Disorders of sexual development (DSD) are associated with atypical chromosomal, gonadal, or phenotypic sex. It is likely that the number of cases of DSD are underestimated in the equine population. Monorchidism in the horse is very rare. This case report describes the clinical assessment of a phenotypic mare with stallion-like behavior which led to the diagnosis of a DSD. A 4-year-old Quarter Horse mare presented in good body condition, with normal external genitalia for a mare, and normal mammary glands with two bilaterally symmetric teats. No uterus, cervix, or gonads were detected on transrectal palpation. Transrectal ultrasonography revealed a single gonad in the right dorsal abdomen with the morphologic appearance of a testicle. Pre-surgical hormonal evaluation revealed elevated serum testosterone and anti-Müllerian hormone (AMH) concentrations. The right gonad was successfully removed via standing exploratory laparoscopy and submitted for histopathology. No gonad was identified on the left side during laparoscopy. Histopathologic examination confirmed that the excised gonad was a testicle. Cytogenetic and molecular analysis revealed a 64,XY, SRY-positive chromosomal constitution. Hormonal evaluation 5 weeks after surgery revealed low serum testosterone and AMH levels. A diagnosis of monorchidism was based on ultrasound examination, laparoscopic exploration of the abdomen, removal of a single gonad, and a subsequent decrease in serum testosterone and AMH concentrations to basal levels. In summary, a combination of clinical signs, endocrine evaluation, chromosomal and molecular analysis, and histopathology can be used in the diagnosis of DSD conditions.

Benzene is a universal ambient pollutant. Population-based studies have shown that benzene exposure affects male fertility. However, the mechanism of benzene-induced reproductive toxicity is unknown. Here, we established a dynamic inhalation model and exposed C57BL/6J mice to 0, 10, and 50 ppm benzene (6 h/day, 6 days/week, 7 weeks). Our study revealed that benzene exposure caused testicular injury, including structural damage to spermatogenic tubules, reduced semen quality, and decreased testosterone levels. In addition, the decrease in the global level of N6-Methyladenosine (m6A) and the change of m6A important regulatory enzymes in mice testes suggested that m6A was involved in the benzene-induced testicular injury. Further genome-wide m6A methylation analysis showed that 1469 differential m6A peaks were present in the testes of control and benzene groups, indicating that benzene exposure modulated m6A methylation in testes. Furthermore, the comprehensive analysis of m6A-sequencing and transcriptome revealed that hypermethylated Rara and its consequent reduced expression impaired the sperm production process. In particular, melatonin alleviated benzene-induced testicular injury by modulating m6A-related genes. Overall, our research provides a new idea and fundamental knowledge into the possible mechanisms of m6A modifications in benzene-induced testicular impairment, as well as a new experimental basis for benzene-induced male fertility therapy.

Testosterone has an anabolic effect on skeletal muscle. The testes produce most of the testosterone in vivo, while the adrenal glands contribute smaller amounts. When testis testosterone is deficient the adrenal gland increases steroid hormone synthesis, which is referred to as compensatory testicular adaptation (CTA).To reveal the effects of testis testosterone deficiency on adrenal steroid hormones synthesis and skeletal muscle development, gene expression related to adrenal steroid hormones synthesis and skeletal muscle development were determined by RNA-seq. The results showed that castrating male ducks had significant effects on their body weight but no significant impact on cross-sectional area (CSA) or density of pectoral muscle fibres. In skeletal muscle protein metabolism, expression levels of the catabolic gene atrogin1/MAFbx and the anabolic gene eEF2 were significantly higher, with concomitant increases after castration. The adrenal glands' alteration of the steroid hormone 11β-hydroxylase (CYP11B1) was significantly lower following castration. Expression pattern analysis showed that the adrenal glands' glucocorticoid receptor (NR3C1/GR) had a potential regulatory relationship with the skeletal muscle-related genes (Pax7, mTOR, FBXO32, FOXO3, and FOXO4). The data showed that castration affected muscle protein metabolism, adrenal steroid and testosterone synthesis. In addition, it was speculated that, after castration, steroid hormones produced by the adrenal gland could have a compensatory effect, which might mediate the changes in skeletal muscle protein metabolism and development.

Aggressive reactions to peer victimization may be tempered by hormone levels. Grounded on the dualhormone hypothesis (DHH), which proposes that testosterone (T) is associated with aggressive behavior only when cortisol (C) is low, this study assessed whether the combination of T and C moderated adolescents' aggressive responses to peer victimization. The study involved 577 adolescents (50.4% girls, aged 12-17 years), who completed measures of online and offline victimization and perpetration of aggressive behavior in three waves over the course of one year. Moreover, they provided salivary samples to measure T and C levels. Multilevel analyses showed a three-way interaction between T, C, and victimization levels for both online and offline aggressive behaviors. In both cases, the adolescents with high T and high C or low T and low C responded with more aggressive behaviors when victimized or provoked by peers. The T/C ratio was only associated with aggressive behavior in the girls' sample. The results are opposite to those predicted by the DHH, but they are consistent with the findings of other studies that examined aggressive behaviors as reactions to provocations. These results suggest that some combinations of T and C predict higher aggressive reactions to peer victimization.

The present study looks for components in seminal plasma (SP) and/or serum that are closely related to in vivo fertility of buffalo bulls. Fourteen healthy mature buffalo bulls were classified according to their in vivo fertility into fertile (n = 10) and subfertile (n = 4) groups. Semen and serum samples were collected from all animals for 12 replicates. The collected ejaculates were examined for sperm characteristics before being centrifuged to collect SP for hormonal (FSH, LH, testosterone, and IGF-1), biochemical [total antioxidant capacity (TAC), catalase (CAT), glutathione peroxidase (GPx), nitric oxide (NO), malondialdehyde (MDA), fructose, total protein, albumin, triglycerides, cholesterol, and high-density lipoprotein (HDL)] and proteomic (SDS-PAGE) analyses. Likewise, serum levels of FSH, LH, testosterone, IGF-1, glucose, total protein, albumin, triglycerides, cholesterol, and HDL were determined. All sperm characteristics and the majority of sperm kinematics were (P < 0.01) different between fertile and subfertile groups. Seminal and serum levels of FSH, LH, testosterone, and IGF-1 were higher (P < 0.01) in the fertile group, but only seminal fructose, total protein, albumin, triglycerides, cholesterol, and HDL were higher (P < 0.01) in the fertile group. Moreover, the fertile group had greater TAC, CAT, GPx, and NO, but the subfertile group had greater MDA. Protein bands of 14, 15, 26, 30, and 55 kDa were larger and denser in the SP of the fertile group but were smaller and faint to absent in that of the subfertile group. Also, the protein fractions of detected protein bands demonstrated a substantial influence of fertility on those of 16, 26, 30, and 55 kDa. In conclusion, sperm characteristics and kinematics with serum, and/or seminal hormonal and biochemical components, should be evaluated for reliable prediction of buffalo bull fertility. Furthermore, protein bands of 26, 30, and 55 kDa may represent fertility-associated proteins in buffalo bull SP.

The present study aimed to investigate whether time-restricted feeding (TRF) ameliorates metabolic and reproductive phenotypes in a letrozole-induced mouse model of polycystic ovary syndrome (PCOS). Sixty female C57BL/6 N mice were randomly divided into two groups according to the type of food received: either a chow or a 60% high-fat diet. Those mice were subcutaneously implanted with letrozole or placebo pellets at four weeks of age. Then, letrozole-treated mice were randomly assigned to different feeding regimens: (1) TRF for 4 h (ZT12-ZT16) or (2) ad libitum diet. After 4 weeks of dietary intervention, estrous cycles were determined with daily vaginal smear examination, and serial tail-tip blood sampling was performed at 5-min intervals for 2 h to measure the luteinizing hormone (LH) pulse frequency, amplitude, and mean LH levels in the diestrus cycle stage. Letrozole-treated mice in the ad libitum group demonstrated multiple PCOS-like phenotypes including ovulatory dysfunction, polycystic ovaries, and increased body weight, parametrial fat weight, adipocyte size and inflammation, and higher expression of Cyp17, Cyp19, and Fshr in the ovary, and Kiss1r and Gnrh in the hypothalamus, elevated serum testosterone levels, and more rapid and elevated LH pulsatility, with increased pulse frequency, amplitude, and mean levels in the diestrus stage, compared with the controls. After TRF for 4 weeks, those phenotypes reverted to normal levels in letrozole-treated mice, except the percentage of diestrus cycles indicating the arrest of estrous cycling which did not differ between the TRF and ad libitum groups. Our results demonstrate that TRF has therapeutic effects on the reproductive and metabolic phenotypes of a letrozole-induced mouse model of PCOS.

Day-to-day coordination of the stress (i.e., hypothalamic-pituitary-adrenal [HPA]) and reproductive (i.e., hypothalamic-pituitary-gonadal [HPG]) axes is central to allostatic regulation, reproductive success, and survival. Reports of positive, within-person testosterone and cortisol relationships (or coupling) suggest cross-talk of a facilitative nature, but longitudinal evidence is scarce and has methodological and analytical limitations. To address this, we used a continuous-time (CT) model to investigate day-to-day, within-person coupling of testosterone and cortisol in two male cohorts. Salivary testosterone and cortisol fluctuations were monitored in 35 athletic men across two international tournaments (M = 19.3 tests) and in 41 healthy men during normal daily living (M = 27.9 tests). Bayesian CT analysis revealed a diminishing effect of each hormone on itself as time-interval length or lag increased. In both groups, cortisol had a negative lagged effect on testosterone that persisted for around three days. The cortisol effect on testosterone peaked after 0.71 and 0.51 days in athletic (standardized estimate = -0.13) and healthy men (standardized estimate = -0.11), respectively. Further estimates of non-lagged, contemporaneous correlations revealed positive testosterone and cortisol relationships (athlete r = 0.04, healthy r = 0.46). In summary, complex within-person HPA and HPG interplay emerged in two independent male cohorts. Specifically, a rising cortisol concentration was linked to a fall in testosterone concentration at later time points, but concurrently these hormones tended to rise and fall together. Our results suggest that inhibitory and facilitatory hormonal actions coexist on varying timescales, thereby expanding knowledge of HPG and HPA cross-talk in everyday life.

Sontag, SA, Cabarkapa, D, and Fry, AC. Testosterone and cortisol salivary samples are stable across multiple freeze-thaw cycles. J Strength Cond Res XX(X): 000-000, 2022-When processing salivary samples for biomarker analysis, avoiding multiple freeze-thaw cycles is generally recommended. However, confusing tissue handling instructions or challenges with collections in the field sometimes makes this problematic. Thus, the purpose of this study was to examine if the stability of salivary testosterone (T) and cortisol (C) hormones remains unchanged when exposed to multiple freeze-thaw cycles. Seven healthy recreationally active adults provided salivary samples at rest (i.e., 1600 hours) for analysis of T and C. Samples were separated into 4 aliquots for each hormone and underwent 4 freeze-thaw cycles (T1-T4 and C1-C4) before being analyzed by enzyme-linked immunosorbent assay. The overall analysis of variance model was significant for T (p = 0.008) and nonsignificant for C (p = 0.820). A follow-up post hoc comparison indicated significant differences in salivary hormonal concentrations between T1 and T4 (p = 0.029), T2 and T4 (p = 0.007), and T3 and T4 (p = 0.032). The findings of this study indicate that salivary steroid hormones seem to be relatively stable following multiple freeze-thaw cycles. However, C seems to be more stable when exposed to multiple freeze-thaw cycles, as T concentrations did reveal a significant decrease by the fourth thaw cycle.

Sex pheromones are chemical substances secreted into the environment that affect the physiology and behavior of recipients. Females use these compounds during oestrus to attract males, which leads to attempts of mating. This study evaluates the influence of manual semen collection in male dogs, in the presence or absence of a female in estrus, on the blood concentrations of cortisol (CRT), oxytocin (OXT), prolactin (PRL) and testosterone (T), as hormones involved both in the physiology of reproduction and stress. Ten male dogs were used in Experiment 1 to measure the serum and plasma concentrations of the aforementioned hormones in the absence of semen collection. Subsequently in the same animals, the concentrations of these hormones were evaluated before and after semen collection in the presence (Exp. 2) or in absence of a female in estrus (Exp. 3). No significant changes in hormone concentration caused by the semen collection were found, either with, or without the presence of female in estrus. Obtained results suggest that the procedure of manual semen collection in dogs, probably due to its passive character, does not stimulate endocrine glands to secrete hormones, and the process of ejaculation is probably controlled by neural pathway. The lack of effect of semiochemical stimulation to the CRT, PRL, OXT and T level, could be caused by a short contact with female during semen collection. Further studies on involvement of the hormones during the process of natural mating, especially preceded by long courtships, similar to that observed under natural conditions, should shed a light on the physiology of mating and the connection between the endocrine system and semiochemical stimulation in dogs.

Hematological parameters and erythropoiesis are known to be influenced by anabolic androgenic steroid (AAS) use. However, little is known in relation to supra-physiological doses of AAS. Therefore, the aim of this study was to evaluate the effect of supra-physiological doses of AAS on serum and urinary erythropoietin (EPO), and blood parameters, from self-reported AAS users. Serum EPO levels were higher in testosterone positive AAS users (11.83 mIU/mL ± 4.19) than non-positive (6.60 mIU/mL ±2.70, p=0.03), while no differences in urinary EPO levels were noted. There were positive correlations between serum EPO and testosterone levels (rs=0.46, p=0.01) and reticulocyte percentage (rs=0.43, p=0.02). Individuals with AAS-induced hypogonadism (ASIH; luteinizing hormone levels <1.4 IU/L) had approximately 75 % higher serum EPO (p<0.05) and 140 % higher high fluorescence reticulocyte fractions (p<0.001), as well as other affected hematological parameters, compared to non-ASIH individuals. The results extend the knowledge of how endocrine and hematological biomarkers are affected by AAS doping.

Over the twentieth century, male reproductive health has suffered a substantial decline, as evidenced by decreases in sperm counts and testosterone levels and increases in reproductive pathologies. At the same time, the prevalence of chronic diseases such as obesity, diabetes, and metabolic syndrome has risen dramatically. Metabolic and reproductive health are highly interconnected, suggesting that their respective trends are intertwined and, given the timeframe of such trends, environmental and not genetic factors are most likely to be the primary causes. Industrialization, which began in Europe in the mid-eighteenth century, has resulted in profound changes to our diet, lifestyle, and environment, many of which are causal factors in the rise in chronic diseases. Industrialization results in a nutrition transition from an agricultural unprocessed to a modern processed diet, incorporating increases in sugar, vegetable oils, ultra-processed foods, linoleic acid, trans-fats, and total energy. This dietary shift has incurred numerous adverse effects on metabolic and reproductive health, characterized by chronic inflammation, oxidative stress, and insulin resistance. Moreover, these effects appear to multiply across subsequent generations via epigenetic inheritance. Men's fertility is markedly affected by obesity and diabetes, with an increase in total energy via processed food intake arguably being the key factor driving the diabesity pandemic. In contrast, wholefoods rich in micronutrients and phytonutrients support male fertility and a healthy body weight. Therefore, men wanting to maximize their fertility should consider making positive dietary changes, such as replacing processed foods with unprocessed foods that support metabolic and reproductive health.

The purpose of this study was to find out whether the seminal testosterone and/or estradiol levels could serve as prognostic criteria for normal spermatogenesis and whether they are able to characterize the sperm pathology. The study involved healthy young male volunteers (n=269); serum and seminal steroid hormones were measured; the sperm concentration, mobility, and morphology were evaluated. The results indicate that the seminal testosterone concentration is lower (p<0.05) and the seminal estradiol is higher than the corresponding parameters in the serum (p<0.05). The seminal testosterone and estradiol concentrations negatively correlated with the sperm concentration, and the seminal estradiol concentration was higher in pathozoospermic than in normospermic men (p<0.05). It is assumed that the seminal estradiol level can be an indicator of sperm quality and serve as a biological predictor of normal spermatogenesis; in addition, this parameter can be used for diagnostic purposes in patients with impaired spermatogenesis induced by excess of estrogens.