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.

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.

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.

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.

15-Hdroxyprostaglandin dehydrogenase activity in vitro in lung and kidney of essential fatty acid-deficient rats.

Weanling rats were fed for 6 months on a diet deficient in essential fatty acids: either fat-free, or with 28% (w/w) partially hydrogenated fish oil. Control rats were fed a diet with 28% (w/w) arachis oil for 6 months. 15-Hydroxyprostaglandin dehydrogenase activity was determined as initial rates of formation of 3H-labelled 15-keto-dihydro-prostaglandin E1 plus 15-keto-prostaglandin E1 in high speed supernatants of lung and kidney from each of the groups of rats. Dehydrogenase activity (expressed as either pmol/min per mg soluble protein, or as nmol/min per g tissue) was decreased 30--40% in the lungs of the two groups on diets deficient in essential fatty acids as compared to the control group. No difference were observed in dehydrogenase activity in the kidneys. The dehydrogenase may be of importance for the regulation of the level of endogenous prostaglandins and, thus, a decrease in activity could result in a slower turnover of prostaglandins.

Metabolism of prostaglandin E1 and of glutathione conjugate of prostaglandin A1 (GSH-prostaglandin A1) by prostaglandin 9-ketoreductase from rabbit kidney.

Rabbit kidney prostaglandin 9-ketoreductase was found to metabolize the glutathione conjugate of prostaglandin A1 (GSH-prostaglandin A1). Apparent Km (GSH-prostaglandin A1) 13 microM and apparent Km (prostaglandin E1) 200 microM. The cytosolic preparation was subjected to gelfiltration and isoelectric focusing, which revealed that metabolism of prostaglandin E1 and GSH-prostaglandin A1 occurs by means of the same fractions. Furthermore, prostaglandin E1 and GSH-prostaglandin A1 are competitive inhibitors of the enzyme, when GSH-prostaglandin A1 and prostaglandin E1 are tested as substrates, respectively. It si concluded, that GSH-prostaglandin A1 is a much better substrate for prostaglandin 9-ketoreductase from rabbit kidney than is prostaglandin E1.

Kinetics of citalopram in elderly patients.

The kinetics of the antidepressant drug citalopram, a specific 5-HT uptake inhibitor, has been investigated in 11 elderly patients (age 73-90) and compared to previous data from younger patients and volunteers. The recorded steady state citalopram levels of 140-545 nM after a once-daily 20-mg dose were up to four times higher than expected from data in younger patients and volunteers. The biological half-life (1.5-3.75 days) and estimated systemic clearance (0.08-0.3 1/min) also differed from data in younger individuals (1.5 days and 0.4 1/min, respectively). Clearance values generally decreased with increasing age. Drug/metabolite ratios were higher in patients with the longest half-lives and lowest citalopram clearance, indicating reduced metabolic activity. No reduction in renal clearance was indicated by urine data, obtained for two of the patients. The study suggests that daily doses of 5-20 mg give approximately the same steady state plasma levels in elderly patients as doses of 40 mg in younger, and that this is due to reduced rates of metabolism in the elderly.

A double-blind, crossover study of a sustained-release tablet of ketoprofen and normal ketoprofen capsules in the treatment of patients with osteoarthritis.

A double-blind, crossover study was carried out to assess the efficacy and tolerance of a sustained-release tablet formulation of ketoprofen given as a single daily 200 mg dose compared with 2 X 50 mg normal formulation capsules of ketoprofen twice daily. Eighty-four patients with osteoarthritis of the hip and/or knee were admitted and received treatment for periods of 3 weeks, preceded by a 1-week placebo wash-out period, with each of the two formulations, in random order. Patients were seen after each study period and clinical objective and subjective assessments made of signs and symptoms of the disease, consumption of rescue analgesic and unwanted effects. Forty-eight of the patients continued, mainly on the sustained-release formulation, in an open long-term tolerance study lasting 3 months. The results were analyzed for 68 patients who completed the double-blind phase and for 33 who completed the open phase of the study. The patients who were withdrawn did so mainly for non-drug related reasons; 19 patients did so because of gastric disorders during the first phase. The incidence of side-effects was low and similar in frequency and nature with both formulations; those that were reported were mild and principally gastro-intestinal. Both active treatment periods afforded similar symptomatic relief and were preferred to placebo by all but 2 patients. No significant differences were found between active treatments, although there was a trend in favour of the sustained-release formulation for most of the parameters studied as there was in patient preference.

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.

[Laparoscopic sterilization during the puerperium employing electrocoagulation. A retrospective study of 76 women]. / Laparoskopisk sterilisation ved hjaelp af elektrokoagulation i puerperiet. En retrospektiv undersøgelse af 76 kvinder.

PIP: 76 women sterilized on the 4th day postpartum by laparoscopic electrocoagulation were contacted 2-6 years after the event. Mean observation time was 4.2 years, mean age was 32.3 years. 2 (2.8%) became pregnant. This does not differ significantly from the frequency of pregnancy after nonpuerperal sterilization by the same method. Both pregnancies were intrauterine and no complications were observed. (author's modified)^ieng