Secretory Phase Endometrium Research Articles

Endometrial pattern in women with abnormal uterine bleeding attending a tertiary care centre of Kathmandu

Background: Abnormal uterine bleeding is one of the most common problems among pre and postmenopausal women. Abnormal uterine bleeding is diagnosed when there is a substantial change in frequency, duration, or amountof bleeding during or between periodsin the absence of any organic pathology. The study was perform to assess the endometrial causes of abnormal uterine bleeding and their incidence in different age groups. Materials and methods: It was a cross sectional and descriptive study carried out at a Manmohan medical College & hospital, over a period of two year from 13th April, 2020 to 15th March, 2022. In the study all the endometrial tissue either obtained by endometrial curettage and biopsy or hysterectomy presenting with abnormal uterine bleeding for more than 3 months were included Results: A total of 88 cases were received during this period, out of which there were 78 endometrial tissue samples and 10 hysterectomy specimens. The patient’s age ranged from 20 years to 73years. The pattern of endometrial biopsy are Proliferative (37.5%) and secretory (20.45%) phase endometrium is the most common pattern seen, followed by disordered proliferative endometrium (13.63%), endometrial hyperplasia without atypia (9.09%), Change consistent with Exogenous Hormone (5.68%), Chronic Endometritis (5.68%), Endometrial Polyp(3.40%), Endometrial Hyperplasia with Atypia(2.27%) and Endometrial Carcinoma (2.27%). Conclusions: Patients with abnormal uterine bleeding show a varying spectrum of endometrial pattern. Cyclic endometrium is commonly seen in the reproductive age group whereas hyperplasia and carcinomas are seen in pre/postmenopausal age group. Endometrial biopsy with histopathological examination is a major diagnostic tool in the evaluation of abnormal uterine bleeding.

Hyperlipidemia negatively impacts implantation by dysregulating tight junction and Claudin-3 and Claudin-4 expression in the endometrium

Clinical observational studies have suggested hyperlipidemia may disturb embryo implantation through endometrium; however, the mechanism has been unclear. With its profound implications for reproductive health, the present study aims to investigate whether hyperlipidemia affects endometrial epithelial cell tight junctions for implantation failures. By constructing hyperlipidemia mice model, the number and distribution of embryo implantation status were investigated after both natural mating and in vitro fertilization and embryo transfer (IVF-ET). Transmission electron microscopy (TEM) was used to compare the ultrastructure of tight junctions in endometrial endothelial cells. Western blot and immunofluorescence were used to explore the expression and localization of tight junction proteins, such as Claudin (CDLN)3, CLDN4, occludin (OCLN), and zonula occludens-1 (ZO1). For women with reproductive failure, mid-luteal phase endometrial tissues were collected, and gene expression of tight junction proteins was investigated using RNA sequencing and qRT-PCR. In hyperlipidemic mice, the number of embryo implantation sites significantly decreased with uneven distribution after natural mating and IVF-ET. Disrupted tight junctions were found, characterized by a decreased number of tight junctions by TEM, downregulated expressions of CDLN4, OCLN, and ZO1, and an increased expression of CLDN3 by western blot. In hyperlipidemic women with reproductive failure, the dysregulated expression of CLDN3 and CLDN4 was also present in the luteal phase endometrium. In this study, evaluation of both animal models and infertile women in vivo demonstrated that hyperlipidemia reduced female fertility, accompanied by disruption of tight junction structures and dysregulation of CLDN3 and CLDN4 expression in the endothelial cells of luteal phase endometrium.

Application of strain elastography ultrasound to the endometrium of normal women

BackgroundWhile there is a scarcity of studies utilizing strain elastography (SE) for the endometrium, commonly used gynecologic ultrasound instruments are equipped with built-in elastography modalities, primarily SE. With the objective of facilitating comprehensive examinations for gynecologic patients on a single ultrasound instrument, we undertook this study. Therefore, our aim was to study the value of SE ultrasonography in the assessment of endometrial elasticity in normal women.MethodsThree hundred and twenty normal women were recruited at our hospitals from November 2021 to December 2022. Each volunteer underwent a transvaginal two-dimensional (2D) and SE ultrasound during either the endometrial proliferative or secretory phase. The 2D ultrasound indices obtained included endometrial thickness, echo type (type A, B, and C), and blood flow grading (grades 0, 1, 2, and 3). SE indices obtained included endometrial strain values, myometrial strain values, and endometrial strain ratios. Differences in endometrial ultrasound indices between different menstrual cycles and different age groups were compared.ResultsComparison of 2D ultrasound parameters revealed that endometrial thickness in the proliferative phase endometrium group was smaller than that in the secretory phase endometrium group, with a statistically significant difference. Additionally, there was a statistically significant difference in endometrial echo types between the two groups, while the disparity in endometrial blood flow grading was not significant. Regarding SE parameters, the median and mean values of endometrial strain ratio in the proliferative phase endometrium group were smaller than those in the secretory phase endometrium group, showing a statistically significant difference. However, there were no significant differences observed between the two groups in endometrial strain and myometrial strain in the fundus. Furthermore, there were no significant differences in any of the endometrial ultrasound indices among the different age groups.ConclusionsSE can reflect changes in endometrial stiffness in different menstrual cycles and is an important tool for assessing endometrial softness.

AI-algorithm training and validation for identification of endometrial CD138+ cells in infertility-associated conditions; polycystic ovary syndrome (PCOS) and recurrent implantation failure (RIF)

BackgroundEndometrial CD138+ plasma cells serve as a diagnostic biomarker for endometrial inflammation, and their elevated occurrence correlates positively with adverse pregnancy outcomes. Infertility-related conditions like polycystic ovary syndrome (PCOS) and recurrent implantation failure (RIF) are closely associated with systemic and local chronic inflammatory status, wherein endometrial CD138+ plasma cell accumulation could also contribute to endometrial pathology. Current methods for quantifying CD138+ cells typically involve laborious and time-consuming microscopic assessments of only a few random areas from a slide. These methods have limitations in accurately representing the entire slide and are susceptible to significant biases arising from intra- and interobserver variations. Implementing artificial intelligence (AI) for CD138+ cell identification could enhance the accuracy, reproducibility, and reliability of analysis. MethodsHere, an AI algorithm was developed to identify CD138+ plasma cells within endometrial tissue. The AI model comprised two layers of convolutional neural networks (CNNs). CNN1 was trained to segment epithelium and stroma across 28,363 mm2 (2.56 mm2 of epithelium and 24.87 mm2 of stroma), while CNN2 was trained to distinguish stromal cells based on CD138 staining, encompassing 7345 cells in the object layers (6942 CD138− cells and 403 CD138+ cells). The training and performance of the AI model were validated by three experienced pathologists. We collected 193 endometrial tissues from healthy controls (n = 73), women with PCOS (n = 91), and RIF patients (n = 29) and compared the CD138+ cell percentages based on cycle phases, ovulation status, and endometrial receptivity utilizing the AI model. ResultsThe AI algorithm consistently and reliably distinguished CD138− and CD138+ cells, with total error rates of 6.32% and 3.23%, respectively. During the training validation, there was a complete agreement between the decisions made by the pathologists and the AI algorithm, while the performance validation demonstrated excellent accuracy between the AI and human evaluation methods (intraclass correlation; 0.76, 95% confidence intervals; 0.36–0.93, p = 0.002) and a positive correlation (Spearman's rank correlation coefficient: 0.79, p < 0.01). In the AI analysis, the AI model revealed higher CD138+ cell percentages in the proliferative phase (PE) endometrium compared to the secretory phase or anovulatory PCOS endometrium, irrespective of PCOS diagnosis. Interestingly, CD138+ percentages differed according to PCOS phenotype in the PE (p = 0.03). On the other hand, the receptivity status had no impact on the cell percentages in RIF samples. ConclusionOur findings emphasize the potential and accuracy of the AI algorithm in detecting endometrial CD138+ plasma cells, offering distinct advantages over manual inspection, such as rapid analysis of whole slide images, reduction of intra- and interobserver variations, sparing the valuable time of trained specialists, and consistent productivity. This supports the application of AI technology to help clinical decision-making, for example, in understanding endometrial cycle phase-related dynamics, as well as different reproductive disorders.

Demystifying the landscape of endometrial immune microenvironment in luteal-phase from cuprotosis: Implications for the mechanism and treatment of RPL

BackgroundAdaptive changes in the endometrial immune microenvironment during the luteal phase are essential for pregnancy, and their abnormalities are associated with recurrent pregnancy loss (RPL). Nevertheless, the specific mechanism is still unknown. Cuprotosis, an innovatively discovered type of programmed cell death, provides us with a pioneering perspective to decipher the landscape of luteal-phase endometrial immune microenvironment in RPL. This study aimed to analyze the immune landscape of luteal-phase endometrial microenvironment in RPL and explore the association of cuprotosis with it through integrative bioinformatics analysis. MethodsThe microarrays involving the luteal phase endometrial tissue of RPL were obtained from the GEO database. Differentially expressed genes (DEGs) of RPL were screened and key modules were detected by WGCNA. GO, KEGG, and GSEA immune enrichment analyses were performed on the DEGs in the most relevant modules to RPL. Then, the endometrial immune microenvironment landscape of RPL was analyzed, including immune infiltration analysis and correlation analysis between immune cells or immune functions. The interaction of cuprotosis-related genes (CRGs), the expression level between groups, the immune localization and their correlation with immune cells and immune function were analyzed. LASSO regression and Nomogram evaluated the diagnostic value of immune-related CRGS in RPL. Functional enrichment analysis was performed on the RPL signature CRGs. And RPL samples were grouped according to the expression of 7 RPL signature CRGs through unsupervised clustering analysis. After that, we analyzed the expression level of CRGs and immune infiltration, as well as performed immune function enrichment analysis in subtypes. In addition, we also screened potential drugs that might act on CRGs to improve the pathological mechanism of RPL. ResultsIn this study, we uncovered that DEGs and genes in key modules derived from weighted gene co-expression network analysis (WGCNA) were involved in immune regulation. And the immune infiltration landscape of RPL was significantly different from healthy controls. Furthermore, six hub genes were screened from CRGs based on Cytohubba, and their expression profilings were verified in RPL and normal mouse samples. Besides, seven CRGs closely associated with the immune regulation of RPL were identified by Spearman correlation analysis, including SLC31A1, LIAS, DLD, DLAT, DBT, ATP7B, and ATP7A, named as immune-related CRGs. Furthermore, three subgroups clustered according to these seven genes showed significant differences in immune landscape, suggesting a remarkable effect of CRGs on immune regulation. Last but not least, we analyzed the regulation network of transcription factors, miRNAs, and CRGs, and screened potential compounds for the treatment of RPL by targeting CRGs. ConclusionsThe abnormal endometrial immune microenvironment in the luteal phase was associated with the pathomechanism of RPL, and cuprotosis was closely involved in the immune microenvironment in the luteal phase endometrium of RPL. Collectively, this study revealed the potential contribution of CRGs to the pathogenesis of RPL, providing a novel breakthroughs in insights into the pathogenesis, diagnosis, and treatment of RPL.

Endometrial receptive status in patients of reproductive age with diffuse and focal adenomyosis

Introduction Endometriosis ranks second place in the structure of infertility. The development of endometrial dysfunction is one of the infertility factors, which affects the degree of decrease in the receptive abilities of endometrium and subsequent implantation.Purpose of the study was to reveal morpho-functional state of receptivity and nidation ability of endometrium in patients of reproductive age with diffuse and nodal forms of adenomyosis.Materials and methods The study included 104 patients, which were divided into 2 groups. The first group consisted of 30 patients with a diffuse form of adenomyosis, second group included 30 patients with a nodal form. The patients underwent aspiration biopsy of endometrium in the middle secretory phase. The level of ER and PR expression in the glandular and stromal compartments of endometrium was assessed, the percentage and degree of maturity of pinopodes were determined.Results The analysis of ER and PR number in the stromal and glandular components of endometrium in patients with diffuse adenomyosis revealed an increase in the secretion of ER in the glandular cells in contrast to patients of the comparison group. Significant differences in the expression of ER and PR in glandular cells were verified among observed with the nodal form of adenomyosis. The immunohistochemical reaction with antibodies to ER and PR in the nodal form of adenomyosis was significantly higher than in diffuse form. Both in the first and second groups, a decrease in the formation ability of pinopodes on the superficial epithelium was noted.Discussion Adenomyosis leads to damage of the morpho-functional state and receptive abilities of the endometrium. In the nodal form the degree of detected disorders was significantly higher.Conclusion The nodal form of adenomyosis is more common in patients of younger reproductive age and is associated with a higher prevalence of endometrial dysfunction. This is especially important for the selection of adequate modern therapy in order to maintain and improve reproductive function.

Computerized histomorphometry in the diagnosis of pre-malignant endometrial lesions

Accurate diagnosis of precursors to endometrial cancer is a major challenge to pathologists. D-score is well reproducible and predictive of prognosis than the WHO schema, however it is not widely available, this study is an attempt to design a simple workstation for estimation of D-score. Fifteen samples of endometrium with low grade hyperplasia, fifteen proliferative and eight secretory phase endometria were analyzed by light microscopy and stereology. Severeal architectural and karyometric parameters were evaluated and the D-score was calculated. The volume density of stroma (VPS) was significantly higher in proliferative than secretory and hyperplastic endometria. Among the three diagnostic groups, secretory phase endometria have -significantly- the least VPS. The mean values of glandular surface density (outSD) were higher in secretory endometria as compared to proliferative and hyperplastic endometria, however, the differences was statistically insignificant. All of evaluated nuclear parameters are insignificantly differ between hyperplastic and proliferative endometria. The ratio of longest nuclear axis to shortest nuclear axis (D/d) was significantly less in secretory than proliferative and hyperplastic endometriia. Secretory endometria also have significantly lower mean values of shape factor and significantly higher mean values of contour index than proliferative and secretory endometria. 67% of hyperplastic endometria show D-score values &lt; 0. it seems feasible to estimate D-score for suspected EIN lesions using a semi-automated workstation based on the simple stereologic and morphometric principles described in this study, preferably, if fortified by a future comparative study in which a reference method such as the QProdit system is used.

Hypoxia-mediated chemotaxis and residence of macrophage in decidua by secreting VEGFA and CCL2 during normal pregnancy.

Hypoxia is vital for the establishment of the maternal-fetal interface during early pregnancy. This study shows that decidual macrophages (dMφ) could be recruited and reside in decidua under the regulation of hypoxia/VEGFA-CCL2 axis. Infiltration and residence of decidual macrophages (dMφ) are of great significance to pregnancy maintenance for their role in angiogenesis, placental development, and inducing immune tolerance. Besides, hypoxia has now been acknowledged as an important biological event at maternal-fetal interface in the first trimester. However, whether and how hypoxia regulates biofunctions of dMφ remain elusive. Herein, we observed increased expression of C-C motif chemokine ligand 2 (CCL2) and residence of macrophages in decidua compared to secretory-phase endometrium. Moreover, hypoxia treatment on stromal cells improved the migration and adhesion of dMφ. Mechanistically, these effects might be mediated by upregulated CCL2 and adhesion molecules (especially ICAM2 and ICAM5) on stromal cells in the presence of endogenous vascular endothelial growth factor-A (VEGFA) in hypoxia. These findings were also verified by recombinant VEGFA and indirect coculture, indicating that the interaction between stromal cells and dMφ in hypoxia condition may facilitate dMφ recruitment and residence. In conclusion, VEGFA derived from a hypoxic environment may manipulate CCL2/CCR2 and adhesion molecules to enhance the interactions between dMφ and stromal cells and thus contribute to the enrichment of macrophages in decidua early during normal pregnancy.

Extracellular vesicles secreted by adenomyosis endometrial organoids contain miRNAs involved in embryo implantation and pregnancy

Research questionDo extracellular vesicles secreted by the endometrium of women with adenomyosis contain miRNAs involved in adenomyosis-related infertility? DesignA descriptive study using organoids from eutopic endometrium of women with adenomyosis (n = 4) generated and differentiated to secretory and gestational phases, in which miRNA cargo from extracellular vesicles secreted by these differentiated organoids in each phase was analysed by next-generation sequencing. miRNAs in secretory-extracellular vesicles and gestational-extracellular vesicles were selected based on the counts per million. miRNAs target genes in each phase were obtained from miRNet and gene ontology was used for enrichment analysis. ResultsmiRNA sequencing identified 80 miRNAs in secretory-phase extracellular vesicles, including hsa-miR-21-5p, hsa-miR-24-3p, hsa-miR-26a-5p, hsa-miR-92a-3p, hsa-miR-92b-3p, hsa-miR-200c-3p and hsa-miR-423a-5p, related to adenomyosis pathogenesis and implantation failure. Further, 60 miRNAs were identified in gestational-phase extracellular vesicles, including hsa-miR-21-5p, hsa-miR-26a-5p, hsa-miR-30a-5p, hsa-miR-30c-5p, hsa-miR-222-3p and hsa-miR-423a-5p were associated with preeclampsia and miscarriage. Among the target genes of these miRNAs, PTEN, MDM4, PLAGL2 and CELF1, whose downregulation (P = 0.0003, P < 0.0001, P = 0.0002 and P = 0.0003, respectively) contributes to adenomyosis pathogenesis, and impaired early embryo development, leading to implantation failure and miscarriage, are highlihghted. Further, functional enrichment analyses of the target genes revealed their involvement in cell differentiation, proliferation, apoptosis, cell cycle regulation and response to extracellular stimuli. ConclusionsEutopic endometrium in secretory and gestational phase from women with adenomyosis releases extracellular vesicles containing miRNAs involved in adenomyosis progression, impaired embryo implantation and pregnancy complications.

TNFα/TNFR1 signal induces excessive senescence of decidua stromal cells in recurrent pregnancy loss

Defects in decidual response are associated with adverse pregnancy outcomes which includes recurrent pregnancy loss (RPL). It is reported that cellular senescence happens during decidualization and pro-senescent decidual response in the luteal phase endometrium is related to RPL. However, the underlying mechanisms of how excessive decidual senescence takes place in RPL decidua cells remain largely unexplored. The senescent phenotype of RPL decidua and tumor necrosis factor receptor 1(TNFR1) expression were analyzed by using our previously published single-cell sequencing dataset of decidua cells from 6 RPL and 5 matched normal decidua, which were further verified by PCR and WB in decidual tissues. Effects of TNFα on the decidual stromal cells (DSCs) senescence and underlying molecular pathways were analyzed using the in vitro decidualization model of human endometrial stromal cells (HESCs). We showed that decidual stroma cells from RPL patients exhibited transcriptomic features of cellular senescence by analysis of single-cell datasets. The TNFα level and TNFR1 expression were increased in RPL decidua tissues. Furthermore, in vitro cell model demonstrated that increased TNFα induced excessive senescence during decidualization and TNFR1/p53/p16 pathway mediates TNFα-induced stromal senescence. In addition, we also found that the expression of IGFBP1 was regulated by TNFα-TNFR1 interaction during decidualization. Taken together, the present findings suggest that the increased secretion of TNFα induced stromal cell excessive senescence in RPL decidua, which is mediated via TNFR1, and thus provide a possible therapeutic target for the treatment of RPL.

A pilot and ex-vivo study of examination of endometrium tissue by catheter based optical coherence tomography

ObjectiveThis study aimed to distinguish ex-vivo normal and abnormal endometrium tissue samples histologically by catheter based optical coherence tomography (OCT).MethodsA total of 72 ex-vivo endometrium specimens were obtained from June 2018 to March 2021 and were imaged fresh after hysterectomy. The scanned region of endometrium was excised for histological examination and endometrium OCT images were precisely compared to corresponding histological images. Meanwhile endometrium OCT images were analyzed quantitatively with intensity of backscattered light in region of interest (ROI) and maximum penetration depth of the OCT signal. Blinded qualitative analysis on endometrium OCT images was performed by 2 assessors to determine accuracy rate and inter-rating reliability on the histopathological diagnosis.ResultsOCT images were performed successfully in 72 endometrium specimens. Five endometrium specimens developed OCT interpretation criteria and the rest 67 endometrium specimens validated qualitatively and analyzed quantitatively. We defined an OCT criteria to distinguish normal endometrium and five different abnormal endometrium phases including proliferative endometrium, secretory phase endometrium, atrophic endometrium, endometrial hyperplasia with atypia and endometrial carcinoma based on OCT imaging features. The overall diagnosis accuracy achieved by the two assessors was 72.4% based on the OCT criteria. The inter-rater reliability between assessors on overall OCT images was substantial (Kendall τb of 0.720, p < 0.05). The changes in ROI minimum intensity, ROI maximum intensity, ROI average intensity and OCT signal maximum penetration depth of five different abnormal endometrium phases were significantly different (all p < 0.001). These parameters of endometrium carcinomas were significantly different from the other four endometrium phases (all p < 0.001).ConclusionOCT has the advantage of noninvasive and rapid diagnosis, which can contribute to the diagnosis of endometrial cancer and will be an indispensable complement to traditional biopsy. Future studies in vivo with larger samples are needed to confirm this conclusion.

Investigating Uterine Natural Killer Cells Through Next Generation Sequencing — A Pilot Gene Expression Study

Background: Uterine natural killer (uNK) cells form the major leucocyte population in the stroma in late secretory phase endometrium. Their actual function is unknown with available data suggesting a potential role in angiogenesis regulation, decidualization and placentation. Next generation sequencing (NGS) can be used to accurately define day of menstrual cycle through endometrial gene expression. Genes uniquely expressed by uNK are currently unknown. However, previous studies of first trimester decidua and blood have identified genes that are uniquely expressed by these NK cells. Aim: To identify potential molecular markers of uNK cells. Method: Endometrial samples were collected from 266 reproductive aged women across menstrual cycle. Gene expression profiles in endometrial tissues were generated using Illumina NovaSeq platforms. Gene expression data was normalized for cycle stage. Expression levels of previously known NK cell genes were examined across menstrual cycle. Results: Our analysis has shown that many NK cell genes (95% decidual, 71% peripheral) are expressed by the endometrium. We have identified 12 common genes that are expressed by both decidual and peripheral NK cells, and the endometrium. Expression levels of these common genes (except one) share a similar pattern across menstrual cycle. Their expression levels increase in secretory phase endometrium with peak expression levels at late secretory phase, as predicted by what is known about changing uNK cell numbers in endometrium. One example is the gene NCAM1 which codes for protein CD56 - a specific marker for uNK cells. Conclusion: This study has shown that it is possible to assess uNK cells through molecular approaches. Further molecular study of uNK cells is warranted and will help to provide evidence regarding uNK cells in human reproduction.

Altered Endometrial Expression of α-Inhibin Subunit and Its Co-Receptor Betaglycan in Infertile Women with Endometriosis

Background: Inhibins and their co-receptor betaglycan are members of the transforming growth factor β superfamily, a group of signaling molecules that control the differentiation of human endometrium in the secretory phase of the menstrual cycle. Objective: Since endometriosis is associated with endometrial dysfunction and infertility, this study aimed at evaluating the expression of α-inhibin and betaglycan mRNA and proteins in endometrial samples of infertile women with and without endometriosis. Design: This was a cross-sectional study. Participants/Materials: Endometrial samples of women with (n = 17) and without (n = 22) endometriosis were subdivided according to the menstrual cycle phase into proliferative and secretory. Setting: University hospital. Methods: We used real-time RT-PCR to quantify mRNA levels and immunohistochemistry to localize the proteins. Results: α-inhibin mRNA levels were significantly increased in the secretory phase (p < 0.01 vs. proliferative phase) only among women with endometriosis. Conversely, betaglycan mRNA levels were downregulated in the secretory endometrium of controls (p < 0.01 vs. proliferative) but failed to change between cycle phases of patients with endometriosis. Both proteins were present in the glandular epithelium and stroma in the endometrium of women with and without endometriosis. Immunostaining analysis showed that while α-inhibin protein expression did not vary significantly, the intensity of betaglycan immunostaining decreased in the secretory phase in the control group (p = 0.038 vs. proliferative phase) but not in the endometriosis group. Limitations: We cannot determine whether endometriosis causes the abnormal expression of α-inhibin and betaglycan in the eutopic endometrium or if this alteration already existed before the establishment of endometriotic lesions. Conclusion: Our findings suggest an abnormally increased expression of α-inhibin mRNA (not protein) and betaglycan (mRNA and protein) in the secretory-phase endometrium of women with endometriosis.

P-413 Non-autologous human platelet lysate stimulates in vitro proliferation of primary human endometrial cells from patients with recurrent implantation failure

Abstract Study question Does patient diagnosis impact induction of primary human endometrial cell proliferation by commercial non-autologous human platelet lysate (HPL)? Summary answer HPL stimulates in vitro proliferation of both primary endometrial epithelial cells (EECs) and stromal cells (ESCs) from patients with and without recurrent implantation failure (RIF). What is known already Inadequate endometrial receptivity and thickness are major causes for RIF. Our preliminary results suggested that non-autologous HPL stimulates in vitro proliferation of primary human EECs and ESCs isolated from patients with a history of RIF, with and without a thin endometrium (TE). In addition, using an in vitro model of embryo attachment, we reported that 48-hour treatment with HPL significantly augments the attachment of trophoblast spheroids (generated with HTR-8/SVneo cells) to EECs, suggesting an increase in endometrial receptivity following treatment. This suggests that HPL may standardize future clinical treatments for a TE and endometrial origins of RIF. Study design, size, duration Endometrial tissue was collected from five patients without RIF (control) and eighteen RIF patients at the CReATe Fertility Centre, Toronto, Canada. The eighteen RIF patients were further classified into three groups (N = 6 each) based on endometrial phenotype: 1) proliferative phase RIF only (without a TE), 2) secretory phase RIF only (without a TE), and 3) secretory phase RIF+TE (with a TE). Primary EECs and ESCs were enzymatically isolated and cultured separately. Participants/materials, setting, methods Primary EECs and ESCs were serum-starved with serum-free culture media (SFM) for 24 hours and then treated for 48 hours with the following treatment media: SFM (negative control), or SFM supplemented with 1% HPL for EECs, or 10% HPL for ESCs. Cell viability and proliferation were assessed using the metabolic assay PrestoBlue reagent and immunocytochemistry to quantify cells actively expressing the nuclear proliferation marker Ki67. Main results and the role of chance The metabolic assay demonstrated that 48-hour treatment with non-autologous HPL stimulates a similar significant increase in EEC viability and proliferation for all patient groups. EECs from patients without RIF (control) had the highest fold increase (1.49-fold, P &amp;lt;0.001), followed by proliferative phase RIF only (1.41-fold, P &amp;lt;0.001), secretory phase RIF+TE (1.39-fold, P &amp;lt;0.001), and secretory phase RIF only (1.24-fold, P &amp;lt;0.05). For ESCs, HPL stimulated a significant 2.69-fold increase in cell viability and proliferation for the secretory phase RIF+TE patient group (P &amp;lt;0.01). Pairwise comparison of the ratio of actively proliferating (Ki67+) cells between SFM and HPL treatment revealed that EECs isolated from control patients had the highest and most significant increase by 27.3% (P &amp;lt;0.05) after HPL treatment. Although not statistically significant, EECs isolated from all RIF patients displayed a similar trend of increased Ki67+ cells after HPL treatment. Whereas HPL treatment significantly increased the ratio of Ki67+ cells for ESCs isolated from both the proliferative phase (by 23.9%, P &amp;lt;0.05) and secretory phase (by 29.9%, P &amp;lt;0.05) endometrium of RIF only patients. Limitations, reasons for caution Although our data suggests that HPL treatment significantly stimulates primary endometrial cell proliferation in vitro, our sample size per group is small (N = 5-6). A larger sample size and future randomized controlled trials are needed to determine the efficacy of HPL as a treatment for a TE and/or RIF. Wider implications of the findings Our study provides the groundwork to improve clinical treatment of a TE and endometrial origins of RIF. We anticipate that in addition to stimulating cell proliferation, HPL will also induce a broad transcriptional response towards improved endometrial receptivity. We will next focus on characterizing the transcriptomic profile following HPL treatment. Trial registration number not applicable

Abnormal Uterine Bleeding among Women Visiting Gynecology Out- patient Department of a Tertiary Care Hospital: A Descriptive Cross- sectional Study.

Abnormal uterine bleeding accounts for more than 70% of complaints in peri- and post-menopausal women. The main purpose of investigating them is to rule out premalignant and malignant endometrial lesions. This study aimed to find out the prevalence of abnormal uterine bleeding among out-patients visiting the gynecology department of a tertiary care hospital. This descriptive cross-sectional study was conducted in the Department of Obstetrics and Gynecology of a tertiary care center from August 1, 2020 to April 23, 2021. Ethical approval was obtained from the Institutional Review Committee of the same institution (IRC-LMC 07-C/020). A total of 2680 women visiting gynecology outpatient departments were included by convenience sampling technique. Women with abnormal uterine bleeding were evaluated by endometrial thickness measurement and endometrial biopsies. Statistical Package for the Social Sciences version 16 was used for data analysis. Point estimate at 95% Confidence Interval was calculated along with frequency, percentage, mean and standard deviation. Among 2680 women visiting the gynecology outpatient department, the prevalence of abnormal uterine bleeding was 240 (8.9%) (7.82-9.98 at 95% Confidence Interval). The mean endometrial thickness was 10.15±6.63mm. Majority of patients 104 (43.3%) had endometrial thickness >10mm. Sixty-six (27.8%) patients had disordered proliferative endometrium while endometrial carcinoma was noted in three (1.2%) patients. Atrophic endometrium was observed in 17 (7.2%) and endometrial hyperplasia in nine (3.8%) patients. The prevalence of abnormal uterine bleeding was found to be higher in comparison to other studies. Disordered proliferative endometrium was the most common histopathological finding followed by secretory phase endometrium.

CD200S-positive granulated lymphoid cells in endometrium appear to be CD56-positive uterine NK cells

The eutopic secretory phase endometrium in endometriosis overproduces and releases a soluble immunosuppressive CD200 molecule (CD200L) and is populated by stromal cells that contain a truncated CD200 (CD200S) that promotes a proinflammatory environment. The CD200S+ cell population persists when pregnancy occurs and are abundant in the early pregnancy decidua of women with missed abortion. In the present study, CD200S+, CD56+, and CD68+ cells were enumerated in formalin-fixed paraffin-embedded tissue sections from women with endometriosis and non-endometriosis controls. CD200S+ cells were more numerous than CD68+ macrophages and were similar in number and location to CD56bright endometrial NK cells. In some endometria, there was an additional population of CD200S- CD56+ NK cells. In ectopic endometrial peritoneal deposits and in ectopic myometrial deposits (adenomyosis), CD200S+ cells were less frequent, consistent with the known paucity of CD56+ NK cells in sites of ectopic deposits. CD200S+ cell frequency was greater in stroma surrounding the smaller ectopic cystic deposits. Dual immunofluorescent antibody staining confirmed CD200S+ cells were CD56+ NK cells. CD200S+ NK cell frequency may be greater in endometriosis patients’ endometrium and may affect embryo survival in early pregnancy. In our opinion, regulation of alternative splicing that results in CD200S rather than CD200L may provide new diagnostic and therapeutic options for women with endometriosis.

“STUDY OF ENDOMETRIAL PATHOLOGY IN ABNORMAL UTERINE BLEEDING”

CONTEXT - Histopathological analysis of endometrial samples in various forms of abnormal uterine bleeding in adjunction with clinical history of patient to identify underlying cause of bleeding. This also helps in early diagnosis of premalignant and malignant lesions leading to their proper follow up and improved prognosis. AIMS - Histopathological analysis of causes of abnormal uterine bleeding with their clinical correlation. SETTINGS AND DESIGN -A prospective observational study MATERIAL AND METHODS - This study was done in department of pathology, GSVM medical college, Kanpur. This was a prospective study from year 2017-2018. A total of 300 cases were studied. Specimen received were xed in 10% formalin, processed and sections of 3-4 micron were prepared, stained with H and E. Histopathological examination was done. RESULTS - It was observed that the highest number of patients were between 41-50 years of age. (44.33%). Menorrhagia is the most common symptom accounting for 135 (45%) patients followed by metrorrhagia accounting for 135 (45%) patients and hypomenoorhea is the least common complaint. It was observed that there were signicantly higher number of patients with proliferative phase – 68 (22.67%) and in endometrial hyperplasia 35 (33.66%) patients. The least common cause were endometrial polyp, seen in 4 (1.33%) patients, gestational endometrium and gestational trophoblastic tumor accounted for 2 ( 0.67%) patients. In patients who presented with metrorrhagia, secretory phase endometrium was the most common histopathological nding accounting for 34.62% followed by proliferative phase. In cases of endometrial hyperplasia, 28 (80%) cases were simple cystic type, 5 ( 14.29%) cases were of simple endometrial hyperplasia with atypia and complex endometrial hyperplasia with atypia was found in 2(5.71%) cases. Among cases of endometrial hyperplasia, 28 (80%) cases were simple cystic type, 5 ( 14.29%) cases were of simple endometrial hyperplasia with atypia and complex endometrial hyperplasia with atypia was found in 2(5.71%) cases. Endometrial carcinoma is commonest complain in 51-60 years and &gt;60 years age group. CONCLUSION - Histopathological analysis of AUB causes helps in identifying the cause and early diagnosis of premalignant and malignant cases.

EXPRESSION PROFILING AND FUNCTIONAL ANALYSIS REVEAL THAT CELL CYCLE PROGRESSION PATHWAYS ARE HIGHLY UPREGULATED IN MURINE ENDOMETRIOSIS LESIONS DEFICIENT IN CD55

To identify differentially expressed pathways in endometriosis lesions originating from CD55 deficient endometrium. Complement pathway genes are often dysregulated in endometriosis, including the regulatory GPI-anchored membrane protein CD55. CD55 is abundant in secretory phase endometrium and suppresses lesion formation in a murine model of endometriosis.

DEVELOPMENT OF SECRETOR AND GESTATIONAL PATIENT-DERIVED ORGANOIDS FROM ADENOMYOSIS DISEASE AS AN IN VITRO MODEL TO STUDY ADENOMYOSIS-RELATED INFERTILITY AND PERSONALIZED DRUG SCREENING

To develop organoids from human endometrium of adenomyosis patients and differentiate them to secretory phase and gestational endometrium for studying mechanisms involved in infertility and pregnancy disorders associated with adenomyosis. This in vitro model would allow the identification of therapeutic targets to improve implantation and effectively avoid miscarriages and consequently, improve ongoing pregnancy and live birth rates in these patients.

RANDOMIZED CONTROLLED DOUBLE BLINDED TRIAL OF GNRHA VS. PLACEBO AT TIME OF EMBRYO TRANSFER: INTERIM ANALYSIS OF THE GRAET (GnRH AGONIST AT TIME OF EMBRYO TRANSFER) STUDY

Embryo implantation requires a receptive endometrium under the influence of tightly regulated endocrine signaling from both the embryo and the corpus luteum. Research has found that after IVF, additional support for the endometrium with exogenous progesterone improves implantation and subsequent live birth rates. The purpose of this study is to investigate the potential benefit of a short acting GnRH agonist (GnRHa), leuprolide acetate, for support of the luteal phase endometrium prior to embryo implantation in IVF.