Average sperm count remains unchanged despite reduction in maternal smoking: results from a large cross-sectional study with annual investigations over 21 years.

STUDY QUESTION: How are temporal trends in lifestyle factors, including exposure to maternal smoking in utero, associated to semen quality in young men from the general population? SUMMARY ANSWER: Exposure to maternal smoking was associated with lower sperm counts but no overall increase in sperm counts was observed during the study period despite a decrease in this exposure. WHAT IS KNOWN ALREADY: Meta-analyses suggest a continuous decline in semen quality but few studies have investigated temporal trends in unselected populations recruited and analysed with the same protocol over a long period and none have studied simultaneous trends in lifestyle factors. STUDY DESIGN, SIZE, DURATION: Cross-sectional population-based study including ~300 participants per year (total number = 6386) between 1996 and 2016. PARTICIPANTS/MATERIALS, SETTING, METHODS: The study is based on men from the Greater Copenhagen area, Denmark, with a median age of 19 years, and unselected with regard to fertility status and semen quality. The men delivered a semen sample, had a blood sample drawn and a physical examination performed and answered a comprehensive questionnaire, including information on lifestyle and the mother's pregnancy. Temporal trends in semen quality and lifestyle were illustrated graphically, and trends in semen parameters and the impact of prenatal and current lifestyle factors were explored in multiple regression analyses. MAIN RESULTS AND THE ROLE OF CHANCE: Throughout the study period, 35% of the men had low semen quality. Overall, there were no persistent temporal trends in semen quality, testicular volume or levels of follicle-stimulating hormone over the 21 years studied. The men's alcohol intake was lowest between 2011 and 2016, whereas BMI, use of medication and smoking showed no clear temporal trends. Parental age increased, and exposure in utero to maternal smoking declined from 40% among men investigated in 1996-2000 to 18% among men investigated in 2011-2016. Exposure to maternal smoking was associated with lower sperm counts but no overall increase in sperm counts was observed despite the decrease in this exposure. LIMITATIONS, REASONS FOR CAUTION: Information of current and prenatal lifestyle was obtained by self-report, and the men delivered only one semen sample each. WIDER IMPLICATIONS OF THE FINDINGS: The significant decline in in utero exposure to maternal smoking, which was not reflected in an overall improvement of semen quality at the population level, suggest that other unknown adverse factors may maintain the low semen quality among Danish men. STUDY FUNDING/COMPETING INTEREST(S): The study has received financial support from the ReproUnion; the Research fund of Rigshospitalet, Copenhagen University Hospital; the European Union (Contract numbers BMH4-CT96-0314,QLK4-CT-1999-01422, QLK4-CT-2002-00603, FP7/2007-2013, DEER Grant agreement no. 212844); the Danish Ministry of Health; the Danish Environmental Protection Agency; A.P. Møller and wife Chastine McKinney Møllers foundation; and Svend Andersens Foundation. None of the funders had any role in the study design, collection, analysis or interpretation of data, writing of the paper or publication decisions. TRIAL REGISTRATION NUMBER: N/A.

Semen quality associated with subsequent hospitalizations - Can the effect be explained by socio-economic status and lifestyle factors?

Semen quality is suggested to be a universal biomarker for future health. Previous studies have mostly been registry based excluding the possibility to address the importance of lifestyle, fertility status, health and socio-economic status. We aimed to investigate whether the association between semen quality and subsequent risk of hospitalization could be explained by differences in occupation, education, fertility, cryptorchidism, BMI or smoking; 1423 men with first semen sample at Fertility Clinic, Frederiksberg Hospital, Denmark, from 1977 to 2010 responded to a questionnaire in 2012 about current health, lifestyle, educational level and occupation. They were followed in the Danish National Patient Registry to first-time hospitalizations using ICD-8 and ICD-10 classification. Data were analysed by Cox proportional hazard regression models to adjust for the possible confounding factors. We found a significant higher risk of being hospitalized with decreasing sperm concentrations (0-15 mill/mL: HR1.78, 95% CI:1.51-2.09; 16-50 mill/mL: HR 1.37 95% CI: 1.17-1.60; 51-100 mill/mL: HR1.25 95% CI: 1.07-1.45). Same significant association of being hospitalized with decreasing total sperm counts was seen. The dose-response increase in risk in hospitalization with decreasing sperm concentration and total sperm count remained constant after further individual adjustment for occupation, marital status, fertility, cryptorchidism, BMI or smoking. The association between semen quality and subsequent morbidity was not explained by differences in lifestyle, behavioural or fertility status. We were unable to adjust for all possible confounders simultaneously due to limited sample size, and reverse causation is a possible explanation as information about education and lifestyle was obtained after semen analysis and hospitalizations occurred and may have changed as consequence of both. Semen quality may be a universal biomarker for future health not explained by lifestyle and socio-economic status, but this needs to be addressed further in future studies.

Possible involvement of the glucocorticoid receptor (NR3C1) and selected NR3C1 gene variants in regulation of human testicular function.

Perceived stress has been associated with decreased semen quality but the mechanisms have not been elucidated. It is not known whether cortisol, the major stress hormone in humans, can act directly via receptors in the testis, and whether variants in the gene encoding the glucocorticoid receptor (NR3C1) can possibly modulate the effect. To address these questions, we investigated the expression of the glucocorticoid receptor in human testicular tissue, including adult and fetal samples (n = 20) by immunohistochemical staining, and in silico analysis of publicly available datasets. In the adult testis NR3C1 protein was detected in peritubular cells, a subset of Leydig cells, Sertoli cells (weak), and spermatogonia, but not in spermatids. The NR3C1 expression pattern in fetal testis samples differed by a notably stronger reaction in Sertoli cells, lack of staining in gonocytes but the presence in a subset of pro-spermatogonia, and the almost absent reaction in nascent peritubular cells. In parallel, we explored the association between adult testicular function and three single nucleotide NR3C1 polymorphisms (BcII [rs41423247], 9ß [rs6198], and Tth111I [rs10052957]) affecting glucocorticoid sensitivity. Testicular function was determined by semen analysis and reproductive hormone profiling in 893 men from the general population. The NR3C1 SNP BclI was associated with semen quality in an over-dominant manner with heterozygotes having better semen parameters compared to both homozygote constellations, and with sperm motility showing the strongest association. This association was supported by a higher inhibin B and inhibin B/FSH ratio, as well as a lower FSH in BclI heterozygotes. The SNPs 9ß and Tth111I were not associated with semen parameters. Although the clinical impact of the findings is limited, the results substantiate a suggested link between stress and testicular function. Hence this investigation should be regarded as a discovery study generating hypotheses for future studies.

Are incident gallstones associated to sex-dependent changes with age? A cohort study.

Age and female sex have repeatedly been identified as gallstone determinants but the underlying mechanisms are not clarified. The objectives of this study were to determine if changes with age in physiology, lifestyle, or reproductive hormones were associated with incident gallstones. A cohort study of a general population random sample (N = 2366) aged 30-60 years was performed. Participants were ultrasound screened for gallstones in 1982-84 and again in 1993-94. Lifestyle data and blood samples were obtained and re-analyzed in 2004. Changes with age in physiology (body mass index, blood pressure, blood lipids, self-rated health), lifestyle (smoking, alcohol and coffee consumption, dietary habits, physical activity level), and indices of reproductive function (number of births, oral contraceptive use, hormone replacement therapy, male reproductive hormones) were explored in females and males separately. Adjusted logistic regression analyses were performed. Incident gallstones (gallstones and cholecystectomy) at ultrasound examination in participants initially free of gallstones at baseline occurred in 9.9% of the study population. In females, increasing alcohol consumption (odds ratio (OR) 0.94, 95% confidence interval (CI) [0.90; 0.98]) and the cessation of hormone replacement therapy (OR 0.29, 95% CI [0.10; 0.83]) inversely determined incident gallstones. In males, increasing levels of SHBG (OR 0.97, 95% CI [0.94; 0.998]) inversely determined incident gallstones. Other changes with age in physiology, lifestyle, or reproductive hormones were not associated. High baseline free testosterone determined incident gallstones in males (OR 1.15, 95% CI [1.02; 1.30]). To conclude, changes with age in alcohol consumption in females and in reproductive hormones determined incident gallstones. Male reproductive hormones seem to have an impact on incident gallstones. Sex differences should be explored further in future studies.

Reference ranges of 17-hydroxyprogesterone, DHEA, DHEAS, androstenedione, total and free testosterone determined by TurboFlow-LC-MS/MS and associations to health markers in 304 men.

We report reference ranges based on LC-MS/MS for testosterone (T), free testosterone (FT) and its precursors, i.e. 17-hydroxyprogesterone (17-OHP), dehydroepiandrosterone (DHEA), DHEA-sulfate (DHEAS) and androstenedione (Adione), in relation to different health markers and lifestyle factors. The study was based on 304 healthy men aged 30-61 years participating in a population-based cross-sectional study (Health2008). Examination program consisted of a clinical examination, completion of a self-administered questionnaire and blood sampling. Steroid metabolites were measured by a validated and sensitive LC-MS/MS method. Older age-groups were significantly associated with decreased concentrations of DHEA, DHEAS, Adione, and FT, while no significant associations with age were shown for 17-OHP or T. Participants with BMI≥30 kg/m(2) had lower age-related steroid metabolite z-scores compared to participants with BMI<30 kg/m(2), i.e. 17-OHP: -0.51 vs. 0.08 (p<0.001); DHEA: -0.27 vs. 0.09 (p=0.014); Adione: -0.29 vs. 0.09 (p=0.012); T: -0.99 vs. 0.14 (p<0.001); and FT -0.55 vs. 0.05 (p<0.001), respectively. In conclusion, this large study on serum steroid metabolites and concomitant assessment of health markers in healthy men provides age-related reference ranges, and furthermore evaluates the impact of lifestyle factors and metabolic syndrome on androgen metabolite levels.

Increasing trends in childlessness in recent birth cohorts - a registry-based study of the total Danish male population born from 1945 to 1980.

The fertility rate has recently declined in many parts of the World, including Europe. To a large extent, this change can be explained by the socio-economic development. However, increasing fertility problems and widespread occurrence of poor semen quality could in part explain the few births. The objective of this registry based study was to investigate birth cohort related trends in fertility and childlessness among Danish men. The study population comprised all 1 616 677 men in Denmark born from 1945 to 1980 of whom 1 359 975 (84.1%) were native Danes. Data were obtained from Statistics Denmark and contained information from The National Danish Birth Registry and The Danish In Vitro Fertilization (IVF) Registry. For consecutive birth cohorts of native Danish men cumulative fertility rates at age 45 declined from 1.91 children per man in the 1945 birth cohort to 1.71 for men born in 1960. The proportion of childless men at age 45 increased from 14.8% to 21.9% in the same birth cohorts. Assisted reproductive technology (ART) seemed to compensate partly for the lower fertility and to reduce the proportion of childless men. In contrast, recent reports on corresponding birth cohorts of Danish women showed that the proportion remaining childless throughout life has been lower than in men and has not shown a similar increase. In conclusion, using unique Danish registries the study showed a birth cohort related decline in fertility rates and an increase in childlessness among men. In the more recent cohorts more than one in five men remained childless. The causes behind the findings are likely multi-factorial. Hitherto, most attention has been given to socio-economic factors which undoubtedly play a major role. Our findings lend support to the hypothesis that the high prevalence of low sperm counts among young Danish men may be a contributing factor.