertility. Although several mechanisms have been proposed to elucidate AM, the exact cause and development of the condition remain unclear. Recent studies have highlighted the significance of macrophage polarization in the microenvironment, which plays a crucial role in AM initiation and progression. However, a comprehensive review regarding the role and regulatory mechanism of macrophage polarization in AM is currently lacking. Therefore, this review aims to summarize the phenotype and function of macrophage polarization and the phenomenon of the polarization of adenomyosis-associated macrophages (AAMs). It also elaborates on the role and regulatory mechanism of AAM polarization in invasion/migration, fibrosis, angiogenesis, dysmenorrhea, and infertility. Furthermore, this review explores the underlying molecular mechanisms of AAM polarization and suggests future research directions. In conclusion, this review provides a new perspective on understanding the pathogenesis of AM and provides a theoretical foundation for developing targeted drugs through the regulation of AAM polarization.

Analysis of the Expression of LSF Transcription Factor in the Regulation of Transcription and TSG101 during the Neoplastic Transformation of Endometrial Cells.

Previous research indicates that carcinogenesis involves disrupting the functions of numerous genes, including factors involved in the regulation of transcription and cell proliferation. For these reasons, in endometrial carcinogenesis, we decided to investigate the expression of TSG101 (a suppressor of tumor transformation) and LSF (a transcription factor involved in numerous cellular processes, such as cell cycle regulation, cell growth, development, and apoptosis). LSF may be involved in the regulation of TSG101 expression. The research material consisted of endometrial cancer samples from 60 patients. The control group consisted of normal endometrium samples donated by 60 women undergoing surgery for benign diseases of the female reproductive organs. The samples were subjected to immunohistochemical staining with antibodies specific to TSG101 and LSF. Specific antibodies were used to identify TSG101 and LSF in the examined histopathological preparations. An approximately 14-fold lower risk of endometrial cancer development was observed in patients with TSG expression in more than 75% of the assessed cells (4% vs. 36%; OR = 0.07; p = 0.0182). There was a four-fold lower risk of endometrial cancer development in patients with LSF expression in more than 50% of the assessed cells (32% vs. 64%; OR = 0.26; p = 0.0262). A more than three-fold lower risk of endometrial cancer development was observed in patients with LSF expression in more than 75% of the assessed cells (24% vs. 52%; OR = 0.29; p = 0.0454). Endometrial cancer was diagnosed in those with a lower level of TSG101 expression than in those with a cancer-free endometrium. Decreased expression of TSG101 may be a marker of endometrial cancer, and increased expression of LSF when diagnosed with endometrial cancer may indicate greater advancement of the disease. These markers might be used as diagnostic and prognostic markers-however, there is a lack of a correlation between them.

Metformin ameliorates endometrial thickness in a rat model of thin endometrium.

Metformin, a well-established anti-diabetic drug, is also used in managing various other metabolic disorders including polycystic ovarian syndrome (PCOS). There are evidences to show that metformin improves endometrial functions in PCOS women. However, fewer studies have explored the direct effects of metformin on endometrium. Previous in vitro studies have shown that therapeutic serum concentrations of metformin enhance endometrial epithelial cell proliferation. The present study was undertaken to investigate in vivo effects of metformin on endometrial proliferation in a rat model of thin endometrium. Toward this, a rat model of thin endometrium was developed. Metformin (0.1% or 1% w/v) was administrated orally for 15 days in rats with thin endometrium. Oral metformin administration for three consecutive estrous cycles (15 days) in the thin endometrium rat model led to an increase in endometrial thickness compared to sham endometrium. Histological analysis showed a significant increase in the number of endometrial glands (P < 0.05), stromal cells (P < 0.01) and blood vessels (P < 0.01) in metformin-treated (n = 10 in each group) uterine horns compared to sham (saline-treated) uterine horns in rats. The expression of proliferating cell nuclear antigen and vascular epithelial growth factor was found to be upregulated on treatment with 1% metformin-treated group (n = 7). However, pregnancy outcomes in the rats treated with metformin remained unaltered despite the restoration of endometrial thickness. In conclusion, the study demonstrated that metformin ameliorates endometrial thickness in a rat model of thin endometrium by increasing endometrial proliferation and angiogenesis, without restoration of embryo implantation.

Differential gene expression in decidualized human endometrial stromal cells induced by different stimuli.

Decidualization can be induced by culturing human endometrial stromal cells (ESCs) with several decidualization stimuli, such as cAMP, medroxyprogesterone acetate (MPA) or Estradiol (E2). However, it has been unclear how decidualized cells induced by different stimuli are different. We compared transcriptomes and cellular functions of decidualized ESCs induced by different stimuli (MPA, E2 + MPA, cAMP, and cAMP + MPA). We also investigated which decidualization stimulus induces a closer in vivo decidualization. Differentially expressed genes (DEGs) and altered cellular functions by each decidualization stimuli were identified by RNA-sequence and gene-ontology analysis. DEGs was about two times higher for stimuli that use cAMP (cAMP and cAMP + MPA) than for stimuli that did not use cAMP (MPA and E2 + MPA). cAMP-using stimuli altered the cellular functions including angiogenesis, inflammation, immune system, and embryo implantation whereas MPA-using stimuli (MPA, E2 + MPA, and cAMP + MPA) altered the cellular functions associated with insulin signaling. A public single-cell RNA-sequence data of the human endometrium was utilized to analyze in vivo decidualization. The altered cellular functions by in vivo decidualization were close to those observed by cAMP + MPA-induced decidualization. In conclusion, decidualized cells induced by different stimuli have different transcriptome and cellular functions. cAMP + MPA may induce a decidualization most closely to in vivo decidualization.

Rate and risk factors of inadequate endometrial tissues after endometrial sampling among Bhutanese women at the national referral hospital of Bhutan: a cross-sectional study.

INTRODUCTION: Women presenting with abnormal uterine bleeding needs careful and thorough assessment including ultrasound examination of endometrium and histopathological assessment of the endometrial tissues. The objective of this cross-sectional study was to determine the rate and the factors associated with inadequate endometrial tissues after endometrial sampling using MedGyn® pipette among Bhutanese women at the colposcopy clinic, Jigme Dorji Wangchuck National Referral Hospital (JDWNRH), Bhutan. METHODS: This cross-sectional study was conducted at the colposcopy clinic, JDWNRH, Thimphu between October, 2021 and March, 2022. Women included in this study underwent endometrial sampling using MedGyn® pipette without anesthesia as an office procedure. Data were collected using an interviewer-administered questionnaire and results extracted into a structured pro forma. The histopathology reports were extracted from the Department of Pathology and Laboratory Medicine, JDWNRH using the unique Bhutanese citizenship identity card number of the study participants. RESULTS: Inadequate endometrial tissues were noted in 27% (33 out of 122 cases). Among 89 patients with an adequate endometrial tissue, histologic results were normal in 30 (33.7%), benign pathology in 22 (24.7%), atrophy in 10 (8.2%), and hyperplasia in 27 (30.3%). In a univariate analysis, menopausal state (OR 1.6, 95% CI 0.708-3.765), overweight and obese (OR 1.6 95% CI 0.640-3.945), unemployed (OR 1.7, 95% CI 0.674-1.140), nulliparous (OR 1.7, 95% CI 0.183-15.816), primipara (OR 5.1, 95% CI 0.635-40.905) and use of hormonal contraception (OR 2.1, 95% CI 0.449-10.049) were associated with increased risk of inadequate endometrial tissues. On multivariate regression analysis, nulliparity (OR 1.1, 95% CI 0.101-12.061), overweight and obesity (OR 1.4, 95% CI 0.490-3.917), use of hormonal contraceptives (OR 2.2, 95% CI 0.347-13.889), and junior surgeons (OR 1.1, 95%CI 0.463-2.443) were found to be associated with inadequate endometrial tissues. However, the above associations were not statistically significant (p > 0.05). CONCLUSION: The rate of inadequate endometrial tissue following endometrial sampling using MedGyn® pipette was 27.0%. Factors associated with an increased risk of inadequate endometrial tissue after endometrial sampling were menopausal state, overweight and obese, unemployed, nulliparous, primipara and use of hormonal contraception.

Isolation and characterization of feline endometrial mesenchymal stem cells.

BACKGROUND: Recently, there has been a growing interest in stem cells for human medicine. Limited feline endometrial mesenchymal stem cell (fEM-MSC) research in veterinary medicine necessitates reporting for future feline disease research and therapy. OBJECTIVES: This study aimed to isolate fEM-MSCs from feline endometrial tissues and evaluate their morphology, proliferative ability, differentiation ability, and immunophenotype. METHODS: Feline endometrial tissues were obtained from the ovariohysterectomies of healthy cats and isolated using an enzymatic method. The morphology and proliferative ability of the isolated cells were assessed using a doubling time (DT) assay from passages 3 to 6 (P3 - P6). We measured pluripotency gene expressions of cells in P2 using quantitative real-time polymerase chain reaction (qRT-PCR). To investigate MSC characteristics, a trilineage differentiation assay was conducted in P4, and cells in P4 were immunophenotyped using flow cytometry. RESULTS: fEM-MSCs showed a typical spindle-shaped morphology under a microscope, and the DT was maintained from P3 to P6. fEM-MSCs could differentiate into adipocytes, osteoblasts, and chondrocytes, and expressed three pluripotency markers (OCT4, SOX2, and NANOG) by qRT-PCR. Immunophenotypic analysis showed that the fEM-MSCs were CD14 -, CD34 -, CD45 -, CD9+, and CD44+. CONCLUSIONS: In this study, the feline endometrium was a novel source of MSCs, and to the best of our knowledge, this is the first report on the isolation method and characteristics of fEM-MSCs.

NK cell receptor profiling of endometrial and decidual NK cells reveals pregnancy-induced adaptations.

Natural killer (NK) cells, with a unique NK cell receptor phenotype, are abundantly present in the non-pregnant (endometrium) and pregnant (decidua) humanuterine mucosa. It is hypothesized that NK cells in the endometrium are precursors for decidual NK cells present during pregnancy. Microenvironmental changes can alter the phenotype of NK cells, but it is unclear whether decidual NK cell precursors in the endometrium alter their NK cell receptor repertoire under the influence of pregnancy. To examine whether decidual NK cell precursors reveal phenotypic modifications upon pregnancy, we immunophenotyped the NK cell receptor repertoire of both endometrial and early-pregnancy decidual NK cells using flow cytometry. We showed that NK cells in pre-pregnancy endometrium have a different phenotypic composition compared to NK cells in early-pregnancy decidua. The frequency of killer-immunoglobulin-like receptor (KIR expressing NK cells, especially KIR2DS1, KIR2DL2L3S2, and KIR2DL2S2 was significantly lower in decidua, while the frequency of NK cells expressing activating receptors NKG2D, NKp30, NKp46, and CD244 was significantly higher compared to endometrium. Furthermore, co-expression patterns showed a lower frequency of NK cells co-expressing KIR3DL1S1 and KIR2DL2L3S2 in decidua. Our results provide new insights into the adaptations in NK cell receptor repertoire composition that NK cells in the uterine mucosa undergo upon pregnancy.

From endometrium to fingernail bed: histological evaluation of a rare cutaneous metastasis.

In the dermatological spectrum of oncologic manifestations, cutaneous metastases from endometrial carcinoma stand as a rarity, given the tumour's predilection for neighbouring uterine regions. We present an exceptional case of a patient in her mid-50s, whereby an endometrial carcinoma, defying conventional pathways, manifested on the skin and nail of her distal fourth finger, an unusual site for cutaneous metastases, with a specific histology of the primary cancer.

Inhibition of IGF2BP1 attenuates the progression of endometriosis through PTBP1.

INTRODUCTION: Endometriosis (EMs), manifested by pain and infertility, is a chronic inflammatory disease. The precise pathophysiology of this disease remains uncertain. Insulin-like growth factor-2 mRNA-binding protein 1 (IGF2BP1) and polypyrimidine tract-binding protein 1 (PTBP1) have both been found to regulate proliferation, apoptosis, and invasion. This study aimed to investigate the effects of IGF2BP1/PTBP1 in treating EMs. MATERIALS AND METHODS: qRT-PCR and western blotting were employed to quantify IGF2BP1 and PTBP1 expression in six patients with EMs (mean age 33.83 years). The correlation analysis, STRING database prediction, and RNA immunoprecipitation were utilized to identify the relationship between IGF2BP1 and PTBP1. Ectopic endometrial volume, weight, HE staining, and IGF2BP1 silencing were utilized to estimate the effects of IGF2BP1 in EMs model rats. qRT-PCR, CCK-8, 5-ethynyl-2'-deoxyuridine (EDU) labeling, Transwell assay, and flow cytometry were utilized to assess the effects of IGF2BP1/PTBP1 on the proliferation, migration, invasion, and apoptosis of ectopic endometrial stromal cells (eESCs). Furthermore, western blotting was employed to evaluate expressions of PCNA, VEGF, and E-cadherin in EMs rats and eESCs. RESULTS: The mRNA and protein levels of IGF2BP1 and PTBP1 in the ectopic and eutopic endometrium of EMs patients were significantly increased. RNA immunoprecipitation revealed a close interaction of IGF2BP1 with PTBP1. Additionally, the endometrial volume, weight, and histopathologic scores in rats were significantly reduced after IGF2BP1 silencing. IGF2BP1 silencing also decreased the expression of PCNA and VEGF, and increased E-cadherin expression in endometrial tissues of EMs rats. Moreover, IGF2BP1 silencing inhibited proliferation, migration, and invasion and promoted apoptosis through PTBP1 in eESCs. CONCLUSIONS: IGF2BP1 exhibits potential beneficial properties in the management of EMs by interacting with PTBP1, thereby highlighting IGF2BP1 as a promising therapeutic target for EMs.

De novo reconstruction of a functional in vivo-like equine endometrium using collagen-based tissue engineering.

To better understand molecular aspects of equine endometrial function, there is a need for advanced in vitro culture systems that more closely imitate the intricate 3-dimensional (3D) in vivo endometrial structure than current techniques. However, development of a 3D in vitro model of this complex tissue is challenging. This study aimed to develop an in vitro 3D endometrial tissue (3D-ET) with an epithelial cell phenotype optimized by treatment with a Rho-associated protein kinase (ROCK) inhibitor. Equine endometrial epithelial (eECs) and mesenchymal stromal (eMSCs) cells were isolated separately, and eECs cultured in various concentrations of Rock inhibitor (0, 5, 10 µmol) in epithelial medium (EC-medium) containing 10% knock-out serum replacement (KSR). The optimal concentration of Rock inhibitor for enhancing eEC proliferation and viability was 10 µM. However, 10 µM Rock inhibitor in the 10% KSR EC-medium was able to maintain mucin1 (Muc1) gene expression for only a short period. In contrast, fetal bovine serum (FBS) was able to maintain Muc1 gene expression for longer culture durations. An in vitro 3D-ET was successfully constructed using a collagen-based scaffold to support the eECs and eMSCs. The 3D-ET closely mimicked in vivo endometrium by displaying gland-like eEC-derived structures positive for the endometrial gland marker, Fork headbox A2 (FOXA2), and by mimicking the 3D morphology of the stromal compartment. In addition, the 3D-ET expressed the secretory protein MUC1 on its glandular epithelial surface and responded to LPS challenge by upregulating the expression of the interleukin-6 (IL6) and prostaglandin F synthase (PGFS) genes (P < 0.01), along with an increase in their secretory products, IL-6 (P < 0.01) and prostaglandin F2alpha (PGF2α) (P < 0.001) respectively. In the future, this culture system can be used to study both normal physiology and pathological processes of the equine endometrium.

Glutaminolysis regulates endometrial fibrosis in intrauterine adhesion via modulating mitochondrial function.

BACKGROUND: Endometrial fibrosis, a significant characteristic of intrauterine adhesion (IUA), is caused by the excessive differentiation and activation of endometrial stromal cells (ESCs). Glutaminolysis is the metabolic process of glutamine (Gln), which has been implicated in multiple types of organ fibrosis. So far, little is known about whether glutaminolysis plays a role in endometrial fibrosis. METHODS: The activation model of ESCs was constructed by TGF-ß1, followed by RNA-sequencing analysis. Changes in glutaminase1 (GLS1) expression at RNA and protein levels in activated ESCs were verified experimentally. Human IUA samples were collected to verify GLS1 expression in endometrial fibrosis. GLS1 inhibitor and glutamine deprivation were applied to ESCs models to investigate the biological functions and mechanisms of glutaminolysis in ESCs activation. The IUA mice model was established to explore the effect of glutaminolysis inhibition on endometrial fibrosis. RESULTS: We found that GLS1 expression was significantly increased in activated ESCs models and fibrotic endometrium. Glutaminolysis inhibition by GLS1 inhibitor bis-2-(5-phenylacetamido-1,2,4-thiadiazol-2-yl) ethyl sulfide (BPTES or glutamine deprivation treatment suppressed the expression of two fibrotic markers, α-SMA and collagen I, as well as the mitochondrial function and mTORC1 signaling in ESCs. Furthermore, inhibition of the mTORC1 signaling pathway by rapamycin suppressed ESCs activation. In IUA mice models, BPTES treatment significantly ameliorated endometrial fibrosis and improved pregnancy outcomes. CONCLUSION: Glutaminolysis and glutaminolysis-associated mTOR signaling play a role in the activation of ESCs and the pathogenesis of endometrial fibrosis through regulating mitochondrial function. Glutaminolysis inhibition suppresses the activation of ESCs, which might be a novel therapeutic strategy for IUA.

Enhancing endometrial receptivity: the roles of human chorionic gonadotropin in autophagy and apoptosis regulation in endometrial stromal cells.

Inadequate endometrial receptivity often results in embryo implantation failure and miscarriage. Human chorionic gonadotropin (hCG) is a key signaling molecule secreted during early embryonic development, which regulates embryonic maternal interface signaling and promotes embryo implantation. This study aimed to examine the impact of hCG on endometrial receptivity and its underlying mechanisms. An exploratory study was designed, and endometrial samples were obtained from women diagnosed with simple tubal infertility or male factor infertile (n = 12) and recurrent implantation failure (RIF, n = 10). Using reverse transcription-quantitative PCR and western blotting, luteinizing hormone (LH)/hCG receptor (LHCGR) levels and autophagy were detected in the endometrial tissues. Subsequently, primary endometrial stromal cells (ESCs) were isolated from these control groups and treated with hCG to examine the presence of LHCGR and markers of endometrial receptivity (HOXA10, ITGB3, FOXO1, LIF, and L-selectin ligand) and autophagy-related factors (Beclin1, LC3, and P62). The findings revealed that the expressions of receptivity factors, LHCGR, and LC3 were reduced in the endometrial tissues of women with RIF compared with the control group, whereas the expression of P62 was elevated. The administration of hCG to ESCs specifically activated LHCGR, stimulating an increase in the endometrial production of HOXA10, ITGB3, FOXO1, LIF and L-selectin ligands. Furthermore, when ESCs were exposed to 0.1 IU/mL hCG for 72 h, the autophagy factors Beclin1 and LC3 increased within the cells and P62 decreased. Moreover, the apoptotic factor Bax increased and Bcl-2 declined. However, when small interfering RNA was used to knock down LHCGR, hCG was less capable of controlling endometrial receptivity and autophagy molecules in ESCs. In addition, hCG stimulation enhanced the phosphorylation of ERK1/2 and mTOR proteins. These results suggest that women with RIF exhibit lower levels of LHCGR and compromised autophagy function in their endometrial tissues. Thus, hCG/LHCGR could potentially improve endometrial receptivity by modulating autophagy and apoptosis.

Tissue-specific nonheritable influences drive endometrial immune system variation.

Although human twin studies have revealed the combined contribution of heritable and environmental factors in shaping immune system variability in blood, the contribution of these factors to immune system variability in tissues remains unexplored. The human uterus undergoes constant regeneration and is exposed to distinct environmental factors. To assess uterine immune system variation, we performed a system-level analysis of endometrial and peripheral blood immune cells in monozygotic twins. Although most immune cell phenotypes in peripheral blood showed high genetic heritability, more variation was found in endometrial immune cells, indicating a stronger influence by environmental factors. Cytomegalovirus infection was identified to influence peripheral blood immune cell variability but had limited effect on endometrial immune cells. Instead, hormonal contraception shaped the local endometrial milieu and immune cell composition with minor influence on the systemic immune system. These results highlight that the magnitude of human immune system variation and factors influencing it can be tissue specific.

The association of endometrial closure during cesarean section to the risk of developing uterine scar defect: a randomized control trial.

PURPOSE: The surgical technique for uterine closure following cesarean section influences the healing of the cesarean scar; however, there is still no consensus on the optimal technique regarding the closure of the endometrium layer. The aim of this study was to compare the effect of closure versus non-closure of the endometrium during cesarean section on the risk to develop uterine scar defect and associated symptoms. METHODS: A randomized prospective study was conducted of women undergoing first elective cesarean section at a single tertiary medical center. Exclusion criteria included previous uterine scar, preterm delivery and dysmorphic uterus. Women were randomized for endometrial layer closure versus non-closure. Six months following surgery, women were invited to the ambulatory gynecological clinic for follow-up visit. 2-D transvaginal ultrasound examination was performed to evaluate the cesarean scar characteristics. In addition, women were evaluated for symptoms that might be associated with uterine scar defect. Primary outcome was defined as the residual myometrial thickness (RMT) at the uterine cesarean scar. Data are presented as median and interquartile range. RESULTS: 130 women were recruited to the study, of them follow-up was achieved in 113 (86.9%). 61 (54%) vs. 52 (46%) of the women were included in the endometrial closure vs. non-closure groups, respectively. Groups were comparable for patient's demographic, clinical characteristics and follow-up time for postoperative evaluation. Median RMT was 5.3 (3.0-7.7) vs. 4.6 (3.0-6.5) mm for the endometrial closure and non-closure groups, respectively (p = 0.38). Substantially low RMT (< 2.5 mm) was measured in four (6.6%) women in the endometrial closure group and three (5.8%) of the women in the non-closure group (p = 0.86). All other uterine scar sonographic measurements, as well as dysmenorrhea, pelvic pain and intermenstrual bleeding rates were comparable between the groups. CONCLUSION: Closure versus non-closure of the endometrial layer during cesarean uterine incision repair has no significant difference in cesarean scar characteristics and symptom rates at 6 months follow-up.

The Dysregulated IL-23/TH17 Axis in Endometriosis Pathophysiology.

Endometriosis is a chronic inflammatory disease in which endometrial-like tissue grows ectopically, resulting in pelvic pain and infertility. IL-23 is a key contributor in the development and differentiation of TH17 cells, driving TH17 cells toward a pathogenic profile. In a variety of inflammatory and autoimmune disorders, TH17 cells secrete proinflammatory cytokines, including IL-17, contributing to disease pathophysiology. Our studies and others have implicated IL-17 and TH17 cell dysregulation in endometriosis, which is associated with disease severity. In this article, we address whether IL-23-driven TH17 cells contribute to cardinal features of lesion proliferation, vascularization, and inflammation in endometriosis using patient samples, representative cell lines, and our established mouse model of endometriosis. The results indicated dysregulated expression of key genes in the IL-23/TH17 axis in patient ectopic and eutopic endometrial samples and increased IL-23 protein in patient plasma compared with controls. In vitro studies using primary human TH cells determined that rIL-23 mixture treatment increased pathogenic TH17 cell frequency. Similarly, rIL-23 treatment of cell lines (12Z cells, EECCs, HUVECs, and hESCs) representative of the endometriotic lesion microenvironment increased cytokines and growth factors, which play a role in lesion establishment and maintenance. In a syngeneic mouse model of endometriosis, rIL-23 treatment altered numbers of myeloid and T cell subsets in peritoneal fluid and increased giant cells within the lesion. Lesions from rIL-23-treated mice did not reveal significant alterations in proliferation/vascularization, although trends of increased proliferation and vascularization were observed. Collectively, these findings provide insights into the impact of the IL-23/TH17 axis on local immune dysfunction and broadly on endometriosis pathophysiology.

An Estrogen-NK Cells Regulatory Axis in Endometriosis, Related Infertility, and Miscarriage.

Endometriosis is a common estrogen-dependent condition that impacts 8-10% of women in their reproductive age, resulting in notable pain, morbidity, and infertility. Despite extensive research endeavors, the precise cause of endometriosis remains elusive, and the mechanisms contributing to its associated infertility are still not well comprehended. Natural killer (NK) cells, vital innate immune cells crucial for successful pregnancy, have been investigated for their potential involvement in the pathogenesis of endometriosis. Prior research has mainly concentrated on the diminished cytotoxicity of NK cells in endometrial fragments that evade the uterus. Interestingly, accumulating evidence suggests that NK cells play multifaceted roles in regulating the biology of endometrial stromal cells (ESCs), promoting local immune tolerance, influencing endometrial receptivity, oocyte development, and embryo implantation, thereby contributing to infertility and miscarriage in patients with endometriosis. In this comprehensive review, our goal is to summarize the current literature and provide an overview of the implications of NK cells in endometriosis, especially concerning infertility and pregnancy loss, under the influence of estrogen.

Simple method for isolation and culture of primary buffalo (Bubalus bubalis) endometrial epithelial cells (pBuEECs) and its characterization using high throughput proteomics approach.

Early embryonic mortality resulting from insufficient interaction between the embryo and the uterus leads to the failure of pregnancy in livestock animals. Thus, it is imperative to comprehend the multifaceted process of implantation at molecular levels, which requires synchronized feto-maternal interaction. The in-vitro models serve as valuable tools to investigate the specific stages of implantation. The present study was undertaken to develop a simple method to isolate and culture the primary buffalo endometrial epithelial cells (pBuEECs), followed by proteome profiling of the proliferating cells. Collagenase I was used to separate uterine epithelial cells (UECs) from the ipsilateral uterine horn, and then the cells were separated using a cell strainer. After being seeded on culture plates, UECs developed colonies with characteristic epithelial shape and expressed important markers such as cytokeratin 18 (KRT18), progesterone receptor (PGR), ß-estrogen receptor (ESR1), and leukemia inhibitory factor (LIF), which were confirmed by PCR. The purity of epithelial cells was assessed using cytokeratin 18 immunostaining, which indicated approximately 99% purity in cultured cells. The proteome profiling of pBuEECs via high-throughput tandem mass spectrometry (MS), identified a total of 3383 proteins. Bioinformatics analysis revealed enrichment in various biological processes, including cellular processes, metabolic processes, biological regulation, localization, signaling, and developmental processes. Moreover, the KEGG pathway analysis highlighted associations with the ribosome, proteosome, oxidative phosphorylation, spliceosome, and cytoskeleton regulation pathways. In conclusion, these well characterized cells offer valuable in-vitro model to enhance the understanding of implantation and uterine pathophysiology in livestock animals, particularly buffaloes.

Establishment of Murine Pregnancy Requires the Promyelocytic Leukemia Zinc Finger Transcription Factor.

Using an established human primary cell culture model, we previously demonstrated that the promyelocytic leukemia zinc finger (PLZF) transcription factor is a direct target of the progesterone receptor (PGR) and is essential for progestin-dependent decidualization of human endometrial stromal cells (HESCs). These in vitro findings were supported by immunohistochemical analysis of human endometrial tissue biopsies, which showed that the strongest immunoreactivity for endometrial PLZF is detected during the progesterone (P4)-dominant secretory phase of the menstrual cycle. While these human studies provided critical clinical support for the important role of PLZF in P4-dependent HESC decidualization, functional validation in vivo was not possible due to the absence of suitable animal models. To address this deficiency, we recently generated a conditional knockout mouse model in which PLZF is ablated in PGR-positive cells of the mouse (Plzf d/d). The Plzf d/d female was phenotypically analyzed using immunoblotting, real-time PCR, and immunohistochemistry. Reproductive function was tested using the timed natural pregnancy model as well as the artificial decidual response assay. Even though ovarian activity is not affected, female Plzf d/d mice exhibit an infertility phenotype due to an inability of the embryo to implant into the Plzf d/d endometrium. Initial cellular and molecular phenotyping investigations reveal that the Plzf d/d endometrium is unable to develop a transient receptive state, which is reflected at the molecular level by a blunted response to P4 exposure with a concomitant unopposed response to 17-ß estradiol. In addition to a defect in P4-dependent receptivity, the Plzf d/d endometrium fails to undergo decidualization in response to an artificial decidual stimulus, providing the in vivo validation for our earlier HESC culture findings. Collectively, our new Plzf d/d mouse model underscores the physiological importance of the PLZF transcription factor not only in endometrial stromal cell decidualization but also uterine receptivity, two uterine cellular processes that are indispensable for the establishment of pregnancy.