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This study aimed to demonstrate the effect of conventional freezing versus cryoprotectant-free vitrification on the recovery of testicular sperm motility. Testicular samples were obtained from 50 patients with azoospermia for testicular biopsy ± potential sperm storage. We retrieved 100 spermatozoa from each patient divided equally into two straws. They were frozen using conventional freezing as a control group and cryoprotectant-free vitrification in micro-capillary system using open–pulled straws. Seven days later, cryopreserved straws were thawed and assessed in duplicate. The mean sperm motility between the original spermatozoa sample and the post warming sample was reduced after conventional freezing compared to cryoprotectant-free vitrification (4.48 ± 2.09% versus 3.25 ± 1.92%, p < 0.001; 4.48 ± 2.09% vs 3.68 ± 1.93%, p < 0.001, respectively). There was a significant difference between the two methods regarding the mean sperm motility after warming (3.38 ± 1.86% versus 3.76 ± 1.88%, p = 0.015). The mean recovery percent of testicular sperm motility from the original sperm sample was lower (p = 0.02) after conventional freezing compared to cryoprotectant-free vitrification (78.4 ± 28.17% versus 85.37 ± 23.63%). Overall, the rate of post-thaw recovery of human testicular sperm motility improved using cryoprotectant-free vitrification compared to conventional freezing. [ABSTRACT FROM AUTHOR]

Background: Congenital hypothyroidism (CH) is a pathology that affects various organs, including the testicles. The mechanisms by which this condition alters fertility is unknown. This study aimed at determining if experimental CH affects gonocyte differentiation and arrests meiosis; and the possible role of the Sertoli cell (SC) in this condition. Material and Methods: Two groups of rats consisting of Control group and Methimazole (MMI) induced CH rats were formed. The induction of CH was achieved by the administration of MMI starting on day 16 postcoitum and continued until euthanized. Euthanasia was performed at 5, 8, 10, 16 and 64 days of age. Following this, the testicular tissue of each animal was extracted and processed for histopathological and ultrastructural analysis. In addition, the tissue was used for the determination of proteins and their transcriptions, events which are characteristics of gonocyte differentiation. The SC functionality proteins was determined immunohistochemically, while sperm parameters of the cauda epididymis were verified. Results: CH caused a delay in the gonocyte differentiation, and arrested meiosis and spermiogenesis. These events had long-term repercussions on the quality of the seminiferous epithelium. The results show that CH induces alterations in the functional state of SCs that may have led to the deficiency in the synthesis and/or in the release of molecules necessary for gonocyte differentiation; as well as disorders in the process of meiosis that resulted in sperm absence. Conclusion: These results suggest that CH affects gonocyte differentiation and arrests meiosis, possibly through altering the functional status of SCs. [ABSTRACT FROM AUTHOR]

The effect of paternal age on fertility remains unclear. This retrospective study aims to examine the impact of male age on semen parameters and the reproductive outcomes of men admitted to an infertility center over a 9-year period. A total of 8046 patients were included in the study. Men were divided into four age groups. The groups were evaluated for semen parameters and reproductive outcome. The 21–30 year group presented lower sperm concentrations in comparison to those aged 31–40 and 41–50, yet shared a similar concentration to those over 50 years of age. Moreover, grades A and B decreased significantly in men aged over 50 years. The highest progressive motility and normozoospermia were observed in the age group 31–40 years while men over 50 years of age had the highest rates of asthenozoospermia and oligoasthenozoospermia. Furthermore, live birth results were reported in 5583 of the patients who underwent intracytoplasmic sperm injection (ICSI) and were found highest between 31–40 years of age. To our knowledge, this is the largest study in Turkey focusing on male age-related semen parameters and ICSI pregnancy outcomes. The study demonstrates that age is a significant factor for semen quality and live birth. [ABSTRACT FROM AUTHOR]

Background: Methotrexate (MTX) is widely administered to manage various cancers. However, MTX induces spermatogenic defects. Objective: This study investigated the protective effects of betaine (BET) against MTX–induced testicular damage. Materials and Methods: Forty‐eight male mice were randomly divided into six groups: control, BET (300 mg/kg), MTX (20 mg/kg), and MTX (20 mg/kg) + BET (100, 200, and 300 mg/kg) groups. Testosterone levels, histological changes, sperm quality, apoptosis, and oxidative stress biomarkers were assessed to evaluate the protective effects of BET. Results: MTX disrupted germinal epithelium, reduced sperm quality, and decreased serum testosterone levels, as well as induced apoptosis and oxidative stress in the testicular tissue. BET dose dependently restored the testosterone levels and catalase (CAT) and superoxide dismutase (SOD). Furthermore, BET reduced lipid peroxidation, as indicated by decreased malondialdehyde (MDA) levels. BET preserved normal spermatogenesis, improved sperm quality, and reduced histological changes by MTX. Moreover, BET reduced apoptosis by decreasing the Bcl‐2 Associated X‐protein (Bax)/B‐cell lymphoma 2 (Bcl‐2) ratio in the testicular tissue of the MTX‐intoxicated mice. Conclusion: The results indicate that BET mitigates testicular harm triggered by MTX by inhibiting apoptosis and decreasing oxidative stress levels. [ABSTRACT FROM AUTHOR]

During the intracytoplasmic sperm injection (ICSI) procedure, 10% polyvinylpyrrolidone (PVP) is commonly used as a selection medium to decrease sperm motility. The study aimed to determine the effect of different concentrations of PVP (5% and 10%) on sperm apoptotic transcripts, sperm DNA fragmentation (SDF), embryo cytokinetic, and clinical characteristics in the ICSI program. In the study, 60 couples with male factor infertility underwent ICSI insemination using either 5% or 10% PVP concentration. Metaphase II (MII) oocytes were divided into two groups, with one group injected with sperm in 5% PVP and the other in 10% PVP. After fertilization, the zygotes were cultured using a time‐lapse microscope to assess morphokinetic parameters also, simultaneously with the ICSI procedure, SDF and apoptotic transcript levels were quantified using reverse transcription quantitative polymerase chain reaction (qPCR). The study found that DNA fragmentation was lower in sperm from the 5% PVP group compared to the 10% PVP group. Additionally, the expression levels of B‐cell lymphoma (BCL2) and the 70 kilodalton heat shock proteins (HSP70) genes were significantly higher in 5% PVP compared to 10% PVP, while the transcript levels of the Bcl‐2‐associated X protein (BAX) gene were lower in 5% PVP. The 5% PVP group also demonstrated higher fertilization and high‐quality embryo formation rates. Cytokinetic variables were significantly different between the two groups, and the rates of fragmentation, uneven blastomere, reverse cleavage (RC), and embryo arrest were higher in the 10% PVP group. The study concluded that a low concentration of PVP (5%) is a suitable replacement for 10% PVP in ICSI cycles for male factor infertility. [ABSTRACT FROM AUTHOR]

Previous studies have shown beneficial effects of empagliflozin (Empa), a selective inhibitor of the sodium-glucose cotransporter 2 (SGLT2), on diabetes and cardiovascular outcomes in patients with diabetes. However, whether Empa could ameliorate diabetes mellitus (DM)-induced male spermatogenesis dysfunction remains unclear. Our study aimed to investigate the effect of Empa in the development of DM-induced male spermatogenesis dysfunction and to reveal the molecular mechanisms. DM mice were orally treated with Empa to investigate the effects of Empa on DM-induced male mice spermatogenesis dysfunction. We employed a cardiac-specific C1q/tumor necrosis factor-related protein 9 (CTRP9)-deficient mouse model and a cardiac-specific CTRP9 overexpression mouse model to investigate its role in the protection of Empa against diabetes-induced male subfertility. We found that Empa treatment could improve DM-induced male mice subfertility. Interestingly, we discovered that cardiac-derived CTRP9 was decreased in DM mice and this decrease was prevented by Empa treatment. A CTRP9 blocking antibody or cardiac-specific depletion of CTRP9 abolished the protection of Empa on DM-induced male subfertility. Cardiac-specific CTRP9 overexpression ameliorated DM-induced male subfertility. Mechanistically, we identified that cardiac-derived CTRP9 increased steroidogenesis in mice with diabetes in a PKA-dependent manner. We also provided direct evidence that activation of AMP activated protein kinase α (AMPKα)/nuclear factor (erythroid-derived 2)-like 2 (Nrf2) signalling pathway by CTRP9 was responsible for the attenuation of ferroptosis in Leydig cells. In conclusions, we supposed that Empa was a potential therapeutic agent against DM-induced male mice spermatogenesis dysfunction. [ABSTRACT FROM AUTHOR]

Aim: In approximately 20% of infertile couples, the male factor is the only main cause. In recent years, it has been hypothesized that sperm DNA integrity may be a better indicator than routine semen analysis. Although the effects of smoking on male infertility have not yet been proven, smoking is considered a reasonable risk factor for infertility. In our study, we aimed to evaluate DNA damage in normospermic infertile men who smoke and do not smoke using the acridine orange method. Material and Methods: This study was conducted on semen samples from 50 male cases, 25 of whom were smokers and non-smokers, who met the infertility criteria and applied to the urology clinic of Dicle University Medical Faculty Hospital. Microscopic examinations and morphological evaluations were performed in accordance with the WHO 2010 criteria. The acridine orange method was applied to evaluate DNA fragmentation and DNA fragmentations were calculated. Statistical analysis of the obtained data was performed using the Mann-Whitney U Test. Results: In our study, in the smoking group, age was significantly associated with the duration of infertility, concentration with total motile sperm count, total sperm count with total motile sperm count, and amorphous head anomaly (p<0.01). In the non-smoking group, age and infertility were significantly associated with the duration, volume and total count, concentration and total count, total motile sperm count, and DNA fragmentation (p<0.01). However, there was no statistical difference between the smoking and non-smoking groups in terms of semen parameters and DNA fragmentation (p>0.05). Conclusion: In order to obtain more reliable results, it is thought that large-scale studies are needed in the same groups and in the general population. [ABSTRACT FROM AUTHOR]

Nowadays, developing countries have seen a reduction in male reproductive parameters, and it has been linked to the exposure of endocrine disrupting chemicals (EDCs), which are able to mimic or disrupt steroid hormone actions. Also, nanoparticles have shown effects on the male reproductive system, in particular the use of TiO2-NPs in drugs, cosmetics, and food as pigment additives, and, thanks to their small size (1–100 nm), provide themselves the opportunity to be internalized by the body and pass the blood–testis barrier (BTB). Therefore, TiO2-NPs can act on spermatogenesis and spermatozoa. In this study, we carried out an in vitro assay on human spermatozoa to evaluate the effects of TiO2-NPs at the concentrations of 500, 250, 100, and 50 ppm. Exposure did not statistically alter sperm parameters (e.g., motility and viability) but induced damage to sperm DNA and the expression of biomarkers by spermatozoa. This immunofluorescence investigation showed a positivity for biomarkers of stress (HSP70 and MTs) on the connecting piece of spermatozoa and also for sex hormone binding globulin (SHBG) biomarkers. The SHBG protein acts as a carrier of androgens and estrogens, regulating their bioavailability; therefore, its expression in the in vitro assay did not rule out the ability of TiO2-NPs to act as endocrine disruptors. [ABSTRACT FROM AUTHOR]

OBJECTIVE To investigate the gastrointestinal (GI) and reproductive risk associated with chronic usage of celecoxib in male albino rats. METHOD Thirty male albino rats weighing 200-230 g were randomly divided into three equal groups of 10 rats each. In group 1, the control group, rats received no drug. In group 2, rats received celecoxib (50 mg/kg/day, orally) for 30 successive days. In group 3, rats received celecoxib (50 mg/kg/day, orally), and royal jelly (300 mg/kg/day, orally) for 30 successive days. Gastric ulcer scoring and semen analysis were employed to elucidate the celecoxib adverse effects on gastric and testicular tissue functions. RESULTSCelecoxib produced no gastric ulcer in both groups 2 and 3. Celecoxib has no appreciable effect on steroidogenesis (testosterone biosynthesis in Leydig cells of the testis) or spermatogenesis (sperms production from germ cells in the seminiferous tubules of the testis), as indicated by normal sperm count, motility, abnormalities, and normal serum testosterone levels. Royal jelly co-administration did not cause any significant change in serum testosterone level, sperm count, and sperm motility percentage, but caused a significant decline in sperm abnormalities level. CONCLUSIONS Upper GI tolerability and reproductive safety were proved in the chronic use of celecoxib in male albino rats, with ameliorative effects of royal jelly against celecoxib-induced oxidative and apoptotic stress. [ABSTRACT FROM AUTHOR]

The ovarian microenvironment plays a crucial role in ensuring the reproductive success of viviparous teleosts. However, themolecular mechanism underlying the interaction between spermatozoa and the ovarian microenvironment has remained elusive. This study aimed to contribute to a better understanding of this process in black rockfish (Sebastes schlegelii) using integrated multi-omics approaches. The results demonstrated significant upregulation of ovarian complement-related proteins and pattern recognition receptors, along with remodeling of glycans on the surface of spermatozoa at the early spermatozoa-storage stage (1 month after mating). As spermatozoa were stored over time, ovarian complement proteins were progressively repressed by tryptophan and hippurate, indicating a remarkable adaptation of spermatozoa to the ovarian microenvironment. Before fertilization, a notable upregulation of cellular junction proteins was observed. The study revealed that spermatozoa bind to ZPB2a protein through GSTM3 and that ZPB2a promotes spermatozoa survival and movement in a GSTM3-dependent manner. These findings shed light on a key mechanism that influences the dynamics of spermatozoa in the female reproductive tract, providing valuable insights into the molecular networks regulating spermatozoa adaptation and survival in species with internal fertilization. [ABSTRACT FROM AUTHOR]

The study was designed to determine the effect of Vitamin E (α-tocopherol) supplimentation into BIOXcellTM and Tris based egg yolk extender on post-thawed quality of Tharparkar cattle bull semen. The research was carried out at Department of Animal Reproduction, Faculty of Animal Husbandry and Veterinary Sciences, Sindh Agricultural University Tandojam. In this study four fertile Tharparkar bulls (A, B, C and D) having age of 4-5 years were used. A total of 52 (n=13) ejaculates were collected with the help of artificial vagina, after the collection macroscopic (color, volume and pH) and microscopic (motility, morphology, viability, concentration and membrane integrity) parameters were observed. The samples having motility, morphology, viability and membrane integrity =70% were pooled and processed. Pooled semen samples were divided into four groups and diluted with Tris, Tris+Vitamin E(a-tocopherol) and BIOXcellTM, BIOXcellTM+ Vitamin E (a-tocopherol). Post-thawed assessment of motility, morphology, viability and membrane integrity with BIOXcellTM+ Vitamin E(a-tocopherol) (58.31±0.86, 76.22±1.04, 71.27±1.44, 64.68±1.43) showed improved quality parameters as compared to Tris+ Vitamin E(α-Tocopherol) (48.68±0.68, 64.45±1.54, 66.22±1.75, 58.25±1.08), BIOXcellTM (45.72±0.53, 67.45±0.84, 64.72±1.67, 56.63±1.39) and Tris (43.54±0.49, 64.68±0.80, 65.18±1.33, 54.41±1.36). On the basis of in vitro results BIOXcellTM+ Vitamin E(a-tocopherol) showed improved post-thawed quality parameters and it was used for artificial insemination. A total of 20 animals were synchronized at Bhens colony; Ghumanrabad with one injection prostaglandin (PGF2a) protocol and 13 animals showed estrus and were inseminated. Meanwhile, 8 were found pregnant through rectal palpation with a pregnancy rate of 61.53%. [ABSTRACT FROM AUTHOR]

This study aims to evaluate the genomic effects of vitamin D (VitD) on steroidogenesis, sperm structure, and function in mice models with artificially induced vitamin D deficiency (VDD), while maintaining calcium and phosphorus homeostasis. qRT‐PCR was utilized to measure the steady‐state RNA levels of key steroidogenic genes, including CYP11A1, CYP19A1, 17βHSDH1, 3βHSDV1, and CYP24A1, involved in VitD metabolism, in testicular extracts from mouse models with varying degrees of VDD severity. Estradiol, testosterone, luteinizing hormone (LH), and follicle‐stimulating hormone (FSH) levels were also assessed. Furthermore, testicular histomorphometric analysis and evaluation of sperm structural and functional parameters were conducted. The study reveals a linear decrease in the expression of monitored steroidogenic genes with increasing severity of VDD. Moreover, in cases of mild and moderate VDD, testicular extracts exhibited significantly reduced levels of LH, estradiol, and testosterone. VDD has substantial effects on testicular steroidogenesis, which directly impacts spermatogenesis, independent of its established role in calcium regulation. Consequently, the reduction in steroidogenesis due to VDD contributes to the diminished efficiency of testicular spermatogenesis in individuals experiencing VDD, a prevalent condition worldwide. [ABSTRACT FROM AUTHOR]

Simple Summary: In recent decades, the practice of artificial insemination has become increasingly prevalent in canine reproduction, thereby underscoring the necessity for improved semen quality assessment routines. While the bacterial microflora of the reproductive system of female dogs has been extensively studied, there is comparatively limited analysis of the male dog's reproductive tract's microbiota. In the field of andrology, a few connections have already been discovered between seminal bacteria and semen's qualitative parameters. The objective of this study was to determine whether the bacteria present in semen samples from breeding dogs can influence the semen's characteristics. The results indicated that multiple bacterial species present in semen samples from breeding dogs are associated with inferior sperm parameters. Moreover, a greater bacterial load was observed to be related with poorer semen quality in breeding dogs. Assessing canine semen quality helps to detect infertility in males, but identifying factors that influence canine semen quality is a complicated task. The objective of this study was the assessment of the potential influence of bacteria found in canine semen samples on the characteristics of dogs' semen. In this study, semen samples were collected manually from 30 dogs and subjected to a comprehensive examination. The results of sperm motility, concentration, viability, and morphology were statistically analysed in relation to the number of bacteria in the semen (CFUs/mL) and the seminal microbiota. Samples with an increased bacterial count per millilitre were associated with lower-quality sperm motility (p < 0.05). The most frequently isolated bacterial genera from the analysed semen samples were Staphylococcus spp. (26.0%), Corynebacterium spp. (17.8%), and Streptococcus spp. (16.4%). The presence of β-haemolytic Escherichia coli bacteria was linked to suboptimal semen samples, characterised by significantly reduced semen viability and a lower proportion of morphologically normal spermatozoa (p < 0.05). Corynebacterium spp. was associated with reduced bacterial load and superior semen quality (p < 0.01). These findings highlight the importance of bacterial cell counts and microbiota diversity in relation to various factors influencing canine semen quality, providing a more comprehensive understanding of canine reproductive well-being. [ABSTRACT FROM AUTHOR]

This study investigates the impact of mulberry leaf extract on the viability, plasma membrane integrity, and motility of spermatozoa from male white rats (Rattus norvegicus) exposed to e-cigarette smoke. A total of twenty-five male rats were divided into five groups: negative control (NC), positive control (PC), T1, T2, and T3. All groups, except for the NC group, were exposed to e-cigarette smoke. Rats in the T1, T2, and T3 groups received mulberry leaf extract in doses of 100, 200, and 400 mg/kg bw, respectively, while the NC and PC groups were given a placebo of 1% Na-CMC. Both the mulberry leaf extract and the placebo were administered daily, beginning three days prior to the start of e-cigarette smoke exposure, which lasted for 28 days. Results showed that spermatozoa motility, plasma membrane integrity, and viability in the experimental groups were significantly lower than those in the NC group (p <0.05). Conversely, rats in the T1, T2, and T3 groups that received mulberry leaf extract demonstrated significantly greater spermatozoa viability, plasma membrane integrity, and motility compared to the PC group (p <0.05). The T3 group exhibited the most pronounced improvements, with significantly enhanced spermatozoa viability, membrane integrity, and motility (p <0.05) relative to the PC group. These results indicate that mulberry (Morus alba L.) leaf extract enhanced spermatozoa viability, plasma membrane integrity, and motility in white rats (Rattus norvegicus) subjected to e-cigarette smoke. [ABSTRACT FROM AUTHOR]

Globozoospermia is a form of male infertility characterized by spermatozoa with spherical heads lacking acrosomes. The aim of this study was to evaluate ultrastructural and molecular defects in different types of globozoospermia. Semen samples from 12 infertile patients (9 with complete globozoospermia and 3 with partial globozoospermia) and 10 normozoospermic men (control) were examined by transmission electron microscopy and immunocytochemistry with antibodies against lamin B1. The presence of lamin A and progerin was assessed by reverse transcription-PCR. Whole exome sequencing was performed in three patients. In semen sampleswith complete and partial globozoospermia, lamin B1 was observed at the periphery of sperm nuclei, whereas lamin A and progerin were absent. Nuclear envelope pores were found in spermatozoa from both patient groups, regardless of morphology and chromatin condensation, in contrast to the control group. Non-condensed chromatin was present in 51%-81% of cases of complete globozoospermia and in 36%-79% of cases of partial globozoospermia. Homozygous DPY19L2 and SPATA16 variants were identified in two patients with partial globozoospermia and one patient with complete globozoospermia. An atypical nuclear membrane with abnormal nuclear pore distribution and lamin B1 localization was observed in spermatozoa from patients with both complete and partial globozoospermia. The genetic defects in the DPY19L2 and SPATA16 genes detected in patients from both globozoospermic groups suggest a generalized disruption of nuclear structure in globozoospermia, highlighting the genetic and phenotypic similarities between complete and partial globozoospermia. [ABSTRACT FROM AUTHOR]

Oxidative stress (OS) and disrupted antioxidant defense mechanisms play a pivotal role in the etiology of male infertility. The alterations in reactive oxygen species (ROS) production and calcium (Ca2+) homeostasis are the main activators for the mitochondrial permeability transition pore (mPTP) opening. The mPTP opening is one of the main mechanisms involved in mitochondrial dysfunction in spermatozoa. This alteration in mitochondrial function adversely affects energy supply, sperm motility, and fertilizing capacity and contributes to the development of male infertility. In human spermatozoa, the mPTP opening has been associated with ionomycin-induced endogenous oxidative stress and peroxynitrite-induced nitrosative stress; however, the effect of exogenous oxidative stress on mPTP opening in sperm has not been evaluated. The aim of this study was to determine the effect of exogenous oxidative stress induced by hydrogen peroxide (H2O2) on mPTP opening, mitochondrial function, motility, and cell death markers in human spermatozoa. Human spermatozoa were incubated with 3 mmol/L of H2O2 for 60 min, and intracellular Ca2+ concentration, mPTP opening, mitochondrial membrane potential (ΔΨm), ATP levels, mitochondrial reactive oxygen species (mROS) production, phosphatidylserine (PS) externalization, DNA fragmentation, viability, and sperm motility were evaluated. H2O2-induced exogenous oxidative stress caused increased intracellular Ca2+, leading to subsequent mPTP opening and alteration of mitochondrial function, characterized by ΔΨm dissipation, decreased ATP levels, increased mROS production, and the subsequent alteration of sperm motility. Furthermore, H2O2-induced opening of mPTP was associated with the expression of apoptotic cell death markers including PS externalization and DNA fragmentation. These results highlight the role of exogenous oxidative stress in causing mitochondrial dysfunction, deterioration of sperm motility, and an increase in apoptotic cell death markers, including PS externalization and DNA fragmentation, through the mPTP opening. This study yielded new knowledge regarding the effects of this type of stress on mitochondrial function and specifically on mPTP opening, factors that can contribute to the development of male infertility, considering that the role of mPTP in mitochondrial dysfunction in human sperm is not completely elucidated. Therefore, these findings are relevant to understanding male infertility and may provide an in vitro model for further research aimed at improving human sperm quality. [ABSTRACT FROM AUTHOR]

The integrity of the deoxyribonucleic acid (DNA) in sperm plays a crucial role in the fertilization process and embryonic development. This study aimed to evaluate the level of the DNA fragmentation of boar sperm in tris-citrate-fructose extender preserved at temperatures between 16-18°C for 3 days. The level of sperm DNA damage was analysed using toluidine blue (TB) staining. Pig semen collected from 4 male pigs aged 1.3-1.8 years was used as the research sample. Semen collected was conducted twice a week. The experimental method using a Completely Randomized Design with 4 storage duration groups as follows: Group I (0 hours of storage), Group II (24 hours of storage), Group III (48 hours of storage), and Group IV (72 hours of storage), each with 5 replications. The data were analysed using analysis of variance (ANOVA) and, if significant differences were found, Duncan's test was performed. The results of the evaluation of the percentage of DNA damage during the storage process at 16-180C at 0 hours, 24 hours, 48 hours and 72 hours was: 0.33 ± 1.66%, 0.80 ± 0.447%, 1.60 ± 1.14% and 2.80 ± 0.447%, respectively. These results demonstrate that liquid boar semen preserved with tris citrate fructose extender at 16-18 °C for 3 days has spermatozoa DNA damage levels within the normal range, making it suitable for artificial insemination and ensuring optimal AI success. [ABSTRACT FROM AUTHOR]

Background: Single sperm cryopreservation (SSC) is a specific technique especially used in individuals with small numbers of sperm who suffered from non-obstructive azoospermia (NOA). Testicular specimens possess poor motility and low population of viable spermatozoa. Therefore, sperm selection methods such as applying pentoxifylline (PTX) may improve motility in these cases. The main aim of this study was to evaluate the protective effects of PTX on testicular spermatozoa before and after performing SSC. Methods: Thirty testicular samples were obtained from men with azoospermia. This study was conducted in two phases. Phase 1 evaluated the effect of PTX for sperm selection before SSC. Twenty testicular samples were divided to two experimental groups: SSC without (I) and with PTX treatment (II). For PTX treatment spermatozoa were incubated with PTX at 37°C for 30 min and only motile spermatozoa were selected for SSC. In phase 2, ten testicular samples were cryopreserved with SSC and warming procedure was carried out in droplet with and without PTX. Motility and viability rates, morphology by motile sperm organelle morphology examination (MSOME), DNA fragmentation by sperm chromatin dispersion test (SCD) and mitochondrial membrane potential (MMP) were evaluated. Results: In phase 1, post warm motility rate was higher in PTX exposed group compared to the unexposed group (25.6 ± 8.13 vs. 0.85 ± 2.1) (p > 0.00). Recovery rate, viability and morphology were not significantly different between groups. DNA integrity and MMP were also similar between both groups. In phase 2 although motility increased in PTX group compared to without PTX group (29.30 ± 12.73 vs. 1.90 ± 2.64) (p > 0.00), the viability rate was not different (70.40 ± 12.12 vs. 65.30 ± 11.87). All above mentioned parameters were similar between the two SSC groups. Conclusions: Supplementation of testicular spermatozoa with PTX before cryopreservation increases motility and did not have adverse effects on viability, morphology, DNA integrity and MMP. PTX could be used as sperm selection method before single sperm cryopreservation, but PTX could not maintain motile the most of viable testicular sperms. [ABSTRACT FROM AUTHOR]

Type 2 diabetes mellitus (T2DM) is a risk factor for male infertility, but the underlying molecular mechanisms remain unclear. Advanced glycation end products (AGEs) are pathogenic molecules for diabetic vascular complications. Here, we investigated the effects of the DNA aptamer raised against AGEs (AGE-Apt) on testicular and sperm abnormalities in a T2DM mouse model. KK-Ay (DM) and wild-type (non-DM) 4- and 7-week-old male mice were sacrificed to collect the testes and spermatozoa for immunofluorescence, RT-PCR, and histological analyses. DM and non-DM 7-week-old mice were subcutaneously infused with the AGE-Apt or control-aptamer for 6 weeks and were then sacrificed. Plasma glucose, testicular AGEs, and Rage gene expression in 4-week-old DM mice and plasma glucose, testicular AGEs, oxidative stress, and pro-inflammatory gene expressions in 7-week-old DM mice were higher than those in age-matched non-DM mice, the latter of which was associated with seminiferous tubular dilation. AGE-Apt did not affect glycemic parameters, but it inhibited seminiferous tubular dilation, reduced the number of testicular macrophages and apoptotic cells, and restored the decrease in sperm concentration, motility, and viability of 13-week-old DM mice. Our findings suggest that AGEs-Apt may improve sperm abnormality by suppressing AGE–RAGE-induced oxidative stress and inflammation in the testes of DM mice. [ABSTRACT FROM AUTHOR]

Background The association between the TDRD6 variants and human infertility remains unclear, as only one homozygous missense variant of TDRD6 was found to be associated with oligoasthenoteratozoospermia (OAT). Methods Whole-exome sequencing and Sanger sequencing were employed to identify potential pathogenic variants of TDRD6 in infertile men. Histology, immunofluorescence, immunoblotting and ultrastructural analyses were conducted to clarify the structural and functional abnormalities of sperm in mutated patients. Tdrd6-knockout mice were generated using the CRISPR-Cas9 system. Total RNA-seq and single-cell RNA-seq (scRNA-seq) analyses were used to elucidate the underlying molecular mechanisms, followed by validation through quantitative RT-PCR and immunostaining. Intracytoplasmic sperm injection (ICSI) was also used to assess the efficacy of clinical treatment. Results Bi-allelic TDRD6 variants were identified in five unrelated Chinese individuals with OAT, including homozygous loss-of-function variants in two consanguineous families. Notably, besides reduced concentrations and impaired motility, a significant occurrence of acrosomal hypoplasia was detected in multiple spermatozoa among five patients. Using the Tdrd6-deficient mice, we further elucidate the pivotal role of TDRD6 in spermiogenesis and acrosome identified. In addition, the mislocalisation of crucial chromatoid body components DDX4 (MVH) and UPF1 was also observed in round spermatids from patients harbouring TDRD6 variants. ScRNA-seq analysis of germ cells from a patient with TDRD6 variants revealed that TDRD6 regulates mRNA metabolism processes involved in spermatid differentiation and cytoplasmic translation. Conclusion Our findings strongly suggest that TDRD6 plays a conserved role in spermiogenesis and confirms the causal relationship between TDRD6 variants and human OAT. Additionally, this study highlights the unfavourable ICSI outcomes in individuals with bi-allelic TDRD6 variants, providing insights for potential clinical treatment strategies. [ABSTRACT FROM AUTHOR]

The biophysics of water, has been debated over more than a century. Although its importance is still underestimated, significant breakthroughs occurred in recent years. The influence of protein condensation on water availability control was documented, new findings on water-transport proteins emerged, and the way water molecules rearrange to minimize free energy at interfaces was deciphered, influencing membrane thermodynamics. The state of knowledge continued to progress in the field of deep-sea marine biology, highlighting unknown effects of high hydrostatic pressure and/or temperature on interactions between proteins and ligands in extreme environments, and membrane structure adaptations. The role of osmolytes in protein stability control under stress is also discussed here in relation to fish egg hydration/buoyancy. The complexity of water movements within the cell is updated, all these findings leading to a better view of their impact on many cellular processes. The way water flow and osmotic gradients generated by ion transport work together to produce the driving force behind cell migration is also relevant to both marine biology and cancer research. Additional common points concern water dynamic changes during the neoplastic transformation of cells and tissues, or embryo development. This could improve imaging techniques, early cancer diagnosis, and understanding of the molecular and physiological basis of buoyancy for many marine species. [ABSTRACT FROM AUTHOR]

Hyperpolarization of the membrane potential (Em), a phenomenon regulated by SLO3 channels, stands as a central feature in sperm capacitation--a crucial process conferring upon sperm the ability to fertilize the oocyte. In vitro studies demonstrated that Em hyperpolarization plays a pivotal role in facilitating the mechanisms necessary for the development of hyperactivated motility (HA) and acrosomal exocytosis (AE) occurrence. Nevertheless, the physiological significance of sperm Em within the female reproductive tract remains unexplored. As an approach to this question, we studied sperm migration and AE incidence within the oviduct in the absence of Em hyperpolarization using a novel mouse model established by crossbreeding of SLO3 knock-out (KO) mice with EGFP/DsRed2 mice. Sperm from this model displays impaired HA and AE in vitro. Interestingly, examination of the female reproductive tract shows that SLO3 KO sperm can reach the ampulla, mirroring the quantity of sperm observed in wild-type (WT) counterparts, supporting that the HA needed to reach the fertilization site is not affected. However, a noteworthy distinction emerges--unlike WT sperm, the majority of SLO3 KO sperm arrive at the ampulla with their acrosomes still intact. Of the few SLO3 KO sperm that do manage to reach the oocytes within this location, fertilization does not occur, as indicated by the absence of sperm pronuclei in the MII-oocytes recovered post-mating. In vitro, SLO3 KO sperm fail to penetrate the ZP and fuse with the oocytes. Collectively, these results underscore the vital role of Em hyperpolarization in AE and fertilization within their physiological context, while also revealing that Em is not a prerequisite for the development of the HA motility, essential for sperm migration through the female tract to the ampulla. [ABSTRACT FROM AUTHOR]

The surgical sterilization of cats and dogs has been used to prevent their unwanted breeding for decades. However, this is an expensive and invasive procedure, and often impractical in wider contexts, for example the control of feral populations. A sterilization agent that could be administered in a single injection, would not only eliminate the risks imposed by surgery but also be a much more cost-effective solution to this worldwide problem. In this study, we sought to develop a targeting peptide that would selectively bind to Leydig cells of the testes. Subsequently, after covalently attaching a cell ablation agent, Auristatin, to this peptide we aimed to apply this conjugated product (LH2Auristatin) to adult male mice in vivo, both alone and together with a previously developed Sertoli cell targeting peptide (FSH2Menadione). The application of LH2Auristatin alone resulted in an increase in sperm DNA damage, reduced mean testes weights and mean seminiferous tubule size, along with extensive germ cell apoptosis and a reduction in litter sizes. Together with FSH2Menadione there was also an increase in embryo resorptions. These promising results were observed in around a third of all treated animals. Given this variability, we discuss how these reagents might be modified in order to increase target cell ablation and improve their efficacy as sterilization agents. [ABSTRACT FROM AUTHOR]

The impact of supplementing glucose, glycine, or a combination of both in Ringer’s lactate egg yolk base extender to preserve the quality of semen from local Indonesian chickens has not been previously investigated. This study aimed to examine the potential of glucose and glycine on chicken semen stored at 5°C for 120 hours. In this study, five local roosters were used. The parameters under observation included semen volume, odor, pH levels, consistency, color, mass movement, concentration, motility, viability, abnormality, plasma membrane integrity, chromatin degeneration, and acrosomal cap integrity. This study was conducted using a completely randomized design (CRD) with four treatments groups and 10 replication, i.e. T1 (control without supplementation), T2 (50 mM glucose), T3 (60 mM glycine), and T4 (a combination of 50 mM glucose and 60 mM glycine), respectively. In result, semen volume was 0.54 ± 0.17 mL/ejaculate, a milky white color, distinctive odor, thick consistency, good mass movement (++/+++), pH of 7.37 ± 0.23, motility of 91.50 ± 2.42%, plasma membrane integrity of 96.85 ± 0.96%, abnormality at 2.88 ± 0.77%, the concentration of 3.04 ± 0.3 billion/mL, and viability of 96.47 ± 1.71%. Following storage at 5°C for 120 hours, the motility, viability, abnormality, and acrosomal cap integrity of local chicken spermatozoa significantly different (p < 0.05) between T3 and T4 compared to T1 and T2 groups. Moreover, the integrity of the plasma membrane and chromatin degeneration in treatment T3 significantly different (p < 0.05) from T1, T2, and T4 groups. In conclusion, local chickens exhibited fair quality fresh semen both in macroscopic and microscopic evaluations. Furthermore, the combination of 60 mM glycine and 50 mM glucose into local chicken semen stored at 5°C for 120 hours effectively preserved motility and viability, minimized abnormality, maintained plasma membrane integrity, minimized chromatin degeneration, and retained acrosomal integrity. [ABSTRACT FROM AUTHOR]

Carob belongs to the Fabaceae family, which has been studied for its medicinal properties, showcasing antioxidative properties and the ability to enhance sexual and immune system functions. This study aimed at examining the effect of adding hydro-alcoholic extracts of carob seeds to the diet on steroid hormones, sperm parameters and blood parameters of Lori-Bakhtiari rams. In this study, carob seeds were extracted using the percolation method. Thirty Lori-Bakhtiari rams were selected with a mean weight of 50 ± 5 kg and mean age of 2.5 years old. They were equally divided into three groups: (1) control (without carob extract), treatment (1) 150 mg/kg body weight (BW), and treatment (2) 300 mg/kg of carob seed extract in the diet for 4 weeks (2021-2022) for adaption process. Then semen and blood were analyzed for four months. Sperm motility, viability and the hypo-osmotic swelling (HOS) tests were evaluated at pre-freezing and post-thawing by the Computer Assisted Semen Analysis (CASA) software, Eosin-Nigrosin staining and a phase-contrast microscope, respectively. Furthermore, blood samples were collected for evaluation of fasting blood sugar (FBS), cholesterol (CH) and triglyceride (TG), serum creatinine (sCr), high-density lipoprotein (HDL), low-density lipoprotein (LDL), and testosterone hormone (T). The results of this study showed that the sperm parameters like motility and viability in the treatment groups increased significantly (P<0.05) in pre-freezing and post-thawing evaluations. Additionally, the results of blood parameters showed that FBS, CH, TG, LDL and sCr were significantly decreased by using the carob extract in a diet (P<0.05), while the testosterone level significantly increased during the experiment (P<0.05). In conclusion, the use of 300 mg/kg carob extract in the diet showed a more positive effect on sperm and blood parameters at Lori-Bakhtiari rams. [ABSTRACT FROM AUTHOR]

STUDY QUESTION Is the Tcte1 mutation causative for male infertility? SUMMARY ANSWER Our collected data underline the complex and devastating effect of the single-gene mutation on the testicular molecular network, leading to male reproductive failure. WHAT IS KNOWN ALREADY Recent data have revealed mutations in genes related to axonemal dynein arms as causative for morphology and motility abnormalities in spermatozoa of infertile males, including dysplasia of fibrous sheath (DFS) and multiple morphological abnormalities in the sperm flagella (MMAF). The nexin–dynein regulatory complex (N-DRC) coordinates the dynein arm activity and is built from the DRC1–DRC7 proteins. DRC5 (TCTE1), one of the N-DRC elements, has already been reported as a candidate for abnormal sperm flagella beating; however, only in a restricted manner with no clear explanation of respective observations. STUDY DESIGN, SIZE, DURATION Using the CRISPR/Cas9 genome editing technique, a mouse Tcte1 gene knockout line was created on the basis of the C57Bl/6J strain. The mouse reproductive potential, semen characteristics, testicular gene expression levels, sperm ATP, and testis apoptosis level measurements were then assessed, followed by visualization of N-DRC proteins in sperm, and protein modeling in silico. Also, a pilot genomic sequencing study of samples from human infertile males (n = 248) was applied for screening of TCTE1 variants. PARTICIPANTS/MATERIALS, SETTING, METHODS To check the reproductive potential of KO mice, adult animals were crossed for delivery of three litters per caged pair, but for no longer than for 6 months, in various combinations of zygosity. All experiments were performed for wild-type (WT, control group), heterozygous Tcte1+/− and homozygous Tcte1−/− male mice. Gross anatomy was performed on testis and epididymis samples, followed by semen analysis. Sequencing of RNA (RNAseq; Illumina) was done for mice testis tissues. STRING interactions were checked for protein–protein interactions, based on changed expression levels of corresponding genes identified in the mouse testis RNAseq experiments. Immunofluorescence in situ staining was performed to detect the N-DRC complex proteins: Tcte1 (Drc5), Drc7, Fbxl13 (Drc6), and Eps8l1 (Drc3) in mouse spermatozoa. To determine the amount of ATP in spermatozoa, the luminescence level was measured. In addition, immunofluorescence in situ staining was performed to check the level of apoptosis via caspase 3 visualization on mouse testis samples. DNA from whole blood samples of infertile males (n = 137 with non-obstructive azoospermia or cryptozoospermia, n = 111 samples with a spectrum of oligoasthenoteratozoospermia, including n = 47 with asthenozoospermia) was extracted to perform genomic sequencing (WGS, WES, or Sanger). Protein prediction modeling of human-identified variants and the exon 3 structure deleted in the mouse knockout was also performed. MAIN RESULTS AND THE ROLE OF CHANCE No progeny at all was found for the homozygous males which were revealed to have oligoasthenoteratozoospermia, while heterozygous animals were fertile but manifested oligozoospermia, suggesting haploinsufficiency. RNA-sequencing of the testicular tissue showed the influence of Tcte1 mutations on the expression pattern of 21 genes responsible for mitochondrial ATP processing or linked with apoptosis or spermatogenesis. In Tcte1−/− males, the protein was revealed in only residual amounts in the sperm head nucleus and was not transported to the sperm flagella, as were other N-DRC components. Decreased ATP levels (2.4-fold lower) were found in the spermatozoa of homozygous mice, together with disturbed tail:midpiece ratios, leading to abnormal sperm tail beating. Casp3-positive signals (indicating apoptosis) were observed in spermatogonia only, at a similar level in all three mouse genotypes. Mutation screening of human infertile males revealed one novel and five ultra-rare heterogeneous variants (predicted as disease-causing) in 6.05% of the patients studied. Protein prediction modeling of identified variants revealed changes in the protein surface charge potential, leading to disruption in helix flexibility or its dynamics, thus suggesting disrupted interactions of TCTE1 with its binding partners located within the axoneme. LARGE SCALE DATA All data generated or analyzed during this study are included in this published article and its supplementary information files. RNAseq data are available in the GEO database (https://www.ncbi.nlm.nih.gov/geo/) under the accession number GSE207805. The results described in the publication are based on whole-genome or exome sequencing data which includes sensitive information in the form of patient-specific germline variants. Information regarding such variants must not be shared publicly following European Union legislation, therefore access to raw data that support the findings of this study are available from the corresponding author upon reasonable request. LIMITATIONS, REASONS FOR CAUTION In the study, the in vitro fertilization performance of sperm from homozygous male mice was not checked. WIDER IMPLICATIONS OF THE FINDINGS This study contains novel and comprehensive data concerning the role of TCTE1 in male infertility. The TCTE1 gene is the next one that should be added to the 'male infertility list' because of its crucial role in spermatogenesis and proper sperm functioning. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by National Science Centre in Poland, grants no.: 2015/17/B/NZ2/01157 and 2020/37/B/NZ5/00549 (to M.K.), 2017/26/D/NZ5/00789 (to A.M.), and HD096723, GM127569-03, NIH SAP #4100085736 PA DoH (to A.N.Y.). The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported. [ABSTRACT FROM AUTHOR]

Simple Summary: In Thailand, the flat-headed cat, fishing cat, and Asiatic golden cat are among nine wild Felidae species requiring urgent conservation efforts. This study assessed routine sperm characteristics and detected protein biomarkers related to fertilization in these species. IZUMO1, involved in gamete interaction, showed the highest levels in the flat-headed cat. CRISP2 and CRISP3, crucial for sperm–egg fusion, had the highest levels in the fishing cat. Cleaved caspase-3, linked to DNA fragmentation, was found in all species and associated with decreased semen quality. These findings suggest that analyzing these biomarkers, alongside routine sperm characteristics, could enhance breeding management for wild Felidae, especially the flat-headed and fishing cats. Effective wild cat conservation programs with assisted reproductive technologies are being developed in different parts of the world. The flat-headed cat, fishing cat, and Asiatic golden cat are three species among nine wild Felidae in Thailand that are in need of urgent conservation efforts. Here, we assessed routine sperm characteristics and we report the detection of protein biomarkers related to the fertilization process, IZUMO1 and the CRISP family, and apoptotic markers, active or cleaved caspase-3, in semen samples collected from these wild cats. IZUMO1 was located in the equatorial segment of the sperm head, which is the region involved in gamete interaction. The highest levels of IZUMO1 were found in both the sperm pellet and the seminal plasma of the flat-headed cat, as determined by immunoblotting. CRISP2, a sperm–egg fusion assisting protein, and CRISP3 were found in both the sperm pellet and the seminal plasma, and the highest levels were observed in the fishing cat. Positive correlations between certain semen parameters and IZUMO1, CRISP2, and CRISP3 expression were also demonstrated. Cleaved caspase-3 was found in all sperm samples in all three species and was associated with an increase in DNA fragmentation and a decrease in certain semen characteristics such as motility, viability, and intact acrosomes. Our results suggest that the analysis of IZUMO1, the CRISP family, and cleaved caspase-3, along with the routine sperm characteristics, may allow for better success in breeding management in wild Felidae, particularly in the flat-headed cat and the fishing cat. [ABSTRACT FROM AUTHOR]

Introduction: Due to their relevant ecological position and well-studied biology, sea urchins are reference organisms for ocean acidification studies, at both within- and trans-generational levels. In this study, we examined gamete quality in specimens exposed to future predicted (-0.4 units) pH conditions during gametogenesis. Methods: Egg physical characteristics, biochemical composition, and fatty acid profiles were assessed after two and six months of exposure, while sperm viability and velocity were analyzed after six months of exposure. Considering the documented intraspecific variability in response to ocean acidification, this study involved two populations of Paracentrotus lividus. One population was sampled from the highly variable lagoon of Venice (Site 1), while the other was obtained from a coastal area (Site 2) characterized by more stable environmental conditions and facing minimal anthropogenic stress. Results: A different response was highlighted in the two sites. Noteworthy trends emerged, especially in the fatty acid profile and sperm traits. Although adults were fed the same diet, Site 1 eggs contained more high-energetic fatty acids than Site 2, potentially boosting the survival odds for the next generation. Moreover, Site 1 sperms displayed higher viability but slower motility compared to those from Site 2. Within sites, a significant difference between time points and a change in the fitness strategy of sea urchin females emerged when comparing eggs spawned after two and six months of exposure to reduced pH. The effects of time and exposure pH are more pronounced in animals from Site 1, suggesting a higher adaptability of this population rather than negative effects of ocean acidification. Discussion: Overall, our findings suggest that sea urchins have the potential to acclimate to reduced pH and to produce gametes of the same quality as controls held at the currently natural pH. Our findings emphasize the relevance of combining investigations of gamete quality characteristics, particularly egg biochemistry and fatty acid composition, and considering site variability to fully understand the transgenerational response potential of sea urchins to ocean acidification. [ABSTRACT FROM AUTHOR]

Silver nanoparticles possess valuable physical, chemical, and biological properties, rendering them widely applied as bioactive agents in the industry. Nonetheless, their influence on the natural environment and on living organisms remains unclear. Therefore, this study aims to investigate the impact of polymer composites containing silver nanoparticles on sperm cells. The nanosilver polymer composites were chemically synthesized, employing sodium alginate as the stabilizer. The reducing agents employed were solutions comprising sodium borohydride and xylose. The concentration of silver nanoparticles obtained after synthesis was 100 parts per million. The examined biological species were rabbit sperm cells. The impact of nanosilver on the sperm was assessed through the elucidation of the toxicity profile, comet test, and analysis of morphological characteristics of the animal cells. The results of the study demonstrate a twofold impact of polymer composites infused with silver nanoparticles on domestic rabbit sperm when obtained through chemical synthesis using two reducing agents (xylose and sodium borohydride) at a 10 ppm concentration. The comet test showed no harmful effect on the DNA integrity of rabbit sperm by the tested compounds. Twenty-four-hour exposure of rabbit spermatozoa to silver nanoparticles, obtained by reducing xylose and borohydride, induced significant secondary changes in the morphological structure of male reproductive cells. These findings indicate the potential reproductive toxicity of silver nanoparticles. [ABSTRACT FROM AUTHOR]

This study aimed to assess the impact of different dilution ratios of rabbit semen to Tris Citrate Glucose (TCG) extender on doe fertility through artificial insemination in the Mekong Delta region of Vietnam. The experiment included three dilution ratios (1:10, 1:15, and 1:20), with each treatment replicated 12 times. Semen quality variables were evaluated and does were artificially inseminated using a standardized technique. Our findings revealed that the 1:20 dilution exhibited the highest percentage of successful insemination (66.67%) and the shortest gestation period (31.6 days). Furthermore, the 1:20 dilution treatment resulted in an average litter size at birth of 5.25 kits, a litter weight at birth of 290.37 g, and a litter weight at weaning of 1963.04 g. These results suggest that the 1:20 dilution ratio holds promise for artificial insemination in rabbit breeding programs in the Mekong Delta of Vietnam. [ABSTRACT FROM AUTHOR]

Simple Summary: Our study aimed to understand how melatonin affects reproductive parameters in Bafra rams during the non-breeding season. One group of rams received melatonin implants, while another served as the control. We measured testicular volume, blood flow, and semen quality over several weeks. The results showed that melatonin increased testicular size and blood flow. This suggests that providing melatonin implants before the non-breeding season may improve reproductive capacity. Our objectives were to investigate the effects of exogenous melatonin on testicular volume (TV), testicular blood flow (TBF), and semen quality in Bafra rams during the non-breeding season. One group of rams (MEL, n = 5) received a 36 mg melatonin implant twice, with 30 days in between, while the other group (CON, n = 5) served as the control. TBF, TV, and semen quality parameters were determined at three-week intervals starting three weeks before until twelve weeks after the first melatonin implant. Testicular blood flow was determined in the supratesticular (STA) and marginal testicular artery (MA) using color Doppler ultrasound. Semen was collected and evaluated, and the total oxidative status (TOS) and total antioxidative status (TAS) was determined using an ELISA. The MEL group had increased (p < 0.05) TV between the sixth and twelfth week after the start of treatment. Overall, the MEL group had lower resistance and pulsatility indexes (p < 0.05) between the third and ninth week, although there was no difference (p > 0.05) between the two groups in most semen quality parameters. However, TAS concentrations increased (p < 0.05) in the MEL group compared with the CON. The results of this study show that exogenous melatonin in the non-breeding season significantly increased both TBF and TV in Bafra rams. Therefore, giving rams implants with 36 mg melatonin twice at least one month prior to the non-breeding season is expected to improve testicular size and function and reproductive capacity. [ABSTRACT FROM AUTHOR]

Canine seminal plasma is a complex fluid containing proteins, peptides, enzymes, hormones as well as extracellular vesicles that are involved in many physiological and pathological processes including reproduction. We examined the expression of the extracellular vesicles surface antigens Aminopeptidase-N (CD13) and Dipeptidyl peptidase IV (CD26) by flow cytometry. For this study, third fraction of the ejaculate, from fertile adult male German Shepherd dogs, was manually collected twice, two days apart. FACS analyses revealed that CD13 and CD26 are co-expressed on the 69.3 ± 3.7% of extracellular vesicles and only a 2.0 ± 0.5% of extracellular vesicles express CD26 alone. On the other hand, 28.6 ± 3.6% of seminal EVs express CD13 alone. Our results agree with the hypothesis that CD26 needs to be co-expressed with other signal-transducing molecules, while CD13, can perform functions independently of the presence or co-expression of CD26. The results obtained in normal fertile dogs could represent physiological expression of these enzymes. Therefore, it would be interesting to carry out further studies to evaluate the expression of CD13 and CD26 on extracellular vesicles as biomarker for prostate pathological condition in dogs. [ABSTRACT FROM AUTHOR]

To understand the impact of sperm speed as they swim against the flow on fertilization rates, we created conditions similar to the female reproductive tract (FRT) on a microfluidic platform for sperm selection. Selected sperm were evaluated based on early development of fertilized embryos. Bovine and human spermatozoa were selected at various fluid flow rates within the device. We found that the speed of bovine spermatozoa increases as the flow rate increases and that the amount of DNA fragmentation index is lowered by increasing the flow rate. Bovine spermatozoa selected by our platform at low (150 μL h−1, shear rate 3 s−1), medium (250 μL h−1, shear rate 5 s−1), and high flow rates (350 μL h−1, shear rate 7 s−1) were used for fertilization and compared to sperm sorted by centrifugation. The samples collected at the highest flow rate resulted in the formation of 23% more blastocysts compared to the control. While selecting for higher quality sperm by increasing the flow rate does result in lower sperm yield, quality improvement and yield may be balanced by better embryonic development. [ABSTRACT FROM AUTHOR]

The present study aimed to examine the protective potential of cholesterol loaded cyclodextrin (CLC) at varying concentrations against sperm morphological abnormalities of cooled and frozen thawed semen of Marwari stallions. A total of forty eight ejaculates (six stallions and eight ejaculates per stallion) were promptly processed following semen collection and subjected to gross assessment of semen quality attributes. The assessment of sperm morphologies in fresh equine semen was conducted using Eosin and Nigrosine staining technique. The semen samples from individual Marwari stallions were evenly divided into five fractions and incubated in a water bath at 37℃ for 15 min after dilution with primary extender to obtain the spermatozoa concentration at 120 × 106 sperm/ml. During incubation, the fractions of every semen sample were exposed to different concentrations of CLC as 0, 1.0, 1.5, 2.0 and 3.0 mg/ml of extender (C, T1, T2, T3, and T4, respectively). All the 5 aliquots were cooled and subsequently subjected to cryopreservation. Each cooled and frozen-thawed (at 37℃ for 30 sec) semen samples were analyzed to assess the morphologies of the spermatozoa. Based on the findings, it was concluded that none of the CLC treatment could render any significant protection against the sperm morphological abnormalities at either pre-freeze and/or post-thaw stage of semen samples from Marwari stallions. [ABSTRACT FROM AUTHOR]

Bumblebees establish annual colonies that start with the emergence of workers in spring and end with the production of sexuals, the majority of which are males, in late summer. To date, the causes responsible for the transition in the production of diploid-female offspring to haploid-male offspring during the decline phase of colonies remain elusive. Using flow cytometry, we tested whether such a caste shift is correlated with a decline in sperm number and quality (i.e., sperm viability and sperm DNA fragmentation) in the queen spermatheca over time, from mating to the emergence of the first males. We found that sperm number and viability significantly decreased, while sperm DNA fragmentation increased in the spermatheca over time. These results suggest that the shift towards male production during the decline phase of a bumblebee colony stems at least partly from a combination of a drop in sperm count and sperm quality in queens' spermatheca. Significance statement: In social Hymenoptera, sex determination is based on the haplodiploidy system in which males develop from unfertilized eggs and are haploid, and females develop from fertilized eggs and are diploid. So far, the proximal mechanisms responsible for the social transition from diploid-female egg production to haploid-male egg production remain unknown. We show that the shift towards male production during the decline phase of bumblebee colonies is associated with a reduction in the quantity and quality of the sperm stored in the queen spermatheca over time. More generally, it suggests that sperm conservation is limited and likely adapted to the relatively short lifespan of bumblebee queens, which produce small and annual colonies. [ABSTRACT FROM AUTHOR]

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Tartrazine is a yellow colouring agent that is commonly used in foods; however, high dosages of Tartrazine affect fertility and create oxidative stress by generating free radicals. A plant species known as Stevia rebaudiana has natural antioxidants that show promise for protecting testicular tissue. Consequently, this study was intended to examine the ameliorative effect of the aqueous extract of S. rebaudiana (Stevia) on the fertility of male Wistar rats induced by the daily oral intake of Tartrazine. Utilizing gas chromatography–mass spectrometry, phytochemical identification was accomplished for Stevia extract. Study groups were separated into several groups: the first group (the control) got distilled water for up to 56 days; the Stevia group (1000 mg/kg), the Tartrazine group (300 mg/kg) and the Stevia and Tartrazine group (the group was given Tartrazine after 1 h of Stevia extract intake). Also, the oxidative damage in testicular tissues was assessed by measuring the levels of malondialdehyde (MDA) and antioxidants (catalase [CAT], superoxide dismutase [SOD] and glutathione reductase [GSH]). Further, histological alterations were examined. In addition, cyclic AMP‐responsive element modulator (Crem) gene expression levels and their relative proteins were measured in the testicular tissues using quantitative real‐time polymerase chain reaction and enzyme‐linked immunosorbent assays, respectively. Sperm analysis and testosterone concentration were also performed. SPSS version 25 was used for the analysis of results while (p <.05) was regarded as significant. Compared with the control group, the results demonstrated that Tartrazine caused a significant reduction (p <.05) in the testosterone hormone level (0.70 ± 0.21) and the Crem protein quantity (1.21 ± 0.23) in the treated Tartrazine group. Also, it had a significant decrease (p <.05) in sperm motility, viability, count and antioxidant levels. Moreover, there was a significant increase (p <.05) in sperm abnormalities, MDA level (7.40 ± 1.10), kidney and liver function parameters, and DNA degradation in the treated Tartrazine group compared with the control group. On the contrary, the Stevia extract intake enhanced the testosterone (2.50 ± 0.60), antioxidants and Crem protein levels (2.33 ± 0.10) with an improvement in sperm quality in the Stevia and Tartrazine‐treated group compared with the Tartrazine group. Stevia also caused a significant decrease (p <.05) in the MDA level (3.20 ± 0.20), and sperm abnormalities with an enhancement of the liver and kidney function parameters in the Stevia and Tartrazine‐treated group compared to the Tartrazine group. Stevia administration has a protective effect on the testicular tissues and sperm quality against toxicity induced by Tartrazine exposure, so it will be a good antioxidant drug to be administered daily before daily administration of Tartrazine. Significance statement: Tartrazine, a yellow food colouring, gradually reduces testosterone and affects fertility, according to recent studies. However, many herbal supplements boost sperm count and male reproductive function. Aqueous Stevia extract (ASE) was tested on male albino rats to see if it could reduce Tartrazine's toxicity. The study found that daily tartrazine consumption induced renal, hepatic and testicular damage. However, ASE treatment ameliorated these abnormalities by enhancing renal and hepatic functions, improving sperm quality, increasing testicular antioxidant indicator levels and decreasing Tartrazine‐induced DNA damage. Thus, the present investigation provided empirical support for the potential use of ASE as a therapeutic agent with dual efficiency, treating infertility and reducing the negative side effects of Tartrazine. [ABSTRACT FROM AUTHOR]

Livestock production requires Sapudi rams, a breed native to Indonesia, to meet meat demand and food security. In artificial high-quality frozen semen is needed to spread Sapudi rams. To maximize the survival of spermatozoa during cryopreservation, semen should be stored in an extender. Green tea leaf extract (GTLE) and skim milk-goat egg yolk (SM-GEY) may be a good cryoprotectants due to their antioxidant properties. This study aimed to determine the effect of adding GTLE to the SM-GEY extender on the quality of Sapudi ram spermatozoa stored at 5°C. The fresh semen sample was divided into four different GTLE treatment groups, which each contained a 0.1 mL semen sample and a 25-mL extender of SM-GEY. Group T0: no GTLE added to SM-GEY; Groups T1, T2, and T3: 0.1 mL semen diluted in 25 mL SM-GEY with 0.05, 0.10, and 0.15 mg GTLE. Extended semen was then stored at 5°C, and its quality was evaluated daily for five days. The variables observed included spermatozoa motility, viability, and membrane integrity. Data were analyzed using one-way analysis of variance followed by Duncan's test using Statistical Program and Service Solution version 23. The result of this study was that adding 0.05 mg GTLE to 25 mL of SMGEY extender significantly maintained the spermatozoa motility, viability, and plasma membrane integrity of Sapudi ram spermatozoa for three days at 5°C (p <0.05). Therefore, it could be concluded that adding 0.05 mg of GTLE to the SM-GEY extender preserved Sapudi ram spermatozoa's motility, viability, and membrane integrity for three days at 5°C. [ABSTRACT FROM AUTHOR]

This study aims to determine the effect of the administration of white guava (Psidium guajava L.) fruit juice on spermatozoa plasma membrane integrity (PMI), morphological abnormality, viability, and motility of lead acetate induced rats (Rattus norvegicus). Twenty-five male rats were divided into five groups: NC (negative control) group, rats were administered with distilled water twice daily at four-hour intervals; T0 (positive control) group, rats were administered daily with lead acetate 50 mg/kg bw and distilled water four hours later; T1, T2, and T3 groups, rats were administered daily with lead acetate 50 mg/kg bw and 0.5 mL of 25, 50, and 100% white guava fruit juice four hours later. The treatment of the rats was conducted for 14 days, and on day 15, all rats were sacrificed to assess the spermatozoa quality. Data was analyzed using ANOVA followed by Duncan's multiple range test at a confidence level of 95%. The results showed that exposure to lead acetate (T0) caused lower spermatozoa PMI, viability, and motility as well as higher spermatozoa morphological abnormalities (p <0.05) compared to those of the T0 group. Administration of white guava fruit juice starting at a dose of 25% (T1) resulted in higher spermatozoa motility, viability, and PMI as well as lower spermatozoa morphological abnormalities (p <0.05) compared to rats in the T0 group. It could be concluded that white guava fruit juice maintained the spermatozoa quality of lead acetate induced rats. [ABSTRACT FROM AUTHOR]

Adding endogenous antioxidants to the diluent is significantly associated with semen quality during the freezing process. This study aimed to investigate the effects of adding glutathione to AndroMed diluent on the preservation of crucial sperm attributes, namely, intact plasma membrane (IPM), intact acrosome membrane (IAM), and progressive motility of Bali cattle spermatozoa. A completely randomized design was used, and spermatozoa samples were obtained from a Bali cattle and divided into two diluent treatment groups (36 diluent samples in each group with six replications), namely pure AndroMed as the control and a group with the addition of glutathione (1 mmol) to AndroMed. Each treatment was replicated six times and evaluated at three freezing stages, including postdilution, post-equilibration, and post-thawing, for crucial sperm properties. The results indicated that fresh Bali cattle spermatozoa had progressive motility, IAM, and IPM of 75%, 89%, and 88%, respectively. During the freezing process, there was a significant decrease in semen quality, including progressive motility, IAM, and IPM of spermatozoa after dilution to post-equilibration and post-equilibration to post-thawing in both treatment groups. Meanwhile, the addition of 1 mmol of glutathione to AndroMed diluent had a significant difference in increasing progressive motility, IAM, and IPM of Bali cattle spermatozoa at each stage of semen freezing, including postdilution, post-equilibration, and post thawing when compared with controls. Based on the results, it can be concluded that adding 1 mmol of glutathione to the AndroMed diluent enhanced the quality and integrity of Bali cattle semen, including progressive motility, IAM, and IPM during the freezing process. [ABSTRACT FROM AUTHOR]

Context: Several assisted reproduction techniques have been proposed for collared peccaries due to the interest in its conservation. However, there is little information about the requirements for sperm capacitation, an initial step in in vitro fertilisation. Aims: We aimed to determine the optimal conditions for collared peccary sperm capacitation by comparing Tyrode's albumin lactate pyruvate (TALP) or the same media plus heparin, caffeine, or their combination at different exposure times (1, 3, and 6 h). Methods: The samples were evaluated for kinetic parameters, membrane functionality and integrity, mitochondrial activity, morphology, DNA integrity, reactive oxygen species (ROS) expression levels, capacitation, and sperm binding ability using swine oocytes. Key results: Samples incubated with caffeine or heparin had a higher percentage of capacitated spermatozoa. The maximum percentage of capacitation was achieved after 3 h of incubation with either agent. Moreover, spermatozoa subjected to heparin capacitation showed better motility than when subjected to caffeine, and lower ROS expression levels after 1 h. No differences were observed among incubation times for the binding ability. Conclusions: In summary, collared peccary spermatozoa can be capacitated with caffeine or heparin; however, heparin better maintains sperm motility and ROS expression levels. The co-incubation of gametes in a medium with heparin for 3 h could be efficient for in vitro fertilisation in collared peccaries. Implications: This result will contribute to the development of assisted reproduction techniques for conservation and productivity of collared peccaries. In vitro fertilisation can contribute to the conservation and knowledge of the reproductive characteristics of collared peccary. There is little information about the requirements for sperm capacitation in the collared peccary, an initial step in in vitro fertilisation. We compared different capacitating media and incubation times and we observed that heparin improved sperm motility and promoted the capacitation of collared peccary spermatozoa. This result will contribute to the development of assisted reproduction techniques for conservation and productivity of collared peccaries. [ABSTRACT FROM AUTHOR]

It is important to find and identify more biological adsorbents that can effectively remove metals from water instead of using the traditional approach. In the present study, a fungal strain resistant to cadmium was isolated from polluted sites. It was identified morphologically and molecularly as Alternaria tenuissima, with an accession number of OQ691600. Heat inactivated biomass of A. tenuissima was evaluated for cadmium (Cd) biosorption. The biosorption process was conducted at a temperature of 28±2°C to remove Cd from water at an initial concentration of 92 mg/L, under different conditions, such as pH (4-6), biosorbent dose (1-3 g/L), and sorption time (15-60 min). The obtained results demonstrated that the dead biomass of A. tenuissima effectively removed Cd, with maximum significant biosorption efficiency of 99.95±0.003% and capacity of 92.33±0.21 mg/g, which were achieved at a pH value of 6 for 30 min and a biosorbent dose of 1 g/L. Fourier transform-infrared (FT-IR) spectroscopy analysis revealed changes in the functional groups present on the surface of A. tenuissima biomass through the biosorption process. Following the biosorption process, three groups of male albino rats were used. The negative control group received only double-distilled water through oral administration. The Cd-polluted water group received a Cd aqueous solution (92 mg/L) through oral administration for 30 days. The treatment group received Cd aqueous solution (92 mg/L) through oral administration after treatment with A. tenuissima as the biosorbent material for a period of 30 days. The results showed that Cd caused a significant drop in weight gain and testicular index weight, as well as total protein, albumin, semen count, and motility. There were significant increases in abnormal sperm, the spleen, and brain index weight, as well as biochemical parameters, including aspartate aminotransferase, alanine aminotransferase, urea, creatinine, and uric acid, showed notable changes. The liver, kidney, testis, and epididymis of male albino rats showed several histopathological changes along with bioaccumulation of Cd traces in liver and kidney tissues. However, in the treatment group, all these findings were noticeably improved. Based on our findings, the dead biomass of A. tenuissima has the potential to be a very effective biosorbent for removing cadmium from water, without causing any hazardous effects on animals. [ABSTRACT FROM AUTHOR]

Background: Sperm deoxyribonucleic acid (DNA) fragmentation is commonly encountered in spermatozoa, and the oocyte assumes responsibility for repairing sperm DNA fragmentation during the oocyte–embryo transition. Objectives: This study aimed to investigate whether the effect of sperm DNA fragmentation on intracytoplasmic sperm injection outcomes depends on the incidence of oocyte dimorphisms. Materials and methods: For the present cohort, 2942 fertilized oocytes from 525 patients submitted to intracytoplasmic sperm injection cycles were assessed. The present study was conducted in a private in vitro fertilization center affiliated to a university from June 2016 to July 2019. Semen samples were divided into the following two groups depending on the sperm DNA fragmentation index: a low fragmentation index group (<30% sperm DNA fragmentation, n = 1468) and a high fragmentation index group (≥30% sperm DNA fragmentation, n = 486). In addition, mature oocytes were examined before sperm injection, and intracytoplasmic and extracytoplasmic defects were recorded. The effect of the sperm DNA fragmentation index on laboratory and clinical intracytoplasmic sperm injection outcomes (depending on the presence of oocyte defects) was evaluated. Results: Significant increases in the rates of fertilization, high‐quality embryo, implantation, and pregnancy were noted for cycles with <30% sperm DNA fragmentation than cycles with ≥30% sperm DNA fragmentation (regardless of the presence of oocyte dimorphisms). The presence of dimorphisms significantly impacted laboratory and clinical outcomes. The lowest fertilization and high‐quality embryo rates were observed when a high sperm DNA fragmentation index was associated with the presence of dark cytoplasm, vacuoles, resistant membrane, and non‐resistant membrane. The lowest implantation and pregnancy rates were observed when a high sperm DNA fragmentation index was associated with the presence of vacuoles, defective perivitelline space, and fragmented polar body. The effect of sperm DNA fragmentation on miscarriage rates was significantly influenced by the presence of centrally located cytoplasmic granulation, a defective perivitelline space and non‐resistant membrane. Conclusion: A high sperm DNA fragmentation index increases the likelihood of miscarriage in intracytoplasmic sperm injection cycles, an effect that may potentially be magnified by the presence of oocyte dysmorphisms. [ABSTRACT FROM AUTHOR]