Generation of transgenic birds can be achieved by temporal suppression of endogenous spermatogenesis in males prior to primordial germ cell implantation. One of many established methods to induce male sterility is the intraperitoneal injection of busulfan, an alkylating agent. Nevertheless, the use of busulfan injections, which may also affect hematopoietic stem cells, carries the risk of potential lethality in animals. Given their safety and non-toxic nature, it has been demonstrated that intratesticular busulfan injections in mammals are less effective than intraperitoneal injections. This study aimed to compare, for the first time, the sterility and toxicity effects of intraperitoneal vs. intratesticular busulfan injections in quail and chickens. Our experimental design involved a previously established single intraperitoneal busulfan injection of 40 mg/kg of body weight (BW). In quail, busulfan was then administered intratesticularly at 3 different concentrations (6, 12, and 20 mg/kg BW), while in chickens, the working concentration was 20 mg/kg BW. We found that a single intraperitoneal busulfan injection of 40 mg/kg of BW resulted in 100% mortality in the treated roosters. In quails, however, this concentration only caused a temporary suppression of fertility for a 15-d period. Moreover, we found that a higher dose of intratesticular injection of busulfan is required to suppress spermatogenesis in quail (20 mg/kg BW) compared to mammals (4 mg/kg BW). Following these findings, we further confirmed that intratesticular injection of 20 mg/kg BW busulfan into roosters did not affect their overall viability. However, it induced a temporary state of male sterility, consistent with the effects observed with intraperitoneal injections. Hence, our data demonstrate that quail and chicken respond differently to busulfan administration. Furthermore, the present study provides evidence that direct injection into the rooster testes causes less physiological stress than intraperitoneal injection.

BACKGROUND: Toxoplasma gondii is an intracellular protozoan parasite that is widely distributed in humans and warm-blooded animals. T. gondii chronic infections can cause toxoplasmic encephalopathy, adverse pregnancy, and male reproductive disorders. In male reproduction, the main function of the testis is to provide a stable place for spermatogenesis and immunological protection. The disorders affecting testis tissue encompass abnormalities in the germ cell cycle, spermatogenic retardation, or complete cessation of sperm development. However, the mechanisms of interaction between T. gondii and the reproductive system is unclear. The aims were to study the expression levels of genes related to spermatogenesis, following T. gondii infection, in mouse testicular tissue.
METHODS: RNA-seq sequencing was carried out on mouse testicular tissues from mice infected or uninfected with the T. gondii type II Prugniaud (PRU) strain and validated in combination with real-time quantitative PCR and immunofluorescence assays.
RESULTS: The results showed that there were 250 significant differentially expressed genes (DEGs) (P < 0.05, |log2fold change| ≧ 1). Bioinformatics analysis showed that 101 DEGs were annotated to the 1696 gene ontology (GO) term. While there was a higher number of DEGs in the biological process classification as a whole, the GO enrichment revealed a significant presence of DEGs in the cellular component classification. The Arhgap18 and Syne1 genes undergo regulatory changes following T. gondii infection, and both were involved in shaping the cytoskeleton of the blood-testis barrier (BTB). The number of DEGs enriched in the MAPK signaling pathway, the ERK1/2 signaling pathway, and the JNK signaling pathway were significant. The PTGDS gene is located in the Arachidonic acid metabolism pathway, which plays an important role in the formation and maintenance of BTB in the testis. The expression of PTGDS is downregulated subsequent to T. gondii infection, potentially exerting deleterious effects on the integrity of the BTB and the spermatogenic microenvironment within the testes.
CONCLUSIONS: Overall, our research provides in-depth insights into how chronic T. gondii infection might affect testicular tissue and potentially impact male fertility. These findings offer a new perspective on the impact of T. gondii infection on the male reproductive system.

UNASSIGNED: With the progress and prevalence of COVID-19, concerns have arisen regarding its impact on men\'s sexual health. Therefore, this study was conducted with the aim of examining the effects of COVID-19 on serum levels of sex hormones and semen.
UNASSIGNED: Sixty participants who met the study inclusion criteria enrolled in this study between January and April 2022. The individuals were divided into three groups (n = 20): healthy, COVID-19 positive, and recovered from COVID-19. Blood and semen samples were collected from the participants. Serum levels of sex hormones and semen were evaluated both macroscopically and microscopically.
UNASSIGNED: Our study results showed that the most common symptoms observed in the COVID-19 group were cough (100 %), fever (100 %), fatigue (95 %), and runny nose (90 %). Serum levels of sex hormones (testosterone, FSH, and prolactin) in the COVID-19 group were significantly decreased compared to the healthy group. Microscopic examination of semen revealed significant differences in vitality, progressive, and motile parameters among the three groups, with a decrease observed in the COVID-19 group.
UNASSIGNED: These results indicate that COVID-19 may have a negative impact on men\'s sexual health, potentially affecting hormone production and sperm quality. Further research is needed to determine the long-term effects of COVID-19 on male fertility and to explore potential treatment options.

OBJECTIVE: The current study aimed to explore the potential protective effect of Passiflora Incarnata L., (PI) in treating IR injury after testicular torsion in rats.
METHODS: This research investigated the impact of PI on IR damage in male Wistar albino rats. Animals were divided to three groups: group 1 (sham), group 2 (IR), and group 3 (IR+PI).
RESULTS: The malondialdehyde (MDA), myeloperoxidase (MPO) and glutathione (GSH) levels did not significantly differ across the groups (p = 0.830, p = 0.153 and p=0.140, respectively). However, Group 3 demonstrated a superior total antioxidant status (TAS) value compared to Group 2 (p = 0.020). Concurrently, Group 3 presented a significantly diminished mean total oxidant status (TOS) relative to Group 2 (p = 0.009). Furthermore, Group 3 showed a markedly improved Johnsen score relative to Group 2 (p < 0.01). IR caused cell degeneration, apoptosis, and fibrosis in testicular tissues. PI treatment, however, mitigated these effects, preserved seminiferous tubule integrity and promoted regular spermatogenesis. Furthermore, it reduced expression of tumor necrosis factor-alpha (TNF-α), Bax, and Annexin V, signifying diminished inflammation and apoptosis, thereby supporting cell survival (p < 0.01, p < 0.01, p < 0.01, respectively).
CONCLUSIONS: This study revealed that PI significantly reduces oxidative stress and testicular damage, potentially benefiting therapies for IR injuries.
OBJECTIVE: Explorar el posible efecto protector de Passiflora incarnata L. (PI) en el tratamiento de la lesión por isquemia-reperfusión (IR) después de una torsión testicular en ratas.
UNASSIGNED: Se estudió el impacto de Passiflora incarnata en el daño por IR en ratas Wistar albinas machos. Los animales se dividieron tres grupos: 1 (simulado), 2 (IR) y 3 (IR+PI).
RESULTS: Los niveles de malondialdehyde (MDA), myeloperoxidase (MPO) y glutathione (GSH) no difirieron significativamente entre los grupos (p = 0.830, p = 0.153 y p = 0.140, respectivamente). Sin embargo, el grupo 3 tuvo un valor de estado antioxidante total (TAS) superior en comparación con el grupo 2 (p = 0.020). Al mismo tiempo, el grupo 3 presentó un estado oxidante total (TOS) medio significativamente disminuido en comparación con el grupo 2 (p = 0.009). El grupo 3 mostró una mejora notable en la puntuación de Johnsen en comparación con el grupo 2 (p < 0.01). La IR causó degeneración celular, apoptosis y fibrosis en los tejidos testiculares. El tratamiento con PI mitigó estos efectos, preservó la integridad de los túbulos seminíferos y promovió la espermatogénesis regular. Además, redujo la expresión de factor de necrosis tumoral alfa, Bax y anexina V, lo que significa una disminución de la inflamación y de la apoptosis, respaldando así la supervivencia celular (p < 0.01, p < 0.01 y p < 0.01, respectivamente).
CONCLUSIONS: Este estudio reveló que PI reduce significativamente el estrés oxidativo y el daño testicular, beneficiando potencialmente las terapias para lesiones por IR.

The current study aimed to investigate the sensitivity of male testis parenchyma cells to chemotherapy agents and the protective effects and mechanisms of Morinda citrifolia (Noni) administration against structural and functional changes before and after chemotherapy (Paclitaxel (PTX)). For this purpose, rats were randomly assigned into four groups (Control = G1, PTX 5 mg/kg = G2; PTX + Noni 10 mg/kg = G3, PTX + Noni 20 mg/kg = G4). PTX was injected intraperitoneally for 4 consecutive weeks, at a dose of 5 mg/kg to all groups except the control group. Then noni was administrated in 10 (G3) and 20 (G4) mg/kg groups orally (gavage) for 14 days. Biochemical analyses, Real-Time Polymerase Chain Reaction (PCR), and immunohistochemical analyses were performed. According to our results, Total Oxidative Stress (TOS) and Malondialdehyde (MDA) were significantly increased in the PTX group (P < 0.01). Superoxide Dismutase (SOD) enzyme activity and Total Antioxidant Capacity (TAC) levels were decreased (P < 0.01). The changes in the rats treated with PTX + Noni 20 mg/kg were noteworthy. The increased levels of IL1-β (Interleukin 1 beta) and TNFα (tumor necrosis factor-alpha) with PTX were down-regulated after treatment with PTX + Noni 20 mg/kg (P < 0.01) (9 % and 5 % respectively). In addition, Noni restored the testicular histopathological structure by reducing caspase-3 expression and significantly (61 %) suppressed oxidative DNA damage and apoptosis (by regulating the Bax (bcl-2-like protein 4)/Bcl-2 (B-cell lymphoma gene-2) ratio). In conclusion, Noni reduced cellular apoptosis and drastically changed Caspase 8 and Bax/Bcl-2 levels. Furthermore, it considerably decreases oxidative damage and can be used in testicular degeneration.

OBJECTIVE: The study aimed to comprehensively analyze testosterone and precursor concentrations in the testicular interstitial fluid (TIF) of men with azoospermia, exploring their significance in the testicular microenvironment and their correlation with testicular sperm retrieval outcomes.
METHODS: We analyzed 37 TIF samples, including 5 from men with obstructive azoospermia (OA) and 32 from men with non-obstructive azoospermia (NOA). Liquid chromatography with tandem mass spectrometry quantified testosterone and precursor levels. Comparative assessments of the outcomes of testicular sperm retrieval were performed between the OA and NOA groups as well as among men with NOA.
RESULTS: Men with NOA who had not undergone hormone treatment exhibited significantly higher intratesticular concentrations of testosterone (median 1,528.1 vs. 207.5 ng/mL), androstenedione (median 10.6 vs. 1.9 ng/mL), and 17-OH progesterone (median 13.0 vs. 1.8 ng/mL) than men diagnosed with OA. Notably, in the subgroup of patients with NOA subjected to medical treatment, men with successful sperm retrieval had significantly reduced levels of androstenedione (median androstenedione 5.7 vs. 18.5 ng/mL, p=0.004). Upon a more detailed analysis of these men who underwent hormone manipulation treatment, the testosterone/androstenedione ratio (indicative of HSD17B3 enzyme activity) was markedly increased in men with successful sperm retrieval (median: 365.8 vs. 165.0, p=0.008) compared with individuals with NOA who had unsuccessful sperm recovery. Furthermore, within the subset of men with NOA who did not undergo medical treatment before microdissection testicular sperm extraction but achieved successful sperm retrieval, the ratio of 17-OH progesterone/progesterone (indicative of CYP17A1 activity) was substantially higher.
CONCLUSIONS: The study suggests distinct testosterone biosynthesis pathways in men with compromised spermatogenesis and those with normal spermatogenesis. Among NOA men with successful retrieval after hormone optimization therapy, there was decreased androstenedione and increased HSD17B3 enzyme activity. These findings have diagnostic and therapeutic implications for the future.

UNASSIGNED: Cryopreservation of semen can give full play to the reproductive advantages of male animals. However, in actual production, due to the poor frost resistance of sheep semen and the low conception rate, the promotion of sheep frozen semen is greatly hindered. Therefore, it is urgent to improve the frost resistance of semen to improve the quality of frozen semen. At present, most studies on improving the quality of frozen semen are based on the improvement of semen dilutions, and few studies on improving the freezing resistance of ram semen by feeding functional amino acids.
UNASSIGNED: Therefore, 24 Turpan black rams were divided into high antifreeze group (HF) and a low antifreeze group (LF) Each of these groups was further randomly divided into control and experimental subgroups. The control subgroup was fed a basal diet, while the experimental subgroup received an additional 12 g/d of L-Cit supplementation based on the control group for a duration of 90 days.
UNASSIGNED: The results showed that Following L-Cit supplementation, the experimental group demonstrated significantly elevated sperm density and VSL (Velocity of straight line), T-AOC, GSH-Px, and NO levels in fresh semen compared to the control group (P < 0.01). After thawing, the experimental group exhibited significantly higher levels of T-AOC, GSH-Px, and NO compared to the control group (P < 0.01). Additionally, the HFT group, after thawing frozen semen, displayed significantly higher HK1 protein expression compared to the control group. The number of spermatogonia, spermatocytes, and sperm cells in the HFT group was significantly higher than that in the HFC group. Moreover, 16S rRNA sequence analysis showed that Candidatus_Saccharimonas, Staphylococcus, Weissella, succinivbrionaceae_UcG_002, and Quinella were significantly enriched in the rumen of the HFT group, while Ureaplasma was significantly enriched in the HFC group. In the duodenum, Clostridiales_bacterium_Firm_14, Butyrivibrio, and Prevotellaceae_NK3831_group were significantly enriched in the HFT group, whereas Desulfovibrio and Quinella were significantly enriched in the HFC group.
UNASSIGNED: Under the conditions employed in this study, L-Cit supplementation was found to enhance the intestinal flora composition in rams, thereby improving semen quality, enhancing the antifreeze performance of semen, and promoting the development of testicular spermatogenic cells.

Ultrasound of the testes is important in the evaluation of breeding dogs, and recently advanced techniques such as Shear Wave Elastography (SWE) have been developed. This study focused on evaluation of normal testicular stiffness in healthy and fertile male dogs, employing both qualitative (2D-SWE) and quantitative (pSWE, 2D-SWE) techniques. Nineteen dogs of various medium-large breeds aged 3.39 ± 2.15 years, and with a history of successful reproduction were included after clinical, B-mode and Doppler ultrasound of testes and prostate, and semen macro and microscopic evaluations. pSWE involved square regions of interest (ROIs) placed at six different points in the testicular parenchyma, while 2D-SWE depicted stiffness with a color scale ranging from blue (soft) to red (stiff), allowing a subsequent quantification of stiffness by the application of 4 round ROIs. The results showed a mean Shear Wave Speed (SWS) of 2.15 ± 0.39 m/s using pSWE, with lower values above the mediastinum compared to below, and in the center of the testis compared to the cranial and caudal poles. 2D-SWE demonstrated a uniform blue pattern in the parenchyma, and a mean SWS of 1.65 ± 0.15 m/s. No significant differences were found between left and right testes, above and below the mediastinum, or among breeds. No correlations were observed between mean SWS and body condition score, age, testicular and prostatic volume. Weight was positively correlated with mean SWS only by 2D-SWE. By performing semen analysis and enrolling only healthy and fertile adult dogs, we ensured both structural and functional integrity of the testes. This pilot study represents a valuable baseline data for testicular stiffness by both pSWE and 2D-SWE with a Mindray US machine in medium-large sized healthy and fertile dogs, pointing out the potential role of SWE in the non-invasive fertility assessment and management of breeding dogs.

Testosterone, an important sex hormone, regulates sexual maturation, testicular development, spermatogenesis and the maintenance of secondary sexual characteristics in males. Testicular Leydig cells are the primary source of testosterone production in the body. Hezuo pigs, native to the southern part of Gansu, China, are characterized by early sexual maturity, strong disease resistance and roughage tolerance. This study employed type IV collagenase digestion combined with cell sieve filtration to isolate and purify Leydig cells from the testicular tissue of 1-month-old Hezuo pigs. We also preliminarily investigated the functions of these cells. The results indicated that the purity of the isolated and purified Leydig cells was as high as 95%. Immunofluorescence analysis demonstrated that the isolated cells specifically expressed the 3β-hydroxysteroid dehydrogenase antibody. Enzyme-linked immunosorbent assay results showed that the testosterone secretion of the Leydig cells cultured in vitro (generations 5-9) ranged between 1.29-1.67 ng/mL. Additionally, the content of the cellular autophagy signature protein microtubule-associated protein 1 light chain 3 was measured at 230-280 pg/mL. Through this study, we established an in vitro system for the isolation, purification and characterization of testicular Leydig cells from 1-month-old Hezuo pigs, providing a reference for exploring the molecular mechanism behind precocious puberty in Hezuo pigs.

Anemia poses a significant healthcare challenge across different socioeconomic groups and can result in reproductive system damage through the generation of free radicals and lipid peroxidation. This study examines the protective effects of quercetin (QUE) and mirtazapine (MIR) against the reproductive damage caused by phenylhydrazine (PHZ) in mice. Fifty NMRI mice, aged 8-10 weeks with an average weight of 27.0 ± 2.0 g, were randomly divided into five groups. The control group (Group 1) received oral administration of 10 mL/kg/day of normal saline. Group 2 (PHZ group) received an initial intraperitoneal dose of 8 mg/100 g body weight of PHZ, followed by subsequent doses of 6 mg/100 g every 48 h. Group 3 received PHZ along with oral QUE at a dosage of 50 mg/kg/day. Group 4 received PHZ along with oral MIR at a dosage of 30 mg/kg/day. Group 5 received PHZ along with oral QUE at a dosage of 50 mg/kg/day and MIR at a dosage of 30 mg/kg/day. The treatment duration was 35 days. Sperm samples were collected from the caudal region of the epididymis post-euthanasia to assess the total mean sperm count, sperm viability, motility, DNA damage, and morphology. Testicular tissue was employed to quantify total antioxidant capacity (TAC), superoxide dismutase (SOD), glutathione peroxidase (GPx), and malondialdehyde (MDA) concentrations, while serum levels of testosterone, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) were analyzed. Additionally, various aspects, including testicular histopathology, oxidative enzyme levels, gene expression related to apoptosis and antiapoptotic pathways, and in vivo fertility index, were evaluated after 35 days. The QUE, MIR, and QUE + MIR groups showed less abnormal morphology and DNA damage, as well as better total and progressive sperm motility, motility characteristics, viability, and plasma membrane function compared to the PHZ group. QUE, MIR, and QUE + MIR administration increased TAC, SOD, and GPx activities in testicular tissue, while reducing MDA levels compared to the PHZ group. Furthermore, QUE, MIR, and QUE + MIR significantly reduced Bax, and caspase-3 expression levels, and increased Bcl-2 expression levels, compared to the PHZ group. Mice treated with QUE, MIR, and QUE + MIR exhibited an increased in vivo fertility index and plasma sex hormone levels compared to the PHZ group. These results show that QUE, MIR, and QUE + MIR might be able to improve the fertility index, boost the testicular antioxidant defense system, and control the death of germ cells. This could mean that they could be used to treat mice with PHZ-induced testicular damage.