Cells In Seminiferous Tubules Research Articles

Phytochemical Evaluation of Lepidium meyenii, Trigonella foenum-graecum, Spirulina platensis, and Tribulus arabica, and Their Potential Effect on Monosodium Glutamate Induced Male Reproductive Dysfunction in Adult Wistar Rats.

Monosodium glutamate (MSG), a sodium salt derived from glutamic acid, is widely used in commercial food products to improve taste, quality, and preservation. However, its consumption may have detrimental effects on male reproductive function. Nevertheless, plant extracts, such as Lepidium meyenii (Maca), Trigonella foenum-graecum (Fenugreek), Spirulina platensis (Spirulina), and Tribulus arabica (Tribulus), may ameliorate these adverse effects. To this effect, the phytochemical properties of Lepidium meyenii, Trigonella foenum-graecum, Spirulina platensis, and Tribulus arabica were assessed, and their potential impact on MSG-induced impairment of reproductive parameters was examined. The phytochemical composition (steroids, terpenes, phenols, flavonoids) of the plants was profiled through spectrophotometry and the antioxidant activity was assessed using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay. Thirty-six male Wistar rats were divided into six groups at random: a control group receiving distilled water, and five experimental groups (MSG, Maca, Fenugreek, Spirulina, and Tribulus) receiving 900 mg/kg/day of MSG dissolved in water for 45 days. Subsequently, the animals in the experimental groups were administered 500 mg/kg/day of the respective plant extract via oral gavage for an additional 35 days, while the MSG group continued to receive water only. Following the treatment period, the animals were sacrificed, and their reproductive tract organs were collected, weighed, and subjected to further analysis. Phytochemical analysis revealed the presence of diverse bioactive elements in the plant extracts, including phenolic and flavonoid compounds. Exposure to MSG negatively impacted total and progressive sperm motility, which was ameliorated by Lepidium meyenii treatment. Sperm morphology showed no significant differences among groups. Treatment of the phytochemical agents diminished histomorphometric alternations of the testicular length, germinal epithelium height, and number of cells in seminiferous tubules, which were caused by the initial administration of MSG. Testosterone and LH levels were reduced in the MSG group but improved in extract-treated groups. The study suggests Lepidium meyenii as a potential remedy for reproductive dysfunction. However, further investigation into its mechanisms and human safety and efficacy is warranted.

Improving Fertility in Non-obstructive Azoospermia: Results from an Autologous Bone Mar-row-Derived Mesenchymal Stromal/Stem Cell Phase I Clinical Trial.

In this phase I clinical trial, our primary objective was to develop an innovative therapeutic approach utilizing autologous bone marrow-derived mesenchymal stromal/stem cells (BM-MSCs) for the treatment of nonobstructive azoospermia (NOA). Additionally, we aimed to assess the feasibility and safety of this approach. We recruited 80 participants in this non-randomized, open-label clinical trial, including patients undergoing NOA treatment using autologous BM-MSCs (n=40) and those receiving hormone therapy as a control group (n=40). Detailed participant characteristics, such as age, baseline hormonal profiles, etiology of NOA, and medical history, were thoroughly documented. Autotransplantation of BM-MSCs into the testicular network was achieved using microsurgical testicular sperm extraction (microTESE). Semen analysis and hormonal assessments were performed both before and six months after treatment. Additionally, we conducted an in-silico analysis to explore potential protein-protein interactions between exosomes secreted from BM-MSCs and receptors present in human seminiferous tubule cells. Our results revealed significant improvements following treatment, including increased testosterone and inhibin B levels, elevated sperm concentration, and reduced levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and prolactin. Notably, in nine patients (22.5%) previously diagnosed with secondary infertility and exhibiting azoospermia before treatment, the proposed approach yielded successful outcomes, as indicated by hormonal profile changes over six months. Importantly, these improvements were achieved without complications. Additionally, our in-silico analysis identified potential binding interactions between the protein content of BM-MSC-derived exosomes and receptors integral to spermatogenesis. Autotransplantation of BM-MSCs into the testicular network using microTESE in NOA patients led to the regeneration of seminiferous tubules and the regulation of hormonal profiles governing spermatogenesis. Our findings support the safety and effectiveness of autologous BM-MSCs as a promising treatment modality for NOA, with a particular focus on the achieved outcomes in patients with secondary infertility (registration number: IRCT20190519043634N1).

The effects of saline water consumption on sperm parameters, testicular histopathology, hormonal and antioxidants concentrations in Barki Rams

The study aimed to assess the effects of water salinity on the sperm parameters, levels of cortisol, LH, FSH, testosterone and antioxidants as well as the testes’ histopathology in Barki rams. Fifteen healthy Barki rams (1–1.5 years) were divided into three equal depending on the type of drinking water for nine months. The rams in the tap water group (TW, water that contained 350 ppm of total dissolved salts (TDS). Males in the high saline water group (HSW) were permitted to consume high saline water with 8,934 ppm TDS, whereas those in the second group were permitted to have moderately saline water (MSW, 4,557 ppm TDS). High salt concentration in drinking water had adverse effect on sperm viability, morphology and sperm cell concertation. Nitric oxide and malondialdehyde concentrations in blood were significantly higher in the MSW and HSW groups than in TW. There was a significant decrease in glutathione concentration as well as superoxide dismutase activity in TDS and HSW. Cortisol was most highly concentrated in the HSW, next in the MSW, and least in TW. The testosterone, LH, and FSH concentrations in the HSW and MSW groups were significantly lower than in TW. As the salt concentration in drinking water increases, damage to testicular tissue. The MSW group demonstrating vacuolation of lining epithelial cells with pyknotic nuclei in the epididymis and necrosis and desquamation of spermatogenic cells in seminiferous tubules while HSW group displaying desquamated necrotic cells and giant cell formation in the epididymis, as well as damage to some of the seminiferous tubules and showed congestion, vacuolation of spermatogenic epithelium of seminiferous tubules, and desquamated necrotic spermatogenic epithelium. In conclusion, the salinity of the water has detrimental impacts on the sperm morphology, viability and concentration, hormones and antioxidant levels in Barki rams.

Granulocyte-macrophage colony-stimulating factor (GM-CSF)-induced maturation of spermatogonial cells from prepubertal mice in vitro is enhanced by testosterone.

Spermatogenesis is the complicated process of sperm generation. During this process, spermatogonial cells proliferate and differentiate via meiotic and post-meiotic stages to produce mature sperm. This process is under the regulation of testicular autocrine/paracrine factors. In addition, endocrine factors are crucial to complete spermatogenesis. We aimed to localize granulocyte-macrophage colony-stimulating factor (GM-CSF) and its receptor (GM-CSFR) in testicular cells and further evaluate its involvement in the development of spermatogenesis in vitro. Weisolated cells from seminiferous tubule cells of seven-day-old mice and cultured them in vitro using a methylcellulose culture system (MCS), in the presence of GM-CSF and/or testosterone for four weeks. The cells were then examined for markers of different stages of spermatogenesis by immunofluorescence staining and/or qPCR analyses. Our results revealed the presence of GM-CSF and GM-CSFR in testicular cells (premeiotic and meiotic cells as well as somatic cells; Leydig and Sertoli cells). We further demonstrated the development of colonies/spheroids in the MCS which contained pre-meiotic, meiotic, and post-meiotic cells. The addition of GM-CSF to the MCS significantly increased the percentage of pre-meiotic and meiotic cells compared to control. Furthermore, the addition of GM-CSF and testosterone together significantly increased the percentage of cells in the post-meiotic stage compared to the addition of each separately. In conclusion, our results indicate that testicular cells express GM-CSF/GM-CSFR, and that GM-CSF is involved in the development of different stages of spermatogenesis in vitro. Furthermore, testosterone enhances the development of spermatogenic cells and potentiates the effect of GMCSF on the development of post-meiotic cells. These findings provide evidence that GM-CSF and testosterone are involved in the development of spermatogenesis in vitro and in vivo. In brief: Testicular somatic and germ cells express GM-CSF and GM-CSFR. Our study suggests that testicular GM-CSF is involved in the development of spermatogenesis, which is potentiated by testosterone.

The Effect of Fermented Honey on Seminiferous Tubule Cells While being Induced by Cisplatin and Islamic Perspective Review

Background: Cisplatin is the most effective cancer treatment used in chemotherapy. Cisplatin works by inducing oxidative stress, but it also has toxic side effects on healthy cells in the male reproductive organs. Therefore, there is a need for a solution to reduce the side effects of cisplatin. Honey fermentation is a natural substance that can reduce the toxicity of cisplatin. Honey is known as “The food of God” which is a gift of sustenance from Allah SWT due to its extraordinary benefits. Methodology: This study utilized primary data with an experimental posttest-only control group design conducted in vivo. The research involved four experimental groups of rats: a control group without any treatment, a cisplatin group, a 5% honey fermentation group exposed to cisplatin, and a 10% honey fermentation group exposed to cisplatin. All rats were weighed before, during, and after the treatment. Subsequently, HE staining was performed for both descriptive and quantitative observations. Results: Honey fermentation can provide a protective effect on primary spermatocytes and Leydig interstitial cells, almost reaching normal levels. The highest improvement was observed in the 5% honey fermentation group for primary spermatocytes and the 10% group for Leydig interstitial cells. Conclusion: Honey fermentation administration can be protective for testicular organ cells against the side effects of cisplatin. From an Islamic perspective, Allah has created honey for consumption, as mentioned in Surah An-Nahl.

Predictions from a mathematical approach to model ionic signaling for rapid responses of Sertoli cells exhibit similarities to pharmacological approaches

Sertoli cells are essential for the male reproduction system as they provide morphological support and nutrients for germ cells to guarantee ongoing spermatogenesis. The aim of this work was to predict the electrical properties at the plasma membrane that trigger Sertoli cell rapid responses by involving ionic channels. The rapid responses of Sertoli cells in culture were monitored using patch clamp electrical measurement and compared to data obtained using pharmacological tools (from intact seminiferous tubules). A mathematical model was used to define the roles of potassium channels and the ATP-dependent Na+/K+ pump in these responses. Mathematical data verification was also performed to determine the resting and hormonal stimulated membrane potentials of Sertoli cells in the intact seminiferous tubules and of Sertoli cells in culture (patch clamp measurements). The prediction of these data based on mathematical modeling demonstrated, for the first time, the involvement of potassium channels and the activation of Na+/K+ pump in the hyperpolarization of Sertoli cells and their consequent rapid responses. Moreover, the mathematical analysis showing the involvement of ionic balance in the rapid responses of these cells to hormones, such as follicle-stimulating hormone, is consistent with previous reports obtained using pharmacological techniques in Sertoli cells. Thus, the validation of such data is reliable and represents a first step in the proposition for a mathematical model to predict rapid responses of Sertoli cells to hormonal stimuli.

Protective effect of paricalcitol in rat testicular damage induced by subchronic 1800 MHz radiofrequency radiation

In the present study, the possible protective effects of paricalcitol (P) were investigated in testicular damage because of 1800 MHz radiofrequency radiation (RFR) exposure. Male Sprague Dawley rats 8–10 weeks old (n = 28) were randomly divided into four groups as control (C) (n = 7), RFR (n = 7, 1800 MHz RFR 1 h/day for 30 days), P (n = 7, 0.2 μg/kg paricalcitol, 3 times a week for 30 days), and RFR + P (n = 7, 1800 MHz RFR 1 h/day for 30 days +0.2 μg/kg paricalcitol, 3 times a week for 30 days). Testicular tissue was evaluated with histological and biochemical methods. No statistically significant differences were detected between the groups in seminiferous tubule diameters and germinal epithelial thicknesses. While ultrastructural changes were observed in the seminiferous tubule and Leydig cells in the RFR group, these changes were decreased in the RFR + P group. It was found that the Johnsen Score, Ki67, and p63 immunoreactivity scores (IRS), superoxide dismutase (SOD), and catalase (CAT) activities in the RFR + P group were statistically increased as compared to the RFR group and the malondialdehyde (MDA) levels were decreased statistically and significantly. These results show that paricalcitol administration may have an ameliorative effect on testicular damage occurring because of 1800 MHz RFR exposure.

Silk Dye Effluent Induced Change in Ultra-Structure of Testis on Swiss Albino Male Mice Mus Musculus and Recovery by Moringa Oleifera Leaves Powder

The silk dye effluent is one of the most potential harmful chemicals liberated in the environment in an unexpected manner. Silk dye waste is widely used as a potent dyeing of yarn and fabrics in many countries and has been shown to produce some adverse health effects. This work focuses primarily on the effects of Moringa oleifera leaf extract on testis of silk dye effluent induced surface ultrastructure in Swiss albino mice Mus musculus. The testis has been taken an account for surface ultrastructural study. The mice were divided into 5 Groups i.e. Group I (Control), Group II (fed with 50% silk dye), Group III (fed with 100% silk dye), Group IV (mice fed with 50% dye treated with M. oleifera leaves powder), Group V (mice fed with 100% dye treated with M. oleifera leaves powder) have been taken for experiment. The dose of silk dye was 2ml/day to both groups II and III and M. oleifera leaf is given as per the standard dose (300mg/kg b.w) to both animals of group IV and V. Administration of silk dye waste result were demonstrated disrupted spermatids, cessation of seminiferous tubule cell, irregular cavity, lump of seminiferous tubules, disrupted interstitial cell and occupied spermatogonial cells but used of Moringa oleifera leafs powder it was significantly recovered the damage tissues has been observed. This study suggested that the extract may have beneficial effect on surface ultrastructural constituents such as Testis.
 Key words: Silk dye Effluent, Moringa oleifera leaf powder, Testis, Mus musculus, SEM for Ultra-Structure, Toxicity assessment.

Chronic levamisole exposure in male rats alters sexual behavior and induces apoptosis in the testis

Levamisole is an anti-helminthic drug developed and introduced in veterinary medicine, and it has been used more frequently after the inclusion of its usage in human medicine regarding disorders with immunomodulatory properties. In recent years, it has started to attract attention since it has beneficial effects on the treatment of COVID-19 due to its immunomodulatory properties. To investigate the effects of levamisole on sexual behavior and the reproductive system in male rats, two groups were formed the vehicle (n = 10) and levamisole (n = 10) groups. The vehicle group was given purified water whereas the levamisole group was administered with levamisole (2 mg/kg) by oral gavage daily for 4 weeks. Levamisole treatment significantly increased the mount latency (ML, P < 0.001) as well as the intromission latency (IL, P < 0.01). It also significantly prolonged postejaculatory interval (PEI, P < 0.01), decreased copulatory rate (CR, P < 0.05), and sexual activity index (SAI, P < 0.05). It significantly decreased serum monoamine oxidase A (MAO-A) levels (P < 0.05). Additionally, levamisole induced disorganizations of germinal epithelial cells of seminiferous tubules, congestion, edema in the interstitial area, and metaphase arrest in some spermatocytes (P < 0.001), and it significantly increased the immunohistochemical expressions of apoptotic Bax and cytochrome c, which is crucial proapoptotic protein, in the testis (P < 0.001). Also, levamisole significantly upregulated the mRNA levels of the apoptosis-related key regulatory genes, including Bax (Bcl-2-associated X protein, P = 0.05) and Bax/Bcl-2 ratio (P < 0.01) in testis. The current research is the first to show that levamisole may decrease sexual performance, potency, sexual motivation, and libido and induce apoptosis in the testis.

In vitro evidence against productive SARS-CoV-2 infection of human testicular cells: Bystander effects of infection mediate testicular injury.

The hallmark of severe COVID-19 involves systemic cytokine storm and multi-organ injury including testicular inflammation, reduced testosterone, and germ cell depletion. The ACE2 receptor is also expressed in the resident testicular cells, however, SARS-CoV-2 infection and mechanisms of testicular injury are not fully understood. The testicular injury could be initiated by direct virus infection or exposure to systemic inflammatory mediators or viral antigens. We characterized SARS-CoV-2 infection in different human testicular 2D and 3D culture systems including primary Sertoli cells, Leydig cells, mixed seminiferous tubule cells (STC), and 3D human testicular organoids (HTO). Data shows that SARS-CoV-2 does not productively infect any testicular cell type. However, exposure of STC and HTO to inflammatory supernatant from infected airway epithelial cells and COVID-19 plasma decreased cell viability and resulted in the death of undifferentiated spermatogonia. Further, exposure to only SARS-CoV-2 Envelope protein caused inflammatory response and cytopathic effects dependent on TLR2, while Spike 1 or Nucleocapsid proteins did not. A similar trend was observed in the K18-hACE2 transgenic mice which demonstrated a disrupted tissue architecture with no evidence of virus replication in the testis that correlated with peak lung inflammation. Virus antigens including Spike 1 and Envelope proteins were also detected in the serum during the acute stage of the disease. Collectively, these data strongly suggest that testicular injury associated with SARS-CoV-2 infection is likely an indirect effect of exposure to systemic inflammation and/or SARS-CoV-2 antigens. Data also provide novel insights into the mechanism of testicular injury and could explain the clinical manifestation of testicular symptoms associated with severe COVID-19.

Evaluation of testicles by sonoelastography in men recovering after Covid-19 disease

IntroductionTesticular cells, seminiferous tubule cells, spermatogonia, Leydig and Sertoli cells showing angiotensin-converting enzyme 2 expression have the potential to be targets and to be damaged by the coronavirus. We aimed to use Two-Dimensional Shear Wave Elastography (2D-SWE) as an effective technique to identify parenchymal damage in the testicles of patients recovering from COVID-19 infection. Methods35 Male patients (group 1) who recovered after COVID-19 infection between 4 and 12 weeks were included in this prospective study. Before 2D-SWE, these male patients were confirmed with control Rt-PCR test negativity. In addition, the first Rt-PCR test positivity of these patients was confirmed. A control group was formed of 31 healthy subjects (group 2). These two groups were compared in terms of age, volume of each testis, and SWE values. Ultrasound including SWE was applied to all the testes. A total of 9 measurements were taken as 3 SWE measurements from each third of the testis (superior, mid, inferior) and the average of these was calculated. Data obtained in the study were analyzed statistically. A value of p < 0.05 was accepted as statistically significant. ResultsThe mean SWE values for the right testis and the left testis were determined to be statistically significantly higher in Group 1 than in Group 2, respectively (p < 0.001, p < 0.001). ConclusionThere is an increase in testicular stiffness in males who have recovered from COVID-19 infection. The underlying cause of testicular damage is changes at the cellular level. The 2D-SWE technique can predict potential testicular parenchymal damage in male patients recovering from COVID-19 infection. Implications for practiceTwo-Dimensional Shear Wave Elastography (2D-SWE) seems to be a promising imaging technique in the evaluation of testis parenchyma.

134 Late-gestation maternal overnutrition alters gene expression and histomorphology in neonatal foal testes

While obesity has deleterious effects on metabolism, performance, lameness, and health in mature horses, little is known about the impacts to reproductive health in foals from overfed dams. The objective of this study was to determine the effects of maternal overnutrition on neonatal foal testicular development. To test this, 16 Quarter horse mares were used in a completely randomized design and stratified by expected foaling date, body weight, and body condition score into one of 2 dietary treatments offered twice daily beginning on gestation d 235: control (CON; n = 8) fed to meet 100% of nutrient requirements and overfed (HIGH; n = 8) fed to 140% of nutrient requirements. At 5 h postpartum, foals were humanely euthanized, and testes were harvested, weighed, and stored for analyses. Testicular weight included the tunica dartos with the epididymis removed. RNA was extracted using the PureLink RNA Mini Kit (Life Technologies, Austin, TX) and submitted for RNA sequencing (Novogene, Sacramento, CA). Histomorphometric evaluations were performed using a hematoxylin and eosin stain. Ten 10X images per animal were analyzed using ImageJ software for individual seminiferous tubule (ST) area, ST number, and % area of STs per 10X field relative to interstitial tissue. Sertoli cell numbers were determined at 40X using 20 seminiferoustubules per animal. Data were analyzed using the MIXED procedure in SAS. Testes from HIGH male foals (n = 4; LSMeans ± SEM; 32.93 ± 1.93 g) were 61% larger than testes from CON male foals (n = 4; 20.50 ± 1.93 g; P < 0.01). RNA sequencing revealed 56 differentially expressed genes between HIGH and CON groups (P-adjusted <0.05). These genes represent biochemical pathways involved in oxidative phosphorylation, fatty acid metabolism, and thermogenesis, which are integral for testicular development and function. Gene ontology analyses revealed differential gene expression was associated with biological processes relevant to reproductive function and spermatogenesis, including cellular respiration and mitochondrial function. Histological analyses revealed no difference in individual ST area (P = 0.76) or Sertoli cell number within STs (P = 0.57); however, testes from CON foals had 2.3x more STs per 10X field (21.68 ± 2.48) than HIGH foals (9.35 ± 2.48; P = 0.01) along with STs representing a greater percentage of the image area relative to interstitial tissue (CON 3.66% ± 0.61%; HIGH 1.60% ± 0.61%; P = 0.05). These data from the neonatal foal indicate that late-gestation maternal overnutrition alters fetal testicular development with potential long-term implications.

Disrupted intercellular bridges and spermatogenesis in fatty acyl-CoA reductase 1 knockout mice: A new model ofether lipid deficiency.

Peroxisomal fatty acyl-CoA reductase 1 (FAR1) is a rate-limiting enzyme for ether lipid (EL) synthesis. Gene mutations in FAR1 cause a rare human disease. Furthermore, altered EL homeostasis has also been associated with various prevalent human diseases. Despite their importance in human health, the exact cellular functions of FAR1 and EL are not well-understood. Here, we report the generation and initial characterization of the first Far1 knockout (KO) mouse model. Far1 KO mice were subviable and displayed growth retardation. The adult KO male mice had smaller testes and were infertile. H&E and immunofluorescent staining showed fewer germ cells in seminiferous tubules. Round spermatids were present but no elongated spermatids or spermatozoa were observed, suggesting a spermatogenesis arrest at this stage. Large multi-nucleated giant cells (MGC) were found lining the lumen of seminiferous tubules with many of them undergoing apoptosis. The immunofluorescent signal of TEX14, an essential component of intercellular bridges (ICB) between developing germ cells, was greatly reduced and mislocalized in KO testis, suggesting the disrupted ICBs as an underlying cause of MGC formation. Integrative analysis of our total testis RNA-sequencing results and published single-cell RNA-sequencing data unveiled cell type-specific molecular alterations underlying the spermatogenesis arrest. Many genes essential for late germ cell development showed dramatic downregulation, whereas genes essential for extracellular matrix dynamics and cell-cell interactions were among the most upregulated genes. Together, this work identified the cell type-specific requirement of ELs in spermatogenesis and suggested a critical role of Far1/ELs in the formation/maintenance of ICB during meiosis.

MP01-11 IN VITRO PROPAGATION OF UNDIFFERENTIATED SPERMATOGONIA ISOLATED FROM A 46, XX MALE TESTIS

You have accessJournal of UrologyCME1 Apr 2023MP01-11 IN VITRO PROPAGATION OF UNDIFFERENTIATED SPERMATOGONIA ISOLATED FROM A 46, XX MALE TESTIS Mark Xu, Aaron Bradshaw, Harpreet Gosal, Abinav Udaiyar, Omar Abdelaal, Kimberly Stogner-Underwood, Stuart Howards, Tamer Yalcinkaya, Anthony Atala, and Hooman Sadri-Ardekani Mark XuMark Xu More articles by this author , Aaron BradshawAaron Bradshaw More articles by this author , Harpreet GosalHarpreet Gosal More articles by this author , Abinav UdaiyarAbinav Udaiyar More articles by this author , Omar AbdelaalOmar Abdelaal More articles by this author , Kimberly Stogner-UnderwoodKimberly Stogner-Underwood More articles by this author , Stuart HowardsStuart Howards More articles by this author , Tamer YalcinkayaTamer Yalcinkaya More articles by this author , Anthony AtalaAnthony Atala More articles by this author , and Hooman Sadri-ArdekaniHooman Sadri-Ardekani More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003212.11AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: The SRY-positive 46, XX genotype is a testicular disorder of sex development found in 1/20,000 males. Patients are phenotypically male with small testes, variable hypogonadism, and azoospermia. Current literature has not yet documented spermatogonia in 46 XX male patients. This study aimed to identify and propagate spermatogonia in vitro in a 46, XX male. METHODS: A 28-year-old Caucasian male was initially suspected of having Klinefelter Syndrome (XXY), but karyotype and FISH analysis found a 46, XX genotype with a Y chromosome translocation containing the SRY gene onto the short distal arm of the X chromosome. He underwent experimental bilateral microscopic testicular biopsy and tissue preservation under IRB protocol. Fresh tissue was sent for histopathologic analysis using H&E staining and immunostaining of undifferentiated spermatogenic markers, including PGP 9.5/UCHL1 and ZBTB16. Biopsy portions from bilateral testes were used for characterization. Digital PCR and Real-time Quantitative (RT-Q) PCR were performed with ZBTB16 (undifferentiated spermatogonia), STAR (Leydig cell), CD34 (Peritubular cell), and SOX9 (Sertoli cell) target primers on testis samples from both sides. Isolated cells from the right testicular biopsy were cultured in the StemPro medium for four weeks. RT-QPCR was performed with target primers for undifferentiated spermatogonia (ZBTB16, UCHL1, THY1, CD9, ITGA6, ITGB1), Sertoli (SOX9, GATA4, CYP19a1), Leydig (STAR, CALB2), and peritubular cells (CD34). Digital PCR was performed using ZBTB16 target primer to quantify undifferentiated spermatogonia. Flow cytometry on cultured cells identified the spermatogonia stem cell (SSC) population as negative for HLA-ABC and positive for CD9 and CD49f. RESULTS: Testes exhibited Leydig cell hyperplasia, seminiferous tubule hyalinization like in Klinefelter Syndrome, and tubules filled mainly with Sertoli cells. Rare cells in seminiferous tubules were weakly positive for PGP9.5 and ZBTB16, representing undifferentiated spermatogonia. Digital and RT-QPCRs of testis tissue and cultured cells found evidence of all major testicular somatic cells and undifferentiated spermatogonia. Digital PCR for ZBTB16 identified 7.61% of cultured cells as undifferentiated spermatogonia and 0.71% as SSCs. Flow cytometry identified 2.52% of cultured cells as SSCs. CONCLUSIONS: This is the first study suggesting the presence of SSCs in 46 XX male testes. Future work will focus on further characterization of cultured testicular cells and In Vitro or In Vivo differentiation of SSCs from XX male testis. Source of Funding: WFIRM © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e6 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Mark Xu More articles by this author Aaron Bradshaw More articles by this author Harpreet Gosal More articles by this author Abinav Udaiyar More articles by this author Omar Abdelaal More articles by this author Kimberly Stogner-Underwood More articles by this author Stuart Howards More articles by this author Tamer Yalcinkaya More articles by this author Anthony Atala More articles by this author Hooman Sadri-Ardekani More articles by this author Expand All Advertisement PDF downloadLoading ...

Male reproductive toxicity of polystyrene microplastics: Study on the endoplasmic reticulum stress signaling pathway.

Microplastics (MPs) have raised health concerns in public for its potential reproductive toxicity. In this study, we subjected the Kunming mice to 0.01, 0.1 and 1.0mg/day polystyrene MPs (10μm, PS-MPs) for 35 days, aiming to investigate the relevant male reproductive toxicity and latent molecular mechanism. The results showed the decreased sperm counts and motility, while the elevated sperm abnormality in PS-MPs-exposed mice. Testicular H&E staining displayed the vacuolization, atrophy, and even shedding of germ cells in seminiferous tubule. And the testosterone content in serum also decreased with PS-MPs treatment. Moreover, molecular analysis indicated that PS-MPs upregulated the expression trait factors for ERS (e.g., immunoglobulin-binding protein [BIP], inositol-requiring protein 1α [IRE1α], X-box-binding protein 1 splicing [XBP1s], Jun kinase [JNK], and the transcription of CCAAT/enhancer-binding protein (C/EBP) homologous protein [CHOP]) and downstream apoptotic modulator (e.g., Caspase-12, -9, and -3) in the testis. The steroidogenic acute regulatory protein (StAR), the testosterone synthetic initiator, was also downregulated. With the supplementation of ERS inhibitor, the MPs-induced testicular damage and decreased testosterone were improved to almost normal level. Overall, this study suggested that PS-MPs generate reproductive toxicity possibly via activating ERS and apoptosis signaling pathway.

Effects of Insulin-degrading enzyme on the proliferation of swine Sertoli cells.

Sertoli cells, the only somatic cells in testis seminiferous tubules, provide a supporting microenvironment for male germ cells and play essential roles in spermatogenesis. The insulin-degrading enzyme (IDE), a ubiquitous zinc peptidase of the inverzincin family, plays crucial role in sperm production, as IDE-knockout mice presented decreased testis weight and impaired sperm viability and morphology. However, whether and how IDE affects swine Sertoli cell proliferation remains unclear. Thus, in the present study, we aimed to evaluate the effects of IDE on the proliferation of swine Sertoli cells, as well as its underlying molecular mechanism. After knocking down IDE expression with small interfering RNA transfection, we analyzed the proliferation of swine Sertoli cells as well as the expression of related regulatory factors (WT1, ERK, and AKT). The results showed that IDE knockdown promoted swine Sertoli cell proliferation and increased WT1 expression, possibly through activating ERK and AKT. Overall, our findings suggest that IDE may be involved in male reproduction by regulating Sertoli cell proliferation, which provides new information to better understand the regulatory mechanisms of swine Sertoli cells and improve the reproductive traits of male pigs.

Preserving the Normal Healthy Testis: Role of Beta-carotene

Background: Many drugs can have significant deleterious side effects on the different organs of the body which can change normal physiology to different pathologies. Recent research have been focusing on valuable methods to preserve the normal healthy testis from the medications’ adverse effects. The study aims to investigate the role of beta-carotene in the protection against potential amikacin’s testicular toxicity. Materials and methods: Male rats were involved in the study. Rats have divided into four groups: the first group was control, the second group has been given amikacin as an inducer of testicular toxicity; the third group has been given both amikacin and beta-carotene, and the fourth group has been given beta-carotene as prophylaxis before induction of the toxicity. The histological architecture of the testis was investigated for all the rats. Results: Amikacin caused a significant deleterious effect which included degeneration and necrosis of cells of seminiferous tubules as well as atrophy and congestion of blood vessels. Co-administration of both beta-carotene and amikacin resulted in a partial improvement in the testicular tissue while using beta-carotene as prophylaxis succeeded in protecting and preserving the normal histological features of the testis completely. Conclusion: The study concluded that beta-carotene can preserve the normal physiological testis and protect it against the amikacin’s deleterious effect on the testis.

Silver nanoparticles induced testicular damage targeting NQO1 and APE1 dysregulation, apoptosis via Bax/Bcl-2 pathway, fibrosis via TGF-β/α-SMA upregulation in rats

In medicine, silver nanoparticles (AgNPs) are employed often. They do, however, have negative impacts, particularly on the reproductive organs. This research aimed to assess AgNP impact on the testis and the possible intracellular mechanisms to induce testicular deteriorations in rats at various concentrations and different time intervals. Sprague Dawley rats (n = 40) were allocated into four equal groups: the control one, and three other groups injected intra-peritoneally with AgNP solution 0.25, 0.5, and 1 mg/kg b.w. respectively for 15 and 30 days. Our findings revealed that AgNPs reduced body and testicular weights, estradiol (E2) and testosterone (T) hormone levels, and sperm parameters while elevating the nitric oxide and malondialdehyde levels with inhibition of reduced glutathione contents in testicular tissue. Interestingly, AgNPs significantly upregulated the testicular inducible nitric oxide synthase, B cell lymphoma 2 (Bcl-2)-associated X, transforming growth factor, and alpha-smooth muscle actin (α-SMA) expression levels. However, apurinic/apyrimidinic endo deoxyribonuclease 1 (APE1), NAD (P) H quinone dehydrogenase 1 (NQO1), and Bcl-2 expression levels were all downregulated indicating exhaustion of body antioxidant and repairing defense mechanisms in testicles in comparison with the control rats. Various histological alterations were also detected which dramatically increased in rats sacrificed after 30 days such as loss of the lining cells of seminiferous tubules with no spermatozoa and tubular irregularities associated with thickening of their basement membranes. Immunolabeling implicated in the apoptotic pathway revealed a negative expression of Bcl-2 and marked immunoreactivity for caspase-3 after 30 days of AgNP treatment in comparison to the control rats. To our knowledge, there have been no previous publications on the role of the α-SMA, APE1, and NQO1 genes in the molecular pathogenesis of AgNP testicular cytotoxicity following AgNP acute and chronic exposure.Graphical

Towards the mechanism of spermatotoxicity of p-tert-butyl-alpha-methylhydrocinnamic aldehyde: inhibition of late stage ex-vivo spermatogenesis in rat seminiferous tubule cultures by para-tert-butyl- benzoic acid.

Molecules metabolized to para-tert-butyl-benzoic acid (p-TBBA) affect male reproduction in rats through effects on spermatogenesis. This toxicity is specific to p-TBBA and not observed in meta-substituted analogues. The underlying mode of action was evaluated by comparing effects of p-TBBA and the position isomer m-TBBA (2-50µM) in an ex vivo 3D primary seminiferous tubule cell culture system from juvenile Sprague Dawley rats (Bio-AlteR®). Treated cultures were evaluated for CoA-conjugate formation, cytotoxicity, blood-testis barrier functionality and different germ cell populations to assess effects on spermatogenesis. In addition, an evaluation of the metabolome of treated cultures was performed by using MxP® Broad Profiling via a LC-MS/MS and GC-MS platform. Para-TBBA decreased germ cell populations of late stages of spermatogenesis and led to the formation of CoA-conjugates in the ex vivo tissue. In addition, p-TBBA had a pronounced effect on the metabolome by affecting lipid balance and other CoA-dependent pathways contributing to energy production and the redox system. Meta-TBBA did not affect germ cell populations and no m-TBBA related CoA-conjugates were detectable. The metabolic profile of m-TBBA treated cells was comparable to vehicle control treated cultures, indicating that formation of CoA-conjugates, inhibition of spermatogenesis, and effects on the metabolome are mechanistically linked events. Thus, for this specific chemical group an adverse outcome pathway can be postulated, including the formation of benzoic acid metabolites, accumulation of CoA-conjugates to a certain threshold and CoA depletion, which affects the metabolic and lipid profile and leads to tissue specific effects with impaired functionalities such as spermatogenesis.

As3MT-mediated SAM consumption, which inhibits the methylation of histones and LINE1, is involved in arsenic-induced male reproductive damage

Studies have demonstrated that arsenic (As) induces male reproductive injury, however, the mechanism remains unknown. The high levels of arsenic (3) methyltransferase (As3MT) promote As-induced male reproductive toxicity. For As-exposed mice, the germ cells in seminiferous tubules and sperm quality were reduced. Exposure to As caused lower S-adenosylmethionine (SAM) and 5-methylcytosine (5 mC) levels, histone and DNA hypomethylation, upregulation of long interspersed element class 1 (LINE1, or L1), defective repair of double-strand breaks (DSBs), and the arrest of meiosis, resulting in apoptosis of germ cells and lower litter size. For GC-2spd (GC-2) cells, As induced apoptosis, which was prevented by adding SAM or by reducing the expression of As3MT. The levels of LINE1, affected by SAM content, were involved in As-induced apoptosis. Furthermore, folic acid (FA) and vitamin B12 (VB12) supplements restored SAM, 5 mC, and LINE1 levels and blocked impairment of spermatogenesis and testes and lower litter size. Exposed to As, mice with As3MT knockdown showed less impairment of spermatogenesis and testes and greater litter size compared to As-exposed wild-type (WT) mice. Thus, the high As3MT levels induced by As consume SAM and block histone and LINE1 DNA methylation, elevating LINE1 expression and evoking impairment of spermatogenesis, which causes male reproductive damage. Overall, we have found a mechanism for As-induced male reproductive damage, which provides biological insights into the alleviation of reproductive injury induced by environmental factors.