Daily levels of sex hormones in 15 subfertile women formulate a menstrual cycle profile predominant with progesterone secretion.

INTRODUCTION: Changes in sex hormone secretions during the menstrual cycle may affect fertility. It has been shown that a prematurely raised progesterone (P4) level after therapeutic injection of human chorionic gonadotropin caused changes in endometrial gene expression and lowered the pregnancy rate. The aim of the present study was to investigate the complete menstrual patterns of P4 together with its derivatives testosterone (T) and oestradiol (E2) in subfertile women during their natural cycles. MATERIAL AND METHODS: Daily serum levels of P4 (ng/mL), T (ng/mL), E2 (pg/mL), and sex hormone binding protein (SHBG, nmol/L) were measured throughout a single 23-28-day menstrual cycle in 15 subfertile women aged 28-40 years with patent oviducts and normospermic partners. Knowing SHBG levels, the free androgen (FAI) and free oestrogen (FEI) indexes were calculated for each cycle day in each patient. RESULTS: Baseline (cycle day one) levels of luteinising hormone (LH), thyroid stimulating hormone (TSH), P4, and T were comparable with reference intervals for a normal cycle, whereas follicle stimulating hormone (FSH), E2, and SHBG exceeded those. During cycles, the levels of P4 correlated positively with E2 levels (r = 0.38, p < 0.05, n = 392) an negatively with T (r = -0.13, p < 0.05, n = 391). T correlated negatively with E2 (r = -0.19, p < 0.05, n = 391). Menstrual cycle phases were hidden. The curve of the mean/median daily levels of P4 rose prematurely, was parallel with the E2 rise, and culminated closely, but with more than 4 times greater amplitude of P4 (2571% of baseline levels in day 16) than of E2 (580% in day 14). In turn, the curve of T declined in a U-shaped manner with a nadir (-27%) on day 16. Averaged daily levels of FEI, but not FAI, varied significantly between 23 and 26 days long and the 27-28-day cycles. CONCLUSIONS: 1. Throughout the entire menstrual cycle length in subfertile women, P4 secretion predominates quantitatively over secretions of the remaining sex hormones when menstrual cycle phases are hidden. 2. The rise of E2 secretion is in parallel with the P4 rise, but with 4 times lower amplitude of E2. 3. T secretion declines and is inversely related to both P4 and E2 secretions. 4. Changes in E2 bioavailability are related to menstrual cycle length.

Expression of G-Protein-Coupled Estrogen Receptor (GPER) in Whole Testicular Tissue and Laser-Capture Microdissected Testicular Compartments of Men with Normal and Aberrant Spermatogenesis.

In this study, we retrospectively investigated GPER expression in biopsies of azoospermic men with complete (obstructive azoospermia-OA) and aberrant spermatogenesis (nonobstructive azoospermia-NOA). Each biopsy was histologically evaluated with morphometry. The testicular GPER expression was analyzed by the immunohistochemistry and RT-PCR technique in the whole testicular tissue and in seminiferous tubules and Leydig cells after laser-capture microdissection. In laser-microdissected compartments, we also analyzed transcriptional expression of selected Leydig (CYP17A1, HSD17B3, StAR) and Sertoli cell (AMH, SCF, BMP4) function markers. Immunohistochemical staining revealed expression of GPER in the cytoplasm of Leydig and Sertoli cells. Its stronger intensity was observed in Sertoli cells of NOA biopsies. The RT-PCR analysis of the GPER mRNA level unequivocally showed its increased expression in seminiferous tubules (i.e., Sertoli cells), not Leydig cells in NOA biopsies. This increased expression correlated positively with the transcriptional level of AMH-a marker of Sertoli cell immaturity, as well as FSH serum level in NOA but not in the OA group. Our results clearly demonstrate altered GPER expression in testes with primary spermatogenic impairment that might be related to Sertoli cell maturity/function.

Changes in prevalence of obesity and high waist circumference over four years across European regions: the European male ageing study (EMAS).

Diversity in lifestyles and socioeconomic status among European populations, and recent socio-political and economic changes in transitional countries, may affect changes in adiposity. We aimed to determine whether change in the prevalence of obesity varies between the socio-politically transitional North-East European (Lódz, Poland; Szeged, Hungary; Tartu, Estonia), and the non-transitional Mediterranean (Santiago de Compostela, Spain; Florence, Italy) and North-West European (Leuven, Belgium; Malmö, Sweden; Manchester, UK) cities. This prospective observational cohort survey was performed between 2003 and 2005 at baseline and followed up between 2008 and 2010 of 3369 community-dwelling men aged 40-79 years. Main outcome measures in the present paper included waist circumference, body mass index and mid-upper arm muscle area. Baseline prevalence of waist circumference ≥ 102 cm and body mass index ≥ 30 kg/m2, respectively, were 39.0, 29.5 % in North-East European cities, 32.4, 21.9 % in Mediterranean cities, and 30.0, 20.1 % in North-West European cities. After median 4.3 years, men living in cities from transitional countries had mean gains in waist circumference (1.1 cm) and body mass index (0.2 kg/m2), which were greater than men in cities from non-transitional countries (P = 0.005). North-East European cities had greater gains in waist circumference (1.5 cm) than in Mediterranean cities (P < 0.001). Over 4.3 years, the prevalence of waist circumference ≥ 102 cm had increased by 13.1 % in North-East European cities, 5.8 % in the Mediterranean cities, 10.0 % in North-West European cities. Odds ratios (95 % confidence intervals), adjusted for lifestyle factors, for developing waist circumference ≥ 102 cm, compared with men from Mediterranean cities, were 2.3 (1.5-3.5) in North-East European cities and 1.6 (1.1-2.4) in North-West European cities, and 1.6 (1.2-2.1) in men living in cities from transitional, compared with cities from non-transitional countries. These regional differences in increased prevalence of waist circumference ≥ 102 cm were more pronounced in men aged 60-79 years than in those aged 40-59 years. Overall there was an increase in the prevalence of obesity (body mass index ≥ 30 kg/m2) over 4.3 years (between 5.3 and 6.1 %) with no significant regional differences at any age. Mid-upper arm muscle area declined during follow-up with the greatest decline among men from North-East European cities. In conclusion, increasing waist circumference is dissociated from change in body mass index and most rapid among men living in cities from transitional North-East European countries, presumably driven by economic and socio-political changes. Information on women would also be of value and it would be of interest to relate the changes in adiposity to dietary and other behavioural habits.

Sperm DNA Fragmentation Index and Hyaluronan Binding Ability in Men from Infertile Couples and Men with Testicular Germ Cell Tumor.

Objective. To investigate sperm DNA fragmentation and sperm functional maturity in men from infertile couples (IC) and men with testicular germ cell tumor (TGCT). Materials and Methods. Semen samples were collected from 312 IC men and 23 men with TGCT before unilateral orchiectomy and oncological treatment. The sperm chromatin dispersion test was performed to determine DNA fragmentation index (DFI) and the ability of sperm to bind with hyaluronan (HA) was assessed. Results. In comparison with the IC men, the men with TGCT had a higher percentage of sperm with fragmented DNA (median 28% versus 21%; p < 0.01) and a lower percentage of HA-bound sperm (24% versus 66%; p < 0.001). Normal results of both analyses were observed in 24% of IC men and 4% of men with TGCT. Negative Spearman's correlations were found between DFI and the percentage of HA-bound sperm in the whole group and in IC subjects and those with TGCT analyzed separately. Conclusions. Approximately 76% of IC men and 96% with TGCT awaiting orchiectomy demonstrated DNA fragmentation and/or sperm immaturity. We therefore recommend sperm banking after unilateral orchiectomy, but before irradiation and chemotherapy; the use of such a deposit appears to be a better strategy to obtain functionally efficient sperms.

Low vitamin D and the risk of developing chronic widespread pain: results from the European male ageing study.

BACKGROUND: The association between low levels of vitamin D and the occurrence of chronic widespread pain (CWP) remains unclear. The aim of our analysis was to determine the relationship between low vitamin D levels and the risk of developing CWP in a population sample of middle age and elderly men. METHODS: Three thousand three hundred sixty nine men aged 40-79 were recruited from 8 European centres for a longitudinal study of male ageing, the European Male Ageing Study. At baseline participants underwent assessment of lifestyle, health factors, physical characteristics and gave a fasting blood sample. The occurrence of pain was assessed at baseline and follow up (a mean of 4.3 years later) by shading painful sites on a body manikin. The presence of CWP was determined using the ACR criteria for fibromyalgia. Serum 25-hydroxyvitamin D (25-(OH) D) was assessed by radioimmunoassay. Logistic regression was used to determine the relationship between baseline vitamin D levels and the new occurrence of CWP. RESULTS: Two thousand three hundred thirteen men, mean age 58.8 years (SD = 10.6), had complete pain and vitamin data available and contributed to this analysis. 151 (6.5%) developed new CWP at follow up and 577 (24.9%) were pain free at both time points, the comparator group. After adjustment for age and centre, physical performance and number of comorbidities, compared to those in upper quintile of 25-(OH) D ( ≥36.3 ng/mL), those in the lowest quintile (<15.6 ng/mL) were more likely to develop CWP (Odds Ratio [OR] = 1.93; 95% CI = 1.0-3.6). Further adjustment for BMI (OR = 1.67; 95% CI = 0.93-3.02) or depression (OR = 1.77; 95% CI = 0.98-3.21), however rendered the association non-significant. CONCLUSIONS: Low vitamin D is linked with the new occurrence of CWP, although this may be explained by underlying adverse health factors, particularly obesity and depression.

Chronic widespread pain is associated with worsening frailty in European men.

BACKGROUND: we hypothesised that chronic widespread pain (CWP), by acting as a potential stressor, may predispose to the development of, or worsening, frailty. SETTING: longitudinal analysis within the European Male Ageing Study (EMAS). PARTICIPANTS: a total of 2,736 community-dwelling men aged 40-79. METHODS: subjects completed a pain questionnaire and shaded a manikin, with the presence of CWP defined using the American College of Rheumatology criteria. Physical activity, smoking, alcohol consumption and depression were measured. Repeat assessments took place a median of 4.3 years later. A frailty index (FI) was used, with frail defined as an FI >0.35. The association between CWP at baseline and the new occurrence of frailty was examined using logistic regression; the association between CWP at baseline and change in FI was examined using negative binomial regression. RESULTS: at baseline, 218 (8.3%) men reported CWP. Of the 2,631 men who were defined as non-frail at baseline, 112 (4.3%) were frail at follow-up; their mean FI was 0.12 (SD 0.1) at baseline and 0.15 (SD 0.1) at follow-up, with a mean change of 0.03 (SD 0.08) P ≤ 0.001. Among men who were non-frail at baseline, those with CWP were significantly more likely to develop frailty. After adjustment for age and centre, compared with those with no pain, those with CWP at baseline had a 70% higher FI at follow-up; these associations remained significant after further adjustment for smoking, body mass index, depression, physical activity and FI at baseline. CONCLUSION: the presence of CWP is associated with an increased risk of frailty in older European men.

The risk of neoplasm associated with dysgenetic testes in prepubertal and pubertal/adult patients.

INTRODUCTION: In patients with Y-chromosome in the karyotype, partial gonadal dysgenesis and disorders of male reproductive sex organs development are usually resected in childhood because of the high risk of germ cell tumours (GCT). In patients with Y-chromosome, complete gonadal dysgenesis and female genitalia gonadectomy is performed markedly later. However, due to the relatively low number of adult patients with preserved dysgenetic gonads, the true risk of neoplasm is unknown. The aim of the study was to evaluate the prevalence of neoplasia in dysgenetic gonads of children and adults with Y-chromosome in a retrospective study. MATERIAL AND METHODS: A review of medical documentation of 94 patients with disorders of sex development (DSD), Y-chromosome and gonadal dysgenesis (GD), aged 1.2-32 years (47 prepubertal, 1.2-10 years; 47 pubertal/adult, 13-32 years), was conducted. Serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH) and testosterone were determined. Bilateral gonadectomy was performed in 73.4% of patients, and unilateral gonadectomy with biopsy of the contralateral gonad in 26.4%. All gonadal tissues were subjected to immunohistochemical evaluation with antibodies against PLAP and OCT3/4 (markers of malignant germ cells, but also foetal multipotent germ cells), while gonads of prepubertal patients were examined by c-KIT, as well. RESULTS: Streak gonads were identified on both sides (complete GD) in 30.8%, a streak gonad on one side and an underdeveloped testis on the other (asymmetric GD) in 38.3%, and underdeveloped testicular structure on both sides (partial GD) in 30.8% of cases. Germ cell neoplasia was found in 53.2% of patients (51.1% in children, 55.3% in pubertal/adults). Invasive GCT were identified in 11.7% of cases, of which 90.9% were in pubertal/adult patients. Other neoplastic lesions included gonadoblastoma (16% prevalence) and testicular carcinoma in situ (25.5%). In younger patients FSH serum levels were increased in 81% of cases (mean 2.82 ± 2.18 IU/L), while LH in 58% (mean 1.82 ± 1.69 IU/L). Hypergonadotropic hypogonadism was diagnosed in most of the pubertal/ /adult patients (mean FSH 54.2 ± 23.3 IU/L, mean LH 21.7 ± 12.1 IU/L, mean testosterone 5.5 ± 4.5 nmol/L). CONCLUSIONS: Dysgenetic gonads in patients with Y chromosome have a high risk of germ cell neoplasia (ca. 50%). If they are preserved until puberty/early adulthood, they may develop overt, invasive GCT. The gonads also have poor hormonal activity (hypergonadotropic hypogonadism) in most of the pubertal/adult patients. Each of these cases must be considered individually and a decision to remove the gonad or not should be based on the comprehensive analysis of the phenotype by a multidisciplinary team of specialists in consultation with the patient and the parents. If dysgenetic gonads are not resected in childhood, these patients need careful ongoing follow-up examination, including biopsy and histopathological evaluation.

Low heel ultrasound parameters predict mortality in men: results from the European Male Ageing Study (EMAS).

BACKGROUND: low bone mineral density measured by dual-energy x-ray absorptiometry is associated with increased mortality. The relationship between other skeletal phenotypes and mortality is unclear. The aim of this study was to determine the relationship between quantitative heel ultrasound parameters and mortality in a cohort of European men. METHODS: men aged 40-79 years were recruited for participation in a prospective study of male ageing: the European Male Ageing Study (EMAS). At baseline, subjects attended for quantitative ultrasound (QUS) of the heel (Hologic-SAHARA) and completed questionnaires on lifestyle factors and co-morbidities. Height and weight were measured. After a median of 4.3 years, subjects were invited to attend a follow-up assessment, and reasons for non-participation, including death, were recorded. The relationship between QUS parameters (broadband ultrasound attenuation [BUA] and speed of sound [SOS]) and mortality was assessed using Cox proportional hazards model. RESULTS: from a total of 3,244 men (mean age 59.8, standard deviation [SD] 10.8 years), 185 (5.7%) died during the follow-up period. After adjusting for age, centre, body mass index, physical activity, current smoking, number of co-morbidities and general health, each SD decrease in BUA was associated with a 20% higher risk of mortality (hazard ratio [HR] per SD = 1.2; 95% confidence interval [CI] = 1.0-1.4). Compared with those in higher quintiles (2nd-5th), those in the lowest quintile of BUA and SOS had a greater mortality risk (BUA: HR = 1.6; 95% CI = 1.1-2.3 and SOS: HR = 1.6; 95% CI = 1.2-2.2). CONCLUSION: lower heel ultrasound parameters are associated with increased mortality in European men.

Autocrine androgen action is essential for Leydig cell maturation and function, and protects against late-onset Leydig cell apoptosis in both mice and men.

Leydig cell number and function decline as men age, and low testosterone is associated with all "Western" cardio-metabolic disorders. However, whether perturbed androgen action within the adult Leydig cell lineage predisposes individuals to this late-onset degeneration remains unknown. To address this, we generated a novel mouse model in which androgen receptor (AR) is ablated from ∼75% of adult Leydig stem cell/cell progenitors, from fetal life onward (Leydig cell AR knockout mice), permitting interrogation of the specific roles of autocrine Leydig cell AR signaling through comparison to adjacent AR-retaining Leydig cells, testes from littermate controls, and to human testes, including from patients with complete androgen insensitivity syndrome (CAIS). This revealed that autocrine AR signaling is dispensable for the attainment of final Leydig cell number but is essential for Leydig cell maturation and regulation of steroidogenic enzymes in adulthood. Furthermore, these studies reveal that autocrine AR signaling in Leydig cells protects against late-onset degeneration of the seminiferous epithelium in mice and inhibits Leydig cell apoptosis in both adult mice and patients with CAIS, possibly via opposing aberrant estrogen signaling. We conclude that autocrine androgen action within Leydig cells is essential for the lifelong support of spermatogenesis and the development and lifelong health of Leydig cells.

Free and bioavailable fractions of sex steroids may influence bones in young men, depending on age and oestradiol level.

INTRODUCTION: Longitudinal bone growth ceases by the end of puberty, and it is thought to be a result, in both sexes, of increased pubertal oestrogen serum concentrations. Since peak bone mass is achieved by the third decade of life or later, the aim of this study was to relate sex steroid hormones and sex hormone binding globulin (SHBG) levels to bone quality in men during their third and fourth decades of life. MATERIAL AND METHODS: Eighty men, healthy volunteers aged between 18 and 39 years, were subjected to an interviewer-administered questionnaire, body mass index (BMI) measurement, blood sample and calcaneal quantitative ultrasound (QUS) (Hologic-SAHARA). Blood was assessed for testosterone (T), oestradiol (E2), dehydroepiandrosterone sulfate (DHEAS), SHBG, luteinising hormone (LH) and follicle stimulating hormone (FSH). Free and bioavailable T and E2 levels were calculated knowing SHBG and albumin levels. RESULTS: While T, E2, DHEAS, LH and FSH levels were not related, free and bioavailable fractions of T and E2 were positively associated with QUS readings. SHBG level was associated negatively. After dichotomisation for age, the associations remained significant only for younger subjects (18-30 years, n = 47). After adjustment for other co-variants, only SHBG in younger subjects retained its negative association with QUS. Older subjects (31-39 years, n = 33) revealed higher BMI and lower serum concentrations of total (-17 %), free (-18.5%) and bioavailable (-22.5%) levels of E2 than younger subjects. CONCLUSION: Free and bioavailable fractions of sex steroids may influence bones in young men, depending on age and E2 level.

Androgen receptor polymorphism-dependent variation in prostate-specific antigen concentrations of European men.

BACKGROUND: Androgens acting via the androgen receptor (AR) stimulate production of PSA, which is a clinical marker of prostate cancer. Because genetic variants in the AR may have a significant impact on the risk of being diagnosed with prostate cancer, the aim was to investigate whether AR variants were associated with the risk of having PSA above clinically used cutoff thresholds of 3 or 4 ng/mL in men without prostate cancer. METHODS: Men without prostate cancer history (n = 1,744) were selected from the European Male Ageing Study cohort of 40 to 79-year-old men from eight different European centers. Using linear and logistic regression models, with age and center as covariates, we investigated whether AR variants (CAG repeat-length and/or SNP genotype) were associated with having serum PSA concentrations above 3 or 4 ng/mL, which often are set as cutoff concentrations for further investigation of prostate cancer. RESULTS: Carriers of the SNP rs1204038 A-allele (16% of the men) were more likely to have PSA>3 and 4 ng/mL (OR; 95% confidence intervals, 1.65; 1.13-2.40 and 1.87; 1.18-2.96, respectively) than G-allele carriers. They also had shorter CAG repeats (median 20 vs. 23, P < 0.0005), but CAG repeat length per se did not affect the PSA concentrations. CONCLUSION: The A-allele of the SNP rs1204038 gives a 65% higher risk of having PSA above 3 ng/mL than the G-allele in men without prostate cancer, and thereby an increased risk of being referred for further examination on suspicion of prostate cancer. IMPACT: Serum PSA as a clinical marker could be improved by adjustment for AR-genotype.

Association of 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D and parathyroid hormone with mortality among middle-aged and older European men.

BACKGROUND: vitamin D deficiency has been associated with an increased risk of mortality, but whether this relationship is causal or linked to co-existent comorbidity and adverse life factors remains uncertain. Our objective was to determine whether endogenous 25-hydroxyvitamin D (25(OH)D), 1,25-dihydroxyvitamin D (1,25(OH)2D) and parathyroid hormone (PTH) levels predicted all-cause, cardiovascular and cancer mortality independently of health and lifestyle factors. SETTING: : prospective cohort analysis within the European Male Ageing Study. PARTICIPANTS: : 2,816 community-dwelling men aged 40-79 years at baseline. METHODS: : Cox regression was used to examine the association of all-cause mortality with 25(OH)D, 1,25(OH)2D and PTH; cardiovascular and cancer mortality were modelled using competing-risks regression. Results were expressed as hazard ratios (HR) and 95% confidence intervals (CIs) for Cox models; sub-hazard ratios (SHR) and 95% CIs for competing-risks models. RESULTS: : a total of 187 men died during a median of 4.3 years of follow-up. Serum levels of 25(OH)D (per 1 SD decrease: HR = 1.45; 95% CI = 1.16, 1.81) and 1,25(OH)2D (per 1 SD decrease: HR = 1.20; 95% CI = 1.00, 1.44) were associated with an increased risk of all-cause mortality after adjusting for age, centre, smoking, self-reported morbidities, physical activity and functional performance. Only levels of 25(OH)D <25 nmol/l predicted cancer mortality (SHR = 3.33; 95% CI = 1.38, 8.04). CONCLUSION: : lower 25(OH)D and 1,25(OH)2D levels independently predicted all-cause mortality in middle-aged and older European men. Associations with cancer mortality were only observed among men with very low levels of 25(OH)D. These associations were only partially explained by the range of adverse health and lifestyle factors measured here.

Estradiol and testosterone inhibit rat seminiferous tubule development in a hormone-specific way.

Follicle stimulating hormone (FSH), testosterone (T) and estradiol (E2) are known to regulate testis maturation, and changes in FSH secretion induced by sex steroid treatment may mediate the effects of sex hormones. The aim of this study was to compare the effects of T and E2 on the pre-meiotic steps of first spermatogenesis and FSH level in rats. Male rat pups were injected daily with 17ß-estradiol benzoate (EB; 12.5 µg) or testosterone propionate (TP; 2.5mg) with the use of one of the two administration modes: 1/transient mode; hormone injections on postnatal days (PND) 1-5 followed by daily vehicle injections until PND 15 (t-EB and t-TP, respectively) or 2/continuous mode; hormone injections on PND 1-15 (c-EB and c-TP, respectively). The control group was injected with vehicle alone. On PND 16, blood was taken for serum hormone measurement and testes were collected for analysis of seminiferous tubule morphometry as well as cell number, proliferation and apoptosis. Testis weight, tubule length, Sertoli and germ cell numbers were reduced, and cell apoptosis in seminiferous epithelium was increased after transient EB and TP treatments. Despite normal or increased FSH secretion, the c-EB treatment inhibited pre-meiotic germ cell development and augmented cell apoptosis, whereas the c-TP treatment reduced the spermatocyte number and inhibited the formation of seminiferous tubule lumen. In conclusion, transient administration of EB or TP during PND 1-5 inhibited testis growth, whereas continuous administration (PND 1-15) impaired pre-meiotic germ cell development in a hormone-specific way.

The ability of three different models of frailty to predict all-cause mortality: results from the European Male Aging Study (EMAS).

Few studies have directly compared the ability of the most commonly used models of frailty to predict mortality among community-dwelling individuals. Here, we used a frailty index (FI), frailty phenotype (FP), and FRAIL scale (FS) to predict mortality in the EMAS. Participants were aged 40-79 years (n=2929) at baseline and 6.6% (n=193) died over a median 4.3 years of follow-up. The FI was generated from 39 deficits, including self-reported health, morbidities, functional performance and psychological assessments. The FP and FS consisted of five phenotypic criteria and both categorized individuals as robust when they had 0 criteria, prefrail as 1-2 criteria and frail as 3+ criteria. The mean FI increased linearly with age (r(2)=0.21) and in Cox regression models adjusted for age, center, smoking and partner status the hazard ratio (HR) for death for each unit increase of the FI was 1.49. Men who were prefrail or frail by either the FP or FS definitions, had a significantly increased risk of death compared to their robust counterparts. Compared to robust men, those who were FP frail at baseline had a HR for death of 3.84, while those who were FS frail had a HR of 3.87. All three frailty models significantly predicted future mortality among community-dwelling, middle-aged and older European men after adjusting for potential confounders. Our data suggest that the choice of frailty model may not be of paramount importance when predicting future risk of death, enabling flexibility in the approach used.

Comparisons of immunoassay and mass spectrometry measurements of serum estradiol levels and their influence on clinical association studies in men.

CONTEXT: Immunoassay-based techniques, routinely used to measure serum estradiol (E2), are known to have reduced specificity, especially at lower concentrations, when compared with the gold standard technique of mass spectrometry (MS). Different measurement techniques may be responsible for the conflicting results of associations between serum E2 and clinical phenotypes in men. OBJECTIVE: Our objective was to compare immunoassay and MS measurements of E2 levels in men and evaluate associations with clinical phenotypes. DESIGN AND SETTING: Middle-aged and older male subjects participating in the population-based Osteoporotic Fractures in Men (MrOS) Sweden study (n = 2599), MrOS US (n = 688), and the European Male Aging Study (n = 2908) were included. MAIN OUTCOME MEASURES: Immunoassay and MS measurements of serum E2 were compared and related to bone mineral density (BMD; measured by dual energy x-ray absorptiometry) and ankle-brachial index. RESULTS: Within each cohort, serum E2 levels obtained by immunoassay and MS correlated moderately (Spearman rank correlation coefficient rS 0.53-0.76). Serum C-reactive protein (CRP) levels associated significantly (albeit to a low extent, rS = 0.29) with immunoassay E2 but not with MS E2 levels. Similar associations of immunoassay E2 and MS E2 were seen with lumbar spine and total hip BMD, independent of serum CRP. However, immunoassay E2, but not MS E2, associated inversely with ankle-brachial index, and this correlation was lost after adjustment for CRP. CONCLUSIONS: Our findings suggest interference in the immunoassay E2 analyses, possibly by CRP or a CRP-associated factor. Although associations with BMD remain unaffected, this might imply for a reevaluation of previous association studies between immunoassay E2 levels and inflammation-related outcomes.

Active vitamin D (1,25-dihydroxyvitamin D) and bone health in middle-aged and elderly men: the European Male Aging Study (EMAS).

CONTEXT: There is little information on the potential impact of serum 1,25-dihydroxyvitamin D [1,25(OH)2D] on bone health including turnover. OBJECTIVE: The objective of the study was to determine the influence of 1,25(OH)2D and 25-hydroxyvitamin D [25(OH)D] on bone health in middle-aged and older European men. DESIGN, SETTING, AND PARTICIPANTS: Men aged 40-79 years were recruited from population registers in 8 European centers. Subjects completed questionnaires that included questions concerning lifestyle and were invited to attend for quantitative ultrasound (QUS) of the heel, assessment of height and weight, and a fasting blood sample from which 1,25(OH)2D, 25(OH)D, and PTH were measured. 1,25(OH)2D was measured using liquid chromatography tandem mass spectrometry. Bone markers serum N-terminal propeptide of type 1 procollagen (P1NP) and crosslinks (ß-cTX) were also measured. Dual-energy x-ray absorptiometry (DXA) of the hip and lumbar spine was performed in 2 centers. MAIN OUTCOME MEASURE(S): QUS of the heel, bone markers P1NP and ß-cTX, and DXA of the hip and lumbar spine were measured. RESULTS: A total of 2783 men, mean age 60.0 years (SD 11.0) were included in the analysis. After adjustment for age and center, 1,25(OH)2D was positively associated with 25(OH)D but not with PTH. 25(OH)D was negatively associated with PTH. After adjustment for age, center, height, weight, lifestyle factors, and season, 1,25(OH)2D was associated negatively with QUS and DXA parameters and associated positively with ß-cTX. 1,25(OH)2D was not correlated with P1NP. 25(OH)D was positively associated with the QUS and DXA parameters but not related to either bone turnover marker. Subjects with both high 1,25(OH)2D (upper tertile) and low 25(OH)D (lower tertile) had the lowest QUS and DXA parameters and the highest ß-cTX levels. CONCLUSIONS: Serum 1,25(OH)2D is associated with higher bone turnover and poorer bone health despite being positively related to 25(OH)D. A combination of high 1,25(OH)2D and low 25(OH)D is associated with the poorest bone health.

Frailty and sexual health in older European men.

BACKGROUND: There has been little research on how late-life frailty interrelates with sexual health. Our objective was to examine the association of frailty with sexual functioning and satisfaction among older men. METHODS: The study population consisted of 1,504 men aged 60 to 79 years, participating in the European Male Aging Study. Self-report questionnaires measured overall sexual functioning, sexual function-related distress, and erectile dysfunction. Frailty status was defined using a phenotype (FP) or index (FI). Associations between frailty and sexual function were explored using regression models. RESULTS: Based on the frailty phenotype, 5% of men were classified as frail, and the mean frailty index was 0.18 (SD = 0.12). Frailty was associated with decreasing overall sexual functioning and increasing sexual function-related distress in multiple linear regressions adjusted for age, smoking, alcohol consumption, living arrangements, comorbidities, and depression. Frailty was also associated with an increased odds of erectile dysfunction after adjustment for the same confounders: odds ratio = 1.99 (95% confidence interval = 1.14, 3.48) and 4.08 (95% confidence interval = 2.63, 6.36) for frailty phenotype and frailty index, respectively. CONCLUSIONS: Frailty was associated with impaired overall sexual functioning, sexual function-related distress, and erectile dysfunction. Individuals assessed for frailty-related deficits may also benefit from an appraisal of sexual health as an important aspect of well-being and quality of life.

Estrogen receptor alpha localization in the testes of men with normal spermatogenesis.

It is known that estrogens act on the male reproductive tract by binding to estrogen receptors (ER) α and ß. However, studies on ER localization in the human testis are discordant. The aim of this study was to investigate the localization of ERα in the testes of adult men with normal spermatogenesis. Semen analysis of ten adult men revealed azoospermia. FSH, LH and testosterone serum concentrations were within normal values, and the volume of the testes was normal, hence obstructive azoospermia was suspected. The tissues from testicular surgical biopsies were fixed in Bouin's fluid and embedded in paraffin. Assessments of the seminiferous epithelium (scoring 10 to -1), the number of Leydig cells (scoring 1 to 5), the areal fraction of intertubular space (IS), measurements of seminiferous tubule diameter, and the thickness of the tubular wall, were performed on microscopic sections. Immunohistochemical staining was applied with monoclonal antibodies against ERα. The mean spermatogenesis score was 10 points; IS - 30.6 ± 8.1%; seminiferous tubule diameter - 193.9 ± 19.4 µm; thickness of tubular wall - 7.44 ± 1.1 µm; number of Leydig cells - 1.6 ± 1.1 points. Immunohistochemical staining showed the localization of ERα to be in the Sertoli and Leydig cell cytoplasm, while ERα was absent in germ cells. The results of testicular tissue analysis confirmed its normal structure and normal, full spermatogenesis. The presence of ERα in Sertoli and Leydig cells in normal human testis demonstrated in this study suggests that estrogens may affect testicular function.

Comparison of serum testosterone and estradiol measurements in 3174 European men using platform immunoassay and mass spectrometry; relevance for the diagnostics in aging men.

BACKGROUND: The limitations of serum testosterone and estradiol (E(2)) measurements using non-extraction platform immunoassays (IAs) are widely recognized. Switching to more specific mass spectrometry (MS)-based methods has been advocated, but directly comparative data on the two methods are scarce. METHODS: We compared serum testosterone and E(2) measurements in a large sample of middle-aged/elderly men using a common platform IA and a gas chromatography (GC)-MS method, in order to assess their limitations and advantages, and to diagnose male hypogonadism. Of subjects from the European Male Aging Study (n=3174; age 40-79 years), peripheral serum testosterone and E(2) were analyzed using established commercial platform IAs (Roche Diagnostics E170) and in-house GC-MS methods. RESULTS: Over a broad concentration range, serum testosterone concentration measured by IA and MS showed high correlation (R=0.93, P<0.001), which was less robust in the hypogonadal range (<11 nmol/l; R=0.72, P<0.001). The IA/MS correlation was weaker in E(2) measurements (R=0.32, P<0.001, at E(2) <40.8 pmol/l, and R=0.74, P<0.001, at E(2) >40.8 pmol/l). Using MS as the comparator method, IA ascertained low testosterone compatible with hypogonadism (<11 nmol/l), with 75% sensitivity and 96.3% specificity. The same parameters with IA for the detection of low E(2) (<40.7 pmol/l) were 13.3 and 99.3%, and for high E(2) (>120 pmol/l) 88.4 and 88.6%. CONCLUSION: A validated platform IA is sufficient to detect subnormal testosterone concentrations in the diagnosis of male hypogonadism. The IA used for E(2) measurements showed poor correlation with MS and may only be suitable for the detection of high E(2) in men.

Association of hypogonadism with vitamin D status: the European Male Ageing Study.

OBJECTIVE: Interrelationships between hormones of the hypothalamic-pituitary-testicular (HPT) axis, hypogonadism, vitamin D and seasonality remain poorly defined. We investigated whether HPT axis hormones and hypogonadism are associated with serum levels of 25-hydroxyvitamin D (25(OH)D) in men. DESIGN AND METHODS: Cross-sectional survey of 3369 community-dwelling men aged 40-79 years in eight European centres. Testosterone (T), oestradiol (E(2)) and dihydrotestosterone were measured by gas chromatography-mass spectrometry; LH, FSH, sex hormone binding globulin (SHBG), 25(OH)D and parathyroid hormone by immunoassay. Free T was calculated from total T, SHBG and albumin. Gonadal status was categorised as eugonadal (normal T/LH), secondary (low T, low/normal LH), primary (low T, elevated LH) and compensated (normal T, elevated LH) hypogonadism. Associations of HPT axis hormones with 25(OH)D were examined using linear regression and hypogonadism with vitamin D using multinomial logistic regression. RESULTS: In univariate analyses, free T levels were lower (P=0.02) and E(2) and LH levels were higher (P<0.05) in men with vitamin D deficiency (25(OH)D <50 nmol/l). 25(OH)D was positively associated with total and free T and negatively with E(2) and LH in age- and centre-adjusted linear regressions. After adjusting for health and lifestyle factors, no significant associations were observed between 25(OH)D and individual hormones of the HPT axis. However, vitamin D deficiency was significantly associated with compensated (relative risk ratio (RRR)=1.52, P=0.03) and secondary hypogonadism (RRR=1.16, P=0.05). Seasonal variation was only observed for 25(OH)D (P<0.001). CONCLUSIONS: Secondary and compensated hypogonadism were associated with vitamin D deficiency and the clinical significance of this relationship warrants further investigation.