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Objectives To investigate whether BclI polymorphism in the glucocorticoid receptor gene influences hypothalamic-pituitary-adrenal (HPA) axis regulation, body composition and metabolic parameters in women with adrenal incidentalomas (AIs). Study design A cross-sectional study. Main outcome measures We analyzed 106 women with AIs. Insulin resistance was assessed using a homeostasis model while HPA activity was assessed using dexamethasone suppression tests (DST), basal ACTH, urinary free cortisol, and midnight serum cortisol level. Body composition was analyzed using dual-energy X-ray absorptiometry. DNA was obtained from peripheral blood leucocytes and BclI polymorphism was detected using PCR, RFLP and DNA sequencing. Results BclI carriers in comparison with those with wild-type BclI had less suppressed cortisol after DST-0.5 mg (126.4 ± 111.4 vs 80.9 ± 75.7 nmol/l, p = 0.026) and had a lower prevalence of impaired glucose tolerance and of type 2 diabetes mellitus (T2DM). BclI carriers had a higher percentage of leg fat mass (FM), lower left-sided limb muscle mass and a decline in total lean body mass. Duration of menopause remained a strong predictor of appendicular lean mass index (ALMI) (β=-0.125, p = 0.034). BclI polymorphism was significantly associated with sum of legs FM percentage (β=0.327, p = 0.048). T2DM was negatively associated with BclI polymorphism, after adjusting for age, truncal FM, ALMI, and sum of legs FM (OR=0.158, 95%CI 0.031–0.806, p = 0.027). Conclusions BclI polymorphism is associated with tissue-specific glucocorticoid sensitivity, relative glucocorticoid resistance of the HPA axis and peripheral adipose tissue, and glucocorticoid hypersensitivity at the muscle level. By modulating glucocorticoid and insulin sensitivity, BclI polymorphism appears to reduce the risk of T2DM in women with AIs.

Introduction: Insulin resistance (IR) is one of the main features of polycystic ovary syndrome (PCOS), and metformin is one of the most commonly prescribed insulin sensitizers in its treatment. Research carried out so far shows heterogeneous results on the effects of metformin on the hormonal and metabolic status of women with PCOS. The aim of the study was to investigate the effect of six-month treatment with metformin (2000 mg per day) on the hormonal and metabolic characteristics of women with PCOS, paired with healthy women by body mass index (BMI) and age. Material and Methods: A number of 20 women with PCOS [age: 23.5 ± 5.9 years, BMI: 24.8 ± 4.2kg / m2] and 20 healthy control group women were tested (age: 24.5 ± 5.3 years, BMI: 23.6 ± 3.0kg / m2). Polycystic ovary syndrome was diagnosed with ESHRE / ASRM criteria. Standard biochemical and hormonal parameters were determined, which were repeated in the PCOS group and after six months of metformin therapy. Insulin resistance was determined using a homeostatic model (HOMA-IR). Results: After six months of metformin therapy in the PCOS group, there was a significant increase in the frequency of menstrual cycle (65% versus 87%, p

Polycystic ovary syndrome is a heterogeneous endocrine and metabolic disorder associated with abdominal obesity, dyslipidemia and insulin resistance. Since abdominal obesity is characterized by low-grade inflammation, the aim of the study was to investigate whether visceral adipose tissue inflammation linked to abdominal obesity and dyslipidemia could lead to impaired insulin sensitivity in the animal model of polycystic ovary syndrome.Female Wistar rats were treated with nonaromatizable 5α-dihydrotestosterone pellets in order to induce reproductive and metabolic characteristics of polycystic ovary syndrome. Glucose, triglycerides, non-esterified fatty acids and insulin were determined in blood plasma. Visceral adipose tissue inflammation was evaluated by the nuclear factor kappa B intracellular distribution, macrophage migration inhibitory factor protein level, as well as TNFα, IL6 and IL1β mRNA levels. Insulin sensitivity was assessed by intraperitoneal glucose tolerance test and homeostasis model assessment index, and through analysis of insulin signaling pathway in the visceral adipose tissue.Dihydrotestosterone treatment led to increased body weight, abdominal obesity and elevated triglycerides and non-esterified fatty acids, which were accompanied by the activation of nuclear factor kappa B and increase in macrophage migration inhibitory factor, IL6 and IL1β levels in the visceral adipose tissue. In parallel, insulin sensitivity was affected in 5α-dihydrotestosterone-treated animals only at the systemic and not at the level of visceral adipose tissue.The results showed that abdominal obesity and dyslipidemia in the animal model of polycystic ovary syndrome were accompanied with low-grade inflammation in the visceral adipose tissue. However, these metabolic disturbances did not result in decreased tissue insulin sensitivity.

Polycystic ovary syndrome (PCOS) is the most common endocrinopathy in women of reproductive age. It is a heterogenous disorder, with hyperandrogenism, chronic anovulation and polycystic ovaries as basic characteristics, and associated metabolic syndrome features. Increased secretion of leptin and leptin resistance are common consequences of obesity. Leptin is a hormone with anorexigenic effects in the hypothalamus. Its function in the regulation of energy intake and consumption is antagonized by glucocorticoids. By modulating leptin signaling and inflammatory processes in the hypothalamus, glucocorticoids can contribute to the development of metabolic disturbances associated with central energy disbalance. The aim of the study was to examine the relationship between hypothalamic leptin, glucocorticoid and inflammatory signaling in the development of metabolic disturbances associated with PCOS. The study was conducted on an animal model of PCOS generated by a continual, 90-day treatment of female rats with 5α-dihydrotestosterone (DHT). The model exhibited all key reproductive and metabolic features of the syndrome. mRNA and/or protein levels of the key components of hypothalamic glucocorticoid, leptin and inflammatory pathways, presumably contributing to energy disbalance in DHT-treated female rats, were measured. The results indicated that DHT treatment led to the development of hyperphagia and hyperleptinemia as metabolic features associated with PCOS. However, these metabolic disturbances could not be ascribed to changes in hypothalamic leptin, glucocorticoid or inflammatory signaling pathways in DHT-treated rats. Keywords: DHT; hypothalamus; leptin; glucocorticoids; inflammation Received: December 14, 2015; Revised: December 23, 2015; Accepted: December 28, 2015; Published online : January 14, 2016

Polycystic ovary syndrome (PCOS) is a cardiometabolic disorder whose features include dyslipidemia, increased oxidative stress (OS, oxy) and chronic inflammation. The aim of this study was to investigate the ability of a summary score for dyslipidemia, OS and inflammation (the DOI score) to discriminate PCOS patients from healthy individuals and to evaluate the effect of obesity on individual scores and the DOI score in patients.Lipid status parameters, OS status parameters (advanced oxidation protein products; total oxidative status; prooxidant-antioxidant balance; malondialdehyde; total protein sulphydryl groups and paraoxonase 1 activity) and CRP were measured in 114 patients and 50 controls using standardised assays. The DOI score was calculated as the sum of dyslipidemia, oxy and inflammation scores, determined as Z-score values for every subject in relation to the controls.PCOS patients had significantly higher oxy-score compared to controls (P0.001). In addition, the DOI score was significantly higher in PCOS patients (P0.001) as the dyslipidemia (P0.05) and inflammatory scores (P0.001) were greater. According to ROC analysis, the oxy-score showed better diagnostic accuracy in discriminating PCOS patients compared to the DOI score (AUC0.9, P0.01). Furthermore, obesity affected the risk scores in patients, especially the DOI score (significantly higher DOI scores in such patients, P0.001).PCOS patients had greater dyslipidemia, chronic inflammation and OS compared to controls and could be segregated using all four scores. Our data suggest that weight gain could be the common factor responsible for induction and propagation of dyslipidemia, OS and inflammation in PCOS patients.Sindrom policističnih jajnika (PCOS) je kardiometabolički poremećaj čije su karakteristike dislipidemija, povišen oksidativni stres (OS) i hronična inflamacija. Cilj ove studije je bio da se ispita sposobnost zbirnog skora dislipidemije, OS i inflamacije (DOI skor) da razlikuje pacijentkinje sa PCOS u poređenju sa zdravim osobama i da se proceni uticaj gojaznosti na pojedinačne skorove i DOI skor kod pacijentkinja.Parametri lipidnog statusa, parametri oksidativnog stresa (uznapredovali produkti oksidacije proteina; ukupni oksidativni status; prooksidativno-antioksidativni balans; malondialdehid; ukupne sulfhidrilne grupe i aktivnost paraoksonaze 1) i CRP su određivani kod 114 pacijentkinja i 50 kontrolnih ispitanica primenom standardizovanih testova. DOI skor je izračunat kao zbir skora dislipidemije, oksidativnog skora i skora inflamacije, određen kao vrednosti Z-skora za svaki subjekt u odnosu na kontrole.PCOS pacijentkinje su imale značajno više vrednosti oksidativnog skora u poređenju sa kontrolnom grupom (P0,001). Zatim, DOI skor je bio značajno viši kod pacijentkinja sa PCOS (P0,001) kao i skorovi dislipidemije (P0,05) i inflamacije (P0,001). Prema rezultatima ROC analize, oksidativni skor je pokazao bolju dijagnostičku tačnost u razlikovanju PCOS pacijentkinja u odnosu na zdrave osobe u poređenju sa DOI skorom (AUC0,9, P0,01). Osim toga, gojaznost je imala uticaj na vrednosti skorova rizika, posebno na DOI skor (vrednosti DOI skora su bile značajno više kod tih pacijentkinja, P0,001).Pacijentkinje sa PCOS su imale izraženiju dislipidemiju, hroničnu inflamaciju i oksidativni stres u poređenju sa kontrolnim ispitanicama i mogu se od njih razlikovati primenom sva četiri skora. Naši rezultati ukazuju da povećanje telesne težine može biti zajednički faktor odgovoran za nastanak i napredovanje dislipidemije, OS i inflamacije kod pacijentkinja sa PCOS.

Androgen excess (AE) was approximated to be present in 7% of the adult population of women. Polycystic ovary syndrome (PCOS) is the most prevalent among them, followed by idiopathic hirsutism (IH), congenital adrenal hyperplasia (CAH), hyperandrogenic insulin-resistant acanthosis nigricans (HAIRAN) syndrome, and androgen-secreting neoplasms (ASNs). Increased cardiovascular risk was implicated in women with AE. Serum testosterone independently increases risk for cardiovascular disease (CVD), and correlates even with indices of subclinical atherosclerosis in various populations of postmenopausal women. Hyperandrogenism in PCOS is closely related to the aggravation of abdominal obesity, and together with insulin resistance forming the metabolic core for the development of CVD. However, phenotypic variability of PCOS generates significant influence on the cardiometabolic risks. Numerous risk factors in PCOS lead to 5-7 times higher risk for CVD and over 2-fold higher risk for coronary heart disease and stroke. However, issue on the cardiometabolic risk in postmenopausal women with hyperandrogenic history is still challenging. There is a significant overlapping in the CVD characteristics of women with PCOS and variants of CAH. Relevant clinical data on the prevalence and cardiometabolic risk and events in women with IH, HAIRAN syndrome or ASNs are scarce. The effects of various oral contraceptives (OCs) and antiandrogenic compounds on metabolic profile are varying, and could be related to the selected populations and different therapy regiments mainly conducted in women with PCOS. It is assumed relation of OCs containing antiandrogenic progestins to the increased risk of cardiovascular and thromboembolic events.

Polycystic ovary syndrome (PCOS) is a frequent endocrine disease in women, with a number of metabolic and reproductive consequences. Obesity, insulin resistance (IR) and type 2 diabetes are prominent metabolic characteristics of PCOS and common factors affecting liver function and generating nonalcoholic fatty liver disease (NAFLD). Multiple genes involved in the synthesis of androgens, cytokines and IR, as well as acquired factors, such as endocrine disruptors, could associate the etiopathogenesis of PCOS and NAFLD. Besides the high prevalence of PCOS in general population, NAFLD was shown to be a frequent condition in transition periods, such as adolescence and menopause. Although liver biopsy is considered to be the gold standard for diagnosing liver damage, its routine use in such a prevalent condition as PCOS can be related to a higher rate of complications. Therefore, it is necessary to be able to diagnose NAFLD using simple and reliable surrogate markers. Recently, fatty liver index and NAFLD fatty liver score analyzed in large cohorts of PCOS women have been shown as accurate markers of liver damage in this metabolically vulnerable population. Lifestyle changes are still the mainstay of the management of NAFLD in PCOS, although prospective randomized controlled clinical studies remain a priority in the field. With regard to medications, metformin may be the drug of choice for treating PCOS patients with NAFLD when pharmacologic therapy is considered. Liraglutide use in obese PCOS has shown favorable effects on the predictors of liver fibrosis. In this review, we aim to summarize the influence of the common risk factors and to discuss the diagnostic approaches and management options for NAFLD in patients with PCOS.

Systemic inflammatory response syndrome (SIRS) changes cortisol dynamics and indicates dissociation between the adrenal cortex and the hypothalamo-pituitary unit. The aim of this study was to assess the cortisol response after stimulation with ACTHFifty-four subjects were included in the study, and SIRS was defined according to the Consensus Conference criteria from 1992. Severity of the disease was determined using the APACHE II score, and organ dysfunction using the SOFA score. Low-dose (1, μg) ACTH test (LDT) was performed in all patients, and cortisol was determined along with basal ACTH. Data were analyzed using parametric and nonparametric tests and regression analysis. The results are presented as mean± standard deviation, and P0.05 was considered statistically significant.There were no differences in cortisol values between the two LDTs. Cortisol increment lower than 250 nmol/L during the LDT was found in 14/54 (25.9%) subjects at the onset of SIRS. Five out of 54 (9.6%)patients died within 7 days from the onset of SIRS. Female sex and maximal cortisol response (▵ max) on LDT predicted the duration of hospitalization in RICU, while APACHE II and SOFA scores best predicted the duration of hospitalization, mortality outcome as well as overall survival outcome.A difference was found in A max at the diagnosis of SIRS and seven days later. ▵ max, and primarily the clinical scores APACHE II and SOFA predicted the outcomes of hospitalization and overall survival.Sindrom sistemskog inflamatornog odgovora (SIRS) menja dinamiku kortizola, ukazujući na disocijaciju između adrenalnog korteksa i hipotalamo-hipofizne jedinice. Cilj ovog rada je da se proceni odgovor kortizola na stimulaciju sa ACTHPedeset četiri osobe su uključene u studiju, a SIRS je definisan prema kriterijumima Konsenzus konferencije iz 1992. Ozbiljnost bolesti je ustanovljena pomoću skora APACHE II, a organska disfunkcija pomoću skora SOFA. Niskodozni (1 μg) ACTH test (LDT) izveden je u svih bolesnika uz određivanje kortizola kao i bazalnog ACTH. Podaci su analizirani pomoću parametarskih i neparametarskih testova i regresione analize. Rezultati su predstavljeni kao srednje vrednosti ± standardna devijacija, a P0,05 je smatrano statistički značajnim.U ovom istraživanju nisu uočene razlike u nivou kortizola između dva LDT-a. Porast kortizola manji od 250 nmol/L tokom LDT-a nađen je kod 14/54 (25,9%) bolesnika prilikom postavljanja dijagnoze SIRS-a. Pet od 54 (9,6%) bolesnika je umrlo unutar sedam dana od početka SIRS-Δ. Ženski pol i maksimalni odgovor kortizola (A max) tokom LDT-a predviđaju trajanje hospitalizacije u RICU, dok skorovi APACHE II i SOFA najbolje predvilaju dužinu hospitalizacije, smrtni ishod kao i opšte preživljavanje.Razlika u Δ max prilikom postavljanja dijagnoze SIRS-a i sedam dana kasnije a pre svega klinickiskorovi APACHE II i SOFA predvilaju ishod hospitalizacije i opšte preživljavanje.

STUDY QUESTION What are the most relevant factors associated with non-alcoholic fatty liver disease (NAFLD) in women with polycystic ovary syndrome (PCOS)? SUMMARY ANSWER Insulin resistance (IR) and lipid accumulation product (LAP) are independently associated with NAFLD in PCOS. WHAT IS KNOWN ALREADY Obesity and IR are frequently present in both women with PCOS and subjects having NAFLD. The coexistence of PCOS and NAFLD might synergistically increase the risk for both type 2 diabetes (T2DM) and cardiovascular disease (CVD). LAP, calculated from waist circumference (WC) and triglycerides (TGs) concentrations [(WC-58) × TGs], has been shown to represent an integrated marker of cardiometabolic risk in women with PCOS. STUDY DESIGN, SIZE, DURATION This cross-sectional study included 600 Caucasian women diagnosed with PCOS by the Rotterdam criteria between May 2008 and May 2013. PARTICIPANTS, SETTINGS, METHODS The study was done at the university hospitals in Belgrade, Serbia and Thessaloniki, Greece. All subjects underwent anthropometric measurements and analyses of fasting blood glucose, insulin, lipids, total testosterone and SHBG, as well as liver tests (transaminases, γ-glutamyltransaminase, total bilirubin and alkaline phosphatase). Calculations for a NAFLD liver fat score (NAFLD-LFS) (with, accordingly, determination of metabolic syndrome and testing for T2DM) as well as homeostasis model assessment of IR (HOMA-IR), LAP as a marker of visceral adiposity, and free androgen index (FAI) were performed. We evaluated the prevance of NAFLD and analyzed associations of the above variables with NAFLD. MAIN RESULTS AND THE ROLE OF CHANCE NAFLD was more prevalent in patients with PCOS than in controls (50.6 versus 34.0%, respectively). Women with PCOS had higher readings for WC, LAP, insulin and HOMA-IR, total cholesterol and TGs than controls (P < 0.001). In PCOS women, the NAFLD-LFS significantly (P < 0.001) correlated with WC, BMI, glucose, HOMA-IR, TGs, LAP and FAI. In multivariate logistic regression, HOMA-IR and LAP were independently associated with NAFLD (P ≤ 0.001). LIMITATIONS, REASONS FOR CAUTION A possible weakness of the study may be the absence of structural confirmation of liver status. Hovewer, liver biopsy is invasive, difficult to perform in large populations and carries some risk of complications while magnetic resonance spectroscopy does not provide any information regarding the presence of fibrosis and is not routinely available. Another possible limitation could be the measurement of total testosterone by radioimmunoassay, which can be inaccurate when determining low levels of testosterone. Finally, fewer controls than subjects in the study group could have affected the significance of the results. WIDER IMPLICATIONS OF THE FINDINGS There is a debate on the most accurate clinical method for diagnosing liver disease as an early predictor of T2DM and CVD in general population and in PCOS women. There current study provided data on this issue from a cohort of Caucasian women with PCOS. STUDY FUNDING/COMPETING INTERESTS The study was supported by a research grant by the Serbian Ministry of Science and Education (grant nos 41009 and 175032). All authors have no competing interests.

Nitric oxide synthases (NOSs) and Na+/K+-ATPase are enzymes essential for regular functioning of the heart. Since both enzymes are under insulin and androgen regulation and since insulin action and androgen level were disturbed in polycystic ovary syndrome (PCOS), we hypothesized that cardiac nitric oxide (NO) production and sodium/potassium transport would be deteriorated in PCOS. To test our hypothesis we introduced animal model of PCOS based on dihydrotestosterone (DHT) treatment of female Wistar rats and analyzed protein expression, phosphorylation or subcellular localization of endothelial NOS (eNOS), inducible NOS (iNOS) and alpha subunits of Na+/K+-ATPase in the heart. Obtained results indicate that DHT treatment significantly decreased cardiac eNOS protein level and activating phosphorylation at serine 1177, while inhibitory phosphorylation at threonine 495 was increased. In contrast to expression of eNOS, iNOS protein level in the heart of DHT-treated rats was significantly elevated. Furthermore, cardiac protein level of alpha 1 subunit of the ATPase, as well as its plasma membrane content, were decreased in rats with PCOS. In line with this, alpha 2 subunit protein level in fraction of plasma membranes was also significantly below control level. In conclusion, DHT treatment impaired effectiveness of NOSs and Na+/K+-ATPase in the female rat heart. Regarding the importance of NO production and sodium/potassium transport in the cardiac contraction and blood flow regulation, it implicates strong consequences of PCOS for heart functioning. Ministry of Education, Science and Technological Development, Republic of Serbia {[}III41009]

Gojaznost se danas smatra uzrokom nastanka kardiovaskularne bolesti, tipa 2 dijabetesa, osteoartritisa, maligniteta, ali i faktorom koji doprinosi nastanku reproduktivnih poremecaja i problema plodnosti. Postoji povecan relativni rizik za nastanak anovulatornog infertiliteta u žena sa izraženom gojaznoscu i produženo vreme do koncepcije. U žena u reproduktivnom periodu gojaznost je povezana sa povecanim rizikom za nastanak hiperandrogenizma i anovulacije, kao sto je slucaj u sindromu policisticnih jajnika (PCOS) kao najcescem hiperandrogenom poremecaju. Postoji veliki broj dokaza u prilog postojanja bliskog odnosa adipokina, gojaznosti, metabolickog sindroma i reproduktivnih posledica. Redukcija težine za 5-10% dovodi do poboljsanja u klinickim, metabolickim i reproduktivnim karakteristikama, kao sto je slucaj u žena sa PCOS. Primena insulinskih senzitajzera vodi sniženju hiperinsulinemije, insulinske rezistencije, uspostavljanju normalne menstrualne ciklicnosti i ovulacije kod znacajnog broja žena sa PCOS. Gojaznost može uticati na stimulaciju ovulacije njenim produžavanjem, povecanjem doze gonadotropina, incidence folikularne asinhronije i prekida stimulacije. Hirursko lecenje gojaznosti predstavlja alternativni vid terapije u slucaju kada ni promena nacina života ni farmakoterapijske mere ne daju povoljne rezultate. Za sada ne postoji dovoljno dokaza u prilog preporuke da se barijatrijska hirurgija koristi u lecenju gojaznih žena sa PCOS.

Polycystic ovary syndrome (PCOS) is a reproductive and metabolic disorder characterized by hyperandrogenism, ovulatory dysfunction, visceral obesity and insulin resistance. We hypothesized that changes in glucocorticoid metabolism and signaling in the visceral adipose tissue may contribute to disturbances of lipid metabolism in the rat model of PCOS obtained by 5 alpha-dihydrotestosterone (DHT) treatment of prepubertal female Wistar rats. The results confirmed that DHT treatment caused anovulation, obesity and dyslipidemia. Enhanced glucocorticoid prereceptor metabolism, assessed by elevated intracellular corticosterone and increased 11 beta-hydroxysteroid dehydrogenase type 1 mRNA and protein levels, was accompanied by glucocorticoid receptor (GR) nuclear accumulation. In concert with the increased expression of GR-regulated prolipogenic genes (lipin-1, sterol regulatory element binding protein 1, fatty acid synthase, phosphoenolpyruvate carboxykinase), histological analyses revealed hypertrophic adipocytes. The results suggest that glucocorticoids influence lipid metabolism in the visceral adipose tissue in the way that may contribute to pathogenesis of metabolic disturbances associated with PCOS. (C) 2014 Elsevier Ireland Ltd. All rights reserved.

Background. Women with polycystic ovary syndrome (PCOS) could develop subclinical atherosclerosis during life.Purpose. To analyze cardiovascular risk (CVR) factors and their relation to clinical markers of cardiovascular disease (CVD) in respect to their age.Material and Methods. One hundred women with PCOS (26.32±5.26years, BMI:24.98±6.38 kg/m2) were compared to 50 respective controls. In all subjects, total cholesterol (TC), HDL-C, LDL-C, triglycerides, TC/HDL-C and TG/HDL-C ratios, glucose, insulin and HOMA index, waist-to-hip ratio (WHR), systolic and diastolic blood pressure (SBP and DBP, resp.), and carotid intima-media thickness (CIMT) were analyzed in respect to their age and level of androgens.Results. PCOS over 30 years had higher WHR (P=0.008), SBP (P<0.001), DBP (P<0.001), TC (P=0.028), HDL-C (P=0.028), LDL-C (P=0.045), triglycerides (P<0.001), TC/HDL-C (P<0.001), and triglycerides/HDL-C (P<0.001) and had more prevalent hypertension and pronounced CIMT on common carotid arteries even after adjustment for BMI (P=0.005and 0.036, resp.). TC/HDL-C and TG/HDL-C were higher in PCOS with the highest quintile of FAI in comparison to those with lower FAI (P=0.045and 0.034, resp.).Conclusions. PCOS women older than 30 years irrespective of BMI have the potential for early atherosclerosis mirrored through the elevated lipids/lipid ratios and through changes in blood pressure.

Polycystic ovary syndrome (PCOS) is associated with an altered plasma lipid profile and increased risk for cardiovascular diseases. We hypothesized that molecular mechanisms underlying cardiac pathology in PCOS involve changes in expression and subcellular localization of several key proteins involved in cardiac lipid transport and metabolism, such as fatty acid transporter CD36, lipin 1, peroxisome proliferator-activated receptor alpha (PPAR alpha), peroxisome proliferator-activated receptor gamma coactivator-1 (PGC1), and carnitine palmitoyltransferase 1 (CPT1). We used the animal model of PCOS obtained by treating female rats with dihydrotestosterone (DHT). Protein levels of CD36, lipin 1, PPAR alpha, PGC1, and antioxidative enzymes were assessed by Western blot in different cardiac cell compartments. Cardiac triglycerides (TG) and lipid peroxidation were also measured. The content of CD36 was decreased in both the cardiac plasma membranes and intracellular pool. On the other hand, total content of cardiac lipin 1 in DHT-treated rats was elevated, in contrast to decreased microsomal lipin 1 content. An increase in nuclear content of lipin 1 was observed together with elevation of nuclear PPAR alpha and PGC1, and an increase in CPT1 expression. However, lipid peroxidation was reduced in the heart, without alterations in antioxidative enzymes expression and cardiac TG content. The results indicate that treatment of female rats with DHT is accompanied by a decrease of fatty acid uptake and a reduction of lipid peroxidation in the heart. The observed elevation of lipin 1, PPAR alpha, PGC1, and CPT1 expression suggests that cardiac fatty acid metabolism is shifted toward mitochondrial beta oxidation. Ministry of Education, Science and Technological Development, Republic of Serbia {[}III41009]

Summary Background: The polycystic ovary syndrome (PCOS) is a frequent endocrine disorder that affects women of reproductive age. As the syndrome is strongly associated with obesity, it is of interest to examine the gene expression diffe rences that accompany its development and the associ a ted metabolic disturbances. Real-time RT PCR is a standard method for studying changes in gene expression. However, to obtain accurate and reliable results, validation of reference genes is obligatory. The aim of this study was to identify a suitable reference for the normalization of gene expression in peripheral blood mononuclear cells (PBMCs) from obese and normal-weight women with PCOS. Methods: The expression stability of four potential reference genes: hypoxanthine guanine phosphoribosyl trans-ferase 1 (HPRT), β-actin (BA), β2-microglobulin (B2M) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), was assessed in PBMCs from healthy women, and from normal-weight and obese women with PCOS. The variability in the expression of potential reference genes was analyzed by the TaqMan real-time RT PCR method, using GeNorm and NormFinder software packages. Results: Direct comparison of cycle threshold (Ct) values showed inter-individual variations for all validated genes, the Ct values of HPRT being less variable than those of BA, GAPDH and B2M. Both software packages pointed to HPRT as the most steadily expressed gene in the PBMCs of women with PCOS and healthy controls. Conclusions: Cross-validation of the expression stability of four potential reference genes identified HPRT as the most stable reference, suitable for further investigations of gene expression in PBMCs from women with PCOS.

It is supposed that women with polycystic ovary syndrome (PCOS) are prone to develop cardiovascular disease as a consequence of multiple risk factors that are mostly related to the state of insulin resistance and consequent hyperinsulinemia. In the present study, we evaluated insulin signaling and glucose transporters (GLUT) in cardiac cells of dihydrotestosterone (DHT) treated female rats as an animal model of PCOS. Expression of proteins involved in cardiac insulin signaling pathways and glucose transporters, as well as their phosphorylation or intracellular localization were studied by Western blot analysis in DHT-treated and control rats. Treatment with DHT resulted in increased body mass, absolute mass of the heart, elevated plasma insulin concentration, dyslipidemia and insulin resistance. At the molecular level, DHT treatment did not change protein expression of cardiac insulin receptor and insulin receptor substrate 1, while phosphorylation of the substrate at serine 307 was increased. Unexpectedly, although expression of downstream Akt kinase and its phosphorylation at threonine 308 were not altered, phosphoiylation of Akt at serine 473 was increased in the heart of DHT-treated rats. In contrast, expression and phosphorylation of extracellular signal regulated kinases 1/2 were decreased. Plasma membrane contents of GLUT1 and GLUT4 were decreased, as well as the expression of GLUT4 in cardiac cells at the end of androgen treatment. The obtained results provide evidence for alterations in expression and especially in functional characteristics of insulin signaling molecules and glucose transporters in the heart of DHT-treated rats with PCOS, indicating impaired cardiac insulin action. (c) 2014 Elsevier Ltd. All rights reserved. Ministry of Education, Science and Technological Development, Republic of Serbia {[}III41009]

OBJECTIVE: There is a need for a simple and accurate method for the assessment of cardiovascular risk in polycystic ovary syndrome (PCOS). Lipid accumulation product (LAP) is based on the assessment of waist circumference and serum triglycerides that yield an estimation of lipid overaccumulation. We aimed to determine whether LAP is associated with metabolic syndrome (MetS) in Caucasian women with PCOS. DESIGN: We studied 222 women with PCOS who were diagnosed using the Rotterdam criteria. In all the subjects and controls, LAP was determined and the MetS was assessed using three different international criteria, NCEP-ATP III, IDF, and JIS. ROC curve and logistic regression analyses were performed to determine and analyze associations with the MetS. RESULTS: In the study population the prevalence of MetS was 16.2–19.4%. The cut-off value of 25.9 determined that LAP has the strongest association with MetS whichever international criteria are used, followed by HDL (NCEP-ATP III and JIS) and glucose (IDF). CONCLUSIONS: LAP is used as an independent clinical indicator for MetS in our PCOS women of Caucasian origin. The high diagnostic accuracy of LAP is superseding the need for the use of multiple clinical indicators for the assessment of lipid accumulation as a prerequisite for diagnosis of metabolic and cardiovascular diseases in PCOS women.

OBJECTIVE: Most women with PCOS have increased adrenal androgen production, enhanced peripheral metabolism of cortisol and elevation in urinary excretion of its metabolites. Increased cortisol clearance in PCOS is followed by a compensatory overdrive of the hypothalamic-pituitary-adrenocortical (HPA) axis. We hypothesized that oral contraceptives containing ethinylestradiol and drospirenone (EE-DRSP) could modulate glucocorticoid receptor (GR) expression and function and thus affect HPA axis activity in PCOS patients. DESIGN: We analyzed 12 women with PCOS (age 24.17 +/- 4.88 years; body mass index 22.05 +/- 3.97 kg/m(2)) treated for 12 months with EE-DRSP and 20 BMI matched controls. In all subjects testosterone, dehydroepiandrosterone sulfate (DHEAS), sex hormone binding globulin (SHBG), cortisol (basal and after dexamethasone), concentrations of GR protein, phospo-GR211 protein, number of GR per cell (B-max) and its equilibrium dissociation constant (K-D) were measured. RESULTS: Before treatment, increased concentrations of testosterone and DHEAS (p