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The accuracy of Follicle Stimulating Hormone as a predictor of azoospermia in adult survivors of childhood cancer is unclear, with conflicting results in the published literature. A systematic review and post hoc analysis of combined data (n = 367) were performed on all published studies containing extractable data on both serum Follicle Stimulating Hormone concentration and semen concentration in survivors of childhood cancer. PubMed and Medline databases were searched up to March 2017 by two blind investigators. Articles were included if they contained both serum FSH concentration and semen concentration, used World Health Organisation certified methods for semen analysis, and the study participants were all childhood cancer survivors. There was no evidence for either publication bias or heterogeneity for the five studies. For the combined data (n = 367) the optimal Follicle Stimulating Hormone threshold was 10.4 IU/L with specificity 81% (95% CI 76%-86%) and sensitivity 83% (95% CI 76%-89%). The AUC was 0.89 (95%CI 0.86-0.93). A range of threshold FSH values for the diagnosis of azoospermia with their associated sensitivities and specificities were calculated. This study provides strong supporting evidence for the use of serum Follicle Stimulating Hormone as a surrogate biomarker for azoospermia in adult males who have been treated for childhood cancer.

Inhibin B has been identified as a potential marker of Sertoli cell function in males. The aim of this study is to produce a normative model of serum inhibin B in males from birth to seventeen years. We used a well-defined search strategy to identify studies containing data that can contribute to a larger approximation of the healthy population. We combined data from four published studies (n = 709) and derived an internally validated model with high goodness-of-fit and normally distributed residuals. Our results show that inhibin B increases following birth to a post-natal peak of 270 pg/mL (IQR 210-335 pg/mL) and then decreases during childhood followed by a rise at around 8 years, peaking at a mean 305 pg/mL (IQR 240-445 pg/mL) at around age 17. Following this peak there is a slow decline to the standard mature adult normal range of 170 pg/mL (IQR 125-215 pg/mL). This normative model suggests that 35% of the variation in Inhibin B levels in young males is due to age alone, provides an age-specific reference range for inhibin B in the young healthy male population, and will be a powerful tool in evaluating the potential of inhibin B as a marker of Sertoli cell function in pre-pubertal boys.

The Sertoli cells are critical regulators of testis differentiation and development. In the adult, however, their known function is restricted largely to maintenance of spermatogenesis. To determine whether the Sertoli cells regulate other aspects of adult testis biology we have used a novel transgenic mouse model in which Amh-Cre induces expression of the receptor for Diphtheria toxin (iDTR) specifically within Sertoli cells. This causes controlled, cell-specific and acute ablation of the Sertoli cell population in the adult animal following Diphtheria toxin injection. Results show that Sertoli cell ablation leads to rapid loss of all germ cell populations. In addition, adult Leydig cell numbers decline by 75% with the remaining cells concentrated around the rete and in the sub-capsular region. In the absence of Sertoli cells, peritubular myoid cell activity is reduced but the cells retain an ability to exclude immune cells from the seminiferous tubules. These data demonstrate that, in addition to support of spermatogenesis, Sertoli cells are required in the adult testis both for retention of the normal adult Leydig cell population and for support of normal peritubular myoid cell function. This has implications for our understanding of male reproductive disorders and wider androgen-related conditions affecting male health.

The diagnosis of hypogonadism in human males includes identification of low serum testosterone levels, and hence there is an underlying assumption that normal ranges of testosterone for the healthy population are known for all ages. However, to our knowledge, no such reference model exists in the literature, and hence the availability of an applicable biochemical reference range would be helpful for the clinical assessment of hypogonadal men. In this study, using model selection and validation analysis of data identified and extracted from thirteen studies, we derive and validate a normative model of total testosterone across the lifespan in healthy men. We show that total testosterone peaks [mean (2.5-97.5 percentile)] at 15.4 (7.2-31.1) nmol/L at an average age of 19 years, and falls in the average case [mean (2.5-97.5 percentile)] to 13.0 (6.6-25.3) nmol/L by age 40 years, but we find no evidence for a further fall in mean total testosterone with increasing age through to old age. However we do show that there is an increased variation in total testosterone levels with advancing age after age 40 years. This model provides the age related reference ranges needed to support research and clinical decision making in males who have symptoms that may be due to hypogonadism.

In rodents, in utero exposure to exogenous estrogens including diethylstilboestrol (DES) results in major suppression of steroidogenesis in fetal testes. Whether similar effects occur in the human fetal testis is equivocal. Based on the results of the rodent studies, we hypothesised that exposure of human fetal testes to DES would result in a reduction in testosterone production. We show, using a xenograft approach, that testosterone production is not reduced in human fetal testis following DES exposure. Human fetal testes (15-19 weeks' gestation, n = 6) were xenografted into castrate male nude mice which were then treated for 35 days with vehicle or 100 µg/kg DES three times a week. For comparison, similar treatment was applied to pregnant rats from e13.5-e20.5 and effects on fetal testes evaluated at e21.5. Xenograft testosterone production was assessed by measuring host seminal vesicle (SV) weights as an indirect measure over the entire grafting period, and single measurement of serum testosterone at termination. Human fetal testis xenografts showed similar survival in DES and vehicle-exposed hosts. SV weight (44.3 v 26.6 mg, p = 0.01) was significantly increased in DES compared to vehicle-exposed hosts, respectively, indicating an overall increase in xenograft testosterone production over the grafting period, whilst serum testosterone at termination was unchanged. In contrast intra-testicular testosterone levels were reduced by 89%, in fetal rats exposed to DES. In rats, DES effects are mediated via Estrogen Receptor α (ESR1). We determined ESR1 protein and mRNA expression in human and rat fetal testis. ESR1 was expressed in rat, but not in human, fetal Leydig cells. We conclude that human fetal testis exposure to DES does not impair testosterone production as it does in rats, probably because ESR1 is not expressed in human fetal Leydig cells. This indicates that DES exposure is likely to pose minimal risk to masculinization of the human fetus.

Reproductive disorders that are common/increasing in prevalence in human males may arise because of deficient androgen production/action during a fetal 'masculinization programming window'. We identify a potentially important role for Chicken Ovalbumin Upstream Promoter-Transcription Factor II (COUP-TFII) in Leydig cell (LC) steroidogenesis that may partly explain this. In rats, fetal LC size and intratesticular testosterone (ITT) increased ~3-fold between e15.5-e21.5 which associated with a progressive decrease in the percentage of LC expressing COUP-TFII. Exposure of fetuses to dibutyl phthalate (DBP), which induces masculinization disorders, dose-dependently prevented the age-related decrease in LC COUP-TFII expression and the normal increases in LC size and ITT. We show that nuclear COUP-TFII expression in fetal rat LC relates inversely to LC expression of steroidogenic factor-1 (SF-1)-dependent genes (StAR, Cyp11a1, Cyp17a1) with overlapping binding sites for SF-1 and COUP-TFII in their promoter regions, but does not affect an SF-1 dependent LC gene (3β-HSD) without overlapping sites. We also show that once COUP-TFII expression in LC has switched off, it is re-induced by DBP exposure, coincident with suppression of ITT. Furthermore, other treatments that reduce fetal ITT in rats (dexamethasone, diethylstilbestrol (DES)) also maintain/induce LC nuclear expression of COUP-TFII. In contrast to rats, in mice DBP neither causes persistence of fetal LC COUP-TFII nor reduces ITT, whereas DES-exposure of mice maintains COUP-TFII expression in fetal LC and decreases ITT, as in rats. These findings suggest that lifting of repression by COUP-TFII may be an important mechanism that promotes increased testosterone production by fetal LC to drive masculinization. As we also show an age-related decline in expression of COUP-TFII in human fetal LC, this mechanism may also be functional in humans, and its susceptibility to disruption by environmental chemicals, stress and pregnancy hormones could explain the origin of some human male reproductive disorders.