Histological aspects of the renal sexual segment of Brazilian snakes of the Boidae family.

Snakes represent a wide and diverse group of species and have anatomical particularities, such as the renal sexual segment (RSS), a structure located in the kidneys and formed from the hypertrophy of the urinary ducts and nephrons. This study aims at describing the histological aspects of the RSS of Boa constrictor, Epicrates cenchria and Corallus hortulanus, all of which are Brazilian snake species from the Boidae family. The reproductive system and kidneys of five male specimens of E. cenchria, three male specimens of C. hortulanus and two male specimens of B. constrictor were obtained. Tissue samples were processed histologically and different stains used (Toluidine Blue, Alcian Blue and Periodic Acid Schiff). The histological evaluation of the RSS of E. cenchria, C. hortulanus and B. constrictor shows that the RSS in these species varies when comparing individuals in the reproductive period with those which are not. It also allows for the observation of the segment's secretory activity in animals in the reproductive stage (mature sperm in the lumen of the seminiferous tubules) as well as in those which are not. Finally, the histological evaluation also reveals the variation of the secretion product in individuals in the reproductive period, in those which are not, and also among individuals within the same reproductive stage.

Effect of IL-1ß on the Development of Spermatogenesis In Vitro in Normal and Busulfan-Treated Immature Mice.

Gonadotoxic agents could impair spermatogenesis and may lead to male infertility. The present study aimed to evaluate the effect of IL-1ß on the development of spermatogenesis from cells isolated from seminiferous tubules (STs) of normal and busulfan-treated immature mice in vitro. Cells were cultured in a 3D in vitro culture system for 5 weeks. We examined the development of cells from the different stages of spermatogenesis by immunofluorescence staining or qPCR analyses. Factors of Sertoli and Leydig cells were examined by qPCR analysis. We showed that busulfan (BU) treatment significantly reduced the expression of testicular IL-1ß in the treated mice compared to the control group (CT). Cultures of cells from normal and busulfan-treated immature mice induced the development of pre-meiotic (Vasa), meiotic (Boule), and post-meiotic (acrosin) cells. However, the percentage of developed Boule and acrosin cells was significantly lower in cultures of busulfan-treated mice compared to normal mice. Adding IL-1ß to both cultures significantly increased the percentages of Vasa, Boule, and acrosin cells compared to their controls. However, the percentage of Boule and acrosin cells was significantly lower from cultures of busulfan-treated mice that were treated with IL-1ß compared to cultures treated with IL-1ß from normal mice. Furthermore, addition of IL-1ß to cultures from normal mice significantly increased only the expression of androgen receptor and transferrin but no other factors of Sertoli cells compared to their CT. However, the addition of IL-1ß to cultures from busulfan-treated mice significantly increased only the expression of androgen-binding protein and the FSH receptor compared to their CT. Adding IL-1ß to cultures of normal mice did not affect the expression of 3ßHSD compared to the CT, but it significantly reduced its expression in cultures from busulfan-treated mice compared to the CT. Our findings demonstrate the development of different stages of spermatogenesis in vitro from busulfan-treated mice and that IL-1ß could potentiate this development in vitro.

The male reproductive cycle of the brown basilisk Basiliscus vittatus (Squamata: Corytophanidae) from Tabasco, Southern Mexico.

We used histological and morphometric methods to study the testis and associated glands, including the epididymis, ductus deferens, and renal sexual segment (RSS), of specimens of Basiliscus vittatus sampled from Tabasco, Mexico (17.5926° N, 92.5816° W). Samples were collected throughout 1 year, which included the dry (February to May) and rainy (June to January) seasons. Spermatogenesis in B. vittatus is active throughout the year, but a significant increase in the testicular volume, diameters of seminiferous tubules, height of the germinal epithelium, spermiogenesis, and released spermatozoa occur in the dry season. During the rainy season, all aforementioned parameters decreased except the secretory activity of the epididymis and the RSS, which increased concomitant with an increase of the spermatozoa population within the ductus deferens. These data strongly suggest that B. vittatus reproduce year-round, but males exhibit a peak in spermatogenic activity during the dry season and a peak in insemination and/or copulation at the beginning of the rainy season. We highlight the importance of analyzing not only the testis but also accessory ducts and glands when determining the reproductive cycles of reptiles. The reproductive cycle of B. vittatus is discussed in relation to the environmental conditions of Southern Mexico and is compared to that of other squamates.

Expression pattern of germ cell markers in cryptorchid stallion testes.

Cryptorchidism affects spermatogenesis and testis development, often resulting in stallion subfertility/infertility. This study aims to identify the specific germ cells impacted by cryptorchism in stallions. In a previous study, we found that PGP9.5 and VASA are molecular markers expressed in different germ cells within stallions. Herein, we assessed the heat stress-induced response of spermatogonial stem cells (SSCs) in the seminiferous tubules (ST) of cryptorchid stallion testes (CST) and normal stallion testes (NST). This goal was accomplished by comparing PGP9.5 and VASA expression patterns through reverse transcription quantitative PCR and immunofluorescence assays. We also compared the cross-sectional ST area between groups. Six post-pubertal Thoroughbred unilateral cryptorchid stallions were used. The relative abundance of the mRNA transcripts of PGP9.5 and VASA was significantly upregulated in the NST group than in the CST group. Additionally, the cross-sectional ST area and localization of PGP9.5 and VASA in germ cells were significantly higher in the NST group than in the CST group. Regarding Leydig cells, PGP9.5 staining was observed in both groups. Spermatogonia, primary spermatocytes and secondary spermatocytes were immunostained with VASA in the NST group, while immunostaining was only observed in spermatogonia in the CST group. These results indicate long-term exposure to heat stress conditions, such as cryptorchidism, directly impacts germ cell proliferation and differentiation, leading to impaired spermatogenesis and compromised fertility in stallions.

Identification of spermatogenesis in individual seminiferous tubules and testicular tissue of adult normal and busulfan-treated mice employing Raman spectroscopy and principal component analysis.

The present study aims to identify spermatogenesis in testicular seminiferous tubules (ST) and testicular tissue of adult normal and busulfan-treated mice utilizing PCA and Raman spectroscopy. Raman measurements were conducted on single tubules and testes samples from adult and immature mice, comparing them with those from busulfan-treated adult mice, with validation through histological examination. The analysis revealed a higher signal variability (30 %-40 % at the peaks), prompting scrutiny of individual Raman spectra as a means of spermatogenesis measurement. However, principal component analysis (PCA) demonstrated significant cluster separation between the ST of mature and immature mice. Similar investigations were performed to compare ST from normal mature mice and those from busulfan-treated (BS-treated) mature mice, revealing substantial separation along PC1 and PC2 for all comparison sets. Additionally, comparing testicular samples from mature and immature mice revealed distinct separation in PCA. The study concludes that the combined approach of PCA and Raman spectroscopy proves to be a noninvasive and potentially valuable method for identifying spermatogenesis in seminiferous tubules and testicular samples.

Grafted Sertoli Cells Exert Immunomodulatory Non-Immunosuppressive Effects in Preclinical Models of Infection and Cancer.

The Sertoli cells (SeCs) of the seminiferous tubules secrete a multitude of immunoregulatory and trophic factors to provide immune protection and assist in the orderly development of germ cells. Grafts of naked or encapsulated SeCs have been proved to represent an interesting therapeutic option in a plethora of experimental models of diseases. However, whether SeCs have immunosuppressive or immunomodulatory effects, which is imperative for their clinical translatability, has not been demonstrated. We directly assessed the immunopotential of intraperitoneally grafted microencapsulated porcine SeCs (MC-SeCs) in murine models of fungal infection (Aspergillus fumigatus or Candida albicans) or cancer (Lewis lung carcinoma/LLC or B16 melanoma cells). We found that MC-SeCs (i) provide antifungal resistance with minimum inflammatory pathology through the activation of the tolerogenic aryl hydrocarbon receptor/indoleamine 2,3-dioxygenase pathway; (ii) do not affect tumor growth in vivo; and (iii) reduce the LLC cell metastatic cancer spread associated with restricted Vegfr2 expression in primary tumors. Our results point to the fine immunoregulation of SeCs in the relative absence of overt immunosuppression in both infection and cancer conditions, providing additional support for the potential therapeutic use of SeC grafts in human patients.

Histological differences between the central and peripheral areas of the testes of busulfan-administered mice.

Busulfan is an anticancer drug known to cause serious damage to seminiferous tubules in the testes and deplete germ cells in human and animal models. The testicular artery is anastomosed with deferential and cremasteric arteries and is divided into capsular arteries, which give rise to the centripetal arteries and then recurrent arteries. The arterial blood in the testicular tissue is supplied by such a consequent system of arterial vessels, in order from the peripheral to the central area. As anticancer drugs are generally distributed throughout the whole body via the bloodstream and the running and distribution of arteries differ among the testicular areas, we hypothesized that the efficacy of busulfan differs in different testicular areas, particularly between the central and peripheral areas. In this study, busulfan was intraperitoneally injected at 40 mg/kg body weight into C57BL/6J male mice. After 28 days, in busulfan-treated mice, the diameters of seminiferous tubules were significantly higher in the central than in the peripheral area of the testes. The seminiferous tubular areas also significantly decreased in the peripheral areas compared with the central areas. The number of germ cells per seminiferous tubule was significantly higher in the central than in the peripheral area. Sertoli cell nuclei were detached into the lumen in the peripheral area. The number of Leydig cells was significantly lower in the peripheral areas. These data suggest that the effects of busulfan differ between the central and peripheral areas of the testis at 4 weeks after busulfan administration.

Deep Learning-Based Spermatogenic Staging in Tissue Sections of Cynomolgus Macaque Testes.

The indirect assessment of adverse effects on fertility in cynomolgus monkeys requires that tissue sections of the testis be microscopically evaluated with awareness of the stage of spermatogenesis that a particular cross-section of a seminiferous tubule is in. This difficult and subjective task could very much benefit from automation. Using digital whole slide images (WSIs) from tissue sections of testis, we have developed a deep learning model that can annotate the stage of each tubule with high sensitivity, precision, and accuracy. The model was validated on six WSI using a six-stage spermatogenic classification system. Whole slide images contained an average number of 4938 seminiferous tubule cross-sections. On average, 78% of these tubules were staged with 29% in stage I-IV, 12% in stage V-VI, 4% in stage VII, 19% in stage VIII-IX, 18% in stage X-XI, and 17% in stage XII. The deep learning model supports pathologists in conducting a stage-aware evaluation of the testis. It also allows derivation of a stage-frequency map. The diagnostic value of this stage-frequency map is still unclear, as further data on its variability and relevance need to be generated for testes with spermatogenic disturbances.

Immunolocalization of apelin receptor (APJ) in mouse seminiferous epithelium.

The apelin receptor (APJ) belongs to the member of the G protein-coupled receptor family, and expression of APJ has been reported in the different cell types of testis. The seminiferous tubules in the testis can be identified as different stages (I-XII). It has been also suggested that different factors could be expressed in stage and cell-specific manner in the seminiferous tubules. Recently, we also shown that expression of APJ is developmentally regulated in the testis from PND1 to PND42. Therefore, we analyzed the expression of APJ in the testis of adult mice by immunohistochemistry. Immunohistochemistry showed that the APJ was highly specific for the round and elongated spermatids with stage-dependent changes. The seminiferous tubules at stages I-VII showed APJ immunostaining in the spermatid steps 1-8, not steps of 13-16. The seminiferous tubules at stages IX-XII showed APJ immunostaining in the spermatid steps 9-12. These results suggested the possible role of APJ in the spermiogenesis process. The intratesticular administration of APJ antagonist, ML221 showed a few round spermatids in the seminiferous tubules and some of the tubules with complete absence of round spermatid. Overall, we present evidence that APJ expression in spermatid is dependent on the stages of the seminiferous epithelium cycle and APJ could be involved in the differentiation of round spermatid to elongated spermatid.

Mouse In Vitro Spermatogenesis on 3D Bioprinted Scaffolds.

Testes have a complex architecture that is compartmentalized into seminiferous tubules with a diameter of approximatively 200 µm in which the germ cells differentiate, surrounded by a basement membrane and interstitium. 3D bioprinting might be used to recreate the compartmentalized testicular architecture in vitro. Directed by a software program, pneumatic microextrusion printers can deposit 3D layers of hydrogel-encapsulated interstitial cells in a controlled manner by applying pressure. Once macroporous-shaped scaffolds resembling seminiferous tubules have been bioprinted with interstitial cells, the epithelial cell fraction can be seeded in the macropores to resemble the in vivo testicular architecture. Moreover, macropores can serve as a delimitation for all testicular cells to reorganize and improve the supply of nutrients to cells through the 3D constructs.

Protective role of hesperidin in finasteride-induced testicular toxicity in adult male Wistar rats: Insights into oxidative stress, apoptosis, and ultrastructure of seminiferous tubules.

A negative impact of finasteride on fertility has been reported, in which over production of reactive oxygen species and apoptosis were implicated. Hesperidin, a plant-derived bioflavonoid with antioxidant and anti-apoptotic effects, may mitigate these adverse effects. In order to investigate the possible protective role of hesperidin against finasteride-induced seminiferous tubules toxicity in adult male Wistar rats, 60 rats were randomized into five groups (I-V) receiving distilled water, 0.5% sodium carboxymethylcellulose solution, hesperidin, finasteride, and combined hesperidin and finasteride respectively. Testicular weight, sperm count and motility were determined. Testicular tissue homogenates were prepared to measure the level of malondialdehyde (MDA), total antioxidant capacity (TAC), reduced glutathione (GSH) and the gene expression of caspase-3 and B-cell lymphoma 2 (Bcl2). Testes were processed for light and electron microscopic evaluation. Johnsen score was calculated. Administration of finasteride resulted in significantly decreased testicular weights, sperm count and motility, Johnsen score, tissue levels of TAC and GSH together with significant increase in tissue MDA. Gene expression revealed significantly increased caspase-3 and decreased Bcl2. Furthermore, finasteride disrupted the seminiferous tubules, causing degenerative changes affecting Sertoli cells and spermatogenic cells. Co-administration of hesperidin with finasteride resulted in improvement in testicular weights, TAC, GSH, Bcl2, Johnsen score, sperm count and motility as well as preservation of the structure of the seminiferous tubules. To conclude, hesperidin was found to have a protective potential on finasteride-induced oxidative stress, apoptosis and testicular structural damage.

Three-dimensional morphometry of the testis in dog using design-unbiased stereology.

Testis is considered the main organ of the male reproductive system. Dogs are used as a suitable experimental model of testicular diseases in humans. From the veterinary aspect, several disorders have been reported to affect the testis in dogs. Thus, the objective of the present study was to investigate the morphometrical features of the dog testis using design-based stereology. The testes of six male dogs were used. Isotropic, uniform random sections were obtained and processed for light microscopy. Testicular total volume and the fractional volume of the seminiferous tubules, interstitial tissue and germinal epithelium were measured using the Cavalieri's estimator and the point counting system. Germinal epithelial surface area was estimated using test lines, and total length of seminiferous tubules was analysed using the counting frames. The total volume of testis was calculated 13.64 ± 1.94 cm3 . The relative volume fractions of the seminiferous tubules, interstitial tissue and germinal layer expressed as a percentage of total testicular volume were found to be 75.87 ± 6.11%, 23.68 ± 5.15% and 64.15 ± 4.82%, respectively. The surface area of the germinal layer was 915.25 ± 150.48 cm2 . The thickness of germinal layer was estimated to be 96.18 ± 10.72 µm. The total length of seminiferous tubules measured 290.8 ± 35.86 m. No statistical difference in investigated parameters was found between the left and right testes (p > 0.05). Our data might contribute to the male reproductive knowledge, help develop experimental studies in this field and possibly lead to advancement in the diagnosis and treatment of testicular diseases in the dog.

Polycaprolactone/Testicular Extracellular Matrix/Graphene Oxide-Based Electrospun Tubular Scaffolds for Reproductive Medicine: Biomimetic Architecture of Seminiferous Tubules.

Numerous scaffolds are developed in the field of testicular bioengineering. However, effectively replicating the spatial characteristics of native tissue, poses a challenge in maintaining the requisite cellular arrangement essential for spermatogenesis. In order to mimic the structural properties of seminiferous tubules, the objective is to fabricate a biocompatible tubular scaffold. Following the decellularization process of the testicular tissue, validation of cellular remnants' elimination from the specimens is conducted using 4',6-diamidino-2-phenylindole staining, hematoxylin and eosin staining, and DNA content analysis. The presence of extracellular matrix (ECM) components is confirmed through Alcian blue, Orcein, and Masson's trichrome staining techniques. The electrospinning technique is employed to synthesize the scaffolds using polycaprolactone (PCL), extracted ECM, and varying concentrations of graphene oxide (GO) (0.5%, 1%, and 2%). Subsequently, comprehensive evaluations are performed to assess the properties of the synthetic scaffolds. These evaluations encompass Fourier-transform infrared spectroscopy, scanning electron microscopy imaging, scaffold degradation testing, mechanical behavior analysis, methylthiazolyldiphenyl-tetrazolium bromide assay, and in vivo biocompatibility assessment. The PCL/decellularized extracellular matrix with 0.5% GO formulation exhibits superior fiber morphology and enhanced mechanical properties, and outperforms other groups in terms of in vitro biocompatibility. Consequently, these scaffolds present a viable option for implementation in "in vitro spermatogenesis" procedures, holding promise for future sperm production from spermatogonial cells.

Ultrastructural study: in vitro and in vivo differentiation of mice spermatogonial stem cells.

Mouse testicular tissue is composed of seminiferous tubules and interstitial tissue. Mammalian spermatogenesis is divided into three stages: spermatocytogenesis (mitotic divisions) in which spermatogonial stem cells (SSCs) turn into spermatocytes, followed by two consecutive meiotic divisions in which spermatocytes form spermatids. Spermatids differentiate into spermatozoa during spermiogenesis. Various factors affect the process of spermatogenesis and the organization of cells in the testis. Any disorder in different stages of spermatogenesis will have negative effects on male fertility. The aim of the current study was to compare the in vitro and in vivo spermatogenesis processes before and after transplantation to azoospermic mice using ultrastructural techniques. In this study, mice were irradiated with single doses of 14 Gy 60Co radiation. SSCs isolated from neonatal mice were cultured in vitro for 1 week and were injected into the seminiferous tubule recipient's mice. Testicular cells of neonatal mice were cultured in the four groups on extracellular matrix-based 3D printing scaffolds. The transplanted testes (8 weeks after transplantation) and cultured testicular cells in vitro (after 3 weeks) were then processed for transmission electron microscopy studies. Our study's findings revealed that the morphology and ultrastructure of testicular cells after transplantation and in vitro culture are similar to those of in vivo spermatogenesis, indicating that spermatogenic cell nature is unaltered in vitro.

Efecto de la Vitamina E en Células Peritubulares de Testículos de Ratones Mus musculus Tratados con Ácido Valproico Durante el Estado Fetal y Pubertad / Effect of Vitamin E on Peritubular Cells from Testis of Mus musculus Mice Treated with Valproic Acid During the Fetal Stage and Puberty

El ácido valproico (VPA) es un fármaco antiepiléptico teratógenico que, al ser administrado durante etapas tempranas del embarazo, puede producir alteraciones en el desarrollo embriofetal, las que se manifiestan tanto a nivel del sistema nervioso como del testículo. No obstante, se ha reportado que la administración de vitamina E (VE) podría revertir dichas alteraciones. El objetivo del presente estudio fue determinar el efecto protector de la VE a nivel testicular en fetos y ratones púberes expuestos a VPA durante la fase embrionaria de su desarrollo. Se utilizó un total de 30 ratones hembra adultas gestantes (Mus musculus) cepa BALB/c, las cuales se dividieron en 6 grupos. El estudio contempló el análisis de fetos machos a los 17,5 días post-coital (dpc) y machos juveniles a las 6 semanas post-natal. A los grupos 1 y 4 se les administró 0,3 mL de solución fisiológica (grupos control para 17,5 dpc y 6 semanas postnatal, respectivamente). A los grupos 2 y 5 se les suministró la cantidad de 600 mg/kg de VPA (grupos VPA), en tanto que a los grupos 3 y 6 se les aplicó la misma dosis de VPA complementada con 200 UI de VE (grupos VPA+VE). Se describió la histología normal y patológica del compartimento peritubular del testículo. En los grupos VPA se evidenció una degeneración de la pared peritubular, y atrofia de túbulos seminíferos, así como exfoliación de las células germinales. Por el contrario, en los grupos VPA+VE tales signos no fueron observados y la morfología presentó aspecto normal solo con algunas alteraciones focales. Estos resultados corroboran el hecho que la administración de VE contrarresta en parte, los efectos deletéreos que ocasiona el VPA.

SUMMARY: Valproic acid (VPA) is a teratogenic antiepileptic drug that, when administered during the early stages of pregnancy, can produce alterations in embryo-fetal development, which manifest both at the level of the nervous system and the testicle. However, it has been reported that the administration of vitamin E (VE) could reverse these alterations. The study aimed to determine the protective effect of VE at the testicular level in fetuses and pubertal mice exposed to VPA during the embryonic phase of their development. 30 pregnant adult female mice (Mus musculus) BALB/c strain were used, which were divided into 6 groups. The study included the analysis of male fetuses at 17.5 days post-coital (dpc) and juvenile males at 6 weeks post-natal. Groups 1 and 4 were administered 0.3 mL of physiological solution. Groups 2 and 5 were given 600 mg/kg of VPA (VPA groups), while groups 3 and 6 were given the same dose of VPA supplemented with 200 IU of VE (VPA+VE). The normal and pathological histology of the peritubular compartment of the testis was described. In the VPA groups, degeneration of the peritubular wall, and atrophy of the seminiferous tubules, as well as exfoliation of the germ cells, were evident. On the contrary, in the VPA+VE groups such signs were not observed and the morphology presented a normal appearance with only some focal alterations. These results corroborate the fact that the administration of VE partially counteracts the deleterious effects caused by VPA.

An Improved Method for Purification of the Residual Bodies from the Seminiferous Tubules of Mice.

Human fertility is declining in Western countries, and it is becoming increasingly clear that male infertility plays a pivotal role in the overall fertility decline. To understand the process that drives successful male germ cell maturation, the study of spermatogenesis of model organisms, such as mice, is essential. Residual bodies (RBs) play an important role in the last stages of spermatogenesis. They are formed at the time when post-meiotic spermatids undergo sequential differentiation steps so that the acrosome and flagellum are developed, the nucleus is markedly condensed, and the cytoplasm is lost. The masses of lost cytoplasm become RBs. Our recent work has shown that RB dynamics are highly sensitive to even small fertility defects. It was also noted that the transcriptome and proteome of RBs changes in response to spermatogenic defects. Thus, RBs represent an excellent and highly sensitive entity for studying male fertility. Previously published protocols for RB purification had some major limitations: they produced an RB fraction that was heavily contaminated with spermatozoa and erythrocytes or required tens of grams of starting material. In addition, most of the available protocols were developed for purification of RBs from rat testes. Here, we present a protocol that allows the isolation of 2.5-3 × 106 RBs from mouse testes with a purity of 98% from only 1 g of starting material. The purified material can be used for various downstream applications to study male fertility, such as transcriptome and proteome analyses, super-resolution microscopy, and electron and cryo-electron microscopy, amongst many others. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: An improved method for purification of the residual bodies from the seminiferous tubules of mice.

Tissue engineering studies in male infertility disorder.

Infertility is an important issue among couples worldwide which is caused by a variety of complex diseases. Male infertility is a problem in 7% of all men. In vitro spermatogenesis (IVS) is the experimental approach that has been developed for mimicking seminiferous tubules-like functional structures in vitro. Currently, various researchers are interested in finding and developing a microenvironmental condition or a bioartificial testis applied for fertility restoration via gamete production in vitro. The tissue engineering (TE) has developed new approaches to treat male fertility preservation through development of functional male germ cells. This makes TE a possible future strategy for restoration of male fertility. Although 3D culture systems supply the perception of the effect of cellular interactions in the process of spermatogenesis, formation of a native gradient of autocrine/paracrine factors in 3D culture systems have not been considered. These results collectively suggest that maintaining the microenvironment of testicular cells even in the form of a 3D-culture system is crucial in achieving spermatogenesis ex vivo. It is also possible to engineer the testicular structures using biomaterials to provide a supporting scaffold for somatic and stem cells. The insemination of these cells with GFs is possible for temporally and spatially adjusted release to mimic the microenvironment of the in situ seminiferous epithelium. This review focuses on recent studies and advances in the application of TE strategies to cell-tissue culture on synthetic or natural scaffolds supplemented with growth factors.

Seasonal variations of testis anatomy and of G-coupled oestrogen receptor 1 expression in Gerbillus gerbillus.

The gerbil, Gerbillus gerbillus, a nocturnal desert rodent of northern Africa, exhibits a seasonal reproductive cycle with marked anatomical and behavioural shifts between breeding season and resting season. The aim of this study is to investigate key elements involved in these seasonal changes, specifically in males: the histology of the testis as well as the expression of the G-protein-coupled oestrogen receptor 1 (GPER1) in the testis. During the breeding season, the seminiferous tubules were full of spermatozoa, and their epithelium contained germinal cells embedded in Sertoli cells. Amidst tubules, well-developed Leydig cells were observed around blood vessels, with peritubular myoid cells providing structural and dynamic support to the tubules. GPER1 was largely expressed throughout the testis. Notably, Leydig cells, spermatogonia and spermatocytes showed strong immunohistochemical signals. Sertoli cells, spermatozoa and peritubular myoid cells were moderately stained. During the resting season, spermatogenesis was blocked at the spermatocyte stage, spermatids and spermatozoa were absent and the interstitial space was reduced. The weight of the testis decreased significantly. At this stage, GPER1 was found in Leydig cells, spermatocytes and peritubular myoid cells. Sertoli cells and spermatogonia were not marked. Overall, the testis of the gerbil, Gerbillus gerbillus, has undergone noticeable histological, cellular and weight changes between seasons. In addition, the seasonal expression pattern of GPER1, with pronounced differences between resting season and breeding season, indicates that this receptor is involved in the regulation of the reproductive cycle.

Isolation of an Insulin-Like Receptor Involved in the Testicular Development of the Mud Crab Scylla paramamosain.

Insulin-like androgenic gland hormone (IAG) is a key regulator of male sexual differentiation in crustaceans that plays important roles in secondary sexual characteristics and testicular development. As a hormone, IAG interacts with its membrane receptor to initiate downstream signal pathways to exert its biological functions. In this study, we isolated a full-length cDNA of an insulin-like receptor (Sp-IR) from the mud crab Scylla paramamosain. Sequence analysis revealed that this receptor consists of a Fu domain, two L domains, three FN-III domains, a transmembrane domain, and a tyrosine kinase domain, classifying it as a member of the tyrosine kinase insulin-like receptors family. Our results also suggested that Sp-IR was highly expressed in the testis and AG in males. Its expression in the testis peaked in stage I but significantly decreased in stages II and III (p < 0.01). Next, both short- and long-term RNA interference (RNAi) experiments were performed on males in stage I to explore Sp-IR function in mud crabs. The results showed that Sp-vasa and Sp-Dsx expression levels in the testis were significantly down-regulated after the specific knockdown of Sp-IR by RNAi. Additionally, the long-term knockdown of Sp-IR led to a considerable decrease in the volume of seminiferous tubules, accompanied by large vacuoles and a reduced production of secondary spermatocytes and spermatids. In conclusion, our results indicated that Sp-IR is involved in testicular development and plays a crucial role in transitioning from primary to secondary spermatocytes. This study provided a molecular basis for the subsequent analysis of the mechanism on male sexual differentiation in Brachyuran crabs.

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.