Endometrial Surface Epithelium Research Articles

Human Chorionic Gonadotropin Enhances Trophoblast–Epithelial Interaction in an In Vitro Model of Human Implantation

Embryo implantation, which is an absolute requirement for reproduction, starts with blastocyst apposition to the uterine endometrium, followed by attachment to the endometrial surface epithelium. Recent clinical studies reported an increase in implantation and pregnancy rates among women receiving intrauterine human chorionic gonadotropin (hCG) prior to embryo transfer suggesting that, at least in some cases, female infertility is a result of inadequate secretion of hCG. In this study, we characterized the effect of hCG on trophoblast-epithelial interaction by further developing our recently described in vitro model of implantation. Here, we confirmed hCG increased attachment of trophoblast to epithelial cells, using a single-cell trophoblast-epithelial coculture system in addition to a blastocyst-like spheroid-epithelial coculture system. Furthermore, we discovered that the source and concentration was pivotal; the first preparation of hCG affected 2 molecules related to implantation, MUC16 and osteopontin, while the second preparation required additional cytokines to mimic the effects. Using this system, we can develop a comprehensive knowledge of the cellular and gene targets of hCG and other factors involved in embryo apposition and implantation and potentially increase the number of therapeutic targets for subfertile patients.

Temporal analysis of prostaglandin F2α receptor, caspase 3, and cyclooxygenase 2 messenger RNA expression and prostaglandin F2α receptor and cyclooxygenase 2 protein expression in endometrial tissue from multiparous Nelore (Bos taurus indicus) cows treated with cloprostenol sodium during puerperium

The use of cloprostenol sodium in puerperium is questionable, as both favorable and unfavorable responses during the uterine involution process have been reported in the literature. This study is based on the hypothesis that cloprostenol sodium promotes modifications in the prostaglandin F2α receptor (FP), caspase 3 (CASP-3), and cyclooxygenase 2 (COX-2) mRNA expression that may favor the process of postpartum uterine involution in multiparous Nelore (Bos taurus indicus) females. Additionally, we aimed to describe the presence and immunolocalization of the FP and COX-2 protein in endometrial tissue at different postpartum time points in these females. Multiparous Nelore cows (n = 24) were treated with cloprostenol sodium (n = 12) or saline solution (n = 12) on postpartum Days 1 and 4 (Day 0 = birth), and endometrial biopsies were performed with a Yomann biopsy instrument and collected on Days 1, 7, 14, 28, and 42 postpartum. The mRNA expression from samples on the Days 1, 7, 14, 28, and 42 and the protein expression from samples on the Days 1, 14, 28, and 42 were then analyzed. The treated cows had altered FP and CASP-3 mRNA expression, and FP and COX-2 protein were observed in the endometrial surface epithelium, the stroma, and the glandular epithelium, with cytoplasmic immunolocalization. Although we attribute the change in CASP-3 mRNA expression to physiological phenomena, the results obtained for FP mRNA expression opens new doors for the study of hormonal protocols associated with cloprostenol sodium in the puerperium of Zebu females.

Immunohistochemical expression of vascular endothelial growth factor receptors Flt1 and KDR in the endometrium during the estrus cycle in albino rats

Background Angiogenesis is an important process in endometrial development and embryonic implantation and is regulated through vascular endothelial growth factor (VEGF); its receptors Flt1 and KDR. Aim This work aimed to study the immunoexpression of VEGF receptors (VEGF-Rs) in the endometrium at different ages and reproductive phases and correlate them with the histological profiles in these phases. Materials and methods Seventy female albino rats were included in this study. They were divided into seven groups of 10 rats each: one group consisted of rats in the prepubertal period at age 4–6 weeks; five groups consisted of rats in the reproductive period at age 6–10 months, which were divided according to estrus cycle phases into proestrus, estrus, metestrus, diestrus, and pregnant groups; and the sixth group consisted of rats in the postmenopausal period at age 15–18 months. The uteri of all rats were removed and processed for staining with H&E and were subjected to immunohistochemical staining for Flt1 and KDR. For morphometric measurements, uterine wall thickness and Flt1 and KDR optical density in the endometrial surface epithelium, glandular epithelium, stromal cells, and endometrial endothelial cells were measured using image analysis. Results were statistically compared. Results The expression of VEGF-Rs was highest in the pubertal age group with marked expression of these receptors in the proestrus phase followed by the estrus phase. This supports the role of sex hormones, especially the estrogen hormone, in regulating VEGF-R expression. The Flt1 receptor was predominantly expressed in endometrial and stromal cells as well as in blastocysts, whereas the KDR receptor was predominantly expressed in endometrial endothelial cells. Comparison among all groups and then between each two groups revealed statistically significant differences in the measured morphometric parameters. Conclusion The upregulation of Flt1 and KDR could be involved in the regulation of endometrial endothelial cell proliferation and in increase in endometrial vascular permeability, especially at implantation sites.

Biosynthesis and Degradation of Canine Placental Prostaglandins: Prepartum Changes in Expression and Function of Prostaglandin F2alpha-Synthase (PGFS, AKR1C3) and 15-Hydroxyprostaglandin Dehydrogenase (HPGD)1

There is no distinct explanation of the mechanism for the prepartal prostaglandin F2alpha (PGF2alpha) increase in pregnant dogs. Although the PGF2alpha-synthase (PGFS [AKR1C3]) mRNA expression and localization profiles have been previously investigated in canine utero/placental compartments, the availability and biochemical activity of the PGFS (AKR1C3) protein remain unknown. In order to better understand the regulation of canine uterine PGF2alpha availability and eventual prepartum release in luteolytic amounts in dogs, canine-specific PGFS (AKR1C3) and 15-hydroxyprostaglandin dehydrogenase (HPGD) antibodies were generated and used to characterize the expression, cellular localization, and biochemical properties of PGFS (AKR1C3) and HPGD in the utero/placental compartments and corpus luteum throughout pregnancy and at prepartum luteolysis. PGFS (AKR1C3) expression was weak or absent in luteal samples. Uterine PGFS (AKR1C3) was up-regulated postimplantation and declined prepartum. The utero/placental expression of PGFS (AKR1C3) was identified in the superficial uterine glands throughout gestation and in the trophoblast cells within the feto-maternal contact zone during placentation, suggesting a possible role for PGFS (AKR1C3) in the trophoblast invasion. Utero-placental HPGD was up-regulated until postimplantation, lower at midgestation, and greatly suppressed at prepartum. Expression was routinely identified in the endometrial surface and glandular epithelia, and positive signals were also observed in the trophoblast cells at the feto-maternal contact zone. The biochemical activity of recombinant PGFS (AKR1C3) and HPGD was confirmed after its expression in a heterologous system. The colocalization of HPGD with PGFS (AKR1C3) expression suggests a modulatory role for HPGD as a gatekeeper of the supply of prostaglandin in the pregnant canine uterus.

Trophinin and integrin β3 expression in the human endometrium. A pilot study

The study involved ten patients with tubal factor infertility. Patients (Group I, age 28–34 years, n=5; Group II, age 35–40 years, n=5) underwent endometrial biopsy on postovulatory days 7–10 of the natural menstrual cycle. Endometrial biopsies were studied by histological methods, presence of trophinin and integrin β<sub>3</sub> in the endometrial surface epithelium and in the glandular epithelium was assessed by immunohistochemistry. Low levels of β<sub>3</sub> integrin and trophinin were estimated in the endometrial epithelial cells in most patients, particularly in Group II, which reflects reduced endometrial capacity for adhesion and points to problems with the implantation of the embryo.

Estrus synchronization affects WNT signaling in the porcine reproductive tract and embryos

The purpose of the study was to investigate an effect of estrus synchronization with prostaglandin (PG) F2α and PMSG/hCG on WNT4, WNT5A, WNT7A, β-catenin (CTNNB1) and E-cadherin (CDH1) gene expression. The weight of the uterus, morphometrical parameters of the endometrium and the number of CL were recorded. The analysis of estradiol (E2), prostaglandin (PG) F2α and E2 content in the uterine luminal flushings (ULFs) and progesterone (P4) level in the blood serum were conducted. RNA was isolated from endometrial, luteal and embryonic tissue of pregnant non-synchronized (Control; n = 15) and pregnant synchronized (PGF2α/PMSG/hCG; n = 15) pigs. Whereas there was no change in uterine weight, differences in height of endometrial surface and glandular epithelium were found. However, height of the endometrium, number of the glands and capillaries were unaffected. The total number of the CLs was higher (P < 0.05) in animals treated with PGF2α/PMSG/hCG. The amount of E2 and P4 was lower (P < 0.05, P < 0.001, respectively) in pregnant gilts administrated with PGF2α/PMSG/hCG. The concentration of PGF2α in ULFs was not affected by hormonal management, while PGE2 was higher (P < 0.01) in hormonally in comparison to non-hormonally treated pigs. The content of WNT4 mRNA in conceptuses increased on particular Days studied in Control and PGF2α/PMSG/hCG administered animals. WNT7A and CTNNB1 were affected by PGF2α/PMSG/hCG treatment in both conceptuses (P < 0.001, P < 0.05) and endometrial tissue (P < 0.001, P < 0.01). The PGF2α/PMSG/hCG treatment resulted in elevated expression of WNT4 (P < 0.001) and CTNNB1 (P < 0.05) in luteal tissue in comparison to the Control gilts. Moreover, luteal amount of WNT5A mRNA was higher in PGF2α/PMSG/hCG animals in comparison to the Control group (P < 0.05). Presented data show that exogenous hormones administration can affect gene expression in the porcine reproductive tract and embryo.

Leptin expression in proliferative, secretory and hyperplastic endometrial tissues

The goal of this study was to detect endometrial leptin expression in proliferative and secretory phases and then to compare the results with that of hyperplastic endometrium. Seventeen proliferative, 23 secretory phase and 18 hyperplastic endometrial tissues diagnosed in our hospital between 2002 and 2007 were included in the study. These samples were stained with leptin antibody using an immunohistochemical method. Endometrial glandular and surface epithelium and stroma were evaluated for staining distribution and intensity. Staining intensity seen in early proliferative phase samples (2.33±0.51) increased significantly throughout the middle (2.40±0.54) and late phases (2.83±0.40) (p<0.05). Early secretory phase samples had the least staining intensity (1.42±0.53), while it increased significantly in later periods (2.38±0.51) (p<0.05). There was no difference in staining intensity among proliferative, secretory and hyperplastic tissues (p>0.05). Although endometrial leptin expression was observed in a differential manner throughout the whole menstrual period, no difference was seen in endometrial hyperplasia. We consider that leptin does not play a role in hyperplastic transformation of the endometrium.

Differential expression of Toll-like receptor 4 (TLR4) in healthy and infected canine endometrium

This study provides the first report into immunohistochemical localization of Toll-like receptor (TLR) in the canine reproductive tract. TLR4 was investigated in endometrium during the estrous cycle and in pyometra. Pyometra is the most important pathological condition of the uterus due to bacterial infection in dogs. To protect against invading pathogens, the female reproductive tract has evolved immune mechanisms. TLRs are the cellular components of the afferent arm of the innate immune system. The expression of TLR4 was significantly higher in the endometrial stroma compared to the endometrial surface epithelium and glandular epithelium in proestrus. The glandular epithelium and stroma at the diestrous stage expressed TLR4 significantly higher than surface epithelium. Furthermore, when compared to other healthy groups, the glandular epithelium at diestrus also higher expressed TLR4 than other stages. The expression of TLR4 in the surface epithelium was higher in dogs with pyometra compared with all other groups. And, the surface epithelium of dogs suffering from pyometra also expressed TLR4 more intensely than the glandular epithelium. The innate immunity of infected canine endometrium response to bacterial infection is intensely extremely increased by the expression of TLR4. Furthermore, the different levels of TLR4 expression seems related to physiological changes in distinct cell types of endometrium, leukocytes populations, cytokines and sex hormones.

Proteomic analysis identifies interleukin 11 regulated plasma membrane proteins in human endometrial epithelial cells in vitro

BackgroundDuring the peri-implantation period, the embryo adheres to an adequately prepared or receptive endometrial surface epithelium. Abnormal embryo adhesion to the endometrium results in embryo implantation failure and infertility. Endometrial epithelial cell plasma membrane proteins critical in regulating adhesion may potentially be infertility biomarkers or targets for treating infertility. Interleukin (IL) 11 regulates human endometrial epithelial cells (hEEC) adhesion. Its production is abnormal in women with infertility. The objective of the study was to identify IL11 regulated plasma membrane proteins in hEEC in vitro using a proteomic approach.MethodsUsing a 2D-differential in-gel electrophoresis (DIGE) electrophoresis combined with LCMS/MS mass spectrometry approach, we identified 20 unique plasma membrane proteins differentially regulated by IL11 in ECC-1 cells, a hEEC derived cell line. Two IL11 regulated proteins with known roles in cell adhesion, annexin A2 (ANXA2) and flotillin-1 (FLOT1), were validated by Western blot and immunocytochemistry in hEEC lines (ECC-1 and an additional cell line, Ishikawa) and primary hEEC. Flotilin-1 was further validated by immunohistochemistry in human endometrium throughout the menstrual cycle (n = 6-8/cycle).Results2D-DIGE analysis identified 4 spots that were significantly different between control and IL11 treated group. Of these 4 spots, there were 20 proteins that were identified with LCMS/MS. Two proteins; ANXA2 and FLOT1 were chosen for further analyses and have found to be significantly up-regulated following IL11 treatment. Western blot analysis showed a 2-fold and a 2.5-fold increase of ANXA2 in hEEC membrane fraction of ECC-1 and Ishikawa cells respectively. Similarly, a 1.8-fold and a 2.3/2.4-fold increase was also observed for FLOT1 in hEEC membrane fraction of ECC-1 and Ishikawa cells respectively. In vitro, IL11 induced stronger ANXA2 expression on cell surface of primary hEEC and ECC-1 whilst, the lipid-raft protein FLOT1 demonstrated punctate staining in the apical surface of ECC-1 plasma membranes and was upregulated in the epithelium in the receptive phase of the menstrual cycle (p lower or equal 0.05).ConclusionsThis is the first study to use a proteomics approach to identify hEEC plasma membrane proteins that may be useful as infertility markers or pharmacological targets for fertility regulation.

Effect of copper intrauterine device on the cyclooxygenase and inducible nitric oxide synthase expression in the luteal phase endometrium

Background To evaluate the effect of copper intrauterine device (IUD) on the expression of cyclooxygenase (COX) and inducible nitric oxide synthase (iNOS) in the luteal phase endometrium. Study Design A prospective clinical study was conducted on 30 women who were willing to use a copper IUD contraception. Endometrial biopsies and blood samples were taken before and 3 months after the insertion of the IUD on Day 3 and Days 20–24 of the cycle. Main outcome measures were to evaluate the effect of copper IUD on uterine artery blood flow using pulsed color Doppler ultrasonography and the relationship of bleeding abnormalities and menstrual pain level with the uterine blood flow, COX-2 and iNOS expression. Results Only the left uterine artery pulsatility and resistance indices decreased statistically significantly (p=.005 and p=.039, respectively). Other Doppler parameters showed no change. Cyclooxygenase-2 expression of both endometrial luminal epithelium (p=.03) and gland epithelium (p=.03) increased significantly. Inducible NOS expression of the endometrial surface epithelium decreased significantly after IUD insertion (p=.01). Conclusions Although COX-2 expression increased 3 months after copper IUD insertion, iNOS expression of the luminal epithelium decreased. Local hypoxia caused by copper and vasoconstrictor prostanoids may play a role in IUD-related menstrual abnormalities.

Thin layer chromatographic fraction of root extract of Polygonum hydropiper induces vaginal epithelial cell maturation in adult ovariectomized albino rat

Dry root powder of Polygonum hydropiper is traditionally used as an antifertility agent by women of north east India. The effect of crude root extract of P. hydropiper on reproductive performance of female albino rat was reported earlier from this laboratory. In the present investigation, attempt was made to find estrogenic effect, if any, of thin layer chromatographic fraction of the crude extract. To achieve the goal, methanolic crude extract was fractionated in thin layer chromatography (TLC). Estradiol-17β was used as the reference compound to identify the estrogenic compound in the crude root extract. The effect of TLC fraction was studied on the vaginal epithelial cell maturation in adult ovariectomized (OVX) rats and uterine histology of OVX and ovary-intact females. The TLC fraction was administered (sc) to adult OVX females at a dose of 5mg/kg/ day for 18 days. Estradiol-17β was injected as reference compound. Vaginal smears were stained with Giemsa's to observe the cell types. Rats were sacrificed by cervical dislocation at the end of the treatment and the uterine epithelium was subjected to histological analysis. The TLC fraction treated OVX females showed characteristic epithelial cell types (parabasal, intermediate, superficial and cornified) in the smear. Histological evidence revealed increased proliferation of the endometrial surface epithelium in the TLC fraction treated females. The results reveal that TLC fraction of root extract of Polygonum hydropiper contains compound(s) which has functional similarities with E2 and ovarian estrogen in female albino rats.

An integrated view of L‐selectin and trophinin function in human embryo implantation

Determining molecular mechanisms of human embryo implantation is an extremely challenging task due to the limitation of materials and significant differences underlying this process among mammalian species. Recently, L-selectin and its ligand carbohydrate have been proposed as a system that mediates initial adhesion of human blastocysts to the uterine epithelia. We have also identified trophinin as a unique apical cell adhesion molecule potentially involved in the initial adhesion of trophectoderm of the human blastocyst to endometrial surface epithelia. In the mouse, the binding between ErbB4 on the blastocyst and heparin-binding epidermal growth factor-like growth factor on the endometrial surface enables the initial step of the blastocyst implantation. The evidence suggests that L-selectin and trophinin are included in human embryo implantation. This review summarizes findings relevant to the functions of L-selectin and trophinin in human embryo implantation, and proposes a model that reconciles these cell adhesion mechanisms.

Induction of trophinin in human endometrial surface epithelia by CGβ and IL-1β

During embryo implantation, trophinin mediates cell adhesion by homophilic binding at the apical surfaces of trophectoderm and endometrium. Trophinin is expressed on the human endometrial epithelia in rare occasions. We developed hCG-coated agarose beads that mimic the physical and physiological features of an implantation-stage human blastocyst. When hCG-coated beads were applied to human endometrial epithelial cells in the presence of IL-1β, endometrial cells acquired strong trophinin expression and the ability for apical cell adhesion with trophinin-expressing human trophoblastic cells. These results provide a mechanism for trophinin-mediated adhesion of human blastocyst to endometrium by a spatially and temporally restricted paracrine effect of hCG derived from the blastocyst.

Class I histone deacetylase expression in the human cyclic endometrium and endometrial adenocarcinomas

Class I histone deacetylases (HDACs) and acetylases (HATs) are members of transcriptional pre-initiation complexes assembled by steroid hormone receptors. Recently, HDAC inhibitors were shown to enhance differentiation of endometrial fibroblasts and endometrial adenocarcinomas. However, there is only rare information on HDAC and HAT expression in the human endometrium. HDAC-1, -2, -3 and HAT (PCAF and GCN5) mRNA expression was studied in tissue from premenopausal women undergoing hysterectomy by real-time or semiquantitative RT-PCR. HDAC protein expression was assessed by Western Blot and immunohistochemistry. In endometrial adenocarcinomas (n = 17), HDAC-1 expression was studied by immunohistochemistry. In the human endometrium, HDAC-1, -2, -3 and PCAF mRNA are expressed without cyclical changes. Western blot analysis demonstrated that HDAC-2 protein expression was slightly, but significantly elevated in the secretory phase (P < 0.01 versus day 5-8), whereas HDAC-1 and -3 protein expression was constitutive throughout the menstrual cycle. By immunohistochemistry, nuclear expression of HDAC proteins was detected in all endometrial cell types. In the case of HDAC-3, immunostaining was significantly reduced in the endometrial surface epithelium on day 6-10 (P < 0.01 versus days 15-18 and 24-28). Compared to normal endometrium, a high proportion of endometrial adenocarcinomas showed impaired HDAC-1 protein expression in the epithelial and stromal compartment. Class I HDACs and HATs are expressed in the human endometrium throughout the menstrual cycle, suggesting the cyclic endometrium as a potential target for HDAC inhibitors. We hypothesis that alterations of HDAC and/or HAT expression are potentially involved in impaired endometrial differentiation.

Mid-luteal endometrial intracrinology following controlled ovarian hyperstimulation involving use of a gonadotrophin releasing hormone antagonist

There are concerns of reduced pregnancy rates with the use of gonadotrophin-releasing hormone antagonists (GnRH antagonists) in IVF/ICSI cycles. Sex steroids and their metabolizing enzymes in the endometrium may play a vital role in embryo implantation. This study has evaluated the levels and localization of sex-steroid receptors and metabolizing enzymes, 3beta-hydroxysteroid dehydrogenases (3betaHSD) and selected 17beta-HSD (17betaHSD), in mid-luteal endometrium of women treated with GnRH antagonist (Cetrorelix) and recombinant FSH (rFSH; Gonal-F) with luteal phase progesterone supplementation. Mid-luteal phase endometrial biopsies were obtained from oocyte donors undergoing ovarian stimulation and from control women with regular periods. Immunohistochemistry and real-time quantitative-polymerase chain reaction (QRT-PCR) were used to compare protein and mRNA expression of progesterone receptor (PR), estrogen receptor alpha (ERalpha), estrogen receptor beta (ERbeta), androgen receptor (AR), 3betaHSD1, 3betaHSD2, 17betaHSD2 and 17betaHSD5. Cetrorelix-rFSH treatment caused a mid-luteal suppression of PR protein expression in the endometrial stroma, surface epithelium and glands, although expression in the glands of control samples was variable. In contrast, the treatment caused an increase in PR staining in perivascular cells. No other significant differences in protein expression were observed between the two groups. mRNA levels of AR, ERalpha, 3betaHSD1 and 17betaHSD2 were significantly reduced in the treatment group. PR mRNA levels were also reduced by GnRH antagonist-rFSH treatment, but the difference was not significant. Changes in the expression of sex-steroid receptors and metabolizing enzymes may lead to alterations in the activity and intracellular availability of estrogens, progestogens and androgens in endometrium of women treated with Cetrorelix and rFSH. Their impact on embryo implantation merits further evaluation.

Immunohistochemical expression of endometrial L-selectin ligand is higher in donor egg recipients with embryonic implantation

To correlate L-selectin ligand (LSL) expression in human endometrium with embryonic implantation. Retrospective cohort analysis. University-based fertility center. Donor egg recipients (DERs) who underwent programmed hormonal replacement for ET with prior mock cycle luteal phase endometrial biopsy. Immunohistochemical expression of LSL using MECA-79 antibody was examined. Slides were scored with a new scoring system, the IHC-Level (range 0-4) as follows: strength of staining-absent (0), weak (1), or strong (2); plus distribution of staining-absent (0), <50% of tissue (1), and >50% (2). Cellular apex and cytoplasm were scored independently in both the endometrial glandular and surface epithelium. Endometrial LSL expression in pregnant versus nonpregnant patients. MECA-79 IHC-Level of the apex of surface epithelium was significantly higher for pregnant versus nonpregnant DERs (3.8 vs. 3.4). When controlling for embryo morphology, there continues to be a significant difference in apex score on surface epithelium (3.8 vs. 3.3, respectively). The new scoring system results correlated with an established scoring system, the HSCORE. We demonstrate significantly higher expression of LSL at the apex of human endometrial surface epithelium obtained from DERs with embryonic implantation. Furthermore, we present the IHC-Level, a method of evaluating immunohistochemistry that may be applied to other markers of endometrial receptivity.

Steroid Receptor Expression and HER‐2/neu (c‐erbB‐2) Oncoprotein in the Uterus of Cats with Cystic Endometrial Hyperplasia–Pyometra Complex

The presence of oestrogen-alpha receptor (ER), progesterone receptor (PR), and HER-2/neu (c-erbB-2) oncoprotein in the uterine walls of 10 healthy cats and 20 subjects with cystic endometrial hyperplasia-pyometra (CEH-P) were evaluated. Lesions were graded according to the severity of cystic dilation, hyperplasia and inflammation, and were classified as normal, mild uterine hyperplasia and severe uterine hyperplasia. The ER, PR and c-erbB-2 expression in the endometrium, glandular epithelium, stromal fibroblasts and myometrial smooth muscle cells was quantified by immunohistochemistry. The ER, PR and c-erbB-2 staining patterns differed between normal uteri and uteri with CEH-P. The ER expression was tended to be higher in the endometrial surface and glandular epithelium in the severe hyperplasia group (P > 0.05) and significantly lower in the mild hyperplasia cases compared with normal endometrium (P < 0.05), whereas the PR expression in both severe and mild hyperplasia cases tended to be higher in stromal cells and glandular epithelium than those in the normal uteri. C-erbB-2 immunoreactivity was observed only in the endometrial surface and glandular epithelium of the uterine wall and immunostaining was found to be highest in cases with severe hyperplasia. As a conclusion, we suggest that c-erbB-2 oncoprotein may play a role in the pathogenesis of the CEH together with the ER and PR in cats, and that ER does not have a role in the mechanism of pyometra, whereas PR plays a role in the pathogenesis of both CEH and pyometra.

Endometrial glandular and stromal breakdown, part 2: Cytomorphology of papillary metaplastic changes

Careful cytomorphologic evaluation of abnormal endometrial lesions has made accurate and reproducible microscopic assessment possible. Histopathology of patients with dysfunctional uterine bleeding due to an anovulatory cycle usually contain endometrial glandular and stromal breakdown (EGBD) and papillary metaplasia on the endometrial surface epithelium, if an appropriate sample has been collected. We often recognized abnormal cell clumps in the cytological smears with EGBD cases. They were composed of metaplastic cells, and some irregular small projection figures were observed from the margins of the cell clumps. We describe the quantitation of metaplastic clumps with irregular protrusions (MCIP) in endometrial endocyte samples. The current study presents the cytomorphological characteristics of the metaplastic changes recognized in EGBD cases. During a 7-yr period, 144 cases for which histopathological diagnoses were obtained following endometrial curettage, after collecting a direct endometrial sample using the endocyte. The material comprised 49 cases of normal proliferative endometrium (NPE) (patients aged 28-51, average 39.9), 32 cases of EGBD (patients aged 30-67, average 49.6), and 63 cases of endometrial hyperplasia without atypia (EH) (patients aged 35-65, average 47.7). The following points were investigated: (1) the occurrence of metaplastic cells; (2) the occurrence and the frequency of MCIP; and (3) the occurrence of MCIP that contains condensed stromal clusters. Metaplastic cells were seen in 93.8% of the EGBD cases. Cytomorphologic pattern identified with MCIP was 90.6%, and its frequency showed 16.1%. The occurrence of MCIP that contain condensed stromal clusters (93.1%) showed a strong association in comparison with other lesions, such as NPE and EH. Our data appear to indicate that the appearance of MCIP with condensed stromal clusters originated from the papillary metaplasia, which occurred on the endometrial surface epithelium. The cytologic observation of those cells may be a useful indicator for providing the nature of EGBD endometrium.

Canine Endothelial Remodelling During the Estrus Cycle: an Overview

The present study in 58 dogs aimed to verify the changes of the canine endometrial surface epithelium during the estrous cycle using electron microscopy and to investigate the roles of apoptosis, cell proliferation and matrix metalloproteinases in canine endometrial remodelling using immunohistochemistry. Possible correlations between these phenomena and serum levels of estradiol‐17β and progesterone were assessed. Prominent scanning electron microscopic changes of the canine endometrium were observed during early metestrus and included a variable height and number of microvilli of the surface epithelium. Using transmission electron microscopy, pronounced features of the surface epithelial cells were noticed in late metestrus and anestrus, during which a highly vacuolized cytoplasm and deformed nuclei due to fat accumulation were present. In these stages no prominent signs of cell death were noticed and also very low apoptotic numbers were observed using immunohistochemistry. Cell proliferation in the endometrial surface epithelium seemed to be in balance with apoptosis. Most prominent features of apoptosis and proliferation were found in the basal glands and to a lesser extent also in the crypts. In contrast, only low amounts of apoptosis and proliferation were observed in the stroma, in which high collagenase‐1 scores coincided with a decreased amount of collagen in the deep part of the endometrium and a massive increase of glandular cells during early metestrus. No cyclic differences were noticed for the expression of TIMP‐1, the collagenase‐1 inhibitor. Finally, no or only low correlations were found between apoptosis, cell proliferation and MMP‐1 expression in the canine endometrium, and the serum levels of estradiol‐17β and progesterone. In conclusion, these findings indicate that there is no desquamation of the canine endometrial surface epithelium at the end of the estrous cycle, but a regular turnover of cells throughout the estrous cycle. Furthermore, our data also suggest a role for MMP‐1 in cellular proliferation.This study was supported by the BOF (#011B4101) of Ghent University.

Morphometric changes in the endometrium and serum leptin levels during the implantation period of the embryo in the rat in response to exogenous ovarian stimulation

To determine the changes in endometrium and serum leptin levels during the periimplantation period of the ovum in the rat in response to exogenous ovarian stimulation by hMG and recombinant FSH (rFSH). Experimental animal study. University hospital. A total of 100 female rats were stimulated with low and high doses of hMG (groups 1 and 2, respectively) and low and high doses of rFSH (groups 3 and 4, respectively) before mating after the regular cycle was observed. The control group of animals was not injected (group 5). The endometrium of the pregnant rats (n = 53) was analyzed by the morphometric method, and serum samples were obtained for leptin and E2 levels at baseline and on the day of implantation. The mitotic index was determined separately for endometrial surface, glandular epithelium, and endometrial stroma by light microscope. Morphometric analysis of endometrium, mitotic index, and serum leptin and E2 levels The mean number of corpora lutea was significantly higher in groups 2 and 4. The measurement of mucosal depth was found to be significantly higher in group 4 compared with the control group (0.361 +/- 0.03 vs. 0.289 +/- 0.04). The depth of the endometrial surface epithelium was found to be significantly higher in all study groups (groups 1-4) compared with the control group. The depth of the surface epithelium was found to be significantly lower in group 1 compared with the other study groups (groups 2, 3, and 4). The depth of the surface epithelium was also significantly lower in group 2 compared with group 3 (21.58 +/- 3.56 vs. 28.21 +/- 3.56). Mitotic indices that were determined at the endometrial surface epithelium and stroma were all significantly lower in all study groups (groups 1-4) compared with the control group. There was no significant increase for the baseline leptin levels compared with the levels measured on the day of implantation. In an animal model, exogenous administration of gonadotropins significantly affects the morphology of the endometrium and the mitotic index in the implantation period of the embryo. These morphological effects become more pronounced as we increase the administered dose of exogenous gonadotropins. However, no differences in leptin levels were observed relative to different gonadotropin type and dose regimens.