DNA Damage Estimation after Chronic and Combined Exposure to Endocrine Disruptors: An In Vivo Real-Life Risk Simulation Approach.

Exposure to chemical substances has always been a matter of concern for the scientific community. During the last few years, researchers have been focusing on studying the effects resulting from combined exposure to different substances. In this study, we aimed to determine the DNA damage caused after chronic and combined exposure to substances characterized as endocrine disruptors using comet and micronuclei assays, specifically glyphosate (pure and commercial form), bisphenol A, parabens (methyl-, propyl- and butylparaben), triclosan and bis (2-ethylhexyl) phthalate. The highest mean tail intensity was observed in the group exposed to a high-dose (10 × ADI) mixture of substances (Group 3), with a mean value of 11.97 (11.26-13.90), while statistically significant differences were noticed between the groups exposed to low-dose (1 × ADI) (Group 2) and high-dose (10 × ADI) (Group 3) mixtures of substances (p = 0.003), and between Group 3 and both groups exposed to high doses (10 × ADI) of the pure and commercial forms of glyphosate (Groups 4 (p = 0.014) and 5 (p = 0.007)). The micronuclei assay results were moderately correlated with the exposure period. Group 5 was the most impacted exposure group at all sampling times, with mean MN counts ranging between 28.75 ± 1.71 and 60.75 ± 1.71, followed by Group 3 (18.25 ± 1.50-45.75 ± 1.71), showing that commercial forms of glyphosate additives as well as mixtures of endocrine disruptors can enhance MN formation. All exposure groups showed statistically significant differences in micronuclei counts with an increasing time trend.

Ageing, a modulator of human endometrial stromal cell proliferation and decidualization: a role for implantation?

RESEARCH QUESTION: Does ageing affect endometrial stromal cell function and gene expression involved in decidualization? DESIGN: Stromal cells were isolated and cultured (98% purity) from pipelle endometrial biopsies obtained from female donors, at the proliferative phase of the menstrual cycle. Initially, 28 samples were collected (age range 25-46 years). These samples were divided into two groups: 25-35 years and 36-46 years. Cell proliferation assays were carried out to determine changes in stromal cell proliferation, in vitro, between the two groups. In addition, mRNA and protein expression of decidualization factors, BMP2 and STAT3, were analysed for the same age groups and samples using real-time polymerase chain reaction and western blot analysis, respectively. Finally, another 12 pipelle endometrial biopsies (age range 25-46 years) were used in separate in-vitro decidualization experiments. The mRNA expression of decidualization markers, prolactin and IGFBP-1 were analysed in cultured stromal cells after adding 8-bromo-cAMP. RESULTS: A statistically significant decrease was observed in stromal cell proliferation with increasing age (P = 0.0006). Messenger RNA expression of BMP2 and STAT3 were found to decrease (P = 0.0167; P = 0.0037, respectively) in the older age group. In addition, BMP2 and STAT3 protein expression between the samples analysed changed significantly (P = 0.0085; P = 0.0463, respectively) with increasing age. In-vitro induced decidualization experiments showed significant changes in stromal cell mRNA expression of decidualization markers (IGFBP1 and prolactin) between different age groups. CONCLUSION: Ageing affected endometrial cell function and gene expression. Changes in cell function and expression of associated molecules that differ in the ageing endometrium can help understand the causes of infertility.

A prospective, cross-sectional study of the protective and risk psychological factors of successful in vitro fertilisation outcome: preliminary results in a Greek sample.

The aim of this prospective, cross-sectional study was to examine the protective and risk psychological factors associated with the successful outcome of In vitro fertilisation (IVF). Various psychological factors that may affect the IVF outcome were measured to a sample of 61 infertile women (mean age 37.2 ± 4.4), who started their first or consecutive IVF treatment cycle in an IVF Unit in Greece. Over half of the participants (50.8%) became pregnant. A binary logistic regression analysis (stepwise) was conducted on pregnancy as the outcome, with various variables as predictors. The model was statistically significant (Omnibus Chi-square = 27.324, df = 5, p < .001), explained 54.7% of the variance, and correctly classified 84.6% of the cases. Life purpose (odds ratio [OR] = 1.35, 95% CI = 1.02-1.78) and negative emotions (e.g. discontent, sorrow) (OR = 1.76, 95% CI = 1.19-2.60) were associated with increased pregnancy rates, whereas autonomy (OR = 0.57, 95% CI = 0.39-0.82), and stress (OR = 0.69, 95% CI = 0.55-0.88) were associated with decreased pregnancy rates. It has been concluded that the relationship between psychological factors and successful IVF outcome is more complex than commonly believed. The identification of the risk and protective psychological factors could contribute to increased pregnancy rates and foster the implementation of tailored therapeutic interventions.Impact statementWhat is already known on this subject? High levels of infertility stress and/or depression have been associated with lower pregnancy rates. However, little is known on the impact of emotions, personality characteristics and other psychological variables on in vitro fertilisation (IVF) outcome.What the results of this study add? A combination of commonly believed 'negative' factors (e.g. stress) and 'positive' ones (e.g. well-being) may diversely affect the IVF outcome. Otherwise believed to be positive aspects of human life (i.e. autonomy) may decrease the likelihood of pregnancy, and other factors believed to be 'negative' (e.g. negative emotions) may increase pregnancy rates.What the implications of these findings are for clinical practice and/or further research? The findings invite researchers to further examine the role of the psychological factors which could potentially affect pregnancy rates. Modifiable factors, such as well-being, stress and emotions, should guide tailored interventions aimed at increasing the pregnancy rates in infertile women.

Title: Repeated implantation failure is associated with increased Th17/Treg cell ratio, during the secretory phase of the human endometrium.

Repeated implantation failure (RIF) is a significant limiting factor in assisted reproduction. Chronic endometrial inflammation has been noted in RIF women, therefore we sought to investigate the potential association of endometrial Th17/Treg ratio and endometrial inflammation in these cases. Endometrial pipelle biopsies were obtained from volunteers, 29 women with RIF (failure to achieve pregnancy following at least 3 transfers of high-grade embryos in IVF-cycles) and 27 fertile women (at least one child) in total, at the secretory phase of the menstrual cycle. Using tissues from 17 fertile and 18 RIF endometrial samples, stromal and immune cells were isolated and flow cytometry analysis was performed to determine Th17 and CD4+ CD25high FOXP3+ cell populations in endometrial stromal cell suspensions. Another group of tissues from 10 fertile and 11 RIF samples were used for mRNA expression levels of Treg and Th17-cell transcription factors, FOXP3 and RORγt respectively. Endometrial inflammatory mediators' mRNA expression was also analyzed. A statistically significant increase in protein flow cytometry analysis of Th17/Treg ratio (p ≤ 0.05) as well as a reduction in absolute Treg cells in the endometrium (p ≤ 0.05) was noted in women with RIF. Additionally, RNA analysis on the same set of women indicated RORγt/FOXP3 significantly increased in women with RIF compared to fertile ones (p ≤ 0.05). Finally, women with RIF exhibited significantly (p ≤ 0.05) elevated mRNA levels of pro-inflammatory mediators (ΤΝF-a, ΙL-6, IL-8 and CCl2). Women with RIF exhibit elevated Th17/Treg ratio, mostly due to endometrial Treg depletion, as well as a pro-inflammatory state in the endometrium.

A combined opposite targeting of p110δ PI3K and RhoA abrogates skin cancer.

Malignant melanoma is the most aggressive and deadly skin cancer with an increasing incidence worldwide whereas SCC is the second most common non-melanoma human skin cancer with limited treatment options. Here we show that the development and metastasis of melanoma and SCC cancers can be blocked by a combined opposite targeting of RhoA and p110δ PI3K. We found that a targeted induction of RhoA activity into tumours by deletion of p190RhoGAP-a potent inhibitor of RhoA GTPase-in tumour cells together with adoptive macrophages transfer from δD910A/D910A mice in mice bearing tumours with active RhoA abrogated growth progression of melanoma and SCC tumours. Τhe efficacy of this combined treatment is the same in tumours lacking activating mutations in BRAF and in tumours harbouring the most frequent BRAF(V600E) mutation. Furthermore, the efficiency of this combined treatment is associated with decreased ATX expression in tumour cells and tumour stroma bypassing a positive feedback expression of ATX induced by direct ATX pharmacological inactivation. Together, our findings highlight the importance of targeting cancer cells and macrophages for skin cancer therapy, emerge a reverse link between ATX and RhoA and illustrate the benefit of p110δ PI3K inhibition as a combinatorial regimen for the treatment of skin cancers.

Association of placental inflammation with fetomaternal hemorrhage and loss of placental mucin-1.

BACKGROUND: Fetomaternal hemorrhage (FMH) poses an immediate risk to the fetus and, in case of Rhesus-immunization, to future pregnancies. Given that altered endothelial permeability is part of the pathophysiology of inflammation, in this study we investigated whether placental inflammatory processes like chorioamnionitis (ChoA) or preeclampsia (PE) lead to increased rates of FMH compared to the established risk factor of placenta previa (PP). Putative accompanying markers of trophoblastic damage were also explored. METHODS: 40 patients (14 PE; 6 ChoA; 9 PP; 11 normal controls) were evaluated for FMH using a flowcytometric test kit, which is able to quantify FMH of 0.06% fetal cells. Placental tissue samples were immunostained for human placental lactogen (hPL), human chorionic gonadotropin (hCG), and mucin-1 (MUC1). MUC1 was evaluated as a potential serum marker of FMH. RESULTS: Patients with ChoA had a mean calculated FMH volume of 29 ml, compared to 4 ml in PE and 1 ml in PP and controls. MUC1 staining was reduced in PE and ChoA placenta samples, while elevated MUC1 serum concentration correlated positively with FMH. CONCLUSION: Diseases of placental inflammation are associated with FMH. Placental MUC1 staining is reduced and serum concentrations are increased in cases of FMH.

Endocrine Disruptors Acting on Estrogen and Androgen Pathways Cause Reproductive Disorders through Multiple Mechanisms: A Review.

Increasing contamination of the environment by toxic compounds such as endocrine disrupting chemicals (EDCs) is one of the major causes of reproductive defects in both sexes. Estrogen/androgen pathways are of utmost importance in gonadal development, determination of secondary sex characteristics and gametogenesis. Most of the EDCs mediate their action through respective receptors and/or downstream signaling. The purpose of this review is to highlight the mechanism by which EDCs can trigger antagonistic or agonistic response, acting through estrogen/androgen receptors causing reproductive defects that lead to infertility. In vitro, in vivo and in silico studies focusing on the impact of EDCs on estrogen/androgen pathways and related proteins published in the last decade were considered for the review. PUBMED and PUBCHEM were used for literature search. EDCs can bind to estrogen receptors (ERα and ERß) and androgen receptors or activate alternative receptors such as G protein-coupled receptors (GPCR), GPR30, estrogen-related receptor (ERRγ) to activate estrogen signaling via downstream kinases. Bisphenol A, dichlorodiphenyltrichloroethane, dichlorodiphenyldichloroethylene, polychlorinated biphenyls and phthalates are major toxicants that interfere with the normal estrogen/androgen pathways leading to infertility in both sexes through many ways, including DNA damage in spermatozoids, altered methylation pattern, histone modifications and miRNA expression.

Pregnancy after breast cancer. A comprehensive review.

Pregnancy after breast cancer treatment has become an important issue since many young breast cancer patients have not completed their family. Generally, these patients should not be discouraged to become pregnant when they want to, since published data suggest no adverse effect of pregnancy on survival. As fertility may be impaired by chemotherapy, different fertility preserving strategies have been developed. Births seem to sustain no adverse effects, while breastfeeding appears to be feasible and safe.

Intrauterine G-CSF Administration in Recurrent Implantation Failure (RIF): An Rct.

This study investigates the effects of intrauterine G-CSF on endometrial thickness, clinical pregnancy rate and live birth rate in a recurrent implantation failure (RIF) group with normal endometrium. This study was designed as a prospective randomized controlled trial with the involvement of 157 RIF group pati; ents. The RIF group was formed on the basis of the RIF criteria: "The failure to achieve a clinical pregnancy after the transfer of at least four good-quality embryos in a minimum of three fresh or frozen cycles to a woman under the age of 40 years. The study sample included 82 patients in the G-CSF group who received G-CSF once a day on hCG. The procedure was performed by administering 30 mIU of Leucostim®(Filgrastim [G-CSF] 30 mIU/mL; DEM Medical, Dong-A; South Korea) through slow infusion into the endometrial cavity using a soft embryo transfer catheter. Normal saline of 1 mL was infused into the endometrial cavity in the same way in 75 patients in the control group. The standard ICSI procedure was used for all patients, and fresh cycle embryos were transferred on the third or fifth day. No statistically significant difference was identified in clinical pregnancy rates, miscarriage rates and live birth rates between the G-CSF group and the control group (p = 0.112, p = 0.171, p = 0.644, respectively), and no difference was observed between the two groups regarding endometrial thickness (p = 0.965). The intervention of administration G-CSF into the uterine cavity in RIF patients with normal endometrium, did not alter the endometrial thickness, clinical pregnancy rates, or live birth rates.

The association of female and male infertility with telomere length (Review).

Telomere length (TL) has long been associated with aging, as telomeres serve as protective caps of chromosomes, and are thus deeply involved in the preservation of genome integrity and are vital to cellular functions. Traditionally, a strong link connects aging and infertility in both sexes, with an earlier onset in females. Over the past decade, telomeres have attracted increasing attention due to the role they play in fertility. In this review, we investigated the potential positive or negative association between relative TL and different factors of female and male infertility. A systematic search of the PubMed database was conducted. Out of the 206 studies identified, 45 were reviewed as they fulfilled the criteria of validity and relevance. Following an analysis and a comparison of the study outcomes, several clear trends were observed. The majority of female infertility factors were associated with a shorter TL, with the exception of endometriosis, premature ovarian failure and clear cell carcinoma that were associated with a longer TL and polycystic ovary syndrome (PCOS), which revealed conflicting results among several studies, leading to ambiguous conclusions. Male infertility factors were associated with a shorter TL. Although this review can provide an outline of general trends in the association of TL with infertility factors, further epidemiological and original research studies are required to focus on investigating the basis of these varying lengths of telomeres.

Expression of the inhibin/activin subunits (-alpha, -betaA and -betaB) in normal and carcinogenic endometrial tissue: possible immunohistochemical differentiation markers.

Inhibins (INH) are dimeric glycoproteins, composed of an alpha-subunit (INH-alpha) and one of two possible beta-subunits (INH-betaA or -betaB), with substantial roles in human reproduction and in endocrine-responsive tumors. The aims of this study were to determine the frequency and tissue distribution of INH-alpha, -betaA and -betaB in normal and malignant endometria. Samples were obtained from normal (n=46), atrophic (n=8) and endometrioid carcinoma tissue (EC; G1=93; G2=32; G3=14). INH-alpha was significantly higher in normal compared to malignant endometrial tissue, showing a cyclical variation throughout the menstrual cycle. EC G3 did not express this subunit. INH-betaA and -betaB showed specific staining reactions within the tumor cells. The highest intensity of INH-betaA was observed in the normal secretory phase compared to adenocarcinomas (p<0.05). For INH-betaB, the significantly highest expression was noted in EC G3 compared to EC G2 (p<0.05) and atrophic endometrial tissue. In conclusion, INH-alpha, -betaA and -betaB were immunolabeled in normal and malignant endometria. INH-alpha was expressed in a declining relationship in the transition from normal to tumor tissue, suggesting a tumor suppressive function in EC. A high expression of INH-betaB was observed in EC G3 compared to G2, suggesting an important role in the progression of endometrial carcinogenesis. However, the utilization of these subunits as specific tumor markers still remains unclear.

Endocrine Disruptors Leading to Obesity and Related Diseases.

The review aims to comprehensively present the impact of exposure to endocrine disruptors (EDs) in relation to the clinical manifestation of obesity and related diseases, including diabetes mellitus, metabolic syndrome, cardiovascular diseases, carcinogenesis and infertility. EDs are strong participants in the obesity epidemic scenery by interfering with cellular morphological and biochemical processes; by inducing inflammatory responses; and by presenting transcriptional and oncogenic activity. Obesity and lipotoxicity enhancement occur through reprogramming and/or remodeling of germline epigenome by exposure to EDs. Specific population groups are vulnerable to ED exposure due to current dietary and environmental conditions. Obesity, morbidity and carcinogenicity induced by ED exposure are an evolving reality. Therefore, a new collective strategic approach is deemed essential, for the reappraisal of current global conditions pertaining to energy management.

3C syndrome with cryptorchidism and posterior embryotoxon.

We report a case of the 3C (cranio-cerebello-cardiac) syndrome, also known as Ritscher-Schinzel syndrome, a rare autosomal recessive disorder characterized by craniofacial, cerebellar, and cardiac anomalies. In addition to features previously reported the child had Wormian bones of the skull, intra-abdominal testes, and posterior embryotoxon that have not previously been reported as part of the 3C syndrome.

Prevalence of human herpes virus types 1-7 in the semen of men attending an infertility clinic and correlation with semen parameters.

OBJECTIVE: To determine the prevalence of herpes viruses in the semen of an asymptomatic male cohort with and without infertility problems and its association with altered semen parameters. DESIGN: A prospective randomized study. SETTING: Medical school and IVF clinic. PATIENT(S): One hundred seventy-two male patients undergoing routine semen analysis: 80 with normal semen parameters (control group) and 92 with abnormal semen parameters. INTERVENTION(S): Semen samples were collected by masturbation. MAIN OUTCOME MEASURE(S): The DNA from the Herpesviridae family (herpes simplex virus 1 [HSV-1], herpes simplex virus 2 [HSV-2], Varicella zoster virus [VZV], Epstein-Barr virus [EBV], cytomegalovirus [CMV], human herpes virus type 6 [HHV-6], human herpes virus type 7 [HHV-7]) and routine semen parameters. RESULT(S): Viral DNA was detected in 143/172 (83.1%) of the total samples for at least one herpes virus: HSV-1, 2.5%; VZV, 1.2%; EBV, 45%; CMV, 62.5%; HHV-6, 70%; HHV-7, 0% in the normal semen samples and HSV-1, 2.1%; VZV, 3.2%; EBV, 39.1%; CMV, 56.5%; HHV-6, 66.3%; HHV-7, 0% in the abnormal semen samples. No association was found between the presence of viral DNA and semen parameters. Interestingly, a statistical significance between leukocytospermia and the presence of EBV DNA was observed. CONCLUSION(S): The DNA of herpes viruses is frequently detected in the semen of asymptomatic fertile and infertile male patients. Further studies are required to investigate the role of herpes viruses in male factor infertility.

Laparoscopic evaluation of infertile patients with chronic pelvic pain.

In this study over a 10-year period, 1584 patients complaining of infertility of more than 1 year duration were evaluated for their laparoscopic findings in relation to the presence or not of chronic pelvic pain (CPP). Infertility was the only complaint in 1215 cases (group 1), whereas 369 patients complained of infertility and CPP (group 2). All cases underwent routine infertility investigation and pelvic ultrasonography, followed by diagnostic laparoscopy, with infertility-only cases acting as a control group. At laparoscopy 76.7% of patients with CPP were found with pelvic pathology, compared with only 42.6% of cases without CPP (P < or = 0.0001). Omental-abdominal wall adhesions, advanced endometriosis, endometriomas with adhesions, pelvic venous congestion, and hydrosalpinges with pelvic adhesions were significantly more frequent in cases with CPP. Dysmenorrhoea was the most frequent type of CPP. Cases with CPP and a negative laparoscopy were further investigated using a multidisciplinary approach. In conclusion, chronic pelvic pain can be the result of several pelvic pathologies. Infertile patients with CPP are much more frequently found with an abnormal pelvis in comparison with cases without CPP. Laparoscopy is an invaluable diagnostic tool especially for symptomatic patients and should be used early in their diagnostic infertility work-up.