An update on semen quality among young Finnish men and comparison with Danish data.

BACKGROUND: Finnish men used to have higher semen quality than Danish men. However, recent studies showed that semen quality in Finland has declined, but it has been relatively stable in Denmark. OBJECTIVE: This study aimed to compare new data on semen quality of the young Finnish men to that of Danish men. MATERIALS AND METHODS: In this cross-sectional study, 18- to 19-year-old men residing in Turku, Finland and Copenhagen, Denmark, were invited to participate in 2008-2011. Each man filled in a questionnaire, provided one semen sample and underwent andrological examination. Semen samples were analyzed according to WHO. Multiway ANOVA was used to adjust semen variables for duration of sexual abstinence and age (and time from ejaculation to the start of semen analysis for sperm motility). RESULTS: Altogether 287 Finnish men and 873 Danish men participated in the study. The adjusted median sperm concentrations were 49 and 47 million/mL for Finnish and Danish men, respectively (p = 0.48). The adjusted median total sperm counts were 148 million in Finland and 146 million in Denmark (p = 0.87). The adjusted median percentages of morphologically normal spermatozoa were 6.9% in Finland and 6.5% in Denmark, p = 0.27. Finnish men had higher adjusted median percentages of motile spermatozoa (A+B+C) than Danish men (80% vs. 69%, p < 0.001). The proportion of men who had low semen quality (sperm concentration, percentage of morphologically normal spermatozoa or percentage of progressively motile spermatozoa below WHO reference limits) was lower in Finland (25.4%) than in Denmark (34.6%), p = 0.004. DISCUSSION: Considerable percentage of men in both countries had low semen quality. The deteriorating semen quality in Finland may result in decreasing fecundity, which is a cause of concern. CONCLUSION: The formerly high semen quality in Finland has converged to the lower Danish levels. Our findings demonstrate the importance of continuing surveillance of semen quality.

Secreted frizzled-related protein 2 (SFRP2) expression promotes lesion proliferation via canonical WNT signaling and indicates lesion borders in extraovarian endometriosis.

STUDY QUESTION: What is the role of SFRP2 in endometriosis? SUMMARY ANSWER: SFRP2 acts as a canonical WNT/CTNNB1 signaling agonist in endometriosis, regulating endometriosis lesion growth and indicating endometriosis lesion borders together with CTNNB1 (also known as beta catenin). WHAT IS KNOWN ALREADY: Endometriosis is a common, chronic disease that affects women of reproductive age, causing pain and infertility, and has significant economic impact on national health systems. Despite extensive research, the pathogenesis of endometriosis is poorly understood, and targeted medical treatments are lacking. WNT signaling is dysregulated in various human diseases, but its role in extraovarian endometriosis has not been fully elucidated. STUDY DESIGN, SIZE, DURATION: We evaluated the significance of WNT signaling, and especially secreted frizzled-related protein 2 (SFRP2), in extraovarian endometriosis, including peritoneal and deep lesions. The study design was based on a cohort of clinical samples collected by laparoscopy or curettage and questionnaire data from healthy controls and endometriosis patients. PARTICIPANTS/MATERIALS, SETTING, METHODS: Global gene expression analysis in human endometrium (n = 104) and endometriosis (n = 177) specimens from 47 healthy controls and 103 endometriosis patients was followed by bioinformatics and supportive qPCR analyses. Immunohistochemistry, Western blotting, primary cell culture and siRNA knockdown approaches were used to validate the findings. MAIN RESULTS AND THE ROLE OF CHANCE: Among the 220 WNT signaling and CTNNB1 target genes analysed, 184 genes showed differential expression in extraovarian endometriosis (P < 0.05) compared with endometrium tissue, including SFRP2 and CTNNB1. Menstrual cycle-dependent regulation of WNT genes observed in the endometrium was lost in endometriosis lesions, as shown by hierarchical clustering. Immunohistochemical analysis indicated that SFRP2 and CTNNB1 are novel endometriosis lesion border markers, complementing immunostaining for the known marker CD10 (also known as MME). SFRP2 and CTNNB1 localized similarly in both the epithelium and stroma of extraovarian endometriosis tissue, and interestingly, both also indicated an additional distant lesion border, suggesting that WNT signaling is altered in the endometriosis stroma beyond the primary border indicated by the known marker CD10. SFRP2 expression was positively associated with pain symptoms experienced by patients (P < 0.05), and functional loss of SFRP2 in extraovarian endometriosis primary cell cultures resulted in decreased cell proliferation (P < 0.05) associated with reduced CTNNB1 protein expression (P = 0.05). LIMITATIONS REASONS FOR CAUTION: SFRP2 and CTNNB1 improved extraovarian endometriosis lesion border detection in a relatively small cohort (n = 20), although larger studies with different endometriosis subtypes in variable cycle phases and under hormonal medication are required. WIDER IMPLICATIONS OF THE FINDINGS: The highly expressed SFRP2 and CTNNB1 improve endometriosis lesion border detection, which can have clinical implications for better visualization of endometriosis lesions over CD10. Furthermore, SFRP2 acts as a canonical WNT/CTNNB1 signaling agonist in endometriosis and positively regulates endometriosis lesion growth, suggesting that the WNT pathway may be an important therapeutic target for endometriosis. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by the Academy of Finland and by Tekes: Finnish Funding Agency for Innovation. The authors have no conflict of interest to declare.

Prevalence of Pain Symptoms Suggestive of Endometriosis Among Finnish Adolescent Girls (TEENMAPS Study).

STUDY OBJECTIVE: To evaluate the prevalence of pain symptoms suggestive of endometriosis among adolescent girls aged 15-19 years. DESIGN: Cross-sectional study. SETTING: University hospital. PARTICIPANTS: Adolescent girls aged 15-19 years attending elementary school, high school, or vocational institute at 3 cities in Southwest Finland in 2010-2011. INTERVENTIONS: The school nurses distributed a detailed questionnaire to 2582 girls who were attending school at the time of the study. Completion of the questionnaire was voluntary and anonymous. MAIN OUTCOME MEASURES: Prevalence and severity of dysmenorrhea, acyclic abdominal pain, dyspareunia, dyschezia, and dysuria. Severity was evaluated with an 11-point numerical rating scale (NRS). RESULTS: A total of 1103 eligible answers were analyzed. The prevalence of dysmenorrhea was 68% (738/1092) with mean NRS of 7.0 (SD = 2.0). Acyclic abdominal pain, dyspareunia, dyschezia, and dysuria were less frequent (19% [207/1085], 12% [53/458], 8% [87/1088] and 5% [50/1084], respectively). The prevalence of severe dysmenorrhea (NRS 8-10) was 33% (355/1089). Severe dysmenorrhea was associated with increased risk of concurrent acyclic abdominal pain (odds ratio [OR] = 2.7; 95% confidence interval [CI], 2.0-3.6), dyschezia (OR = 2.5; 95% CI, 1.6-3.9), and regular absenteeism from school or hobbies (OR = 10.0; 95% CI, 4.2-23.6). Using different criteria, 2%-10% (21-106/1103) of all girls could be identified as having symptoms suggestive of endometriosis. Five percent of girls (n = 53/1103) had severe dysmenorrhea, used oral contraceptive pills, and reported inadequate relief from pain medication. CONCLUSION: One-third (355/1089) of 15- to 19-year-old girls had severe menstrual pain and 14% (49/355) of them were regularly absent from school or hobbies. Five percent of all teenage girls (53/1103) were poor responders to conventional therapy for primary dysmenorrhea.

A cohort effect on serum testosterone levels in Finnish men.

OBJECTIVE: To investigate whether a population-level decline in serum testosterone exists in Finnish men. In comparison with other European populations, Finnish men have compared well in the studies of reproductive health (i.e. semen quality, incidence of cryptorchidism and testicular cancer); thus, we expected no significant cohort-dependent decrease in serum testosterone. METHODS: We analysed serum levels of testosterone, gonadotrophin and sex hormone binding globulin (SHBG) in 3271 men representing different ages (25-74 years) and birth cohorts within three large Finnish population surveys conducted in 1972, 1977 and 2002. RESULTS: Serum testosterone levels decreased (from 25.3 nmol/l in 25- to 29-year-old men gradually to 16.9 nmol/l in 70- to 74-year-old men), whereas SHBG and gonadotrophin levels increased with increasing age. In addition, a significant secular trend in testosterone (total and free), SHBG and gonadotrophin levels was observed with lower levels in more recently born age-matched men. Serum testosterone level decreased in men aged 60-69 years from 21.9 nmol/l (men born 1913-1922) to 13.8 nmol/l (men born 1942-1951). These decreases remained significant following adjustment for BMI. An age-independent birth cohort effect existed on reproductive hormones measured in the Finnish men. In concert with the lower free testosterone levels, we observed lower gonadotrophin levels, suggesting that while there may be detrimental changes at the gonad level, the hypothalamus-pituitary-axis is not responding appropriately to this change. CONCLUSIONS: The more recently born Finnish men have lower testosterone levels than their earlier born peers. This study offers no explanation for this substantial recent adverse development.

Serum HE4 concentration is not dependent on menstrual cycle or hormonal treatment among endometriosis patients and healthy premenopausal women.

OBJECTIVE: Human epididymal secretory protein E4 (HE4) is a new promising tumor marker developed for the diagnostics and follow up of ovarian cancer. It has yet to become widely accepted in clinical practice, and its biological properties have not been inclusively studied. The aim of this study was to investigate whether serum HE4 concentration varies within the normal menstrual cycle and whether common gynecological hormonal treatments have an effect on HE4 values. METHODS: Our study population consisted of 180 women, including 126 endometriosis patients and 54 healthy women. We measured their serum HE4 and CA125 concentrations and evaluated the effect of the menstrual cycle and the possible hormonal medication on these marker concentrations. RESULTS: We found no significant variation in serum HE4 concentrations in samples taken at different phases of the menstrual cycle. The median HE4 concentrations in proliferative, secretory and menstrual phase were 41.5, 45.1 and 35.3 pM in healthy women, and 43.4, 44.3 and 43.0 pM in endometriosis patients, respectively. The use of combined estrogen and progestin contraceptives did not affect serum HE4 levels significantly. CONCLUSIONS: The present study shows that the HE4 measurement in healthy premenopausal women as well as in women with endometriosis can be carried out at any phase of the menstrual cycle, and irrespective of hormonal medication, extending the benefits of HE4 use in clinical practice.

Recent adverse trends in semen quality and testis cancer incidence among Finnish men.

Impaired semen quality and testicular cancer may be linked through a testicular dysgenesis syndrome of foetal origin. The incidence of testis cancer has been shown to increase among Finnish men, whereas there is no recent publication describing temporal trends in semen quality. Therefore, we carried out a prospective semen quality study and a registry study of testis cancer incidence among Finnish men to explore recent trends. A total of 858 men were investigated in the semen quality study during 1998-2006. Median sperm concentrations were 67 (95% CI 57-80) million/mL, 60 (51-71) and 48 (39-60) for birth cohorts 1979-81, 1982-83 and 1987; total sperm counts 227 (189-272) million, 202 (170-240) and 165 (132-207); total number of morphologically normal spermatozoa 18 (14-23) million, 15 (12-19) and 11 (8-15). Men aged 10-59 years at the time of diagnosis with testicular cancer during 1954-2008 were included in the registry study, which confirmed the increasing incidence of testicular cancer in recent cohorts. These simultaneous and rapidly occurring adverse trends suggest that the underlying causes are environmental and, as such, preventable. Our findings necessitate not only further surveillance of male reproductive health but also research to detect and remove the underlying factors.

Serum HE4 concentration differentiates malignant ovarian tumours from ovarian endometriotic cysts.

Human epididymal secretory protein E4 (HE4, also known as WAP four-disulphide core domain protein 2) is a new promising biomarker for ovarian cancer but its specificity against ovarian endometriotic cysts is only superficially known. We, thus, analysed serum HE4 concentrations together with a tumour marker CA125 in serum samples of women diagnosed with various types of endometriosis, endometrial cancer or ovarian cancer, and in samples from healthy controls. The mean serum concentration of HE4 was significantly higher in serum samples of patients with both endometrial (99.2 pM, P<0.001) and ovarian (1125.4 pM, P<0.001) cancer but not with ovarian endometriomas (46.0 pM) or other types of endometriosis (45.5 pM) as compared with healthy controls (40.5 pM). The serum CA125 concentrations were elevated in patients with ovarian cancer, advanced endometriosis with peritoneal or deep lesions, or ovarian endometriomas, but not in the patients with endometrial cancer. The microarray results revealed that the mRNA expression of the genes encoding HE4 and CA125 reflected the serum protein concentrations. Taken together, measuring both HE4 and CA125 serum concentrations increases the accuracy of ovarian cancer diagnosis and provides valuable information to discriminate ovarian tumours from ovarian endometriotic cysts.

Evaluation of factor V Leiden, prothrombin and methylenetetrahydrofolate reductase gene mutations in patients with severe pregnancy complications in northern Finland.

BACKGROUND: Thrombosis in placenta may lead to severe pregnancy complications. Most important inherited thrombophilias are factor V Leiden mutation, prothrombin mutation, and methylenetetrahydrofolate reductase mutation. The aim of our research was to evaluate the prevalence of inherited thrombophilias in severe pregnancy complications and in normal pregnancies. MATERIAL AND METHODS: The study subjects with severe preeclampsia, intrauterine growth restriction, placental abruption or fetal death were collected during the period 1999-2004 from Oulu University Hospital. We also collected during the same period voluntary parturients with normal pregnancy outcome as the control group. FVL, FII, and MTHFR gene mutations of the patients and controls were analyzed. RESULTS: We found a significant difference in the prevalence of FVL mutation between the groups. There were 9.5% FVL mutations in the study group compared to 1.8% in the control group; the observed difference between prevalences was 7.7% (95% CI 2.0-13.4). No statistical difference was found in the FII or MTHFR mutations between the groups. All FV and FII mutations were heterozygous and all the MTHFR mutations homozygous. CONCLUSION: Women with thrombophilia have a risk for severe pregnancy complications. Randomized controlled trials are needed to assess the influence of low-molecular-weight heparin in pregnant women with thrombophilia.

A multicentre study investigating subcutaneous etonogestrel implants with injectable testosterone decanoate as a potential long-acting male contraceptive.

BACKGROUND: The combination of etonogestrel implants with injectable testosterone decanoate was investigated as a potential male contraceptive. METHODS: One hundred and thirty subjects were randomly assigned to three treatment groups, all receiving two etonogestrel rods (204 mg etonogestrel) and 400 mg testosterone decanoate either every 4 weeks (group I, n = 42), or every 6 weeks (group II, n = 51) or 600 mg testosterone decanoate every 6 weeks (group III, n = 37) for a treatment period of 48 weeks. RESULTS: One hundred and ten men completed 48 weeks of treatment. Sperm concentrations of <1 x 10(6)/ml were achieved in 90% (group I), 82% (group II) and 89% (group III) of subjects by week 24. Suppression was slower in group II, which also demonstrated more frequent escape from gonadotrophin suppression than groups I and III. Peak testosterone concentrations remained in the normal range throughout in all groups. Mean trough testosterone concentrations were initially subphysiological but increased into the normal range during treatment. Mean haemoglobin levels increased in group I, and a non-significant increase in weight and decline in high-density lipoprotein cholesterol was observed in all groups. Fourteen subjects discontinued treatment due to adverse events. CONCLUSIONS: Subcutaneous etonogestrel implants in combination with injectable testosterone decanoate resulted in profound suppression of spermatogenesis that could be maintained for up to 1 year. Efficacy of suppression was less in group II, probably due to inadequate testosterone dosage. This combination has potential as a long-acting male hormonal contraceptive.

Maternal serum beta-HCG and alpha-fetoprotein concentrations in singleton pregnancies following assisted reproduction.

BACKGROUND: The reason for the elevated levels of HCG in assisted reproduction pregnancies remains unknown. Our hypothesis was that this increase is caused by the ovarian superovulation therapy. METHODS: We compared the beta-HCG and alpha-fetoprotein (AFP) multiples of the median (MoM) in singleton pregnancies after IVF or ICSI with those achieved by frozen embryo transfer (FET) in spontaneous cycles. RESULTS: The HCG and AFP MoMs (plus minus SEMs) of 59 FET pregnancies were compared with 144 IVF (including 48 ICSI) pregnancies. The maternal HCG of pregnancies following ovarian stimulation was 1.31 plus minus 0.08 MoM compared with 1.35 plus minus 0.12 MoM in the unstimulated ones. The values for AFP were 1.06 plus minus 0.05 versus 1.11 plus minus 0.05 respectively. No significant differences could be observed between pregnancies following stimulated IVF/ICSI and unstimulated FET cycles. CONCLUSIONS: Our results show that second trimester maternal serum HCG is also elevated in singleton pregnancies following spontaneous FET cycles. The increased maternal serum HCG in IVF pregnancies is thus not related to superovulation therapy. Because of the elevated maternal serum HCG levels, serum screening cannot be performed reliably in pregnancies following assisted reproduction technology. Ultrasonographic detection of the nuchal translucency is unaffected and should be used for this group of women undergoing assisted reproduction.

Evaluation of first trimester maternal serum and ultrasound screening for Down's syndrome in Eastern and Northern Finland.

The current trend in prenatal diagnosis is that trisomy screening is being moved to the first trimester and ultrasonographic nuchal translucency measurement is included in risk calculation. It is likely that biochemical screening in the second trimester will gradually be given up. In Eastern and Northern Finland, during the year 1999 we offered first-trimester ultrasonographic and serum screening for trisomy 21, with measurements of maternal serum PAPP-A and beta-hCG. A total of 2515 pregnant women participated in the screening, yielding the detection of eight foetuses with Down's syndrome. Six affected foetuses (75%) were detected by means of first-trimester serum screening. Since we were in the phase of collecting data for the Finnish medians for PAPP-A and beta-hCG, the women were not given the estimates of risk for trisomy 21. Only 1602 of the 2515 enrolled women had the combination of first-trimester ultrasonographic and serum screening performed, and in that group there were five foetuses with Down's syndrome. The combination ultrasonographic and serum approach yielded a Down's syndrome detection rate of 80% (four out of five) with a 5% false positive rate, whereas in nuchal translucency based-screening the detection rate was 60%, with a 5% false positive rate.

Enhanced sensitivity to steroid-negative feedback during breast-feeding: low-dose estradiol (transdermal estradiol supplementation) suppresses gonadotropins and ovarian activity assessed by inhibin B.

Breast-feeding reduces fertility, and this seems to be related, in part, to an enhancement of the sensitivity of the GnRH system to the negative feedback effects of estradiol related to suckling. Previously, we showed that short-term treatment with small doses of estradiol delivered transdermally suppress plasma gonadotropin concentrations in breast-feeding women. We have now monitored the effects on ovarian function of longer-term low-dose estradiol treatment using plasma inhibin B and inhibin A concentrations and ultrasonography. Breast-feeding women (n = 45) using barrier methods of contraception were enrolled at 6 weeks postpartum and followed up to 18 weeks PP. Nineteen women agreed to being randomized to wear either an estrogen [transdermal estradiol supplementation (TES); n = 10; Estraderm, 50 microg/24 h] or a placebo (PL; n = 9) patch for 12 weeks, whereas the remaining 26 women acted as untreated controls. TES did not significantly increase plasma estradiol concentrations. Plasma FSH levels decreased from 6.1+/-0.8 U/L to 3.3+/-0.6 U/L after 2 weeks of treatment (P < 0.01) and were lower in the TES group compared with the PL group at all times during the treatment (at least P < 0.05). Plasma LH concentrations in the TES group were lower than in the PL group after 4, 6, 8, and 10 weeks of estrogen treatment (at least P < 0.05). Throughout the study, no ovarian follicles detected by ultrasound were greater than 10 mm in diameter. Nevertheless, after 2 weeks of treatment, plasma inhibin B concentrations were significantly lower in the TES group than in the PL group (15.5+/-5.8 vs. 64.9+/-11.1 ng/L; P < 0.01) and remained significantly (P < 0.01) suppressed throughout the treatment, suggesting a suppression of the functional ovarian activity during TES. Inhibin A levels remained low in all groups (3-45 ng/L) but were suppressed further by TES treatment with no levels greater than 7 ng/L. We conclude that low-dose estradiol treatment given as TES suppresses ovarian activity as measured by inhibins B and A by reducing the secretion of LH and FSH during breast-feeding for several weeks. This supports the concept that suckling-induced suppression of the GnRH system is associated with an enhancement of the negative effects of estradiol on the hypothalamic GnRH system. Furthermore, because the contraceptive efficacy of breast-feeding is complicated by the unpredictable early return of ovarian activity in some women, TES could be the basis for the development of a novel contraceptive for breast-feeding women.

Biphasic increase in serum inhibin B during puberty: a longitudinal study of healthy Finnish boys.

We investigated the concentrations of serum inhibin B and the pro-alphaC-containing inhibins (pro-alphaC inhibin) and their relations to serum FSH levels in 38 healthy boys during their progression through puberty. Furthermore, we studied the effect of recombinant human FSH (rhFSH) on pro-alphaC inhibin production in three prepubertal gonadotropin-deficient boys. The serum inhibin B level increased between Tanner stages G1 and G2 (p < 0.02), simultaneously with the serum LH and testosterone concentrations. In advanced puberty, inhibin B changed less, and at stage G4, correlated negatively with serum FSH level (r = -0.57, p < 0.001, n = 37). The serum pro-alphaC inhibin level changed differently and increased also in advanced puberty. In prepubertal healthy subjects, the serum pro-alphaC inhibin and FSH levels correlated positively (r = 0.50, p = 0.051, n = 16), and during the rhFSH treatment of gonadotropin-deficient boys, serum pro-alphaC inhibin levels increased. These findings suggest that an increased production of inhibin B is an early event in puberty and that inhibin B can inhibit pituitary FSH secretion in the course of sexual maturation. Furthermore, they suggest that during puberty free inhibin (pro)-alpha-subunits, detected by the pro-alphaC inhibin assay, are present in large amounts and that FSH stimulates their production by prepubertal testes.

Modulation of gonadotropin secretion at the pituitary level by testosterone in gonadotropin-releasing hormone-treated male rats during food deprivation.

Testosterone (T) inhibits the synthesis and secretion of FSH and LH by decreasing the secretion of GnRH from the hypothalamus. However, T is also able to stimulate FSH gene expression and synthesis at the pituitary level when the release or action of GnRH is blocked. In the present study, we analyzed whether the positive effect of T on pituitary FSH could also be brought about during food restriction, which represents a model of suppressed GnRH secretion. We also wanted to learn whether this positive effect could be detected if GnRH pulsatility is maintained by exogenous injections. Adult male rats were subjected to various combinations of the following treatments: 1) implantation of silastic capsules containing T for Days 0-4 of the experiment, 2) starvation for Days 1-4 of the experiment, and 3) GnRH-treatment at 2-h intervals (500 ng/kg BW) for Days 3-4. The combined treatments were as follows: 1) control, 2) only starvation, 3) only GnRH, 4) starvation+GnRH, 5) only T, 6) starvation+T, 7) GnRH+T, and 8) starvation+GnRH+T (n = 12/group; two independent experiments). Serum FSH level was decreased 20% by starvation (p < 0.01), but no decrease was observed when the starving animals were treated with T. GnRH treatment increased serum FSH in both ad libitum-fed and starving animals to 266% and 333% of the respective control values (both p < 0.01). When T was added to these treatments, the increases in serum FSH were smaller, 219% and 272%, respectively (p < 0.01 vs. respective groups without T).(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of epitestosterone on gonadotrophin synthesis and secretion.

The effects of 3-week treatment with increasing doses of epitestosterone (ET) on gonadotrophin gene expression and secretion, on testosterone and 5 alpha-dihydrotestosterone (DHT) levels, and on the weight of testes and prostates, were studied in intact adult male rats. The hormones were delivered by means of silastic capsules of different lengths filled with the steroid. One group of rats received testosterone (T) instead of ET, to compare the results with previous studies concerning the testosterone effect. The controls were given capsules with glucose only. Treatment with ET, as well as with T, significantly reduced the weights of prostates. When the data from ET-treated rats and controls were combined, a significant negative correlation (P < 0.001) was found between the weight of prostates and serum ET. T, in contrast to ET, also decreased significantly the weights of testes, ET treatment caused a significant reduction of serum T levels but only an insignificant decline of DHT levels, independent of the dose. Serum and pituitary (p) luteinizing hormone (LH) levels in the ET-treated rats did not change. Pituitary mRNA contents for the beta LH subunit (beta LH-mRNA) showed a dose-dependent significant increase, up to 170% (P < 0.01), with ET treatment. pFSH decreased with the lowest ET (2 cm) dose (P < 0.05), but no change was observed with the other doses. The mRNA for the common alpha-subunit also increased with the ET load. In conclusion, ET acts at several sites in the regulation of gonadotrophin formation and release. It enhances the steady-state mRNA levels of both gonadotrophins in the pituitary.(ABSTRACT TRUNCATED AT 250 WORDS)

Biphasic effect of exogenous testosterone on follicle-stimulating hormone gene expression and synthesis in the male rat.

Effects of 2-week treatments with increasing doses of testosterone (T) on gonadotropin gene expression and secretion were studied in intact and acutely castrated male rats. T was administered in silastic capsules with lengths of 2, 4, 8 or 16 cm, and control animals received empty capsules (eight per treatment). The treatments increased serum T up to 3-fold of control levels. In intact animals, the 2-8 cm capsules suppressed pituitary follicle-stimulating hormone-beta (FSH beta) mRNA contents by 40-50% (p < 0.01), but 16 cm of T returned the levels back to control range. Castration alone increased the FSH beta mRNA level 2.3-fold (p < 0.01) and, after T treatment, the FSH beta message returned to control levels indistinguishable from intact controls but higher than in intact animals receiving the same T dose. Pituitary luteinizing hormone-beta (LH beta) mRNA displayed a dose-dependent suppression in response to T, to 32-35% of controls (p < 0.01) with the 8 and 16 cm capsules. Castration increased this message 10-fold, and additional T treatment suppressed the levels to the range of T-treated intact animals. Pituitary common-alpha mRNA decreased to 30-31% of controls by 2, 4 and 8 cm of T (p < 0.01), but the highest dose of T increased the common-alpha contents, in comparison to the other doses, to 54% of controls (p < 0.01). Castration alone increased the common-alpha contents 4.4-fold, and there was a dose-dependent suppression of this parameter by T down to the range of T-treated intact rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Differential regulation of FSH and inhibin gene expression and synthesis by testosterone in immature and mature male rats.

Direct effects of testosterone on gonadotrophins at the pituitary level were studied in intact and castrated immature (age 10 days) and mature (70 days) male rats. Gonadotrophin-releasing hormone action was blocked by treatment with a potent GnRH antagonist, Ac-D-pClPhe-D-pClPhe-D-Trp-Ser-Tyr-D-Arg-Leu-Arg-Pro-D-Ala-+ ++NH2CH3COOH (Ant; Organon 30276; 1.0 mg/kg body weight per day) injected subcutaneously. Silicone elastomer capsules were used for the testosterone treatment. Both treatments commenced on the day of orchiectomy and lasted for 7 days. In adult male rats Ant treatment suppressed serum testosterone from 9.5 +/- 2.5 (S.E.M.) nmol/l to below the limit of detection (< 0.10 nmol/l; P < 0.01), and the testosterone implants reversed the decrease. Treatment with Ant decreased the pituitary content of FSH-beta subunit mRNA in intact and orchiectomized rats to 14% of their respective controls (P < 0.01). These levels were increased to 80-81% of controls (not significant) in both groups by combined treatment with testosterone and Ant. Orchiectomy alone increased FSH-beta subunit mRNA by 202% (P < 0.01). In intact immature rats Ant treatment decreased the level of pituitary FSH-beta subunit mRNA to 21% (P < 0.01), and a partial recovery (P < 0.01) to 42% of controls was observed with combined Ant+testosterone treatment. In contrast, in orchiectomized immature rats, where ANT decreased FSH-beta subunit levels to 48% of controls (P < 0.01), testosterone was able to reverse these mRNA levels completely (114% of controls). No evidence for the direct pituitary effects of testosterone were found in the mRNA of the common alpha or LH-beta subunits. In adult rats, the testicular inhibin alpha and beta A subunit mRNA levels were increased (P < 0.01) by Ant+testosterone compared with Ant-treated animals, but there were no differences in serum immunoreactive inhibin between any of the uncastrated adult groups. In intact immature rats, Ant+testosterone treatment increased (P < 0.01) inhibin beta A subunit mRNA levels compared with controls and Ant-treated animals. Ant decreased the level fo peripheral inhibin immunoreactivity from 8.3 +/- 2.0 U/ml to 2.1 +/- 0.4 U/ml (P < 0.01) and testosterone reversed it to 5.8 +/- 0.6 U/ml (not significant). In conclusion, our observations indicated that testosterone is able to stimulate FSH gene expression and secretion directly in immature and adult rats, but the testosterone response is enhanced at both ages by orchiectomy, even more so in the immature rat.(ABSTRACT TRUNCATED AT 400 WORDS)

A supersensitive immunofluorometric assay for rat luteinizing hormone.

We have developed an immunofluorometric assay (IFMA) for rat (r) LH, which is based on two monoclonal antibodies, one to bovine and the other to human LH. Signal detection occurs by time-resolved fluorescence evoked by a europium label (Delfia, Wallac). The method is fast in comparison to the standard RIA with the NIDDK reagents (4 h vs. 3 days). The sensitivity of the IFMA assay (0.75 pg/tube; NIDDK rLH RP-2) is over 30-fold higher than that of the NIDDK RIA (usual detection limit, 20-30 pg/tube). Using 25-microliters serum samples, the sensitivity of IFMA is 0.03 micrograms/liter; with 100-microliters samples, it is 0.0075 micrograms/liter. The cross-reactivity of the IFMA assay is 0.3% with rFSH, 3% with rTSH, and less than 0.05% with rGH, rPRL, and the rat alpha-subunit. A linear correlation between IFMA and RIA values is seen at serum levels above 0.4 micrograms/liter. Below this level, only IFMA is able to detect concentration differences between samples. In practice, this means that only IFMA is able to provide meaningful measurements of suppressed levels of serum LH. The correlation coefficient between IFMA and the mouse interstitial cell in vitro bioassay for LH in randomly selected rat pituitary homogenates was 0.93 (n = 47). The serum concentration of LH determined by IFMA is 0.57 +/- 0.10 micrograms/liter in intact adult male rats (mean +/- SEM; n = 12) and 0.41 +/- 0.10 micrograms/liter (n = 10) in randomly cycling females. The level in hypophysectomized rat serum is 0.035 +/- 0.0033 micrograms/liter (n = 8), if the limit of sensitivity (0.03 microgram/liter) is assigned to unmeasurable levels. One-week treatment of male rats with 2-cm Silastic implants containing testosterone suppressed serum LH, measured by IFMA, from 0.56 +/- 0.057 to 0.086 +/- 0.057 micrograms/liter (P < 0.01). The suppression of LH measured in the same samples by RIA was lower, from 0.73 +/- 0.057 to 0.44 +/- 0.048 micrograms/liter (P < 0.01). A 5-day starvation of intact male rats suppressed serum LH from 0.57 +/- 0.10 to 0.30 +/- 0.05 microgram/liter by IFMA (P < 0.01), whereas the decrease determined by RIA was not significant (0.80 +/- 0.07 vs. 0.66 +/- 0.13 micrograms/liter).(ABSTRACT TRUNCATED AT 400 WORDS)

Starvation-induced suppression of pituitary-testicular function in rats is reversed by pulsatile gonadotropin-releasing hormone substitution.

This study was carried out to test the hypothesis that reduced hypothalamic GnRH release is responsible for the suppression of reproductive functions during starvation. Adult male rats were kept for 4 days under total fasting (only water allowed) and injected during this time at 2-h intervals with 100 or 500 ng/kg BW of GnRH or vehicle. Serum levels of LH and FSH decreased by 30% during starvation (p less than 0.05), and these effects were fully reversed by either dose of GnRH treatment. Starvation reduced the pituitary mRNA contents of the gonadotropin common alpha- and FSH beta-subunits by 30% and 35% in starved animals (p less than 0.05 for both), but the LH beta-subunit mRNA was unaffected. The GnRH treatments partly or totally reversed these changes, but up-regulation of the mRNA levels by GnRH was seen only in controls fed ad libitum. Starvation reduced the testicular and serum levels of testosterone by 84% (p less than 0.01) and 42% (p less than 0.05), respectively. These changes were fully reversed by the 500-ng/kg dose of GnRH treatment during fasting, but only serum T was completely reversed by the 100-ng/kg GnRH treatment. To elucidate whether fasting per se had direct effects at the gonadal level, we blocked the secretion of gonadotropins by treatment with a GnRH antagonist, and replaced the gonadotropins by injecting of hCG (10 IU/kg BW once daily) and hFSH (75 IU/kg BW once daily). No differences were observed between starved and control animals in either testicular or serum levels of T, or in accessory sex gland weights.(ABSTRACT TRUNCATED AT 250 WORDS)