Sexual dimorphism in a mouse model of Friedreich's ataxia with severe cardiomyopathy.

Friedreich's ataxia (FA) is an autosomal recessive disorder caused by reduced frataxin (FXN) expression in mitochondria, where the lethal component is cardiomyopathy. Using the conditional Fxnflox/null::MCK-Cre knock-out (Fxn-cKO) mouse model, we discovered significant sex differences in the progression towards heart failure, with Fxn-cKO males exhibiting a worse cardiac phenotype, low survival rate, kidney and reproductive organ deficiencies. These differences are likely due to a decline in testosterone in Fxn-cKO males. The decrease in testosterone was related to decreased expression of proteins involved in cholesterol transfer into the mitochondria: StAR and TSPO on the outer mitochondrial membrane, and the cholesterol side-chain cleavage enzyme P450scc and ferredoxin on the inner mitochondrial membrane. Expression of excitation-contraction coupling proteins (L-type calcium channel, RyR2, SERCA2, phospholamban and CaMKIIδ) was decreased significantly more in Fxn-cKO males. This is the first study that extensively investigates the sexual dimorphism in FA mouse model with cardiac calcium signaling impairment.

The impact of furan exposure on steroidogenesis in Leydig cells: cellular and molecular observations.

BACKGROUND: Furan is an organic compound that occurs as a result of heat treatment during the processing and cooking of many food products. Furthermore, the environment contains furan in tobacco smoke and vehicle exhaust gases, and it serves as an intermediate molecule in the synthesis of various pharmaceutical and chemical agents, pesticides, and stabilizers. Studies on the male reproductive system have not been able to elucidate the pathway through which furan exerts its negative effects. METHODS AND RESULTS: In this study, the TM3 Leydig cell line was exposed to various furan concentrations (0.03, 0.3, and 3 mM) for 24 h. In order to assess the cytotoxic effects of furan on Leydig cells, we examined cell viability, cell proliferation, and lactate dehydrogenase enzyme levels. To investigate the detrimental effects of furan on testosterone biosynthesis, quantitative analyses were conducted on cAMP and testosterone levels, as well as the expression levels of key genes and transcription factors implicated in the steroidogenic pathway. The results indicate that furan inhibited the viability and proliferation of Leydig cells and enhanced the activity of lactate dehydrogenase. Leydig cells administered to furan exhibited notable reductions in cAMP and testosterone levels. Additionally, while the expression levels of steroidogenic genes were downregulated, significant changes were detected in the expression levels of the transcription factors responsible for the regulation of these genes. CONCLUSIONS: Consequently, our findings suggest that furan exerts inhibitory effects on steroidogenesis in Leydig cells through multiple mechanisms, ultimately leading to infertility by inducing dysfunction in Leydig cells.

Meconium as an Analyte for Androgen Exposure: Analysis Through Varying Maternal-Fetal Biomarkers.

Meconium, the first stool produced by neonates, has been used as an analyte for exogenous fetal exposures. However, few studies have investigated the relationship between meconium and androgen exposure in utero. Here, we examine the associations of testosterone and dehydroepiandrosterone (DHEA) across maternal antenatal salivary testosterone, cord blood, meconium, and infant salivary testosterone. A total of 47 women with singleton, uncomplicated pregnancies, and their infants were included in this study. Participants were recruited from an academic obstetric clinic. Maternal saliva was collected at 36-weeks' gestation. Cord blood and meconium were collected at birth. Infant salivary testosterone was collected at 1 and 4 weeks of age. Multivariate model results showed that meconium testosterone was associated with neonatal testosterone at 1 (F = 5.62, p = 0.029) and 4 weeks (F = 4.28, p = 0.048) postnatal age; no sex differences were detected. This study suggests meconium is a valuable tool for evaluating endogenous androgen exposure and should be used in future studies to investigate the fetal hormonal milieu.

Oocyte-derived growth differentiation factor 9 suppresses the expression of CYP17A1 and androgen production in human theca cells.

OBJECTIVE: To investigate the direct effect of growth differentiation factor 9 (GDF9) on androgen production in human theca cells. DESIGN: Experimental study. SETTING: Tertiary hospital-based research laboratory. PATIENT(S): Women who underwent in vitro fertilization and intracytoplasmic sperm injections at our clinic were included in this study. INTERVENTION(S): Primary cultured human theca cells from women undergoing in vitro fertilization and intracytoplasmic sperm injection treatment were treated with GDF9, an activin receptor-like kinase 5 (ALK5) inhibitor, and a SMAD4 agonist. MAIN OUTCOME MEASURE(S): The expression of androgen synthesis-related genes StAR, CYP17A1, and LHCGR, levels of androstenedione and testosterone, phosphorylation of SMAD2/3, and the interaction between bone morphogenic protein-activated type II receptor and ALK5 were evaluated using reverse transcription-quantitative polymerase chain reaction, Western blot, enzyme-linked immunosorbent assays, and coimmunoprecipitation assays, respectively. RESULT(S): Growth differentiation factor 9 decreased StAR, CYP17A1, and LHCGR expression levels in human theca cells, which was prevented by treatment with the ALK5 inhibitor, and suppressed production of androgen in human theca cells. Growth differentiation factor 9 increased SMAD2/3 phosphorylation, and the ALK5 inhibitor also suppressed this effect. Bone morphogenic protein-activated type II receptor and ALK5 bound to each other after GDF9 stimulation. The SMAD4 agonist kartogenin also decreased messenger RNA levels of StAR and CYP17A1 and protein levels of StAR in human theca cells. CONCLUSION(S): Growth differentiation factor 9 can activate the bone morphogenic protein-activated type II receptor-ALK5-SMAD2/3 signaling pathway, suppress CYP17A1 expression, and decrease androgen production in human theca cells.

Strategies found not to be suitable for stabilizing high steroid hydroxylation activities of CYP450 BM3-based whole-cell biocatalysts.

The implementation of biocatalytic steroid hydroxylation processes plays a crucial role in the pharmaceutical industry due to a plethora of medicative effects of hydroxylated steroid derivatives and their crucial role in drug approval processes. Cytochrome P450 monooxygenases (CYP450s) typically constitute the key enzymes catalyzing these reactions, but commonly entail drawbacks such as poor catalytic rates and the dependency on additional redox proteins for electron transfer from NAD(P)H to the active site. Recently, these bottlenecks were overcome by equipping Escherichia coli cells with highly active variants of the self-sufficient single-component CYP450 BM3 together with hydrophobic outer membrane proteins facilitating cellular steroid uptake. The combination of the BM3 variant KSA14m and the outer membrane pore AlkL enabled exceptionally high testosterone hydroxylation rates of up to 45 U gCDW-1 for resting (i.e., living but non-growing) cells. However, a rapid loss of specific activity heavily compromised final product titers and overall space-time yields. In this study, several stabilization strategies were evaluated on enzyme-, cell-, and reaction level. However, neither changes in biocatalyst configuration nor variation of cultivation media, expression systems, or inducer concentrations led to considerable improvement. This qualified the so-far used genetic construct pETM11-ksa14m-alkL, M9 medium, and the resting-cell state as the best options enabling comparatively efficient activity along with fast growth prior to biotransformation. In summary, we report several approaches not enabling a stabilization of the high testosterone hydroxylation rates, providing vital guidance for researchers tackling similar CYP450 stability issues. A comparison with more stable natively steroid-hydroxylating CYP106A2 and CYP154C5 in equivalent setups further highlighted the high potential of the investigated CYP450 BM3-based whole-cell biocatalysts. The immense and continuously developing repertoire of enzyme engineering strategies provides promising options to stabilize the highly active biocatalysts.

Mouse testicular macrophages can independently produce testosterone and are regulated by Cebpb.

BACKGROUND: Testicular macrophages (TM) have long been recognized for their role in immune response within the testicular environment. However, their involvement in steroid hormone synthesis, particularly testosterone, has not been fully elucidated. This study aims to explore the capability of TM to synthesize and secrete testosterone de novo and to investigate the regulatory mechanisms involved. RESULTS: Transcriptomic analysis revealed significant expression of Cyp11a1, Cyp17a1, Hsd3b1, and Hsd17b3 in TM, which are key enzymes in the testosterone synthesis pathway. qPCR analysis and immunofluorescence validation confirmed the autonomous capability of TM to synthesize testosterone. Ablation of TM in mice resulted in decreased physiological testosterone levels, underscoring the significance of TM in maintaining testicular testosterone levels. Additionally, the study also demonstrated that Cebpb regulates the expression of these crucial genes, thereby modulating testosterone synthesis. CONCLUSIONS: This research establishes that TM possess the autonomous capacity to synthesize and secrete testosterone, contributing significantly to testicular testosterone levels. The transcription factor Cebpb plays a crucial role in this process by regulating the expression of key genes involved in testosterone synthesis.

Professional football training and recovery: A longitudinal study on the effects of weekly conditioning session and workload variables.

The main purpose of this longitudinal study was to investigate football players' recovery status, through hormonal response, in relation to accumulated workload at two comparable time points of the first (T1) and second half (T2) of the competitive season. Moreover, this study investigated athletes' hormonal response to a typical weekly conditioning session (5 days before match: MD-5), at T1 and T2, to detect changes in players' recovery capability over time. Salivary cortisol (sC) and testosterone (sT) of 24 professional players (27.8 ± 4.1 years of age) were collected before, after, and 24 hours following MD-5 in two comparable microcycles of T1 and T2. GPS training data (total and high-intensity distance) of the 7 and 28 days before sampling were used to obtain athletes' acute and chronic workloads. Results showed a pre-training significant decrease of sT and an increase of sC (p<0.05) in T2, compared to T1. Moreover, athletes showed high sC and low sT levels before, after and 24 hours following MD-5 in T2. Workload analysis revealed significant correlations of chronic load with sC (r = 0.45, p = 0.056) and T/C ratio (r = -0.59; p = 0.007). These results suggested that, in professional football, chronic workload has a greater impact on players' recovery time than acute workload over the sport season. Moreover, athletes' hormonal response to the weekly conditioning session at T2 revealed an altered anabolic/catabolic balance, highlighting the key role of continuous internal and external workload monitoring during the season.

Regulation of the Organic Anion Transporting Polypeptide 1B3 (OATP1B3) by Sex Hormones.

The mechanisms of regulation of the organic anion transporting polypeptide OATP1B3 by sex hormones were studied using HepG2 cells. Estradiol, progesterone, and testosterone were added to cells at concentrations of 1, 10, 100 µM for 24 h. The relative content of OATP1B3 was evaluated by Western blotting. Estradiol at concentrations of 10 and 100 µM increased the level of OATP1B3 acting through the farnesoid X-receptor, testosterone at concentrations of 1, 10, and 100 µM increased the expression of the transporter protein due to its effect on the liver X-receptor subtype α (LXRα), and progesterone did not affect the expression of OATP1B3.

Involvement of the AMPK Pathways in Muscle Development Disparities across Genders in Muscovy Ducks.

The differences in muscle development potential between male and female ducks lead to variations in body weight, significantly affecting the growth of the Muscovy duck meat industry. The aim of this study is to explore the regulatory mechanisms for the muscle development differences between genders. Muscovy ducks of both sexes were selected for measurements of body weight, growth traits, hormone levels, and muscle gene expression. The results show that male ducks compared to females had greater weight and growth traits (p < 0.05). Compared to male ducks, the level of serum testosterone in female ducks was decreased, and the estradiol levels were increased (p < 0.05). The RNA-seq analysis identified 102 upregulated and 49 downregulated differentially expressed genes. KEGG analysis revealed that among the top 10 differentially enriched pathways, the AMPK signaling pathway is closely related to muscle growth and development. Additionally, the mRNA and protein levels of CD36, CPT1A, LPL, and SREBP1 were increased and the P-AMPK protein level decreased in the female ducks compared to the male ducks (p < 0.05). In conclusion, muscle development potential difference between male and female ducks is regulated by sex hormones. This process is likely mediated through the activation of the AMPK pathway.

Metabolic benefits afforded by estradiol and testosterone in both sexes: clinical considerations.

Testosterone (T) and 17ß-estradiol (E2) are produced in male and female humans and are potent metabolic regulators in both sexes. When E2 and T production stops or decreases during aging, metabolic dysfunction develops and promotes degenerative metabolic and vascular disease. Here, we discuss the shared benefits afforded by E2 and T for metabolic function human females and males. In females, E2 is central to bone and vascular health, subcutaneous adipose tissue distribution, skeletal muscle insulin sensitivity, antiinflammatory immune function, and mitochondrial health. However, T also plays a role in female skeletal, vascular, and metabolic health. In males, T's conversion to E2 is fundamental to bone and vascular health, as well as prevention of excess visceral adiposity and the promotion of insulin sensitivity via activation of the estrogen receptors. However, T and its metabolite dihydrotestosterone also prevent excess visceral adiposity and promote skeletal muscle growth and insulin sensitivity via activation of the androgen receptor. In conclusion, T and E2 are produced in both sexes at sex-specific concentrations and provide similar and potent metabolic benefits. Optimizing levels of both hormones may be beneficial to protect patients from cardiometabolic disease and frailty during aging, which requires further study.

Supraphysiological Dose of Testosterone Impairs the Expression and Distribution of Sex Steroid Receptors during Endometrial Receptivity Development in Female Sprague-Dawley Rats.

This study aims to investigate the effect of a supraphysiological dose of testosterone on the levels of sex steroid hormones and the expression and distribution of sex steroid receptors in the uterus during the endometrial receptivity development period. In this study, adult female Sprague-Dawley rats (n = 24) were subcutaneously administered 1 mg/kg/day of testosterone alone or in combination with the inhibitors (finasteride or anastrozole or both) from day 1 to day 3 post-coitus, while a group of six untreated rats served as a control group. The rats were sacrificed on the evening of post-coital day 4 of to measure sex steroid hormone levels by ELISA. Meanwhile, gene expression and protein distribution of sex steroid receptors were analysed by quantitative polymerase chain reaction (qPCR) and immunohistochemistry (IHC), respectively. In this study, treatment with a supraphysiological dose of testosterone led to a significant reduction in oestrogen and progesterone levels compared to the control. The mRNA expression of the androgen receptor increased significantly in all treatment groups, while the mRNA expression of both the progesterone receptor and the oestrogen receptor-α decreased significantly in all treatment groups. The IHC findings of all sex steroid receptors were coherent with all mRNAs involved. This study shows that a supraphysiological dose of testosterone was able to interrupt the short period of the implantation window. This finding could serve as a basis for understanding the role of testosterone in endometrial receptivity in order to develop further therapeutic approaches targeting androgen-mediated disorders of endometrial receptivity.

Characterization of prostate macrophage heterogeneity, foam cell markers, and CXCL17 upregulation in a mouse model of steroid hormone imbalance.

Benign prostatic hyperplasia (BPH) is a prevalent age-related condition often characterized by debilitating urinary symptoms. Its etiology is believed to stem from hormonal imbalance, particularly an elevated estradiol-to-testosterone ratio and chronic inflammation. Our previous studies using a mouse steroid hormone imbalance model identified a specific increase in macrophages that migrated and accumulated in the prostate lumen where they differentiated into lipid-laden foam cells in mice implanted with testosterone and estradiol pellets, but not in sham animals. The current study focused on further characterizing the cellular heterogeneity of the prostate in this model as well as identifying the specific transcriptomic signature of the recruited foam cells. Moreover, we aimed to identify epithelia-derived signals that drive macrophage infiltration and luminal translocation. Male C57BL/6J mice were implanted with slow-release testosterone and estradiol pellets (T + E2) or sham surgery was performed and the ventral prostates were harvested two weeks later for scRNA-seq analysis. We identified Ear2 + and Cd72 + macrophages that were elevated in response to steroid hormone imbalance, whereas a Mrc1 + resident macrophage population did not change. In addition, an Spp1 + foam cell cluster was almost exclusively found in T + E2 mice. Further markers of foam cells were also identified, including Gpnmb and Trem2, and GPNMB was confirmed as a novel histological marker with immunohistochemistry. Foam cells were also shown to express known pathological factors Vegf, Tgfb1, Ccl6, Cxcl16 and Mmp12. Intriguingly, a screen for chemokines identified the upregulation of epithelia-derived Cxcl17, a known monocyte attractant, in T + E2 prostates suggesting that it might be responsible for the elevated macrophage number as well as their translocation to the lumen. Our study identified macrophage subsets that responded to steroid hormone imbalance as well as further confirmed a potential pathological role of luminal foam cells in the prostate. These results underscore a potential pathological role of the identified prostate foam cells and suggests CXCL17-mediated macrophage migration as a critical initiating event.

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.

Dipeptide metabolite, glutamyl-glutamate mediates microbe-host interaction to boost spermatogenesis.

The decrease in sperm count and infertility is a global issue that remains unresolved. By screening environmental bacterial isolates, we have found that a novel lactic acid bacterium, Lactiplantibacillus plantarum SNI3, increased testis size, testosterone levels, sperm count, sexual activity and fertility in mice that have consumed the bacteria for four weeks. The abundance of L. plantarum in the colon microbiome was positively associated with sperm count. Fecal microbiota transplantation (FMT) from L. plantarum SNI3-dosed mice improved testicular functions in microbiome-attenuated recipient animals. To identify mediators that confer pro-reproductive effects on the host, untargeted in situ mass spectrometry metabolomics was performed on testis samples of L. plantarum SNI3-treated and control mice. Enrichment pathway analysis revealed several perturbed metabolic pathways in the testis of treated mice. Within the testis, a dipeptide, glutamyl-glutamate (GluGlu) was the most upregulated metabolite following L. plantarum SNI3 administration. To validate the pro-reproductive feature of GluGlu, systemic and local injections of the dipeptide have been performed. γ-GluGlu increased sperm count but had no effect on testosterone. These findings highlight the role of γ-GluGlu in mediating spermatogenetic effects of L. plantarum on the male mouse host and -following relevant human clinical trials- may provide future tools for treating certain forms of male infertility.

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.

Adolescent exposure to bisphenol-a antagonizes androgen regulation of social behavior in male mice.

Social behavior is sexually dimorphic, which is regulated by gonadal hormones in the brain. Our recent study found that exposure to low doses of bisphenol-A (BPA) during adolescence, permanently alters social behavior in adult male mice, but the underlying mechanisms remain unclear. Using adolescent gonadectomy (GDX) male mice with testosterone propionate (TP, 0.5 mg/kg) supplement (TP-GDX), this study showed that BPA antagonized promoting effects of TP on social interaction, sexual behavior, and aggression in GDX mice. BPA eliminated the reversal effects of TP on GDX-induced decrease in the number of immunoreactive to arginine vasopressin (AVP-ir) neurons in the medial amygdala (MeA) and the levels of AVP receptor 1a (V1aR) in the MeA and the nucleus accumbens (NAc). In addition, BPA removed down-regulation in the levels of dopamine (DA) transporter (DAT) and DA receptor 1 (DR1) in the NAc of TP-GDX mice. BPA exposure reduced testosterone (T) levels in the brain and serum and the expression of androgen receptor (AR) protein in the amygdala and striatum of sham-operated and TP-GDX males. These results suggest that adolescent exposure to BPA inhibits regulation of androgen in AVP and DA systems of the brain regions associated with social behavior, and thus alters social behaviors of adult male mice.

Chitinase 3-Like 1 and C-X-C motif chemokine ligand 5 proteins and the hair cycle.

Dermal papilla cells (DPCs) exhibit self-recovery ability, which may be involved in hair growth. Therefore, we tested whether DPCs subjected to temporary growth-inhibiting stress (testosterone, 17ß-estradiol, mitomycin C, or undernutrition) treatments exhibit self-recovery behavior that can activate hair follicle growth, and examined the changes in cell proliferation capacity and gene expression. Related proteins were identified and their relationships with the hair cycle was examined using a mouse model. Recovery-period DPCs (i.e., from day 3 after loading) were subjected to microarray analysis to detect genetic variations common to each stress treatment. Co-culture of recovery-period DPCs and outer root sheath cells (ORSCs) confirmed the promotion of ORSC proliferation, suggesting that the activation of hair follicle growth is promoted via signal transduction. Chitinase 3-like 1 (CHI3L1) and C-X-C motif chemokine 5 (CXCL5) exhibited ORSC proliferation-promoting effects. Measurement of protein content in the skin during each phase of the hair cycle in mice revealed that CHI3L1 and CXCL5 secretion increased immediately after anagen transition. In a hair-loss mouse model treated with testosterone or 17ß-estradiol, CHI3L1 and CXCL5 secretion was lower in treated telogen skin than in untreated skin. Our results suggest that CHI3L1 and CXCL5 secreted by recovery-state DPCs promote hair growth.

Hexavalent chromium reduces testosterone levels by impairing lipophagy and disrupting lipid metabolism homeostasis: Based on a metabolomic analysis.

Hexavalent chromium (Cr(VI)) causes testicular damage and reduces testosterone secretion. Testosterone synthesis relies on cholesterol as a raw material, and its availability can be affected by lipophagy. However, the role of lipophagy in Cr(VI)-induced testicular damage and reduced testosterone secretion remains unclear. In this study, we investigated the effect of Cr(VI) on lipid metabolism and lipophagy in the testes of ICR mice. Forty mice were randomly divided into four groups and exposed to different doses of Cr(VI) (0, 75, 100, 125 mg/kg) for thirty days. Cr(VI) increased the rate of sperm abnormalities, decreased testosterone level, and decreased the levels of testosterone synthesis-related proteins, namely steroidogenic acute regulatory (StAR) and 3ß-hydroxysteroid dehydrogenase (3ß-HSD) proteins. Through metabolomic analysis, Oil Red O staining, and biochemical indicator (triglyceride and total cholesterol) analysis, Cr(VI) was found to disrupt testicular lipid metabolism. Further investigation revealed that Cr(VI) inhibited the AMP-activated protein kinase (AMPK)/sterol regulatory element-binding protein 1 (SREBP1) pathway, elevated levels of the autophagy-related proteins microtubule-associated protein 1 light chain 3B (LC3B) and sequestosome 1 (SQSTM1)/P62 and lipophagy-related proteins Rab7 and Rab10, while increasing colocalization of LC3B and Perilipin2. These findings suggest that Cr(VI) exposure leads to abnormal lipid metabolism in the testes by suppressing the AMPK/SREBP1 pathway and disrupting lipophagy, ultimately reducing testosterone level and inducing testicular damage.

New Insights into D-Aspartate Signaling in Testicular Activity.

D-aspartate (D-Asp) is an amino acid found in high concentrations in the testis and pituitary gland. Increasing evidence suggests that D-Asp promotes spermatogenesis by activating testosterone production in the Leydig cells via LH release from the pituitary gland. In vitro studies indicate that D-Asp may also influence steroidogenesis and spermatogenesis through autocrine and paracrine signals. D-Asp enhances StAR and steroidogenic enzyme expressions, facilitating testicular cell proliferation via the GluR/ERK1/2 pathway. Moreover, it supports spermatogenesis by enhancing the mitochondrial function in spermatocytes, aiding in the metabolic shift during meiosis. Enhanced mitochondrial function, along with improved MAM stability and reduced ER stress, has been observed in Leydig and Sertoli cells treated with D-Asp, indicating potential benefits in steroidogenesis and spermatogenesis efficiency. Conversely, D-Asp exerts a notable anti-apoptotic effect in the testis via the AMPAR/AKT pathway, potentially mediated by antioxidant enzyme modulation to mitigate testicular oxidative stress. This review lays the groundwork for future investigations into the molecules promoting spermatogenesis by stimulating endogenous testosterone biosynthesis, with D-amino acids emerging as promising candidates.

Screening persistent organic pollutants for effects on testosterone and estrogen synthesis at human-relevant concentrations using H295R cells in 96-well plates.

Many persistent organic pollutants (POPs) are suspected endocrine disruptors and it is important to investigate their effects at low concentrations relevant to human exposure. Here, the OECD test guideline #456 steroidogenesis assay was downscaled to a 96-well microplate format to screen 24 POPs for their effects on viability, and testosterone and estradiol synthesis using the human adrenocortical cell line H295R. The compounds (six polyfluoroalkyl substances, five organochlorine pesticides, ten polychlorinated biphenyls and three polybrominated diphenyl ethers) were tested at human-relevant levels (1 nM to 10 µM). Increased estradiol synthesis, above the OECD guideline threshold of 1.5-fold solvent control, was shown after exposure to 10 µM PCB-156 (153%) and PCB-180 (196%). Interestingly, the base hormone synthesis varied depending on the cell batch. An alternative data analysis using a linear mixed-effects model that include multiple independent experiments and considers batch-dependent variation was therefore applied. This approach revealed small but statistically significant effects on estradiol or testosterone synthesis for 17 compounds. Increased testosterone levels were demonstrated even at 1 nM for PCB-74 (18%), PCB-99 (29%), PCB-118 (16%), PCB-138 (19%), PCB-180 (22%), and PBDE-153 (21%). The MTT assay revealed significant effects on cell viability after exposure to 1 nM of perfluoroundecanoic acid (12%), 3 nM PBDE-153 (9%), and 10 µM of PCB-156 (6%). This shows that some POPs can interfere with endocrine signaling at concentrations found in human blood, highlighting the need for further investigation into the toxicological mechanisms of POPs and their mixtures at low concentrations relevant to human exposure.