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Aim: : To measure obestatin levels in the blood and saliva samples of normal-weight patients with polycystic ovary syndrome (PCOS) in comparison with normal-weight healthy controls, and to determine whether there were relationships between blood and/ or salivary obestatin levels and other measured parameters. Materials and Methods: Fifteen healthy women and 15 patients with PCOS, all of which had normal weight, were included in the study. Participants' age, height, weight, menstrual characteristics, hormone levels, body mass index, waist/hip ratio, and modifed Ferriman-Gallwey (FG) scores were recorded. Obestatin levels were measured in both fasting blood and saliva samples. Homeostasis Model Assessment (HOMA-IR) was used to predict insulin resistance. Results: In the PCOS group, menstrual cycle duration and FG scores were signifcantly higher (P-value, <0.001, <0.001, respectively). The levels of luteinizing hormone, total testosterone and androstenedione were signifcantly higher in the PCOS group than in the control group (P-value, 0.001, 0.009, 0.004, respectively). In the PCOS group, blood obestatin level was 1265.2 ± 221.9 pg/ml, salivary obestatin level was 3095.33 ± 310.2 pg/ml; whereas the control group demonstrated lower levels of 939.66 ± 72.3 pg/ml and 2611.20 ± 217.1 pg/mL, respectively. However, no statistically signifcant difference was found between the PCOS and control groups when comparing obestatin levels in blood or saliva (P-value, 0.218, 0.369, respectively). No correlation was found between blood and salivary obestatin levels in either group. Finally, obestatin levels were not associated with any of the other measured parameters. Conclusion: Our results suggest that increased levels of obestatin, albeit in a small amount that would not be statistically signifcant in normal weight patients with PCOS, may have signifcant effects on weight control in these patients. In addition, our fndings show that saliva sample can be used as an alternative to blood sample in the measurement of obestatin level in patients with PCOS. [ABSTRACT FROM AUTHOR]

Objective. To determine the effects of Mullerian inhibiting substance (MIS) treatment on endometriosis cells through study of apoptosis and autophagy. Design. Experimental in vitro study. Setting. University research laboratory. Cell Line. CRL-7566 endometriosis cell line. This line was established from a benign ovarian cyst taken from a patient with endometriosis. Interventions. In vitro treatment with MIS. Main Outcome Measures. The main outcome measures were cellular viability, proliferation, cell-cycle arrest, and induction of apoptosis and autophagy in endometriotic cells. Results. MIS treatment inhibited proliferation of endometriosis cells and induced apoptosis, as indicated by Annexin V staining, and induced caspase-9 cleavage and cell-cycle arrest, as evidenced by increased expression of p27 CDK-inhibitor. MIS treatment also induced autophagy in endometriosis cells as demonstrated by a significant increase in LC3-II induction, a hallmark of autophagy. Conclusions. MIS inhibits cell growth and induces autophagy, as well as apoptosis, in ectopic endometrial cell lines. Our results suggest that MIS may have a potential as a novel approach for medical treatment of endometriosis. Further studies may be needed to test the efficacy of MIS treatment in animal models and to develop MIS treatment specifically targeted to the endometriosis. [ABSTRACT FROM AUTHOR]

Objective: To investigate ghrelin and obestatin expression in serous ovarian tumours. Materials and methods: Preparations of deparaffinized blocks obtained from the pathology archives of a total of 47 previously diagnosed cases of benign serous tumour ( n = 20), borderline serous tumour ( n = 7) and malignant serous tumour ( n = 20) were subjected to immunohistochemical examination to find out ghrelin and obestatin expressions. Results: Mean ghrelin expressions decreased significantly in the benign group, relative to the malignant group ( p < 0.05), while there was no significant change in mean obestatin expression. It was established that rates of preparations with moderate and severe ghrelin and obestatin expression displayed a significant increase from benign to malignant ones ( p < 0.05). Conclusion: The fact that rates of preparations with severe expression correlated with an increase in malignancy suggests that ghrelin and obestatin may be effective in the malignant transformation in at least some cases. [ABSTRACT FROM AUTHOR]

Abstract: Objectives: The present study aims to establish the levels of acylated ghrelin, desacylated ghrelin, obestatin and preptin, during pregnancy and the postpartum period in pregnant women with Gestational Diabetes Mellitus (GDM) and healthy pregnancy women. Design and methods: The study registered 20 pregnant women with GDM and 20 healthy pregnant women. Fasting venous blood samples were collected from all cases between weeks 24 and 28 of pregnancy and after 24h postpartum. Hormones were analyzed using ELISA method. Results: Serum acylated ghrelin (p:0.001), desacylated ghrelin (p:0.001), obestatin (p:0.006) and preptin (p:0.001) levels were all found statistically higher in both groups during the postpartum period, when compared to the pregnancy period. A positive correlation was established between desacylated ghrelin and acylated ghrelin (p:0.008), desacylated ghrelin and preptin (p:0.012) and preptin and insulin (p:0.039) in the GDM group during pregnancy. Conclusions: The studied hormones (especially desacylated ghrelin and obestatin) are critical in GDM pathophysiology based on the comparison of measure after and before the delivery. [Copyright &y& Elsevier]

Background. Endometrium carcinoma ranks fourth among female carcinomas. Therefore, early diagnosis of endometrium pre-malignant lesions is emphasised, and attempts are made to identify the risk factors. Since hyperplasias, particularly those with atypia, are held responsible for the development of the most common endometrium carcinomas, it is important to definitely distinguish between well-differentiated carcinomas and hyperplasia with atypia. In the present study, we aimed to explore whether ghrelin expression had a role in distinguishing between benign, pre-malignant and malignant lesions of endometrium. Methods. Tissue ghrelin expressions of a total of 60 cases, who were diagnosed in the Pathology Department Laboratory of Fıırat University Medical School, and of whom 10 were in the proliferation phase, 10 had simple hyperplasia without atypia, 10 had simple hyperplasia with atypia, 10 had complex hyperplasia without atypia, 10 had complex hyperplasia with atypia and 10 had endometrioid carcinoma cases, were examined using immunohistochemical method. Additionally, tissue samples were homogenised to analyse tissue ghrelin levels in the supernatants according to RIA method. Samples from the parotid glands were used as positive control for ghrelin. Cells that exhibited cytoplasmic staining with ghrelin antibody were evaluated as positive. Results. Immunohistochemical examination showed that ghrelin expression increased markedly in the proliferation phase, relative to hyperplasias and carcinoma. These results were parallel to ghrelin levels in tissue supernatants. Immunohistochemical and RIA analysis results indicate that ghrelin expression either markedly decreases or is entirely depleted in endometrial carcinomas. Conclusions. Therefore, we think that ghrelin expression can be useful in differentiating not only endometrium carcinomas from benign lesions but also complex hyperplasias with atypia, which pose diagnostic difficulties. [ABSTRACT FROM AUTHOR]

Aim. The present study was designed to investigate the effects of two aromatase inhibitors on steroid hormone levels, bone mineral density (BMD) and bone turnover markers in intact female rats. Methods. Letrozole and anastrazole at two different dose levels were investigated for their effect on serum levels of estradiol, androstenedione, testosterone, dehydroepiandrosterone and dehydroepiandrosterone sulfate, BMD of femur and dorsal spine, and osteocalcin and pyridinoline levels as bone turnover markers. Fifty intact female rats were randomly divided in five groups (group 1 (n = 10): control, 2 ml saline; group 2 (n = 10): letrozole 1 mg/kg; group 3 (n = 10): letrozole 2 mg/kg; group 4 (n = 10): anastrazole 0.1 mg/kg; group 5 (n = 10): anastrazole 0.2 mg/kg), and oral gavages were applied for a period of 16 weeks. Results. Both doses of letrozole and anastrazole did not change femoral and vertebral BMD. Serum estradiol levels were reduced significantly at all dose levels by both agents (p < 0.001); all androgen levels were significantly elevated by letrozole (p < 0.05), although anastrazole increased only androstenedione (p < 0.05). The higher dose of letrozole increased osteocalcin levels (p < 0.05), while pyridinoline levels were increased (p < 0.05) by the higher dose of anastrazole. Conclusion. Our results indicate that short-term use of letrozole and anastrazole had no clear effects on BMD in intact rats. Further investigations are needed to understand their effects on bone metabolism in intact females. [ABSTRACT FROM AUTHOR]

Binding activity of steroidogenic factor-1 (SF-1) to promoters of the majority of steroidogenic genes in response to gonadotropins is a critical mechanism that regulates steroidogenesis in gonads. Thus, the modulation of SF-1 action may be essential for the differential regulation of formation of sex steroids in the ovary. Aromatase P450 (P450arom) is the rate-limiting enzyme for estrogen formation. In this study, we characterize another nuclear receptor half site in the gonadal aromatase promoter which we show to be important for aromatase regulation. We also show herein that the stimulation of P450arom promoter activity by SF-1 in ovarian granulosa, testicular Sertoli and JEG-3 choriocarcinoma cells is inhibited by two transcription factors, Wilms’ tumor suppressor gene (WT1) and dosage sensitive sex reversal adrenal hypoplasia congenita critical region on the X chromosome gene 1 (DAX-1). Given the characterized roles of these transcription factors in gonadal development and function, modulation of SF-1 action by WT1 and DAX-1 may represent an important key mechanism in steroidogenesis. [Copyright &y& Elsevier]

Abstract: Objectives: The goal of this study was to compare levels of acyl and des-acyl ghrelin, obestatin, nesfatin-1 and leptin in healthy gravidas to hyperemesis gravidarum (HG) patients. Design and methods: Twenty pregnant women with HG and twenty healthy pregnant women all of similar ages, BMI and all at similar pregnancy development comprised the study cohort. Fasting serum samples were obtained and measured for acyl and des-acyl ghrelin, leptin, obestatin and nesfatin-1. Results: Nesfatin-1 concentrations in the HG group were higher compared to the control group whereas; leptin concentrations during pregnancy were lower in the HG group as compared to the control group. The two groups did not differ with regard to acyl and des-acyl ghrelin and obestatin. Conclusion: This pilot study suggests a possible role of leptin and nesfatin-1, which might be involved in the pathology of the disease. [Copyright &y& Elsevier]

A letter to the editor is presented which discusses a medical case study of a 72-year-old woman who has a basal cell carcinoma and its treatment.

Abstract: Endometriosis is a common and chronic disease characterized by persistent pelvic pain and infertility. Estradiol is essential for growth and inflammation in endometriotic tissue. The complete cascade of steroidogenic proteins/enzymes including aromatase is present in endometriosis leading to de novo estradiol synthesis. PGE2 induces the expression of the genes that encode these enzymes. Upon PGE2 treatment, coordinate recruitment of the nuclear receptor SF-1 to the promoters of these steroidogenic genes is the key event for estradiol synthesis. SF-1 is the key factor determining that an endometriotic cell will respond to PGE2 by increased estradiol formation. The presence of SF-1 in endometriosis and its absence in endometrium is determined primarily by the methylation of its promoter. The key steroidogenic enzyme in endometriosis is aromatase encoded by a single gene because its inhibition blocks all estradiol biosynthesis. Aromatase inhibitors diminish endometriotic implants and associated pain refractory to existing treatments in affected women. [Copyright &y& Elsevier]

Abstract: Pathogenesis and growth of three common women''s cancers (breast, endometrium and ovary) are linked to estrogen. A single gene encodes the key enzyme for estrogen biosynthesis named aromatase, inhibition of which effectively eliminates estrogen production in the entire body. Aromatase inhibitors successfully treat breast cancer, whereas their roles in endometrial and ovarian cancers are less clear. Ovary, testis, adipose tissue, skin, hypothalamus and placenta express aromatase normally, whereas breast, endometrial and ovarian cancers overexpress aromatase and produce local estrogen exerting paracrine and intracrine effects. Tissue-specific promoters distributed over a 93-kb regulatory region upstream of a common coding region alternatively control aromatase expression. A distinct set of transcription factors regulates each promoter in a signaling pathway- and tissue-specific manner. In cancers of breast, endometrium and ovary, aromatase expression is primarly regulated by increased activity of the proximally located promoter I.3/II region. Promoters I.3 and II lie 215bp from each other and are coordinately stimulated by PGE2 via a cAMP-PKA-dependent pathway. In breast adipose fibroblasts exposed to PGE2 secreted by malignant epithelial cells, PKC is also activated, and this potentiates cAMP-PKA-dependent induction of aromatase. Thus, inflammatory substances such as PGE2 may play important roles in inducing local production of estrogen that promotes tumor growth. [Copyright &y& Elsevier]

Abstract: Endometrial tissue from uterine disease-free women does not exhibit aromatase activity. In contrast, aromatase enzyme activity and mRNA levels are readily detectable in endometriosis. PGE2 stimulates both aromatase expression and activity in endometriotic stromal cells via promoter II region of the aromatase gene. This results in local production of estradiol, which induces PGE2 formation and establishes a positive feedback cycle. This mechanism seems to contribute to continuous production of estradiol and PGE2. Aromatase mRNA levels and enzyme activity are also present in uterine leiomyomata that are estrogen-dependent benign tumors of the myometrium. Successful treatment of endometriosis and uterine leiomyomata using aromatase inhibitors by recent pilot trials underscores the clinical significance of these molecular studies. [Copyright &y& Elsevier]

: ObjectiveTo determine the role of an aromatase inhibitor, letrozole, in the treatment of reproductive-age women with endometriosis and associated chronic pelvic pain.: DesignPhase 2, open-label, nonrandomized proof-of-concept study.: SettingOutpatient tertiary-care center.: Patient(s)Ten patients with endometriosis, all previously treated both medically and surgically, with unsatisfactory results.: Intervention(s)Endometriosis was diagnosed by biopsy and scored from an initial diagnostic laparoscopy performed within 1 month before treatment was begun. Oral administration of letrozole (2.5 mg), the progestin norethindrone acetate (2.5 mg), calcium citrate (1,250 mg), and vitamin D (800 IU) was done daily for 6 months. Within 1–2 months after completion of the treatment, a second-look laparoscopy was performed to score and biopsy endometriosis.: Main outcome measure(s)Changes in American Society for Reproductive Medicine (ASRM) scores for endometriosis, pelvic pain assessed by visual analog scale, serum hormone levels (FSH, LH, E2, and estrone [E1]), and bone density (DEXA scan).: Result(s)No histologically demonstrable endometriosis was present in any patient during the second-look laparoscopy. ASRM and pelvic pain scores decreased significantly in response to treatment. Overall, no significant change in bone density was detected. Gonadotropin levels were not significantly altered by treatment, and although circulating E2 and E1 levels were reduced, the decrease was not statistically significant.: Conclusion(s)The combination of letrozole and norethindrone acetate achieved marked reduction of laparoscopically visible and histologically confirmed endometriosis in all 10 patients and significant pain relief in nine out of 10 patients who had not responded previously to currently available treatments. On this basis, letrozole should be a candidate for the medical management of endometriosis. [Copyright &y& Elsevier]

Background: The objective of this study is to investigate the effect of pyrrolidine dithiocarbamate [PDTC; a nuclear factor-kappaB (NF-kappaB) inhibitor] and bortezomib (Velcade; a proteasome inhibitor) on the development of experimental endometriotic implants in rats., Methods: Endometriosis was surgically induced in 30 rats using the method of Vernon and Wilson. Three weeks later the viability and volume of the implants were recorded and classified. Afterwards, rats were put into three groups with equal numbers. The groups were labelled as the control, the PDTC and the bortezomib groups. Seven days after treatment, a third laparotomy was done and the volume of implants was measured again. The animals were then sacrificed, and the implants were stained with Ki67, proliferating cell nuclear antigen (PCNA), CD34, CD31 and Masson's trichrome histochemical staining., Results: In 80% of the implanted rats, vesicles at the suture region were observed, and the rats graded according to average vesicle diameter (D) as: Grade 1 (no vesicle, 20% of rats), Grade 2 (D < 2 mm, 33.3% of rats), Grade 3 (2 mm 4.5 mm, 26.7% of rats) and Grade 4 (D > 4.5 mm, 20% of rats). After treatment with PDTC or bortezomib, these percentages were decreased for Grades 3 and 4, and increased in Grade 1. The post-treatment implant volumes were decreased in the PDTC and bortezomib groups (P < 0.002 and P < 0.001), and slightly increased in the control group (P = 0.279). In the PDTC and bortezomib groups, CD34, CD31, PCNA and Ki67 expression levels were similar but were significantly reduced compared with the control group., Conclusions: PDTC and bortezomib may represent a novel therapeutic strategy for treatment of endometriosis.

Objective: To investigate the effect of circadian rhythm changes on fetal-placental development in pregnant rats., Material and Method: Pregnant rats were randomly allocated to 4 groups with 10 rats in each group. Group 1: 12 hours light 12 hours dark period (n:10, Control Group); Group 2: 6 hours light 6 hours dark period (n:10); Group 3: constant light group (n:10); Group 4: constant dark group (n:10). Rats were made to deliver by cesarean birth at the end of their pregnancy and their placenta, number and weight of the offspring, and placental histopathological changes were examined., Finding: Fetal number in the constant dark group was found significant in comparison to constant light group (p<0.01). Fetal weight in all groups with altered circadian rhythm was significantly lower than that in the control group (p<0.01). The decrease in fetal weight was more evident in the constant light group than in the constant dark group (p<0.01). Comparison of the histopathological findings of the placenta showed that placental edema, fibrin accumulation and leukocyte infiltration were significantly higher in constant light and constant dark groups than in other groups (p<0.01)., Conclusion: Circadian rhythm change in pregnancy adversely affects the fetal-placental development. More detailed studies are needed to clarify this mechanism.

A single gene encodes the key enzyme for estrogen biosynthesis termed aromatase, inhibition of which effectively eliminates estrogen production. Aromatase inhibitors successfully treat breast cancer and endometriosis, whereas their roles in endometrial cancer, uterine fibroids, and aromatase excess syndrome are less clear. Ovary, testis, adipose tissue, skin, hypothalamus, and placenta express aromatase normally, whereas breast and endometrial cancers, endometriosis, and uterine fibroids overexpress aromatase and produce local estrogen that exerts paracrine and intracrine effects. Tissue-specific promoters distributed over a 93-kilobase regulatory region upstream of a common coding region alternatively control aromatase expression. A distinct set of transcription factors regulates each promoter in a signaling pathway- and tissue-specific manner. Three mechanisms are responsible for aromatase overexpression in a pathologic tissue versus its normal counterpart. First, cellular composition is altered to increase aromatase-expressing cell types that use distinct promoters (breast cancer). Second, molecular alterations in stromal cells favor binding of transcriptional enhancers versus inhibitors to a normally quiescent aromatase promoter and initiate transcription (breast/endometrial cancer, endometriosis, and uterine fibroids). Third, heterozygous mutations, which cause the aromatase coding region to lie adjacent to constitutively active cryptic promoters that normally transcribe other genes, result in excessive estrogen formation owing to the overexpression of aromatase in many tissues.

We investigated the regulation of prostaglandin production in normal endometrial stromal cells (ESC) by malignant endometrial epithelial cells. We found that cyclooxygenase (COX)-2 mRNA and protein levels and prostaglandin (PG)E(2) production in ESC were significantly increased by Ishikawa malignant endometrial epithelial cell conditioned medium (MECM). By using transient transfection assays, we found that the -360/-218-bp region of the COX-2 promoter gene was critical for MECM induction of promoter activity. This MECM-responsive region contained a variant nuclear factor (NF)-kappa B site at -222 to -213 that, when mutated, completely abolished COX-2 promoter activation by MECM. Employing electrophoretic mobility shift assays, we further demonstrated that binding of NF-kappa B p65 to this NF-kappa B-binding site is, in part, responsible for the COX-2 promoter activation by MECM. To investigate further the potential effects of MECM on COX-2 mRNA stability, ESC were treated with MECM in the absence or presence of actinomycin D, a general transcription inhibitor. We found that MECM significantly increased COX-2 mRNA stability. Intriguingly, we found that PGE(2) was one of the major factors in MECM, which was responsible for up-regulating COX-2 expression in ESC. ECC-1 and HEC-1A malignant endometrial epithelial cell lines also produced significantly increased quantities of PGE(2). In conclusion, malignant endometrial epithelial cells secrete PGE(2) that induces COX-2 expression in normal endometrial stromal cells in a paracrine fashion through activation of transcription and stabilization of COX-2 mRNA.

We investigated the effects of vascular endothelial growth factor (VEGF) on cyclooxygenase-2 (COX-2) expression and prostaglandin E(2) (PGE(2)) synthesis in human microvascular endothelial cells (HMEC-1). Treatment of HMEC-1 with VEGF resulted in a dose- and time-dependent up-regulation of COX-2 mRNA and protein levels. This up-regulation was accompanied by a 1.6-fold increase in PGE(2) synthesis. Pretreatment of HMEC-1 with a selective COX-2 inhibitor, NS-398, abolished VEGF-induced PGE(2) synthesis, suggesting specific up-regulation of COX-2 activity by VEGF in HMEC-1. Transient transfection assays using deletion mutants of the human COX-2 promoter fused to the luciferase reporter gene indicated critical requirement of a regulatory region spanning -828/-123 bp for VEGF induction of COX-2 promoter activity in HMEC-1. Site-directed mutation analysis demonstrated that a GATA cis-acting element at -685/-680 bp was essential for VEGF- induced COX-2 promoter activity in HMEC-1. These observations are of particular importance given the recent demonstrations of critical requirement of COX-2 isoenzyme for tumor growth and angiogenesis.