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BACKGROUND: Disorders/differences of sex development (DSD) are congenital conditions in which the development of chromosomal, gonadal, or anatomical sex is atypical. With overlapping phenotypes and multiple genes involved, poor diagnostic yields are achieved for many of these conditions. The current DSD diagnostic regimen can be augmented by investigating transcriptome/proteome in vivo, but it is hampered by the unavailability of affected gonadal tissue at the relevant developmental stage. We try to mitigate this limitation by reprogramming readily available skin tissue-derived dermal fibroblasts into Sertoli cells (SC), which could then be deployed for different diagnostic strategies. SCs form the target cell type of choice because they act like an organizing center of embryonic gonadal development and many DSD arise when these developmental processes go awry. METHODS: We employed a computational predictive algorithm for cell conversions called Mogrify to predict the transcription factors (TFs) required for direct reprogramming of human dermal fibroblasts into SCs. We established trans-differentiation culture conditions where stable transgenic expression of these TFs was achieved in 46, XY adult dermal fibroblasts using lentiviral vectors. The resulting Sertoli like cells (SLCs) were validated for SC phenotype using several approaches. RESULTS: SLCs exhibited Sertoli-like morphological and cellular properties as revealed by morphometry and xCelligence cell behavior assays. They also showed Sertoli-specific expression of molecular markers such as SOX9, PTGDS, BMP4, or DMRT1 as revealed by IF imaging, RNAseq and qPCR. The SLC transcriptome shared about two thirds of its differentially expressed genes with a human adult SC transcriptome and expressed markers typical of embryonic SCs. Notably, SLCs lacked expression of most markers of other gonadal cell types such as Leydig, germ, peritubular myoid or granulosa cells. CONCLUSIONS: The trans-differentiation method was applied to a variety of commercially available 46, XY fibroblasts derived from patients with DSD and to a 46, XX cell line. The DSD SLCs displayed altered levels of trans-differentiation in comparison to normal 46, XY-derived SLCs, thus showcasing the robustness of this new trans-differentiation model. Future applications could include using the SLCs to improve definitive diagnosis of DSD in patients with variants of unknown significance.

The increased LH levels resulting from the absence of negative feedback after castration has been linked to long-term health issues. A need exists for an alternative contraceptive agent that functions without interfering the LH pathways. This study aimed to develop antibody fragments against the follicular-stimulating hormone receptor (anti-FSHr) using phage-display technology and evaluate its effects on Sertoli cell functions. Phage clones against the extracellular domain of dog and cat FSHr selected from an antibody fragment phagemid library were analyzed for binding kinetics by surface plasmon resonance. Sertoli cells were isolated from testes of adult animals (five dogs and five cats). Efficacy test was performed by treating Sertoli cell cultures (SCCs) with anti-FSHr antibody fragments compared with untreated in triplicates. Expressions of androgen binding protein (ABP), inhibin subunit beta B (IHBB) and vascular endothelial growth factor A (VEGFA) mRNA in SCCs were quantified by RT-qPCR. The results demonstrated that the molecular weight of the purified dog and cat anti-FSHr antibody fragment was 25 kDa and 15 kDa, respectively. Based on protein molecular weight, the antibody fragment of dogs and cats was therefore, so-called single-chain variable fragments (scFv) and nanobody (nb), respectively. The binding affinity with dissociation constant (K D) was 2.32 × 10−7 M and 2.83 × 10−9 M for dog and cat anti-FSHr antibody fragments, respectively. The cross-binding kinetic interactions between the dog anti-FSHr scFv and the cat ECD of FSHr could not be fitted to the curves to determine the binding kinetics. However, the cross-binding affinity K D between the cat anti-FSHr nb and the dog ECD FSHr was 1.75 × 10−4 M. The mRNA expression of ABP , IHBB and VEGFA in SCCs was less (P < 0.05) in both dogs (12.26, 4.07 and 5.11 folds, respectively) and cats (39.53, 14.07 and 20.29 folds, respectively) treated with anti-FSHr antibody fragments, indicating the Sertoli cell functions were suppressed. In conclusion, this study demonstrated the establishment of species-specific antibody fragments against FSHr in SCCs for dogs and cats. The fragment proteins illustrate potential to be developed as non-surgical contraceptive agent targeting FSHr in companion animals. • The anti-follicular stimulating hormone receptor (anti-FSHr) antibody fragment was developed in dogs and cats by biopanning. • The anti-FSHr antibody fragments demonstrated inhibitory function in the Sertoli cell cultures. • The anti-FSHr antibody fragments demonstrated species-spefic binding. [ABSTRACT FROM AUTHOR]

Background: Inflammation-induced testicular damage is a significant contributing factor to the increasing incidence of infertility. Traditional treatments during the inflammatory phase often fail to achieve the desired fertility outcomes, necessitating innovative interventions such as cell therapy. Methods: We explored the in vivo properties of intravenously administered Sertoli cells (SCs) in an acute lipopolysaccharide (LPS)-induced inflammatory mouse model. Infiltrating and resident myeloid cell phenotypes were assessed using flow cytometry. The impact of SC administration on testis morphology and germ cell quality was evaluated using computer-assisted sperm analysis (CASA) and immunohistochemistry. Results: SCs demonstrated a distinctive migration pattern, importantly they preferentially concentrated in the testes and liver. SC application significantly reduced neutrophil infiltration as well as preserved the resident macrophage subpopulations. SCs upregulated MerTK expression in both interstitial and peritubular macrophages. Applied SC treatment exhibited protective effects on sperm including their motility and kinematic parameters, and maintained the physiological testicular morphology. Conclusion: Our study provides compelling evidence of the therapeutic efficacy of SC transplantation in alleviating acute inflammation-induced testicular damage. These findings contribute to the expanding knowledge on the potential applications of cell-based therapies for addressing reproductive health challenges and offer a promising approach for targeted interventions in male infertility. [ABSTRACT FROM AUTHOR]

Background: In polygynous species, the development of secondary sexual characters is usually decisive for male reproductive success. However, our understanding about the links between the growth of these traits and reproductive efficiency is still elusive. Most research efforts in this topic have been also focused on adult males, although the development of some secondary sexual characters, like bovid horns, typically starts after birth, continues during the puberty and in some species, such as the common eland, slows or even stops during adulthood. In this study, we investigated the relationships between horn size and testicular function during sexual development in common elands using a comprehensive approach that considers both spermatogenic and sperm parameters. Methods: Twenty-two non-sexually mature common elands were used for the present study. Horn size, body mass, testes mass, and gonadosomatic index were assessed. Spermatogenic activity was determined by cytological and histological analyses. Sperm concentration, morphology, morphometry, and intramale variation in sperm size were evaluated on epididymal sperm samples. Cluster analysis was performed to explore the influence of age on relationships between horn size and reproductive function. Results: We found that bigger horns are associated with increased Sertoli cell efficiency and reduced intramale variation in sperm size. Both parameters were not related to one another while they have shown to be associated with enhanced sperm quality in ungulates. Moreover, horn size was positively linked to the testis mass, sperm concentration, and testicular investment in the seminiferous epithelium. Spiral length and basal circumference were the horn traits most strongly correlated with spermatogenic and sperm parameters as well as those responsible for the sexual dimorphism in this species. Cluster analysis rendered two groups: the first one including males ≤30 months old, while the second one those >30 months old. Horn development and reproductive function were still correlated within age groups, with the strongest relationship found between horn size and sperm size homogeneity in males >30 months old. Conclusion: Taken together, our results indicate that horn size can be regarded as a good index of male reproductive potential during sexual development and provide insights into the role of secondary sexual characters in sexual selection dynamics. [ABSTRACT FROM AUTHOR]

Testicular sex cord-stromal tumors are clonal neoplasms, with the majority being of Leydig cell followed by Sertoli cell origins. In Leydig cell tumors, adipocytic differentiation has been previously reported as a possible distinguishing feature, which has not been reported in other sex cord-stromal tumors. Herein, we report a case of a 48-year-old man who presented with an incidentally discovered 1.1 cm testicular mass, for which he underwent partial orchiectomy. Microscopically, the tumor showed features consistent with sex cord-stromal tumor with strong and diffuse nuclear and cytoplasmic reaction for B-catenin immunohistochemistry, supporting the diagnosis of Sertoli cell tumor. A novel adipocytic differentiation, reported previously in Leydig cell tumors, was present in this tumor. [ABSTRACT FROM AUTHOR]

This article discusses the use of gonadotropin treatment as an alternative to surgery for adults with congenital hypogonadotropic hypogonadism (CHH) and cryptorchidism. CHH is a condition where there is a deficiency of certain hormones, leading to absent or incomplete puberty. Cryptorchidism, or undescended testis, is more common in individuals with CHH. The article presents three case studies of adult males with CHH and persistent cryptorchidism who were successfully treated with gonadotropin therapy. The findings suggest that this treatment can be a safe and effective alternative to surgery for correcting cryptorchidism in young adult males with CHH. Further research is needed to explore this treatment in larger patient groups. The report is authored by experts in the field and provides valuable insights for library patrons conducting research in this area. [Extracted from the article]

We evaluated the influence of different media plus various concentrations of Glial cell line‐derived neurotrophic factor (GDNF) during the in vitro culture (IVC) of testicular tissues from prepubertal collared peccary. Testes from 5 individuals were collected, fragmented and cultured for 28 days (34°C and 5% CO2). Culture media were Dulbecco's modified essential medium (DMEM) or stem cell serum free media (StemPro‐34™ SFM), both supplemented with various concentrations of GDNF (0, 10, or 20 ng/mL). Fragments were cultured on the flat surface of 0.75% agarose gel and were evaluated every 7 days for fragment area, histomorphology, cellular viability, and proliferative activity. Data were expressed as mean ± standard error and analyzed by Kruskal–Wallis's and Tukey test. Fragments area decreased over the 28 days‐culture, regardless of the treatment. For morphology, the StemPro‐37 SFM medium plus 10 ng/mL GDNF provided higher scores at all time points in comparison to DMEM using any GDNF concentration (p <.05). After 28 days, similar cellular viability (~70%) was observed in all treatments (p >.05). For proliferating cell nuclear antigen assay, only DMEM plus 10 ng/mL GDNF improved (p <.05) cellular proliferation on Days 14 and 28. Looking at argyrophilic nucleolar organizing regions, after 28 days, there were no differences among treatments regarding cell proliferative capacity for both spermatogonia and Sertoli cells (p >.05). In summary, the DMEM and StemPro‐34 SFM are adequate medium for IVC of prepubertal peccary testicular tissue. Supplementation with GDNF, especially at a 10 ng/mL concentration, appears to be essential for the maintenance of cell survival and proliferation. [ABSTRACT FROM AUTHOR]

Background Objective Materials and methods Results Discussion and conclusion Reproductive aging can adversely affect male fertility and the health of offspring. The aging process is accompanied by impaired autophagy. Recent studies have shown that Trehalose plays an important role in the prevention of various diseases by regulating autophagy. However, the roles of Trehalose in testicular aging and reproductive decline remain to be clarified.The present study aimed to evaluate the protective effects of Trehalose on testes in an aging mouse model.In this study, an in vivo aging model in mice by administering D‐galactose was established to explore the protective effect of Trehalose on testicular aging. We examined histological changes and related indicators of apoptosis, autophagy, mitochondrial biogenesis, and sperm quality.D‐galactose treatment induced oxidative stress, apoptosis, and impairment of autophagy of testicular cells in mouse testes. Trehalose administration significantly reduced germ cell apoptosis and DNA damage caused by D‐galactose‐induced oxidative stress. Notably, Trehalose activated autophagy activity and improved mitochondrial function in testicular cells. Furthermore, Trehalose treatment increased the expression level of the tight junction protein ZO‐1, and accelerated clearance of damaged mitochondria in Sertoli cells, indicating that Trehalose ameliorated Sertoli cell function in D‐galactose‐induced aging testes.These findings suggest that Trehalose administration activated the autophagy activity in testicular cells and improved mitochondrial function, thereby effectively preventing testicular aging. Trehalose and its activated autophagy are crucial for preventing testicular aging, thus restoring autophagy activity by administering Trehalose could be a promising means to delay aging. [ABSTRACT FROM AUTHOR]

Yaks live in the Qinghai-Tibet Plateau for a long time where oxygen is scarce, but can ensure the smooth development of testis and spermatogenesis. The key lies in the functional regulation of the Sertoli cells under hypoxia. In this study, we sequenced yak Sertoli cells cultured in normal oxygen concentration (Normoxia) and treated with low oxygen concentration (Hypoxia) by whole transcriptomics, and screened out 194 differentially expressed mRNAs (DEmRNAs), 934 differentially expressed LncRNAs (DELncRNAs) and 129 differentially expressed miRNAs (DEmiRNAs). GO and KEGG analysis showed that these differential genes were mainly concentrated in PI3K-AKT, MAPK, RAS, and other signaling pathways, and were associated with glucose metabolism, tight junction, steroid hormone synthesis, cell fusion, and immunity of yak Sertoli cells. We constructed the gene interaction network of yak Sertoli cells in hypoxia and screened out the relationship pairs related to glucose metabolism and tight junction. The results suggested that the changes in energy metabolism, tight junction, and immune regulation of yak Sertoli cells under hypoxia might provide favorable conditions for spermatogenesis. This study provides data for further study on the role of non-coding RNA in testis development and spermatogenesis of yak. [ABSTRACT FROM AUTHOR]

Aims Methods and results Conclusions ST11 adnexal tumour is a rare, recently described malignant neoplasm that is associated with Peutz–Jeghers syndrome. It predominantly originates from the para‐adnexal soft tissues and often shows secondary involvement of the fallopian tube and ovary. STK11 adnexal tumours have a broad differential diagnosis due to their variable morphology and non‐specific immunoprofile, and diagnostic confirmation currently requires sequencing to identify an STK11 mutation. We investigate the diagnostic utility of STK11 (LKB1) immunohistochemistry (IHC) in a cohort of STK11 adnexal tumours and morphological mimics.IHC for STK11 was performed on 122 tumours, including 17 STK11 adnexal tumours and 105 morphological mimics (10 female adnexal tumours of Wolffian origin, 22 adult granulosa cell tumours, 10 juvenile granulosa cell tumours, four Sertoli–Leydig cell tumours, two Leydig cell tumours, one Sertoli cell tumour, one steroid cell tumour, four extra‐ovarian sex cord‐stromal tumours, 16 ovarian endometrioid carcinomas, eight tubo‐ovarian high‐grade serous carcinomas, five ovarian mesonephric‐like adenocarcinomas, 14 ovarian carcinosarcomas, five peritoneal malignant mesotheliomas, two pelvic plexiform leiomyomata and one ovarian solid pseudopapillary tumour). All STK11 adnexal tumours showed complete loss of cytoplasmic staining for STK11. All other tumour types showed retained cytoplasmic staining, except for one endometrioid carcinoma with mucinous differentiation which showed complete loss of STK11 expression and a high‐grade serous carcinoma with subclonal loss.STK11 is a highly sensitive and specific immunohistochemical marker for distinguishing STK11 adnexal tumour from its histological mimics, and can obviate the need for confirmatory molecular studies in the appropriate morphological context. [ABSTRACT FROM AUTHOR]

A 27-year-old man was admitted to the hospital after a year of marriage due to infertility. During laparoscopic exploratory surgery, the presence of a retrovesical uterus was clearly observed, and the gonadal organs were visible on both sides. However, the testicles or ovaries were not identifiable, nor were the spermatic vessels and fallopian tubes at the joint. Intraoperative bilateral gonad biopsy was performed. Cryopreservation of the right gonadal gland revealed gonadoblastoma and malignant germinoma (asexual tumor/seminoma) with sclerosis and atrophy of testicular tissue. No proliferation of germ cells and sertoli cells was observed in spermatic tubule. The left gonad was diagnosed as a gonadoblastoma. Finally, total hysterectomy and bilateral gonadal tumor organectomy were performed to seal the vaginal stump. Local radiotherapy was administered after surgery. In general, tumors were found on both sides of the gonads, especially gonadoblastoma and malignant germinoma on the right side and gonadoblastoma on the left side. [ABSTRACT FROM AUTHOR]

Fumonisin B1 (FB1), a mycotoxin produced by Fusarium species, is prevalent in crops and animal feed, posing significant health risks to livestock and humans. FB1 induces oxidative stress in Sertoli cells, destroys testicular structure, and affects spermatogenesis. However, methods to mitigate the reproductive toxicity of FB1 in testes remain unknown. Quercetin, a natural flavonoid antioxidant, may offer protective benefits. This study investigated the protective effects and mechanisms of quercetin against FB1-induced reproductive toxicity in TM4 cells (a Sertoli cell line). The results indicated that 40 μM quercetin improved cell viability, reduced apoptosis, and preserved cell functions. Quercetin also decreased reactive oxygen species (ROS) levels in TM4 cells exposed to FB1, enhanced the expression of antioxidant genes, and improved mitochondrial membrane potential. Compared with FB1 alone, the combination of quercetin and FB1 increased ATP levels, as well as pyruvate and lactic acid, the key glycolysis products. Furthermore, this combination elevated the mRNA and protein expression of glycolysis-related genes, including glucose-6-phosphate isomerase 1 (Gpi1), hexokinase 2 (Hk2), aldolase (Aldoa), pyruvate kinase, muscle (Pkm), lactate dehydrogenase A (Ldha) and phosphofructokinase, liver, B-type (Pfkl). Quercetin also boosted the activity of PKM and LDHA, two crucial glycolytic enzymes. In summary, quercetin mitigates FB1-induced toxicity in TM4 cells by reducing ROS levels and enhancing glycolysis. This study offers new insights into preventing and treating FB1-induced toxic damage to the male reproductive system and highlights the potential application of quercetin. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Acrolein (ACR) is a ubiquitous environmental pollutant and byproduct of lipid peroxidation that has been implicated in male infertility. However, the molecular mechanisms underlying ACR-induced toxicity in Sertoli cells remain unclear. Given its role in inducing oxidative stress, we examined whether ferroptosis, an iron-dependent form of regulated cell death, could mediate ACR toxicity in Sertoli cells. We also tested if hydrogen sulfide (H2S), which has antioxidant and ACR detoxifying properties, could protect Sertoli cells from ACR-induced ferroptosis. ACR exposure decreased Sertoli cell viability, increased protein carbonylation and p38 MAPK phosphorylation, indicating oxidative injury. ACR also depleted glutathione (GSH), downregulated the cystine importer SLC7A11, increased intracellular ferrous iron (Fe2+) and lipid peroxidation, suggesting activation of ferroptosis. Consistently, the ferroptosis inhibitor deferoxamine (DFO) markedly attenuates ACR-induced cell death. Further studies revealed that ACR-induced ferroptotic changes were prevented by exogenous H2S and exaggerated by inhibition of endogenous H2S production. Furthermore, H2S also suppressed GPX4 inhibitor RSL3-induced intracellular ACR accumulation and ferroptosis. In summary, our study demonstrates that ACR induces ferroptotic cell death in Sertoli cells, which can be prevented by H2S through multiple mechanisms. Targeting the H2S pathway may represent a therapeutic strategy to mitigate ACR-induced Sertoli cell injury and preserve male fertility. [ABSTRACT FROM AUTHOR]

Introduction: Androgens play a pivotal role in shaping male sexual characteristics, with testosterone being an essential hormone in orchestrating various developmental processes. Testosterone biosynthesis involves a series of enzymatic reactions, among which the 17ß-hydroxysteroid dehydrogenase type 3 (HSD17B3) holds significance. While its role in adult Leydig cells is well established, its localization and importance during the fetal period remain less known, especially in humans. This study aims to delineate the dynamics of HSD17B3 expression in human fetal testes to clarify the contribution of specific cell types to testosterone biosynthesis. Methods: Using immunofluorescence staining, we investigated the expression pattern of HSD17B3 in human fetal and adult testicular tissues. Results and discussion: The findings of this study revealed a distinct temporal and cellular expression pattern of HSD17B3 protein in the fetal period. We detected its expression exclusively in Sertoli cells, the highest during the second trimester. This unique localization suggests the inclusion of fetal Sertoli cells in testosterone production during the critical masculinization-programming window. Furthermore, we demonstrated a shift in HSD17B3 expression from Sertoli cells to Leydig cells in adulthood, corroborating findings from rodent studies. This study sheds light on the intricate, still underexplored regulation of steroidogenesis during fetal development, whose disturbance might lead to testicular dysgenesis. Further research is warranted to elucidate the regulatory pathways governing the expression of HSD17B3 and its transition between Sertoli and Leydig cells, potentially paving the way for novel therapeutic interventions in disorders of sexual development. [ABSTRACT FROM AUTHOR]

Bacterial testicular inflammation is one of the important causes of male infertility. Using plant-derived compounds to overcome the side effects of antibiotics is an alternative treatment strategy for many diseases. Schizandrin B (SchB) is a bioactive compound of herbal medicine Schisandra chinensis which has multiple pharmacological effects. However its effect and the mechanism against testicular inflammation are unknown. Here we tackled these questions using models of lipopolysaccharide (LPS)-induced mice and -Sertoli cells (SCs). Histologically, SchB ameliorated the LPS-induced damages of the seminiferous epithelium and blood-testicular barrier, and reduced the production of pro-inflammatory mediators in mouse testes. Furthermore, SchB decreased the levels of pro-inflammatory mediators and inhibited the nuclear factor kB (NF-κB) and MAPK (especially JNK) signaling pathway phosphorylation in LPS-induced mSCs. The bioinformatics analysis based on receptor prediction and the molecular docking was further conducted. We targeted androgen receptor (AR) and illustrated that AR might bind with SchB in its function. Further experiments indicate that the AR expression was upregulated by LPS stimulation, while SchB treatment reversed this phenomenon; similarly, the expression of the JNK-related proteins and apoptotic-related protein were also reversed after AR activator treatment. Together, SchB mitigates LPS-induced inflammation and apoptosis by inhibiting the AR-JNK pathway. [ABSTRACT FROM AUTHOR]

Plastic pollution is an emerging environmental issue, with microplastics and nanoplastics raising health concerns due to bioaccumulation. This work explored the impact of polystyrene nanoparticle (PS-NPs) exposure during prepuberty on male reproductive function post maturation in rats. Rats were gavaged with PS-NPs (80 nm) at 0, 3, 6, 12 mg/kg/day from postnatal day 21 to 95. PS-NPs accumulated in the testes and reduced sperm quality, serum reproductive hormones, and testicular coefficients. HE staining showed impaired spermatogenesis. PS-NPs disrupted the blood-testis barrier (BTB) by decreasing junction proteins, inducing inflammation and apoptosis. Transcriptomics identified differentially expressed genes related to metabolism, lysosome, apoptosis, and TLR4 signaling. Molecular docking revealed Cordycepin could compete with polystyrene for binding to TLR4. Cordycepin alleviated oxidative stress and improved barrier function in PS-NPs treated Sertoli cells. In conclusion, prepubertal PS-NPs exposure induces long-term reproductive toxicity in male rats, likely by disrupting spermatogenesis through oxidative stress and BTB damage. Cordycepin could potentially antagonize this effect by targeting TLR4 and warrants further study as a protective agent. This study elucidates the mechanisms underlying reproductive toxicity of PS-NPs and explores therapeutic strategies. [ABSTRACT FROM AUTHOR]

D-Aspartic Acid (D-Asp) affects spermatogenesis by enhancing the biosynthesis of the sex steroid hormones acting either through the hypothalamu-spituitary-testis axis or directly on Leydig cells. Recently, in vitro studies have also demonstrated the direct effects of D-Asp on the proliferation and/or activity of germ cells. However, although D-Asp is present in Sertoli cells (SC), the specific role of the amino acid in these cells remains unknown. This study investigated the effects of D-Asp on the proliferation and activity of TM4 SC, focusing on the mitochondrial compartment and its association with the endoplasmic reticulum (ER). We found that D-Asp enhanced the proliferation and activity of TM4 cells as evidenced by the activation of ERK/Akt/PCNA pathway and the increase in the protein levels of the androgen receptor. Furthermore, D-Asp reduced both the oxidative stress and apoptotic process. An increase in mitochondrial functionality and dynamics, as well as a reduction in ER stress, were also found in D-Asptreated TM4 cells. It is known that mitochondria are closely associated with ER to form the Mitochondrial-Associated Endoplasmic Reticulum Membranes (MAM), the site of calcium ions and lipid transfer from ER to the mitochondria, and vice versa. The data demonstrated that D-Asp induced stabilization of MAM in TM4 cells. In conclusion, this study is the first to demonstrate a direct effect of D-Asp on SC activity and to clarify the cellular/molecular mechanism underlying these effects, suggesting that D-Asp could stimulate spermatogenesis by improving the efficiency of SC. [ABSTRACT FROM AUTHOR]

Male fertility depends on normal pubertal development. Di-(2-ethylhexyl) phthalate (DEHP) is a potent antiandrogen chemical, and exposure to DEHP during peripuberty can damage the developing male reproductive system, especially the testis. However, the specific cellular targets and differentiation processes affected by DEHP, which lead to testicular toxicity, remain poorly defined. Herein, we presented the first single-cell transcriptomic profile of the pubertal mouse testis following DEHP exposure. To carry out the experiment, 2 groups (n  = 8 each) of 3-week-old male mice were orally administered 0.5% carboxymethylcellulose sodium salt or 100 mg/kg body weight DEHP daily from postnatal day 21–48, respectively. Using single-cell RNA sequencing, a total of 31 distinct cell populations were identified, notably, Sertoli and Leydig cells emerged as important targets of DEHP. DEHP exposure significantly decreased the proportions of Sertoli cell clusters expressing mature Sertoli markers (Sox9 and Ar), and selectively reduced the expression of testosterone synthesis genes in fetal Leydig cells. Through cell–cell interaction analyses, we observed changed numbers of interactions in Sertoli cells 1 (SCs1), Leydig cells 1 (LCs1), and interstitial macrophages, and we also identified cell-specific ligand gene expressions in these clusters, such as Inha , Fyn , Vcam1 , and Apoe. Complementary in vitro assays confirmed that DEHP directly reduced the expression of genes related to Sertoli cell adhesion and intercellular communication. In conclusion, peripubertal DEHP exposure reduced the number of mature Sertoli cells and may disrupt testicular steroidogenesis by affecting the testosterone synthesis genes in fetal Leydig cells rather than adult Leydig cells. [ABSTRACT FROM AUTHOR]

Background Objectives Materials and methods Results Discussion and conclusion Severe acute respiratory syndrome coronavirus 2, the virus responsible for coronavirus disease 2019, affects multiple organs. The virus enters cells through angiotensin‐converting enzyme‐2 and host factors present in genital organs, leading to concern over virus shedding in semen and reproductive function.To investigate severe acute respiratory syndrome coronavirus 2 in semen from patients with a mild infection, identify the seminal infected cells, and explore the effect of the infection on sex hormones and semen parameters.Prospective study of 54 men with mild severe acute respiratory syndrome coronavirus 2 infection. Semen was collected at 7, 15, 30, 60, 90, 180, and 365 days after symptom onset, and severe acute respiratory syndrome coronavirus 2 RNA was measured in serum, saliva, urine, and semen. The presence of infectious severe acute respiratory syndrome coronavirus 2 in semen was assessed using Vero cell culture. Infected semen cells were identified using immunofluorescence against severe acute respiratory syndrome coronavirus 2 nucleoprotein antigen and cell markers. Semen characteristics as well as testosterone, inhibin B, luteinizing hormone, and follicle‐stimulating hormone levels were determined.11% of patients had at least one severe acute respiratory syndrome coronavirus 2 RNA‐positive semen. One patient had viral semen shedding up to day 90 after infection onset, with replication‐competent virus isolated from semen and 40% cell fraction at day 7. After sperm preparation, 90% fraction was severe acute respiratory syndrome coronavirus 2 RNA‐positive at days 7 and 15. The swim‐up fraction was positive only on day 7. In semen, nucleoprotein antigen was detected mainly in exfoliated epithelial cells and less frequently in Sertoli cells. Sperm count and motile sperm count were lower at day 30 than at day 7. Round cells in semen were increased during the acute phase. At days 7 and 15, sperm count and motile sperm count were lower in severe acute respiratory syndrome coronavirus 2 RNA‐positive semen compared with negative semen, while semen volume and follicle‐stimulating hormone levels were increased. Long‐term follow‐up shows no evidence of a detrimental effect on hormonal or semen characteristics.11% of patients with mild coronavirus disease 2019 who were not hospitalized had severe acute respiratory syndrome coronavirus 2 excretions in semen, which persisted for up to 90 days in one patient. No germ cells appeared infected by the virus, but the detection of nucleoprotein antigen‐positive epithelial semen cells and Sertoli cells suggests genital tract infection. Albeit infrequent, semen may contain the replication‐competent virus during the acute phase with potential risk of severe acute respiratory syndrome coronavirus 2 transmissions during sexual contact and assisted reproduction procedures. The effect of mild coronavirus disease 2019 on spermatogenesis and reproductive hormones was moderate and reversible. [ABSTRACT FROM AUTHOR]

In all vertebrates, reproductive strategies are achieved by modulation of the neuroendocrine system in a similar manner and with minor variations among the different classes. Most of the available information on amphibian testicular cycles derive from anurans, and among these, water frogs have been extensively studied in terms of reproductive mechanisms and sex steroid correlation. Adiponectin (AdipoQ) and its receptors—AdipoR1 and AdipoR2—are essential for most of the normal testicular and sperm functions. In this study, the identification of AdipoQ and its two receptors was carried out by immunohistochemistry in the testis of adult males of Pelophylax bergeri. The AdipoQ system was observed in the frog spermatogenic cysts, in both germinal and Sertoli cells, as well as in the rete testis. AdipoQ and AdipoR1 were localized in germ-line cells, from spermatogonia to round spermatids, while AdipoR2 was detected in the elongated spermatids, spermatozoa, and Sertoli cells. AdipoR1 was also observed in the intratesticular canals of the rete testis. This preliminary study shows the AdipoQ system's presence in the anurans' testis. The results obtained could be a starting point for future functional studies aimed at defining the physiological role of the AdipoQ system in frog testicular functions. [ABSTRACT FROM AUTHOR]

Ribavirin, an antiretroviral agent targeting the hepatitis C virus, causes male reproductive toxicity. This study investigated the mechanism of ribavirin transport at the blood–testis barrier (BTB). In vivo mouse integration plot analysis after intravenous administration revealed that the net influx clearance of [3H]ribavirin in the testis was 3.6-fold greater than that of [14C]D-mannitol, a paracellular transport marker, implying transcellular transport of ribavirin across the BTB. Moreover, [3H]ribavirin uptake by TM4 cells, mouse-derived Sertoli cells, was time- and concentration-dependent, with a K m value of 2.49 mM. S -[(4-nitrophenyl)methyl]-6-thioinosine, an inhibitor of Na+-independent equilibrative nucleoside transporters (ENTs), strongly inhibited the [3H]ribavirin uptake by TM4 cells at 100 µM. Compared to the uptake of [3H]adenosine, a typical endogenous nucleoside, [3H]ribavirin uptake was relatively similar to ENT2 transport. [3H]Ribavirin uptake was also observed in mouse ENT2-expressing Xenopus laevis oocytes, and gene silencing via the transfection of ENT2 small interfering RNA significantly reduced the [3H]ribavirin transport into TM4 cells by 13%. Taken together, these results suggest that ENT2 partially contributes to ribavirin transport at the BTB. [Display omitted] • Ribavirin, an anti-viral drug, undergoes the facilitative blood-to-testis transport in vivo. • Carrier-mediated processes contribute to the ribavirin transport at the BTB. • The characteristics of ribavirin transport at the BTB is different from ENT1-mediated adenosine transport. • ENT2 partially contributes to ribavirin transport at the BTB. [ABSTRACT FROM AUTHOR]

Sertoli cell (SC) proliferation plays an important role in sperm production and quality; however, the regulatory mechanism of SC proliferation is not well understood. This study investigated the role of adenosine monophosphate-activated protein kinase (AMPK) in the regulation of immature boar SC activity. Cell counting kit-8, Seahorse XFe96, mitochondrial respiratory enzyme-related assay kits, and transmission electron microscopy were used to detect SC proliferative viability, oxygen consumption rate (OCR), mitochondrial respiratory enzyme activity, and the ultrastructure of primary cultured SCs in vitro from the testes of 21-day-old boars. A dual luciferase reporter assay was performed to determine the miRNA-mRNA target interaction. Western blotting was used to analyze cell proliferation-related protein expression of p38, p21, proliferating cell nuclear antigen (PCNA), Cyclin-dependent kinase 4 (CDK4), Cyclin D3, and phosphorylated retinoblastoma protein (Rb). Each experiment had a completely randomized design, with three replicates in each experiment. The results showed that the AMPK inhibitor (Compound C, 20 μM-24 h) increased cell proliferation viability, ATP production, and maximal respiration of SCs by 0.64-, 0.12-, and 0.08-fold (p < 0.05), respectively; increased the SC protein expression of PCNA, CDK4, Cyclin D3, and p-Rb by 0.13-, 0.09-, 0.88-, and 0.12-fold (p < 0.05), respectively; and decreased the SC protein expression of p38 and p21 by 0.36- and 0.27-fold (p < 0.05), respectively. The AMPK agonist AICAR (2 mM-6 h) significantly inhibited SC ultrastructure, OCR, mitochondrial respiratory enzyme activity, and cell proliferation-related protein levels. AMPK was validated to be a target gene of miR-1285 based on the result in which the miR-1285 mimic inhibited the luciferase activity of wild-type AMPK by 0.54-fold (p < 0.001). MiR-1285 mimic promoted the OCR of SCs, with 0.45-, 0.15-, 0.21-, and 0.30-fold (p < 0.01) increases in ATP production, basal and maximal respiration, and spare capacity, respectively. MiR-1285 mimic increased the mitochondrial respiratory enzyme activity of SCs, with 0.63-, 0.70-, and 0.97-fold (p < 0.01) increases in NADH-Q oxidoreductase, cytochrome c oxidase, and ATP synthase, respectively. Moreover, the miR-1285 mimic increased the protein expression of PCNA, CDK4, Cyclin D3, and p-Rb by 0.24-, 0.30-, 0.22-, and 0.13-fold (p < 0.05), respectively, and reduced the protein expression of p38 and p21 by 0.58- and 0.66-fold (p < 0.001). MiR-1285 inhibitor showed opposite effects on the above indicators and induced numerous autophagosomes and large lipid droplets in SCs. A high dose of estradiol (10 μM-6 h, showed a promotion of AMPK activation in a previous study) significantly inhibited SC ultrastructure, mitochondrial function, and proliferation-related pathways, while these adverse effects were weakened by Compound C treatment or miR-1285 mimic transfection. Our findings suggest that the activation and inhibition of AMPK induced by specific drugs or synthesized targeted miRNA fragments could regulate immature boar SC proliferative activity by influencing the CDK4/Cyclin D3 pathway and mitochondrial function; this helps to provide a basis for the prevention and treatment of male sterility in clinical practice. • Compound C increases immature boar SC proliferative activity. • MiR-1285 mimic can target and inhibit AMPK. • AMPK inhibition increases SC mitochondrial function and CDK4/Cyclin D3 pathway. • AMPK activation damages SC ultrastructure. • AMPK inhibition reduces adverse effects of estradiol on SCs. [ABSTRACT FROM AUTHOR]

In mammals, glial cell derived neurotrophic factor (GDNF) plays a critical role in the self-renewal and maintenance of spermatogonial stem cells (SSCs) in testis and oogenesis in ovary, whilst retinoic acid (RA), the key factor of meiosis initiation, can downregulate its expression. Unlike mammals, two Gdnf replication genes are widely present in teleost fishes, however, our understanding of them is still poor. In the present study, two paralogous gdnf from Nile tilapia (Oreochromis niloticus), namely as Ongdnfa and Ongdnfb , were characterized, and then their cellular expression profiles in testis and ovary and responsiveness to RA treatment at the tissue and cellular levels were investigated. In phylogenetic tree, the Gdnfa and Gdnfb from teleost fishes were clustered into two different subclasses, respectively, and then clustered with the homologs from cartilaginous fish and tetrapods, suggesting that OnGdnfa and OnGdnfb are orthologous to GDNF and paralogous to each other. Ongdnfa is expressed in Sertoli cells and Leydig cells in testis and oocytes in ovary. The expression pattern of Ongdnfb is similar to Ongdnfa. In the ex vivo testicular organ culture, RA down-regulated the expression of Ongdnfa , whereas up-regulated the expression of Ongdnfb (P < 0.05), suggesting that they have differential responsiveness to RA signaling. RA treatment of the cultured cells derived from adult Nile tilapia testis which have the expression of RA receptors (RAR), Ongdnfa and Ongdnfb further confirmed the above result. Collectively, our study suggests that Ongdnfa and Ongdnfb have non-germline expression patterns in testis and germline expression patterns in ovary; furthermore, they have differential responsiveness to RA signaling, implying that they might have differential biological functions. This study broadens and enriches our understanding of fish GDNF homologs and lays foundation for the study of their biological functions in the future. • Two paralogous gdnf (Ongdnfa and Ongdnfb) were characterized from Nile tilapia. • Ongdnfa and Ongdnfb had non-germline expressions in testis. • Ongdnfa and Ongdnfb had germline expressions in ovary. • Ongdnfa and Ongdnfb had differential responsiveness to RA treatment. [ABSTRACT FROM AUTHOR]

To improve the mechanistic screening of reproductive toxicants in chemical-risk assessment and drug development, we have developed a three-dimensional (3D) heterogenous testicular co-culture model from neonatal mice. Di-n-butyl phthalate (DBP), an environmental contaminant that can affect reproductive health negatively, was used as a model compound to illustrate the utility of the in vitro model. The cells were treated with DBP (1 nM to 100 µM) for 7 days. Automated high-content imaging confirmed the presence of cell-specific markers of Leydig cells (CYP11A1 +), Sertoli cells (SOX9 +), and germ cells (DAZL +). Steroidogenic activity of Leydig cells was demonstrated by analyzing testosterone levels in the culture medium. DBP induced a concentration-dependent reduction in testosterone levels and decreased the number of Leydig cells compared to vehicle control. The levels of steroidogenic regulator StAR and the steroidogenic enzyme CYP11A1 were decreased already at the lowest DBP concentration (1 nM), demonstrating upstream effects in the testosterone biosynthesis pathway. Furthermore, exposure to 10 nM DBP decreased the levels of the germ cell-specific RNA binding protein DAZL, central for the spermatogenesis. The 3D model also captured the development of the Sertoli cell junction proteins, N-cadherin and Zonula occludens protein 1 (ZO-1), critical for the blood–testis barrier. However, DBP exposure did not significantly alter the cadherin and ZO-1 levels. Altogether, this 3D in vitro system models testicular cellular signaling and function, making it a powerful tool for mechanistic screening of developmental testicular toxicity. This can open a new avenue for high throughput screening of chemically-induced reproductive toxicity during sensitive developmental phases. [ABSTRACT FROM AUTHOR]

Azoospermia, the absence of sperm cells in semen, affects around 15% of infertile males. Sertoli cell-only syndrome (SCOS) is the most common pathological lesion in the background of non-obstructive azoospermia and is characterised by the complete absence of germinal epithelium, with Sertoli cells exclusively present in the seminiferous tubules. Studies have shown a correlation between successful spermatogenesis and male fertility with lipid composition of spermatozoa, semen, seminal plasma or testis. The aim of this research was to discover the correlation between the Johnsen scoring system and phospholipid expressions in testicular cryosections of SCOS patients. MALDI imaging mass spectrometry is used to determine spatial distributions of molecular species, such as phospholipids. Phosphatidylcholines (PCs), phosphatidylethanolamines (PEs) and sphingomyelins (SMs) are the most abundant phospholipids in mammalian cells and testis. SMs, the structural components of plasma membranes, are crucial for spermatogenesis and sperm function. Plasmalogens, are unique PCs in testis with strong antioxidative properties. This study, using imaging mass spectrometry, demonstrates the local distribution of phospholipids, particularly SMs, PCs, plasmalogens and PEs in human testicular samples with SCOS for the first time. This study found a strong relationship between the Johnsen scoring system and phospholipid expression levels in human testicular tissues. Future findings could enable routine diagnostic techniques during microTESE procedures for successful sperm extraction. [ABSTRACT FROM AUTHOR]

Although testicular tumors are the most common genital system tumors in dogs, Sertoli cells tumors (SCT) are the least occurring of them all to warrant their continuous evaluations. More so, SCTs may present some microscopic diagnostic difficulties requiring the use of other diagnostic tools. The present study focused on hematological, histomorphological and immunohistochemical evaluation of bilateral testicular growths in a Mongrel dog. All vital parameters were within optimum ranges and all investigations were according to standard procedures. The disease condition was characterized by lymphocytosis and thrombocytopenia with the rare bilateral grey to yellowish multilobulated testicular growths composed of diffuse haphazardly arranged uniform small round cells with scanty cytoplasm and hyperchromatic round to oval nuclei in a nest-like formation separated by little intercellular fibrillar septa, which reacted positively to only vimentin and NSE and also α-SMA (fibrillar septa only) but negatively to CD3, CD20, CD99, CK A1/A3, CD117, LCA, desmin, S-100, HMB45, β-HCG, E-cadherin, synaptophysin, and NFP antibodies, respectively. A diagnosis of bilateral canine SCT with tendency towards metastasis became imperative based on the study findings. Surgical excision of the mass was performed and managed. The used large battery of immunohistochemical antibodies highlighted their diagnostic potentials in aiding histopathological diagnosis of canine SCTs in veterinary practice. The findings further highlighted the need for the continuous monitoring of sexually active dogs with descended testis for cases of SCTs. [ABSTRACT FROM AUTHOR]

The yak (Bos grunniens) is a valuable livestock animal endemic to the Qinghai–Tibet Plateau in China with low reproductive rates. Cryptorchidism is one of the primary causes of infertility in male yaks. Compared with normal testes, the tight junctions (TJs) of Sertoli cells (SCs) and the integrity of the blood–testis barrier (BTB) in cryptorchidism are both disrupted. MicroRNAs are hairpin-derived RNAs of about 19–25 nucleotides in length and are involved in a variety of biological processes. Numerous studies have shown the involvement of microRNAs in the reproductive physiology of yak. In this study, we executed RNA sequencing (RNA-seq) to describe the expression profiles of mRNAs and microRNAs in yaks with normal testes and cryptorchidism to identify differentially expressed genes. GO and KEGG analyses were used to identify the biological processes and signaling pathways which the target genes of the differentially expressed microRNAs primarily engaged. It was found that novel-m0230-3p is an important miRNA that significantly differentiates between cryptorchidism and normal testes, and it is down-regulated in cryptorchidism with p < 0.05. Novel-m0230-3p and its target gene CSF1 both significantly contribute to the regulation of cell adhesion and tight junctions. The binding sites of novel-m0230-3p with CSF1 were validated by a dual luciferase reporter system. Then, mimics and inhibitors of novel-m0230-3p were transfected in vitro into SCs, respectively. A further analysis using qRT-PCR, immunofluorescence (IF), and Western blotting confirmed that the expression of cell adhesion and tight-junction-related proteins Occludin and ZO-1 both showed changes. Specifically, both the mRNA and protein expression levels of Occludin and ZO-1 in SCs decreased after transfection with the novel-m0230-3p mimics, while they increased after transfection with the inhibitors, with p < 0.05. These were achieved via the CSF1/CSF1R/Ras signaling pathway. In summary, our findings indicate a negative miRNA-mRNA regulatory network involving the CSF1/CSF1R/Ras signaling pathway in yak SCs. These results provide new insights into the molecular mechanisms of CSF1 and suggest that novel-m0230-3p and its target protein CSF1 could be used as potential therapeutic targets for yak cryptorchidism. [ABSTRACT FROM AUTHOR]

Per- and polyfluoroalkyl substances (PFASs) are a family of "forever chemicals" including perfluorooctane sulfonate (PFOS). These toxic chemicals do not break down in the environment or in our bodies. In the human body, PFOS and perfluoroctanoic acid (PFOA) have a half-life (T1/2) of about 4–5 yr so low daily consumption of these chemicals can accumulate in the human body to a harmful level over a long period. Although the use of PFOS in consumer products was banned in the United States in 2022/2023, this forever chemical remains detectable in our tap water and food products. Every American tested has a high level of PFAS in their blood (https://cleanwater.org/pfas-forever-chemicals). In this report, we used a Sertoli cell blood-testis barrier (BTB) model with primary Sertoli cells cultured in vitro with an established functional tight junction (TJ)-permeability barrier that mimicked the BTB in vivo. Treatment of Sertoli cells with PFOS was found to perturb the TJ-barrier, which was the result of cytoskeletal disruption across the cell cytoplasm, disrupting actin and microtubule polymerization. These changes thus affected the proper localization of BTB-associated proteins at the BTB. Using RNA-Seq transcriptome profiling, bioinformatics analysis, and pertinent biochemical and cell biology techniques, it was discovered that PFOS -induced Sertoli cell toxicity through the c-Jun N-terminal kinase (JNK; also known as stress-activated protein kinase, SAPK) and its phosphorylated/active form p-JNK signaling pathway. More importantly, KB-R7943 mesylate (KB), a JNK/p-JNK activator, was capable of blocking PFOS-induced Sertoli cell injury, supporting the notion that PFOS-induced cell injury can possibly be therapeutically managed. NEW & NOTEWORTHY: PFOS induces Sertoli cell injury, including disruption of the 1) blood-testis barrier function and 2) cytoskeletal organization, which, in turn, impedes male reproductive function. These changes are mediated by JNK/p-JNK signaling pathway. However, the use of KB-R7943, a JNK/p-JNK activator was capable of blocking PFOS-induced Sertoli cell injury, supporting the possibility of therapeutically managing PFOS-induced reproductive dysfunction. [ABSTRACT FROM AUTHOR]

Simple Summary: The goal of this research was to determine if the neurotrophin β-Nerve growth factor is present in the reproductive tract of stallions. This protein has been identified in the semen of camelids and functions to stimulate the release of a hormone from the pituitary of female camelids that is ultimately responsible for the induction of ovulation. β-Nerve growth factor (β-NGF) is a protein produced in the reproductive tract of camelids (camels, llamas, and alpacas) that has been identified as the ovulation inducing factor in seminal plasma. β-NGF from seminal plasma deposited into the reproductive tract of the female camelid acts systemically to stimulate the secretion of luteinizing hormone (LH) from the anterior pituitary, which in turn induces follicle maturation and ovulation. The objectives of the present study were to determine if β-NGF is present in the reproductive tract of the stallion and identify the specific site(s) of production. The hypotheses were that β-NGF would be present in the stallion reproductive tract and would primarily be localized in Sertoli cells of the testes and the prostate gland. Immunohistochemistry on paraffin-embedded paraformaldehyde-fixed tissues was performed using a rabbit polyclonal anti-β-NGF antibody on a total of six male equine reproductive tracts, including a one-day old colt, a one-year-old colt, and four adult stallion tracts. Strong immunostaining was observed in the efferent ducts of the testes and the epithelial cells of the prostate, seminal vesicles, bulbourethral glands, and ampullae. Weaker β-NGF staining was noted in Leydig cells, Sertoli cells, and spermatogonia within the testes and in epithelial cells of the epididymis. In conclusion, immunohistochemistry revealed that β-NGF is present in the stallion reproductive tract, and the protein is primarily present in the efferent ducts of the testes and in all accessory sex glands. [ABSTRACT FROM AUTHOR]

Background and Aim: Marburg virus (MARV) is a highly virulent virus of animal origin and the cause of a lethal infection (known as Marburg virus disease [MVD]) with a case‐fatality ratio ranging from 24% to 90%. While the potential nonzoonotic routes of virus spread are plausible, the risk is not yet fully determined. Here, we described the ways by which MARV spreads within the human population focusing mainly on the potential of sexual transmission. In addition, we addressed some measures that should be taken to minimize the risk of sexual spread of the virus and proposed a future research agenda on the risk of sexual transmission. Methods: For this perspective, we searched four electronic databases (i.e., PubMed, Scopus, Web of Science, and Google Scholar) and included the most relevant studies published since the first identification of the virus in 1967. We used "Marburg virus," "Marburg virus disease," "Seminal fluid," "Sexually‐transmitted virus," "Sexual transmission," and "Emerging infectious disease" as keywords. Results: MARV is transmitted to humans via both direct and indirect contact with infected animals (most importantly bats) and individuals who have recently been diagnosed with or died of the disease. The virus transmission through sexual contact has been previously suspected (exclusively from men to their sexual partners). Studies suggest that this virus persists predominantly in testicular Sertoli cells within seminiferous tubules over a relatively long period and is released through seminal fluid (in some reports >200 days post onset of infection) both could potentially threaten sexual health. In addition to men, women could theoretically, although less probably contribute to the sexual transmission of the disease. Conclusion: MVD, however, rarely, could be passed through sex, and men appear to be the main carriers in this regard. Taking preventive countermeasures and practicing safe sex are recommended to reduce the risk of interhuman transmission. Key points: Marburg virus disease (MVD) is a severe and often lethal viral infection with epidemic potential recorded over the past few decades.Marburg virus (MARV) spreads to humans via animal‐to‐human and human‐to‐human transmission.MARV infects the male and likely female reproductive system with testes being the main site of virus persistence.MARV sheds into genital fluids, particularly in the seminal fluid of male survivors, and probably could be passed, however rarely, through sex, even after the disease has resolved.Sexual transmission of MVD is preventable by early detection, taking precautions, and practicing safer sex. [ABSTRACT FROM AUTHOR]

There are 3 physiological waves of central hypothalamic-pituitary-gonadal (HPG) axis activity over the lifetime. The first occurs during fetal life, the second—termed "mini-puberty"—in the first months after birth, and the third at puberty. After adolescence, the axis remains active all through adulthood. Congenital hypogonadotropic hypogonadism (CHH) is a rare genetic disorder characterized by a deficiency in hypothalamic gonadotropin-releasing hormone (GnRH) secretion or action. In cases of severe CHH, all 3 waves of GnRH pulsatility are absent. The absence of fetal HPG axis activation manifests in around 50% of male newborns with micropenis and/or undescended testes (cryptorchidism). In these boys, the lack of the mini-puberty phase accentuates testicular immaturity. This is characterized by a low number of Sertoli cells, which are important for future reproductive capacity. Thus, absent mini-puberty will have detrimental effects on later fertility in these males. The diagnosis of CHH is often missed in infants, and even if recognized, there is no consensus on optimal therapeutic management. Here we review physiological mini-puberty and consequences of central HPG axis disorders; provide a diagnostic approach to allow for early identification of these conditions; and review current treatment options for replacement of mini-puberty in male infants with CHH. There is evidence from small case series that replacement with gonadotropins to mimic "mini-puberty" in males could have beneficial outcomes not only regarding testis descent, but also normalization of testis and penile sizes. Moreover, such therapeutic replacement regimens in disordered mini-puberty could address both reproductive and nonreproductive implications. [ABSTRACT FROM AUTHOR]

The sequential culture expansion has been shown to affect the favorable characteristics of human mesenchymal stem/stromal cells (hMSCs), including their immunomodulatory potential. Hereby, we have investigated the immunomodulatory potential of Yazd human testicular Cells (YhTCs) regarding gene expression profile assessment during their lifespan. Morphological examination, immunostaining of vimentin and fibronectin markers, and flow cytometry analysis were used to confirm cultured cells' MSC properties. Quantitative RT-PCR was used to investigate the target genes, COX-1, TGF-β, IL-6, VCAM-1, and FAS, at different passage numbers (P3, P6, and P9). Our findings confirmed the MSC properties of YhTCs over the lifespan. The results of COX-1 and IL-6 indicated significantly higher expression levels at the late passages (p‑value = 0.03 and p-value = 0.01, respectively). Despite the similar expression pattern, VCAM-1 expression altered significantly at different passages (p-value = 0.0006), but the changes in TGF-β expression levels were non-significant across the lifespan. The YhTCs maintained the main characteristics of MSCs over the sequential culture expansion. Although some of the target genes in the study were sensitive to the culture expansion in the unstimulated YhTCs (IL-6, COX-1, and VCAM1), it seems that FAS and TGF-β need external stimuli or intracellular signals to be induced in YhTCs. [ABSTRACT FROM AUTHOR]

Spermatogenesis is a multi-step biological process where mitotically active diploid (2n) spermatogonia differentiate into haploid (n) spermatozoa via regulated meiotic programming. The alarming rise in male infertility has become a global concern during the past decade thereby demanding an extensive profiling of testicular gene expression. Advancements in Next- Generation Sequencing (NGS) technologies have revolutionized our empathy towards complex biological events including spermatogenesis. However, despite multiple attempts made in the past to reveal the testicular transcriptional signature(s) either with bulk tissues or at the single-cell, level, comprehensive reviews on testicular transcriptomics and associated disorders are limited. Notably, technologies explicating the genome-wide gene expression patterns during various stages of spermatogenic progression provide the dynamic molecular landscape of testicular transcription. Our review discusses the advantages of single-cell RNA-sequencing (Sc-RNA-seq) over bulk RNA-seq concerning testicular tissues. Additionally, we highlight the cellular heterogeneity, spatial transcriptomics, dynamic gene expression and cell-tocell interactions with distinct cell populations within the testes including germ cells (Gc), Sertoli cells (Sc), Peritubular cells (PTc), Leydig cells (Lc), etc. Furthermore, we provide a summary of key finding of single-cell transcriptomic studies that have shed light on developmental mechanisms implicated in testicular disorders and male infertility. These insights emphasize the pivotal roles of Sc-RNA-seq in advancing our knowledge regarding testicular transcriptional landscape and may serve as a potential resource to formulate future clinical interventions for male reproductive health. [ABSTRACT FROM AUTHOR]

Heat stress reduces the number of Sertoli cells, which is closely related to an imbalanced redox status. Glutamate functions to maintain the equilibrium of redox homeostasis. However, the role of glutamate in heat treated Sertoli cells remains unclear. Herein, Sertoli cells from 3-week-old piglets were treated at 44 °C for 30 min (heat stress). Glutamate levels increased significantly following heat stress treatment, followed by a gradual decrease during recovery, while glutathione (GSH) showed a gradual increase. The addition of exogenous glutamate (700 μM) to Sertoli cells before heat stress significantly reduced the heat stress-induced apoptosis rate, mediated by enhanced levels of antioxidant substances (superoxide dismutase (SOD), total antioxidant capacity (TAC), and GSH) and reduced levels of oxidative substances (reactive oxygen species (ROS) and malondialdehyde (MDA)). Glutamate addition to Sertoli cells before heat stress upregulated the levels of glutamate-cysteine ligase, modifier subunit (Gclm), glutathione synthetase (Gss), thioredoxin (Trx1) and B-cell leukemia/lymphoma 2 (Bcl-2), and the ratio of phosphorylated Akt (protein kinase B)/total Akt. However, it decreased the levels of Bcl2-associated X protein (Bax) and cleaved-caspase 3. Addition of the inhibitor of glutaminase (Gls1), Bptes (Bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide, 30 μM)to Sertoli cells before heat stress reversed these effects. These results inferred that glutamate rescued heat stress-induced apoptosis in Sertoli cells by enhancing activity of antioxidant enzymes and activating the Trx1-Akt pathway. Thus, glutamate supplementation might represent a novel strategy to alleviate the negative effect of heat stress. • Heat treatment could cause apoptosis and oxidate stress of boar SCs. • Exogenous glutamate reduced apoptosis by enhancing the levels of antioxidant substances. • Glutamate improves oxidation resistance by regulating the expression of Gclm/Gss to increase GSH. • Protective effects of glutamate are mediated through the Trx1-Akt pathway. [ABSTRACT FROM AUTHOR]

Context: A population of sperm progenitor cells, known as Asingle spermatogonia, has been described in mammalian testes. During division cycles in spermatogenesis, some cells will form part of the Asingle spermatogonia group, while others form primary spermatocytes. Thus, during spermatogenesis, spermatogonia are the progenitor cells of spermatozoa. Aims: In this study, we characterise the spermatogonial stem cells (SSCs) in the testicles of Artibeus jamaicensis and Sturnira lilium bats. The knowledge generated from this will contribute to the understanding of the biology of germ cells and the mechanisms of spermatogenesis in mammals, generating information on wildlife species that are important for biodiversity. Methods: Testes were analysed by light and electron microscopy. Likewise, the expression of specific factors of stem cells (Oct4 and C-kit), germ cells (Vasa), cell proliferation (pH3 and SCP1) and testicular somatic cells (MIS, 3βHSD and Sox9) was characterised by immunofluorescence and western blot. Key results: The histological analysis enabled the location of type Asingle, Apaired and Aaligned spermatogonia in the periphery of the seminiferous tubules adjacent to Sertoli cells. The expression of genes of stem and germ cells made it possible to corroborate the distribution of the SSCs. Conclusions: Results indicate that type Asingle spermatogonia were not randomly distributed, since proliferative activity was detected in groups of cells adjacent to the seminiferous tubules membrane, suggesting the localisation of spermatogonial niches in a specific region of testes. Implications: This study provides evidence for the existence of SSCs in the testis of chiropterans that contribute to the renewal of germline progenitor cells to maintain the reproduction of the organisms. A population of sperm progenitor cells, known as Asingle spermatogonia, has been described in mammalian testes, but these cells have not been studied in bat testes. Bats are a good model due to their human-like reproductive characteristics. The results show the existence of spermatogonial stem cells that constantly renew spermatogenesis in these chiropterans, similar to that described in other mammals such as humans. Image by Tania Porras-Gómez and Norma Moreno-Mendoza. [ABSTRACT FROM AUTHOR]

In studying the molecular underpinning of spermatogenesis, we expect to understand the fundamental biological processes better and potentially identify genes that may lead to novel diagnostic and therapeutic strategies toward precision medicine in male infertility. In this review, we emphasized our perspective that the path forward necessitates integrative studies that rely on complementary approaches and types of data. To comprehensively analyze spermatogenesis, this review proposes four axes of integration. First, spanning the analysis of spermatogenesis in the healthy state alongside pathologies. Second, the experimental analysis of model systems (in which we can deploy treatments and perturbations) alongside human data. Third, the phenotype is measured alongside its underlying molecular profiles using known markers augmented with unbiased profiles. Finally, the testicular cells are studied as ecosystems, analyzing the germ cells alongside the states observed in the supporting somatic cells. Recently, the study of spermatogenesis has been advancing using single-cell RNA sequencing, where scientists have uncovered the unique stages of germ cell development in mice, revealing new regulators of spermatogenesis and previously unknown cell subtypes in the testis. An in-depth analysis of meiotic and postmeiotic stages led to the discovery of marker genes for spermatogonia, Sertoli and Leydig cells and further elucidated all the other germline and somatic cells in the testis microenvironment in normal and pathogenic conditions. The outcome of an integrative analysis of spermatogenesis using advanced molecular profiling technologies such as scRNA-seq has already propelled our biological understanding, with additional studies expected to have clinical implications for the study of male fertility. By uncovering new genes and pathways involved in abnormal spermatogenesis, we may gain insights into subfertility or sterility. [ABSTRACT FROM AUTHOR]

Background: Testicular Leydig cell tumours (LCTs) are the most common type of sex cord–stromal tumour in men, representing 1%–3% of all testicular neoplasms. Among testicular sex cord–stromal tumours, CTNNB1 mutations and nuclear expression of β‐catenin have been typically associated with Sertoli cell tumour. Recent genomic analyses have shown that CTNNB1 variants are also identified in a subset of LCTs; however, the frequency and clinicopathologic associations of β‐catenin alterations remain incompletely understood in this tumour type. Methods: In this study, we evaluated 32 LCTs (five malignant/metastasizing, 27 nonmetastasizing) using β‐catenin immunohistochemistry and DNA sequencing. Results: Immunohistochemistry revealed focal or multifocal nuclear β‐catenin expression in 47% of the tumours. Diffuse nuclear β‐catenin expression (in >50% of the tumour cells) was not detected in any of the cases analysed herein. Comparison of β‐catenin‐positive and β‐catenin‐negative cases did not show significant differences in the frequency of adverse histopathologic findings or malignant clinical behaviour. DNA sequencing performed de novo on a subset of seven cases revealed the presence of exon 3 CTNNB1 variants in four of them (4/7, 57%), with variant allele frequencies (VAF) ranging from 7 to 33%. Two additional β‐catenin‐positive cases that had been sequenced as part of a previous study harboured exon 3 CTNNB1 variants at VAF of 28% and 7%, respectively. Conclusion: These results demonstrate that β‐catenin alterations are relatively common in LCT, most likely occurring as subclonal events that are not enriched in cases with aggressive features. Further studies are needed to clarify the oncogenic role of β‐catenin in this tumour type. [ABSTRACT FROM AUTHOR]

Context: Testicular regression syndrome (TRS), clinically termed "vanishing testes," is a condition with atrophy of testes in late fetal or early neonatal life. The spermatic cord ends blindly or into a small fibrovascular nubbin. Histopathology reveals a fibrovascular nodule with associated hemosiderin-laden macrophages and dystrophic calcification. Few cases show the presence of seminiferous tubules and germ cells. The presence of germ cells is associated with the risk of developing a testicular germ cell tumor. Aims: To study the histopathological features of testicular nubbins and to determine the presence of germ cells and associated neoplasm. Settings and Design: It is a retro-prospective observational study carried out on patients referred to a pediatric surgical pathology center in Miraj, who were clinically diagnosed with undescended testis, over a period of 1 year. Data were collected after the study was approved by the IEC-MGCH on December 19, 2022. Materials and Methods: Twenty patients diagnosed with cryptorchidism, undescended testis, or atrophic testis were included. Findings of hematoxylin and eosin-stained slides of each case were noted, and Prussian Blue (special staining) was performed in four cases. Statistical analysis used: As this is a descriptive observational study, simple arithmetic calculations were performed. No unique statistical analysis was performed. Results: Among twenty cases of vanishing testes syndrome, 20 cases showed fibrovascular tissue only, and 1 case showed the presence of seminiferous tubules with Sertoli cells. Conclusions: Vanishing testes syndrome is a rare condition that is difficult to diagnose on clinical examination, and a definitive diagnosis may be missed if the pathologist is unaware of the condition. Also, there is a risk of germ cell tumor if there are residual germ cells in the testicular nubbin. [ABSTRACT FROM AUTHOR]

Gallbladder paraganglioma is an exceptionally uncommon tumour. Paragangliomas are a subset of Neuroendocrine Neoplasms (NENs) called neurogenic NENs that arise from neural crest cells of the neuroectoderm. They present diagnostic challenges due to their uncommon occurrence and non specific clinical manifestations. The paraganglion system consists of two cell types, called chief and sustentacular cells. The most common location of a paraganglioma is the adrenal medulla, defined as pheochromocytoma. Pheochromocytomas, found in the adrenal medulla, are known for their high Catecholamine (CA) production, leading to significant clinical manifestations like hypertension and metabolic disturbances. A 36-year-old female, otherwise asymptomatic, underwent cholecystectomy due to persistent abdominal discomfort in the right hypochondrium for two months. Preoperative imaging demonstrated a gallstone. Laparoscopic cholecystectomy was performed. The pathological analysis following surgery documented the presence of gallbladder paraganglioma and chronic cholecystitis. Immunohistochemically, chief cells displayed widespread positivity for chromogranin, synaptophysin, and sustentacular cells displayed S100 positivity. There are no definitive guidelines for the management of gallbladder paragangliomas due to their rarity. Reporting instances of paraganglioma in the gallbladder is critical to raising clinical awareness, enhancing diagnostic criteria, optimising treatment protocols, and unravelling the underlying molecular mechanisms. All of these efforts are critical for improving patient care and outcomes in treating this uncommon and enigmatic tumour entity [ABSTRACT FROM AUTHOR]

Objective The process of spermatogenesis is complex involving multiple regulatory mechanisms and requiring complex interactions between germ cells and somatic cells. The mechanisms of Sertoli cells involving in spermatogenesis remain to be explored. Methods We collected single-cell RNA sequencing data from a high-throughput gene expression database (GEO: GSE113293) from wild-type C57BIL6J male mice with normal spermatogenesis and four different mutant mouse strains (knockout Mlh3, Hormadl, and Cul4a; Cnp transgene). Then bioinformatics analyses such as gene function enrichment analysis (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) enriched pathway analysis, protein- protein interaction (PPI) network analysis, transcription factor regulation analysis (SCENIC), and gene set variation analysis (GSVA) on the Sertoli cells were further indentified. Results We identified nine subtypes of spermatogenic cell clusters (undiferentiated spermatogonia, diferentiating sperrmatogonia, leptotene/zygotene, pachytene, diplotene, round sperrmatids, elongating spermatids, and spermatozoa), and two types of somatic cell clusters (Leydig cells, and Sertoli cells), and one unknown type of cell cluster. Compared to the normal spermatogenesis model, 276 genes were upregulated and 234 genes were downregulated in the Sertoli cells of the mouse spermatogenesis defect model. GO analysis showed that differential genes in Sertoli cells were significantly enriched in spermatogenesis and energy metabolism. KEGG enrichment analyses showed that the homogeneous ribosomal pathway, glycolytic pathway, and oxidative phosphorylation pathway were significantly enriched in Sertoli cells. Protein-protein interaction (PPI) network analysis of diferentially expressed genes identified 20 hub genes, all of which belonged to ribosomal genes, and GSVA analysis revealed that the upregulated genes in Sertoli cells were closely related to the Notch signaling pathway. Conclusion The hub genes and the Notch signaling pathway in Sertoli cells are involved in the process of spermatogenesis. This study provides new perspectives on infertility caused by spermatogenesis dis orders. [ABSTRACT FROM AUTHOR]

Male infertility is a major public health concern globally with unknown etiology in approximately half of cases. The decline in total sperm count over the past four decades and the parallel increase in childhood obesity may suggest an association between these two conditions. Here, we review the molecular mechanisms through which obesity during childhood and adolescence may impair future testicular function. Several mechanisms occurring in obesity can interfere with the delicate metabolic processes taking place at the testicular level during childhood and adolescence, providing the molecular substrate to hypothesize a causal relationship between childhood obesity and the risk of low sperm counts in adulthood. Pediatric obesity is increasing worldwide and its contribution to the decline is sperm count has been questioned. By comprehensively reviewing the literature, the authors herein report molecular mechanisms that may suggest the causality of this association. [ABSTRACT FROM AUTHOR]

Nonylphenol (NP), an endocrine-disrupting chemical, is an environmental contaminant, and many notorious effects on male fertility have been reported in animal models and wild-type species. Here, we evaluated the effects of NP in follicle-stimulating hormone (FSH) signal transduction pathways and lipid metabolism using an in vitro model of rat Sertoli cell (SC) primary culture. Results show that an acute (1 h) SC exposure to NP (10 µM) increased the intraand extra-cellular concentrations of free fatty acids (FFAs), mainly arachidonic acid (20:4n-6). Phosphatidylinositol seemed to be the major phospholipid source of this 20:4n-6 release by activation of the protein kinase A (PKA)/cytoplasmic phospholipase A2 (cPLA2) pathway. NP also increased diacylglycerols (DAG) levels and the expression (mRNA) of cyclooxygenase 2 (Cox2) and prostaglandin E2 (PGE2) levels. It is noteworthy that accumulation of lipid droplets took place after 24 h NP exposition, which was prevented by both a PKA inhibitor and a PLA2 inhibitor. Like FSH, NP triggers the release of 20:4n-6, which is a substrate for PGE2 synthesis via PKA/PLA2 activation. In addition, NP induces the formation of DAG, which could be required as a cofactor of the PKC-mediated activation of the COX2 inflammatory pathway. Our findings suggest that NP alters lipid homeostasis in SCs by inducing the activation of pro-inflammatory pathways that may trigger adverse effects in testis physiology over time. Concomitantly, the SC enhances the acylation of surplus FFAs (including 20:4n-6) in neutral lipids as a protective mechanism to shield itself from lipotoxicity and pro-inflammatory signals. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV), a mosquito-borne flavivirus, is prominently associated with microcephaly in babies born to infected mothers as well as Guillain-Barré Syndrome in adults. Each cell type infected by ZIKV—neuronal cells (radial glial cells, neuronal progenitor cells, astrocytes, microglia cells, and glioblastoma stem cells) and non-neuronal cells (primary fibroblasts, epidermal keratinocytes, dendritic cells, monocytes, macrophages, and Sertoli cells)—displays its own characteristic changes to their cell physiology and has various impacts on disease. Here, we provide an in-depth review of the ZIKV life cycle and its cellular targets, and discuss the current knowledge of how infections cause neuropathologies, as well as what approaches researchers are currently taking to further advance such knowledge. A key aspect of ZIKV neuropathogenesis is virus-induced neuronal apoptosis via numerous mechanisms including cell cycle dysregulation, mitochondrial fragmentation, ER stress, and the unfolded protein response. These, in turn, result in the activation of p53-mediated intrinsic cell death pathways. A full spectrum of infection models including stem cells and co-cultures, transwells to simulate blood–tissue barriers, brain-region-specific organoids, and animal models have been developed for ZIKV research. [ABSTRACT FROM AUTHOR]

Abstract Background Inflammation-induced testicular damage is a significant contributing factor to the increasing incidence of infertility. Traditional treatments during the inflammatory phase often fail to achieve the desired fertility outcomes, necessitating innovative interventions such as cell therapy. Methods We explored the in vivo properties of intravenously administered Sertoli cells (SCs) in an acute lipopolysaccharide (LPS)-induced inflammatory mouse model. Infiltrating and resident myeloid cell phenotypes were assessed using flow cytometry. The impact of SC administration on testis morphology and germ cell quality was evaluated using computer-assisted sperm analysis (CASA) and immunohistochemistry. Results SCs demonstrated a distinctive migration pattern, importantly they preferentially concentrated in the testes and liver. SC application significantly reduced neutrophil infiltration as well as preserved the resident macrophage subpopulations. SCs upregulated MerTK expression in both interstitial and peritubular macrophages. Applied SC treatment exhibited protective effects on sperm including their motility and kinematic parameters, and maintained the physiological testicular morphology. Conclusion Our study provides compelling evidence of the therapeutic efficacy of SC transplantation in alleviating acute inflammation-induced testicular damage. These findings contribute to the expanding knowledge on the potential applications of cell-based therapies for addressing reproductive health challenges and offer a promising approach for targeted interventions in male infertility.

Abstract Yaks live in the Qinghai-Tibet Plateau for a long time where oxygen is scarce, but can ensure the smooth development of testis and spermatogenesis. The key lies in the functional regulation of the Sertoli cells under hypoxia. In this study, we sequenced yak Sertoli cells cultured in normal oxygen concentration (Normoxia) and treated with low oxygen concentration (Hypoxia) by whole transcriptomics, and screened out 194 differentially expressed mRNAs (DEmRNAs), 934 differentially expressed LncRNAs (DELncRNAs) and 129 differentially expressed miRNAs (DEmiRNAs). GO and KEGG analysis showed that these differential genes were mainly concentrated in PI3K-AKT, MAPK, RAS, and other signaling pathways, and were associated with glucose metabolism, tight junction, steroid hormone synthesis, cell fusion, and immunity of yak Sertoli cells. We constructed the gene interaction network of yak Sertoli cells in hypoxia and screened out the relationship pairs related to glucose metabolism and tight junction. The results suggested that the changes in energy metabolism, tight junction, and immune regulation of yak Sertoli cells under hypoxia might provide favorable conditions for spermatogenesis. This study provides data for further study on the role of non-coding RNA in testis development and spermatogenesis of yak.

Although bitter receptors, known as Tas2Rs, have been identified in the testes and mature sperm, their expression in testicular Sertoli cells (SCs) and their role in recognizing harmful substances to maintain the immune microenvironment remain unknown. To explore their potential function in spermatogenesis, this study utilized TM4 cells and discovered the high expression of the bitter receptor Tas2R143 in the cells. Interestingly, when the Tas2R143 gene was knocked down for 24 and 48 h, there was a significant downregulation (P < 0.05) in the expression of tight junction proteins (occludin and ZO-1) and NF-κB. Additionally, Western blot results demonstrated that the siRNA-133+NF-κB co-treatment group displayed a significant downregulation (P < 0.05) in the expression of occludin and ZO-1 compared to both the siRNA-133 transfection group and the NF-κB inhibitors treatment group. These findings suggest that Tas2R143 likely regulates the expression of occludin and ZO-1 through the NF-κB signaling pathway and provides a theoretical basis for studying the regulatory mechanism of bitter receptors in the reproductive system, aiming to attract attention to the chemical perception mechanism of spermatogenesis. • Tas2R143 affected to varying degrees the expression of BTB tight junction proteins (occludin, ZO-1) and NF-κB. • Tas2R143 likely regulates the expression of occludin and ZO-1 through the NF-κB signaling pathway in vitro. • This study proposes for the first time that the bitter receptor Tas2R143 is expressed in mouse Sertoli cells (TM4). • This study provides a theoretical basis for studying the regulatory mechanism of bitter receptors in the reproductive system. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate the signaling pathways involved in the proliferation and differentiation of pig Sertoli cells (SCs) mediated by thyroid hormone (T3) to provide a theoretical and practical basis for enhancing pig semen production. The effects of different concentrations of T3 on the proliferation of pig SCs were evaluated using the CCK8 assay. The impact of T3 on the proliferation and differentiation of pig SCs was further examined using RNA-seq, qPCR, and Western Blotting techniques. Additionally, the involvement of the p38 MAPK and NFκB pathways in mediating the effects of T3 on SCs proliferation and differentiation was investigated. Our findings revealed a strong correlation between the dosage of T3 and the inhibition of pig SCs proliferation and promotion of maturation. T3 regulated the activation state of the NFκB signaling pathway by upregulating IKKα, downregulating IKKβ, and promoting IκB phosphorylation. Furthermore, T3 facilitated SCs maturation by upregulating AR and FSHR expression while downregulating KRT-18. In conclusion, T3 inhibits pig SCs proliferation and promote pig SCs maturation through the IKK/NFκB and p38 MAPK pathways. These findings provide valuable insights into the mechanisms by which T3 influences the proliferation and maturation of pig SCs. • T3 can affect the phosphorylation status of IκB and NFκB by regulating the expression levels of IKKα and IKKβ, to regulate the proliferation of porcine testicular Sertoli cells. • T3 treatment of Sertoli cells reduces the expression of p38 protein to stimulate signals and help promote the BTB and testicular microenvironment construction. [ABSTRACT FROM AUTHOR]

Abstract Background Testis is an immune privileged organ, which prevents the immune response against sperm antigens and inflammation. Testicular cells responsible for immune tolerance are mainly Sertoli cells, which form the blood-testis barrier and produce immunosuppressive factors. Sertoli cells prevent inflammation in the testis and maintain immune tolerance by inhibiting proliferation and inducing lymphocyte apoptosis. It has been shown that 9-cis-retinoic acid (9cRA) blocks ex vivo apoptosis of peripheral blood lymphocytes and promotes the differentiation of Treg cells in the gut. However, the role of retinoid signaling in regulating the immune privilege of the testes remains unknown. Objective The aim of this study was to determine whether 9cRA, acting via the retinoic acid receptors (RAR) and the retinoic X receptors (RXR), controls the immunomodulatory functions of Sertoli cells by influencing the secretion of anti-inflammatory/pro-inflammatory factors, lymphocyte physiology and Treg cell differentiation. Methods Experiments were performed using in vitro model of co-cultures of murine Sertoli cells and T lymphocytes. Agonists and antagonists of retinoic acid receptors were used to inhibit/stimulate retinoid signaling in Sertoli cells. Results Our results have demonstrated that 9cRA inhibits the expression of immunosuppressive genes and enhances the expression of pro-inflammatory factors in Sertoli cells and lymphocytes, increases lymphocyte viability and decreases apoptosis rate. Moreover, we have found that 9cRA blocks lymphocyte apoptosis acting through both RAR and RXR and inhibiting FasL/Fas/Caspase 8 and Bax/Bcl-2/Caspase 9 pathways. Finally, we have shown that 9cRA signaling in Sertoli cells inhibits Treg differentiation. Conclusion Collectively, our results indicate that retinoid signaling negatively regulates immunologically privileged functions of Sertoli cells, crucial for ensuring male fertility. 9cRA inhibits lymphocyte apoptosis, which can be related to the development of autoimmunity, inflammation, and, in consequence, infertility.