Endometrial Sensitivity Research Articles

"All-In-One" Genetic Tool Assessing Endometrial Receptivity for Personalized Screening of Female Sex Steroid Hormones.

Endometrium is the uterine lining that undergoes hundreds of cycles of proliferation, differentiation, and desquamation throughout a woman's reproductive life. Recently, much attention is paid to the appropriate endometrial functioning, as decreased endometrial receptivity is stated to be one of the concerns heavily influencing successes of embryo implantation rates and the efficacy of in vitro fertilization (IVF) treatment. In order to acquire and maintain the desired endometrial receptivity during IVF cycles, luteal phase support by various progestagens or other hormonal combinations is generally recommended. However, today, the selection of the specific hormonal therapy during IVF seems to be empirical, mainly due to a lack of appropriate tools for personalized approach. Here, we designed the genetic tool for patient-specific optimization of hormonal supplementation schemes required for the maintenance of endometrial receptivity during luteal phase. We optimized and characterized in vitro endometrial stromal cell (ESC) decidualization model as the adequate physiological reflection of endometrial sensitivity to steroid hormones. Based on the whole transcriptome RNA sequencing and the corresponding bioinformatics, we proposed that activation of the decidual prolactin (PRL) promoter containing ancient transposons MER20 and MER39 may reflect functioning of the core decidual regulatory network. Furthermore, we cloned the sequence of decidual PRL promoter containing MER20 and part of MER39 into the expression vector to estimate the effectiveness of ESC decidual response and verified sensitivity of the designed system. We additionally confirmed specificity of the generated tool using human diploid fibroblasts and adipose-derived human mesenchymal stem cells. Finally, we demonstrated the possibility to apply our tool for personalized hormone screening by comparing the effects of natural progesterone and three synthetic analogs (medroxyprogesterone 17-acetate, 17α-hydroxyprogesterone caproate, dydrogesterone) on decidualization of six ESC lines obtained from patients planning to undergo the IVF procedure. To sum up, we developed the “all-in-one” genetic tool based on the MER20/MER39 expression cassette that provides the ability to predict the most appropriate hormonal cocktail for endometrial receptivity maintenance specifically and safely for the patient, and thus to define the personal treatment strategy prior to the IVF procedure.

193 Melatonin implants in spring improve embryo production of aged ewes after superovulation regardless of endometrial progesterone receptor expression

Twenty-three Rasa Aragonesa aged ewes (average age: 10.3±0.3 years) were used to determine the effect of melatonin on ovulatory response, embryo production, and endometrial expression of progesterone receptors (PR) after superovulation. Ewes were treated (M, n=13) or not (control, C, n=10) with melatonin implants in March (Day 0, Northern Hemisphere autumn), and received intravaginal progestogen sponges for 14 days on Day 77. Superovulatory treatments consisted of 8 doses in decreasing concentrations (2 mL×2 and 1 mL×6) of 176 NIH-FSH-S1 units of NIADDK-oFSH-17 (Ovagen, ICPbio Reproduction, Auckland, New Zealand) administered twice daily starting 72h before sponge removal. Seven days after oestrus, embryos were recovered by laparotomy, ewes were killed, and uterine horns were processed to study PR expression by immunohistochemistry. The amount of PR was estimated subjectively by 2 independent observers in 5 endometrial compartments: luminal epithelium (LE), superficial (sGE) and deep (dGE) glandular epithelia, and superficial (sS) and deep (dS) stroma. The extent of staining was expressed on a scale from 0 to 100. Data were analysed with a 2×2 factorial ANOVA. Melatonin implants improved fertilization (92v. 57%, for M and C groups, respectively; P<0.01), blastocyst (47v. 9%; P<0.01), viability (88v. 31%; P<0.0001), and freezability (69v. 21%; P<0.001) rates. Specifically, melatonin induced a significant reduction of the number of non-viable (degenerate and retarded) embryos (0.3v. 1.5; P<0.05) and increased blastocysts (2.8v. 0.8; P<0.05) per ewe. Melatonin treatment decreased PR staining intensity (47v. 55%; P<0.05), but this effect was not observed when the individual cell types were compared (Table 1). Because the number of corpora lutea (CL) was responsible for different PR expression in both groups (P<0.0001), animals were divided into 2 ovulation rate categories: <10 CL and ≥10 CL, with lesser PR expression in the ≥10 CL group (P<0.0001); this lower PR immunostaining in ≥10 CL is consistent with progesterone down-regulation of its own receptor. An interaction among number of CL and treatment was found for embryo quality (P<0.05); thus, the positive effect of melatonin on this parameter was particularly effective in the low-ovulation-rate group. These results demonstrate that melatonin treatment in the autumn improves embryo quality in aged ewes, and that this effect is not explained by a differential endometrial sensitivity to progesterone. Table 1.Embryo production (mean±s.e.m.) in melatonin-treated (M) and control (C) ewes after superovulation in autumn, and staining intensity of progesterone receptors in the endometrium (CL=corpora lutea)

Abstract 248: Metformin sensitizes endometrial cancer cells to chemotherapy by inhibiting IDH1-TET1 pathway

Abstract Chemotherapy-resistance is the major obstacle for endometrial carcinoma patient administration. Accumulated evidences indicate that metformin sensitizes cancers from several different sites to chemotherapy drugs and reverses progestin resistance in endometrial cancer. Our group has demonstrated that metformin reverses progestin resistance and enhances the rate of cell-killing induced by chemotherapeutic agents in endometrial cancer cells by downregulating GloI expression. Recent study has revealed that aberrant expression of IDH1 contribute to chemotherapy resistance. However, the role of IDH1 in metformin induced chemotherapy sensitivity is not well characterized. As a critical enzyme of the tricarboxylic acid (TCA) cycle, IDH1 catalyzes isocitrate to produce α-ketoglutarate (α-KG), a substrate of TET1, which control TET1-mediated epigenetic modulation of the target genes, including the chemotherapy-resistant genes. In the present study, we have found that IDH1 is overexpressed in endometrial cancers compared with benign endometrial lesions and normal endometria by immunohistochemical analysis of specimens (200 cases of endometrial cancers, 10 cases of benign endometrial lesions and 10 cases of normal endometria). Tissue microarray immunohistochemical detection showed the expression of IDH1, TET1 and 5hmc was positively correlated in different pathological types of endometrial tissue. Interestingly, we found that metformin reduces expression of IDH1 at protein levels with a time- and dose-manners in endometrial cancer cells. Furthermore, overexpression of IDH1 increased TET1 at protein level and blocked metformin-sensitized endometrial cancer to chemotherapy drugs, whereas decrease IDH1 expression lessened TET1 expression and paralleled with the attenuated chemotherapy-resistance. Mechanistic investigations revealed that TET1 involved in metformin-enhanced chemotherapy sensitivity via IDH1 by western blotting and DNA dot blotting assays, this may attribute to that reduced production of α-KG resulted from metformin-suppressing IDH1 is not enough to start TET1-mediated epigenetic modulation of chemotherapy resistant genes. Overall, our findings demonstrate that IDH1 regulates α-KG level and TET1 expression, which in turn affecting metformin mediated endometrial cancer chemotherapeutic sensitivity. Our study suggests IDH1 is a potential target to overcome cisplatin and paclitaxel resistance in endometrial cancer. Citation Format: Mingzhu Bai, Zhenbo Zhang, Yinhua Yu, Youji Feng. Metformin sensitizes endometrial cancer cells to chemotherapy by inhibiting IDH1-TET1 pathway. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 248.

Does experience in hysteroscopy improve accuracy and inter-observer agreement in the management of abnormal uterine bleeding?

Hysteroscopic reliability may be influenced by the experience of the operator and by a lack of morphological diagnostic criteria for endometrial malignant pathologies. The aim of this study was to evaluate the diagnostic accuracy and the inter-observer agreement (IOA) in the management of abnormal uterine bleeding (AUB) among different experienced gynecologists. Each gynecologist, without any other clinical information, was asked to evaluate the anonymous video recordings of 51 consecutive patients who underwent hysteroscopy and endometrial resection for AUB. Experts (>500 hysteroscopies), seniors (20-499 procedures) and junior (≤19 procedures) gynecologists were asked to judge endometrial macroscopic appearance (benign, suspicious or frankly malignant). They also had to propose the histological diagnosis (atrophic or proliferative endometrium; simple, glandulocystic or atypical endometrial hyperplasia and endometrial carcinoma). Observers were free to indicate whether the quality of recordings were not good enough for adequate assessment. IOA (k coefficient), sensitivity, specificity, predictive value and the likelihood ratio were calculated. Five expert, five senior and six junior gynecologists were involved in the study. Considering endometrial cancer and endometrial atypical hyperplasia, sensitivity and specificity were respectively 55.5% and 84.5% for juniors, 66.6% and 81.2% for seniors and 86.6% and 87.3% for experts. Concerning endometrial macroscopic appearance, IOA was poor for juniors (k=0.10) and fair for seniors and experts (k=0.23 and 0.22, respectively). IOA was poor for juniors and experts (k=0.18 and 0.20, respectively) and fair for seniors (k=0.30) in predicting the histological diagnosis. Sensitivity improves with the observer's experience, but inter-observer agreement and reproducibility of hysteroscopy for endometrial malignancies are not satisfying no matter the level of expertise. Therefore, an accurate and complete endometrial sampling is still needed.

Gene x environment interactions impact endometrial function and the menstrual cycle: PROGINS, life history, anthropometry, and physical activity

We assessed the impact of a high frequency, functionally significant allelic variant of the progesterone receptor gene (PROGINS) on endometrial function and menstrual cycle characteristics. Further we asked whether PROGINS moderates the impact of life history characteristics, anthropometric measures, and physical activity on endometrial function. Fifty-two women were genotyped for the PROGINS variant, provided life history information, and had anthropometric measurements made. Women monitored their menstrual bleeding for three cycles, performed mid-cycle urinary ovulation tests, and recorded physical activity. A subset of women provided daily saliva samples and had mid-luteal endometrial thickness measurements taken during the third menstrual cycle. Salivary progesterone was assayed using ELISAs. The direct impact of PROGINS on endometrial and menstrual cycle characteristics was determined via independent t-tests with Bonferroni correction. Interactions between PROGINS and covariates were assessed by moderated regression. PROGINS did not directly impact any indicator of endometrial function. However, PROGINS caused an increase in menstrual cycle length with increasing mid-luteal progesterone levels; the opposite relationship was present in noncarriers (P < 0.05). Additionally, PROGINS interacted with four of six anthropometric measures (BMI, waist circumference, height, and waist-hip ratio) to impact endometrial function, however, interactions between PROGINS and life history variables, or physical activity was limited. The gene x environment interactions we report suggest that PROGINS alters endometrial sensitivity to maternal energetic condition. Thus, the possibility of genetically-based variation in sensitivity to energetic stress should be considered in future adaptive models of women's reproduction.

Abstract 1522: Gene expression profiling (GEP) reveals Epithelial Mesenchymal Transition (EMT) genes selectively differentiating eribulin sensitive breast cancer cell lines.

Abstract Objectives: Eribulin mesylate (ERI) is a simplified synthetic macrocyclic ketone analog of the marine sponge natural product halichondrin B and an inhibitor of microtubule dynamics. In this study, we investigated whether selective signal pathways were associated with ERI activity compared to paclitaxel (PTX), which stabilize microtubules, based on GEP using 3 cancer cell line panels in vitro. Methods: In vitro anti-proliferation activity (IC50) was determined in 27 breast, 19 endometrial and 21 ovarian cancer cell lines treated with ERI and PTX. We performed GEP for the same cancer panels treated for 24 hours at conc. of 10 x IC50 using Affimetrix Human Genome U133 Plus 2.0 arrays. We applied paired t-test (p&amp;lt;0.01,FC&amp;gt;1.5) to identify genes significantly altered between treatments. Pathway enrichment was performed using Metacore and Ingenuity Pathway Analysis software. Cell lines were clustered by hierarchical clustering based on Pearson correlation. Results: We determined the sets of genes differentially altered between ERI and PTX treatments in 3 cancer panels. Regardless of cancer panels, several tubulins (TUBA1B, TUBA1C, TUBA3D, TUBA4A, TUBB2C, TUBBG1, TUBB3, TUBB2A, TUBB2C, TUBB and TUBB6) had significantly lower expression in cell lines treated with ERI, compared to PTX. Our analysis revealed that expression profiles of the gene sets correlated with drug sensitivity in cancer panels (p=0.004 for ERI in breast cancer, p=0.06 for PTX in endometrial cancer, p=0.05 for ERI in ovarian cancer). Pathway enrichment analysis of the gene revealed that the common pathways altered between 2 treatments in 3 cancer panels were related to cytoskeleton remodeling (p&amp;lt;0.002) and role of NEK in cell cycle regulation (p&amp;lt;0.0005). We also identified pathways specific to cancer types. EMT was enriched in genes with significantly altered expression between the two drugs for breast (p=3.4e-07) and endometrial cancer (p=0.001), but not for ovarian cancer. Expression of genes from EMT pathway correlated with breast cancer ERI sensitivity (p=0.1) and endometrial cancer PTX sensitivity (p=0.07). In breast cancer most genes from the EMT signature were up-regulated in ERI resistant cluster, allowing us differentiate cell line sensitivity based on expression. In addition, we found selective signaling pathways were associated to drug sensitivity in breast cancer panels among clustered 394 pathways based on GEP (13 and 4 pathways for ERI and PTX respectively, p&amp;lt;0.01). Conclusion: GEP analysis showed that gene sets altered with ERI and PTX were correlated to drug sensitivity in vitro anti-proliferative activity among 3 cancer panels. Breast cancer is the best target to differentiate ERI sensitivity based on GEP compared to PTX and EMT genes are selectively related to ERI sensitivity. Citation Format: Zoltan Dezso, Judith Oestreicher, Amy Weaver, Tadashi Kadowaki, Mamoru Yanagimachi, Junji Matsui, Sergei I. Agoulnik, Yasuhiro Funahashi. Gene expression profiling (GEP) reveals Epithelial Mesenchymal Transition (EMT) genes selectively differentiating eribulin sensitive breast cancer cell lines. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1522. doi:10.1158/1538-7445.AM2013-1522

Can endometrial arylsulfatase A activity predict the onset of endometrial polyps over the years?

To assess if arylsulfatase A activity (ASA) and sulfatide (SL) concentration in the human endometrium can be predictive of the development of endometrial polyps over the years, since ASA activity reflects the endometrial sensitivity to hormones. ASA activity and SL concentration were determined by biochemical procedures on endometrial samples collected between 1990 and 1994 in non-menopausal women. These women underwent a new endometrial sampling following the clinical indication some years after the first endometrial sampling. The histological assessment of the second endometrial specimens found four patients with normal endometrial pattern and 10 patients with one or more endometrial polyps. ASA activity/years elapsed and SL concentration/years elapsed were compared using two tailed Mann-Whitney test for unpaired data between patients with normal pattern and patients with endometrial polyps. Median ASA activities were 2.62 (normal pattern) versus 1.85 (endometrial polyps) nmol hydrolized substrate/min. Median activity/years elapsed is higher in patients with second endometrial sample presenting normal pattern (p=0.006) and median SL concentration/years elapsed does not differ significantly among groups, even if median SL concentration seems to be higher in patients who subsequently developed polyps (1031 µg/g of fresh tissue versus 341,5 µg/g of fresh tissue). ASA activity can predict the onset of endometrial polyps over the years.

It’s time to pay attention to the endometrium

The physiologic importance of the endometrium in assisted reproductive technology is now receiving much research attention. The historical importance of ultrasound for determination of endometrial pattern, thickness and contractility is reviewed and speculation provided for a possible mechanism of the adverse effect of thin endometrium on implantation.

KSR1 Is Overexpressed in Endometrial Carcinoma and Regulates Proliferation and TRAIL-Induced Apoptosis by Modulating FLIP Levels

The Raf/MEK/extracellular signal-regulated kinase (ERK) pathway participates in many processes altered in development and progression of cancer in human beings such as proliferation, transformation, differentiation, and apoptosis. Kinase suppressor of Ras 1 (KSR1) can interact with various kinases of the Raf/MEK/extracellular signal-regulated kinase pathway to enhance its activation. The role of KSR1 in endometrial carcinogenesis was investigated. cDNA and tissue microarrays demonstrated that expression of KSR1 was up-regulated in endometrial carcinoma. Furthermore, inhibition of KSR1 expression by specific small hairpin RNA resulted in reduction of both proliferation and anchorage-independent cell growth properties of endometrial cancer cells. Because inhibition of apoptosis has a pivotal role in endometrial carcinogenesis, the effects of KSR1 in regulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis were investigated. KSR1 knock-down sensitized resistant endometrial cell lines to both TRAIL- and Fas-induced apoptosis. Sensitization to TRAIL and agonistic anti-Fas antibody was caused by down-regulation of FLIP (FLICE-inhibitory protein). Also investigated was the molecular mechanism by which KSR1 regulates FLIP protein levels. It was demonstrated that KSR1 small hairpin RNA did not affect FLIP transcription or degradation. Rather, FLIP down-regulation was caused by Fas-associated death domain protein-dependent inhibition of FLIP translation triggered after TRAIL stimulation in KSR1-silenced cells. Re-expression of heterologous KSR1 in cells with down-regulated endogenous KSR1 restored FLIP protein levels and TRAIL resistance. In conclusion, KSR1 regulates endometrial sensitivity to TRAIL by regulating FLIP levels.

Effect of medroxy-progesterone acetate on follicular growth and endometrial cycloxygenase-2 (COX-2) expression during the bovine estrous cycle

The objective of this study was to evaluate the effect of medroxy-progesterone acetate (MAP) with or without estradiol benzoate (EB) on follicular growth during the estrous cycle in cattle. In the first experiment, Hereford cows were synchronized with a synthetic analogue of PGF2 alpha and were treated with two different doses of MAP (250 or 500 mg) with or without EB for 7 days starting on day 8 of the estrous cycle. Follicular growth was inhibited (P&lt;0.05) in all cows except controls and those receiving 250mg MAP without EB. Seventy-five percent of the animals (15/20) showed estrus on days 21 and 22 of the cycle rather than at MAP withdrawal, demonstrating that these treatments did not induce estrus. To determine whether the EB treatment altered endometrial sensitivity to oxytocin and thus the luteolytic cascade, multiparous pre-synchronized cows received 5 mg of EB followed 6 hours later with 50 IU of oxytocin (OT; n=9). Eight hours after EB injection, endometrial fragments were collected from the cows on days 4, 13 and 17 of the estrous cycle and COX-2 gene expression was measured by PCR. EB increased COX-2 mRNA levels only on day 17 of the estrous cycle (P&lt;0.05). In conclusion, MAP alone or associated with EB is able to suppress bovine follicular growth. However, EB in the presence of MAP is not efficient to induce luteolysis in cows when injected on day 8 of the estrous cycle.

Abstract 1645: The phosphorylation status of BAD pathway influence to chemo-sensitivity

Abstract Objective: Patients with advanced or recurrent endometrial cancer have a poor prognosis in part due to lack of efficacy of currently available chemotherapies and incomplete understanding of the biology that underlies chemo-response. In the current study we used genomic expression data to identify molecular signaling pathways associated with endometrial cancer cell resistance to cisplatin, doxorubicin chemotherapy and further characterized the role of the BAD apoptosis pathway as a therapeutic target and determinant of chemo-response. Methods: Eight endometrial cell lines were subject to genome-wide expression analysis using Affymetrix HG-U133Plus GeneChips and in parallel, ranked by relative sensitivity to cisplatin and doxorubicin using MTS cell proliferation assays. Gene array data from sensitive versus resistant cell lines was compared by student's t-test and Pearson's correlation to identify those genes associated with drug resistance and further evaluated by GeneGo Metacore software for representation of molecular signaling pathways. Furthermore, the role of the BAD pathway in chemo-resistance was evaluated by selective inhibition of BAD pathway members, PKA, PP2C, AKT1, AKT2 and AKT3 by siRNA or pharmacologic agents in the presence and absence of cisplatin. Cell death and growth arrest in the presence of cisplatin were evaluated by Western blot for PARP cleavage as well as nuclear condensation and fragmentation and MTS assays. Results: Student's t-test and Pearson's correlation identified the expression of 325 and 1136 genes to be associated with cisplatin (p&amp;lt;0.01) and doxorubicin (p&amp;lt;0.01) resistance, respectively. Pathway analysis of genes associated with both cisplatin and doxorubicin resistance indicated significant representation of the BAD apoptosis pathway (p&amp;lt;0.002). Selective depletion of the phosphatase, PP2C by siRNA resulted in increased phosphorylation of BAD, serine 118 and increased resistance to cisplatin induced cell growth arrest and cell death. In contrast, depletion of the kinase, PKA resulted in decreased phosphorylation of BAD, serine 118 and increased cell death in the presence of cisplatin. In addition, depletion of AKT1 using siRNA as wells as treatment of cells with the AKT inhibitor, triciribine, increased cisplatin-induced growth arrest and cell death, whereas depletion of AKT2 or AKT3 did not affect cisplatin sensitivity. Conclusions: The phosphorylation status BAD may influence endometrial cancer cell sensitivity to chemotherapy and represents a compelling target for future therapeutic development. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1645.

Favorable influence of local injury to the endometrium in intracytoplasmic sperm injection patients with high-order implantation failure

Local injury to the endometrium prior to controlled ovarian stimulation may considerably improve implantation rates and pregnancy outcomes in intracytoplasmic sperm injection patients with high-order implantation failure (> or =4 IVF trials and > or =12 transferred embryos).

The effect of undernutrition on the establishment of pregnancy in the ewe

The relationship between nutrition and reproduction in sheep has been the subject of research in several international groups. This review will particularly focus on the effects of undernutrition on the potential causes of reproductive failure including abnormalities of the ovum or the embryo, luteal inadequacy and failure of the supply of progesterone to the uterus, or the mechanisms involved in maternal recognition of pregnancy. The level of nutrition and peripheral progesterone concentrations are inversely related, and increased rates of embryo loss, associated with higher progesterone concentrations in ewes with low levels of nutrition have been reported. Undernutrition may act through changes in the distribution of progesterone in the endometrium. Thus, lower endometrial levels on day 5 of the cycle in ewes fed half of their maintenance requirements have been observed, providing a link between the known role of progesterone in embryo survival by the modulation of uterine function and the higher embryo losses found in undernourished ewes. The evidence of an effect of maternal nutrition on IFNtau secretion from the conceptus and of PGF2alpha production from the uterus is presented. Moreover, undernutrition provokes a reduction in the sensitivity of the endometrium to progesterone that may affect embryo survival. Finally, a state of undernutrition induces changes in the endometrial sensitivity to steroid hormones at early stages of pregnancy that could adversely alter uterine environment to the detriment of embryo survival.

Androgen and estrogen receptors and co-regulators levels in endometria from patients with polycystic ovarian syndrome with and without endometrial hyperplasia

Objective. To study if the endocrinological status of PCOS women affects the endometrial sensitivity to steroids by evaluating the expression of androgen receptor (AR), estrogen receptor α (ERα), estrogen receptor β (ERβ), co-activators AIB1 and ARA70, and co-repressor NCoR. Methods. Gene and/or protein expression of steroid receptors and co-regulators was measured in 17 samples of normal endometrium (NE), 23 PCOS endometrium without treatment (PCOSE), 11 endometria from PCOS women and with endometrial hyperplasia (HPCOSE), and 10 endometria from patients with endometrial hyperplasia (HE), using RT-PCR and/or immunohistochemistry and Western blot. Results. Gene and protein expression of AR was relatively elevated in PCOSE and HPCOSE compared with NE. A significant increase in ERα protein expression was observed in PCOSE, preferentially in the nucleus of endometrial cells, whereas ERβ gene and protein expression increased gradually from PCOSE to HPCOSE and HE, mainly in the epithelial compartment. Importantly, we found a gradual increase in the ERβ/ERα gene and protein expression ratio in endometria from the four groups of women. AIB1 showed increased nuclear protein expression in PCOSE compared to NE, in the presence of a high expression of ARA70 in all groups. High expression of ARA70 together with a normal expression level of AIB1 was observed in HPCOSE. The cytoplasmic immunostaining of NCoR was similar between the four groups of patients. Conclusion. The PCOS endometrium exhibits a higher sensitivity to steroid action. We can inferred that these alterations could deregulate the transcription of genes involved in the cell cycle, which may lead to the development of endometrial hyperplasia in PCOS women.

Antifertility Activity of Artabotrys odoratissimus Roxb. and Couroupita guianensis Aubl.

Objective: To study antifertility activity of various extracts of A. odoratissimus Roxb. and C. guianensis Aubl. Materials and methods: Antifertility activity of benzene, ethanol and water extracts of leaves of A. odoratissimus and bark and flowers of C. guianensis was studied for their effect on duration of various stages of oestrus cycle in adult female rats and on the number implantation sites in the pregnant rats. Results: All the extracts of A. odoratissimus leaf and ethanol and water extracts of C. guianensis bark and flower showed significant prolongation of dioestrus stage (p<0.05). Ethanol and water extracts of A. odoratissimus leaf showed anti-implantation effect. Ethanol extract of C. guianensis bark and all the extracts of its flower reduced the number of implantations. Conclusion: The disturbances in the reproductive cycle indicate effect of these plants on the sex hormones in the animals. Prevention of implantation may be due their interference in the receptive stage of the uteri and endometrial sensitivity for decidualisation. The results suggest that the both plants are having antifertility activity and possible therapeutic use.

A mechanistic assessment of 1,3-butadiene diepoxide-induced inhibition of uterine deciduoma proliferation in pseudopregnant rats

Butadiene diepoxide (BDE), a reactive metabolite of 1,3-butadiene that is an important industrial chemical used in synthetic rubber production causes a dose-dependent inhibition of deciduoma development in pseudopregnant Sprague-Dawley rats. This study used 4 daily i.p. BDE doses of 0.20, 0.25, 0.30, 0.35, or 0.40 to characterize mechanisms that may be responsible for the antideciduoma effect. Pseudopregnant rats were treated either before (pseudopregnancy [PPG] days 1–4) or after (PPG days 5–9) deciduoma induction by endometrial trauma with a blunt needle. Animals were killed on PPG day 9 and evaluated for serum progesterone and endometrial protein and DNA. RT-PCR was used to measure message for estrogen receptor (ER) α and pituitary adenylate cyclase-activating polypeptide (PACAP). Substrate zymography and Western blotting were used respectively to measure matrix metalloproteinase (MMP)-9 and inducible nitric oxide synthase. The antideciduoma effects of BDE were associated with decreases in endometrial weight, protein, and DNA, with decreases in serum progesterone, and with decreases in PACAP message and MMP-9. A reduction in NOS was identified at the highest dose of BDE. Message for estrogen receptor (ER) α was not affected at any dose. We conclude that the reduction in decidual proliferation was direct and appeared to be associated with either 1) a decrease in the effectiveness of the deciduogenic stimulation and/or a weakened endometrial sensitivity to the stimulus; or 2) an effect on deciduoma development. Molecular mechanisms that apparently contributed to BDE inhibition of decidual metabolism included the synthesis of protein and DNA involved in decidual growth, the synthesis and activation of a matrix metalloproteinase for degradation of the extracellular matrix that is essential for tissue remodeling during deciduoma development, and the nitric oxide/nitric oxide synthase and pituitary adenylate cyclase-activating peptide systems that are involved in promoting vasodilation and increased vascular permeability to enhance the availability of substrates for maximal deciduoma growth. The ovotoxicity of BDE, which has previously been established, may indirectly affect decidual proliferation by reducing progesterone, the preeminent endocrine regulator of deciduoma development. The findings also suggest that BDE may possess no estrogenic action since it was associated with endometrial weight loss and unaltered levels of the estrogen receptor α mRNA expression.

Uterine luminal epithelial alkaline phosphatase activity and pinopod development in relation to endometrial sensitivity in the rat

The period of maximal endometrial sensitivity in the rat was characterized by high alkaline phosphatase activity in uterine luminal and glandular epithelium and endometrial stroma. The activity in endometrial stroma increased following decidualization. Pinopod development on the endometrial surface was first observed during the presensitivity period. Their number increased, apparently more so on the antimesometrial rather than the mesometrial segment of the uterus, on the day of maximal sensitivity. Inhibition in endometrial sensitivity by single anti-implantation (1.25 mg/kg, po) dose of centchroman on day 1 post-coitum (p.c.), although it did not affect alkaline phosphatase activity on days 2 and 3 p.c., caused complete inhibition in its activity in uterine luminal and glandular epithelium and pinopod development on days 4 and 5—coinciding, respectively, with time of entry of preimplantation embryos into the uterus and period of maximal endometrial sensitivity in this species. Significant decrease in enzyme activity was also evident in the entire endometrial stroma and myometrium, except blood capillaries, on these days. In comparison, prevention of entry of native embryos into the uterus by placing a ligature at the utero–tubal junction had no effect on pinopod development, but caused marked decrease in enzyme activity in luminal and glandular epithelium only during the immediate postimplantation period. The uterine lumen on the day of maximal sensitivity in centchroman-treated rats appeared highly distended and was lined with tall columnar epithelium, in comparison to low cuboidal epithelium in controls. The findings demonstrate: (a) a correlation between uterine luminal epithelial alkaline phosphatase activity and endometrial sensitivity; (b) complete inhibition in enzyme activity in luminal and glandular epithelium following centchroman treatment might be related to altered permeability characteristics of epithelial cells, which together with the absence of pinopods and highly distended uterine lumen on the day of maximal sensitivity, suggest inhibition of endocytosis/pinocytosis of luminal fluid, luminal closure, apposition of blastocyst trophoblast to luminal epithelium, and secretory activity of glandular epithelium; (c) pinopod development on the endometrial surface was independent of presence of viable blastocysts in utero; and (d) complete absence of pinopods suggests lack of endometrial sensitivity, but their presence might not necessarily indicate a sensitized endometrium in the rat.

Uterine estradiol and progesterone receptor concentration, activities of certain antioxidant enzymes and dehydrogenases and histoarchitecture in relation to time of secretion of nidatory estrogen and high endometrial sensitivity in rat.

Alterations in uterine nuclear and cytosolic estradiol (ER) and progesterone (PR) receptor concentration, activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), glucose-6-phosphate dehydrogenase (G-6-PDH) and lactate dehydrogenase (LDH), surface and transmission electron microscopy and histology in relation to the time of secretion of nidatory estrogen and the onset of endometrial sensitivity in the rat were investigated. A significant increase in plasma estradiol (E2) concentration in control rats was observed at 22.00 h on day 4 post-coitum, whereas progesterone (P) concentration increased at 17.00 h on day 4 and was maintained until 17.00 h on day 5. The period of high endometrial sensitivity (10.00 h on day 5) was characterized by elevated uterine cytosolic ER and nuclear and cytosolic PR concentration and POD activity, low columnar luminal epithelium with undulating surface and intercellular membranes, covered with short microvilli and pinopods, and containing numerous electron-transparent apical vesicles, mitochondria, polyribosomes, rough (RER) and smooth (SER) endoplasmic reticulum, well developed Golgi, few lysosomes and lipid droplets and loose edematous antimesometrial stroma. Inhibition in endometrial sensitivity by post-coital centchroman was associated with a marked depletion in uterine cytosolic ER and an increase in nuclear ER concentration, a decrease in POD and G-6-PDH activities, compact fibroblastic stroma, an increase in luminal epithelial cell height with decreased RER, SER, polyribosomes, Golgi, straightening of intercellular membranes, reduced surface undulations and absence of pinopods. Electron-transparent vesicles appeared flattened and clumped in the apical portion of cells, tight junctions were more prominent and lipid droplets were translucent. Nuclear and cytosolic PR and the pattern of secretion or plasma E2 and P remained unaffected. CAT, SOD and LDH activities, although high throughout pre-implantation, did not vary in relation to the secretion of nidatory estrogen, endometrial sensitivity or centchroman treatment.

Correlation of pinopod development on uterine luminal epithelial surface with hormonal events and endometrial sensitivity in rat.

Intrinsic role of preovulatory and nidatory estrogen and progesterone and presence of viable blastocysts in utero in pinopod development on the uterine luminal epithelial surface and correlation between time of their development and onset of endometrial sensitivity were investigated. In adult rats, pinopods were observed on the entire epithelium even before secretion of nidatory estrogen, i.e. at 14.00 h on day 4 post-coitum (p.c.). Apparently, their number increased, more so on the antimesometrial than the mesometrial side, at 10.00 h on day 5, but were fewer and mostly collapsed at 10.00 h on day 6. Pinopods on day 4 were located within epithelial depressions and foldings, but protruded from the surface on days 5 and 6. Normal pinopods were also present on day 8 p.c. in rats under delayed implantation, but an implantation-inducing dose of estradiol-17 beta administered about 18 h earlier caused their collapse like that on day 6 in intact rats. Development and appearance of pinopods in intact or delayed rats was unaffected when native preimplantation embryos were prevented from entering the uterus. Normal pinopods were seen in immature rats receiving progesterone for at least 3 days or cyproterone acetate for 4 days, but not after estradiol alone. In animals receiving progesterone or priming/sensitizing estradiol in addition to progesterone, the decidual response was suboptimal, irrespective of the presence of pinopods on the day of stimulation. In animals in which a condition mimicking preimplantation had been produced by suitable hormone supplementation, optimal endometrial sensitivity and decidual response were elicited, even though most pinopods appeared collapsed, resembling those on day 6 in intact rats and about 18 h after estradiol in implantation-delayed rats. Findings confirm that pinopod development on uterine luminal epithelium was dependent on progesterone alone and demonstrate that: (i) preovulatory (priming) or nidatory (endometrial sensitizing) estrogen or viable blastocysts in utero have no role in their development. Nidatory estrogen, instead, appears to limit pinopod development by causing their collapse; (ii) pinopod development/presence on the endometrial surface might indicate the uterus coming into a period of sensitivity rather than actually being in it and might thus serve as a useful marker of "transfer window" rather than "implantation window"; (iii) in the rat, pinopod development might serve as an alternate assay for evaluation of progestational activity of newer test agents.

Time-related effects of a triphenylethylene antiestrogen on estrogen-induced changes in uterine weight, estrogen receptors, and endometrial sensitivity in rats

Time-related estrogen antagonistic action of a single oral contraceptive (1.25 mg/kg) dose of the triphenylethylene antiestrogen centchroman was determined in ovariectomized immature rats. Tamoxifen and nafoxidine were used for comparison. A single oral administration of centchroman followed by three doses of estradiol-17β (1 μg/d, s.c.) caused significant dose-dependent inhibition in estradiol-17β-induced increase in uterine weight and nuclear and cytosolic estrogen receptors. But the inhibition at anti-implantation dose was evident only if estradiol-17β treatment was initiated not later than 48 h post-antiestrogen. Alternatively, when antiestrogen treatment was followed by a single dose of estradiol-17β between days 2–7, a synergistic action, typical of antiestrogens possessing weak estrogen agonistic activity, was observed. In immature rats in which a condition mimicking preimplantation was produced by estradiol-17β (0.5 μg/d, s.c.) priming on days -2 and -1, followed by progesterone (1 mg/d, s.c.) and an endometrial sensitizing dose (0.5 μg/d, s.c.) of estradiol-17β at 1600 h on day 4, anti-implantation dose of centchroman administered on day 1, too, failed to inhibit uterine weight gain induced by sensitizing dose of estradiol-17β, but caused marked inhibition in endometrial sensitivity to a deciduogenic stimulus and decidualization and weight gain of traumatized uterine horn 96 h post-traumatization over non-traumatized horn was only about 150% (725% in controls). Inhibition in endometrial sensitivity and decidualization was evident when the interval between antiestrogen treatment and sensitizing estradiol was <126 h. Pinopods were present on endometrial surface on day 5 whether or not priming and/or sensitizing doses of estradiol were administered, but decidual response was mild if either of these doses of estradiol-17β was deferred. Findings suggest that: (a) duration of antiestrogenic action of single anti-implantation dose of centchroman in rat was about 126 h, which in ovariectomized immature rats was evident only when a condition mimicking preimplantation was produced and the antiestrogenic response was based on inhibition in estradiol-induced endometrial sensitivity and not uterine weight gain; (b) priming as well as sensitizing estrogen were essential to get optimal decidual responses; (c) appearance of pinopods on endometrial surface may not be related to endometrial sensitivity; and (d) tamoxifen and nafoxidine appear slightly longer acting with duration of antiestrogenic action of ∼150 h.