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Self-sampling for human papillomavirus (HPV) testing is an alternative to physician sampling particularly for cervical cancer screening nonattenders. The GRECOSELF study is a nationwide observational cross-sectional study aiming to suggest a way to implement HPV-DNA testing in conjunction with self-sampling for cervical cancer screening in Greece, utilizing a midwifery network. Women residing in remote areas of Greece were approached by midwives, of a nationwide network, and were provided with a self-collection kit (dry swab) for cervicovaginal sampling and asked to answer a questionnaire about their cervical cancer screening history. Each sample was tested for high-risk (hr) HPV with the Cobas HPV test. HrHPV-Positive women were referred to undergo colposcopy and, if needed, treatment according to colposcopy/biopsy results. Between May 2016 and November 2018, 13,111 women were recruited. Of these, 12,787 women gave valid answers in the study questionnaire and had valid HPV-DNA results; hrHPV prevalence was 8.3%; high-grade cervical/vaginal disease or cancer prevalence was 0.6%. HrHPV positivity rate decreased with age from 20.7% for women aged 25–29 years to 5.1% for women aged 50–60 years. Positive predictive value for hrHPV testing and for HPV16/18 genotyping ranged from 5.0% to 11.6% and from 11.8% to 27.0%, respectively, in different age groups. Compliance to colposcopy referral rate ranged from 68.6% (for women 25–29) to 76.3% (for women 40–49). For women residing in remote areas of Greece, the detection of hrHPV DNA with the Cobas HPV test, on self-collected cervicovaginal samples using dry cotton swabs, which are provided by visiting midwives, is a promising method for cervical cancer secondary prevention.

Suppl. 1. Comparison of different storage and transportation conditions

Background: Human papillomavirus (HPV)-DNA testing combined with self-sampling could increase cervical cancer screening effectiveness, utilizing a sensitive screening modality and an easy sampling method with minimal pain or discomfort. Self-sampling acceptability, though, is pivotal. Materials and Methods: This study is a nested cross-sectional survey within GRECOSELF, a cross-sectional study on HPV-based screening with self-sampling, aiming at investigating self-sampling acceptability among Greek women residing in rural areas, and the factors affecting it. Women between 25 and 60 years old were recruited by midwives participating in a nationwide midwifery network. Participants, after self-sampling, filled out a questionnaire with three sections, one regarding demographic characteristics, a second with questions pertaining to the participants' cervical cancer screening history, and a third with questions regarding the self-sampling process per se. Results: The sample included 13,111 women. Most participants (67.9%), including those screened or not in the past, would prefer self-sampling if assured that the results are not inferior to standard testing. Discomfort or pain during self-sampling was absent or minimal in 97.1% and 96.5% of the cases, respectively, and 74.4% of the women felt adequately confident that they followed the instructions correctly. Women mostly preferred self-sampling at home compared with health care facilities. Pain and discomfort during the procedure, although rare, were significant factors against acceptance. Most of the women reporting a negative impression had a negative experience with conventional sampling in the past. Conclusion: Self-sampling is highly acceptable. Acceptance can be further improved with proper communication of the process and its noninferiority compared with conventional screening.

Background Although Candida is a commensal of the urogenital tract, intrauterine fungal infections are extremely uncommon in clinical practice. Aims In the present work we evaluated whether amniotic fluid (AF) possesses direct antifungal activity against clinical isolates of Candida albicans and other Candida species. Methods A total of 23 AF samples from pregnant women with gestational age of 38–41 weeks were obtained under aseptic conditions by the aspiration of the amniotic sac during cesarean section. Different Candida species were inoculated in amniotic fluid and Sabouraud broth, used as control, and were incubated at 37 °C for 48 h. Quantitative cultures of test samples and controls were performed at 0, 4, 8, 12, 24, and 48 h. Results AF collected from 23 pregnant women had consistent and significant inhibitory activity against all Candida isolates tested. Nonetheless, a complete inhibition of growth by all 23 AF samples tested was observed only against Candida glabrata. Conclusions It is likely that the antifungal activity of the AF against C. albicans, C. glabrata and Candida parapsilosis observed in vitro also exists in vivo, contributing to protect against intrauterine fungal infections.

Endometriosis-related chronic pelvic pain is a major component of the disease that affects quality of life in women of reproductive age suffering from endometriosis. The present review summarizes current evidence upon pathophysiology and its impact on mental health. It seems that endometriosis-related chronic pelvic pain is the result of chronic stress on the central nervous system as a consequence of chronic pelvic inflammation. Mental health issues may rise as a result of central nervous system derangement and further aggravate pain perception and therefore quality of life. Further properly designed studies are needed in order to elucidate the connection between mental disorders and endometriosis-related chronic pelvic pain.

Self-sampling for human papillomavirus (HPV) testing is an alternative to physician sampling particularly for cervical cancer screening nonattenders. The GRECOSELF study is a nationwide observational cross-sectional study aiming to suggest a way to implement HPV-DNA testing in conjunction with self-sampling for cervical cancer screening in Greece, utilizing a midwifery network. Women residing in remote areas of Greece were approached by midwives, of a nationwide network, and were provided with a self-collection kit (dry swab) for cervicovaginal sampling and asked to answer a questionnaire about their cervical cancer screening history. Each sample was tested for high-risk (hr) HPV with the Cobas HPV test. HrHPV-Positive women were referred to undergo colposcopy and, if needed, treatment according to colposcopy/biopsy results. Between May 2016 and November 2018, 13,111 women were recruited. Of these, 12,787 women gave valid answers in the study questionnaire and had valid HPV-DNA results; hrHPV prevalence was 8.3%; high-grade cervical/vaginal disease or cancer prevalence was 0.6%. HrHPV positivity rate decreased with age from 20.7% for women aged 25–29 years to 5.1% for women aged 50–60 years. Positive predictive value for hrHPV testing and for HPV16/18 genotyping ranged from 5.0% to 11.6% and from 11.8% to 27.0%, respectively, in different age groups. Compliance to colposcopy referral rate ranged from 68.6% (for women 25–29) to 76.3% (for women 40–49). For women residing in remote areas of Greece, the detection of hrHPV DNA with the Cobas HPV test, on self-collected cervicovaginal samples using dry cotton swabs, which are provided by visiting midwives, is a promising method for cervical cancer secondary prevention.

Recent advances in Spatial Temporal Image Correlation (STIC) 4 D fetal echocardiography include the application of eSTIC based on electronic probe image acquisition. We aimed to directly compare the performance of conventional STIC versus eSTIC technique (B-Mode and color Doppler imaging) during off-line reconstruction of STIC/eSTIC fetal heart volume pairs.Pairs of B-Mode and Color Doppler STIC volumes were acquired sequentially by firstly conventional (STIC) followed by electronic (eSTIC) probes during 33 consecutive obstetric scans at median 23 (range 13-31) gestational weeks. The resulting 66 fetal heart volume pairs were assessed blindly off-line by a fetal cardiologist who documented feasibility of reconstruction, presence of motion artifacts, subjective image quality on a 4-level scale: 1-best to 4-non-diagnostic and morphological diagnosis, to enable a paired comparison of STIC and eSTIC in the same fetus under similar scanning conditions.eSTIC volumes had higher temporal resolution (37 vs. 24 frames per second,eSTIC was associated with more effective 4 D fetal heart reconstruction due to reduced motion artifacts and superior image quality in all planes, when compared to STIC. Early gestation reconstructions were not generally successful using either technology. Further study is needed to define the cost-effectiveness and diagnostic impact of eSTIC over conventional STIC and their role over, or in addition to, screening 2 D fetal echocardiography by appropriately trained sonographers.

Primary splenic pregnancy is a rare entity, usually treated with splenectomy. We report the first case of conservative management of splenic pregnancy with selective embolization and intramuscular methotrexate administration. Postoperative treatment was uneventful. We have further systematically reviewed the literature upon the treatment options in case of splenic pregnancy. It has been evident that most cases were treated with spleenectomy, especially in case of active hemorrhage. Spleen preservation is rarely achieved, only in cases of hemodynamically stable, uncomplicated patients. Splenic selective embolization could be a treatment option in case of uncomplicated splenic pregnancy with the benefit of splenic preservation.

A randomized control trial of 64 hyperprolactinemic women with Recurrent Pregnancy Loss (RPL) treated with bromocriptine showed a higher incidence of live births, while PRL levels were significantly higher in women who miscarried. Progesterone is essential for successful implantation and maintenance of a normal pregnancy. Therefore, disorders related to inadequate progesterone secretion by the corpus luteum may affect the outcome of the pregnancy. One plausible explanation for the relationship between hypothyroidism and pregnancy loss is luteal phase defect linked to a hypofunctioning thyroid. Untreated hypothyroidism in pregnancy is associated with a greater risk for adverse pregnancy complications, such as miscarriage, premature birth, low birth weight, and detrimental effects on fetal neurocognitive development. Treatment of endometriosis has been shown to be beneficial for future fertility and improved pregnancy outcomes. Studies from in vitro fertilization have documented decreased pregnancy rates that can be improved with gonadotropin-releasing hormone agonist suppression, surgery, or aromatase inhibitor therapy.

Endometrial corticotropin-releasing hormone (CRH) has been described as a mediator of decidualisation and as a contributor of maternal-fetal immunotolerance. Deregulation of the CRH expression pattern has been associated with unfavourable reproductive outcomes as well as chronic endometrium-derived inflammatory disorders, such as endometriosis and adenomyosis. The current review summarises the evidence produced regarding the role of CRH in endometrial physiology and pathophysiology and highlights recent clinical data regarding the role of CRH in improving clinical pregnancy rates in women with repeated implantation failures following in vitro fertilisation and embryo transfer.

Insulin-like growth factor (IGF)-axis is involved in human oncogenesis and metastasis development for various solid tumors including breast cancer. Aim of this study was to assess the association between IGF-1, IGF-binding protein-3 (IGFBP-3) serum levels and the presence of circulating tumor cells (CTCs) in the peripheral blood of women diagnosed with early breast cancer (EBC), before and after adjuvant chemotherapy.171 patients with early-stage breast adenocarcinomas were retrospectively evaluated. Immunoradiometric (IRMA) assays were employed for the in-vitro determination of IGF-1 and IGFBP-3 serum levels in blood samples collected after surgical treatment and before initiation of adjuvant chemotherapy. CTCs' presence was assessed through detection of cytokeratin-19 (CK-19) mRNA transcripts using quantitative real time reverse transcription polymerase chain reaction (RT-PCR). IGF-1, IGFBP-3 serum levels were correlated with CTCs' presence before and after adjuvant chemotherapy as well as with tumor characteristics including tumor size, axillary lymph node status, oestrogen (ER)/progestorene (PR) and human epidermural growth factor receptor 2 (HER2) receptor status. Log-rank test was applied to investigate possible association between IGF-1, IGFBP-3 serum levels and disease-free interval (DFI) and overall survival (OS).Before initiation of adjuvant therapy IGF-1, IGFBP-3 serum levels were moderately associated (Spearman's rho=0.361, p0.001) with each other, while presenting significant differences across age groups (all p values0.05). IGF-1 serum levels did not correlate with the presence of CTCs before initiation (p=0.558) or after completion (p=0.474) of adjuvant chemotherapy. Similarly, IGFBP-3 serum levels did not show significant association with detectable CTCs either before (p=0.487) or after (p=0.134) completion of adjuvant chemotherapy. There was no statistically significant association between the clinical outcome of patients in terms of DFI, OS and IGF-1(DFI: p=0.499; OS: p=0.220) or IGFBP-3 (DFI: p=0.900; OS: p=0.406) serum levels.IGF-1 and IGFBP-3 serum levels before initiation of adjuvant chemotherapy are not indicative of CTCs' presence in the blood and do not correlate with clinical outcome of women with early-stage breast cancer.

Peroxisome proliferator-activated receptors (PPARα, PPARβ/δ, and PPARγ) belong to the transcription factor family, and they are highly expressed in all types of trophoblast during pregnancy. The present review discusses currently published papers that are related to the regulation of PPARs via lipid metabolism, glucose metabolism, and amino acid metabolism to affect trophoblast physiological conditions, including differentiation, maturation, secretion, fusion, proliferation, migration, and invasion. Recent pieces of evidence have proven that the dysfunctions of PPARs in trophoblast lead to several related pregnancy diseases such as recurrent miscarriage, preeclampsia, intrauterine growth restriction, and gestational diabetes mellitus. Moreover, the underlying mechanisms of PPARs in the control of these processes have been discussed as well. Finally, this review’s purposes are to provide more knowledge about the role of PPARs in normal and disturbed pregnancy with trophoblast, so as to find PPAR ligands as a potential therapeutic target in the treatment and prevention of adverse pregnancy outcomes.

Patient selection for PI3K-targeted solid cancer treatment was based on the PIK3CA/PTEN mutational status. However, it is increasingly clear that this is not a good predictor of the response of breast cancer cells to the anti-proliferative effect of PI3K inhibitors, indicating that isoform(s) other than p110α may modulate cancer cells sensitivity to PI3K inhibition. Surprisingly, we found that although no mutations in the p110δ subunit have been detected thus far in breast cancer, the expression of p110δ becomes gradually elevated during human breast cancer progression from grade I to grade III. Moreover, pharmacological inactivation of p110δ in mice abrogated the formation of tumours and the recruitment of macrophages to tumour sites and strongly affected the survival, proliferation and apoptosis of grafted tumour cells. Pharmacological inactivation of p110δ in mice with defective macrophages or in mice with normal macrophages but grafted with p110δ-lacking tumours suppressed only partly tumour growth, indicating a requisite role of p110δ in both macrophages and cancer cells in tumour progression. Adoptive transfer of δD910A/D910A macrophages into mice with defected macrophages suppressed tumour growth, eliminated the recruitment of macrophages to tumour sites and prevented metastasis compared with mice that received WT macrophages further establishing that inactivation of p110δ in macrophage prevents tumour progression. Our work provides the first in vivo evidence for a critical role of p110δ in cancer cells and macrophages during solid tumour growth and may pave the way for the use of p110δ inhibitors in breast cancer treatment.

We propose a study protocol capable of improving clinical outcomes following medically assisted reproduction (MAR) in infertile women with endometriosis and polycystic ovary syndrome (PCOS). The proposed research derives from the published evidence on the positive impact from endometrial injury (EI) and the beneficial nature of the intervention towards improved implantation rates. We primarily refer to the cluster of events and hypotheses, such as the mechanical cascade, the inflammatory response per se, the events accompanying wound healing, the immune cell recruitment and protein involvement, alterations in gene expression and the neo-angiogenesis theories, which have been previously investigated for this purpose. We are also exploring the possible problems in MAR cycles with negative outcomes in PCOS and endometriosis patients and we are proposing potential mechanisms on how this intervention might work. Our hypothesis states that the EI before the initiation of the MAR cycle can affect clinical pregnancy rates in patients with the aforementioned pathologies.

Acetylcholinesterase inhibitors are widely used for a variety of medical, agricultural and public health purposes. Consequently, exposure is highly possible during lifetime. However, their systematic use raises concerns for the potential impact on the fetus and newborn since these substances may affect angiogenesis, the neonatal and maternal intensive care, neuroimmune function and response, mammary growth/lactation via cholinergic/non-cholinergic central and peripheral neuroendocrine pathways. New methodologies, neuroscientific technologies and research studies are needed to harness existing knowledge along with the proper management, availability for new acetylcholinesterase inhibitors, with stable pharmacodynamics and clinical outcomes.

Objectives: To investigate the effect of metformin on endometrial receptivity in women with polycystic ovary syndrome (PCOS). Methods: Twenty volunteer women with polycystic ovaries and oligomenorrhea were prospectively investigated. All women were treated with exogenous estradiol and progesterone to simulate a normal menstrual cycle (28-day duration) after GnRH-induced pituitary desensitization. Ten of the women received no other medication (group A, control), while the remaining 10 received metformin (group B, metformin). Endometrial biopsy was performed in all women on day 21 of the 2 simulated cycles. Results: The expression of corticotropin – releasing hormone and urocortin in the endometrium was investigated. There was no significant difference between the 2 groups. A 3-day delay in the secretory maturation of the glandular epithelium relatively to the stroma was observed in 7 out of 10 women of group B (70%) as compared to only 1 out of 10 women of group A (10%, p = 0.02). Conclusions: It is shown for the first time that metformin administration to women with PCOS did not affect the expression of endometrial receptivity markers but delayed histological glandular maturation. It is suggested that metformin may have an impact on the function of the endometrium in PCOS.

PURPOSE: To evaluate whether ICSI offers any benefit compared with IVF in different ovarian response categories in case of non-male factor infertility. METHODS: This is a retrospective multicenter analysis using individual patient data, conducted in 15 tertiary referral hospitals in Europe (1 center in Belgium and 14 in Spain). The study included the first cycle of all patients undergoing ovarian stimulation for IVF or ICSI in a GnRH antagonist protocol. Only patients having either IVF or ICSI for non-male factor infertility were included. Patients were divided into 4 groups based on their ovarian response as follows: group A, poor responders (1–3 oocytes); group B, suboptimal responders (4–9 oocytes); group C, normal responders (10–15 oocytes); group D, high responders (> 15 oocytes). RESULTS: In total, 4891 patients were analyzed, of whom 4227 underwent ICSI and 664 IVF. There was no significant difference for the insemination method (ICSI vs. IVF) used among the different ovarian response categories: 87% vs. 13%, 87% vs. 13%, 86% vs. 14%, 84% vs. 16%, for groups A, B, C, and D, respectively, p value = 0.35. Mean fertilization rates and embryo utilization rates were comparable between IVF and ICSI in the whole cohort. Fresh and cumulative LBR did not differ significantly for IVF and ICSI in poor, suboptimal, normal, and high responders. CONCLUSION: There is no advantage of ICSI over IVF as insemination method for non-male factor infertility, irrespective of the ovarian response. The number of oocytes retrieved has no value for the selection of the insemination procedure in case of non-male infertility. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s10815-019-01563-1) contains supplementary material, which is available to authorized users.

Background Previous studies have shown that the intrauterine administration of peripheral blood mononuclear cells (PBMC) may improve pregnancy outcome of women with repeated implantation failure (RIF). We have demonstrated that, during implantation, corticotropin-releasing hormone (CRH) plays a key role in facilitating endometrial decidualization and maternal–foetal immunotolerance. In the present preliminary study, we investigated whether the intrauterine administration of autologous CRH-treated PBMC can improve clinical pregnancy rates of women with RIF. Methods Forty-five (n = 45) women with at least three failed in vitro fertilization (IVF) attempts and no previously reported clinical pregnancy were included in this crossover study. All women underwent controlled ovarian stimulation using the long GnRH agonist protocol. PBMC were isolated at day of oocyte retrieval, treated with CRH and administered in the uterine cavity at day 2, following oocyte retrieval. Blastocyst transfer was performed on day 5. Results Following the CRH-PBMC intrauterine administration, a significant increase was observed in the clinical pregnancy rate of this cohort of women with RIF (20/45 women had a clinical pregnancy; 44·44%, P

BACKGROUND The intrauterine administration of activated autologous peripheral blood monocytes (PBMC) prior to embryo transfer seems to improve reproductive outcomes in women with repeated implantation failure (RIF). We have previously shown that the intrauterine administration of PBMC treated with corticotropin-releasing hormone (CRH) prior to blastocyst transfer (day 5) improves significantly the clinical pregnancy rate of women with RIF. In the present crossover pilot study, we have investigated whether CRH-PBMC treatment could be of benefit in case of fresh early cleavage stage embryo transfer (day 3) in women with RIF. METHODS Twenty-six (n = 26) women with at least three previous failed IVF attempts and no history of clinical pregnancy in the past were recruited in this study. Ovarian stimulation was performed following either the long or the short protocol. PBMC were collected during the oocyte retrieval, were treated with CRH, and transferred in the uterine cavity 2 days later. Good quality cleavage stage embryos were transferred at day 3, following oocyte retrieval. RESULTS Following the intrauterine administration of CRH-treated autologous PBMC, 15/26 clinical pregnancies occurred (57.69%). Compared to the null result of the same women prior to recruitment, this observation was considered significant (P

Objective To investigate the effect of metformin administration on the expression of endometrial corticotrophin-releasing hormone (CRH) and urocortin (UCN) in the midluteal phase of the cycle. Design Experimental study, performed in 2010–2011. Setting University hospital. Patient(s) Eight healthy, normally cycling and parous women volunteered for the study. Intervention(s) All women were investigated in two nonconsecutive cycles (control cycle, untreated and after one cycle break; trial cycle, oral administration of metformin [850 mg × 2]). Endometrial pipelle biopsies were obtained on day LH+7. Main Outcome Measure(s) The endometrial biopsies were immunohistochemically assessed for CRH and UCN expression. Evaluation of positivity was performed by applying the immunoreactive score. Result(s) Compared with samples from control cycles, CRH and UCN were significantly reduced in endometrial samples obtained during metformin treatment. This down-regulation was significant both in the endometrial cells and in the endometrial stroma. Conclusion(s) This is the first study showing that during the midluteal phase of the cycle, metformin may decrease the production of CRH and UCN in the endometrium. Metformin interference to decidualization could happen by CRH/UCN modification.

Phytoestrogens have a controversial effect on hormone-dependent tumours. Herein, we investigated the effect of the pumpkin seed extract (PSE) on estradiol production and estrogen receptor (ER)-α/ER-β/progesterone receptor (PR) status on MCF7, Jeg3, and BeWo cells. The PSE was prepared and analyzed by mass spectrometry. MCF7, Jeg3, and BeWo cells were incubated with various concentrations of PSE. Untreated cells served as controls. Supernatants were tested for estradiol production with an ELISA method. Furthermore, the effect of the PSE on ER-α/ER-β/PR expression was assessed by immunocytochemistry. The PSE was found to contain both lignans and flavones. Estradiol production was elevated in MCF7, BeWo, and Jeg3 cells in a concentration-dependent manner. In MCF7 cells, a significant ER-α downregulation and a significant PR upregulation were observed. The above results after properly designed animal studies could highlight a potential role of pumpkin seed's lignans in breast cancer prevention and/or treatment.

Macrophages hold a key role in both regulating and executing the body's own immune response under various conditions. Hence, although endometriosis, preeclampsia and abortions are clinically different, all three are regarded to involve highly complex immunological processes. The aim of our current work was to evaluate the role of macrophages within these gynaecological disorders. Macrophages have been shown to invade endometriosis lesions and to mediate propagation of endometriotic cyst growth. However this is the first time that significant GPER up-regulation in macrophages is demonstrated. This highlights a potential alternative way through which estrogen may modulate immune response of macrophages in endometriosis. In addition, during spontaneous miscarriages the macrophage population increases significantly. This deregulation may possibly support an inflammatory scheme further triggering abortive procedures. Macrophage-mediated apoptosis of extravillous trophoblasts (EVT) has been associated with preeclampsia. Larger numbers of apoptotic EVT were detected in preeclamptic placentas compared with normal. In preeclamptic placentas, decidual macrophages were found to be Fas ligand (FasL)-positive. Our results highlight a new aspect of macrophage biology in endometriosis and pregnancy physiology and patho-physiology. Further studies with larger samples are needed to verify the current results and evaluate their clinical impact. Our data strongly indicate that macrophages hold key roles in various gynaecological disorders and might be crucial to further elucidate their patho-physiology.

In vitro growth systems of preantral follicles allow studying the effect of various endocrine, paracrine, and autocrine factors on follicular growth and oocyte maturation. CRH is a 41-amino-acid neuropeptide responsible for endocrine, autonomic, immunological, and behavioral responses of mammals to stress and has two receptors, CRH receptor type 1 (CRH-R1) and CRH-R2. Antalarmin, a CRH-R1 antagonist, has been used to elucidate the role of CRH in stress, inflammation, and reproduction. The present study describes in vitro growth of mouse preantral follicles, early embryo development, and steroidogenesis in the presence of CRH and its antagonist antalarmin. We cultured 732 follicles in control media, 1306 in CRH 10(-7) mol/liter, and 1202 in CRH 10(-7) plus antalarmin 10(-6) mol/liter. The culture medium was assayed on alternate days for 17β-estradiol, progesterone, and β-human chorionic gonadotropin. Total RNA was extracted from preantral follicles as well as early preimplantation embryos and was assessed by real-time RT-PCR for the expression of CRH-R1 and CRH-R2 mRNAs. Hormone analysis showed that the CRH group had lower levels of 17β-estradiol, progesterone, and β-human chorionic gonadotropin as the culture progressed, in comparison with the other two groups. RT-PCR demonstrated the presence of CRH-R1 and CRH-R2 in all stages of preantral follicle culture. Morula/blastocyst-stage embryos expressed only CRH-R1. In conclusion, CRH has an inhibitory effect on in vitro fertilized oocytes, resulting from cultured preantral follicles at all stages of preimplantation embryo development. Furthermore, the presence of CRH in the culture medium inhibits steroidogenesis by preantral mouse follicles cultured in vitro.

Corticotropin-releasing hormone (CRH) and its receptors are expressed in human placenta. Recently, the impaired function of this system has been associated with a number of complications of pregnancy, including pre-eclampsia. The aim of the study was to test the hypothesis that CRH participates in the pathophysiology of pre-eclampsia through the induction of macrophage-mediated apoptosis of extravillous trophoblasts (EVTs). We found that the expression of CRH was increased in the EVT of the placental bed biopsy specimens from pre-eclamptic pregnancies (1.8-fold increase; P < 0.05). In addition, significantly larger numbers of apoptotic EVT were detected in pre-eclamptic placentas compared with normal ones (P < 0.05), and only in pre-eclamptic placentas, decidual macrophages were found to be Fas ligand (FasL)-positive. In vitro studies on the effect of CRH on human macrophages suggested that CRH induced the expression of the FasL protein in human macrophages and potentiated their ability to induce the apoptosis of a Fas-expressing EVT-based hybridoma cell line in co-cultures. These findings demonstrate a possible mechanism by which the aberrant expression of CRH in pre-eclampsia may activate the FasL-positive decidual macrophages, impair the physiological turnover of EVT and eventually disturb placentation.

Class IA PI3K isoforms have divergent, nonredundant cell biological roles. In untransformed cells and tissues, p110α and p110β are ubiquitously expressed, whereas p110δ expression is highly enriched in leukocytes. High levels of p110δ expression have been documented in some solid tumor cell lines, but the functional role is unknown. This study aimed to elucidate the link between elevated expression of p110δ PI3K and cancer. We report that in breast and prostate cancer cells that contain leukocyte levels of p110δ, p110δ activity dampens the activity of the PTEN tumor suppressor. Indeed, inactivation of p110δ in these cells led to PTEN activation, suppression of Akt phosphorylation, and inhibition of cell proliferation, with inhibition of PTEN activity being able to counterbalance p110δ inactivation. Likewise, forced overexpression of p110δ in cells with low p110δ expression reduced PTEN activity, resulting in increased Akt phosphorylation. Our data indicate that the oncogenic potential of p110δ PI3K overexpression might at least partially act through PTEN inactivation, and that p110δ-selective PI3K inhibitors can have a dual antitumor mechanism, namely by directly inhibiting p110δ signaling and by a broader inhibition of class I PI3K activity through PTEN activation. These data may have important implications in the intervention of breast cancer.

AIM AND SETTING: To test the effects of crude extracts from flax (Linum usitatissimum) on progesterone and estradiol and ERα and β/PR production in choriocarcinoma cell lines Jeg 3 and BeWo. Tumor trophoblast cells (Jeg 3 and BeWo) were incubated in the presence of different concentrations of the flax crude extracts. Estradiol and progesterone production was measured. Estrogen receptor α and β as well as progesterone receptor expressions were also assessed.In Jeg 3 cells, progesterone production was downregulated by flax root and leaves extract, while in BeWo cells only flax root extract did manage to downregulate progesterone production. ERβ expression was significantly downregulated by flax root and flax leaves extract in both cell lines; on the contrary, ERα expression was increased by flax leaves extract in BeWo cells. PR expression was downregulated by flax leaves extract in Jeg 3 and by flax root extract in BeWo cells.Flax extracts derived from leaves and especially from roots can modify progesterone and possibly estradiol production, while at the same time they seem to alter ERβ expression. Further studies on animal models and adequately designed retrospective epidemiological studies are imperative to clarify this role upon progesterone.

Colony stimulating factor-1 (CSF-1) and its receptor (CSF-1R) are key regulators of macrophage biology, and their elevated expression in cancer cells has been linked to poor prognosis. CSF-1Rs are thought to function at the plasma membrane. We show here that functional CSF-1Rs are present at the nuclear envelope of various cell types, including primary macrophages, human cancer cell lines, and primary human carcinomas. In response to CSF-1, added to intact cells or isolated nuclei, nucleus-associated CSF-1R became phosphorylated and triggered the phosphorylation of Akt and p27 inside the nucleus. Extracellularly added CSF-1 was also found to colocalize with nucleus-associated CSF-1Rs. All these activities were found to depend selectively on the activity of the p110δ isoform of phosphoinositide 3-kinase (PI3K). This finding was related to the p110δ-dependent translocation of exogenous CSF-1 to the nucleus-associated CSF-1Rs, correlating with a prominent role of p110δ in activation of the Rab5 GTPase, a key regulator of the endocytic trafficking. siRNA-silencing of Rab5a phenocopied p110δ inactivation and nuclear CSF-1 signaling. Our work demonstrates for the first time the presence of functional nucleus-associated CSF-1Rs, which are activated by extracellular CSF-1 by a mechanism that involves p110δ and Rab5 activity. These findings may have important implications in cancer development.

Recent developments in reproductive medicine address oocyte morphology, sperm analysis and embryo selection. However, in a subgroup of infertile couples, it is the embryo implantation process that is disrupted. Diagnostic tools to identify patients at risk of implantation failure are limited and therapeutic options are far away from being established. In this review we focus on selected possible causes and treatments of failed implantation. Reproductive surgery allows a proper first step diagnosis and therapy of recurrent implantation failure (RIF). Possible anatomical malformations and associated diseases with treatment options are mentioned. Diagnostic procedures in patients often focus on defining gene polymorphisms (like hereditary thrombophilia and p53) and determinants of endometrial receptivity including endometrial gene expression profiles. Although significant differences in gene expression have been identified, the study populations are quite small, some of the data conflicting and clinical significance has yet to be proven. Implantation requires a close interaction between the fetal trophoblast and the maternal endometrium with natural killer cells (NK cells) playing a main part at the feto-maternal interface during early pregnancy. Therefore this review also focuses on NK cell receptor expression and new immunomodulatory treatment options like G-CSF in RIF patients.

Objective To compare the effect of two different insulin sensitizers, metformin and pioglitazone, on endothelial function in women with polycystic ovary syndrome (PCOS). Design Prospective randomized study. Setting University Hospital endocrinology outpatient clinic. Patient(s) Young women with PCOS (aged 23.3 ± 4.9 years). Intervention(s) Patients were assigned randomly to no treatment (n = 14), metformin 850 mg two times per day (n = 15), and pioglitazone 30 mg daily (n = 14) for 6 months. Healthy age- and body mass index–matched women served as controls (n = 14). Main Outcome Measure(s) Brachial artery flow-mediated dilation was studied at baseline and 6 months. Result(s) Women with PCOS had higher insulin resistance and hyperandrogenism indices and lower flow-mediated dilation compared with controls. The three groups of women with PCOS did not differ at baseline. No differences were observed at follow-up in women who received no treatment. Metformin and pioglitazone improved flow-mediated dilation to a similar extent, restoring it to normal values at 6 months. Both insulin sensitizers induced favorable changes in insulin resistance and hyperandrogenism indices in women with PCOS. Independent predictors of flow-mediated dilation improvement at 6 months were treatment with insulin sensitizers and reduction in insulin resistance. Conclusion(s) In young women with PCOS, treatment with metformin or pioglitazone for 6 months induces a similar beneficial effect on endothelial function; this may be partially attributed to an improvement in insulin resistance. Further research is needed to investigate whether treatment with insulin sensitizers in women with PCOS also reduces cardiovascular risk.

Citation Toth B, Haufe T, Scholz C, Kuhn C, Friese K, Karamouti M, Makrigiannakis A, Jeschke U. Placental Interleukin-15 expression in recurrent miscarriage. Am J Reprod Immunol 2010; 64: 402–410 Problem Cytokines play a fundamental role at the feto–maternal interface. A dysregulation of the cytokine profile could be linked with fetal rejection. This study investigated differences in the expression of the Th1 cytokine interleukin-15 (IL-15) of normal and disturbed pregnancy. Method of study Analysis of IL-15 expression by immunohistochemistry and real-time RT-PCR in placental tissue from normal pregnancies, spontaneous miscarriage and recurrent miscarriage (RM) was performed. Results Expression of IL-15 was upregulated on mRNA and protein level in the placenta of miscarriage patients, especially in patients with RM. Immunohistochemistry and immunofluorescence double staining revealed extravillous trophoblast cells (EVT) as IL-15-expressing cells. Staining results were confirmed by real-time PCR (TaqMan). Conclusion Increased expression of IL-15 in the decidua of patients with RM may be linked to disturbed implantation and vascularization with consecutive placental and fetal rejection.

Life exists by establishing a balanced equilibrium, called homeostasis, constantly challenged by adverse stimuli, called stressors. In response to these stimuli, a complex neurohormonal reaction exerted by the activation of the so-called stress system is initiated. The latter is activated in a coordinated fashion, leading to behavioral and peripheral changes that improve the ability of the organism to adjust homeostasis and increase its chance for survival. The stress system suppressive effects on female reproduction involve suppression of the hypothalamic-pituitary-ovarian axis at the hypothalamic, pituitary, ovarian, and uterine levels. Experimental and human data suggest that adverse prenatal stimuli, of either maternal or fetal origin, acting in the developing embryo in utero, can lead to the development of short- and long-term health disorders. These include preterm birth of the offspring, low birth weight, and the development of adult diseases ranging from the metabolic syndrome to several neurodevelopmental disorders.

Although recurrent miscarriage (RM) affects only 1-3% of couples, it has a major influence on the wellbeing and psychosocial status of patients. Therefore, research into improved diagnosis and development of new treatment strategies is essential. In this review, we summarize current concepts on diagnosis and treatment in RM, drawing upon research reports and international guidelines to provide insights into the pathophysiology of pregnancy disrupted by repeated miscarriage. Anatomical malformations, infectious diseases, endocrine disorders, autoimmune defects as well as acquired and inherited thrombophilia are established risk factors in RM. In addition, our recent findings indicate an impact on miscarriage incidence of glycoproteins such as glycodelin, and nuclear hormone receptors such as the peroxisome proliferator-activated receptors (PPARs). Significantly reduced glycodelin expression is associated with miscarriage, whereas up-regulation of PPARs appears to compensate for either the activated immune response or the disturbed cytotrophoblast differentiation in RM patients. There is also evidence that circulating placental microparticles are increased in a subgroup of RM patients, indicating an acquired procoagulant state even outside pregnancy. Treatment strategies like aspirin and low molecular weight heparin (LMWH) are standard medications in RM, although only a few placebo-controlled trials have proven their benefit in respect to live birth rate. There is emerging evidence that new treatment options, including drugs like TNFalpha inhibitors and granulocyte colony-stimulating factor (G-CSF) might be beneficial in some cases of RM. However, larger clinical trials must be completed to further prove or disprove benefits of these drugs in the treatment of RM patients.

Glycoproteins expressed at the fetal-maternal interface have been shown to exert immunomodulating effects. Glycodelin, hCG and transferrin have been used in in vitro experiments as ligands to block E-selectin-mediated cell adhesion. We found that glycodelin is a strong inhibitor of the E-selectin-mediated cell adhesion with a 10(3)-fold increase in potency compared to the monovalent tetrasaccharide sialyl Lewis X. HCG with distinct carbohydrate expression is also an effective selectin antagonist, whereas the potency of transferrin is low. This could indicate a possible role of glycodelin, hCG and transferrin in preventing leukocyte adhesion to the fetal trophoblast. In decidual tissue of abortion patients, glycodelin expression was significantly reduced compared to normal gestation. These results were confirmed by in situ hybridization. Moreover, glycodelin expression in endometrial cells in vitro could be stimulated by addition of hCG. Because hCG is down-regulated in women with abortion, we speculate that hCG could be one of the factors regulating glycodelin expression. Galectins are structurally related proteins with the ability to bind beta-galactosides through a conserved carbohydrate recognition domain. Galectin-1 (gal-1) expression in the syncytiotrophoblast is down-regulated in early pregnancy loss. Gal-1 recognizes the Thomsen-Friedenreich disaccharide (Galbeta1-3GalNAc-) on the syncytiotrophoblast and extravillous trophoblast. Gal-1 also inhibited trophoblast cell proliferation but did not induce apoptosis in BeWo cells. Ligation of Gal-1 on trophoblast cells may have regulatory effects on trophoblast cell differentiation. Decreased expression of Gal-1 may partly explain disturbed trophoblast differentiation during early placentation leading to early pregnancy loss.

Objective Reestablishing immune tolerance and long-term suppression of disease represent major therapeutic goals in rheumatoid arthritis (RA). Dendritic cells (DCs) likely play a central role in such regulation via the expansion and/or induction of Treg cells. The present study was undertaken to explore the contribution of DCs to the development of Treg cells in a human autoimmune disease setting. Methods DC subsets were characterized by flow cytometry in the peripheral blood and synovial fluid of patients with RA. Proliferation of and cytokine release by naive CD4+CD25− T cells were measured in cocultures of these cells with DCs from patients with RA and healthy controls. The suppressive capacity of DC-polarized T cells was explored in vitro by a standard suppression assay. Results Only very low numbers of both plasmacytoid DCs (CD303+) and myeloid DCs (CD1c+) were present in the peripheral blood of patients with active RA. In contrast, patients with therapy-induced remission of RA exhibited higher numbers of circulating plasmacytoid DCs. Mature plasmacytoid DCs from RA patients with low disease activity, but not those from healthy controls, expressed high levels of indoleamine 2,3-dioxygenase and promoted the differentiation of allogeneic naive CD4+CD25− T cells into interleukin-10–secreting Treg cells, or Tr1 cells, that showed poor proliferation in vitro. Importantly, these plasmacytoid DC–primed Treg cells potently suppressed the proliferation of autologous naive CD4+ T cells, in a dose-dependent manner. Conclusion These results demonstrate, for the first time, that human plasmacytoid DCs may be educated within the rheumatoid microenvironment to acquire a tolerogenic phenotype. Modulation of the immune response by plasmacytoid DCs might provide novel immune-based therapies in autoimmunity and transplantation.

CRH and Urocortins 1, 2 and 3 comprise the, so far identified, members of the CRH family of peptides in humans. Their actions are mediated through two distinct receptors, CRHR1 and CRHR2, encoded by different genes. CRH-like peptides and their receptors have been identified in reproductive tissues, such as the ovary, uterus as well as fetal and placental membranes. The participation of the "CRH family" of peptides and receptors in the physiology of these organs is currently under intense investigation. During the estrus cycle, endometrial CRH acts as a fine tuner of stromal cells decidualization. CRH is produced by embryonic trophoblast and maternal decidual cells and plays important roles in implantation. CRH also participates in the control of trophoblast invasion. Furthermore, placental CRH and Urocortin are involved in the mechanisms controlling maintenance of pregnancy and the onset of labor. The level of participation of urocortins 2 and 3 in these phenomena is currently under investigation. This review will focus on existing data on the expression and regulation of the CRH family of peptides and their receptors in the female reproductive system, as well as in their potential biologic role(s) in human reproductive functions. Curr Med Chem

Leptin and insulin are secreted into the maternal and to a lesser extent into the fetal bloodstream where they act as placental signals and nourish the fetus, making them possible candidates for the endocrine control of the placenta. We investigated differences in leptin (LR) and insulin receptors (IR) expression in normal and disturbed first trimester human pregnancy at protein level by immunohistochemistry and at mRNA level by real-time reverse transcriptase-polymerase chain reaction (TaqMan). Highest expression of LR and IR was present in villous (VT) and extravillous trophoblasts (EVT). In hydatidiform mole trophoblasts, significantly higher LR and IR expression was observed when compared with normal pregnancy. In addition, LR and IR were also expressed in glandular epithelial cells of the decidua, again to the highest extent in hydatidiform mole when compared with normal pregnancy. With regard to abortive placentas, significant differences were also present when compared with normal first trimester placenta in the expression of LR and IR in VT, EVT and in glandular epithelial cells of the decidua. Results at protein expression of LR and IR were confirmed at mRNA level. The majority of IR and LR are expressed on struc- tures that are currently assumed to drive placental growth. LR and IR are strongly up-regulated in placentas of hydatidiform mole and abor- tion. Our findings may suggest IR and LR as possible new candidates for the endocrine control of human pregnancy.

Pregnancy represents a major challenge to the maternal immune system since it has to tolerate the semi-allograft fetus. Indeed, the success of pregnancy requires an appropriate immunological interaction between the mother and the fetus. Furthermore, evidence suggests that some pregnancy complications, such as spontaneous abortions, preeclampsia and intrauterine growth restriction, are associated with abnormal maternal-fetal immune responses. Corticotropin-releasing hormone (CRH), a 41-amino acid peptide originally found in the hypothalamus, appears to have a fundamental role in the mechanisms responsible for the implantation and maintenance of human pregnancy. Reproductive CRH is a form of tissue CRH (CRH found in peripheral tissues), analogous to the immune CRH detected in peripheral inflammatory sites. Reproductive CRH has been identified in the endometrial glands, the decidualized endometrial stroma and the placental trophoblast, synctiotrophoblast and decidua. Reproductive CRH participates in various reproductive functions with an inflammatory component, where it serves as an autocrine/paracrine modulator. The immunological mechanisms contributing to the establishment and maintenance of pregnancy are not fully understood. The present article focuses on the potential roles of CRH on the physiology and pathophysiology of pregnancy and highlights its participation in implantation, early fetal immunotolerance and parturition.

Objectives: To investigate if skewed X-chromosome inactivation (XCI) is associated with unexplained recurrent miscarriage (RM) in Greek women. Methods: This was a prospective case-control study. A methylation-sensitive assay was used to investigate the X-inactivation pattern of women with unexplained RM and controls. Results: Fifty-six of the 74 patients (75.7%) and 55 of 80 controls (68.8%) were informative. Among the informative cases, 6/56 (10.7%) women showed extreme XCI (>90%) and among the informative controls, 2/55 (3.6%) showed extreme XCI. Conclusions: In the present study, women with unexplained RM showed a statistically nonsignificant increase in skewed XCI prevalence (10.7%) compared with control women (3.6%; p = 0.271).

Lewis antigens belong to the blood group of antigens and mediate cellular adhesion through interaction with selectins. Invasive trophoblasts use an array of adhesion molecules to facilitate cell-cell and cell-extracellular matrix interactions. Here, we examined immunohistochemically the expression of Sialyl Lewis a (sLe(a)), Sialyl Lewis x (sLe(x)) and Lewis y (Le(y)) in term placentas obtained from cases of normal, intrauterine growth retardation (IUGR), preeclamptic (PE) and hemolysis, elevated liver enzymes and low platelets syndrome (HELLP) pregnancies. We report the expression of sLe(x) in third trimester extravillous trophoblasts (EVT). sLe(x) was significantly decreased in IUGR and moderately decreased in PE compared to normal placentas. sLe(x) was additionally found in syncytiotrophoblast, without however any significant differences in staining intensity between normal and pathological cases. sLe(a) was restricted to amnion epithelium. Finally, Le(y) was expressed in cytotrophoblasts and villous endothelial cells. Le(y) expression was significantly upregulated in IUGR and HELLP, whereas there was a trend toward increase in PE compared to normal placentas. The present study suggests that downregulation of sLe(x) in EVT might be associated with IUGR and PE. Furthermore, Le(y), which was recently described as a potent angiogenic factor, is upregulated in placental villi in conditions associated with placental malperfusion.

The formation of the corpus luteum (CL) is critical for the establishment of a successful pregnancy. After ovulation, the CL develops from the remnants of the ovulated ovarian follicle. This process, which involves varying cell-matrix interactions, is poorly characterized. To understand the role and potential regulation of cell-matrix interactions in the formation of the CL, we investigated the expression and activity of the matrix protein fibronectin (FN) and several of its integrin receptors on luteinized granulosa cells (GCs). In situ, FN and several FN-binding integrins were detected around luteinizing GCs during the early luteal phase, although expression declined in the late luteal phase. In vitro, GCs released FN, and stimulation of these cells with human chorionic gonadotropin increased the surface expression of FN, alpha(5)beta(1), and alpha(v)beta(3). Up-regulation of these proteins on GCs was reproduced by stimulation with vascular endothelial growth factor (VEGF) and was inhibited by anti-VEGF antibody. Lastly, expression of alpha(5)beta(1) and alpha(v)beta(3) mediated adhesion to FN, facilitated migration, and prevented apoptosis. These data suggest that in vivo luteogenic hormones, in part through a VEGF-dependent mechanism, stimulate selected integrin-matrix adhesive interactions that promote the motility and survival of GCs and thus contribute to the formation and preservation of the CL.

To investigate whether corticotropin-releasing hormone (CRH) and corticotropin (ACTH) plasma concentrations in women diagnosed with preterm labor are of potential clinical value in the assessment of the risk of preterm birth.Plasma samples of 79 women diagnosed with preterm labor were used in this study. Samples were divided into three groups based on the week of gestation (24th-28th, 29th-32nd, 33rd-37th). CRH and ACTH values were determined by ELISA.Mean maternal peripheral plasma values of CRH and ACTH were significantly higher (p0.001) in women who were initially diagnosed with preterm labor and finally delivered a preterm birth, compared to women with the same diagnosis but with term birth.CRH and ACTH serum levels in women diagnosed with preterm labor could be used as predictors for the timing of parturition.

The fundamental process of implantation involves a series of steps leading to effective cross-talk between invasive trophoblast cells and the maternal endometrium. The molecular interactions at the embryo-maternal interface during the time of blastocyst adhesion and subsequent invasion are not fully understood. Embryonic trophoblast and maternal decidual cells produce corticotropin-releasing hormone (CRH) and express Fas ligand (FasL), a proapoptotic cytokine. Fas and its ligand are pivotal in the regulation of immune tolerance. Trophoblast and decidual CRH play crucial roles in implantation, as well as in the anti-rejection process that protects the fetus from the maternal immune system, primarily by killing activated T cells through Fas-FasL interaction. The potential use of CRH antagonists is presently under intense investigation. CRH antagonists have been used experimentally to elucidate the role of CRH in blastocyst implantation and invasion, early fetal immunotolerance, and premature labor.