Leydig Cells Of Testis Research Articles

Amyloids facilitate DNA transfection in vivo

Amyloid fibril deposits are a main source of pathology in neurodegenerative diseases. Normal proteins such as tau, alpha-synuclein, TDP-43 and others could form specific conformational fibrils called amyloid, which deposited in the brains of neurodegenerative diseases. Although the pathological roles of amyloids in cell death have been discussed a lot, their other functions have not been investigated well. Here, we studied the effect of amyloids on DNA transfection in vivo. We injected quantum dot labeled or non-labeled amyloid-preformed fibrils (PFFs) and a green fluorescent protein (EGFP) expression vector into organs including brain, testis, liver and calf muscle. GFP expression patterns were examined by immunohistochemistry and western blotting. At 24 h after injection, EGFP was predominantly expressed in the neurons in the cortex and the striatum, Leydig cells in testis, hepatocytes in the liver and muscle cells. EGFP expression was inhibited by an endocytosis inhibitor, sertraline in the brain and testis. The amyloid-PFFs potentiated Ca2+ transients shown by calcium imaging and EGFP expression in the brain was blocked by Ca blocker, cilnidipine. Our results show that amyloid-PFFs facilitate DNA transfection and can be used for a new gene delivery system in vivo.

Ontogeny of mouse Sertoli, Leydig and peritubular myoid cells from embryonic day 10 to adulthood

We present a comprehensive description of the differentiating somatic cell types (Sertoli, Leydig, and peritubular myoid cells) of the mouse testis from embryonic day 10.5 (E10.5) to adulthood, postnatal day 60 (P60). Immunohistochemistry was used to analyze expression of: Sox9 (a Sertoli cell marker), 3βHSD-1 (a fetal Leydig cell marker), 3βHSD-6 (an adult Leydig cell marker), α-actin (a peritubular myoid cell marker), and androgen receptor (a marker of all three somatic cell types). The temporal-spatial expression of these markers was used to interrogate findings of earlier experimental studies on the origin of Sertoli, Leydig and peritubular myoid cells, as well as extend previous descriptive studies across a broader developmental period (E10.5-P60). Such comparisons demonstrate inconsistencies that require further examination and raise questions regarding conservation of developmental mechanisms across higher vertebrate species.

Reproductive Function of Male Rats and Motor Activity of Their Offspring in Fire Emissions Modeling.

We studied the reproductive function of albino male rats after long-term exposure to wildfire smoke and the species-specific behavior of their sexually mature offspring. After the end of exposure, the mean number of spermatogonia and the relative number of Leydig cells in rat testes decreased and the index of spermatogenesis decreased. The offspring of exposed males mated with intact females demonstrated impaired cognitive abilities and disturbances of species-specific behavior. The severity of these changes depended on time interval between the end of the inhalation exposure and mating.

Transient Hypogonadism Induced by SARS-CoV-2 during COVID-19 in Men Aged under 65 Years

Background: The presence of the angiotensin-converting enzyme 2 (ACE2) receptor, the main entry receptor for SARS-CoV-2 in lung alveolar tissue, in Sertoli and Leydig cells of the adult testis, may suggest possible testicular involvement during SARS-CoV-2 infection. Our aim was to investigate the impact of COVID-19 on gonadal function in men. Methods: This was a cross-sectional descriptive and analytical study in a population of men aged below 65 years of age with SARS-CoV-2. Not included in the study were any subjects on testosterone replacement therapy or with a known condition that could create hypogonadism. We recruited patients through a questionnaire and then performed total testosterone and SHBG analysis at 8 hours and 2 months post infection by ELISA. We used the Spearman Rho test for statistical analysis of correlations. The significance level was set at 0.05. Results: The sample consisted of 40 male COVID positive patients with a mean age of 46.4 ± 11.8 years. Eight patients were reviewed after infection. The mean total testosterone was 11 ng/ml ± 2.4 and the SHBG was 113 nmol/l ± 66.9 during infection. In the 8 patients reviewed after infection, median total testosterone decreased during infection (11 ng/ml) and increased after infection (12.7 ng/ml), this result was statistically significant (P = 0.028). Median SHBG during infection was increased (115.7 nmol/l) and after infection was decreased (82 nmol/l). There was a statistically significant (P = 0.04) and positive correlation between serum testosterone and SHBG levels in patients with severe infection. Conclusion: There could be a transient relative hypogonadism during SARS-CoV-2 infection, more marked in the severe forms of the disease with a tendency to improve after infection.

MeCP2 duplication causes hyperandrogenism by upregulating LHCGR and downregulating ROR\u03b1

Duplication of MECP2 (methyl-CpG-binding protein 2) gene causes a serious neurological and developmental disorder called MECP2 duplication syndrome (MDS), which is usually found in males. A previous clinical study reported that MDS patient has precocious puberty with hyperandrogenism, suggesting increased MeCP2 may cause male hyperandrogenism. Here we use an MDS mouse model and confirm that MECP2 duplication significantly upregulates androgen levels. We show for the first time that MeCP2 is highly expressed in the Leydig cells of testis, where androgen is synthesized. Mechanistically, MECP2 duplication increases androgen synthesis and decreases androgen to estrogen conversion through either the upregulation of luteinizing hormone receptor (LHCGR) in testis, as a result of MeCP2 binds to G-quadruplex structure of Lhcgr promoter and recruits the transcription activator CREB1 or the downregulation of the expression of aromatase in testis by binding the CpG island of Rorα, an upstream regulator of aromatase. Taken together, we demonstrate that MeCP2 plays an important role in androgen synthesis, supporting a novel non-CNS function of MeCP2 in the process of sex hormone synthesis.

Quercetin ameliorates testosterone secretion disorder by inhibiting endoplasmic reticulum stress through the miR-1306-5p/HSD17B7 axis in diabetic rats.

Testicular damage and testosterone secretion disorder are associated with diabetes mellitus. Quercetin, a common flavonoid, has antioxidant, anti-cancer, and blood sugar lowering effects. Therefore, this study aims to investigate the effect of quercetin on the reproductive system of male rats with diabetes in vivo and in vitro and elucidate its mechanism. Streptozotocin (STZ) induction was used to establish a diabetes model in 40 male Sprague Dawley rats, which were subsequently administered with 20 or 50 mg/kg of quercetin. Leydig cells of rat testes were treated by high glucose (HG) followed by 5 or 10 μM quercetin. Two doses of quercetin increased rat body weight and testicular weight, decreased blood glucose, and inhibited oxidative stress. Quantitative real-time polymerase chain reaction and Western blotting revealed that quercetin alleviated STZ-induced testicular damage and promoted testosterone synthesis. Both doses of quercetin reduced reactive oxygen species and malondialdehyde levels, and increased superoxide dismutase level in HG-treated cells. Both, in vivo and in vitro results confirmed that a high dose of quercetin was more effective. MiR-1306-5p was upregulated in testicular tissue of diabetic rats and HG-treated cells. 17β-hydroxysteroid dehydrogenase (HSD17B7) was a target of miR-1306-5p and HSD17B7 was downregulated in STZ-induced rat tissues and HG-treated cells. HSD17B7 overexpression reversed the increase of C/EBP homologous protein (CHOP) and glucose-regulated protein 78 (Grp78) protein levels as well as eIF2α phosphorylation level and promotion of cell apoptosis caused by miR-1306-5p overexpression. Moreover, overexpression of HSD17B7 activated the Janus kinase 2/signal transducer and activator of transcription 3 axis in HG-treated cells. In conclusion, quercetin inhibits endoplasmic reticulum stress and improves testosterone secretion disorder through the miR-1306-5p/HSD17B7 axis in diabetic rats.

DAPL1 is a novel regulator of testosterone production in Leydig cells of mouse testis

Leydig cells in the testes produce testosterone in the presence of gonadotropins. Therefore, male testosterone levels must oscillate within a healthy spectrum, given that elevated testosterone levels augment the risk of cardiovascular disorders. We observed that the expression of death-associated protein-like 1 (DAPL1), which is involved in the early stages of epithelial differentiation and apoptosis, is considerably higher in the testes of sexually mature mice than in other tissues. Accordingly, Dapl1-null mice were constructed to evaluate this variation. Notably, in these mice, the testicular levels of steroidogenic acute regulatory protein (StAR) and serum testosterone levels were significantly elevated on postnatal day 49. The findings were confirmed in vitro using I-10 mouse testis-derived tumor cells. The in vivo and in vitro data revealed the DAPL1-regulated the expression of StAR involving altered transcription of critical proteins in the protein kinase A and CREB/CREM pathways in Leydig cells. The collective findings implicate DAPL1 as an important factor for steroidogenesis regulation, and DAPL1 deregulation may be related to high endogenous levels of testosterone.

O-037 The impact of COVID-19 on male fertility: what do we know?”

Abstract text Since the very early days of the COVID-19 pandemic, concern has been raised about the possibility of damage to the male reproductive system in those men who are infected with SARS-CoV-2. This was first raised by the early observation (January 2020) that the route of infection was via the Angiotensin Converting Enzyme 2 (ACE-2) receptor present on host cells. ACE-2 was first described in the year 2000 and subsequently shown in 2004 to be selectively expressed in the Leydig cells of the adult testis. This opens up the possibility that although COVID-19 is primarily a respiratory infection, it may also infect the male reproductive system. The authors of several review articles have proposed that male fertility may be theoretically impacted by SARS-CoV-2 in a number of ways. This includes alteration of: (i) testicular architecture; (ii) reproductive hormone profiles (LH/FSH); (iii) spermatogenesis as evidenced by changes to ejaculate quality; (iv) sperm function (e.g., DNA damage); (v) sexual/erectile function; or a combination of all five. Clearly each of these individually may impact on the chance of pregnancy or live birth either in natural or medically assisted reproduction. There is also the possibility that SARS-CoV-2 may be transmitted sexually if sufficient numbers of SARS-CoV-2 are found in semen. Reassuringly, of 14 studies published to date, there is little evidence to suggest that SARS-CoV-2 is present in semen and so the possibility of sexual transmission in patient or donor samples can probably be discounted. However, there is currently an incomplete picture of whether semen quality is affected by SARS-CoV-2 infection as studies are often limited by the fact that no pre-infection control samples are available for direct comparison or they are too short to identify any long-term effects. Nevertheless, the few case-controlled studies published which compare semen quality infected with non-infected (control) individuals suggest that there may be a statistically significant alteration in sperm concentration and motility, although it is not clear whether this is linked to infection by the SARS-CoV-2 virus or simply a consequence of febrile illness and fever (or medication given to combat the infection). There is currently a lack of long-term data on any impact of COVID-19 on male reproductive hormones (although much speculation about the role that testosterone might play in the severity of disease). There are also emerging reports of increased risk of erectile dysfunction in men following SARS-CoV-2. In terms of birth rates, it is simply too early to tell whether these have been affected by the pandemic, given the possibility of lockdown affecting sexual behaviour in fertile couples and IVF clinic closures in infertile couples. In conclusion, although the COVID-19 pandemic has infected over 130 million people worldwide we still know too little about the impact of SARS-CoV-2 on the male reproductive system. Given the incidence of long-COVID, and the asymptomatic nature of the infection for some, it is important to commission and conduct long-term studies which can monitor the reproductive outcomes of young men who have survived a SARS-CoV-2 infection.

Marsilea crenata ethanol extract prevents monosodium glutamate adverse effects on the serum levels of reproductive hormones, sperm quality, and testis histology in male rats

Background and Aim:Marsilea crenata is an aquatic plant that contains high antioxidants level and could prevent cell damages caused by free radicals. The present study aimed to investigate the effect of M. crenata ethanol extract on luteinizing hormone (LH), testosterone levels, sperm quality, and testis histology of adult male rats induced by monosodium glutamate (MSG).Materials and Methods:This study randomly divided 48 male rats into eight groups (n=6): control group; MSG group (4 mg/g body weight [b.w.] for 30 days); MS1, MS2, and MS3 groups (4 mg/g b.w. MSG and M. crenata ethanol extract at dose 0.216, 0.432, and 0.648 mg/g b.w., respectively, for 30 days); and S1, S2, and S3 groups (M. crenata ethanol extract at dose of 0.216, 0.432, and 0.648 mg/g b.w., respectively, for 30 days). The blood sample was collected on days 0 and 30 to determine the LH and testosterone levels. The animals were dissected on day 30, and the testes were isolated for morphometric, histology (spermatogenic cell number), and malondialdehyde (MDA) examination. Moreover, semen was collected to determine the sperm quality parameter.Results:The LH and testosterone levels significantly increased (p<0.05) after M. crenata administration at all doses. The higher dose of M. crenata ethanol extract demonstrated a high decrease in MDA level in MSG-treated rat testis; increase of spermatogonia, spermatocytes, spermatids, and Leydig cells number; and increase of seminiferous tubular diameter and germinal epithelium thickness.Conclusion:The ethanol extract of M. crenata can improve the levels of LH, testosterone, sperm quality, number of testis morphometric, spermatogenic, and Leydig cells in MSG-treated male rats.

Histopathological changes associated with exposure of some viscerals and testicular tissues to bisephenol A and di(2-ethylhexyl)phthalate

Purpose: To evaluate the pathological effect associated with the exposure of viscerals and testicular tissues of albino rats to BPA and DEHP.&#x0D; Methods: A total of sixty adult male Wistar rats weighing 200 – 250 g were divided into four (4) groups of 15 rats per cage in a sanitized environment. Group I animals received normal rodent pellet and water and served as untreated group. Thereafter, the rats in the second group (II) were administered 5 mg/kg per day of BPA mixed with rodent pellet orally; group III received 0.5 mg/kg daily dose of DEHP mixed with rodent pellet; while group IV received orally mixture of 0.5 mg/kg of BPA and 0.5 mg/kg DEHP + pelleted rodent feed. The rats were fed and water provided regularly for 30 days; finally, the animals were weighed and sacrificed by cranial dislocation.&#x0D; Result: BPA and DEHP resulted in significant (p &lt; 0.05) weight losses, inflammation and fatty deposits in the liver, degenerated bowman capsule and glomeruli of the kidneys, lung oedema, and deteriorated leydig cells in testes.&#x0D; Conclusion: The results suggest that BPA and DEHP cause significant weight loss and are injurious to the cellular make-up of rat tissues, which may be the same in higher animals.

Expression and localisation of ephrin-B1 and EphB4 in steroidogenic cells in the naturally cycling mouse ovary

Ephrin receptors and ligands are membrane-bound molecules that modulate diverse cellular functions such as cell adhesion, epithelial–mesenchymal transition, motility, differentiation and proliferation. We recently reported the co-expression of ephrin-B1 and EphB4 in adult and foetal Leydig cells of the mouse testis, and thus speculated that their co-expression is a common property in gonadal steroidogenic cells. Therefore, in this study we examined the expression and localisation of ephrin-B1 and EphB4 in the naturally cycling mouse ovary, as their expression patterns in the ovary are virtually unknown. We found that ephrin-B1 and EphB4 were co-expressed in steroidogenic cells of all kinds, i.e. granulosa cells and CYP17A1-positive steroidogenic theca cells as well as in 3β-HSD-positive luteal cells and the interstitial glands; their co-expression potentially serves as a good marker to identify sex steroid-producing cells even in extra-gonadal organs/tissues. We also found that ephrin-B1 and EphB4 expression in granulosa cells was faint and strong, respectively; ephrin-B1 expression in luteal cells was weak in developing and temporally mature corpora lutea (those of the current cycle) and likely strong in regressing corpora lutea (those of the previous cycle) and EphB4 expression in luteal cells was weak in corpora lutea of the current cycle and likely faint/negative in the corpora lutea of the previous cycle. These findings suggest that a luteinising hormone surge triggers the upregulation of ephrin-B1 and downregulation of EphB4, as this expression fluctuation occurs after the surge. Overall, ephrin-B1 and EphB4 expression patterns may represent benchmarks for steroidogenic cells in the ovary.

Effects of SARS-CoV-2 on Cardiovascular System: The Dual Role of Angiotensin-Converting Enzyme 2 (ACE2) as the Virus Receptor and Homeostasis Regulator-Review.

Angiotensin-converting enzyme 2 (ACE2) is the entry receptor for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the cause of Coronavirus Disease-2019 (COVID-19) in humans. ACE-2 is a type I transmembrane metallocarboxypeptidase expressed in vascular endothelial cells, alveolar type 2 lung epithelial cells, renal tubular epithelium, Leydig cells in testes and gastrointestinal tract. ACE2 mediates the interaction between host cells and SARS-CoV-2 spike (S) protein. However, ACE2 is not only a SARS-CoV-2 receptor, but it has also an important homeostatic function regulating renin-angiotensin system (RAS), which is pivotal for both the cardiovascular and immune systems. Therefore, ACE2 is the key link between SARS-CoV-2 infection, cardiovascular diseases (CVDs) and immune response. Susceptibility to SARS-CoV-2 seems to be tightly associated with ACE2 availability, which in turn is determined by genetics, age, gender and comorbidities. Severe COVID-19 is due to an uncontrolled and excessive immune response, which leads to acute respiratory distress syndrome (ARDS) and multi-organ failure. In spite of a lower ACE2 expression on cells surface, patients with CVDs have a higher COVID-19 mortality rate, which is likely driven by the imbalance between ADAM metallopeptidase domain 17 (ADAM17) protein (which is required for cleavage of ACE-2 ectodomain resulting in increased ACE2 shedding), and TMPRSS2 (which is required for spike glycoprotein priming). To date, ACE inhibitors and Angiotensin II Receptor Blockers (ARBs) treatment interruption in patients with chronic comorbidities appears unjustified. The rollout of COVID-19 vaccines provides opportunities to study the effects of different COVID-19 vaccines on ACE2 in patients on treatment with ACEi/ARB.

The Presence of Colony-Stimulating Factor-1 and Its Receptor in Different Cells of the Testis; It Involved in the Development of Spermatogenesis In Vitro.

Spermatogenesis is a complex process, in which spermatogonial cells proliferate and differentiate in the seminiferous tubules of the testis to generate sperm. This process is under the regulation of endocrine and testicular paracrine/autocrine factors. In the present study, we demonstrated that colony stimulating factor-1 (CSF-1) is produced by mouse testicular macrophages, Leydig, Sertoli, peritubular cells and spermatogonial cells (such as CDH1-positively stained cells; a marker of spermatogonial cells). In addition, we demonstrated the presence of CSF-1 and its receptor (CSF-1R) in testicular macrophages, Leydig, Sertoli, peritubular cells and spermatogonial cells of human testis. We also show that the protein levels of CSF-1 were the highest in testis of 1-week-old mice and significantly decreased with age (2–12-week-old). However, the transcriptome levels of CSF-1 significantly increased in 2–3-week-old compared to 1-week-old, and thereafter significantly decreased with age. On the other hand, the transcriptome levels of CSF-1R was significantly higher in mouse testicular tissue of all examined ages (2–12-week-old) compared to 1-week-old. Our results demonstrate the involvement of CSF-1 in the induction the proliferation and differentiation of spermatogonial cells to meiotic and postmeiotic stages (BOULE- and ACROSIN-positive cells) under in vitro culture conditions, using methylcellulose culture system (MCS). Thus, it is possible to suggest that CSF-1 system, as a testicular paracrine/autocrine system, is involved in the development of different stages of spermatogenesis and may be used in the development of future therapeutic strategies for treatment of male infertility.

Systematic evaluation of CdSe/ZnS quantum dots toxicity on the reproduction and offspring health in male BALB/c mice

Increased applications of quantum dots (QDs) in the biomedical field have aroused attention for their potential toxicological effects. Although numerous studies have been carried out on the toxicity of QDs, their effects on reproductive and development are still unclear. In this study, we systematically evaluated the male reproductive toxicity and developmental toxicity of CdSe/ZnS QDs in BALB/c mice. The male mice were injected intravenously with CdSe/ZnS QDs at the dosage of 2.5 mg/kg BW or 25 mg/kg BW, respectively, and the survival status, biodistribution of QDs in testes, serum sex hormone levels, histopathology, sperm motility and acrosome integrity was measured on Day 1, 7, 14, 28 and 42 after injection. On Day 35 after treatment, male mice were housed with non-exposed female mice, and then offspring number, body weight, organ index and histopathology of major organs, blood routine and biochemical tests of offspring were measured to evaluate the fertility and offspring health. The results showed that CdSe/ZnS QDs could rapidly distribute in the testis, and the fluorescence of QDs could still be detected on Day 42 post-injection. QDs had no adverse effect on the structure of testis and epididymis, but high-dose QDs could induce apoptosis of Leydig cells in testis at an early stage. No significant differences in survival of state, body weight organ index of testis and epididymis, sex hormones levels, sperm quality, sperm acrosome integrity and fertility of male mice were observed in QDs exposed groups. However, the development of offspring was obviously influenced, which was mainly manifested in the slow growth of offspring, changes in organ index of main organs, and the abnormality of liver and kidney function parameters. Our findings revealed that CdSe/ZnS QDs were able to cross the blood-testis barrier (BTB), produce no discernible toxic effects on the male reproductive system, but could affect the healthy growth of future generations to some extent. In view of the broad application prospect of QDs in biomedical fields, our findings might provide insight into the biological safety evaluation of the reproductive health of QDs.

Secretome improves testosterone and androgen-binding protein production in testicular dysfunction rats induced by cisplatin.

Objective:This study was determined to see the effects of secretome on cellular production of testosterone and androgen-binding protein (ABP) using immunohistochemistry in testicle dysfunction due to cisplatin.Materials and Methods:Forty-eight rats were divided into four groups: the healthy group, the group with testicular dysfunction, the secretome-treated group with 0.2 ml/kg body weight (BW), and the secretome-treated group with 0.3 ml/kg BW. The immunohistochemistry staining method is used to find out testosterone and ABP reactivity in tissue organs.Results:Very strong testosterone and ABP immunoreactivity were found in Leydig cells of normal testes. While in the Leydig cells of cisplatin-induced testicles, testosterone and ABP immunoreactivity were not observed. Testosterone and ABP were observed 1 week after the second secretome injection. The number of testosterone-immunoreactive cells in the low dose group from week 1 to 3 was 0, 19, and 32, respectively. From week 1 until week 4, the high dose group was 0, 29, 33, and 65, respectively. The number of ABP-immunoreactive cells from the first week until the third week in the low dose group was 0, 28, and 34, respectively. The high dose group from the first week until the fourth week was 0, 26, 58, and 83, respectively. The number of cells that produce testosterone and ABP increased from week 2 to 4.Conclusion:The administration of secretome could increase the number of immunoreactive cells toward testosterone and ABP in testicular dysfunction.

Fshr: a fish sex-determining locus shows variable incomplete penetrance across flathead grey mullet populations.

SummaryWhole-genome sequencing data were produced from a single flathead grey mullet female and assembled into a draft genome sequence, whereas publicly available sequence data were used to obtain a male draft sequence. Two pools, each consisting of 60 unrelated individuals, respectively, of male and female fish were analyzed using Pool-Sequencing. Mapping and analysis of Pool-Seq data against the draft genome(s) revealed >30 loci potentially associated with sex, the most promising locus of which, encoding the follicle-stimulating hormone receptor (fshr) and harboring two missense variants, was genotyped on 245 fish from four Mediterranean populations. Genotype data showed that fshr represents a previously unknown sex-determining locus, although the incomplete association pattern between fshr genotype and sex-phenotype, the variability of such pattern across different populations, and the presence of other candidate loci reveal that a greater complexity underlies sex determination in the flathead grey mullet.

Zearalenone perturbs the circadian clock and inhibits testosterone synthesis in mouse Leydig cells

ABSTRACT Zearalenone (ZEA), a mycotoxin, is known to impair reproductive capability by disrupting the synthesis and secretion of testosterone by Leydig cells (LCs), although the mechanism is unknown. Robust rhythmicity of circadian clock and steroidogenic genes were identified in LCs. The aim of this study was to examine whether ZEA significantly attenuated the transcription of core clock genes (Bmal1, Dbp, Per2, and Nr1d1) as well as steroidogenic genes (StAR, Hsd3b2, and Cyp11a1) in mouse testis Leydig cell line (TM3). Western blotting confirmed declines in BMAL1, NR1D1, and StAR protein levels. ZEA also suppressed secreted testosterone levels. In primary LCs, isolated from PER2::LUCIFERASE reporter gene knock in mice, ZEA diminished the amplitude of PER2::LUC expression, and induced a phase shift and period extension. In primary LCs, ZEA also suppressed the expression levels of core clock and steroidogenic genes, reduced protein levels of BMAL1, and decreased testosterone secretion. In vivo expression of core clock and steroidogenic genes were reduced in testes of mice exposed to ZEA for 1 week leading to decreased serum testosterone levels. In summary, data suggest that ZEA may impair testosterone synthesis through attenuation of the circadian clock in LCs culminating in reproductive dysfunction in male mammals .

Neurotrophin-3 stimulates stem Leydig cell proliferation during regeneration in rats.

Neurotrophin‐3 (NT‐3) acts as an important growth factor to stimulate and control tissue development. The NT‐3 receptor, TRKC, is expressed in rat testis. Its function in regulation of stem Leydig cell development and its underlying mechanism remain unknown. Here, we reported the role of NT‐3 to regulate stem Leydig cell development in vivo and in vitro. Ethane dimethane sulphonate was used to kill all Leydig cells in adult testis, and NT‐3 (10 and 100 ng/testis) was injected intratesticularly from the 14th day after ethane dimethane sulphonate injection for 14 days. NT‐3 significantly reduced serum testosterone levels at doses of 10 and 100 ng/testis without affecting serum luteinizing hormone and follicle‐stimulating hormone levels. NT‐3 increased CYP11A1‐positive Leydig cell number at 100 ng/testis and lowered Leydig cell size and cytoplasmic size at doses of 10 and 100 ng/testis. After adjustment by the Leydig cell number, NT‐3 significantly down‐regulated the expression of Leydig cell genes (Lhcgr, Scarb1, Star, Cyp11a1, Hsd3b1, Cyp17a1, Hsd17b3, Hsd11b1, Insl3, Trkc and Nr5a1) and the proteins. NT‐3 increased the phosphorylation of AKT1 and mTOR, decreased the phosphorylation of 4EBP, thereby increasing ATP5O. In vitro study showed that NT‐3 dose‐dependently stimulated EdU incorporation into stem Leydig cells and inhibited stem Leydig cell differentiation into Leydig cells, thus leading to lower medium testosterone levels and lower expression of Lhcgr, Scarb1, Trkc and Nr5a1 and their protein levels. NT‐3 antagonist Celitinib can antagonize NT‐3 action in vitro. In conclusion, the present study demonstrates that NT‐3 stimulates stem Leydig cell proliferation but blocks the differentiation via TRKC receptor.

Bisphenol A alters differentiation of Leydig cells in the rabbit fetal testis

The endocrine disruptor Bisphenol A (BPA) crosses the placental barrier and reaches the fetal organs, including the gonads. In the testis, fetal Leydig cells (FLC) produce testosterone required for the male phenotype and homeostatic cell-cell signaling in the developing testis. Although it is known that BPA affects cell proliferation and differentiation in FLC, results concerning the mechanism involved are contradictory, mainly due to differences among species. Fast developing fetal gonads of rodents lack cortex and medulla, whereas species with more extended gestation periods form these two tissue compartments. The rabbit provides a good subject for studying the disruptive effect of BPA in fetal Leydig and possible postnatal endocrine consequences in adult Leydig cells. Here, we investigated the impact of BPA administered to pregnant rabbits on the FLC population of the developing testes. Using qRT-PCR, we assessed the levels of SF1, CYP11A1, 3β-HSD, and androgen receptor genes, and levels of fetal serum testosterone were measured by ELISA. These levels correlated with both the mitotic activity and the ultrastructural differentiation of the FLC by confocal and electron microscopy, respectively. Results indicate that BPA alters the expression levels of essential genes involved in androgen paracrine signaling, modifies the proliferation and differentiation of the FLCs, and alters the levels of serum testosterone after birth. Thus, BPA may change the postnatal levels of serum testosterone due to the impaired FLC population formed by the proliferating stem and non-proliferating cytodifferentiated FLC.

Expression of the Insulin-like Growth Factor System in First- and Second-Trimester Human Embryonic and Fetal Gonads.

Insulin-like growth factor (IGF) signaling is crucial for sex differentiation and development of Leydig and Sertoli cells in fetal mice testes. No such information is available for human embryonic and fetal testes and ovaries. To investigate presence and activity of the IGF signaling system during human embryonic and fetal ovarian and testicular development. Human embryonic and fetal gonads were obtained following legal terminations of pregnancies. Gene expression was assessed by microarray and qPCR transcript analyses. Proteins of the IGF system components were detected with immunohistochemistry and immunofluorescence analyses. Specimens were included from 2010 to 2017. University Hospital. Ovaries and testes from a total of 124 human embryos and fetuses aged 5 to 17 postconception weeks were obtained from healthy women aged 16 to 47 years resident in Denmark or Scotland. Gene expression analysis using microarray was performed in 46 specimens and qPCR analysis in 56 specimens, both sexes included. Protein analysis included 22 specimens (11 ovaries, 11 testes). IGF system members were detected in embryonic and fetal testes and ovaries, both at gene transcript and protein level. A higher expression of IGF regulators was detected in testes than ovaries, with a preferred localization to Leydig cells. These data indicate that the IGF system is active during very early gestation, when it may have a regulatory role in Leydig cells.