Sperm Research Articles

Human sperm RNA in male infertility.

The function and value of specific sperm RNAs in apparently idiopathic male infertility are currently poorly understood. Whether differences exist in the sperm RNA profile between patients with infertility and fertile men needs clarification. Similarly, the utility of sperm RNAs in predicting successful sperm retrieval and assisted reproductive technique (ART) outcome is unknown. Patients with infertility and fertile individuals seem to have differences in the expression of non-coding RNAs that regulate genes controlling spermatogenesis. Several RNAs seem to influence embryo quality and development. Also, RNA types seem to predict successful sperm retrieval in patients with azoospermia. These findings suggest that sperm RNAs could influence decision-making during the management of patients with infertility. This evidence might help to identify possible therapeutic approaches aimed at modulating the expression of dysregulated genes in patients with infertility. Performing prospective studies with large sample sizes is necessary to investigate cost-effective panels consisting of proven molecular targets to ensure that this evidence can be translated to clinical practice.

The Effect of Metallic Nanoparticles Supplementation in Semen Extender on Post-thaw Quality and Fertilizing Ability of Egyptian Buffalo (Bubalus bubalis) Spermatozoa.

Nanomaterials offer several promising prospects in the field of farm animal reproduction, encompassing a broad range of applications such as transgenesis and the precise delivery of substances to sperm cells, antimicrobial, antioxidants properties as well as their potent role in improving cryopreservation methods. The aim of the present study is to explore the effect of supplementing the semen extender with 10µg/mL nano gold (Au-NPs10), 10µg/mL nano silver (Ag-NPs10), 1µg/mL nano selenium (Se-NPs1), and 100µg/mL nano zinc oxide (ZnO-NPs100) on sperm characteristics and kinematics parameters, acrosome integrity, oxidative biomarkers, morphological and apoptosis-like changes of frozen-thawed buffalo bull sperm, and, ultimately, their fertilizing capacity. The results revealed that all aforementioned nano materials significantly improved viability, progressive motility, membrane integrity, acrosome integrity, and kinematic parameters as well as apoptosis-like changes of post-thawed buffalo bull sperm compared to the control (p < 0.05). No discernible effects were observed on sperm ultrastructure morphology measures as a response to the addition of these metallic nanoparticles to the extender. The values of caspase 3 significantly decreased by 64.22, 45.99, 75.59, and 49.39% in Au-NPs10, Ag-NPs10, Se-NPs1, and ZnO-NPs100treated groups, respectively, compared to the control. The addition of 100µg ZnO-NPs to the extender significantly decreased the total count of bacteria, fungi, and yeast compared to the control (p < 0.05). The AuNPs10 and SeNPs1 treated groups showed lower content of malondialdehyde, hydrogen peroxide, and nitric oxide concentrations and higher values of total antioxidant capacity of post-thawed extended semen (p < 0.05). Pregnancy rates increased by 17.5, 20, and 30% in buffalo cows inseminated with sperm treated with ZnO-NPs100, Se-NPs1, and Au-NPs10, respectively, compared to the control group. The present results indicate that the freezing extender supplemented with metallic nanoparticles can be an effective strategy to enhance the cryotolerance and fertility potential of buffalo bull sperm.

A modified U-Net to detect real sperms in videos of human sperm cell.

This study delves into the crucial domain of sperm segmentation, a pivotal component of male infertility diagnosis. It explores the efficacy of diverse architectural configurations coupled with various encoders, leveraging frames from the VISEM dataset for evaluation. The pursuit of automated sperm segmentation led to the examination of multiple deep learning architectures, each paired with distinct encoders. Extensive experimentation was conducted on the VISEM dataset to assess their performance. Our study evaluated various deep learning architectures with different encoders for sperm segmentation using the VISEM dataset. While each model configuration exhibited distinct strengths and weaknesses, UNet++ with ResNet34 emerged as a top-performing model, demonstrating exceptional accuracy in distinguishing sperm cells from non-sperm cells. However, challenges persist in accurately identifying closely adjacent sperm cells. These findings provide valuable insights for improving automated sperm segmentation in male infertility diagnosis. The study underscores the significance of selecting appropriate model combinations based on specific diagnostic requirements. It also highlights the challenges related to distinguishing closely adjacent sperm cells. This research advances the field of automated sperm segmentation for male infertility diagnosis, showcasing the potential of deep learning techniques. Future work should aim to enhance accuracy in scenarios involving close proximity between sperm cells, ultimately improving clinical sperm analysis.

Human sperm mitochondrial DNA copy numbers and deletion rates: Comparing persons living in two urban industrial agglomerations differing in sources of air pollution

Persons living in industrial environments are exposed to levels of air pollution that can affect their health and fertility. The Czech capital city, Prague, and the Ostrava industrial agglomeration differ in their major sources of air pollution. In Prague, heavy traffic produces high levels of nitrogen oxides throughout the year. In the Ostrava region, an iron industry and local heating are sources of particulate matter (PM) and benzo[a]pyrene (B[a]P), especially in the winter. We evaluated the effects of air pollution on human sperm mitochondrial DNA (mtDNA). Using real-time PCR, we analysed sperm mtDNA copy number and deletion rate in Prague city policemen in two seasons (spring and autumn) and compared the results with those from Ostrava. In Prague, the sperm mtDNA deletion rate was significantly higher in autumn than in spring, which is the opposite of the results from Ostrava. The sperm mtDNA copy number did not show any seasonal differences in either of the cities; it was correlated negatively with sperm concentration, motility, and viability, and with sperm chromatin integrity (assessed with the Sperm Chromatin Structure Assay). The comparison between the two cities showed that the sperm mtDNA deletion rate in spring and the sperm mtDNA copy number in autumn were significantly lower in Prague vs. Ostrava. Our study supports the hypothesis that sperm mtDNA deletion rate is affected by the composition of air pollution. Sperm mtDNA abundance is closely associated with chromatin damage and standard semen characteristics.

How do taurine and ergothioneine additives improve the parameters of high- and low-quality turkey semen during liquid preservation?

Abstract The study aimed to examine the parameters of turkey ejaculates (n=40) of high and low quality (HQ and LQ, respectively), preserved with the addition of taurine (TAU; 5 mM, 10 mM) and ergothioneine (EGT; 5 mM, 10 mM) for 48 h at 5°C in a liquid state. The motility, plasma membrane integrity (PMI), mitochondrial function, apoptotic and necrotic spermatozoa percentage, and sperm cells that generated NO were determined after 2, 24, and 48 h of storage. The preserved semen was also analysed for the activity of superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), glutathione (GSH) content, and malondialdehyde (MDA) production. Taurine, in both doses, may improve the antioxidant status of stored turkey semen as well as sperm motility, viability and functionality regardless of ejaculate quality, as manifested by increased SOD and CAT activities, reduced MDA levels, and enhanced sperm characteristics i.e. plasma membrane integrity (PMI), mitochondrial membrane potential (MMP), total motility (TMOT), and progressive motility (PMOT). In turn, the addition of EGT increased GSH content in the external environment and suppressed lipid peroxidation in turkey spermatozoa, in particular those from low-quality (LQ) ejaculates. However, it appears that EGT could deliver more satisfactory results when added at lower concentrations.

TUBB4B is essential for the expansion of differentiating spermatogonia

Microtubules, polymers of αβ-tubulin heterodimers, are essential for various cellular processes. The incorporation of different tubulin isotypes, each encoded by distinct genes, is proposed to contribute to the functional diversity observed in microtubules. However, the functional roles of each tubulin isotype are not completely understood. In this study, we investigated the role of the β4B-tubulin isotype (Tubb4b) in spermatogenesis, utilizing a Tubb4b knockout mouse model. We showed that β4B-tubulin is expressed in the germ cells throughout spermatogenesis. β4B-tubulin was localized to cytoplasmic microtubules, mitotic spindles, manchette, and axonemes of sperm flagella. We found that the absence of β4B-tubulin resulted in male infertility and failure to produce sperm cells. Our findings demonstrate that a lack of β4B-tubulin leads to defects in the initial stages of spermatogenesis. Specifically, β4B-tubulin is needed for the expansion of differentiating spermatogonia, which is essential for the subsequent progression of spermatogenesis.

D4Z4 Hypomethylation in Human Germ Cells.

Expression of the double homeobox 4 (DUX4) transcription factor is highly regulated in early embryogenesis and is subsequently epigenetically silenced. Ectopic expression of DUX4 due to hypomethylation of the D4Z4 repeat array on permissive chromosome 4q35 alleles is associated with facioscapulohumeral muscular dystrophy (FSHD). In peripheral blood samples from 188 healthy individuals, D4Z4 methylation was highly variable, ranging from 19% to 76%, and was not affected by age. In 48 FSHD2 patients, D4Z4 methylation varied from 3% to 30%. Given that DUX4 is one of the earliest transcribed genes after fertilization, the D4Z4 array is expected to be unmethylated in mature germ cells. Deep bisulfite sequencing of 188 mainly normozoospermic sperm samples revealed an average methylation of 2.5% (range 0.3-22%). Overall, the vast majority (78%) of individual sperm cells displayed no methylation at all. In contrast, only 19 (17.5%) of 109 individual germinal vesicle (GV) oocytes displayed D4Z4 methylation <2.5%. However, it is not unexpected that immature GV oocytes which are not usable for assisted reproduction are endowed with D4Z4 (up to 74%) hypermethylation and/or abnormal (PEG3 and GTL2) imprints. Although not significant, it is interesting to note that the pregnancy rate after assisted reproduction was higher for donors of sperm samples and oocytes with <2.5% methylation.

Evaluation of Known Markers of Ferroptosis in Semen of Patients with Different Reproductive Pathologies and Fertile Men.

This study aims to investigate the role of ferroptosis, an iron-dependent form of regulated cell death, in male infertility. The motivation behind this research stems from the increasing recognition of oxidative stress and iron metabolism dysregulation as critical factors in male reproductive health. In this study, 28 infertile patients (grouped by the presence of urogenital infections or varicocele) and 19 fertile men were selected. Spermiograms were performed by light microscopy (WHO, 2021). Testosterone, ferritin, transferrin-bound iron, transferrin, and F2-isoprostanes (F2-IsoPs) were detected in seminal plasma. Glutathione peroxidase 4 (GPX4) and acyl coenzyme A synthetase long chain family member 4 (ACSL4) were also assessed in sperm cells using enzyme-linked immunosorbent assays (ELISA). All the variables were correlated (statistically significant Spearman's rank correlations) in the whole population, and then the comparison between variables of the different groups of men were carried out. Seminal ferritin and transferrin positively correlated with seminal F2-IsoPs, which had positive correlations with ACSL4 detected in sperm cells. Ferritin and ACSL4 negatively correlated with the seminal parameters. No correlation was detected for GPX4. Comparing the variables in the three examined groups, elevated levels of ACSL4 were observed in infertile patients with urogenital infections and varicocele; GPX4 levels were similar in the three groups. These results suggested a mechanism of ferroptosis, identified by increased ACSL4 levels and the occurrence of lipid peroxidation. Such events appear to be GPX4-independent in reproductive pathologies such as varicocele and urogenital infections.

The efficacy and functional consequences of interactions between human spermatozoa and seminal fluid extracellular vesicles.

Seminal fluid extracellular vesicles (SFEVs) have previously been shown to interact with spermatozoa and influence their fertilisation capacity. Here, we sought to extend these studies by exploring the functional consequences of SFEV interactions with human spermatozoa. SFEVs were isolated from the seminal fluid of normozoospermic donors prior to assessing the kinetics of sperm-SFEV binding in vitro, as well as the effects of these interactions on sperm capacitation, acrosomal exocytosis, and motility profile. Biotin-labelled SFEV proteins were transferred primarily to the flagellum of spermatozoa within minutes of co-incubation, although additional foci of SFEV biotinylated proteins also labelled the mid-piece and head domain. Functional analyses of high-quality spermatozoa collected following liquefaction revealed that SFEVs did not influence sperm motility during incubation at pH 5, yet SFEVs induced subtle increases in total and progressive motility in sperm incubated with SFEVs at pH 7. Additional investigation of sperm motility kinematic parameters revealed that SFEVs significantly decreased beat cross frequency and increased distance straight line, linearity, straightness, straight line velocity, and wobble. SFEVs did not influence sperm capacitation status or the ability of sperm to undergo acrosomal exocytosis. Functional assessment of both high- and low-quality spermatozoa collected prior to liquefaction showed limited SFEV influence, with these vesicles inducing only subtle decreases in beat cross frequency in spermatozoa of both groups. These findings raise the prospect that, aside from subtle effects on sperm motility, the encapsulated SFEV cargo may be destined for physiological targets other than the male germline, notably the female reproductive tract. A male's influence over the biological processes of pregnancy extends beyond the provision of sperm. Molecular signals present in the ejaculate can influence the likelihood of pregnancy and healthy pregnancy progression, but the identity and function of these signals remain unclear. In this study, we wanted to understand if nano-sized particles present in the male ejaculate, called seminal fluid extracellular vesicles, can assist sperm in traversing the female reproductive tract to access the egg. To explore this, we isolated seminal fluid extracellular vesicles from human semen and incubated them with sperm. Our data showed that seminal fluid extracellular vesicles act to transfer molecular information to sperm, but this resulted in only subtle changes to the movement of sperm.

Aquaporin-3a Dysfunction Impairs Osmoadaptation in Post-Activated Marine Fish Spermatozoa.

Spermatozoon volume regulation is an essential determinant of male fertility competence in mammals and oviparous fishes. In mammals, aquaporin water channels (AQP3, -7 and -8) have been suggested to play a role in spermatozoon cell volume regulatory responses in the hypotonic female oviduct. In contrast, the ejaculated spermatozoa of marine teleosts, such as the gilthead seabream (Sparus aurata), experience a high hypertonic shock in seawater, initially resulting in an Aqp1aa-mediated water efflux, cell shrinkage and the activation of motility. Further regulatory recovery of cell volume in post-activated spermatozoa is mediated by Aqp4a in cooperation with the Trpv4 Ca2+ channel and other ion channels and transporters. Using a paralog-specific antibody, here, we show that seabream spermatozoa also express the aquaglyceroporin AQP3 ortholog Aqp3a, which is highly accumulated in the mid posterior region of the spermatozoon flagella, in a similar pattern to that described in mouse and human sperm. To investigate the role of Aqp3a in seabream sperm motility, we used a recently developed AQP3 antagonist (DFP00173), as well as the seabream Aqp3a-specific antibody (α-SaAqp3a), both of which specifically inhibit Aqp3a-mediated water conductance when the channel was heterologously expressed in Xenopus laevis oocytes. Inhibition with either DFP00173 or α-SaAqp3a did not affect sperm motility activation but did impair the spermatozoon motion kinetics at 30 s post activation in a dose-dependent manner. Interestingly, in close resemblance to the phenotypes of AQP3-deficient murine sperm, electron microscopy image analysis revealed that both Aqp3a inhibitors induce abnormal sperm tail morphologies, including swelling and angulation of the tail, with complete coiling of the flagella in some cases. These findings suggest a conserved role of Aqp3a as an osmosensor that regulates cell volume in fish spermatozoa under a high hypertonic stress, thereby controlling the efflux of water and/or solutes in the post-activated spermatozoon.

HLA-DQB1 as a potential prognostic biomarker of hormonal therapy in patients with non-obstructive azoospermia

The gonadotropin treatment of infertile men may improve spermatogenesis and lead to sperm cell production, however, only a small fraction of treated patients positively responds to such therapy. To identify individual treatment prognostic biomarkers associated with responsiveness to gonadotropins, we compared the gene expression profiles of testicular oligobiopsies from 3 patients with non-obstructive azoospermia (NOA) who positively responded to therapy with a combination of human chorionic gonadotropin and recombinant follicle-stimulating hormone (hCG/rFSH) to those of 3 non-responders. We used Affymetrix Human Gene 1.0 ST microarrays. The results of the microarray evaluation were validated by the qPCR technique while gene variants of the HLA-DQB1 (major histocompatibility complex, class II, DQ beta 1) were subsequently sequenced. In our microarrays, we have identified most significantly 5 transcripts with different expression levels in responders versus non-responders groups. Our interest has been primarily focused on the transcript associated with the HLA-DQB1 gene. Because the expression of this gene was up-regulated in the non-responding patients and only patients with heterozygotic alleles of HLA-DQB1 turned out to be positive to gonadotropin therapy, we suggest that this gene may be a biomarker of potential significance for the gonadotropin treatment of male infertility. We also compared the testicular gene expression profile in one individual before and after gonadotropin treatment. In the re-biopsied sample, we have identified over 600 genes that showed differences in testicular expression; some of these genes are critical for spermiogenesis. Thus, we documented that the applied gonadotropins successfully stimulated the spermatogenetic wave in patients with NOA.

Supplementation of sperm cooling medium with Eurycoma longifolia extract enhances native Thai chicken sperm quality and fertility potential.

Cooled semen storage methods result in oxidative stress generated by an imbalance between oxidation rates, specifically reactive oxygen species production, and sperm cell antioxidants, leading to degradation of semen quality. We aimed to investigate the impact of adding Eurycoma longifolia (EL) extract as an antioxidant supplement in semen storage medium (IGGKPh semen extender) on semen quality and fertility potential. EL extract at concentrations of 5, 10, 15, and 20 mg/mL was assessed for its antioxidant capacity in IGGKPh semen extender. Our findings revealed that the total phenolic content in the EL extract did not vary significantly across the various concentrations and temperatures tested. However, incubation at 5°C was found to be the most effective temperature for increasing the EL extract antioxidant capacity as assessed via the 2,2-diphenyl-1-picrylhydrazyl inhibition assay in a dose-dependent manner. Supplementation of the IGGKPh semen extender with 15 mg/mL EL extract was found to enhance semen quality during cold storage for up to 48 h (p < 0.05), as evidenced by decreased malondialdehyde levels in cooled semen (p < 0.05). However, antioxidant enzyme activities showed no significant differences among the various experimental groups (p > 0.05). The fertility test showed that the 15 mg/mL EL extract group stored for 24 h had a higher percentage than the control group (p < 0.05). However, there was no significant difference in percentage between the two groups at 48 h of storage (p > 0.05). The hatchability showed no significant difference in both 24 and 48-h storage periods (p > 0.05). Our results indicated that supplementing the IGGKPh semen extender with 15 mg/mL EL extract may positively influence semen quality during storage, suggesting potential applications for enhancing semen quality.

The BNB-GLID module regulates germline fate determination in Marchantia polymorpha.

Germline fate determination is a critical event in sexual reproduction. Unlike animals, plants specify the germline by reprogramming somatic cells at the late stages of their development. However, the genetic basis of germline fate determination and how it evolved during the land plant evolution are still poorly understood. Here, we report that the plant homeodomain finger protein GERMLINE IDENTITY DETERMINANT (GLID) is a key regulator of the germline specification in liverwort, Marchantia polymorpha. Loss of the MpGLID function causes failure of germline initiation, leading to the absence of sperm and egg cells. Remarkably, the overexpression of MpGLID in M. polymorpha induces the ectopic formation of cells with male germline cell features exclusively in male thalli. We further show that MpBONOBO (BNB), with an evolutionarily conserved function, can induce the formation of male germ cell-like cells through the activation of MpGLID by directly binding to its promoter. The Arabidopsis (Arabidopsis thaliana) MpGLID ortholog, MALE STERILITY1 (AtMS1), fails to replace the germline specification function of MpGLID in M. polymorpha, demonstrating that a derived function of MpGLID orthologs has been restricted to tapetum development in flowering plants. Collectively, our findings suggest the presence of the BNB-GLID module in complex ancestral land plants that has been retained in bryophytes, but rewired in flowering plants for male germline fate determination.

CORRELATION BETWEEN BMI AND GONADOTROPIN HORMONES IN THE FEMALE GENDER

Introduction: The pituitary gland affects growth and development. Its main function is in the activity of sexual/reproductive hormones and in the functioning of the glands (thyroid gland, adrenal glands and gonads). Gonadotropin hormones (gonadostimulins) are so named because they stimulate the function of the gonads - male and female sex glands (testicles and ovaries) in which sex cells - sperm and egg cells are created. Overweight is a state of metabolic disorders where excess body fat accumulates to the extent that it can negatively affect human health. Obesity as a BMI factor is related to the functioning of the thyroid gland with its hormones and the pituitary gland with gonadotropin hormones. Purpose of the study: This paper aims to present the correlation between BMI and gonadotropin hormones in reproductive and postmenopausal women. The aim of the paper will also consist in the status of gonadotropin hormones and the functional state of these hormones by comparing them with metabolic disorders such as BMI, obesity and the synthesis of follicle stimulating hormone and gonadotropin hormone. Material and method: A total of 20 patients will be included in the research, divided according to age group into two categories. The first group will include patients aged 10-40 years, the second group patients over 41 years old. Blood (serum) samples will be taken from patients for analyzing gonadotropin hormones: FSH, LH, PRL and thyroid hormones: TSH, FT4, FT3. They will be analyzed with the modern automatic fluorescent enzyme immunoassay method (Vidas-Biomerie). The BMI of the body will be measured with a metric formula where the weight of the patient (in kg) divided by the height of the patient (in m2) set to the square power will give us the measure of body index-BMI (%). SPSS – Software package version 20.0 Numerical series were analyzed using central tendency measures (average, median, minimum values, maximum values), as well as distribution measures (standard deviation); To determine the regularity of the frequency distribution of the examined variables, the Shapiro-Wilk W Test was used; Non-parametric tests for two independent parameters were used to test the significance and difference between some numerical parameters with irregular frequency distribution (Mann Whitney U test); To determine statistical significance, a two-way analysis was used with a significance level (means) of p&lt;0.05. Results: The results of the research will give a real insight into the data of gonadotropic hormones FSH, LH, PRL and thyroid hormones TSH, FT4, FT3 of the female sex in the Tetova region. The results of the research will also provide information about the correlation of BMI and gonadotropin and thyroid hormones. Conclusion: From the analyzes performed on patients of different age groups we come to the conclusion that: • Gonadotropin hormonal disorders can lead to the phenomenon of metabolic diseases such as BMI. • These disorders can have consequences such as: polycystic ovary diseases in women, Hashimoto's genetic syndrome, Graves, infertility in women, etc.

The involvement of lysine glutarylation and mitochondria in spermatic movement deficiency induced by hexavalent chromium

Chromium (Cr) and hexavalent chromium Cr(VI) compounds are extensively utilized in various industrial processes, leading to significant environmental pollution. Animal studies have indicated that Cr(VI) can easily penetrate into male germ cells, thereby impairing male fertility. However, there is limited information on how Cr(VI) impacts human spermatozoa functions and the mechanisms involved. This study aims to elucidate the impact of Cr(VI) on human spermatozoa functions and the underlying mechanisms involved. We observed that K2CrO4 inhibited human spermatozoa motility in vitro, with the inhibition being positively correlated with both the concentration of K2CrO4 and the duration of exposure. Additionally, K2CrO4 was found to reduce lysine glutarylation (Kglu) levels and increase the accumulation of reactive oxygen species (ROS) within the cells, leading to mitochondrial depolarization and a subsequent decrease in adenosine triphosphate (ATP) content. Interestingly, sodium glutarate (Na-glu), a precursor of glutaryl-CoA, demonstrated the ability to partially ameliorate the detrimental effects of K2CrO4 on spermatozoa motility. These results suggest that the environmental pollutant Cr(VI) may impair human spermatozoa motility by disrupting lysine glutarylation and mitochondrial functions.

Development of a Novel Male Reproductive Toxicity Assessment Method to Predict Male-mediated Effects on the Next Generation

Less than 10% of the candidate drug compounds are associated with male reproductive toxicity. Genetic and/or epigenetic information on sperm may be crucial for fetal development. Therefore, developmental toxicity, such as paternally transmitted birth defects, is possible if genetic abnormalities in the male germ line persist and accumulate in the sperm during spermatogenesis. First, this study provides an overview of chemical and male reproductive toxicity, which may lead to developmental toxicity from the perspective of male reproduction. Second, we demonstrate methods for evaluating male reproductive toxicity to anticipate male-mediated developmental toxicity. We developed a novel staining technique for evaluating sperm quality, as well as a noninvasive imaging analysis of male reproductive toxicity. The former is a mammalian male germ cell-specific staining method using reactive blue 2 dye (RB2), as previously confirmed in human sperm, and a method for detecting the early-stage DNA fragmentation in a single nucleus from mouse spermatozoa using single-cell pulsed-field gel electrophoresis. The latter is a new, ready-to-use, and compact magnetic resonance imaging (MRI) platform utilizing a high-field permanent magnet to evaluate male reproductive toxicity. The histopathological analysis supported the suitability of the MRI platform. The present study, for the first time, revealed a rapid, noninvasive evaluation of male reproductive toxicity in vivo using compact MRI. These novel toxicity assessments can help predict male-mediated developmental toxicity, contributing to accelerated drug discovery and drug repositioning.

Immunofluorescence and biochemical investigation of the protective effects of naringin and diosmin on the freezability of Merino ram semen

BACKGROUND: Various antioxidant substances are added to sperm extenders to protect spermatozoa against oxidative stress and cryodamage. OBJECTIVE: To investigate the effects of the flavonoid diosmin (DIO) and a flavanone glycoside naringin (NAR) on the freezability of ram semen. MATERIALS AND METHODS: In this study, six Merino rams were used during the breeding season. The ejaculates were pooled after collection from the rams. Pooled ejaculates were divided into six groups: control, NAR 1 mM, NAR 2 mM, NAR 4 mM, DIO 2 mM, and DIO 4 mM, and then diluted with a TRIS-based diluent. The pooled semen was equilibrated, placed in 0.25 mL pipettes with 10 × 107 sperm cells in each pipette, and frozen in liquid nitrogen vapor. After 24 h, the pipettes were thawed at 37°C for 25 s and analyzed in terms of spermatological parameters. RESULTS: The highest plasma membrane integrity ratio was found in the DIO 4 mM group, whereas a statistically significant difference was found between the NAR 1 mM and NAR 2 mM groups (p&lt;0.05). While the DIO 4 mM group had the highest acrosome integrity rate, a statistically significant difference was found between the other groups (p&lt;0.05). Mitochondrial activity was the highest in the NAR 4 mM, DIO 4 mM and DIO 2 mM groups (p&lt;0.05). In the analysis of the sperm membrane lipid profile, it was observed that the DIO group had the highest lipid-phospholipid ratio. In sperm membrane protein profile analysis, it was found that both additives exerted protective effects at different levels. The highest total protein content was seen in the DIO 4 mM and NAR 4 mM groups. 8-hydroxydeoxyguanosine (8-OhDG) positivity was more common in the control group than in the DIO and NAR groups. Cu-Zn superoxide dismutase (SOD) expression was lower in the control group and more intense in all other groups. Positive results were especially observed in the acrosome of the sperm cells. CONCLUSION: The addition of NAR and DIO to the ram semen extender increased the quality of sperm parameters after the freeze-thaw process.

Protective Effect of Aloe vera (L.) on Diabetes-Induced Oxidative Stress Linked Spermiological Co-Morbidity in Human and Rat: An In-Vitro Analysis.

Diabetes linked reproductive complications are rising problems nowadays. The study focused on the protective efficacy of Aloe vera (L.) on sperm cell damage in an oxidative stress milieu encumbered by a chronic diabetes in human and streptozotocin treated Wistar rat (Rattus norvegicus). Spermatozoa from rat's epididymal washing, and human semen after 3-4days of abstinence of mating or masturbation were collected from control and diabetes groups. Spermatozoa of human and rat were incubated for 1 or 2h at 370C in an in-vitro medium separately and considered as normo-glycemic control and diabetes sub-groups. Dose of 1 or, 2 or, 4mg/ml of Aloe vera (L.) hydro-ethanolic (40:60) extract (AVHE) was given to diabetes samples, considered as sub-sub-group for assessing its protective effect on spermiological and oxidative stress parameters. The motility, viability, plasma membrane integrity, nuclear chromatin decondensation for DNA fragmentation, acrosome cap status, and antioxidative status of human and rat spermatozoa were decreased whereas spermatozoal apoptosis was elevated significantly (p < 0.05), noted by TUNEL assay in diabetes samples compared to the duration-matched control group. Exposure of AVHE to diabetes samples resulted significant rectification (p < 0.05) in the said parameters than the unexposed diabetes group. In control group, AVHE exposure has significant protective effect from spermiological deterioration compared to unexposed control group. Identification of major phytomolecules in AVHE was done by LC-MS study. Diabetes-induced oxidative stress-mediated spermatozoal injuries can be protected by AVHE extract, raise the possibility for potentiating sperm of human for increasing the success rate of in-vitro fertilization-blastocyst implantation.

Male Gamete Cryopreservation for Adolescent Fertility Preservation: Our Experience to Date

Male Gamete Cryopreservation for Adolescent Fertility Preservation: Our Experience to Date

The Sperm-Associated Antigen 11A (Spag11a) Knockout Mice Display Sub-Fertility and Perturbations in the Sperm Proteome.

Spermatogenesis and sperm maturation that occur in the testis and epididymis respectively are crucial for fertility. Factors secreted by the testicular and epididymal epithelial cells govern the processes of gametogenesis and maturation. Proteins encoded by the sperm-associated antigen 11a (Spag11a) gene are implicated as having a possible role in sperm maturation. However, studies that demonstrate their definite role in fertility and sperm function using knockout models have not yet reported. In this study, Spag11a knockout mice were generated, genotyped and the reproductive parameters (fecundity, sperm count, capacitation, and acrosome reaction) and sperm proteome were determined. Litter size and sperm count were decreased in the Spag11a knockout mice when compared to the wild-type controls. Spermatozoa from the knockout mice were able to undergo capacitation. However, acrosome reaction did not occur in sperm obtained from knockout mice. Structural abnormalities in the head and tail structures were evident in the spermatozoa of knockout mice. Perturbations in the expression of sperm proteins that are involved in gametogenesis were evident. The subfertility observed in Spag11a knockout mice could be a manifestation of lower sperm count, impaired acrosome reactions, and disturbances in the sperm proteome. The results of this study lend further support to the role of Spag11a gene in male gamete function.