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Ethanol consumption is increasingly prevalent in communities and has several side effects for humans. Chronic alcohol consumption is often associated with decreased libido and infertility. This study aimed to evaluate the impact of beer on spermatogenesis, proteins, and gene expression of P21 and cyclin D1 in testicular tissue. Twenty-four male mice were assigned to 4 groups. The control group received normal saline, and the experimental groups received alcoholic beer (3 g/kg BW as 20% v/v). After 7, 15, and 35 days, mice were sacrificed, and a part of testicular tissues was stored at −70 °C to measure the protein and gene expression of P21 and cyclin D1 by Western blot and real-time PCR. Moreover, this assay measured the levels of malondialdehyde (MDA), superoxide dismutase (SOD), and catalase (CAT). Histomorphometry and histopathology were done. The consumption of alcoholic beer led to increased protein and gene expression of P21 and decreased expression of cyclin D1, as well as reduced levels of SOD and CAT, increased levels of MDA, and subsequent tissue damage in the testicular tissue in a time-dependent manner. Also, the diameter of seminiferous tubules and the thickness of the germinal epithelium were reduced in a time-dependent manner, and the percentage of seminiferous tubules with tubular differentiation, replacement, and negative spermiogenesis coefficients increased significantly. Alcoholic beer disrupted the process of cell division in the testes by reducing the expression of cyclin D1, increasing p21 expression, and inducing oxidative stress, which in turn reduced sperm production and quality. • Alcoholic beer consumption increased protein and gene expression of P21. • Alcoholic beer consumption decreased protein and gene expression of cyclin D1. • Alcoholic beer consumption increased levels of MDA and subsequent tissue damage in the testicular tissue. [ABSTRACT FROM AUTHOR]

Diabetes mellitus (DM) is a widespread metabolic disease presenting with various complications, including spermatogenic dysfunction. However, the underlying mechanisms are still unclear. Ferroptosis, a novel type of programmed cell death, is associated with much metabolic diseases. Here, we investigated the role of ferroptosis in spermatogenic dysfunction of streptozotocin (STZ)-induced type 1 diabetic mice (diabetic mice), high glucose (HG)-treated GC-2 cells (HG cells) as well as testicular tissues of diabetic patients. We found an accumulation of iron, elevated malondialdehyde level and reduced glutathione level in the testis tissues of diabetic mice and HG cells. Histological examination showed a decrease in spermatogenic cells and spermatids within the seminiferous tubules as well as mitochondrial shrinkage in the testis tissues of diabetic mice. Ferrostatin-1 (Fer-1), the inhibitor of ferroptosis, mitigated ferroptosis-associated iron overload, lipid peroxidation accumulation and spermatogenic dysfunction of diabetic mice. Furthermore, we observed a downregulation of GPX4, FTL and SLC7A11 in diabetic mice and HG cells. Fer-1 treatment and GPX4 overexpression counteracted the effects of HG on cell viability, reactive oxygen species, lipid peroxidation and glutathione via inhibition of ferroptosis. Moreover, we found an elevation of ferroptosis in testicular tissues of diabetic patients. Taken together, our results identify the crucial role of ferroptosis in diabetic spermatogenic dysfunction and ferroptosis may be a promising therapeutic target to improve spermatogenesis in diabetic patients. [Display omitted] • Increased ferroptosis caused spermatogenic dysfunction in type 1 diabetic mice and patients. • Ferrostatin-1 treatment alleviated spermatogenic dysfunction in diabetic mice. • Ferrostatin-1 treatment and GPX4 overexpression attenuated ferroptosis in high glucose treated GC-2 cells. • Inhibition of ferroptosis may be a potential therapy to alleviate diabetic male infertility. [ABSTRACT FROM AUTHOR]

• Chlorophytum borivilianum Santapau & R.R.Fern. (C. borivillianum) used for the treatment of male sexual dysfunction and oligospermia. • Ethanolic extract and saponin enriched n-butanol fraction was evaluated to evolve the underlying biochemical mechanism for spermatogenic potential of C. borivilianum. • Ethanolic extract and saponin enriched n-butanol fraction evaluated to alter fold shift in the mRNA expression of the StAR and 3β-HSD enzymes. • Saponin enriched n-butanol fraction might have interacted with CREB and additionally stimulated LH receptors to amplify the expression of 3β-HSD and StAR mRNA. Traditional applications of the roots of Chlorophytum borivilianum Santapau & R.R.Fern. include spermatogenic potential. Using rats as test subjects, the potential of alcoholic extract (AE-SM) and n-Butanol fraction (NB-SM), which is rich in saponin, to alter the levels of testicular testosterone and cholesterol as well as mRNA expression of the 3β-Hydroxy-∆5- steroid dehydrogenase (3β-HSD) and Acute Steroid Regulatory Protein (StAR) were determined. Histological analyses of testicular tissues were performed, and the quantity of sperm in the seminal fluid was measured. The testosterone concentration increased in AE-SM and NB-SM in a dose-dependent and statistically significant manner. In-vivo mRNA expression of the StAR and 3β-HSD enzymes was also enhanced by the test samples. When compared to control animals, AE-SM and NB-SM increased testicular testosterone by 11.46 and 12.98 fold, respectively, while the extract and fractions revealed a 5.72 and 7.70 fold alteration in StAR mRNA expression. In testicular tissues, AE-SM and NB-SM increased 3β-HSD activity by 4.98 and 9.05 fold, respectively, while changing the expression of the 3β-HSD mRNA by 6.46 and 15.54 factor, respectively. The fluid sampled from the caudal region of the AE-SM and NB-SM indicated an increased sperm concentration, and histological examinations revealed dense Sertoli cell cytoplasm and an increase in the number of sperms in the luminal part of the seminiferous tubules. The components of AE-SM and NB-SM might stimulated testosterone biosynthesis in-vivo by modifying mRNA expression of selected steroidogenic enzymes. By modifying the mRNA expression of the StAR and 3β-HSD, AE-SM and NB-SM can raise the level of testosterone in testicular tissues. As evidenced by an elevated sperm concentration discovered within the seminiferous fluid and histological studies, overexpression of steroidogenic enzymes increased testicular testosterone levels and may have stimulated the spermatogenesis process.Saponin enriched n-butanol fraction might have interacted with CREB and additionally stimulated LH receptors to elevate the StAR and 3β-HSD mRNA expression. [ABSTRACT FROM AUTHOR]

The branched-chain amino acids (BCAAs: leucine, isoleucine and valine) are essential for animal growth and metabolic health. However, the effect of valine on male reproduction and its underlying molecular mechanism remain largely unknown. Here, we showed that l -valine supplementation (0.30% or 0.45%, water drinking for 3 weeks) did not change body and testis weights, but significantly altered morphology of sertoli cells and germ cells within seminiferous tubule, and enlarged the space between seminiferous tubules within mouse testis. l -valine treatment (0.45%) increased significantly the Caspase3/9 mRNA levels and CASPASE9 protein levels, therefore induced apoptosis of mouse testis. Moreover, gene expression levels related to autophagy (Atg5 and Lamb3), DNA 5 mC methylation (Dnmt1, Dnmt3a, Tet2 and Tet3), RNA m6A methylation (Mettl14, Alkbh5 and Fto), and m6A methylation binding proteins (Ythdf1/2/3 and Igf2bp1/2) were significantly reduced. Protein abundances of ALKBH5, FTO and YTHDF3 were also significantly reduced, but not for ATG5 and TET2. Testis transcriptome sequencing detected 537 differentially expressed genes (DEGs, 26 up-regulated and 511 down-regulated), involved in multiple important signaling pathways. RT-qPCR validated 8 of 9 DEGs (Cd36, Scd1, Insl3, Anxa5, Lcn2, Hsd17b3, Cyp11a1, Cyp17a1 and Agt) to be decreased significantly, consistent with RNA-seq results. Taken together, l -valine treatment could disturb multiple signaling pathways (autophagy and RNA methylation etc.), and induce apoptosis to destroy the tissue structure of mouse testis. • l -valine intake changed cell morphology and tissue structure of mouse testis in a dose-dependent manner. • l -valine intake induced apoptosis via the Caspase pathway. • l -valine affected autophagy and nucleic acid methylation pathways. • RNA-seq identified 537 DEGs altered by l -valine, most of which down-regulated and involved in multiple signaling pathways. [ABSTRACT FROM AUTHOR]

5-Fluorouracil (5-FU) is one of the most broadly chemotherapeutic compounds employed in treating solid tumors. Quercetin (QU) as a flavonoid has diverse positive clinical applications including antioxidative, antiproliferative, and anti-inflammatory. The current study aims to assess the histopathological changes and anti-oxidative effects of QU loaded with chitosan nanoparticles (QU-NPs) on the testicular injury triggered by 5-FU in mice. Twenty male albino mice were randomly divided into 4 groups as follows: the control, 5-FU, 5-FU + QU (5 mg/kg), and 5-FU + QU-NPs (5 mg/kg) for 14 days. Oxidative marker [malondialdehyde (MDA)] and antioxidant markers [superoxide dismutase (SOD) and catalase (CAT)] were examined in serum. Testicular damage stemmed from a single dose injection of 5-FU intraperitoneally (i.p.). The control group received normal saline and the treatment groups received QU with i.p. injection for 14 days. Serum MDA level was lower in the QU and QU-NPs treated groups than in the 5-FU group. Moreover, serum CAT levels were remarkably increased in both the QU- and QU-NPs treated groups compared to the 5-FU group. In the case of SOD, the most considerable difference was found between the 5-FU group and the QU-NPs, but not the QU group. Further, both QU and QU-NPs, particularly the QU-NPs, ameliorated 5-FU-induced testicular tissue damage, as evidenced by a decrease in hyperemia, edema, and vacuolation, and an improvement in the area of seminiferous tubules, and spermatocyte and seminiferous tubule counts. Nanotechnology and fabrication of QU-NPs can ameliorate the testicular damage caused by 5-FU. [Display omitted] • 5-fluorouracil exposure could induce spermatogenesis disorder in mice. • 5-fluorouracil caused testicular and epididymal histopathological lesions in mice. • 5-fluorouracil decreased serum testosterone levels in mice. • Quercetin improves the oxidative damage induced by 5-FU. [ABSTRACT FROM AUTHOR]

Eif2s3y (eukaryotic translation initiation factor 2, subunit 3, structural gene Y-linked, Eif2s3y) is an essential gene for spermatogenesis. Early studies have shown that Eif2s3y can promote the proliferation of spermatogonial stem cells (SSCs) and can replace the Y chromosome together with sex-determining region Y (Sry) to transform SSCs into round spermatozoa. We injected lentiviral particles into the seminiferous tubules of mouse testes by sterile surgery surgically to establish overexpressing Eif2s3y testes. And then the mice were intraperitoneally injected with LPS to established the model of testis inflammation. Through RNA sequencing, qRT–PCR analysis, Western blot, co-culture etc., we found that Eif2s3y alleviated LPS-induced damage in mouse testes and maintained spermatogenesis. In testes with Eif2s3y overexpression, the seminiferous tubules were more regularly organized after exposure to LPS compared with the control. Eif2s3y performs its function by negatively regulating Adamts5 (a disintegrin and metalloproteinase containing a thrombospondin-1 motif), an extracellular matrix-degrading enzyme. ADAMTS5 shows a disruptive effect when the testis is exposed to LPS. Overexpression of Eif2s3y inhibited the TLR4/NFκB signaling pathway in the testis in response to LPS. Generally, our research shows that Eif2s3y protects the testis from LPS and maintains spermatogenesis by negatively regulating A damts5. • Overexpression of Eif2s3y alleviates testicular injury induced by LPS. • Eif2s3y is a negative regulator of Adamts5. • Eif2s3y alleviates LPS-induced germ cell damage by negatively regulating Adamts5. • Eif2s3y maintains testicular immune homeostasis through negative regulation of Adamts5. [ABSTRACT FROM AUTHOR]

Lysine-specific histone demethylase 2 (Kdm2a) is a regulatory factor of histone modifications that participates in gametogenesis and embryonic development. The mis-regulation of Kdm2a can lead to aberrant gene expression, thereby contributing to abnormal cell proliferation, differentiation, apoptosis, and tumorigenesis. However, due to the potential confounding effects that are secondary to the loss of Kdm2a function from the soma in existing whole-animal mutants, the in vivo function of Kdm2a in spermatogenesis for male fertility remains unknown. Herein, we focus on exploring the spatiotemporal expression profile and biological functions of Kdm2a in the spermatogenesis and fertility of male mice. A testis-specific knockout Kdm2a model (Kdm2a cKO) was established by using the Stra8 -Cre/loxP recombinase system to explore the roles of Kdm2a in male fertility. Our results showed that Kdm2a was ubiquitously expressed and dynamically distributed in multiple tissues and cell types in the testis of mice. Surprisingly, Kdm2a -deficient adult males were completely fertile and comparable with their control (Kdm2a flox/flox ) counterparts. Despite the significantly reduced total number of sperm and density of seminiferous tubules in Kdm2a cKO testis accompanied by the degeneration of spermatogenesis, the fertilization ability and embryonic developmental competence of the Kdm2a cKO were comparable with those of their control littermates, suggesting that Kdm2a disruption did not markedly affect male fertility, at least during younger ages. Furthermore, Kdm2a homozygous mutants exhibited a lower total number and motility of sperm than the control group and showed notably affected serum 17β-estradiol concentration. Interestingly, the transcriptome sequencing revealed that the loss of Kdm2a remarkably upregulated the expression level of Kdm2b. This effect, in turn, may induce compensative effects in the case of Kdm2a deficiency to maintain normal male reproduction. Together, our results reveal that Kdm2a shows spatiotemporal expression during testicular development and that its loss is insufficient to compromise the production of spermatozoa completely. The homologous Kdm2b gene might compensate for the loss of Kdm2a. Our work provides a novel Kdm2a cKO mouse allowing for the efficient deletion of Kdm2a in a testis-specific manner, and further investigated the biological function of Kdm2a and the compensatory effects of Kdm2b. Our study will advance our understanding of underlying mechanisms in spermatogenesis and male fertility. • The testis-specific knockout Kdm2a model was constructed through Kdm2a floxed mice bred with Stra8 -Cre transgenic mice. • The loss of Kdm2a is insufficient to completely compromise the production of spermatozoa. • Knockout Kdm2a gene led to a decrease the sperm concentration and motility, accompanied by degeneration of seminiferous tubules. • Deletion of Kdm2a changes gene expression profiles and significantly upregulates the mRNA expression of Kdm2b. [ABSTRACT FROM AUTHOR]

Ultrasound elastography was proposed for the evaluation of testicular focal lesions, but no studies verified the agreement between the whole histological architecture of the testis and the stiffness measured by elastography. The present study explored the use of strain elastography in the evaluation of testis with normal or abnormal spermatogenesis, classified based on epididymal sperm attributes, and the consistency between elastographic parameters and the testicular histological feature. Strain elastography was performed during the routine andrological examination in 22 dogs presented for elective orchiectomy. Epididymal sperm attributes and testicular histology were analyzed after orchiectomy. Based on the epididymal sperm characteristics, testes were classified according to normal or abnormal spermatogenesis, and strain elastographic attributes were compared between groups. Possible correlations between strain elastography and histological features were also explored. Consistent with the literature in humans, testes with abnormal spermatogenesis were stiffer (mean strain elastographic index 3.6 ± 0.6) compared with normal testes (mean strain elastographic index 1.9 ± 0.2; P < 0.01). The strain elastographic index was negatively correlated with the area occupied by seminiferous tubules (Pearson's rho = −0.716; P = 0.0003), the mean diameter (Pearson's rho = -0.742; P = 0.0002), and thickness of the seminiferous tubule (Pearson's rho = −0.728; P = 0.0002). Surprisingly, no correlations were found between the area occupied by connective tissue in histological sections and elastographic attributes, suggesting that the increased stiffness was not related to the increased amount of connective tissue. This study demonstrated that strain elastography could be used to support the andrological examination, but measurements should be acquired in specific regions to be reliable. • Strain elastography estimates tissue stiffness semi-quantitatively. • Abnormal testes were stiffer when compared with normal testis. • Strain elastography index was negatively correlated with seminiferous tubule area, diameter, and thickness. • No correlations were found between stiffness and the amount of connective tissue. • Strain elastography index was correlated with sperm primary abnormalities. [ABSTRACT FROM AUTHOR]

The goals of this study were to determine whether contrast-enhanced ultrasound (CEUS) imaging could be used for assessment of chronic alcohol-induced testicular damage (CAITD) and to explore the relationships between the laboratory and pathological findings of CAITD and the quantitative parameters of CEUS. Thirty-six rabbits were randomly divided into a chronic ethanol exposure (CEE) group and negative control (NC) group, which were further randomly divided into six groups with equal numbers of rabbits by period of exposure (30 d, 60 d, 90 d). All rabbits underwent conventional US and CEUS imaging at the end of the induction period. Blood and histological specimens were collected for laboratory and pathological examination. The peak intensity (PI) and area under the curve (AUC) for the CEUS parameters decreased as CAITD progressed (p < 0.05). Both PI and AUC were positively correlated with the Johnsen score (r = 0.945 and 0.898, respectively, all p values <0.001) and the mean epithelium thickness of the seminiferous tubule (METST) (r = 0.927 and 0.881, respectively, all p values <0.001) of the testis, and negatively correlated with the serum levels of endothelin-1 (ET-1) (r = –0.940 and –0.899, respectively, all p values <0.001) and nitric oxide (NO) (r = –0.894 and –0.954, respectively, all p values <0.001), as well as the testicular tissue content of malondialdehyde (MDA) (r = –0.894 and –0.945, respectively, all p values <0.001). CEUS imaging can be used for monitoring organ perfusion of the testis to quantify the development of CAITD. [ABSTRACT FROM AUTHOR]

Spermatogenesis is a highly complicated biological process that occurs in the epithelium of the seminiferous tubules. It is regulated by a complex network of endocrine and paracrine factors. Sertoli cells (SCs) play a key role in spermatogenesis due to their production of trophic, differentiation, and immune-modulating factors. However, many of the molecular pathways of SC action remain controversial and unclear. Recently, many studies have focused on exosomes as an important mechanism of intercellular communication. We found that the exosomes derived from mouse SCs inhibited the apoptosis of primary spermatogonia. A total of 1016 miRNAs in SCs and 556 miRNAs in exosomes were detected using miRNA high-throughput sequencing. A total of 294 miRNAs were differentially expressed between SCs and exosomes. Furthermore, 19 tsRNA families appeared in SCs, while 6 tsRNA families appeared in exosomes. A total of 57 and 1 miRNAs (RPM >4) and 14 and 1 tsRNAs were exclusively expressed in SCs and exosomes, respectively. MiR-10b is one of the top ten exosomes with a relatively large enrichment of miRNA. Overexpression of miR-10b downregulates the expression of the target KLF4 to reduce spermatogonial apoptosis in primary spermatogonia or the C18-4 cell line. • Mouse SCs delivered exosomes protected the spermatogonia by decreasing cell apoptosis. • Total of 1016 miRNAs in SCs and 556 miRNAs in exosomes were detected using miRNA high-throughput sequencing. • 19 tsRNAs families were appeared in SCs, and 6 tsRNAs families contained in exosomes. • GO and KEGG analyses showed that SC-exosomes regulated the apoptosis of spermatogonia by PI3K/AKT and MAPK signal pathways. • MiR-10b down-regulated expression of the target KLF4 to reduce spermatogonial apoptosis. [ABSTRACT FROM AUTHOR]

Simulated microgravity during hindlimb unloaded (HU) induces multi-organ pathologies in mice, including testicular dysfunction. However, a detailed characterization of testicular histology and its driving molecular mechanisms remains elusive. We investigated the potential contribution of elevated endoplasmic reticulum (ER) stress to testicular atrophy in HU mice. We divided male c57BL/6j mice into a control group (C) or unloaded mice without treatment (U), or treated with 4PBA (UP), an ER stress inhibitor, for three weeks. At the end of the experiment, mice were euthanized, and testes tissues were dissected for histopathology and mRNA sequencing. HU was associated with significant testicular atrophy (p = 0.03) and several histopathological alterations, including a reduction in the diameter of seminiferous tubules (p < 0.001), epithelial thinning (p < 0.001), reduced sperm density (p < 0.001), and thickening of the epididymal epithelium (p < 0.001). mRNA sequencing revealed alterations in several pathways associated with oxidative stress, protein catabolism, and inflammation induction (all p < 0.05). Three weeks of treatment with 4PBA prevented testicular atrophy (p = 0.214), partly restored testicular microarchitecture, and reversed changes in the transcriptomic profiling of HU mice. Our novel findings indicate a mechanistic role of elevated ER stress in causing testicular pathology during HU conditions. Based on findings, we report a therapeutic potential of 4PBA in reversing testicular pathology in conditions mimicking prolonged bed rest and spaceflight. [Display omitted] • Hindlimb unloaded (HU) mice exhibit testicular disruption in histology. • Testes of HU mice also show elevated endoplasmic reticulum (ER) stress. • Pharmacological inhibition of ER stress reverses testicular histology during HU. • Signature transcriptomic changes accompany ER stress-induce testicular disruption. • Inhibiting ER stress may be a molecular target to restore testes during HU. [ABSTRACT FROM AUTHOR]

This study was aimed to investigate the effects of dietary curcumin supplementation on the hydrogen peroxide (H 2 O 2)-induced testicular oxidative damage in breeder roosters. Thirty-two 20-week roosters were randomly divided into four groups: (1) basal diet (CON); (2) basal diet with H 2 O 2 challenge (H 2 O 2); (3) basal diet with 200 mg/kg curcumin (CUR); (4) basal diet with 200 mg/kg curcumin and H 2 O 2 challenge (CUR + H 2 O 2). The trial lasted for 8 weeks, H 2 O 2 challenged groups got an intraperitoneal injection of H 2 O 2 at the 50 and 53 days, while the CON and CUR groups received an injection of saline. The results showed that dietary curcumin supplementation significantly decreased abnormal sperm rates in the semen, notably improved seminiferous tubules, increased testis scores, and serum testosterone levels. Curcumin supplementation could also ameliorate the redox damage caused by H 2 O 2 , by enhancing the capacities of antioxidant enzymes (CAT, GSH-Px, SOD, and T-AOC), and reducing MDA levels. In addition, curcumin normalized the H 2 O 2 -induced negative effects, which included downregulations in spermatogenesis-related genes (STAR , HSD3-β1 , SYCP3, AKT1) and antioxidant genes (HMOX-1 , NQO-1), reduced protein expressions of Nrf2, PCNA, and Bcl-2, and increased protein expressions of Caspase 3 and Bax. Moreover, H 2 O 2 -induced decreased mRNA expressions of EIF2AK3, Caspase3, and BCL-2 were all reversed by dietary curcumin supplementation. In summary, dietary curcumin supplementation could relieve H 2 O 2 -induced oxidative damage and reproduction decline through the Nrf2 signaling pathway and anti-apoptotic effects in roosters. • Intraperitoneal injection of hydrogen peroxide can build a model of oxidative damage of testes in roosters. • Curcumin can improve the reproduction of roosters by antioxidant and anti-apoptotic effects under oxidative damage conditions. • Curcumin can be utilized as a feed additive to prevent oxidative damage and enhance the reproductive performance of roosters. [ABSTRACT FROM AUTHOR]

Turner syndrome (TS) is caused by the absence of part of or a whole X-chromosome and occurs in 1 in 2500 live female births. The recommendation to perform prophylactic gonadectomy at diagnosis has been called into question based on reports of spontaneous puberty and pregnancy in some individuals with TS and Y. Because the decision to forego early prophylactic gonadectomy to preserve fertility must be balanced with avoidance of gonadal malignancy, our study sought to investigate gonadal histology from individuals who underwent prophylactic gonadectomy due to a diagnosis of TS with Y material. Ovarian tissue was obtained during prophylactic bilateral gonadectomy of patients with TS and Y at a pediatric tertiary care center. Histological data (i.e., follicle number, morphology) was collected from H&E slides and immunohistochemically stained slides of patients. Chart review was used to identify associations between histological and clinical data and evaluate presence of germ cell neoplasia in situ (GCNIS), gonadal malignancy, and seminiferous tubules. This study was approved by the Institutional Review Board. Twelve TS with Y patients were included in the analysis, all of which had undergone bilateral gonadectomy with salpingectomy. The demographics, clinical outcomes, and gonadal histological characteristics of the patients are included in Table 1. One patient had menarche prior to gonadectomy. None of the patients had follicles or seminiferous tubules. GCNIS was present in 5 patients, and these were not identified in the pathology reports. There was no evidence of malignancy in the pathology reports or the histological examination of the patients. Our study revealed the absence of germ cells, indicating a low likelihood of fertility in individuals with Turner syndrome and Y-chromosome material. Furthermore, the presence of GCNIS, a precursor to cancer, suggests a non-negligible risk. Our findings revealed no instances of malignancy, which suggests that prophylactic gonadectomy need not be immediately administered upon diagnosis, although delaying the procedure appears to offer limited, if any, fertility benefits. While isolated case reports have suggested the potential for fertility in some instances, such occurrences remain rare. Our limited sample size emphasizes the need for further research to clarify the optimal timing and criteria for prophylactic gonadectomy in these cases. [ABSTRACT FROM AUTHOR]

Citrate-coated silver nanoparticles were synthesized in one step method using tri-sodium citrate and silver nitrate at pH 6.1. After synthesis, the resulting silver nano-suspension was characterized using UV–visible spectroscopy, dynamic light scattering, high resolution-scanning electron microscopy, energy dispersive x-ray spectroscopy and fourier transform infrared-spectroscopy. The particles were intraperitoneally injected into Swiss albino male mice for a period of one complete spermatogenic cycle. The LD 50 was determined following the procedure of Dixon's Up-and-Down method. The intra-testicular level of silver was measured using the technique of inductively coupled plasma-mass spectrometry. The intra-testicular location of accumulated nanoparticles was observed using auto-metallography. The cytology and volume of Leydig cells were assessed and analysed. Following the exposure to silver nanoparticles, it was found that accumulation of nanoparticles inside the seminiferous tubules is a dose-dependent process. The deposition of silver agglomerate induced morphometric changes in the lumen of seminiferous tubules and Leydig cells. The exposure also caused significant changes at the level of structural integrity of sperm chromatin material and variable damages to sperm DNA. • Citrate coated silver nanomaterial intraperitoneally injected into the body of Swiss albino mice for the duration of one complete spermatogenic cycle • Intra-testicular silver level was measured by the technique of ICP-MS • The seminiferous tubules were examined to locate the accumulation of nanoparticles and Leydig cells were morphometrically analysed • Loss of sperm nuclear integrity leading to cell death • Citrate capped silver nanoparticle can cause significant adverse changes in seminiferous tubules and leydig cell architecture and structural integrity of sperm nuclear material leading to cell death. [ABSTRACT FROM AUTHOR]

The busulfan, an alkylating agent, suppresses endogenous spermatogenesis in recipient testes. However, considering a wide variation in the effects of busulfan among animal species, its dosage and route of infusion need optimization to prepare effective and safe recipients. Thus, the current study aimed to create a suitable recipient goat model for germ cell (Gc) transplantation through a single intra-testicular (i.t.) busulfan infusion under ultrasonographic (USG) guidance. As observed through the infusion of trypan blue under USG guidance into mediastinum testis (MT) of pre-pubertal Barbari bucks, 3–5 mL of trypan blue solution could fill almost 80% of seminiferous tubules. Thereafter, in Experiment-1, the effect of different busulfan doses (mg/kg) i.e. 0 [negative control, Group (Gr) 1; 0 mg/kg-MT], 1 (Gr 2; 1 mg/kg-MT), 2 (Gr 3; 2 mg/kg-MT), and 3 (Gr 4; 3 mg/kg-MT) were studied. Further, in Experiment-2, sterilizing effects of busulfan infusion through two different routes [MT or cavum vaginale (CV)] were compared. Following i.t. busulfan treatment, no adverse physiological effects or body weight loss were detected. The histological analyses demonstrate a dose-dependent depletion of Gc with almost complete loss of Gc and spermatogenic activities in Gr 3 and 4, and extensive fibrosis in Gr 4. A considerable suppression of spermatogenesis marked with devoid of endogenous spermatogonial population and absence of significant (P > 0.05) effect on key hematological variables were observed in 2 mg/kg-MT Gr. These findings coupled with the results of significant (P < 0.05) down-regulation of marker genes of undifferentiated spermatogonia (THY-1 and PLZF), Gc pluripotency (UCHL-1 , OCT-4, and DDX-4), and adhesion (E-cadherin and β -integrin); up-regulation of apoptotic genes (ID - 4 and BCL-6), and unchanged expression of Sertoli cell marker (vimentin), confirmed the safe and efficient depletion of endogenous Gc in 2 mg/kg-MT Gr. Furthermore, the effect of busulfan infusion on scrotal-testicular biometry, endocrine variables (plasma cortisol and testosterone), and Gc removal was more evident when busulfan was infused into MT than into CV. Overall, the results demonstrated that 2.0 mg/kg is an optimal single dose of busulfan when infused into the MT under USG guidance for the preparation of pre-pubertal recipient bucks. Overall, this study provides a basis to prepare suitable recipients through providing an available niche for efficient Gc transplantation in goats. • Intra-testicular busulfan infusion resulted into a dose-dependent depletion of germ-cells. • Busulfan infusion triggered down-regulation of pluripotency and adhesion marker genes in the testis. • Scrotal-testicular biometry variables affected more when busulfan is infused into MT. • Hematological variables mostly remain unaffected after intra-testicular busulfan infusion. • 2 mg/kg is the most optimal busulfan dose when infused into the MT with USG guidance. [ABSTRACT FROM AUTHOR]

Given the importance of males as semen donors for artificial insemination (AI) and the high incidence of low birthweight piglets at commercial farms, the impact of birthweight on fertility in boars deserves special attention. The aim of this study was to evaluate testicular morphofunctional parameters and semen characteristics in different birthweight boars. Forty littermate males were selected at birth and divided into two experimental groups, according to birthweight: high (HW, birthweight ranging from 1.80 to 2.15 kg, n = 20) and low (LW, birthweight ranging from 0.75 to 1.10 kg, n = 20). At 170 days of age, a sub-set of 24 littermate boars (n = 12 HW and n = 12 LW) were randomly selected for semen collection, which was performed once a week, at a 15-day interval, during five weeks. At 300 days of age, boars were orchiectomized, and the testis processed for histological and molecular analyses. The HW group was heavier and presented larger testes compared to LW animals (P < 0.05). Despite that, birthweight did not significantly affect semen volume or sperm quality parameters (concentration, motility, vigor or morphology), although LW boars produced 38.2% fewer total sperm and 24% lower semen concentration, leading to 36.8% less inseminating doses. The histomorphometrical evaluation showed that seminiferous tubules diameter and germinal epithelium height were similar between experimental groups. However, LW boars presented shorter seminiferous tubules and, consequently, fewer Sertoli cells per testis (P 0.05). Even though plasma testosterone levels were equivalent in both birthweight groups, LW testis presented less androgen receptors (P < 0.05). Additionally, birthweight was positively correlated with total seminiferous tubule length and number of Sertoli cells (P < 0.01), and with body and testis weights (P < 0.01). Taken together, even though adult LW boars showed no evident semen pathologies or spermatogenesis commitment, mature HW males have the potential to produce more spermatozoa, consequently more semen doses per ejaculate, being more valuable to an industry that relies on AI. • Larger testicles. • Longer seminiferous tubules. • More Sertoli cells per testis. • Higher expression of androgen receptors. • Higher sperm production capacity. [ABSTRACT FROM AUTHOR]

Dysfunction of spermatogenesis is a major complication of diabetes mellitus (DM). This study characterized the protective effects of Dendrobium nobile -derived polysaccharides (DNP) against spermatogenetic dysfunction in mice with streptozotocin (STZ)-induced diabetes. The diabetic mice had lower body and testicular mass, and fewer spermatozoa with a higher incidence of malformation. The testicular histology showed disordered narrow seminiferous tubules covering a smaller area, and fewer spermatogenic cells. Moreover, the qRT-PCR analysis indicated that DM was associated with high expression of the pro-apoptotic factor Bax and low expression of the anti-apoptotic factor Bcl-2 in the testes. The qRT-PCR and immunohistochemical analysis clarified that DM was also associated with low testicular expression of the Sertoli cell (SC) markers GATA-4, WT1, and vimentin, and genes encoding the glycolytic rate-limiting enzymes LDHA, PKM2, and HK2. DNP treatment increased the body and testicular masses, sperm count, and number of spermatogenic cells of the mice, and reduced the proportion of abnormal sperm. DNP also reduced the expression of Bax, and increased that of Bcl-2, GATA-4, WT1, vimentin, LDHA, PKM2, and HK2, in the testes of the diabetic mice. Thus, DNP protects against spermatogenic dysfunction in diabetic mice by inhibiting apoptosis and activating the glycolytic pathway in their testes. • DNP treatment alleviates the histology of the testis and epididymis of diabetic mice. • DNP ameliorates the apoptosis of spermatogenic cells in testes of diabetic mice. • DNP recovers the number and function of testicular Sertoli cells in diabetic mice. • DNP enhances the expression of genes encoding rate-limiting glycolytic enzymes in testes of diabetic mice. [ABSTRACT FROM AUTHOR]

[Display omitted] • PCA and Raman spectroscopy were used to identify spermatogenesis in mouse testicular seminiferous tubules and tissue. • Raman measurements for single tubules and testes samples of adult and immature mice compared with busulfan-treated adults. • A higher signal of variability is obtained which questions the individual Raman spectra as a measurement for spermatogenesis. • PCA of these spectra shows a significant cluster separation between the ST of mature and immature mice. • The combination of PCA and Raman spectroscopy could be a noninvasive and potentially useful tool for spermatogenesis identification. 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. [ABSTRACT FROM AUTHOR]

The adipokines, visfatin, chemerin, and its receptor are expressed in the testis. It has also been shown that heat-stress alters the secretion and expression of other adipokines. Testicular heat-stress is now well known to cause the impairment in the testis. It has also been documented that heat-stress changes the expression of genes and proteins in the testis. To the best of our knowledge, the expression and localization of visfatin chemerin and its receptor have not been investigated in the heat-stressed testis. Therefore, the present study has investigated the expression and localization of these proteins in the heat-stressed testis. The expression of visfatin and chemerin and receptor exhibits a differential repossess against the heat stress. Visfatin expression was up-regulated while chemerin and chemerin receptor was down-regulated in the heat-stressed testis as shown by western blot analysis. The immunolocalization of visfatin and chemerin showed increased abundance in the seminiferous tubules of heat-stressed mice testis. Furthermore, abundance of visfatin, chemerin, and its receptor showed a decrease in abundance in the Leydig cells of heat-stressed testis. The decreased abundance of these proteins in the Leydig cells coincides with decreased 3β-HSD immunostaining along with decreased testosterone levels. These results suggest that heat-stress might decrease testosterone secretion by modulating visfatin and chemerin in the Leydig cells. The increased abundance of visfatin and chemerin in the primary spermatocytes, round spermatid, and multinucleated germ cells also coincides with increased immunostaining of active caspase-3. Moreover, expression of Bcl-2 was down-regulated, and expression of active caspase-3 and HSP70 were up-regulated along with increased oxidative stress in the heat-stressed testis, suggesting stimulated apoptosis. In conclusion, our results showed that visfatin, chemerin, and its receptor are differentially expressed in the testis under heat-stress and within the testis also it might differentially regulate testosterone biosynthesis in the Leydig cells and apoptosis in the seminiferous tubules. • Heat-stress up-regulates visfatin and down-regulates chemerin and its receptor (CMKLR1) in the testis. • However, abundance of visfatin, chemerin and CMKLR1 decreases in the Leydig cells of heat-stressed testis. • The decreased abundance of visfatin and chemerin in Leydig cells coincides with decreased testosterone. • Elevated immunostaining of visfatin and chemerin in the seminiferous tubules coincides with increased apoptosis. • Heat-stressed differentially regulates the expression of visfatin and chemerin system. [ABSTRACT FROM AUTHOR]

Glyphosate (GLY) is a widely used herbicide that has been revealed to inhibit testosterone synthesis in humans and animals. Melatonin (MET) is an endogenous hormone that has been demonstrated to promote mammalian testosterone synthesis via protecting mitochondrial function. However, it remains unclear whether MET targets mitochondria to alleviate GLY-inhibited testosterone synthesis in avian. In this study, an avian model using 7-day-old rooster upon chronic exposure to GLY with the treatment of MET was designed to clarify this issue. Data first showed that GLY-induced testicular Leydig cell damage, structural damage of the seminiferous tubule, and sperm quality decrease were mitigated by MET. Transcriptomic analyses of the testicular tissues revealed the potentially critical role of mitophagy and steroid hormone biosynthesis in the process of MET counteracting GLY-induced testicular damage. Also, validation data demonstrated that the inhibition of testosterone synthesis due to GLY-induced mitochondrial dynamic imbalance and concomitant Parkin-dependent mitophagy activation is alleviated by MET. Moreover, GLY-induced oxidative stress in serum and testicular tissue were significantly reversed by MET. In summary, these findings demonstrate that MET effectively ameliorates GLY-inhibited testosterone synthesis by inhibiting mitophagy activation, which provides a promising remedy for the application of MET as a potential therapeutic agent to antagonize reproductive toxicity induced by GLY and similar contaminants. [Display omitted] • Melatonin (MET) prevents glyphosate (GLY)-induced testicular damage in roosters. • Mitophagy and steroid hormone biosynthesis are revealed by transcriptomic analysis. • MET alleviates GLY-inhibited testosterone synthesis via inhibiting mitophagy. • MET mitigates GLY-activated Parkin-dependent mitophagy by inhibiting oxidative stress. [ABSTRACT FROM AUTHOR]

Spermatogonial stem cells (SSCs) provide a foundation for spermatogenesis, but the mechanism of SSC proliferation is still poorly understood. To investigate whether and how ascorbic acid (AA) regulates the growth of mouse SSCs in vitro , the SSCs were cultured in different concentration AA medium for 14 days. The proliferation, apoptosis and the reactive oxygen species (ROS) levels of SSCs in different AA groups were respectively detected. Moreover, the SSC activity in 40 μg/mL AA group and the control was tested by a transplantation assay. To explore the mechanism of AA regulating mouse SSCs proliferation, the dishevelled homolog 2 (DVL2) and nucleoredoxin (NRX) protein levels, the expression of axis inhibition protein 2 (Axin2), leucine-rich G-protein coupled receptor 5 (Lgr5), B-cell lymphoma-2 (Bcl-2), BCL2-Associated X (Bax), c-myc and cyclin D1 genes in Wnt/ β -catenin pathway were respectively confirmed. The results showed that the adding concentration of AA did not affect the main shape of SSCs. A 40 μg/mL AA in culture medium promoted the proliferation, and decreased the ROS production and apoptosis rate of SSCs. Moreover, colonization efficiency in the seminiferous tubules of the recipient testis in 40 μg/mL AA group was higher compared with the control group by a transplantation assay. Finally, the appropriate ROS in the 40 μg/mL AA group further adjust the levels of DVL2 and NRX protein in the Wnt/ β -catenin pathway to maintain the nuclear intensity of β -catenin, in turn, the expression of apoptosis gene Bax decreased, while the expression of Bcl2, Axin2 , Lgr5 , c-myc and cyclin D1 genes increased. The study confirmed that AA adjusts the endogenous ROS level to impact on SSC proliferation in a dose-dependent manner by Wnt/ β -catenin signaling pathway. [Display omitted] • The effect of the different concentrations of AA on mouse SSC proliferation in an AA concentration-dependent way. • AA-mediated intracellular ROS levels of SSCs adjusted the expression DVL2 and NRX protein expression levels, and then changed the nuclear accumulation of β-catenin in Wnt/β-catenin signal pathway. • a moderate AA-mediated ROS metabolism maintain the accumulation of certain level nuclear β-catenin to further regulate the expression of downstream target genes and apoptosis-related genes to control the proliferation of mouse SSCs by Wnt/β-catenin signaling pathway. [ABSTRACT FROM AUTHOR]

Gonadal white adipose tissue (gWAT) can regulate gametogenesis via modulation of neuroendocrine signaling. However, the effect of gWAT on the local microenvironment of the gonad was largely unknown. Herein, we ruled out that gWAT had a neuroendocrine effect on gonad function through a unilateral lipectomy strategy, in which cutting off epididymal white adipose tissue could reduce seminiferous tubule thickness and decrease sperm counts only in the adjacent testis and epididymis of the affected gonad. Consistent with the results in males, in females, ovary mass was similarly decreased by lipectomy. We determined that the defects in spermatogenesis were mainly caused by augmented apoptosis and decreased proliferation of germ cells. Transcriptome analysis suggested that lipectomy could disrupt immune privilege and activate immune responses in both the testis and ovary on the side of the lipectomy. In addition, lipidomics analysis in the testis showed that the levels of lipid metabolites such as free carnitine were elevated, whereas the levels of glycerophospholipids such as phosphatidylcholines and phosphatidylethanolamines were decreased, which indicated that the metabolic niche was also altered. Finally, we show that supplementation of phosphatidylcholine and phosphatidylethanolamine could partially rescue the observed phenotype. Collectively, our findings suggest that gWAT is important for gonad function by not only affecting whole-body homeostasis but also via maintaining local metabolic and immune niches. [ABSTRACT FROM AUTHOR]

Zearalenone (ZEN) is a worldwide fusarotoxin that poses a threat to the consumer due to its chronic toxicity. Herein we examined the effects of ZEN on adult mouse testis, focusing on oxidative stress, biochemical and morphological parameters. In addition, since cytoskeletal remodeling is a key event for the production of good quality gametes, the expression and localization of two proteins, Dishevelled-associated activator of morphogenesis 1 (DAAM1) and Prolyl endopeptidase (PREP), involved in cytoskeletal dynamics during spermatogenesis were evaluated. To ameliorate the testicular dysfunction induced by ZEN we tested the eventual protective effects of lactic bacteria Lactobacillus plantarum MON03 (LP) on its reprotoxicity. Adult male mice were then treated daily for 2 wks by oral gavage with ZEN and/or LP. The results confirmed that ZEN altered sperm parameters, generated oxidative stress and provoked structural alteration, evidenced by the increased number of abnormal seminiferous tubules and of apoptotic cells, particularly Leydig cells. Interestingly, at molecular level we evaluated, for the first time, the ability of ZEN to alter DAAM1 and PREP protein level and localization. Moreover, the co-treatment with LP, thanks to its capacity to reduce ZEN bioavailability in the gastrointestinal tract, ameliorated all the considered parameters. These results suggest the use of this probiotic as food supplement to prevent/counteract ZEN-induced reprotoxicity. [Display omitted] • Zearalenone damage testicular structure and function. • Lactobacillus plantarum protects male mice against Zearalenone reprotoxicity. • Lactobacillus plantarum can be used as dietary supplement against fusarotoxins. [ABSTRACT FROM AUTHOR]

[Display omitted] • Pubertal acrylamide exposure decrease sperm concentration by interfering spermatogenesis in male mice. • Pubertal acrylamide exposure induces apoptosis of germ cells in the testes. • Pubertal acrylamide exposure delays the meiotic prophase I progression. • Pubertal acrylamide exposure disrupts H2AX phosphorylation pattern of meiotic prophase I. Teenagers are a major group likely to love junk foods, such as potato chips and bread items, which contain high levels of acrylamide (AA). The increasing evidence suggests that AA exposure may be associated with decreased reproductive capacity in humans and animals. However, the reproductive toxicity of AA in pubertal males has not been fully elucidated. In this study, we evaluated the effects of pubertal AA exposure on adult spermatogenesis in male mice. Mice were exposed to AA at 0, 5, 10, 20, and 40 mg/kg/day by gavage from postnatal day 28 (PND28) to PND56. Our results showed that pubertal AA exposure increased apoptosis of germ cells in seminiferous tubules, decreased sperm concentration, and caused defects in sperm of adult mice. To explore the possible mechanisms of AA on spermatogenesis, the meiotic process was analyzed. The ratio of leptotene and zygotene spermatocytes increased, while the pachytene and diplotene spermatocytes decreased in AA-treated mice. Further analysis revealed that AA exposure disrupted the pattern of H2AX phosphorylation expansion, synapsis, and the crossover formation during meiotic prophase I (MPI). Taken together, these results indicate that pubertal AA exposure affects the spermatogenesis may be by disrupting the MPI progression of male mice. [ABSTRACT FROM AUTHOR]

• Prenatal DBP 100 mg/kg exposure did not cause malformation in next generation rats • Prenatal DBP exposure did not decrease the body weight or AGD of male rats • Prenatal DBP exposure induced accumulation of spermatocytes at stages IX to X • Prenatal DBP exposure significantly delayed spermatogenesis at puberty and adulthood • Modified BrdU injection method can quantitatively show delay in rat spermatogenesis Di(n-butyl) phthalate (DBP) esters are plasticizers that are used to provide transparency and flexibility in household plastic products but can easily leach out to contaminate organisms and the environment. We investigated whether prenatal DBP exposure affects spermatogenesis in rats. Pregnant Sprague–Dawley rats were injected with DBP 10, 50, and 100 mg/kg, or vehicle, administered intragastrically, on gestation days 12–21. At 9 or 17 weeks, 5-bromodeoxyuridine (BrdU) 50 mg/kg was injected intraperitoneally, and one testis was removed 3 h later. The remaining testis was excised 12.95 days + 3 h after the BrdU injection. Immunohistochemical analysis of BrdU was performed with periodic acid-Schiff and hematoxylin counterstaining for a quantitative analysis of the delay in one cycle of spermatogenesis. The DBP 100 mg group showed that the ratio of the appearance of seminiferous tubules in stages VII and VIII were significantly decreased, but those of stages IX and X were significantly increased compared to the Vehicle group. The reference value for the duration of spermatogenesis per cycle was set at 310.8 h. The DBP 100 mg group showed a significant delay in the duration of one cycle of spermatogenesis (16.95 h at puberty and 19.01 h at adulthood) compared with the Vehicle group. This study determined that F1-generation rats with prenatal DBP 100 mg exposure revealed significant accumulation of spermatogenic cells at stages IX to X in the second and third cycles, and the significant delay in the duration of spermatogenesis was more prominent at adulthood than in puberty. [ABSTRACT FROM AUTHOR]

Human infertility has become a global medical and social health problem. Mice deficient in type 3 adenylyl cyclase (AC3), a key enzyme that synthesizes cyclic adenosine monophosphate (cAMP), develop male infertility, although the underlying molecular mechanisms remain unknown. We performed a label-free quantitative (LFQ) proteomics analyses to identify testicular differentially expressed proteins (DEPs) and their respective biological processes. Furthermore, histological examination demonstrated that AC3 deficiency in mice led to mild impairment of spermatogenesis, including the thinning of seminiferous epithelium and local lesions in the testis. We further identified that the integrity of the blood–testis barrier (BTB) was impaired in AC3 knockout (AC3 −/−) mice accompanied with the reduction in the expression of tight junctions (TJs) and ectoplasmic specialization (ESs)–related proteins. In addition, the deletion of AC3 in mice also reduced the germ cell proliferation, increased apoptosis, and decreased lipid deposition in the seminiferous tubules. Collectively, our results revealed a role of AC3 in regulating the BTB integrity during spermatogenesis. Thus, our findings provide new perspectives for future research in male infertility. Male AC3 −/− mice displays mild impairment of spermatogenesis including BTB dysfunction, reduced proliferation of the germ cells, increased apoptosis of the germ cells, and decreased lipid deposition. [Display omitted] • AC3 deletion disrupts the BTB integrity. • AC3 deficiency in mice leads to mild impairment of spermatogenesis and deformity of the sperm head. • AC3 deletion suppresses the proliferation of germ cells and increases germ cells apoptosis. • Deletion of AC3 led to decrease in lipid deposition around seminiferous tubules. [ABSTRACT FROM AUTHOR]

A comprehensive understanding of gonad development in the Pacific whiteleg shrimp, Litopenaeus vannamei , is vital for cultivating high-quality male broodstock and seedlings. The aim of this study was to clarify the characteristic changes that transpire during discrete developmental phases in male L. vannamei gonads via morphological, histological, biochemical, and molecular analyses. The gonadal development of male L. vannamei is categorized into four distinct phases during sexual maturation based on observations of external morphological traits. During phase I, male gonads complete gonadal differentiation but have yet to initiate development. Phase II signifies rapid development and initiation of reproductive functions; concurrently, a subtle whitish ampulla emerges at the base of the fifth pereiopod, concomitant with structural differentiation in the vas deferens. Seminiferous tubules and germ cells appear in the testis, and a primary spermatophore layer forms in the medial vas deferens, although mature sperm masses are absent. Phases III and IV primarily involve enhancing individual reproductive abilities. In phase III, a white spermatophore structure develops at the base of the fifth pereiopod, and the first pair of pleopods form the petasma. In Phase IV, the ampulla becomes fuller, and the spermatophore can be expelled artificially. Our analysis indicated that in phase II, the mRNA expression levels of the crustacean hyperglycemic hormone family (CHHs), which includes CHH isoforms (CHH1 and CHH2), a gonadal-inhibiting hormone (GIH), and a molting-inhibiting hormone (MIH), decreased within the eyestalk. During phase III, despite the presence of immature sperm in the vas deferens and a reduction in serum hyperglycemic hormone levels, gonadal development approached a state of maturity. In phase IV, mature sperm are present in the medial vas deferens, indicating that during gonad maturation, the mRNA expression levels of CHHs and insulin-like androgenic gland hormone (IAG) significantly increase. Commencing from phase II, a prominent shift occurs in the transcription levels of testicular genes. Analysis via weighted gene coexpression network (WGCNA) and functional enrichment demonstrated the upregulation of pathways pertinent to the cell cycle, RNA transport, and meiosis, juxtaposed with the downregulation of pathways linked to lysosomes. This study provides key insights into the reproductive biology and maturation of L. vannamei males. • First identification of four male Penaeus vannamei gonad maturation stages. • Revealed hormone regulation across male P. vannamei 's developmental phases. • Phase II marks onset of gonadal development, with significant testicular transcript shifts and CHHs gene decrease. • Phase III is marked by enhanced reproductive potential, crucial for spermatophore maturation [ABSTRACT FROM AUTHOR]

The biological effects of simulated photoperiod and melatonin on the control of reproduction of guinea fowls (Numida meleagris) are not well understood. Herein, thirty (30) sexually mature guinea fowl cocks were randomly assigned to 1–6 groups (n = 5) and subjected to different photoperiodic regimes in the presence or absence of exogenous melatonin (Mel; 1 mg/kgBW/day, i/m) for eight weeks. Testes of the euthanized cocks were processed for gross morphology, histological, histochemical, and oxidative stress markers. Testosterone concentration was determined in serum samples using the enzyme-linked immunosorbent assay (ELISA) technique. We observed an increase in testicular size in the Mel and Non-Mel groups under long-day (LD) photoperiods, and in the Non-Mel group under short-day (SD) photoperiod. Conversely, the testicular size was drastically reduced in the Mel group for SD. Seminiferous tubules in the Mel and Non-Mel groups of the SD showed cytomorphological changes, including degenerated cells, focal vacuolations, and depletion of germinal epithelium. However, the germinal epithelium appeared to be complete and active in both the Mel and Non-Mel groups for the LD. In all groups, the testes showed positive staining for PAS with varying intensities. There was a significant difference in PAS-staining intensity between different photoperiodic regimes and exogenous melatonin. The study observed the interaction between photoperiods and exogenous melatonin on glutathione reductase (GSH), malondialdehyde (MDA), and serum testosterone. Overall, the results indicated that a long-day (LD) photoperiod, combined with exogenous melatonin, enhanced reproductive activity in male guinea fowl by increasing testicular size and serum testosterone concentration. [Display omitted] • Testicular size and serum testosterone concentration decreased in male guinea fowl exposed to short-day (8 L:16D) photoperiod and exogenous melatonin • Enhanced reproductive performance was observed in male guinea fowl exposed to long-day (16 L:8D) photoperiod and exogenous melatonin. • Increased levels of testicular malondialdehyde (MDA) were observed following exposure to a short-day (8 L:16D) photoperiod and exogenous melatonin. [ABSTRACT FROM AUTHOR]

• Coenzyme Q10 (CoQ10) mitigated MTX-induced testicular toxicity, primarily via its anti-inflammatory, anti-oxidant, and anti-apoptotic effects. • The novel protective effect of Co-Q10 was mediated via Nrf2-PPAR-γ signaling loop and its downstream iNOS, NF-κB, and the Bcl-2 family proteins. • The novel protective effect of Co-Q10 was mediated via Nrf2-PPAR-γ signaling loop and its downstream iNOS, NF-κB, and the Bcl-2 family proteins. Studies have identified Coenzyme Q10 (CoQ10) as a promising agent in improving idiopathic male infertility; however, its role in chemically or environmentally induced testicular dysfunction is not well-established. We investigated the potential of CoQ10 to attenuate methotrexate (MTX)-induced testicular damage and to identify molecular targets of CoQ10 effects. Wistar rats received a single intraperitoneal dose of 20 mg/kg MTX on the fifth day of the 10-day experimental protocol. 100 mg/kg CoQ10 was given orally daily for ten days, alone or combined with MTX. The testes of MTX-treated animals showed thickened tunica albuginea, distortion of seminiferous tubules with a marked reduction of germinal lining, a few primary spermatocytes with no spermatozoa, apoptotic cells, congested sub-capsular and interstitial blood vessels, and interstitial edema. Reduction of reproductive hormones and increased oxidative, inflammatory, and apoptotic biomarkers levels were also seen in the MTX-treated rats. CoQ10 + MTX-treated rats were protected against MTX-induced testicular histological changes and showed improvement in testosterone, luteinizing-, and follicle-stimulating hormone serum levels compared to the MTX group. The testes of the CoQ10 + MTX-treated rats showed reduced malondialdehyde, myloperoxidase, tumor necrosis factor -α, interleukin-6 and -1β and Bax: Bcl2 ratio and enhanced glutathione, and catalase compared to MTX alone. CoQ10 enhanced MTX-induced downregulation of Nrf2 and PPAR-γ signaling and modulated its downstream targets, the inducible nitric oxide synthase, NF-κB, Bax, and Bcl2. In conclusion, CoQ10 targeted the Nrf2-PPAR-γ signaling loop and its downstream pathways, mitigating MTX-induced oxidative stress-related damages and alleviating the testicular dysfunction MTX caused. Our data suggest Nrf2-PPAR-γ signaling as a potential therapeutic target in testicular toxicity, where oxidative stress, inflammation, and apoptosis trigger damage. [ABSTRACT FROM AUTHOR]

Testicular tissue culture in vitro is considered an important tool for the study of spermatogenesis and the treatment of male infertility. Although agarose hydrogel is commonly used in testicular tissue culture, the efficiency of spermatogenesis in vitro is limited. In this study, testicular tissues from adult mice were cultured using a gas-liquid interphase method based on agarose (Agarose), gelatin methacryloyl (GelMA), alginate methacryloyl (AlgMA), dextran methacryloyl (DexMA), and mixture GelMA-Agarose, AlgMA-Agarose, and DexMA-Agarose hydrogels, respectively, for 32 days in vitro. The integrity of the seminiferous tubules, the density and proportions of spermatogonia, spermatocytes, Sertoli cells, and testosterone concentrations were quantified and compared between groups. Properties of different hydrogels including compression modulus, Fourier Infrared Spectroscopy (FITR) spectra, pore size, water absorption, and water retention were tested to investigate how biochemical and physical properties of hydrogels affect the results of testicular tissue culture. The results indicate that testicular tissues cultured on AlgMA exhibited the highest seminiferous tubule integrity rate (0.835 ± 0.021), the presence of a high density of spermatocytes (2107.627 ± 232.082/mm2), and a high proportion of SOX9-positive well-preserved seminiferous tubules (0.473 ± 0.047) compared to all remaining experimental groups on day 32. This may be due to the high water content of AlgMA reducing the toxic effect of oxygen on testicular tissue. In the later period of culture, testicular tissues cultured on DexMA, not DexMA-Agarose, produced significantly more testosterone (18.093 ± 3.302 ng/mL) than the other groups, suggesting that DexMA is friendly to Leydig cells. Our study provides a new idea for the optimization of the gas-liquid interphase method for achieving in vitro spermatogenesis, facilitating the future achievement of efficient in vitro spermatogenesis in more species, including humans. [ABSTRACT FROM AUTHOR]

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. • Finasteride exhibited a negative influence on testicular weight and semen analysis. • Finasteride induced ultrastructural seminiferous tubules changes. • Finasteride increased apoptotic and oxidative stress markers. • Hesperidin ameliorated finasteride-induced deleterious effects. [ABSTRACT FROM AUTHOR]

The aim of the study was to explore the regulatory mechanism of differences in embryonic gonadal development between intergeneric distance hybrid offspring Mulard ducks and parent ducks. The morphological differences gonadal tissues of Muscovy ducks, Pekin ducks and Mulard ducks at 12.5-day embryonic age were observed by sectioning and hematoxylin-eosin (HE) staining. Then followed by transcriptome sequencing to screen for gonadal development-related differentially expressed circRNAs and mRNAs to construct a competitive endogenous RNA (ceRNA) regulatory network. Finally, qRT-PCR and luciferase reporter system were used to verify the sequencing data and targeting relationship of ceRNA pairs. The results showed that the seminiferous tubule lumen of Mulard ducks was not obvious, while there were obvious seminiferous tubules and tubular structures in testis of Pekin ducks and Muscovy ducks, with number and shape indicating maturity. There were 18 upregulated circRNAs and 16 downregulated circRNAs in Mulard ducks and Pekin ducks, respectively, and 39 upregulated circRNAs and 1 downregulated circRNA in Mulard ducks and Muscovy ducks, respectively. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis found that genes involves in dorso-ventral axis formation, for example, neurogenic locus notch homolog protein 1 (NOTCH1), were significantly enriched (P < 0.05). The novel_circ_0002265-gga-miR-122-5p-PAFAH1B2 regulatory network was constructed. The qRT-PCR results showed that the sequencing results were reliable. The dual-luciferase reporter assay showed that gga-miR-122-5p exists binding site of circ_0002265 and PAFAH1B2, indicating circ_0002265-gga-miR-122-5p-PAFAH1B2 targeting relationship. In summary, the embryonic gonadal development of intergeneric hybrid Mulard ducks may be regulated by differentially expressed circRNAs and genes, such as novel_circ_0000519, novel_circ_0003537, NOTCH1, FGFR2, PAFAH1B1, and PAFAH1B2, among which circ_0002265-gga-miR-122-5p-PAFAH1B2 may participate in the targeted regulation of gonadal development in Mulard ducks. The findings of this study are helpful for analyzing the mechanism of embryonic gonadal development differences in avians. [ABSTRACT FROM AUTHOR]

Obesity is a global health issue that affects more than one-third of the population and is linked to several chronic consequences, including reproductive abnormalities. Chitosan is gaining popularity as a natural anti-obesity agent in various medical applications. As a result, this study aimed to evaluate the protective effect of crawfish chitosan on the reproductive capacity of male rats fed a high-fat diet. The crystallinity of chitosan particles was determined by X-ray diffraction (XRD). Twenty-one male Wistar rats were divided into three groups (n = 7/group); group I (control) was fed a standard pellet diet and water; group II, a high-fat diet (untreated), received 0.5 ml of 1% glacial acetic acid; and group III, a high-fat diet treated with 0.5 ml of 30 mg/kg/day of chitosan dissolved in 1% glacial acetic acid via oral gavage for 28 days. Many parameters were determined in this study, including sperm characteristics, hormonal profiles LH, FSH, and testosterone, as well as liver and kidney functions, testicular oxidative stress, and histological alterations. Results revealed that chitosan reduced body weight, enhanced sperm count and viability, and boosted reproductive hormone levels. It also increased antioxidant activity and improved kidney and liver functions compared to the HFD group. Histopathological observations of treated rats showed a normal appearance of seminiferous tubules with regular structure, displaying entire stages of spermatogenesis and presence of spermatids in the lumen. Conclusively, crawfish chitosan improved testicular dysfunction in HFD-fed rats by reducing oxidative stress. [ABSTRACT FROM AUTHOR]

Nanoplastics have been demonstrated to be reproductively toxic to mammals. However, the mechanisms of nanoplastics induce reproductive damage in mammals, especially their effects on spermatogenesis, remain elusive. Herein, we explored the effects and underlying mechanisms of polystyrene nanoplastics (PS-NPs) on the testicular development of male mice after 28 days of exposure, representing the first systematic study of PS-NPs-induced male reproductive injury by integrating histomorphology, transcriptomics and proteomics. PS-NPs decreased the sperm concentration, sperm motility, and disrupted the structure of the seminiferous tubules of the mice. Besides, transcriptome and proteome analyses revealed that PS-NPs disrupted spermatogenesis by inhibiting the transcription of Prm3 / Tnp1 / Aurkc / Mea1 / Mettl14 and the expression of Pmfbp1/Ggn/Fsip2. Furthermore, PS-NPs enabled Hsd3b5 protein expression to reduce dihydrotestosterone levels, and affected sperm flagellar assembly by decreasing the expression of Dnah8 / Tekt5 / Rsph6a. Moreover, PS-NPs induced testicular cell apoptosis by up-regulating the expression of cathepsins (B/F/H). In addition, PS-NPs destroyed tight junctions by reducing the expression of the Claudin family (3/5/15). In conclusion, PS-NPs can disrupt spermatogenesis by altering the expression patterns of transcriptome and proteome, inducing testicular cell apoptosis and destroying tight junctions. [Display omitted] • PS-NPs disrupt spermatogenesis by inhibiting the expression of Prm3/Aurkc/Pmfbp1. • PS-NPs enable Hsd3b5 protein expression to reduce dihydrotestosterone levels. • PS-NPs affect sperm flagellum by decreasing the expression of Dnah8/Tekt5 / Rsph6a. • Cathepsins (B/F/H) are involved in PS-NPs-induced testicular cell apoptosis. • PS-NPs destroy tight junction via suppressing the expression of Claudins (3/5/15). [ABSTRACT FROM AUTHOR]

Mammalian spermatogenesis is a temperature-sensitive process, and an increase in testicular temperature impairs spermatogenesis. Leptin modulates testicular activity, but the effect of leptin or its synthetic analogue on heat-induced testicular impairment is unclear. We investigated the effects of synthetic leptin peptide (116–130 amides) on testicular activity in heat-stressed mice model. 15 adult mice (25.54 ± 1.43 g) were selected for the study. Ten mice were subjected to a single heat stress treatment (HS) at 43 °C for 15 min by submerging the lower half of the body in a thermostatic water bath. After heat treatment, mice were divided into two groups, the heat-stressed HS group (n = 5) and the second group as HSL, treated with leptin peptide (116–130 amide) for 14 days. The HS group showed a significant (p < 0.05) decline in the GSI (0.25 ± 0.018), Johnsenscore (4.5 ±.19), seminiferous tubule diameter (160.75 ± 10.18 μm), germinal epithelium height, (GEH) (37.5 ± 1.59 μm) compared to the CN (GSI-0.37 ± 0.015; Johnsen score-7.9 ± 0.20; GEH- 73.25 ± 1.29 μm; tubule diameter-230.25 ± 1.39 μm) and the HSL groups (GSI-0.38 ± 0.014; Johnsen' score-8.0 ± 0.32; GEH- 37.5 ± 1.59 μm; tubule diameter-160.75 ± 10.18 μm) groups. Heat treatment significantly (p < 0.05) increased the intra-testicular levels of leptin (HS-20.11 ± 2.1 pg/mg protein; CN-10.50 ± 0.17 pg/mg protein; HSL-12.99 ± 0.52 pg/mg protein) with a reduced level of pStat3, suggesting leptin resistance during testicular hyperthermia. Furthermore, heat treatment was associated with significantly (p < 0.05) decreased germ cell proliferation and reduced circulating testosterone levels (HS-2.69 ± 2.01 ng/mL; CN-7.69 ± 0.32 ng/mL; HSL-5.36 ± 0.73 ng/mL). However, the circulating androstenedione levels showed a significant (p < 0.05) increase in the HS group (0.75 ± 0.03 ng/mL) compared to the CN (0.51 ± 0.02 ng/mL) and HSL (0.57 ± 0.07 ng/mL) groups. Immunolocalisation of 3β−HSD showed moderate to faint staining in the Leydig cells in the HS group compared to the CN and HSL groups. Treatment with leptin peptide resulted in decrease in the intra-testicular leptin levels with increased phosphorylation of Stat3, suggesting improved leptin resistance, which was positively associated with increased germ cell proliferation, elevated testosterone levels, and improved testicular histoarchitecture. Testicular hyperthermia may cause leptin resistance and impaired leptin signalling, decreased testosterone biosynthesis and suppressed spermatogenesis, which could be a manifestation of leptin resistance. Treatment with leptin peptide improves leptin signalling and testicular activity in heat-stressed mice, but the underlying mechanism is still unclear. • Scrotal hyperthermia increases the intra-testicular leptin levels. • Intra-testicular hyperleptinemia leads to leptin resistance. • Leptin c-fragment peptide (116–130) improves leptin signaling and spermatogenesis. [ABSTRACT FROM AUTHOR]

A component from ginseng in which sugars and proteins are covalently bound is named Panax ginseng glycoproteins (PGG). The contents of neutral carbohydrate, acid carbohydrate, and protein were 45.4%, 4.3% and 51.1%. The average molecular weight was 12,690 Da. The structure analysis showed that PGG had more than 1100 glycoproteins with molecular weight between 308.13 Da and 9991.52 Da, it was divided into two parts: long chain structure and short chain structure. These two parts were compared in molecular mass, number of amino acids, theoretical pI, instability index, aliphatic index and GRAVY. The in vivo distribution test of mice showed that PGG was enriched in mice testis, testicular tissue sections showed strong fluorescence signal expression on the surface of seminiferous tubules. We used cyclophosphamide (CP) to establish a mice model of oligoasthenozoospermia to investigate the anti-oligoasthenozoospermic effect of PGG. The results showed that PGG increased the levels of sex hormones T, FSH, PRL and sperm quality. Histopathology demonstrated that PGG promoted the differentiation process. The organ coefficient indicated that PGG had no obvious toxic and side effects. And the mechanism may be to affect the expression of protein levels such as p-ERK/ERK, p-AKT/AKT, Caspase-3, Bcl-2 and Bax. Therefore, PGG has the potential to develop into drugs for improving spermatogenic disorders. [ABSTRACT FROM AUTHOR]

• Dose effect of CBNPs exposure to three life stages of mice were observed. • Testicular oxidative stress and inflammation had time/dose-dependent effects. • Testicular damage structurally happened with exposure dose effects. • Sperm qualities functionally decreased with exposure dose effects. Environmental carbon black nanoparticles (CBNPs) can enter into various organs including testes through the respiratory tract. However, there are few studies describing reproductive toxicity of CBNPs after respiratory exposure. In this study, male KM mice were exposed to CBNPs in their natural breathing state. Four-, 8-, and 12-week-old mice were exposed to 0, 9, 18 and 27 mg/m3 of CBNPs for 4 weeks in order to examine the relationship between CBNP exposure and age. Eight-week-old mice were exposed to CBNPs at the same four concentrations for 1–4 weeks in order to examine the effects of CBNP exposure time. After CBNP exposure, testicular oxidative stress and inflammation increased significantly, and these effects varied with exposure time. Seminiferous tubule diameter (STD), seminiferous epithelium height (SEH), the number of spermatogenic and Leydig cells, sperm motility, and sperm speed decreased significantly, and these effects varied with exposure dose. Data analyses suggested that increased oxidative stress and inflammation in testes damaged testicular morphology, spermatogenesis, and testosterone secretion, and decreased sperm quality. Morphological damage to the testes was also closely related to decreased the sperm quantity. These findings are of significance for evaluating the reproductive toxicity of CBNPs. [ABSTRACT FROM AUTHOR]

Is vitrification with microinjection of single seminiferous tubules an efficient cryopreservation approach for limited testicular tissue? Testicular tissue from 10 patients with normal spermatogenesis were assigned to a fresh control group or one of the following cryopreservation procedures: uncontrolled slow freezing (USF) using either 1.5 or 2.1 M DMSO combined with sucrose and vitrification with or without single seminiferous tubules microinjection. Single seminiferous tubules microinjected with cryoprotective agents (CPA) enhanced the penetration of CPA compared with CPA-treated testicular tissue fragments. Microinjection of seminiferous tubules (VLP) maintained tubule structural integrity and germ cell numbers, and reduced spermatogonial apoptosis after cryopreservation compared with vitrification without microinjection (apoptosis rate: VLP versus vitrification without microinjection, P = 0.047; VLP versus USF, P= 0.049). Freezing of single seminiferous tubules using 0.25-ml straws and traditional sperm freezing methods protected sperm retrieval and recovery rates, and the progressive motility index. Vitrification of single seminiferous tubule with microinjection of low CPA concentration is an effective approach to testicular cryopreservation. [ABSTRACT FROM AUTHOR]

Meiotic recombination is key to the repair of DNA double-strand break damage, provide a link between homologs for proper chromosome segregation as well as ensure genetic diversity in organisms. Defects in recombination often lead to sterility. The ubiquitously expressed Rad51 and the meiosis-specific DMC1 are two closely related recombinases that catalyze the key strand invasion and exchange step of meiotic recombination. This study cloned and sequenced the coding region of cattle-yak Rad51 and determined its mRNA and protein expression levels, evaluated its molecular and evolutionary relationship as well as evaluated the histo-morphological structure of testes in the yellow cattle, yak and the sterile cattle-yak hybrid. The Rad51 gene was amplified using PCR, cloned and sequenced using testicular cDNA from yak and cattle-yak. Real-time PCR was used to examine the expression levels of Rad51/DMC1 mRNA in the cattle, yak and cattle-yak testis while western blotting, immunofluorescence and immunohistochemistry were used to assess the protein expression and localization of Rad51/DMC1 protein in the testicular tissue sections. The results revealed that the mRNA and protein expression of Rad51 and DMC1 are extremely low in the male cattle-yak testis with a corresponding higher incidence of germ cell apoptosis. There was also thinning of the germinal epithelium possibly due to the depletion of the germ cells leading to the widening of the lumen area of the cattle-yak seminiferous tubule. Our findings provide support for the hypothesis that the low expression of Rad51 and DMC1 may contribute to the male hybrid sterility in the cattle-yak. • F1 male hybrids between male yellow cattle and yak referred to as cattle-yak exhibits sterility while the female is fertile. • Male cattle–yak sterility is attributed to meiotic arrest at prophase I. • Rad51 which plays a crucial role in recombination is under-expressed in the male cattle-yak testis. • There is a higher incidence of germ cell apoptosis in the cattle-yak. • There is thinning of the germinal epithelium and widening of the seminiferous tubule lumen area in the cattle-yak. [ABSTRACT FROM AUTHOR]

The aim of this study was to determine the cellular and molecular mechanisms of leptin (LEP) and the leptin receptor (LEPR) in testicular development of prepubertal ganders. In an in vivo animal experiment, active immunization against LEPR severely depressed prepubertal testicular development by significantly reducing testicular weights at 200 and 227 days of age. The number of elongated spermatids in the seminiferous tubules was also significantly decreased by immunization with LEPR at ages of 200 and 227 days. Inhibition of testicular development by LEPR immunization was associated with decreases in LHR , StAR , 3β-HSD , CYP11A1 , CYP17A1, and PRLR mRNA expression levels in testicular tissue, which resulted in a significant decrease in testosterone synthesis. In the in vitro experiments, the addition of LEP combined with anti-LEPR antibodies strengthened LEPR signal transduction, and inhibited significantly testosterone production in cultured Leydig cells isolated from prepubertal gander testes. The mRNA expression of LHR, StAR , 3β-HSD , CYP11A1 , CYP17A1 also decreased significantly after treatment with LEP combined with anti-LEPR antibodies in cultured Leydig cells. These results suggest that anti-LEPR antibodies strengthen LEPR signaling transduction in the presence of LEP, and immunization against LEPR inhibited testes development and testosterone secretion in prepubertal ganders. • Immunization against leptin receptor severely depressed prepubertal testicular development in ganders. • Combined treatment with leptin and anti-LEPR antibodies inhibit steroidogenesis gene expression and testosterone secretion in cultured Leydig cells isolated from prepubertal gander testes. [ABSTRACT FROM AUTHOR]

The objective of this study was to evaluate the effect of exposure to different colors of light during egg incubation on the reproductive parameters of male and female Japanese quails. A total of 1776 eggs were incubated under four lighting conditions for 24 h a day during the entire incubation period: white LEDs, green LEDs, red LEDs and darkness (control). The experimental design was a randomized block (incubation time) with four treatments of six replicates of two cages each. After hatching, the birds were housed in brood cages with 18 birds each to 35 days of age, when they were sexed and transferred to production cages for evaluation of reproductive parameters. After the onset of laying, the number of eggs in each cage was recorded daily, and the values were used to obtain the age of the females at first egg and at 80% laying. At 35 and 60 days of age, several birds from each cage were euthanized for anatomical and histological evaluation of the gonads. Two females from each cage were weighed every three days until 60 days of age to determine the growth curve. After 60 days, eggs from each cage were collected and assessed for external and internal quality. At 70, 74 and 78 days of age, semen collection was performed and seminal quality was evaluated. Then, the males were transferred to cages containing 9 females for the fertility test. Hatchability was higher (P < 0.05) in eggs incubated in the dark and under the red LED. The age of maximum growth was higher (P < 0.05) in birds from eggs incubated in the dark and under the white LED. There was no difference (P > 0.05) in the anatomical and histological characteristics of the testicles between the groups incubated under different light colors, except for the diameter of the seminiferous tubules, which was greater (P = 0.05) in the dark and in the white LED groups. There was no effect (P > 0.05) of light color during incubation on the productive index or egg quality of adult birds. There was also no effect (P > 0.05) on sperm quality, except for sperm motility, the values of which were higher (P < 0.05) in birds from eggs incubated in different colors of light. However, this difference was not sufficient to significantly (P > 0.05) influence bird fertility. It is concluded that under the studied conditions, the incubation of quail eggs under white, red, and green LED lamps does not influence the reproductive characteristics of the quails. • Light has a strong influence on the reproductive activity of birds. • Light colors during embryonic development of the reproductive axis were studied. • Hatchability was higher in eggs incubated in the dark and under the red LED. • Light colors during embryonic development increased the sperm motility. • Light colors does not affect the fertility and characteristics of quail eggs. [ABSTRACT FROM AUTHOR]

• We evaluated the effects of NMS on mouse testes in prepuberty. • The results revealed a significant decrease in Sertoli cell number at PND 16. • The number of p27-positive Sertoli cells was significantly high at PND 10. • A significant increase in serum corticosterone was detected at PND 10. • Further studies are needed to clarify the mechanism by which p27 levels elevate. Neonatal maternal separation (NMS) is an experimental model for early life stress, which affects the growth and development of various organs, resulting in adverse health effects in humans and animals. In our previous study, we demonstrated that NMS [(0.5-, 1-, 2-h/day NMS, from postnatal day (PND) 1–10] induced morphological changes to the male reproductive system, including decreased Sertoli cell numbers in mouse testes at PND 70. To clarify the mechanism by which NMS decreases Sertoli cell numbers, we evaluated the effects of NMS on mouse testes at PNDs 10 and 16. At PND 10, the Sertoli cell number was not significantly different among experimental groups; however, it decreased in 0.5- and 2-h/day NMS mice at PND 16. The termination of Sertoli cell proliferation in prepuberty can be induced by p27, a cyclin-dependent kinase inhibitor. At PND 10, we observed an increase in the number of p27-positive Sertoli cells in 2-h/day NMS mice. The seminiferous tubule diameters decreased significantly in 1- and 2-h/day NMS mice, and the relative interstitial area increased in 2-h/day NMS mice. Serum corticosterone level significantly increased, and serum testosterone level significantly decreased in the 2-h/day NMS mice. At PND 16, the tubule diameters and height of seminiferous epithelium were significantly higher in 0.5- and 2-h/day NMS mice. Our results suggest that NMS disturbs serum corticosterone and testosterone levels and increases the number of p27-positive Sertoli cells at PND 10, resulting in a decrease in the number of Sertoli cells at PND 16. [ABSTRACT FROM AUTHOR]

Diabetes mellitus (DM) is a global health burden, affecting about 463 million of the adult population worldwide. Approximately 94% of diabetic male individuals develop varying degrees of testicular disorders (TDs), which usually result in hypogonadism, hypotestosteronemia and defective spermatogenesis and steroidogenesis. Short chain fatty acids (SCFAs) have shown potential benefits in metabolic health. However, its effect on TD associated with DM is not clear. Howbeit, the present study investigated the hypothesis that SCFAs, acetate would ameliorate TD accompanying DM, possibly by suppressing proprotein convertase subtilisin/kexin type 9 (PCSK9). Male Wistar rats (210–240 g) were allotted into groups (n = 6/group): control (vehicle; po), DM with/without 200 mg/kg (po) of sodium acetate (SAc). Diabetes was induced by streptozotocin 65 mg/kg (iv) after a dose of nicotinamide (110 mg/kg). Semen/biochemical and histological analyses were performed with appropriate methods. In addition to hyperglycemia, hyperinsulinemia and reduced insulin sensitivity, DM led to increased serum and testicular triglyceride or total cholesterol/high-density lipoprotein cholesterol ratio, low-density lipoprotein cholesterol, malondialdehyde, TNF-α, IL-6 and PCSK9 as well as reduced high-density lipoprotein cholesterol and glutathione. Moreover, DM caused TD which is characterized by altered sperm parameters, disrupted tissue architecture, atrophied seminiferous tubules, deleterious spermatogonia, disappearance of lumen and cellular degeneration as well as decreased luteinizing hormone and testosterone. However, the administration of SAc attenuated these alterations. The study demonstrates that DM-induced TD is accompanied by elevated PCSK9. The results however suggest that SAc rescues testicular disorder/dysfunction associated with DM by suppression of PCSK9 and improvement of insulin sensitivity. [Display omitted] • Diabetes mellitus (DM) induces testicular dysfunction (TD). • DM-induced TD is associated with elevated PCSK9. • Sodium acetate ameliorates TD induced by DM. • Sodium acetate's ameliorative effect is inversely correlated to PCSK9. [ABSTRACT FROM AUTHOR]

We evaluated the effects of feeding 6%, 12% or 18% crude glycerin, containing 80.5% glycerol, on testicular histomorphometry and markers of oxidative stress and on plasma testosterone concentrations in lambs. Body weight, testicular biometric measurements, gonadosomatic index and net weight of the testicles were higher for the treated groups (P <0.05) compared with a control group that did not receive dietary glycerin. The mean total length of seminiferous tubules was higher in the 6% group (P <0.05), while the mean total tubular and seminiferous epithelium volumes increased in all treated groups (P <0.05). The volume of Leydig cells increased in the 12% group, while their number per gram of testicle decreased (P <0.05). There was a decrease in mean nuclear diameter and mean volume of Leydig cells, and an increase in the mean number of these cells per gram of testicle, in the 18% group (P <0.05). Plasma testosterone concentrations were unaffected. There was desquamation of seminiferous epithelium and vacuolation of Sertoli cells in the treated groups. Variable degrees of spermatocyte necrosis and the presence of giant cells were seen in all groups and there was intense vacuolation of Sertoli cells in the 12% and 18% groups. Superoxide dismutase and catalase production increased most in the 12% and 18% groups (P <0.05), while glutathione production was higher in the 18% group (P <0.05). Mean nitric oxide concentration decreased in all treated groups (P <0.05), while malondialdehyde production was higher in the 18% group than in the control and 6% groups (P <0.05). We conclude that the inclusion of 6% glycerin in the diet of lambs results in changes in testicular morphology that have been previously associated with improved reproductive function, but without evidence of oxidative stress. [ABSTRACT FROM AUTHOR]

• The paternal exposure to BaP compromised the puberty onset in males and females. • The male offspring presented a decrease in important sperm parameters. • The histology of the testis was altered. • The estrous cyclicity and some fertility parameters were displayed in the female offspring. Benzo(a)pyrene (BaP) is an ubiquitous environmental pollutant which can lead to adverse effects on male reproduction. However, the persistence of these changes on a multigenerational scale has not been sufficiently explored. This study evaluated if peripubertal exposure to BaP in male rats can induce reproductive impairment in offspring. Male rats received BaP at environmentally relevant doses (0, 0.1, 1, or 10 μg/kg/day) orally from post-natal (PND) 23–53. On PND 90, treated males were mated with non-treated females for obtaining the next generation (F1). The paternal exposure to BaP decreased the body weight of offspring on PND 1, 13 and 22, as well as it provoked a reduction in the relative anogenital distance of the males. This exposure also brought forward the onset of puberty, evidenced by an earlier vaginal opening and first estrous in females of the lowest dose group and by a delay in the testicular descent and preputial separation ages in males. The males presented a decrease in the daily sperm production and a disrupted sperm morphology. Furthermore, the testicular histology was altered, evidenced by a reduction in the Leydig cell numbers and in the seminiferous tubules diameter, as well as a disrupted seminiferous tubules staging. The estrous cyclicity and some fertility parameters were changed in the females, as well as alterations in the ovary and uterus histology were observed. BaP compromised several reproductive parameters of the F1 generation, suggesting that peripubertal exposure to this compound provokes permanent modifications in male germ line of F0 generation. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate the sperm head morphometry and chromatin condensation at different regions of the reproductive tract in bulls. Sperm smears of seminiferous tubules (ST), epididymis head (EH), body (EB), and tail (ET), and ductus deferens (DD) were stained with toluidine blue. Afterwards, the sperm head morphometry and chromatin alteration types were evaluated by a computational image analysis. Overall, spermatozoa of ST had lower (P < 0.05) area (A), perimeter (P), width (W), length (L), ellipticity (E), and Fourier harmonics (F0, F1, and F2). The chromatin decondensation (CD) and heterogeneity (CH) were higher (P < 0.05) in the ST region and decreased (P < 0.0001) during the migration along the reproductive tract (ST − DD direction). Considering the factors extracted (Factors 1 and 2) by the principal component analysis, the parameters A, P, W, L, and F0 were responsible for ∼36% of the Factor 1, while the E, F0, F1, and anterior-posterior symmetry (APS) contributed ∼27% to Factor 2. Both, CD and CH were associated with Factor 1 in the EH and ET regions and Factor 2 in the ST. Also, a well-defined difference between sperm heads collected from the ST and DD regions was observed by canonical analysis. The distribution of each chromatin alteration type was recorded. The proportion of normal sperm was lower (P < 0.05) in ST compared to other regions. Moreover, the chromatin influenced the morphometry and sperm heads with whole chromatin alteration type showed a smaller (P < 0.05) A, P, W, L, and E. In summary, the epididymal maturation is important for chromatin compaction and final morphometry of the sperm head. Also, the identification and quantification of the sperm chromatin condensation in different regions of reproductive tract can be used as potential biomarkers to predict the fertility in bulls. • Chromatin alteration types were measured in different regions of the reproductive tract in bulls. • Sperm head area, perimeter, width, length, ellipticity, and Fourier harmonics were lower in the seminiferous tubules. • Sperm chromatin decondensation and heterogeneity were higher in the seminiferous tubules. • The influence of the chromatin alteration on the sperm morphometric parameters was observed. [ABSTRACT FROM AUTHOR]

Compared to human cases, potential applications of Doppler imaging technology in farm animals' practice are still limited and are mostly concerned with females. Testicular blood perfusion (TBP) is the pathway through which all nutrients, oxygen, and regulatory agents pass through to the testis. TBP is crucial to the testis function because of the high metabolic rate and very low concentration of oxygen in the seminiferous tubules, which constitute about 70%–80% of the testicular mass. Color Doppler ultrasonography is one of the recent non-invasive technologies, which plays a vital role in the assessment of blood flow to the testis in various animal species. In the past 20 years, a significant number of studies have highlighted the importance of assessments of TBP using color Doppler ultrasonography to illustrate its potential uses for breeding soundness examination, diagnosis, and monitoring of infertility problems in farm animals. This review was aimed at providing, for the first time, updated knowledge on the potential clinical uses of color Doppler ultrasonography for the assessment of TBP in domestic animals. Additionally, a few essential details on the relationship between TBP and male fertility are discussed in this review. Furthermore, a number of caveats relating to the use of color Doppler ultrasonography are discussed, which collectively appear to constrain its routine, on-farm application for breeding soundness assessments, at least at the present. The new perspectives related to animal reproduction that researchers claim to do in further investigations are also discussed. Image 1 • Testicular blood flow has crucial roles for the testis function. • Color Doppler is a valid noninvasive tool to assess testicular hemodynamics in farm animals. • Close relationships between the testicular blood perfusion and semen quality of male animals. • Diagnosis of various reproductive disorders in the males by color Doppler ultrasonography. • There are challenges for future uses of color Doppler technologies in animal reproduction. [ABSTRACT FROM AUTHOR]

Cattleyak is a typically male sterile species. The meiosis process is blocked and the scarcity of spermatogenic stems cells are both contributing factors to the inability of male cattleyak to produce sperm. While Glial cell line-derived neurotrophic factor (GDNF) is the first discovered growth factor known to promote the proliferation and self-renewal of spermatogenic stem cells, its relationship to the spermatogenesis arrest of cattleyak remains unclear. In this report, we studied the differential expression of GDNF in the testis of yak and cattleyak, and discussed the optimal concentration of GDNF in the culture medium of undifferentiated spermatogonia (UDSPG) of cattleyak in vitro and the effect of GDNF on the proliferation of cattleyak UDSPG. The results indicated that GDNF expression in the testicular tissue of cattleyak was inferior to that of yak. Moreover, the optimum value for the UDSPG in vitro culture was determined to be 20–30 ng/mL for cattleyak. In vitro, the proliferation activity of UDSPG was observed to increase with additional GDNF due to the up-regulation of proliferation-related genes and the down-regulation of differentiation-related genes. We hereby report that the scarcity of cattleyak UDSPG is due to insufficient expression of GDNF, and that the addition of GDNF in vitro can promote the proliferation of cattleyak UDSPG by regulating the expression of genes related to proliferation and differentiation. This work provides a new insight to solve the issue of spermatogenic arrest in cattleyak. [Display omitted] • Spermatogenesis is impacted by the aberrant expression of GDNF in cattleyak testis. • Abnormal seminiferous tubule is one of the reasons for cattleyak's male sterility. • In vitro proliferation of cattleyak UDSPG can be enhanced by the addition of GDNF. [ABSTRACT FROM AUTHOR]

• Heating pad devices can cause acute damage to the seminiferous tubules in stallions. • Intratesticular MSCs therapy alleviates testicular damage after testicular heat stress. • The testicular insulation with the heating pad device did not obliterate testicular germ cells. Testicular degeneration (TD) is the most frequent cause of sub or infertility in stallions. Currently, mesenchymal stem cells (MSC) have been studied as a therapeutic option for several diseases including induced-TD in laboratory animals. Therefore, this study aimed to evaluate the effect of intratesticular MSC therapy on the testicular histology of stallions submitted to scrotal heat stress. Ten healthy Miniature-horse stallions were submitted to testicular heat stress induced by a heating wrap device (42-45°C). Afterward, the stallions were divided into two groups and treated seven days later. MSCs-treated stallions were treated with an intratesticular injection of 10 × 106 of MSCs diluted in 5 mL of PBS, whereas placebo-treated stallions had 5 mL of PBS intratesticular injected. All stallions had testicular biopsies collected seven days before and one- and 14-days post-heat stress and were castrated 30 days after testicular insult. Tissue sections were stained with H&E and evaluated for the tubular and luminal diameter, epithelial thickness, seminiferous tubules (STs) integrity, the number of spermatozoa in the STs, and the percent of abnormal STs. Significance was set at P≤0.05. In both groups, testicular heat stress damaged the STs (P<0.05). However, STs' parameters were improved in MSCs-treated stallions compared to placebo-treated stallions 30 days after the testicular insult (P<0.05). In conclusion, the results of the present study suggest that intratesticular MSC therapy provided a therapeutic advantage in rescuing acute TD in stallions. However, further studies are essential to evaluate the benefits of this therapy on semen parameters and stallions with idiopathic TD. [ABSTRACT FROM AUTHOR]

Rats-treated with Acecl showed marked-atrophy of seminiferous-tubules, arrested-spermatogenesis and sperm-deformation, while animals-treated with Zr(IV)-complex showed testicular-histo-architecture improvement back towards healthy-tissues. Regarding molecular-docking, binding affinities of Zr(IV)-complex to anti-oxidative-enzymes; SOD, CAT and pro-apoptotic; P53 higher than corresponding parent-ligand Acecl , giving the opportunity to Zr(IV)-complex to attenuating oxidative-damages (testicular-tissue-atrophy) caused by Acecl. [Display omitted] • Synthesis of new mixed complexes of Cu(II), Zr(IV), La(III), Th(IV) and U(VI) with Acecl in presence of Gly. • Characterization for Acecl , Gly and their metal-complexes with different techniques. • Acecl -treated rats showed higher levels of oxidants with inhibition of anti-oxidants activities. • Acecl -treatment triggered testicular cell apoptosis by rising P53 and lowering BCL2 proteins. • Acecl-metal complexes ameliorate testicular oxidative toxicity back towards healthy rats. New nano metal complexes of Cu(II), Zr(IV), La(III), Th(IV) and U(VI) ions using Acecl as primary ligand and glycine (Gly) as secondary ligand were synthesized and characterized. Physico-chemical and spectroscopic techniques namely, elemental (CHN), molar-conductance (Λ), magnetic susceptibility (µeff), ultraviolet–visible (UV–Vis.), Fourier-transform-infrared (FT-IR), proton-nuclear-magnetic-resonance (1H NMR), x-ray-diffraction (XRD), mass-spectrometry (m/z), and thermal-analyses (TG-DTG/DTA), were utilized to determine the binding mode and composition of the complexes. FT-IR spectra revealed that Acecl chelated as negative bidentate mode to the metal ions via oxygen of carboxylic and keto groups while, Gly chelated as negative bidentate via nitrogen of amino group and deprotonated carboxylic oxygen. Λ values revealed that La(III) and Th(IV) complexes were 1:1 and 1:2 electrolytes. XRD designated the formation of metal complexes, in which their structures changed from ligands due to presence of metal ions and all complexes found as crystalline in phase. Forty-two male-albino-rats categorized into seven groups, comprising healthy, Acecl treated and its metal complexes groups were examined. Treatment with Acecl showed severe testicular-toxicity, manifested by severe atrophy in seminiferous tubules, arrested spermatogenesis and negatively-altered semen parameters. Such oxidative-damages recognized by elevated oxidants levels, reduced anti-oxidant activities and up-regulated testicular cell pro-apoptotic-P53. Contrarily, metals complexation, particularly Zr(IV) complex, protected testes from Acecl -induced testicular-apoptosis through increasing pro-survival-BCL2, lowering pro-apoptotic-P53, reducing oxidant levels and stimulating anti-oxidants activities. Consequently, marked improvements in testicular histo-architecture, spermatognial maturation and sperm characteristics towards the healthy were noticed. Regarding molecular docking, Zr(IV)-complex experienced the best binding affinities with superoxide-dismutase, catalase and P-53 targets. ADME-Tox prediction of novel metal complexes revealed their acceptable drug-likeness properties. [ABSTRACT FROM AUTHOR]