Evaluation of cell proliferation and endometrial thickness of bitches in different periods of diestrus

ABSTRACT This study aimed to determine the histological features of the endometrium of bitches, as well as the cell proliferation at specific moments of diestrus, 10, 20, 30, 40, 50 and 60 days post ovulation, correlating the endometrial thickness with the uterine cell proliferation and the metabolic state (weight, blood glucose and plasma cholesterol) of the animals. Therefore, the right and left uterine horns of 26 clinically healthy bitches submitted to ovariohysterectomy were histologically analyzed 10, 20, 30, 40, 50 and 60 days post ovulation. The hematoxylin-eosin and AgNOR staining techniques were performed. All parameters were evaluated by ANOVA and post-hoc Tukey test (p<0.05). The correlation between endometrial thickness and uterine cell proliferation, weight, blood glucose and plasma cholesterol of animals was observed using the Pearson method (p<0.05). In the present study, it is concluded that endometrial thickness does not differ at any of the moments analyzed in diestrus. The endometrial thickness is not influenced by hormones, weight, blood glucose or serum cholesterol of bitches in this phase of the estrous cycle. However, there is greater cell proliferation in the endometrium at day 40 compared to day 60 post ovulation under the influence of the endocrine profile.

Differential expression of upstream stimulatory factor (USF) 2 variants in eutopic endometria from women with endometriosis: estradiol regulation

BACKGROUND: Endometriosis, pro-inflammatory and invasive benign disease estrogen dependent, abnormally express in endometria the enzyme P450Arom, positively regulated by steroid factor-1 (SF-1). Our objective was to study the nuclear protein contents of upstream stimulating factor 2 (USF2a and USF2b), a positive regulator of SF-1, throughout the menstrual cycle in eutopic endometria from women with and without (control) endometriosis and the involvement of nuclear estrogen receptors (ER) and G-coupled protein estrogen receptor (GPER)-1. RESULTS: Upstream stimulating factor 2 protein contents were higher in mid (USF2b) and late (USF2a and USF2b) secretory phase in eutopic endometria from endometriosis than control (p < 0.05). In isolated control epithelial cells incubated with E2 and PGE2, to resemble the endometriosis condition, the data showed: (a) significant increase of USF2a and USF2b nuclear protein contents when treated with E2, PPT (specific agonist for ERa) or G1 (specific agonist for GPER1); (b) no increase in USF2 binding to SF-1 E-Box/DNA consensus sequence in E2-treated cells; (c) USF2 variants protein contents were not modified by PGE2; (d) SF-1 nuclear protein content was significantly higher than basal when treated with PGE2, E2 or G1, stimulation unaffected by ICI (nuclear ER antagonist); and (e) increased (p < 0.05) cytosolic protein contents of P450Arom when treated with PGE2, E2, PPT or G1 compared to basal, effect that was additive with E2 + PGE2 together. Nevertheless, in endometriosis cells, the high USF2, SF-1 and P450Arom protein contents in basal condition were unmodified. CONCLUSION: These data strongly suggest that USF2 variants and P450Arom are regulated by E2 through ERa and GPER1, whereas SF-1 through GPER1, visualized by the response of the cells obtained from control endometria, being unaffected the endogenously stimulated cells from endometriosis origin. The lack of E2 stimulation on USF2/SF-1 E-Box/DNA-sequence binding and the absence of PGE2 effect on USF2 variants opposite to the strong induction that they exert on SF1 and P450 proteins suggest different mechanisms and indirect regulations. The sustained USF2 variants protein expression during the secretory phase in eutopic endometria from women with endometriosis may participate in the pathophysiology of this disease strongly associated with infertility and its characteristic endometrial invasion to ectopic sites in the pelvic cavity.

A Study of Endometrial Pathology by Transvaginal Color Doppler Ultrasonography and its Correlation with Histopathology in Post-menopausal Women.

Objective: Endometrial carcinoma is the most common gynaecological malignancy. Approximately 80% of endomentrial carcinoma occur in post-menopausal women. Present study aimed to evaluate the role of transvaginal ultrasonography and colour flow imaging in diagnosing endometrial pathologies especially endometrial carcinoma and later its confirmation by histopathology. Methods: 38 women presenting with history of at least 6 months amenorrhoea followed by bleeding per vaginum were included in the study. Transvaginal colour Doppler (TVS) followed by fractional curettage was done in all cases and cervical biopsy was done in selected cases. Uterine size, endometrial thickness and blood flow indices (RI, PI) were measured. Analysis of data was done using ‘z’ test and ‘t’ test. Results: Out of 38 women maximum number of cases (39.47%) were between 50 – 55 years. Using 4 mm of endometrial thickness as cut off value for discriminating normal and abnormal endometrium, sensitivity, specificity, PPV and NPV were 94.12%, 50%, 95.12% and 50% respectively (p < 0.05). No case of endometrial carcinoma was detected when the endometrium was <4 mm, making the sensitivity as 100%, NPV 100%, specificity 13.33% and PPV 23.53%. Using RI = 0.81 as cut off value for discriminating benign and malignant endometrium, sensitivity was 62.5%, specificity 53.33%, PPV – 26.3% and NPV as 84.2%. Conclusion: Transvaginal sonographic (TVS) evaluation of endometrial thickness (ET) is a reliable method of screening women with post-menopausal bleeding. Conservative approach may be offered to women showing ET of less than 4 mm and high impedance to flow in uterine and endometrial vessels.

Curcumin inhibits endometriosis endometrial cells by reducing estradiol production

Endometriosis is a complex estrogen-dependent disease that is defined as the presence of endometrial gland and stroma outside the uterine cavity. Although the exact mechanism for the development of endometriosis remains unclear, there is a large body of research data and circumstantial evidence that suggests a crucial role of estrogen in the establishment and maintenance of this disease. This study is an attempt to assess the effect of curcumin on inhibiting endometriosis endometrial cells and to investigate whether such an effect is mediated by reducing estradiol production. Endometriotic stromal cells, normal endometrial stromal cells, endometriotic epithelial cells and normal endometrial epithelial cells were isolated and cultured. E[2] value of cells and the effect of curcumin on cell proliferation were evaluated. Finally, effect of curcumin on E[2] assay was detected. Electrochemiluminescence immunoassay results showed that E[2] value of endometriotic epithelial cells was higher than the endometriotic stromal cells [p=0.037], while the expression of E[2] in normal endometrial stromal and epithelial cells was extremely low. WST-8 result showed, compared with endometrial stromal cells, ectopic endometriotic stromal cells had a higher growth rate. After intervene with curcumin [10micromol/L, 30micromol/L and 50micro mol/L] for 0-96h, the number of endometriotic stromal cells was reduced and cells growth slowed, compared with 0micromol/L group. Compared with 0micromol/L group, E[2] level was lower after treatment with curcumin, especially in 30micromol/L and 50micromol/L group. In summary, in this study we found that E[2] is important in ectopic endometrium, and epithelial cell is in dominant position with E[2] secretion. Curcumin was able to suppress the proliferation of endometrial cells by reducing the E2 value.

Detecção imunoistoquímica de receptores de estrógeno e progesterona no endométrio de vacas Nelore (Bos taurus indicus) durante o anestro pós-parto / Immunohistochemical detection of receptors for estrogen and progesterone in endometrial glands and stroma during the pospartum anestrus in Nelore (Bos taurus indicus) cows

Foram utilizadas 24 vacas Nelore P.O., em anestro pós-parto, diagnosticado pelo histórico reprodutivo, por avaliações ultrassonográficas transretais e por dosagem de progesterona plasmática, que foram submetidas à colheita de fragmento uterino via transcervical. Os animais foram divididos em dois grupos conforme o máximo diâmetro folicular: grupo 1: folículos <6mm (n=12); grupo 2: folículos >6mm (n=12). Para avaliar receptor de estrógeno e receptor de progesterona no epitélio glandular e no estroma, foi utilizada a técnica de imunoistoquímica. Altas contagens relativas e alta intensidade de marcação para receptor de estrógeno e progesterona no epitélio glandular e no estroma foram observadas nos dois grupos. No entanto, a intensidade de marcação para o receptor de progesterona no epitélio glandular foi mais alta no grupo 2 comparado ao grupo 1. Quando o epitélio glandular e o estroma foram comparados, o número relativo de receptor de estrógeno no grupo 1 foi mais alto no epitélio glandular comparado ao estroma, e a intensidade de marcação para o receptor de progesterona no grupo 2 foi mais alta no epitélio glandular comparado ao estroma. Os resultados sugerem que os mecanismos que controlam a expressão de receptores no anestro são semelhantes aos observados durante o ciclo estral.

Twenty-four postpartum anestrous Nelore purebred cows were used in the study. Anestrous was determined based on the reproductive history which was confirmed in each cow based on plasma progesterone concentration and by transrectal ultrasonography. Endometrial biopsies were collected. The animals were separated into two groups according to maximum follicular diameter- Group 1: follicles <6mm (n=12) and Group 2: follicles >-6mm follicles (n=12). The immunohistochemistry technique was employed to evaluate the presence of estrogen and progesterone receptors in the uterine glandular epithelium and stroma. High counts of positive nuclei and high intensity of immunostain for estrogen and progesterone receptors in the glandular epithelium and stroma were observed in the two groups. However, the immunostain intensity of progesterone receptors in the glandular epithelium was higher in Group 2 compared to Group 1. When glandular epithelium and stroma were compared within each group, the relative number of estrogen receptors in the Group 1 was higher in the glandular epithelium compared to stroma and the immunostain intensity for the progesterone receptor in Group 2 was higher in the glandular epithelium compared to stroma. The results suggest that the mechanisms that control the expression of endomerial receptors in the anestrus are similar to the ones observed during the estrus cycle.

Histochemical study of early embryo implantation in rats / Estudio histoquímico de la implantación temprana en ratas

Embryo implantation is the process that results in attachment of the conceptus to the uterine wall. In this histochemical study, we investigated the early stage of embryo implantation in rats by morphological analysis and by the detection of total proteins and glycosaminoglycans using hematoxylin-eosin, toluidine blue at pH 4.0 (TB), and Xylidine Ponceau at pH 2.5 (XP). In non-pregnant females, the uterine layers could be clearly distinguished and showed the normal histology of the organ. In pregnant females, an increase in the number of cells and a reduction in the interstitial space were observed in the endometrium close to the implantation sites. The blastocyst was partially inserted in the endometrium, with the observation of the inner cells mass around the blastocyst cavity surrounded by trophoblastic cells. TB staining revealed mild metachromatic basophilia, which was more evident in the endometrial stroma around the implantation site. Histochemical staining with XP was also more intense in the stroma close to the site of implantation. On the other hand, histochemical staining with either TB or XP was more discrete at sites distant from the conceptus. This study demonstrated changes in the endometrial stroma in areas adjacent to the site of embryo implantation, with variations in glycosaminoglycans and proteins as demonstrated by the detection of anionic and cationic radicals, respectively.

La implantación embrionaria es el proceso que resulta en la unión del embrión a la pared uterina. En este estudio histoquímico, se determinó la fase inicial de implantación del embrión en ratas mediante el análisis morfológico y por la detección de proteínas totales y glicosaminoglicanos con hematoxilina-eosina, azul de toluidina a pH 4,0 (TB) y xilidina Ponceau a un pH de 2,5 (XP). En las hembras no preñadadas, las capas del útero pueden ser claramente distinguidas y mostraron la histología normal del órgano. En las hembras preñadas, se observa un incremento en el número de células y una reducción en el espacio intersticial del endometrio para cerrar los sitios de implantación. El blastocisto se implanta parcialmente en el endometrio, con presencia de masa de células internas en torno a la cavidad del blastocisto, rodeado por las células trofoblásticas. La tinción TB reveló leve basofilia metacromática, lo cual fue más evidente en el estroma endometrial alrededor del sitio de implantación. Tinción histoquímica con XP también fue más intensa próximo del estroma en el sitio de implantación. Por otro lado, la tinción histoquímica, ya sea con la tuberculosis o XP fue más leve en los lugares distantes del embrión. Este estudio demostró cambios en el estroma del endometrio en las zonas adyacentes al sitio de la implantación del embrión, con variaciones en glucosaminoglicanos y proteínas, como lo demuestra la detección de radicales aniónicos y catiónicos, respectivamente.

Sildenafil effect on nitric oxide secretion by normal human endometrial epithelial cells cultured in vitro

Sildenafil is a selective inhibitor of cyclic-guanosine monphosphat-specific phosphodiesterase type 5. It increases intracellular nitric oxide [NO] production in some cells. There are reports on its positive effect on uterine circulation, endometrial thickness, and infertility improvement. Endometrial epithelial cells [EEC] play an important role in embryo attachment and implantation. The present work investigates the effect of sildenafil on human EEC and their NO secretion in vitro. In this experimental in vitro study, endometrial biopsies [n=10] were washed in a phosphate buffered solution [PBS] and digested with collagenase I [2 mg/ml in DMEM/F12 medium] at 37°C for 90 minutes. Epithelial glands were collected by sequential filtration through nylon meshes [70 and 40 micro m pores], respectively. Epithelial glands were then treated with trypsin to obtain individual cells. The cells were counted and divided into four groups: control and 1, 10, and 20 micro M sildenafil concentrations. Cells were cultured for 15 days at 37°C and 5% CO[2]; the media were changed every 3 days, and their supernatants were collected for the NO assay. NO was measured by standard Greiss methods. Data were analyzed by one way ANOVA. There was no significant difference between groups in cell count and NO secretion, but the level of NO increased slightly in the experimental groups. The 10 micro M dose showed the highest cell count. EEC morphology changed into long spindle cells in the case groups. Sildenafil [1, 10, and 20 micro M] showed a mild proliferative effect on human EEC numbers, but no significant change was seen in NO production

Detection of PTEN immunoreactivity in endometrial hyperplasia and adenocarcinoma.

OBJECTIVE: To investigate PTEN (phosphatase and tensin homolog deleted on chromosome 10) expression in endometrial hyperplasia and adenocarcinoma as analyzed by immunohistochemistry. MATERIAL AND METHOD: PTEN protein expression was evaluated by immunohistrochemical study of 70 paraffin-embedded curettage endometrial tissue samples (10 normal endometrium, 55 endometrial hyperplasia, and 15 endometrial adenocarcinomas) selected from surgical pathology files of the Division of Gynecologic Pathology, Department of Obstetrics and Gynecology, Faculty of Medicine, Chulalongkorn University, from 2001 to 2004. Intensity of epithelial staining of PTEN immunoreactivity in different histologic types was determined. RESULTS: Absence of PTEN protein expression was detected in 60% of endometrial carcinoma, 60% of atypical endometrial hyperplasia, and 24% of typical endometrial hyperplasia. In endometrial hyperplasia without atypia group, the majority of cases revealed moderate to strong PTEN expression, with 70% in simple hyperplasia and 47% in complex hyperplasia. There is a significant statistical difference of PTEN immunoreactivity among proliferative endometrium, endometrial hyperplasia and endometrial carcinoma group (p = 0.004). CONCLUSION: Complete loss of PTEN protein expression was most commonly found in endometrial carcinoma and hyperplasia with cytologic atypia.

N-myc downstream-regulated gene 1 as a downregulated target gene of PTEN in the controlling of tumourigenesis in endometrioid carcinoma.

BACKGROUND & OBJECTIVE: Mutation/deletion of PTEN has been known to be involved in the development of many cancers including endometrial carcinoma. NDRG1 (N-myc downstream-regulated gene 1) is reported to be associated with tumourigenesis. PTEN expression has been shown to be correlated with NDRG1 in both prostate and breast cancer. In this study, we explored the possibility that PTEN alteration may cause carcinogenesis of endometrioid carcinoma by regulating the expression of the NDRG1 gene. METHODS: Tissue blocks of 103 patients with pathologically confirmed endometrioid carcinoma were included. All the carcinoma tissues were accompanied with varied degree of necrosis. Using two-step method and avidin-biotin peroxidase complex immunohistochemistry method, the correlation of the two genes expression in ischaemic area and the relationship of NDRG1 expression between ischaemic and non-ischaemic area in endometrioid carcinomas was evaluated. RESULTS: PTEN alteration and NDRG1 expressions were significantly increased in the ischaemic area of endometrioid carcinoma compared with their expressions in the normal endometrium respectively (P<0.001, P<0.001). A positive correlation was found between PTEN alteration and NDRG1 expression in the ischaemic area of endometrioid carcinoma. INTERPRETATION & CONCLUSION: We suggest that NDRG1 may be an important candidate gene in facilitating endometrium carcinogenesis in the adaptation of hypoxia for survival. Alteration of PTEN may upregulate NDRG1 expression, which plays an important role in the process leading to endometrial carcinogenesis.

Effect of human chorionic gonadotropin (hCG) on expression of vascular endothelial growth factor a (VEGF-a) in human mid-secretory endometrial cells in three-dimensional primary culture.

Several lines of evidence suggest that human uterine endometrial cells can bind human chorionic gonadotropin (hCG) which, in turn, influences the physiology of implantation stage endometrium. Vascular endothelial growth factor (VEGF) appears to be a candidate mediator in this process. However, our knowledge about hCG action on VEGF in human endometrial cells is very thin. In the present study, we have examined microscopically hCG binding to dissociated human endometrial cells collected from mid-luteal phase and maintained in three-dimensional primary co-culture on rat-tail collagen type I biomatrix and examined the effect of different concentrations (0, 1, 10, 100 and 1000 IU/ML) of hCG on VEGF expression and secretion by endometrial cells maintained in the above system. We report that both cytokeratin positive epithelial cells as well as vimetin positive stromal cells from human mid luteal phase endometrium could bind hCG and that their number increased (P < 0.01) steadily with time. Administration of hCG enhanced (P < 0.05) immunoreactive VEGF protein expression in dose dependent manner in endometrial cells retrieved from mid-luteal phase of cycle, and co-cultured in a three-dimensional cell culture system, but with no marked change in VEGF secretion. Collectively, it appears that hCG influences VEGF protein synthesis in human midluteal phase endometrial cells, but has little effect on post-translational regulation and secretion. From physiological homeostasis point of view, it is likely that synthesis and secretion of VEGF exhibits a modular and factorial regulation to achieve a fine tuning of this potent vasotropic agent in receptive stage endometrium.

Passagem de células endometriais para a cavidade peritoneal durante histeroscopia diagnóstica / Dissemination of endometrial cells into the peritoneal cavity during diagnostic hysteroscopy

OBJETIVO: avaliar a passagem de células endometriais para a cavidade peritoneal durante histeroscopia diagnóstica. MÉTODOS: estudo descritivo, prospectivo, envolvendo 61 pacientes sem afecção endometrial maligna e 15 com câncer do endométrio. Duas amostras de lavado peritoneal foram colhidas, uma antes (LP-1) e outra (LP-2), imediatamente após a realização da histeroscopia diagnóstica. A passagem para a cavidade peritoneal foi definida como a presença de células endometriais no LP-2, devendo tais células estarem ausentes no LP-1. Utilizou-se histeroscópio com 5 mm de diâmetro (Storz). O meio de distensão foi o CO2 com pressão de fluxo de 80 mmHg controlada eletronicamente. O LP foi fixado em álcool absoluto (1:1). As lâminas foram preparadas pelo método de Papanicolaou e todas as leituras feitas pelo mesmo observador. RESULTADOS: foram excluídas quatro pacientes (5,26 por cento) por apresentarem células endometriais no LP-1, sendo duas em cada grupo. Nas 72 restantes, não houve passagem de células para a cavidade peritoneal. No grupo sem afecção maligna endometrial, 88,1 por cento (52/59) apresentaram endométrio secretor, com correlação de 80 por cento entre o diagnóstico histeroscópico e a biópsia do endométrio. No grupo com afecção maligna endometrial, a maioria das pacientes encontrava-se no estádio I (92,3 por cento). A correlação entre histeroscopia/biópsia endometrial e exame anatomopatológico da peça cirúrgica foi de 100 por cento. CONCLUSÕES: a realização de histeroscopia diagnóstica com CO2 e pressão de fluxo de 80 mmHg não determinou passagem de células endometriais para a cavidade peritoneal em ambos os grupos, sugerindo que a histeroscopia é método seguro nas pacientes com suspeita de câncer endometrial.

PURPOSE: to evaluate the spreading of endometrial cells to the peritoneal cavity during diagnostic hysteroscopy. METHODS: a prospective, descriptive study involving 76 patients divided in two groups: one with 61 patients without malignant endometrial cancer, and the other with 15 patients with endometrial cancer. Two samples of peritoneal fluid were collected, one before (PF-1) and the other immediately after (PF-2) the diagnostic hysteroscopy. Spread to the peritoneal cavity was defined by the presence of endometrial cells in PF-2, with the absence of such cells in PF-1. The 5 mm diameter Storzs hysteroscopy was used. Distention was obtained by CO2 with electronically controlled flow pressure of 80 mmHg. The PF was fixated in absolute alcohol (ratio1:1). The PF samples were centrifuged and aliquots were smeared and stained using the Papanicolaou method. Analyses were performed by the same observer. RESULTS: during the study, four patients (5.26 percent) were excluded for presenting endometrial cells in PF-1. In the remaining 72 patients, there was no spread of cells to the peritoneal cavity. In the non-endometrial cancer group, 88.1 percent (52/59) presented secretory endometrial phase, with correlation of 80 percent between the hysteroscopy and the biopsy. In the group with endometrial cancer, most of the patients were in stage I (92.3 percent). There was a 100 percent correlation between the hysteroscopy/biopsy and histopathology of the surgical sample. CONCLUSIONS: the diagnostic hysteroscopy with CO2 at flow pressure of 80 mmHg did not cause spread of endometrial cells to the peritoneal cavity in both groups, thus suggesting that the diagnostic hysteroscopy is safe for patients at high risk for endometrial cancer.

Evaluation of the macroscopic growth degree of experimental endometriosis in rats

PURPOSE: To evaluate macroscopically the growth degree of self-transplantation of endometriosis in rats. METHODS: Forty female rats, after a 7-day period for adpating and evaluating of the estrous cycle regularity, underwent tail abdominal midline laparotomy with 3-cm cuts. The average third of the left uterine horn was removed, 4mm x 4mm patches in liquid environment were made, and self-transplanted in the rat mesenterium with a single stitch, and the endometrial surface of the endometriotic implant facing the lumen of the peritoneal cavity. The rats were programmed to die after three weeks. The abdominal cavity displaying was held and self-transplants were identified and classified. RESULTS: The results achieved were: one case for degree 0 (2,5 percent), three cases for degree 1 (7,5 percent), eleven cases for degree II (27,5 percent) and twenty-five cases for degree III (62,5 percent). CONCLUSION: The experimental endometriosis development, through the self-transplantation technique, showed to be most common in degrees 3 and 2 of development.

OBJETIVO: Avaliar macroscopicamente o grau de crescimento de autotransplantes de endometriose em ratos. MÉTODOS: Quarenta ratos fêmeas, após período de sete dias para adaptação e avaliação da regularidade do ciclo estral, foram submetidas à laparotomia mediana abdominal caudal com incisões de três cm. Foi retirado o terço médio do corno uterino esquerdo, feito retalhos de quatro mm x quatro mm em meio líquido, sendo em seguida autotransplantado no mesentério da rata com ponto simples, tomando o cuidado de manter a superfície mucosa voltada para luz abdominal. Após o período de três semanas as mortes das ratas foram programadas. Realizou-se a exposição da cavidade abdominal com identificação e classificação dos autotransplantes. RESULTADOS: os resultados encontrados foram: grau 0 obteve um caso (2,5 por cento), o grau I foi observado em três casos (7,5 por cento), o grau II com onze casos (27,5 por cento) e o grau III foi visto em vinte cinco casos (62,5 por cento). CONCLUSÃO:Desenvolve-se a endometriose experimental pela técnica do autotransplante com a maioria dos casos em grau 3 e 2 de desenvolvimento.

Endometrial biopsy during induction of ovulation with tamoxifen polycystic ovary syndrome

The dichotomy between ovulation rates and pregnancy rates for women with polycystic ovary syndrome [PCOS] treated with tamoxifen prompted the present study to determine the effect of tamoxifen on endometrial maturity. To compare endometrial histology of PCOS patients on induction of ovulation with tamoxifen with controls. Prospective case-control study. Bab El-Shareya University Hospital, Cairo, Egypt. Sixty-seven anovulatory women with PCOS on their third ovulatory cycle of tamoxifen were compared with controls [n=70] healthy but subfertile regularly ovulating women whose husbands had abnormal semen parameters. All subjects had an endometrial biopsy in the late luteal phase. Endometrial histology classified according to the classical Noyes criteria revealed out-of-phase endometrium in 11/67 [16%] of the tamoxifen group compared with 2/70 [3%] in controls [p<0.001]. The incidence of out-of-phase endometrium seems to increase with higher pre-induction LH levels. Tamoxifen treatment significantly increases the prevalence of out-of-phase endometrium and this could explain, in part, the large difference between ovulation and pregnancy rates. The higher the pre-induction LH levels, the more is the possibility of the endometrium being out-of-phasestandard 12 lead ECG, laboratory investigations including CPK, IMA, CRP, lipogram, kidney and liver functions were done. Twenty two patients were diagnosed as non ischaemic chest pain [NICP] and 51 cases as ACS. CPK and troponin levels were normal in all groups at presentations but 6 hours later CPK levels were significantly higher in ACS patients if compared with NICP or control groups [p<0.000]. on the other hand, IMA levels were statistically significantly high in ACS group only [p<0.000] with a good negative predictive value to diagnose NICP [86.3%]. Moreover, IMA levels were statistically significantly higher in patients finally diagnosed as unstable angina [UA] than those who diagnosed as non ST elevation myocardial infarction [NSTEMI] [p<0.04] meanwhile, it is insignificantly higher than that in STEMI patients. The sensitivity of IMA to predict ACS cases [94.1%] was higher than that of ECG [78.4%] and CPK at presentation [14.7%] and after 6 hours [54.7%]. On the other hand, the specificity of IMA, ECG, CPK at presentation and 6 hours later [86.4%, 90.9%, 86.4% and 97% respectively]. IMA can be used at the emergency setting to exclude the diagnosis of ADS. Moreover, the use of IMA as a diagnostic biomarker in addition to standard markers of myocardial injury is very useful for the evaluation of patients with suspected ACSMedical personnel are without doubt exposed to many biological and chemical agents [e.g., ionizing radiation, ultraviolet light rays, ultrasound, alkylating agents, anesthetic gases and antineoplastic agents] which are efficient inducers of chromosomal aberrations. X-ray and ultrasound are used extensively in many branches of modern medicine. X-rays, together with gamma rays and UV light are all part of the electromagnetic spectrum. Their interaction with biologic material depends on the frequency of wavelength of the radiation. At the short wavelengths of X-rays, electromagnetic radiation has sufficient energy to produce ionization as a result of the removal of electrons from atoms, and are thus called ionizing radiation. Ultrasound [term used to describe mechanical vibrations at frequencies above the human limit of audibility] has experienced phenomenal growth in medical diagnosis in recent years and it is usable in many sensitive circumstances where X-rays might be damaging. The aim of the present study is to assess genomic damage produced by occupational exposure to X-rays and ultrasound equipment using FISH technique using the whole chromosome paint probes for chromosome 1 and 2. Our study revealed chromosomal aberration in 2 personnel exposed to ionizing radiation, continuously for more than 10 years, in the form of absent signaling of chromosomes 2 which denotes total loss [monosomy] of this chromosome. In conclusion, our results may not provide significant proof of the cytogenetic damage caused by occupational exposure to ionizing radiation and ultrasound, however it may prove useful for future human population studies in this field and surely concluded that the necessity of following safety rules in fields dealing with radiations

Effect of matrigel on function and morphology of human endometrial epithelial cell in vitro

The importance of extra cellular matrix [ECM] in development and function of different cells has been reported but little is known about its role in human endometrial epithelial cells. The aim of the present study was to examine effects of artificial ECM [Matrigel] and progesterone on the function and morphology of human endometrial epithelial cells in vitro. Endometrial samples were removed, with informed patients consent and Ethics Committee approval, from 17 previously fertile women undergoing total abdominal hysterectomy. The tissue was dissociated and centrifuged to provide an epithelial rich suspension which was cultured either on plastic or seeded into Matrigel to produce polarized cells and then supplemented with or without progesterone [10-6 M]. The amount of nucleic acid content of the cells in both in vitro model systems was examined by DNA, RNA extraction methods. The DNA and RNA content were later measured by spectrophotometry. The amount of total RNA in cells grown on Matrigel [23 +/- 1.5 pg/cell] was more than double that in cells grown on pl1astic [9.1 +/- 1.4 pg/cell]. Cells cultured on both in vitro model systems had RNA induced by steroid hormones, but the extent of induction was greater in cells grown on Matrigel [30 +/- 2 pg/cell] than those on plastic [12 +/- 1.9 pg/cell]. Cells cultured on Matrigel were differentiated and became polarized but cells grown on plastic proliferated to full confluency. Cells grown on Matrigel with progesterone supplementation were highly polarized, euchromatic and had greater mitochondria and accumulation of glycogen, when compared to unsupplemented cultures. These results suggest that ECM plays an important role in gene expression, polarization and differentiation of human endometrial epithelial cells in vitro. Endometrial cells grown on ECM responded to steroid hormone in a manner to that reported in endometrial cells in vivo

Blastocyst-endometrium interaction: intertwining a cytokine network

The successful implantation of the blastocyst depends on adequate interactions between the embryo and the uterus. The development of the embryo begins with the fertilized ovum, a single totipotent cell which undergoes mitosis and gives rise to a multicellular structure named blastocyst. At the same time, increasing concentrations of ovarian steroid hormones initiate a complex signaling cascade that stimulates the differentiation of endometrial stromal cells to decidual cells, preparing the uterus to lodge the embryo. Studies in humans and in other mammals have shown that cytokines and growth factors are produced by the pre-implantation embryo and cells of the reproductive tract; however, the interactions between these factors that converge for successful implantation are not well understood. This review focuses on the actions of interleukin-1, leukemia inhibitory factor, epidermal growth factor, heparin-binding epidermal growth factor, and vascular endothelial growth factor, and on the network of their interactions leading to early embryo development, peri-implantatory endometrial changes, embryo implantation and trophoblast differentiation. We also propose therapeutical approaches based on current knowledge on cytokine interactions.

In vitro action of leukemia inhibitory factor (LIF) on mid-secretory stage endometrial stromal cells collected from hormone-simulated, ovariectomized monkey and maintained in three-dimensional primary culture.

Leukemia inhibitory factor (LIF) is a pleiotropic cytokine that is known to play an important role in blastocyst implantation. The putative action of LIF in the regulation of uterine function has been examined using mid-secretory stage monkey endometrial stromal cells cultured on rat-tail collagen type I and treated with recombinant human LIF (rhLIF) or immunoneutralized LIF (in LIF) under serum-free condition. Long-term ovariectomized rhesus monkeys (n=8) underwent simulation of their menstrual cycles with steroid hormones and endometrial tissue samples were collected on cycle day 18; stromal cells were isolated and grown in primary culture on three-dimensional collagen matrix. Significant decline in cellular protein synthesis (P < 0.01) and cell proliferation index (P < 0.05) was observed in cells with increasing doses (0-1000 ng/ml) of rhLIF under serum-free in vitro condition. JAK1 expression in cultured cells increased (P < 0.01) in response to rhLIF as revealed from Western blot and confocal laser scanning microscopic examination, STAT1 and STAT2 expressions were unchanged, while pSTAT3 expression increased (P < 0.01) with increased concentration of rhLIF in culture medium. Autophosphorylation of JAK1 in endometrial stromal cells showed no change with increasing concentration (0.01 to 100 ng/ml) of rhLIF in vitro, but significant (P < 0.05) increase was observed with the time of exposure to rhLIF. Immunoneutralization of LIF or no addition of rhLIF to cultured cells led to significant (P < 0.01) increase in stromal cell proliferation index and significant (P < 0.01) decrease in the level of JAK1 and its autophosphorylation as compared to cells exposed to rhLIF alone. From the present set of experiments we conclude that rhLIF affects the physiological behaviour of monkey mid secretory stage endometrial stromal cells in vitro via the JAK-STAT signaling pathway.

Histologia endometrial, densidade, calibre microvascular e matriz-metaloproteinase-3 em usuarias do sistema intra-uterino liberador de levonorgestrel ou do implante subdermico liberador de nestorone com e sem sangramento endometrial / Endometrium, density, microvascular caliber and matrix-metalloproteinase-3 in users of the intrauterine system releasing levonorgestrel-releasing subdermal implant or Nestorone with and without endometrial bleeding

Atualmente, os métodos anticoncepcionais somente com progestógenos estão sendo usados por um número cada vez maior de mulheres. Estes são de alta eficácia e de longa duração. Dentre eles encontram-se o sistema intra-uterino liberador de levonorgestrel (SIU-LNG) e o implante subdérmico liberador de nestorone (NES). Nas usuárias destes métodos, as mudanças no padrão do fluxo menstrual são quase universais e a imprevisibilidade do sangramento uterino constitui a principal causa de descontinuação dos mesmos. O objetivo deste estudo de corte transversal foi verificar diferenças quanto aos aspectos histológicos em hematoxilina-eosina (HE) e através de reações imunoistoquímicas em biópsias endometriais nas usuárias do SIU-LNG ou do implante subdérmico liberador de NES, em grupos com e sem sangramento uterino, após seis meses de uso dos métodos. Sujeitos e Métodos: Foram avaliadas e submetidas à biópsia endometrial 58 usuárias do SIU-LNG, subdivididas em 29 casos em cada grupo com e sem sangramento e 20 usuárias do implante subdérmico liberador de NES, sendo 14 no grupo com sangramento e 6 sem sangramento. Após a avaliação morfológica em HE o material foi submetido a reações imunoistoquímicas para os marcadores CD34 (avaliação dos vasos da mucosa) e MMP-3 (matriz-metaloproteinase-3), enzima relacionada aos processos de colapso do endométrio. Resultados: Dentre as usuárias do SIU-LNG, o aspecto histológico mais freqüente foi o endométrio progestacional. Não houve correlação significativa entre a presença do sangramento e a idade, paridade, cor, índice de massa corpórea, tempo de uso do SIU-LNG e o padrão menstrual prévio ao uso do SIU-LNG. Também não houve diferença estatisticamente significante com relação aos dados morfológicos analisados de endometrite, necrose estromal, erosão de superfície com reepitelização focal, colapso estromal, pseudoestratificação glandular,...

Progestogen-only contraception is being used by an increasing number of women. It has a long term and high efficacy. Among these methods, there is the levonorgestrel intrauterine system (LNG-IUS) and the Nestoroneâ (NES)-releasing contraceptive implant. However, the changes of menstrual patterns are almost universal and unpredictable and remains the main reason for discontinuation of the method. Objective: This was a cross sectional study. It aimed to investigate the endometrial histology and immunohistochemical reations in users of LNG-IUS or NES-releasing contraceptive implant, for more than six months, in women with and without endometrial bleeding. Methods: In users of LNG-IUS endometrial biopsy was obtained in a total of 58 healthy volunteers, twenty-nine women in each group: with or without endometrial bleeding. In users of (NES)-releasing contraceptive implant in 20 womens, 14 with and 6 without endometrial bleeding. Results: In users of LNG-SIU, histological analysis revealed that the majority of samples displayed a progestin-modified appearance. There was no significant difference between the two groups comparing age, body mass index, duration of contraceptive use, parity, ethnicity and menstrual pattern previous to use of LNG-IUS. There was no significant difference in the evaluation of endometritis, stroma collapse, superficial erosion, glandular pseudo stratification, oedema, density and caliber microvascular. The perimeter and the major glandular diameter were the only characteristics significantly higher in the group without bleeding. A number significantly higher of leukocytes was found in the group with bleeding. This was the only histological characteristic correlated with the endometrial bleeding in these groups. MMP-3 showed a significantly higher...

Endocrine and paracrine correlates of endometrial receptivity to blastocyst implantation in the human.

Synchronous attainment of maternal endometrial receptivity allows implantation-stage adhesive blastocyst to undertake apposition, attachment and invasion. In the present essay, we propose a model according to which luteal phase progesterone induces a basic drive in endometrium toward receptivity and as a result, adequately primed endometrium differentiates through certain steps in a fixed action pattern. The implantation-stage embryo senses such endometrial responsiveness circumstantially by the factors secreted by maternal endometrium and undertakes differentiation to implant by secreting factors which act on maternal endometrial cells to further potentiate them to implantation stage-specific changes. Such a dynamic temporo-spatial manner of interaction involving a set of specific factors acting synchronously leads to the activation of innate releasing process in both compartments towards embryo attachment followed by successful intrusion and controlled invasion of trophoblast cells into maternal endometrium. In the present review we discuss the potential role of various endocrine and paracrine factors in the process of blastocyst implantation in the human.

Apoptosis in endometrium of mouse during estrous cycle.

The present study was carried out to evaluate apoptosis in endometrium and to correlate these changes with the circulating levels of estradiol and progesterone in the mouse. Apoptosis was observed in various compartments of mouse uterus i.e. stroma, glandular epithelium and luminal epithelium depending on the stage of cycle. Stromal cell apoptosis was observed during various stages of cyclicity except on estrus day. Luminal epithelial cells showed apoptotic changes during all stages of cyclicity except on diestrus day. During metestrus, apoptosis was observed in glandular and luminal epithelia as well as stromal cells. Steroid antagonists such as tamoxifen and onapristone altered the apoptotic changes in the uterus. The results suggest that epithelial cell apoptosis is regulated by estrogen while stromal cell apoptosis is under the control of progesterone.