Influence of Nodal signalling on pluripotency factor expression, tumour cell proliferation and cisplatin-sensitivity in testicular germ cell tumours.

BACKGROUND: Testicular germ cell tumours (TGCTs) are characterised by an overall high cisplatin-sensitivity which has been linked to their continued expression of pluripotency factors. Recently, the Nodal signalling pathway has been implicated in the regulation of pluripotency factor expression in fetal germ cells, and the pathway could therefore also be involved in regulating expression of pluripotency factors in malignant germ cells, and hence cisplatin-sensitivity in TGCTs. METHODS: We used in vitro culture of the TGCT-derived cell line NTera2, ex vivo tissue culture of primary TGCT specimens and xenografting of NTera2 cells into nude mice in order to investigate the consequences of manipulating Nodal and Activin signalling on pluripotency factor expression, apoptosis, proliferation and cisplatin-sensitivity. RESULTS: The Nodal signalling factors were markedly expressed concomitantly with the pluripotency factor OCT4 in GCNIS cells, seminomas and embryonal carcinomas. Despite this, inhibition of Nodal and Activin signalling either alone or simultaneously did not affect proliferation or apoptosis in malignant germ cells in vitro or ex vivo. Interestingly, inhibition of Nodal signalling in vitro reduced the expression of pluripotency factors and Nodal pathway genes, while stimulation of the pathway increased their expression. However, cisplatin-sensitivity was not affected following pharmacological inhibition of Nodal/Activin signalling or siRNA-mediated knockdown of the obligate co-receptor CRIPTO in NTera2 cells in vitro or in a xenograft model. CONCLUSION: Our findings suggest that the Nodal signalling pathway may be involved in regulating pluripotency factor expression in malignant germ cells, but manipulation of the pathway does not appear to affect cisplatin-sensitivity or tumour cell proliferation.

Positive association between cholesterol in human seminal plasma and sperm counts: results from a cross-sectional cohort study and immunohistochemical investigations.

BACKGROUND: Cholesterol is essential for cell membrane stability, permeability, and fluidity. Cholesterol is present in seminal plasma, but whether a relationship between the level of cholesterol in seminal plasma and semen quality exists remains to be elucidated. OBJECTIVES: To explore the association between cholesterol levels in seminal plasma and serum cholesterols, semen quality, and serum reproductive hormones. Secondly, to explore whether the associations are biologically plausible. MATERIALS AND METHODS: An association study between cholesterol levels in seminal plasma and semen quality in 403 men, median age 19 years, from the general population. Additionally, an immunohistochemical evaluation of proteins involved in cholesterol metabolism and transport in tissues from the male reproductive tract (testis, epididymis, prostate, and seminal vesicle). Tissue specimens were investigated by immunohistochemistry for markers of cholesterol metabolism and transport (ABCA1, ABCG1, CYP11A1, CYP51A1, HMGCR, LAL, LCAT, LDLR, and SOAT1). RESULTS: Trend analyses showed that total amount of total cholesterol in seminal plasma was positively associated with sperm concentration, total sperm count, sperm motility, and morphology (all p < 0.008, adjusted). Cholesterol concentrations in seminal plasma were neither associated with serum cholesterol and lipid levels nor serum reproductive hormone (FSH, LH, testosterone, estradiol, sex-hormone-binding globulin, inhibin b) levels. All investigated markers of cholesterol metabolism and transport were expressed in the investigated tissue specimens to varying degrees. DISCUSSION: Seminal plasma level of cholesterol was positively associated with semen parameters. The presence of proteins and enzymes involved in cholesterol metabolism in Leydig cells, Sertoli cells, and maturing germ cells in the seminiferous tubules supports the view that cholesterol may be important for spermatogenesis. CONCLUSION: Cholesterol level in seminal plasma may be an indicator of semen quality. Investigations are needed to corroborate or refute our findings and to clarify the exact role of cholesterols for semen quality.

Danish Prostate Cancer Registry - methodology and early results from a novel national database.

BACKGROUND: Systematized Nomenclature of Medicine (SNOMED) codes are computer-processable medical terms used to describe histopathological evaluations. SNOMED codes are not readily usable for analysis. We invented an algorithm that converts prostate SNOMED codes into an analyzable format. We present the methodology and early results from a new national Danish prostate database containing clinical data from all males who had evaluation of prostate tissue from 1995 to 2011. MATERIALS AND METHODS: SNOMED codes were retrieved from the Danish Pathology Register. A total of 26,295 combinations of SNOMED codes were identified. A computer algorithm was developed to transcode SNOMED codes into an analyzable format including procedure (eg, biopsy, transurethral resection, etc), diagnosis, and date of diagnosis. For validation, ~55,000 pathological reports were manually reviewed. Prostate-specific antigen, vital status, causes of death, and tumor-node-metastasis classification were integrated from national registries. RESULTS: Of the 161,525 specimens from 113,801 males identified, 83,379 (51.6%) were sets of prostate biopsies, 56,118 (34.7%) were transurethral/transvesical resections of the prostate (TUR-Ps), and the remaining 22,028 (13.6%) specimens were derived from radical prostatectomies, bladder interventions, etc. A total of 48,078 (42.2%) males had histopathologically verified prostate cancer, and of these, 78.8% and 16.8% were diagnosed on prostate biopsies and TUR-Ps, respectively. FUTURE PERSPECTIVES: A validated algorithm was successfully developed to convert complex prostate SNOMED codes into clinical useful data. A unique database, including males with both normal and cancerous histopathological data, was created to form the most comprehensive national prostate database to date. Potentially, our algorithm can be used for conversion of other SNOMED data and is available upon request.

Screening for carcinoma in situ in the contralateral testicle in patients with testicular cancer: a population-based study.

BACKGROUND: Screening programmes for contralateral carcinoma in situ (CIS) testis in patients with unilateral germ-cell cancer (GCC) have never been evaluated. We investigated the effect of screening for contralateral CIS in a large nation-wide, population-based study. PATIENTS AND METHODS: A contralateral single-site biopsy was offered to 4130 patients in whom GCC had been diagnosed in 1984-2007 (screened cohort); 462 patients in whom GCC was diagnosed in 1984-1988 comprised the unscreened cohort. Cases with CIS were offered radiotherapy. Initially CIS-negative biopsies in patients with metachronous GCC were revised according to today's standards. Risk for metachronous GCC was estimated using cumulative incidence and the Cox proportional hazards model. RESULTS: In the screened cohort, contralateral CIS was found in 181 (4.4%) patients. The cumulative incidence of metachronous GCC after 20 years was 1.9% in the screened cohort and 3.1% in the unscreened cohort (P = 0.097), hazard ratio (HR) for the unscreened cohort: 1.59 (P = 0.144). Expert revision with contemporary methodology of CIS-negative biopsy samples from patients with metachronous cancer revealed CIS in 17 out of 45 (38%) cases. Decreased risks for metachronous GCC were related to older age at diagnosis (HR 0.52 per 10 years, P < 0.001) and chemotherapy (HR 0.35, P = 0.002). Limitations include the small number of patients in the unscreened cohort and the retrospective study design. CONCLUSIONS: Our evaluation of a national population-based screening programme for contralateral CIS in patients with testicular cancer showed no significant difference in the risk for metachronous GCC between a screened and an unscreened cohort. Single-site biopsy including modern immunohistochemistry does not identify all cases of CIS.

Hanging drop cultures of human testis and testis cancer samples: a model used to investigate activin treatment effects in a preserved niche.

BACKGROUND: Testicular germ cell tumours of young adults, seminoma or non-seminomas, are preceded by a pre-invasive precursor, carcinoma in situ (CIS), understood to arise through differentiation arrest of embryonic germ cells. Knowledge about the malignant transformation of germ cells is currently limited by the lack of experimental models. The aim of this study was to establish an experimental tissue culture model to maintain normal and malignant germ cells within their niche and allow investigation of treatment effects. METHODS: Human testis and testis cancer specimens from orchidectomies were cultured in 'hanging drops' and effects of activin A and follistatin treatment were investigated in seminoma cultures. RESULTS: Testis fragments with normal spermatogenesis or CIS cells were cultured for 14 days with sustained proliferation of germ cells and CIS cells and without increased apoptosis. Seminoma cultures survived 7 days, with proliferating cells detectable during the first 5 days. Activin A treatment significantly reduced KIT transcript and protein levels in seminoma cultures, thereby demonstrating a specific treatment response. CONCLUSIONS: Hanging drop cultures of human testis and testis cancer samples can be employed to delineate mechanisms governing growth of normal, CIS and tumorigenic germ cells retained within their niche.

Individual versus standard dose of rFSH in a mild stimulation protocol for intrauterine insemination: a randomized study.

BACKGROUND: Controlled ovarian stimulation (COS) and intrauterine insemination (IUI) are often used as the first-line treatment for subfertile couples. To minimize the variability in ovarian response in patients' first treatment cycle, we recently developed a recombinant follicle-stimulating hormone (rFSH) dosage nomogram. The nomogram has now been tested. METHODS: Multicentre randomized controlled trial (RCT) including 228 ovulatory patients scheduled for COS and IUI. Patients were randomized to 'individual' (50-100 IU rFSH/day, n = 113) or 'standard' (75 IU rFSH/day, n = 115) dose. 'Individual' dose was prescribed according to the nomogram, which was based on patients' body weight and antral follicle count. The primary end-point was the proportion of patients with two to three follicles > or = 14 mm (maximum two follicles > or = 18 mm) on the day of hCG (leading follicle = 18 mm). Primary analysis was made by intention-to-treat. RESULTS: In the 'individual' group, 79/113 (70%) of the patients developed two to three follicles versus 64/115 (56%) in the 'standard' group [absolute difference = 14.3 percentage points; 95% confidence interval (CI) 2-26, P = 0.03; absolute difference = 14.4; 95% CI 2-27, P = 0.02, when adjusting for centre]. Among patients with two to three follicles, the proportion of patients with two follicles was 46/79 (58%) in the 'individual' group versus 34/64 (53%) in the 'standard' group, P = 0.54. Ongoing pregnancy rate was 23/113 (20%) in the 'individual' group and 21/115 (18%) in the 'standard' group and the rate of multiple gestations was 1/113 (1%) versus 5/115 (4%), P = 0.21. CONCLUSIONS: This RCT is the first to clinically test a dosage nomogram in ovulatory IUI patients' first rFSH treatment cycle. Dosing according to the nomogram was superior to standard dosing. TRIAL REGISTRATION: ClinicalTrials.gov Identifier NCT00374634.

Involuntary automaticity: a potential legal defence against an allegation of clinical negligence?

BACKGROUND: It has been argued that the safety of patients can be put at risk because the verbal challenge--response protocols, frequently used by healthcare professionals--can be compromised by the system of work in use and the operational environment. Individuals who constantly undertake verbal double-checking safety protocols, when subjected to adverse operational conditions such as high workloads, strict time constraints and high levels of stress, can be captured by a so-called "involuntary automaticity." CONCLUSION: Evidence suggests that involuntary automaticity could be used as a potential legal defence, against an allegation of clinical negligence, where the healthcare professional concerned had warned their management that the conditions in which they worked could promote the phenomenon, and no remedial actions had been taken by them.

Predictors of ovarian response in intrauterine insemination patients and development of a dosage nomogram.

The objective of this prospective study was to identify predictors of ovarian response in ovulatory patients treated with low-dose recombinant FSH (rFSH), gonadotrophin-releasing hormone antagonist and intrauterine insemination (IUI), and to develop an rFSH dosage nomogram based on the findings. Patients (n = 159) were stimulated with a starting dose of 75 IU rFSH/day. Ten parameters were investigated as possible predictors of the number of mature follicles >or=15 mm: age, spontaneous cycle length, body weight, body mass index, smoking status, total ovarian volume, total number of antral follicles, total Doppler score of the ovarian stromal blood flow, baseline FSH and oestradiol. Simple and multiple linear regressions were used for the statistical analysis. Appropriate ovarian response was defined as two to three mature follicles. Body weight (P = 0.001) and the number of antral follicles (P = 0.004) were the strongest independent predictive factors of the number of mature follicles. In conclusion, body weight and antral follicle count may be used to achieve appropriate ovarian response for IUI in ovulatory patients. Based on this, a simple rFSH dosage nomogram was developed for individual ovarian stimulation prior to IUI.

Assisted hatching improves implantation rates on cryopreserved-thawed embryos. A randomized prospective study.

BACKGROUND: Focus on the hatching process has so far been in the field of fresh embryos. Cryopreserved-thawed embryos have a lower rate of pregnancy than fresh embryos, which might be due to hardening of the zona pellucida. METHODS: During a 2 year period, a prospective randomized study enrolling 253 cryopreserved-thawed cycles was performed on day 2 embryos. Pseudorandomization to assisted hatching or a control group was done on the basis of even and odd dates for thawing. One hour before embryo transfer, hatching was carried out using acidic Tyrode's solution. RESULTS: Among 136 embryos exposed to assisted hatching, 11.4% (30) were implanted compared with only 5.8% (13) of 117 embryos not exposed to assisted hatching (P<0.05, chi(2) test). No difference in the rate of clinical pregnancy and positive serum HCG was observed between the two groups. Very few women >38 years old were included in the study, and no significant difference according to age could be found between the groups. CONCLUSIONS: These results show that assisted hatching using acidic Tyrode's solution increases the implantation rate of cryopreserved-thawed embryos (P<0.05).

Progesterone supplementation during early gestations after IVF or ICSI has no effect on the delivery rates: a randomized controlled trial.

BACKGROUND: The aim was to study whether prolongation of luteal support during early pregnancy influences the delivery rate after IVF. METHODS: Dual centre study including 303 women who achieved pregnancy after IVF or ICSI was used. All were treated with the long protocol using GnRH agonists and given luteal support with 200 mg vaginal progesterone three times daily during 14 days from the day of transfer until the day of a positive HCG test. The study group (n = 150) withdrew vaginal progesterone from the day of positive HCG. The control group (n = 153) continued administration of vaginal progesterone during the next 3 weeks of pregnancy. RESULTS: The number of miscarriages prior to and after week 7 of gestation was seven (4.6%) and 15 (10.0%) in the study group and five (3.3%) and 13 (8.5%) in the control group respectively. The number of deliveries was 118 (78.7 %) in the study group and 126 (82.4 %) in the control group. The differences were not significant. CONCLUSIONS: Prolongation of progesterone supplementation in early pregnancy has no influence on the miscarriage rate, and thus no effect on the delivery rate. Progesterone supplementation can safely be withdrawn at the time of a positive HCG test.

Serological investigation of Mycoplasma genitalium in infertile women.

BACKGROUND: The role of Mycoplasma genitalium in the pathogenesis of pelvic inflammatory disease has not been characterized. METHODS: Sera from 308 infertile women were investigated for antibodies to M. genitalium by immunoblotting. Women with tubal factor infertility (TFI) made up 132 of the patients, 67 of the women had an infertile male partner and 109 were infertile for unknown reasons. RESULTS: Of the TFI patients 29 (22.0%) were seropositive to the major adhesin, MgPa, of M. genitalium versus 11 (6.3%) in the group of women with normal tubes. No cross-reactions between MgPa and P1 of the related Mycoplasma pneumoniae were found. Besides, MgPa positive sera were confirmed by immunoblotting using a cloned fragment of the C-terminal part of MgPa specific to M. genitalium. Chlamydia trachomatis is known to be able to cause infertility as a result of salpingitis. Therefore, the sera were tested against C. trachomatis using a commercial ELISA test. Seventy-five (56.8%) of the TFI patients were seropositive to C. trachomatis. Eight (27.6%) TFI patients seropositive to MgPa were negative to C. trachomatis. CONCLUSIONS: This study indicates that M. genitalium may be an independent risk factor in the development of an inflammatory process leading to scarring of the uterine tubes in women and thereby causing infertility.

Birth of a healthy girl after ICSI with ejaculated spermatozoa from a man with non-mosaic Klinefelter's syndrome.

Klinefelter's syndrome is a major contributor to male infertility. Recent reports of births after ICSI with especially testicular spermatozoa from infertile men with this syndrome are promising. The birth of a healthy girl after ICSI treatment with ejaculated spermatozoa from a man with non-mosaic Klinefelter's syndrome is reported. The non-mosaic karyotype was confirmed by chromosome analysis of both peripheral blood leukocytes and fibroblasts from a skin biopsy. In conclusion, in a very few cases, men with apparently non-mosaic Klinefelter's syndrome have ejaculated spermatozoa that can result in a birth of a healthy child following ICSI.

Comparison of two ibuprofen formulations in the treatment of shoulder tendonitis.

In order to compare the efficacy and tolerance of two drug formulations of ibuprofen, conventional tablets 600 mg QID (CI) and sustained-release tablets 1200 mg BID (SRI), a total of 147 patients in 7 centres in Denmark with nontraumatic shoulder pain were included in a double-blind dummy study. Initially all patients received a local injection of corticosteroid and local anaesthetic, and were randomly allocated either drug (CI or SRI) for a period of 3 weeks. Complete relief was recorded from significantly more of the patients in the CI group (21%) than in the SRI group (7%) while a similar number of patients improved viz., 67% of the SRI treated group and 77% of the CI treated group. Based on doctor's assessment improvement in the two groups was equal. 44% of the patients recorded side effects, the number and pattern being the same in the two groups. No serious side effects were recorded. It is concluded that the two treatment regimens can be rated as clinically equivalent.

Medical treatment of ectopic pregnancy.

Methotrexate is an alternative to surgical treatment of unruptured tubal pregnancy. Among 39 women with the diagnosis of tubal pregnancy there were ten10 cases without rupture of salpinx. Nine9 out of these treated with Methotrexate. In one1 case there was rupture immediately after the termination of the treatment. There were no experiences of major side effects from the treatment with Methotrexate. The passage through the tubes was examined laparoscopically 3 months after the treatment. There was passage through all affected tubes.

Medical treatment of ectopic pregnancy.

Methotrexate is an alternative to surgical treatment of unruptured tubal pregnancy. Among 39 women with the diagnosis of tubal pregnancy there were ten (10) cases without rupture of salpinx. Nine (9) out of these treated with Methotrexate. In one (1) case there was rupture immediately after the termination of the treatment. There were no experiences of major side effects from the treatment with Methotrexate. The passage through the tubes was examined laparoscopically 3 months after the treatment. There was passage through all affected tubes.

Medical treatment of ectopic pregnancy.

Methotrexate is an alternative to surgical treatment of unruptured tubal pregnancy. Among 39 women with the diagnosis of tubal pregnancy there were ten cases without rupture of the salpinx. Nine out of these were treated with methotrexate. In one case there was a tubal rupture immediately after the termination of the treatment. There were no major side effects from the treatment with methotrexate. Tubal patency was tested for laparoscopically three months after the treatment. There was patency of all previously affected tubes.

Epithelial atypia in ovarian mucinous cystadenoma during the puerperium.

A papillary mucinous cystadenoma of the ovary with unique benign epithelial atypia is presented. The tumor was removed from a 33-year-old puerperal woman. Morphologically, the lining epithelium of the cystadenoma strikingly resembled the so-called Arias-Stella reaction of the endometrial epithelium, a benign cellular atypia produced by chorionic tissue during pregnancy. The atypical epithelium of the endometrium is known to persist several weeks after parturition or abortion, and should not be misinterpreted as malignancy.