Immune system adaptation during gender-affirming testosterone treatment.

Infectious, inflammatory and autoimmune conditions present differently in males and females. SARS-CoV-2 infection in naive males is associated with increased risk of death, whereas females are at increased risk of long COVID1, similar to observations in other infections2. Females respond more strongly to vaccines, and adverse reactions are more frequent3, like most autoimmune diseases4. Immunological sex differences stem from genetic, hormonal and behavioural factors5 but their relative importance is only partially understood6-8. In individuals assigned female sex at birth and undergoing gender-affirming testosterone therapy (trans men), hormone concentrations change markedly but the immunological consequences are poorly understood. Here we performed longitudinal systems-level analyses in 23 trans men and found that testosterone modulates a cross-regulated axis between type-I interferon and tumour necrosis factor. This is mediated by functional attenuation of type-I interferon responses in both plasmacytoid dendritic cells and monocytes. Conversely, testosterone potentiates monocyte responses leading to increased tumour necrosis factor, interleukin-6 and interleukin-15 production and downstream activation of nuclear factor kappa B-regulated genes and potentiation of interferon-γ responses, primarily in natural killer cells. These findings in trans men are corroborated by sex-divergent responses in public datasets and illustrate the dynamic regulation of human immunity by sex hormones, with implications for the health of individuals undergoing hormone therapy and our understanding of sex-divergent immune responses in cisgender individuals.

Testosterone Treatment and Fractures in Men with Hypogonadism.

BACKGROUND: Testosterone treatment in men with hypogonadism improves bone density and quality, but trials with a sufficiently large sample and a sufficiently long duration to determine the effect of testosterone on the incidence of fractures are needed. METHODS: In a subtrial of a double-blind, randomized, placebo-controlled trial that assessed the cardiovascular safety of testosterone treatment in middle-aged and older men with hypogonadism, we examined the risk of clinical fracture in a time-to-event analysis. Eligible men were 45 to 80 years of age with preexisting, or high risk of, cardiovascular disease; one or more symptoms of hypogonadism; and two morning testosterone concentrations of less than 300 ng per deciliter (10.4 nmol per liter), in fasting plasma samples obtained at least 48 hours apart. Participants were randomly assigned to apply a testosterone or placebo gel daily. At every visit, participants were asked if they had had a fracture since the previous visit. If they had, medical records were obtained and adjudicated. RESULTS: The full-analysis population included 5204 participants (2601 in the testosterone group and 2603 in the placebo group). After a median follow-up of 3.19 years, a clinical fracture had occurred in 91 participants (3.50%) in the testosterone group and 64 participants (2.46%) in the placebo group (hazard ratio, 1.43; 95% confidence interval, 1.04 to 1.97). The fracture incidence also appeared to be higher in the testosterone group for all other fracture end points. CONCLUSIONS: Among middle-aged and older men with hypogonadism, testosterone treatment did not result in a lower incidence of clinical fracture than placebo. The fracture incidence was numerically higher among men who received testosterone than among those who received placebo. (Funded by AbbVie and others; TRAVERSE ClinicalTrials.gov number, NCT03518034.).

Hormonal basis of sex differences in anesthetic sensitivity.

General anesthesia-a pharmacologically induced reversible state of unconsciousness-enables millions of life-saving procedures. Anesthetics induce unconsciousness in part by impinging upon sexually dimorphic and hormonally sensitive hypothalamic circuits regulating sleep and wakefulness. Thus, we hypothesized that anesthetic sensitivity should be sex-dependent and modulated by sex hormones. Using distinct behavioral measures, we show that at identical brain anesthetic concentrations, female mice are more resistant to volatile anesthetics than males. Anesthetic sensitivity is bidirectionally modulated by testosterone. Castration increases anesthetic resistance. Conversely, testosterone administration acutely increases anesthetic sensitivity. Conversion of testosterone to estradiol by aromatase is partially responsible for this effect. In contrast, oophorectomy has no effect. To identify the neuronal circuits underlying sex differences, we performed whole brain c-Fos activity mapping under anesthesia in male and female mice. Consistent with a key role of the hypothalamus, we found fewer active neurons in the ventral hypothalamic sleep-promoting regions in females than in males. In humans, we demonstrate that females regain consciousness and recover cognition faster than males after identical anesthetic exposures. Remarkably, while behavioral and neurocognitive measures in mice and humans point to increased anesthetic resistance in females, cortical activity fails to show sex differences under anesthesia in either species. Cumulatively, we demonstrate that sex differences in anesthetic sensitivity are evolutionarily conserved and not reflected in conventional electroencephalographic-based measures of anesthetic depth. This covert resistance to anesthesia may explain the higher incidence of unintended awareness under general anesthesia in females.

A chemical signal in human female tears lowers aggression in males.

Rodent tears contain social chemosignals with diverse effects, including blocking male aggression. Human tears also contain a chemosignal that lowers male testosterone, but its behavioral significance was unclear. Because reduced testosterone is associated with reduced aggression, we tested the hypothesis that human tears act like rodent tears to block male aggression. Using a standard behavioral paradigm, we found that sniffing emotional tears with no odor percept reduced human male aggression by 43.7%. To probe the peripheral brain substrates of this effect, we applied tears to 62 human olfactory receptors in vitro. We identified 4 receptors that responded in a dose-dependent manner to this stimulus. Finally, to probe the central brain substrates of this effect, we repeated the experiment concurrent with functional brain imaging. We found that sniffing tears increased functional connectivity between the neural substrates of olfaction and aggression, reducing overall levels of neural activity in the latter. Taken together, our results imply that like in rodents, a human tear-bound chemosignal lowers male aggression, a mechanism that likely relies on the structural and functional overlap in the brain substrates of olfaction and aggression. We suggest that tears are a mammalian-wide mechanism that provides a chemical blanket protecting against aggression.

Understanding One Half of the Sex Difference Equation: The Modulatory Effects of Testosterone on Diabetic Cardiomyopathy.

Diabetes is a prevalent disease, primarily characterized by high blood sugar (hyperglycemia). Significantly higher rates of myocardial dysfunction have been noted in individuals with diabetes, even in those without coronary artery disease or high blood pressure (hypertension). Numerous molecular mechanisms have been identified through which diabetes contributes to the pathology of diabetic cardiomyopathy, which presents as cardiac hypertrophy and fibrosis. At the cellular level, oxidative stress and inflammation in cardiomyocytes are triggered by hyperglycemia. Although males are generally more likely to develop cardiovascular disease than females, diabetic males are less likely to develop diabetic cardiomyopathy than are diabetic females. One reason for these differences may be the higher levels of serum testosterone in males compared with females. Although testosterone appears to protect against cardiomyocyte oxidative stress and exacerbate hypertrophy, its role in inflammation and fibrosis is much less clear. Additional preclinical and clinical studies will be required to delineate testosterone's effect on the diabetic heart.

Androgenic steroids induce pathologic scarring in a preclinical porcine model via dysfunctional extracellular matrix deposition.

Hypertrophic scarring is a major source of morbidity. Sex hormones are not classically considered modulators of scarring. However, based on increased frequency of hypertrophic scarring in patients on testosterone, we hypothesized that androgenic steroids induce abnormal scarring and developed a preclinical porcine model to explore these effects. Mini-swine underwent castration, received no testosterone (noT) or biweekly testosterone therapy (+T), and underwent excisional wounding. To create a delayed wound healing model, a subset of wounds were re-excised at 2 weeks. Scars from postoperative day 42 (POD42) and delayed wounds (POD28) were harvested 6 weeks after initial wounding for analysis via histology, bulk RNA-seq, and mechanical testing. Histologic analysis of scars from +T animals showed increased mean fibrosis area (16 mm2noT, 28 mm2+T; p = .007) and thickness (0.246 mm2noT, 0.406 mm2+T; p < .001) compared to noT. XX+T and XY+T scars had greater tensile burst strength (p = .024 and p = .013, respectively) compared to noT swine. Color deconvolution analysis revealed greater deposition of type I and type III collagen as well as increased collagen type I:III ratio in +T scars. Dermatopathologist histology scoring showed that +T exposure was associated with worse overall scarring (p < .05). Gene ontology analysis found that testosterone exposure was associated with upregulation of cellular metabolism and immune response gene sets, while testosterone upregulated pathways related to keratinization and laminin formation on pathway analysis. In conclusion, we developed a preclinical porcine model to study the effects of the sex hormone testosterone on scarring. Testosterone induces increased scar tissue deposition and appears to increase physical strength of scars via supraphysiologic deposition of collagen and other ECM factors. The increased burst strength seen in both XX and XY animals suggests that hormone administration has a strong influence on scar mechanical properties independent of chromosomal sex. Anti-androgen topical therapies may be a promising future area of research.

Female-biased embryonic death from inflammation induced by genomic instability.

Genomic instability can trigger cellular responses that include checkpoint activation, senescence and inflammation1,2. Although genomic instability has been extensively studied in cell culture and cancer paradigms, little is known about its effect during embryonic development, a period of rapid cellular proliferation. Here we report that mutations in the heterohexameric minichromosome maintenance complex-the DNA replicative helicase comprising MCM2 to MCM73,4-that cause genomic instability render female mouse embryos markedly more susceptible than males to embryonic lethality. This bias was not attributable to X chromosome-inactivation defects, differential replication licensing or X versus Y chromosome size, but rather to 'maleness'-XX embryos could be rescued by transgene-mediated sex reversal or testosterone administration. The ability of exogenous or endogenous testosterone to protect embryos was related to its anti-inflammatory properties5. Ibuprofen, a non-steroidal anti-inflammatory drug, rescued female embryos that contained mutations in not only the Mcm genes but also the Fancm gene; similar to MCM mutants, Fancm mutant embryos have increased levels of genomic instability (measured as the number of cells with micronuclei) from compromised replication fork repair6. In addition, deficiency in the anti-inflammatory IL10 receptor was synthetically lethal with the Mcm4Chaos3 helicase mutant. Our experiments indicate that, during development, DNA damage associated with DNA replication induces inflammation that is preferentially lethal to female embryos, because male embryos are protected by high levels of intrinsic testosterone.

Decreased AMPK/SIRT1/PDK4 induced by androgen excess inhibits human endometrial stromal cell decidualization in PCOS.

Polycystic ovary syndrome (PCOS) is a complex common endocrine disorder affecting women of reproductive age. Ovulatory dysfunction is recognized as a primary infertile factor, however, even when ovulation is medically induced and restored, PCOS patients continue to experience reduced cumulative pregnancy rates and a higher spontaneous miscarriage rate. Hyperandrogenism, a hallmark feature of PCOS, affects ovarian folliculogenesis, endometrial receptivity, and the establishment and maintenance of pregnancy. Decidualization denotes the transformation that the stromal compart of the endometrium must undergo to accommodate pregnancy, driven by the rising progesterone levels and local cAMP production. However, studies on the impact of hyperandrogenism on decidualization are limited. In this study, we observed that primary endometrial stromal cells from women with PCOS exhibit abnormal responses to progesterone during in vitro decidualization. A high concentration of testosterone inhibits human endometrial stromal cells (HESCs) decidualization. RNA-Seq analysis demonstrated that pyruvate dehydrogenase kinase 4 (PDK4) expression was significantly lower in the endometrium of PCOS patients with hyperandrogenism compared to those without hyperandrogenism. We also characterized that the expression of PDK4 is elevated in the endometrium stroma at the mid-secretory phase. Artificial decidualization could enhance PDK4 expression, while downregulation of PDK4 leads to abnormal decidualization both in vivo and in vitro. Mechanistically, testosterone excess inhibits IGFBP1 and PRL expression, followed by phosphorylating of AMPK that stimulates PDK4 expression. Based on co-immunoprecipitation analysis, we observed an interaction between SIRT1 and PDK4, promoting glycolysis to facilitate decidualization. Restrain of AR activation resumes the AMPK/SIRT1/PDK4 pathway suppressed by testosterone excess, indicating that testosterone primarily acts on decidualization through AR stimulation. Androgen excess in the endometrium inhibits decidualization by disrupting the AMPK/SIRT1/PDK4 signaling pathway. These data demonstrate the critical roles of endometrial PDK4 in regulating decidualization and provide valuable information for understanding the underlying mechanism during decidualization.

Combined repetitive inhalant endotoxin and collagen-induced arthritis drive inflammatory lung disease and arthritis severity in a testosterone-dependent manner.

Respiratory-related diseases are a leading cause of death in rheumatoid arthritis (RA) and are disproportionately higher in men, which may be attributable to environmental risk factors. Animal studies have demonstrated potentiated autoimmunity, arthritis, and profibrotic/inflammatory lung disease with a combination of airborne exposures and collagen-induced arthritis (CIA). This study aimed to determine whether hormone-dependent differences explained these observations. Arthritis-prone male intact and castrated DBA/1J mice received intranasal inhalation of lipopolysaccharide (LPS) daily for 5 wk and CIA induction. Arthritis scores and serum pentraxin-2 levels were increased in castrated versus intact mice. In contrast, airway cell influx, lung tissue infiltrates, and lung levels of proinflammatory and profibrotic markers (C5a, IL-33, and matrix metalloproteinases) were reduced in castrated versus intact mice. CIA + LPS-induced lung histopathology changes and the expression of lung autoantigens including malondialdehyde acetaldehyde (MAA)- and citrulline (CIT)-modified proteins and vimentin were reduced in castrated animals. There were no differences in serum anti-MAA or anti-CIT protein antibody (ACPA) levels or serum pentraxin levels between groups. Testosterone replacement led to a reversal of several lung inflammatory/profibrotic endpoints noted earlier in castrated male CIA + LPS-treated mice with testosterone supplementation promoting neutrophil influx, MAA expression, and TNF-α, IL-6, and MMP-9. These findings imply that testosterone contributes to lung and arthritis inflammatory responses following CIA + LPS coexposure, but not to systemic autoantibody responses. The CIA + LPS model provides a paradigm for investigations focused on the mechanistic underpinnings for epidemiologic and phenotypic sex differences in RA-related lung disease.NEW & NOTEWORTHY Our study shows that testosterone acts as a key immunomodulatory hormone contributing to critical features of rheumatoid arthritis (RA)-associated lung disease in the setting of airborne endotoxin (lipopolysaccharide; LPS) exposures and concomitant arthritis induction in mice. The exaggerated airway inflammation observed following combined exposures in male mice was accompanied by increases in profibrotic mediators, netosis, and increased expression of lung autoantigens, all relevant to the pathogenesis of lung disease in arthritis.

NADPH oxidase 4-derived hydrogen peroxide counterbalances testosterone-induced endothelial dysfunction and migration.

High levels of testosterone (Testo) are associated with cardiovascular risk by increasing reactive oxygen species (ROS) formation. NADPH oxidases (NOX) are the major source of ROS in the vasculature of cardiovascular diseases. NOX4 is a unique isotype, which produces hydrogen peroxide (H2O2), and its participation in cardiovascular biology is controversial. So far, it is unclear whether NOX4 protects from Testo-induced endothelial injury. Thus, we hypothesized that supraphysiological levels of Testo induce endothelial NOX4 expression to attenuate endothelial injury. Human mesenteric vascular endothelial cells (HMECs) and human umbilical vein endothelial cells (HUVEC) were treated with Testo (10-7 M) with or without a NOX4 inhibitor [GLX351322 (10-4 M)] or NOX4 siRNA. In vivo, 10-wk-old C57Bl/6J male mice were treated with Testo (10 mg/kg) for 30 days to study endothelial function. Testo increased mRNA and protein levels of NOX4 in HMECs and HUVECs. Testo increased superoxide anion (O2-) and H2O2 production, which were abolished by NOX1 and NOX4 inhibition, respectively. Testo also attenuated bradykinin-induced NO production, which was further impaired by NOX4 inhibition. In vivo, Testo decreased H2O2 production in aortic segments and triggered endothelial dysfunction [decreased relaxation to acetylcholine (ACh)], which was further impaired by GLX351322 and by a superoxide dismutase and catalase mimetic (EUK134). Finally, Testo led to a dysregulated endothelial cell migration, which was exacerbated by GLX351322. These data indicate that supraphysiological levels of Testo increase the endothelial expression and activity of NOX4 to counterbalance the deleterious effects caused by Testo in endothelial function.NEW & NOTEWORTHY By inducing ROS formation, high levels of testosterone play a major role in the pathogenesis of cardiovascular disease. NOXs are the major sources of ROS in the vasculature of cardiovascular diseases. Herein, we describe a novel compensatory mechanism by showing that NOX4 is a protective oxidant enzyme and counterbalances the deleterious effects of testosterone in endothelial cells by modulating hydrogen peroxide formation.

METTL14 mediates nerve growth factor-stimulated testosterone synthesis in porcine theca cells†.

Ovarian theca cells produce testosterone, which acts as a vital precursor substance for synthesizing estrogens during follicular development. Nerve growth factor (NGF) has been shown to participate in reproductive physiology, specifically to follicular development and ovulation. There is currently no available data on the impact of NGF on testosterone synthesis in porcine theca cells. Furthermore, m6A modification is the most common internal modification in eukaryotic mRNAs that are closely associated with female gametogenesis, follicle development, ovulation, and other related processes. It is also uncertain whether the three main enzymes associated with m6A, such as Writers, Erasers, and Readers, play a role in this process. The present study, with an in vitro culture model, investigated the effect of NGF on testosterone synthesis in porcine theca cells and the role of Writers-METTL14 in this process. It was found that NGF activates the PI3K/AKT signaling pathway through METTL14, which regulates testosterone synthesis in porcine theca cells. This study will help to further elucidate the mechanisms by which NGF regulates follicular development and provide new therapeutic targets for ovary-related diseases in female animals. Summary Sentence  The present study investigated the effect of NGF on testosterone synthesis in porcine theca cells. It was found that NGF activates the PI3K/AKT signaling pathway through METTL14, which regulates testosterone synthesis in porcine theca cells.

Long noncoding RNA XIST inhibition promotes Leydig cell apoptosis by acting as a competing endogenous RNA for microRNA-145a-5p that targets SIRT1 in late-onset hypogonadism.

Leydig cell (LCs) apoptosis is responsible for decreased serum testosterone levels during late-onset hypogonadism (LOH). Our study was designed to illustrate the regulatory effect of lncRNA XIST on LCs and to clarify its molecular mechanism of action in LOH. The Leydig cells (TM3) was treated by 300 µM H2O2 for 8 h to establish Leydig cell oxidative stress model in vitro. The expression levels of lncRNA XIST in the testicular tissues of patients with LOH were measured using fluorescence in situ hybridization (FISH). The interaction between lncRNA XIST/SIRT1 and miR-145a-5p was assessed using starBase and dual-luciferase reporter gene assays. Apoptotic cells and Caspase3 activity were determined by flow cytometry (FCM) assay. Testosterone concentration was determined by ELISA. Moreover, histological assessment of testicles in mice was performed by using HE staining and the TUNEL assay was used to determine apoptosis. We found that the lncRNA XIST was downregulated in the testicular tissues of LOH patients and mice and in H2O2-induced TM3 cells. XIST siRNA significantly promoted apoptosis, enhanced Caspase3 activity and reduced testosterone levels in H2O2-stimulated TM3 cells. Further studies showed that the miR-145a-5p inhibitor reversed the effect of XIST-siRNA on H2O2-induced Leydig cell apoptosis. MiR-145a-5p negatively regulated SIRT1 expression, and SIRT1-siRNA reversed the effects of the miR-145a-5p inhibitor on H2O2 stimulated TM3 cells. The in vivo experiments indicated that silencing of the lncRNA XIST aggravated LOH symptoms in mice. Inhibition of lncRNA XIST induces Leydig cell apoptosis through the miR-145a-5p/SIRT1 axis in the progression of LOH.

Stored platelet hemostatic phenotype and function is not altered when donors are on testosterone replacement therapy.

BACKGROUND: Critical shortages in the national blood supply have led to a re-evaluation of previously overlooked donor sources for blood products. As a part of that effort, red blood cells collected from therapeutic phlebotomy of donors on testosterone replacement therapy (TRT) have been conditionally approved for transfusion. However, platelets from TRT donors are not currently approved for use due to limited data on effects of supraphysiologic testosterone on recipient safety and platelet function. The objective of this study was to provide a comprehensive profile of phenotype and function in platelets from TRT and control donors. STUDY DESIGN AND METHODS: Platelets in plasma were collected from TRT and control donors (N = 10 per group; age- and sex-matched) and stored at room temperature for 7 days. On storage Day 1 (D1) and Day 7 (D7), platelet products were analyzed for platelet count, metabolic parameters (i.e., glucose, lactate, mitochondrial function), surface receptor expression, aggregation, thrombin generation, and thrombus formation under physiological flow conditions. RESULTS: TRT donor platelets were not significantly different than control donor platelets in terms of count, surface phenotype, metabolic function, ability to aggregate, thrombin generation, or ability to form occlusive thrombus under arterial flow regimes. Both groups were similar to each other by D7, but had significantly lost hemostatic function compared to D1. DISCUSSION: Platelets derived from donors undergoing TRT have similar phenotypic and functional profiles compared to those derived from control donors. This suggests that therapeutic phlebotomy of TRT donors may provide a useful source for platelet products.

Impact of continuous testosterone exposure on reproductive physiology, activity, and pain-related behavior in young adult female rats.

Testosterone may reduce pain in cisgender women and transgender men. Rodents can provide a useful model for investigating physiological effects of hormone therapy. To this end, continuous-release testosterone or blank (placebo) capsules were implanted s.c. into young adult female rats, and three weeks later rats were either ovariectomized or sham-ovariectomized. Testosterone treatment that mimicked previously reported endogenous levels in males eliminated estrous cycling and decreased uterine weight. Testosterone also significantly increased body weight and suppressed the increases in daily wheel running observed in placebo controls over time. Subsequent ovariectomy or sham-ovariectomy decreased wheel running in all groups, but testosterone-treated rats recovered significantly more quickly than did placebo-treated rats. Neither testosterone nor ovariectomy significantly altered hindpaw mechanical threshold. Two weeks after sham/ovariectomy surgery, injection of Complete Freund Adjuvant (CFA) into one hindpaw reduced wheel running and mechanical threshold in all groups; running significantly decreased from the first to second day after CFA in testosterone- but not in placebo-treated rats. Morphine 1.0 but not 3.2 mg/kg increased CFA-suppressed wheel running similarly in all groups, whereas both doses of morphine increased CFA-suppressed mechanical threshold. These data suggest that weeks-long testosterone treatment with or without ovariectomy may provide a useful physiological model of testosterone therapy as used in human gender transition. Although testosterone administered at levels similar to those in gonadally intact males tended to hasten female rats' recovery from surgery, it did not decrease maximal pain-related behaviors after surgery or hindpaw inflammatory insult, nor did it alter opioid antinociception.

Activational and organizational effects of testosterone on the number of mating partners and reproductive success in males of a social rodent.

The timing of exposure to the steroid hormone, testosterone, produces activational and organizational effects in vertebrates. These activational and organizational effects are hypothesized to relate with the number of female mating partners and reproductive success in males. We tested this hypothesis by examining 151 wild degu (Octodon degus) males across a 10-year study. We quantified the association between adult serum testosterone levels (i.e., an indirect index of adult activational effects) and anogenital distance (AGD) length (i.e., a direct index of fetal organizational effects), and their interaction on the number of female mating partners and reproductive success. We found no evidence of an association between adult male serum testosterone levels and the number of female mating partners, or between adult male serum testosterone levels and reproductive success. However, male AGD was positively associated with reproductive success, but not so with the number of female mating partners. Additionally, the positive association between male AGD and male reproductive success was mediated by the number of mates. Our findings do not support major roles of activational or organizational effects of testosterone on the number of female mating partners and its consequences on male reproductive success. Instead, our results suggest that compared with individual male attributes, the female social environment plays a more important role in driving male reproductive success.

Androgen signaling in LMAN regulates song stereotypy in male canaries.

During breeding when testosterone concentrations are high, male songbirds that are open-ended vocal learners like canaries (Serinus canaria) tend to produce a stable, stereotyped song that facilitates mate attraction or territory defense. Outside breeding contexts, song becomes more variable. The neuroendocrine mechanisms controlling this vocal variability across seasons are not entirely clear. We tested whether androgen signaling within the lateral magnocellular nucleus of the anterior nidopallium (LMAN), a cortical-like brain region of the vocal control system known as a vocal variability generator, plays a role in seasonal vocal variability. We first characterized song in birds housed alone on a short day (SD) photoperiod, which simulates non-breeding conditions. Then, cannulae filled with the androgen receptor (AR) blocker flutamide or left empty as control were implanted bilaterally in LMAN. Birds were then transferred to long days (LD) to simulate the breeding season and song was analyzed again. Blocking AR in LMAN increased acoustic variability of song and the acoustic variability of syllables. However, blocking AR in LMAN did not impact the variability of syllable usage nor their sequencing in LD birds, song features that are controlled by androgen signaling in a somatosensory brain region of the vocal control system called HVC. These findings highlight the multifactorial, non-redundant actions of steroid hormones in controlling complex social behaviors such as birdsong. They also support the hypothesis that LMAN is a key brain area for the effects of testosterone on song plasticity both seasonally in adults and during the song crystallization process at sexual maturity.

The role of male hormones in bacterial infections: enhancing Staphylococcus aureus virulence through testosterone-induced Agr activation.

Staphylococcus aureus is a notorious pathogen predominantly involved in skin and soft tissue infections, exhibiting a distinct innate sex bias. This study explores the influence of testosterone on the virulence of S. aureus and elucidates its underlying mechanisms. Utilizing a skin abscess model in intact and castrated male mice, we assessed the effects of testosterone on S. aureus pathogenicity. Compared to controls, castrated mice showed significantly reduced abscess sizes and decreased bacterial loads, highlighting the role of testosterone in modulating the severity of S. aureus infections. In vitro experiments revealed that testosterone enhances the hemolytic activity, cytotoxicity, and oxidative stress resistance of S. aureus. Real-time quantitative PCR analysis showed a significant upregulation of the genes encoding α-hemolysin (hla) and phenol-soluble modulin (psmα). Importantly, testosterone treatment significantly enhanced the expression of the accessory gene regulator (Agr) quorum-sensing system components (agrC, agrA, agrB, agrD), while the SaeRS system (saeR, saeS, and sbi) exhibited only slight changes. Gene knockout experiments revealed that deletion of agrC, rather than saeRS and agrBD, abolishes the testosterone-induced enhancement of hemolysis and gene expression, underscoring the key role of AgrC. Molecular docking simulations indicated a direct interaction between testosterone and AgrC protein, with a strong binding affinity at the active site residue SER201. This study provides new insights into the mechanistic basis of how testosterone enhances the pathogenicity of S. aureus, potentially contributing to increased male susceptibility to S. aureus infections and offering a targeted approach for therapeutic interventions.

Effects of maternal androgens and their metabolite etiocholanolone on prenatal development in birds.

Offspring phenotypes can be affected by maternal testosterone and androstenedione (A4), which are considered a tool of mothers to adjust offspring to a fluctuating environment. Yet testosterone and A4 are very rapidly metabolized by developing avian embryos, suggesting that either the maternal testosterone and A4 have potent organizational effects on the embryos extremely early before being metabolized or it is the metabolites that evoke phenotypic variation in the offspring. One of the metabolites, etiocholanolone, increases substantially during early embryonic development and is a likely candidate for mediating maternal effects as it can promote erythropoiesis. To investigate and compare the effects of testosterone and A4 with the possible effects of etiocholanolone during prenatal embryonic development, we increased their levels in black-headed gull eggs (Larus ridibundus), and used sham-injected eggs as controls. This species usually has 3-egg clutches in which maternal androgen levels increase with the egg-laying sequence. We analysed embryonic heart rate, peri-hatching biometric traits, the ratio of white to red blood cells (W/R ratio) and bursa development. We found that testosterone and A4 treatment increased embryonic heart rate irrespective of egg-laying sequence and decreased bill length and W/R ratio, whereas etiocholanolone did not mimic these effects. Instead, etiocholanolone treatment decreased tarsus length and brain mass. Our finding that etiocholanolone does not mimic the effects induced by testosterone and A4 suggests that the embryonic metabolism of maternal testosterone and A4 can potentially diversify the function of these maternal androgens.

Penile erection and cardiovascular function: effects and pathophysiology.

Penile erection (PE) is a hemodynamic event that results from a neuroendocrine process, and it is influenced by the cardiovascular status of the patient. However, it may also modulate an individual's cardiovascular events. The present study provides the mechanisms involved in the association of PE and cardiovascular function. Erection upsurges the cardiac rate, blood pressure, and oxygen uptake. Sex-enhancing strategies, such as phosphodiesterase inhibitors, alprostadil, and testosterone also promote vasodilatation and cardiac performance, thus preventing myocardial infarction. More so, drugs that are used in the treatment of hypertensive heart diseases (such as angiotensin system inhibitors and ß-blockers) facilitate vasodilatation and PE. These associations have been linked with nitric oxide- and testosterone-dependent enhancing effects on the vascular endothelium. In addition, impaired cardiovascular function may negatively impact PE; therefore, impaired PE may be a pointer to cardiovascular pathology. Hence, evaluation of the cardiovascular status of an individual with erectile dysfunction (ED) is essential. Also, employing strategies that are used in maintaining optimal cardiac function may be useful in the management of ED.