Co-cultivation of human granulosa cells with ovarian cancer cells leads to a significant increase in progesterone production.

PURPOSE: In humans, granulosa cells (GCs) are part of the follicle and nourish the growing oocyte. GCs produce estrogen and, after ovulation, progesterone. They are embedded in a multicellular tissue structure of the ovary, which consists of a variety of different cell types that are essential for the physiological function of the ovary. However, the extent to which individual ovarian cell types contribute to overall functionality has not yet been fully elucidated. In this study, we aim to investigate the effects of co-culturing human granulosa cells with ovarian cancer cells on their progesterone and estrogen production in an in vitro model. METHODS: After seeding, the cells were stimulated with 200 µM forskolin in DMEM for 72 h and the medium of the different cell culture experiments was collected. Subsequently, progesterone and oestradiol concentrations were determined using an Elisa assay. RESULTS: Morphologically, it was striking that the cells self-organize and form spatially separated areas. Compared to culturing granulosa cells alone, co-culturing human granulosa cells together with the ovarian cancer cell line OvCar-3 resulted in a significant increase in progesterone production (20.3 ng/ml versus 50.2 ng/ml; p < 0.01). CONCLUSIONS: Using a simple in vitro model, we highlight the importance of cellular crosstalk between different ovarian cells in a complex cellular network and that it strongly influences granulosa cell hormone production. This could have potential implications for the procedure of transplanting endocrine tissues after cryopreservation, as it highlights the importance of survival of all cells for the functionality of the transplanted tissue.

Different Impacts of Cryopreservation in Endothelial and Epithelial Ovarian Cells.

The aim of our laboratory-based study was to investigate the extent of delayed-onset cell death after cryopreservation in endothelial and epithelial cell lines of ovarian origin. We found differences in percentages of vital cells directly after warming and after cultivation for 48 to 72 h. A granulosa cell line of endothelial origin (KGN) and an epithelial cell line (OvCar-3) were used. In both DMSO-containing and DMSO-free protocols, significant differences in vitality rates between the different cell lines when using open and closed vitrification could be shown (DMSO-containing: KGN open vs. OvCar open, p = 0.001; KGN closed vs. OvCar closed, p = 0.001; DMSO-free: KGN open vs. OvCar open, p = 0.001; KGN closed vs. OvCar closed, p = 0.031). Furthermore, there was a marked difference in the percentage of vital cells immediately after warming and after cultivation for 48 to 72 h; whereas the KGN cell line showed a loss of cell viability of 41% using a DMSO-containing protocol, the OvCar-3 cell loss was only 11% after cultivation. Using a DMSO-free protocol, the percentages of late-onset cell death were 77% and 48% for KGN and OvCar-3 cells, respectively. Our data support the hypothesis that cryopreservation-induced damage is cell type and cryoprotective agent dependent.

Basal and dynamic relationships between serum anti-Müllerian hormone and gonadotropins in patients with functional hypothalamic amenorrhea, with or without polycystic ovarian morphology.

BACKGROUND: To evaluate in women with functional hypothalamic amenorrhea (FHA), whether there is a difference between patients with and without polycystic ovarian morphology (PCOM) concerning the response to a gonadotropin releasing hormone (GnRH) stimulation test and to pulsatile GnRH treatment. METHODS: In a retrospective observational study, 64 women with FHA who underwent a GnRH stimulation test and 32 age-matched controls without PCOM were included. Pulsatile GnRH treatment was provided to 31 FHA patients and three-month follow-up data were available for 19 of these. RESULTS: Serum levels of gonadotropins and estradiol were lower in FHA women than in controls (p < 0.05). FHA patients revealed PCOM in 27/64 cases (42.2%). FHA patients without PCOM revealed lower anti-Müllerian hormone (AMH) levels than controls (median 2.03 ng/mL, IQR 1.40-2.50, versus 3.08 ng/mL, IQR 2.24-4.10, respectively, p < 0.001). Comparing FHA patients with and without PCOM, the latter revealed lower AMH levels, a lower median LH increase after the GnRH stimulation test (240.0%, IQR 186.4-370.0, versus 604.9%, IQR 360.0-1122.0; p < 0.001) as well as, contrary to patients with PCOM, a significant increase in AMH after three months of successful pulsatile GnRH treatment (median 1.69 ng/mL at baseline versus 2.02 ng/mL after three months of treatment; p = 0.002). CONCLUSIONS: In women with FHA without PCOM, the phenomenon of low AMH levels seems to be based on relative gonadotropin deficiency rather than diminished ovarian reserve. AMH tended to rise after three months of pulsatile GnRH treatment. The differences found between patients with and without PCOM suggest the former the existence of some PCOS-specific systemic and/or intra-ovarian abnormalities.

Comparison of a practice-based versus theory-based training program for conducting vacuum-assisted deliveries: a randomized-controlled trial.

PURPOSE: Vacuum-assisted deliveries (VAD) are complex procedures that require training and experience to be performed proficiently. We aimed to evaluate if a more resource intensive practice-based training program for conducting VAD is more efficient compared to a purely theory-based training program, with respect to immediate training effects and persistence of skills 4-8 weeks after the initial training. METHODS: In this randomized-controlled study conducted in maternity staff, participants performed a simulated low-cavity non-rotational vacuum delivery before (baseline test) and immediately after the training (first post-training test) as well as 4-8 weeks thereafter (second post-training test). The study's primary endpoint was to compare training effectiveness between the two study groups using a validated objective structured assessment of technical skills (OSATS) rating scale. RESULTS: Sixty-two participants were randomized to either the theory-based group (n = 31) or the practice-based group (n = 31). Total global and specific OSATS scores, as well as distance of cup application to the flexion point improved significantly from baseline test to the first post-training test in both groups (pall < 0.007). Skill deterioration after 4-8 weeks was only found in the theory-based group, whereas skills remained stable in the practice-based group. CONCLUSION: A practice-based training program for conducting VAD results in comparable immediate improvement of skills compared to a theory-based training program, but the retention of skills 4-8 weeks after training is superior in a practice-based program. Future studies need to evaluate, whether VAD simulation training improves maternal and neonatal outcome after VAD.

Markers of vitality in ovaries of transmen after long-term androgen treatment: a prospective cohort study.

BACKGROUND: Gender-affirming hormone therapy has been hypothesized to reduce the patient's reproductive potential in transmen, although the exact long-term effects on future fertility are unknown. METHODS: In this prospective cohort study we aimed to evaluate ovaries of 20 transmen by using hormone serum levels, histomorphological analysis and fluorescence activated cells sorting (FACS) analysis - in order to assess the amount of vital cells. RESULTS: The median total number of follicles per field of view was 39 (IQR 12-122). Of all follicles (n = 1661), the vast majority was primordial (n = 1505, 90.6%), followed by primary (n = 76, 4.6%), abnormal (n = 63, 3.8%) and secondary follicles (n = 17, 1.0%). FACS analysis was available for 13 samples (65.0%) and the median frequency of vital cells was 87.5% (IQR, 77.7-95.4%). Both a higher age (p = 0.032) and a lower BMI (p = 0.003) were significantly associated with a higher frequency of vital cells. CONCLUSION: The majority of ovarian cells after long-term androgen treatment were vital in FACS analysis and histomorphological evaluation revealed a normal cortical follicle distribution. These results are currently exploratory, but might be promising for issues on fertility preservation. TRIAL REGISTRATION: The study was approved by the ethics committee of the Medical University of Vienna (EK 2240/2016) and was retrospectively registered in the Current Controlled Trials Register (registration number NCT03649087 , date of registration: 28.08.2018).

Assessment of Tubal Patency: A Prospective Comparison of Diagnostic Hysteroscopy and Laparoscopic Chromopertubation.

STUDY OBJECTIVE: To evaluate whether the presence of a visualizable "flow" effect in the fallopian tube ostia in hysteroscopy was predictive of tubal patency. DESIGN: A prospective cohort study. SETTING: In a prospective study, infertile women who underwent surgery because of infertility between March and November 2018 were included. The main outcome parameter was fallopian tube patency assessed by laparoscopic chromopertubation. The predictive parameter tested was the presence of hysteroscopic tube flow. PATIENTS: Seventy-two infertile women. INTERVENTIONS: Combined hysteroscopy and laparoscopy with chromopertubation. RESULTS: One-hundred forty-four fallopian tubes were evaluated, with 88 (61.1%) patent tubes at laparoscopic chromopertubation. A positive hysteroscopic flow effect was recorded for 94 (65.3%) ostia and was accurate in predicting patency (p < .001), with a sensitivity of 85.3% (95% confidence interval [CI], 76.1-91.9) and a specificity of 66.1% (95% CI, 52.2-78.2). A multivariate binary regression model revealed that the presence of a hydrosalpinx (odds ratio = 8.216; 95% CI, 1.062-63.574; p = .044) and peritubal adhesions (odds ratio = 3.439; 95% CI, 1.142-10.353; p = .028) were associated with a false-normal flow result. A hazy hysteroscopic picture was found in 15 of 21 (71.4%) and 5 of 51 (9.8%) cases with and without bilateral tubal occlusion, respectively (p < .001, sensitivity = 71.4% [95% CI, 47.8-88.7], specificity = 90.2% [95% CI, 78.6-96.7]). CONCLUSIONS: The presence of hysteroscopic tubal flow was a reliable indicator of tubal patency. A hydrosalpinx or peritubal adhesions increase the risk for a false-normal result. A hazy hysteroscopic picture suggests bilateral tubal occlusion. Using the hysteroscopic flow effect, one can provide additional information for the patient.

The impact of repetitive oocyte retrieval on the ovarian reserve: a retrospective cohort study.

PURPOSE: To investigate a possible influence of repetitive micro-traumata on the ovaries in the course of oocyte retrieval during IVF/ICSI treatment on serum anti-Müllerian hormone (AMH) levels. METHODS: The study included retrospectively collected data from women who underwent three or more consecutive IVF/ICSI treatments between 2007 and 2017. The primary endpoint of the study was to evaluate changes in serum AMH levels on cycle days 1-3 during the course of repetitive IVF/ICSI treatments. RESULTS: A total of 125 patients were included in this study. Median AMH levels before the first, second and third IVF/ICSI cycles were 3.8 ng/mL (IQR 1.8-7.1), 3.3 ng/mL (IQR 1.8-6.1) and 3.0 ng/mL (IQR 1.6-5.3), respectively (p = n.s.). In patients who underwent IVF/ICSI due to polycystic ovary syndrome (PCOS), we found a significant decrease in AMH serum levels between the first [AMH 9.7 ng/mL (IQR 7.4-14.4)] and the third [AMH 5.3 ng/mL (IQR 3.3-10.4)] IVF/ICSI cycles (p = 0.026). When performing a generalized linear model, we found PCOS to be an independent predictor for serum AMH decrease during the course of three oocyte retrievals (p < 0.001). CONCLUSIONS: When comparing the indications for IVF/ICSI, we observed a significant decrease in AMH serum levels after repetitive oocyte retrievals only in women with PCOS, while the decrease in AMH was not significant in patients with tubal factor, endometriosis, male factor and unexplained infertility. This finding leads us to hypothesize that repetitive micro-traumata on the ovarian cortex might diminish/normalize functional ovarian reserve in women with PCOS. Further prospective studies are highly warranted to allow firm conclusions.

Impact of vitrification on granulosa cell survival and gene expression.

INTRODUCTION: Cryopreservation of ovarian tissue is an essential step in Ovarian Tissue Banking. In order to prevent the formation of ice crystals, typically the tissue is slowly frozen using a cryoprotectant. As an alternative the method of ultra-fast freezing by vitrification becomes more attention for freezing ovarian tissue because it has successfully been used for oocytes, embryos and sperm. However the impact of vitrification on granulosa cells, which are an essential part of ovarian tissue is uncertain. AIM: In this study, we have therefore analysed the influence of vitrification on the survival rates of granulosa cells, the impact of DMSO or ethylenglycol containing vitrification protocols and investigated to what extent the gene expression of apoptosis- and temperature-sensitive genes changes. MATERIAL AND METHODS: We used the human granulosa cell line KGN as a model for human granulosa cells and determined the survival rate and cell cycle stages by FACS analyses. The change in gene expression was determined by quantitative PCR analyses. RESULTS: Our results show that vitrification is possible in granulosa cells but it reduces cell viability and leads to fluctuations in the cell cycle. The DMSO containing protocol results in a lower amount of dead cells than the ethylenglycol containing protocol. Gene expression analysis reveals that TNF-alpha expression is strongly increased after vitrification, while other apoptosis or temperature-related genes seem to stay unaffected. CONCLUSION: We conclude that vitrification influences the viability of human granulosa cells. Furthermore, our results suggest that this could be mediated by a change in TNF-alpha gene expression.

Risk and protective factors for obstetric anal sphincter injuries: A retrospective nationwide study.

BACKGROUND: In view of the reported increase in obstetric anal sphincter injuries, the objective of this study was to evaluate the incidence of such injuries over time and the associated risk and protective factors. METHODS: This was a retrospective cohort study from a national database of 168 137 primiparous women with term, singleton, cephalic, vaginal delivery between 2008 and 2014. The main outcome measure was obstetric anal sphincter injury. A multivariate regression model was used to identify risk and protective factors. RESULTS: Age >19 years, birthweight >4000 g, and operative vaginal delivery were independent risk factors for obstetric anal sphincter injuries. Mediolateral episiotomy increased the risk for obstetric anal sphincter injuries in spontaneous vaginal birth (number needed to harm 333), whereas it was protective in vacuum delivery (number needed to treat 50). From 2008 to 2014, there was an increase in the rate of obstetric anal sphincter injuries (2.1% vs 3.1%, P < .01), vacuum deliveries (12.1% vs 12.8%, P < .01), and cesarean delivery after labor (17.1% vs 19.4%, P < .01), while forceps deliveries (0.4% vs 0.1%, P < .01) and episiotomy rate decreased (35.9% vs 26.4%, P < .01). CONCLUSIONS: Episiotomy may be a risk or protective factor depending on the type of episiotomy and the clinical setting in which it is used. Our study supports a restrictive use of mediolateral episiotomy in spontaneous vaginal deliveries. In vacuum deliveries mediolateral episiotomy may help prevent obstetric anal sphincter injuries.

The effect of slow release insemination on pregnancy rates: report of two randomized controlled pilot studies and meta-analysis.

PURPOSE: A modified application technique of intrauterine insemination (IUI) is slow release insemination (SRI), first described by Muharib et al. (Hum Reprod 7(2):227-229, 1992), who postulated higher pregnancy rates with a slow release of spermatozoa for 3 h. METHODS: To investigate this approach, two randomized controlled, cross-over pilot studies were performed from 2004 to 2006 in Israel and Germany to compare SRI with the standard bolus IUI. We aimed to present the results and perform a meta-analysis on available data for SRI. Univariate comparisons of pregnancy rates were performed using one-tailed z tests for method superiority. For meta-analysis, a fixed-effect Mantel-Haentzel weighted average of relative risk was performed. RESULTS: Fifty treatment cycles (IUI: n = 25, SRI: n = 25) were performed in Germany, achieving four pregnancies (IUI: 4%, SRI: 12%, p > 0.05). Thirty-nine treatment cycles (IUI: n = 19, SRI: n = 20) were performed in Israel achieving six pregnancies (IUI: 10.5%, SRI: 20%; p > 0.05). Meta-analysis of all eligible studies for SRI (n = 3) revealed a combined relative risk for pregnancy after SRI of 2.64 (95% CI 1.04-6.74), p = 0.02). CONCLUSIONS: In conclusion, these results lend support to the hypothesis that the pregnancy rate might be improved by SRI compared to the standard bolus technique.

Risk Factors for De Novo Malignancies in Women After Kidney Transplantation: A Multicenter Transversal Study.

OBJECTIVE: Transplantation results in a 5-time elevated risk for a variety of malignancies (Kaposi sarcoma, skin, liver, lung, gastrointestinal cancer). A patient's risk for malignancies could be of particular interest for the follow-up programs of patients and risk adaption after kidney transplantation. The aim of this study was to identify independent risk factors for de novo malignancies in women after renal transplantation. METHODS AND MATERIALS: This is a multicenter transversal study, conducted at the Medical University of Vienna and Hospital Rudolfstiftung, Vienna, Austria. We included female kidney graft recipients who were transplanted between 1980 and 2012 and followed-up at our institutions (N = 280). Clinical data of patients were extracted from hospital charts and electronic patient files. Patients were interviewed using a standardized questionnaire regarding their medical history, history of transplantation, and malignant diseases. Detailed information about present and past immunosuppressive regimens, rejection episodes and therapies, renal graft function, and information about primary disease was obtained. Diagnostic work-up and/or surgical exploration was performed if any presence of malignancy was suspected during routine follow-up. Histological specimens were obtained from all patients. MAIN OUTCOME MEASURES: the presence of de novo malignancy after kidney transplantation. RESULTS: Two hundred sixty-two women were included for statistical analysis. Median (interquartile range) follow-up period after transplantation was 101.1 (27.3-190.7) months. Thirty-two patients (12.2%) developed a malignancy: dermatologic malignancies (5.7%), breast cancer (3.4%), cervical cancer (0.8%), lung cancer (0.4%), gastrointestinal malignancies (1.5%), vulvar cancer (0.4%), and unclassified malignancies (1.9%). Median (interquartile range) time to malignancy after transplantation was 185.9 (92.0-257.6) months. Cumulative cancer rates were 4.9% (1 year), 14.4% (3 years), 16.4% (5 years), and 21.8% (10 years). Second transplantations were identified as independent risk factor for development of malignancy after transplantation. CONCLUSIONS: Long-term risk of developing a malignancy after kidney transplantation is high, which might justify a follow-up of more than 10 years.

Pelvic floor symptoms in female cyclists and possible remedies: a narrative review.

INTRODUCTION AND HYPOTHESIS: Cycling has become a popular athletic activity worldwide and can lead to genital and pelvic floor dysfunction. This review summarizes the current body of evidence about the epidemiology of genital and pelvic floor symptoms in female cyclists, the therapy, and preventive interventions. METHODS: Two electronic meta-databases, OvidSP™ and Deutsches Institut für Medizinische Dokumentation und Information (DIMDI), comprising 40 individual databases, were searched for studies that described genital and pelvic floor symptoms in association with cycling and studies that tested possible therapies and prophylactic measures. For the literature search we explored the search terms "female", "bicycling", "pelvic floor", "lower urinary tract symptoms", and "vulvar diseases". RESULTS: The search retrieved 1,219 articles, leaving 763 articles after removal of duplicates, and finally 12 articles eligible for review. We assessed 10 observational and 2 experimental studies. Genital and pelvic floor symptoms related to bicycling were pain, tenderness, neuropathy, urological dysfunction and skin lesions. Broader and conventionally shaped saddles were associated with fewer symptoms compared with cut-out saddle designs. CONCLUSION: The quality of existing studies is generally low, but there is evidence that female cyclists suffer from similar problems to male cyclists, ranging from minor skin lesions to severe sequelae such as pain and neurological deficiencies.

The usefulness of CYFRA 21-1 to diagnose and predict preeclampsia: a nested case-control study.

BACKGROUND: The ability to identify patients at risk for developing preeclampsia is important for preventing morbidity and mortality in both the mother and child. Although CYFRA 21-1 (a fragment of Cytokeratin 19) is considered a promising biomarker for diagnosing preeclampsia, little is known regarding the levels of CYFRA 21-1 during pregnancy. Here, we measured serum CYFRA 21-1 levels in women with an uneventful pregnancy and in women whose pregnancy was complicated by preeclampsia. Furthermore we evaluated whether maternal CYFRA 21-1 levels can be used to predict and/or diagnose preeclampsia. METHODS: Longitudinal, sequential blood samples were collected prospectively at seven predetermined visits during pregnancy. Maternal CYFRA 21-1 levels were measured in 50 women with an uneventful pregnancy (control group) and in 10 asymptomatic women whose pregnancy was later complicated by preeclampsia (PE_long group). In addition, CYFRA 21-1 levels were measured from a single sample collected from a separate group of 50 pregnant women with symptomatic preeclampsia (PE_state group). RESULTS: The CYFRA 21-1 levels were significantly higher in the PE_state group compared to the control group (p < 0.001). In the PE_long group, CYFRA 21-1 levels were lower from gestational week 11 through 17, but were higher than the control group from gestational weeks 18 through 36. Out of the ROC curves that were calculated to investigate the predictive and diagnostic properties of CYFRA 21-1 levels for preeclampsia, the ROC curve for diagnosing preeclampsia in gestational week 28-32 showed the largest AUC of 0.92, at a cut-off point of 3.1 ng/ml, leading to sensitivity of 92 % and specificity of 80 %. CONCLUSIONS: The elevated serum levels of CYFRA 21-1 observed in both groups of women with preeclampsia may reflect endothelial damage and/or dysfunction. Our results suggest that maternal serum CYFRA 21-1 is a promising biomarker for diagnosing preeclampsia. Although its value for predicting the long-term occurrence of subsequent preeclampsia may be limited, our findings indicate a trend towards elevated maternal CYFRA 21-1 levels preceding the short-term occurrence of preeclampsia in asymptomatic women. Additional prospective longitudinal studies are needed in order to determine the value of measuring maternal serum CYFRA 21-1 in predicting preeclampsia.

Prevalence of genital dysplasia after kidney transplantation--a retrospective, non-interventional study from two centers.

OBJECTIVE: Renal transplant patients are at increased risk for human papillomavirus-related malignancies of the lower genital tract. Our aim was to describe the incidence of genital dysplasia, assess the most common cervical cancer screening intervals and identify independent risk factors for the development of genital dysplasia in renal transplant patients. DESIGN: Retrospective, non-interventional study from two centers. SETTING: Post-transplant nephrologic follow-up visit at the Medical University of Vienna and a Viennese teaching hospital. POPULATION: 262 consecutive female renal transplant patients with renal transplant performed between 1980 and 2012 at the Medical University of Vienna. METHODS: Sociodemographic patient characteristics, frequency of gynecological examinations, histo- and cytopathological test results were collected. MAIN OUTCOME MEASURES: Dysplasia rates in renal transplant patients. RESULTS: 16 patients (6.2%) with genital dysplasia after renal transplant were observed. The 1-year, 3-year and 10-year proportional incidence rates for genital dysplasia in general and cervical dysplasia in particular were 1.3 and 1.3%, 3.3 and 2.7%, and 13.6 and 12.0%, respectively. Patients attended cervical cancer screening on a regular basis once a year in 82.7% of cases. In multivariate analysis re-transplantation [odds ratio 12.1 (1.5-96.3)], and renal transplant at a young age [odds ratio 0.6 (0.4-0.9)] were identified as independent risk factors for the development of female genital dysplasia. CONCLUSIONS: Female renal transplant patients have an increased risk for the development of genital dysplasia in general and of cervical dysplasia in particular. Within this cohort, women at a young age at the time of transplantation and after re-transplantation are at highest risk for the development of genital dysplasia.

Routine Use of an Antenatal Infection Screen-and-Treat Program to Prevent Preterm Birth: Long-Term Experience at a Tertiary Referral Center.

BACKGROUND: Vaginal infection in early pregnancy is associated with preterm birth. This study evaluates long-term results after integrating an antenatal screen-and-treat program for asymptomatic vaginal infections into routine pregnancy care. METHODS: We retrospectively analyzed data of all women with singleton high-risk pregnancies delivering at our tertiary referral center between 2005 and 2014. The intervention group included women who presented for a prenatal visit for a planned birth between 10 + 0 and 16 + 0 gestational weeks. Women were routinely screened for asymptomatic infections using Gram stain. In cases of bacterial vaginosis, candidiasis or trichomoniasis, women were treated according to our clinical protocol. The control group included women who did not undergo the program. Prenatal care was equal in both groups. Preterm birth served as the primary outcome variable. RESULTS: Of the 20,052 women with singleton pregnancies, 8,490 (42.3%) participated in the antenatal prevention program. The mean gestational age at birth was 38.8 ± 2.6 weeks and 37.5 ± 4.3 weeks in the intervention and control groups, respectively (p < 0.001). The incidence of preterm birth was significantly lower in the intervention group than in the control group (9.7% vs 22.3%; p < 0.001). Low-birthweight neonates, stillbirths, and late miscarriages were less frequent in the intervention group (p < 0.001). CONCLUSIONS: Long-term results support the use of an antenatal infection screen-and-treat program to prevent preterm birth. If integrated into routine pregnancy care at a high-risk obstetrical setting, this simple public health intervention could lead to a significant reduction in preterm birth, low infant birthweight, and adverse pregnancy outcomes.

Rectal Lactobacillus species and their influence on the vaginal microflora: a model of male-to-female transsexual women.

INTRODUCTION: Based on Lactobacillus species co-colonizing the vagina and rectum, it has been hypothesized that the rectum may be an important reservoir for vaginal colonization by lactobacilli. There are no data on this issue in male-to-female transsexual women. AIM: We undertook this observational study to characterize the Lactobacillus species present in the neovagina and rectum of male-to-female transsexual women and to determine the degree of neovaginal-rectal co-colonization in order to gain a better understanding of the potential role of the gut as a reservoir for genital lactobacilli. METHODS: Sixty-one male-to-female transsexual women with penile skin lined neovagina without clinical signs of infection were recruited on an ongoing basis from among male-to-female transsexual outpatients. Neovaginal and rectal smears were taken for molecular Lactobacillus species profiling by denaturing gradient gel electrophoresis (PCR-DGGE). MAIN OUTCOME MEASURES: Matching Lactobacillus species between neovagina and rectum. RESULTS: Forty-three of the 61 male-to-female transsexual women (70.5%) simultaneously harbored the same lactobacilli in both the neovagina and rectum. We found 276 neovaginal and 258 rectal DGGE bands representing 11 Lactobacillus species, with 201 matches of the same Lactobacillus species in neovagina and rectum. 37 of the 61 women (61%) had two or more matching Lactobacillus species. CONCLUSION: These data support the hypothesis that the rectum may play an important role as source of Lactobacillus species that colonies neovagina of male-to-female transsexual women. In view of the specific anatomical circumstances of the study population, these findings may be extended to the general population of women.

A case-control study about markers of stress in normal-/overweight women with polycystic ovary syndrome and in controls.

Background: Polycystic ovary syndrome (PCOS) is linked to an elevated risk of psychological disorders, decreased quality of life and emotional distress. Serum cortisol as a potential stress marker has been found to be increased in women with PCOS. The aim of this study was to evaluate both saliva stress markers and subjective psychological distress in women with PCOS. Methods: In a prospective case-control study, 31 PCOS women and 31 healthy controls were included. Salivary cortisol, and metanephrines were collected in the morning and in the evening. Emotional distress and quality of life were assessed by means of the Perceived Stress Scale (PSS-10) and the Short Form-36 (SF-36). Multivariable generalized linear models were applied to test the influence of various parameters on numerical outcome parameters. Results: After correction for age and body mass index (BMI), there were no statistically significant differences of salivary biomarkers between PCOS women and healthy controls (p>0.05). PCOS patients revealed significantly higher increased PSS total scores and lower quality of life in all SF-36 modules apart from pain (p< 0.05). The PSS total score was positively correlated to prolactin in PCOS women (r= 0.450; p= 0.011). In overweight/obese PCOS patients, a higher BMI, a higher Ferriman Gallwey score and higher age significantly predicted the PSS total score (p< 0.05). Conclusion: Stress measured by salivary biomarkers did not differ between PCOS women and healthy controls, whereas stress scores evaluated by questionnaires were significantly greater in women with PCOS. A higher BMI, hirsutism and a higher age seem to be the main modulators of subjective stress in PCOS. Prolactin might serve as a biomarker for chronic stress in PCOS women.

Glutamate decarboxylase 67 is expressed in hippocampal mossy fibers of temporal lobe epilepsy patients.

Recently, expression of glutamate decarboxylase-67 (GAD67), a key enzyme of GABA synthesis, was detected in the otherwise glutamatergic mossy fibers of the rat hippocampus. Synthesis of the enzyme was markedly enhanced after experimentally induced status epilepticus. Here, we investigated the expression of GAD67 protein and mRNA in 44 hippocampal specimens from patients with mesial temporal lobe epilepsy (TLE) using double immunofluorescence histochemistry, immunoblotting, and in situ hybridization. Both in specimens with (n = 37) and without (n = 7) hippocampal sclerosis, GAD67 was highly coexpressed with dynorphin in terminal areas of mossy fibers, including the dentate hilus and the stratum lucidum of sector CA3. In the cases with Ammon's horn sclerosis, also the inner molecular layer of the dentate gyrus contained strong staining for GAD67 immunoreactivity, indicating labeling of mossy fiber terminals that specifically sprout into this area. Double immunofluorescence revealed the colocalization of GAD67 immunoreactivity with the mossy fiber marker dynorphin. The extent of colabeling correlated with the number of seizures experienced by the patients. Furthermore, GAD67 mRNA was found in granule cells of the dentate gyrus. Levels, both of GAD67 mRNA and of GAD67 immunoreactivity were similar in sclerotic and nonsclerotic specimens and appeared to be increased compared to post mortem controls. Provided that the strong expression of GAD67 results in synthesis of GABA in hippocampal mossy fibers this may represent a self-protecting mechanism in TLE. In addition GAD67 expression also may result in conversion of excessive intracellular glutamate to nontoxic GABA within mossy fiber terminals.

Maternal smoking and fetal lung signals--an in utero MRI investigation.

OBJECTIVE: To investigate whether fetal lung signals and fetal lung signal progression over gestation observed on magnetic resonance imaging are different in mothers who reported smoking during pregnancy compared with nonsmoking controls. METHOD: Cross-sectional retrospective study of 100 consecutive singleton pregnancies that underwent magnetic resonance imaging. Fetal lung-liver signal intensity ratios of 18 fetuses of mothers who reported smoking during pregnancy were compared with 82 fetuses of nonsmoking controls. RESULTS: Average gestational age at magnetic resonance imaging was 26.4 ± 5.2 weeks (Range 18.4-38.2 weeks). Cases reported smoking between 2 and 15 cigarettes per day. The mean number of cigarettes per day for cases was 9.2 ± 3.4. Mean fetal lung-liver signal intensity ratios did not differ significantly between the two groups (p = 0.8). They showed a linear increase with gestational age (r(2) = 0.3). Multiple regression analysis of lung-liver signal intensity ratios using gestational age and smoking status as predictors revealed a significant influence of gestational age (p < 0.0001) but not maternal smoking status (p = 0.8) on fetal lung-liver signal intensity ratios. CONCLUSIONS: Fetuses of mothers who reported smoking during pregnancy show similar lung signals and lung signal progression over gestation on magnetic resonance imaging as nonsmoking controls.

Analyses of human granulosa cell vitality by fluorescence activated cell sorting after rapid cooling.

In reproductive medicine, the technique of rapid cooling becomes increasingly important for the preservation of tissue and cells. In order to protect the cells, incubation in different cryopreservation solutions is essential. The speed of the cooling process also makes a pivotal contribution to the success of this method. Using Flourescence Activated Cell Sorting (FACS), we investigated the impact of an open rapid and a closed rapid cooling technique on the vitality of human granulosa cells. Furthermore, we examined effects of the different solutions used for rapid cooling and warming before and after rapid cooling. We found a significant lower proportion of vital cells after rapid cooling compared to untreated controls independently of the technique and the tube size. However, we did not find any significant differences between open and closed rapid cooling. In both, a lower proportion of vital granulosa cells were found after incubation in rapid cooling solution only compared to warming solution only. Our results lend support to the conclusion that the difference of cooling-speed between open and closed rapid cooling is, in our settings, not crucial for the success of the procedure and that cryoprotective agents in the rapid cooling solutions have a higher potential to cause severe cell damage than agents used for warming.