Polymorphism detection and characterization of sperm cells chromatin remodeling associated genes in Murrah buffalo.

Seasonal variations significantly impact buffalo bull semen production and quality, particularly during the summer months. Understanding the genetic basis of these changes is important for managing bull fertility and improving sperm quality. The present study focused on characterizing and identifying polymorphisms in chromatin remodeling genes, protamines (PRMs) and Transition Nuclear Proteins (TNPs) in Murrah buffalo bulls with varying semen quality due to seasonal effects. Our findings revealed none of the coding region variation in PRM1, PRM2, TNP1, and TNP2, these genes are highly conserved in buffalo. Two intronic variants were identified, including G16C in PRM1 intron 1 and intronic SNP in PRM2 intron 1 (G96A). The complete CDS of consensus sequence of bubaline PRM1 was 86.3% identical and 94.1% similar to the bovine PRM1. Whereas the complete CDS of consensus sequence of bubaline TNP2 was 78.2% identical and 91.0% similar to bovine TNP2. Further, no statistically significant differences in the fold change of TNP1, TNP2, PRM1, and PRM2 levels between the hot summer SNA and SA groups and the winter SNA and SA groups This study represents the first comprehensive report on the characterization of bubaline PRM1 (complete CDS), PRM2 (partial CDS), TNP1 (partial CDS), and TNP2 (complete CDS) genes in buffalo sperm cells. Results of the study, clearly indicate that the genes associated with protamine (PRM1 and TNP2) are highly conserved in Bubalus bubalis. Understanding these genetic underpinnings can have implications for improving buffalo bull fertility and semen quality.

Improvement of Post-Thaw Quality and In Vivo Fertility of Simmental Bull Spermatozoa Using Ferulic Acid.

BACKGROUND: Artificial insemination and semen cryopreservation have significantly improved the quality and quantity of cattle production. Through cryopreserved semen and artificial insemination, top-breeding bull sperm can be used to inseminate thousands of cows worldwide. OBJECTIVES: Our study aimed to determine the effect of adding ferulic acid (FA) to a Tris-based semen extender on frozen and thawed Simmental bull sperm. METHODS: Semen samples were collected from three Simmental bulls. Pooled Simmental semen (n = 34 ejaculations) were diluted with a Tris-base extender containing varying FA concentrations (0.1, 0.15, 0.25, 0.35, and 0.45 mM). After the samples were frozen and thawed, the samples were tested for malondialdehyde (MDA), total antioxidant capacity (TAC), superoxide dismutase (SOD), glutathione peroxidase (GPx), total motility, progressive motility, motility characteristics, and plasma membrane functionality. RESULTS: The control and the groups with the best FA concentrations, 0.25 and 0.35, were compared for in vivo fertility. Fifty-one cows were inseminated 24 h after the onset of oestrus. A rectal examination was used to diagnose pregnancies at least 60 days after fertilization. Results showed that adding FA-0.45, FA-0.35, FA-0.25, and FA-0.15 to the semen of Simmental bulls improved total and progressive motility, motility characteristics, and plasma membrane functionality. It also increased GPx and TAC levels, reducing MDA and DNA damage after freezing. The addition of FA did not affect SOD values. The fertility rate in the FA-0.25 and FA-0.35 groups was higher than in the control group, 35.29%, with rates of 76.47% and 70.58%, respectively. CONCLUSIONS: In conclusion, adding FA (0.15, 0.25, 0.35, and 0.45 mM) to Tris-based semen extenders can improve the quality parameters of cryopreserved Simmental bull semen and increase in vivo fertility using 0.25 and 0.35 concentrations of FA.

Role of Leukocytospermia in the Management of Male Infertility: Decoding a Mystery for the Busy Clinicians.

Leukocytospermia, defined as a leukocyte concentration in semen exceeding 1×106 leukocytes/mL, significantly impacts male reproductive health by affecting sperm functionality and fertility outcomes. This condition arises from various etiological factors, including infections, autoimmune responses, lifestyle factors, and other physiological conditions. Adverse effects on sperm associated with leukocytospermia include acrosome damage and abnormalities in the sperm midpiece and tail. The review explores the complex interplay between leukocytospermia and oxidative stress, emphasizing the harmful effects on sperm DNA integrity and overall sperm quality. Due to the multifactorial nature of leukocytospermia, diagnosing this condition presents several challenges. Effective management strategies discussed include the use of antibiotics, anti-inflammatory agents, and assisted reproductive technologies. Diagnostic methods range from traditional peroxidase staining to more advanced techniques such as immunocytochemistry and flow cytometry, which offer higher sensitivity and specificity. Infections of the male genital tract, particularly male accessory gland infection and male genital tract infection, play a significant role in the etiology of leukocytospermia. These infections lead to an inflammatory response, resulting in leukocyte infiltration into the semen. Systemic conditions like diabetes mellitus and autoimmune disorders also contribute to leukocytospermia by provoking inflammatory responses that facilitate leukocyte presence in semen. This review underscores the importance of a comprehensive diagnostic approach that includes patient history, physical examination, and advanced laboratory tests. Treatment is tailored to the identified underlying cause, whether infectious or non-infectious. Lifestyle modifications, such as reducing stress, improving diet, and avoiding environmental toxins, are also recommended to enhance semen quality. For clinicians, this review provides a concise yet thorough overview of leukocytospermia, integrating the latest research findings and clinical insights to aid in the effective management of this condition, ultimately aiming to improve patient care in male reproductive health.

Cross-ancestry meta-genome-wide association studies provide insights to the understanding of semen traits in pigs.

Semen traits play a crucial role in pig reproduction and fertility. However, limited data availability hinder a comprehensive understanding of the genetic mechanisms underlying these traits. In this study, we integrated 597 299 ejaculates and 3 596 sequence data to identify genetic variants and candidate genes related to four semen traits, including sperm progressive motility (MOT), semen volume, sperm concentration (CON), and effective sperm count (SUM). A cross-ancestry meta-genome-wide association study was conducted to detect 163 lead single nucleotide polymorphisms (SNPs) associated with MOT, CON, and SUM. Subsequently, transcriptome-wide association studies and colocalisation analyses were integrated to identify 176 candidate genes, many of which have documented roles in spermatogenesis or male mammal semen traits. Our analysis highlighted the potential involvement of CSM5, PDZD9, and LDAF1 in regulating semen traits through multiple methods. Finally, to validate the function of significant SNPs, we performed genomic feature best linear unbiased prediction in 348 independent pigs using identified trait-related SNP subsets as genomic features. We found that integrating the top 0.1, 1, and 5% significant SNPs as genomic features could enhance genomic prediction accuracy for CON and MOT compared to traditional genomic best linear unbiased prediction. This study contributes to a comprehensive understanding of the genetic mechanisms of boar semen traits and provides insight for developing genomic selection models.

Dietary alpha-linolenic acid supplementation enhances semen quality, antioxidant capacity, and sperm survival in aging breeder roosters.

Aging in breeder roosters is often accompanied by a decline in semen quality, negatively impacting reproductive performance. This study aimed to investigate the effect of dietary alpha-linolenic acid (ALA), an essential omega-3 polyunsaturated fatty acid, on semen quality, antioxidant capacity, and sperm survival in aging breeder roosters. Roosters were divided into 4 groups and fed diets supplemented with 0%, 0.5%, 1%, and 2% ALA for 6 wk. Results indicated significant improvements in semen volume, sperm viability, and sperm density in ALA-supplemented groups compared to the control (P < 0.05). The 1% ALA group exhibited the most notable enhancements in sperm viability and density. Additionally, ALA supplementation increased the activities of antioxidant enzymes such as superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and reduced malondialdehyde (MDA) levels, indicating enhanced antioxidant capacity (P < 0.05). Furthermore, ALA improved mitochondrial membrane potential (MMP) and reduced early and late sperm apoptosis, with the 2% ALA group showing the highest MMP and the lowest ROS-positive rate (P < 0.05). These findings suggest that dietary ALA supplementation enhances semen quality and antioxidant defenses, and mitigates oxidative stress, thus supporting the reproductive health of aging breeder roosters. This study underscores the potential of ALA as a dietary strategy to improve reproductive efficiency in poultry production.

The Effects of Adding Hempseed Cake on Sperm Traits, Body Weight, Haematological and Biochemical Parameters in Rabbit Males.

Incorporating of agro-industrial co-products into animal nutrition could represent an opportunity to lessen the environmental impact of the food production chain. One such co-product is a hempseed cake originating from cold pressing hemp seeds to extract oil for human consumption. The aim of the present study was to evaluate the action of hempseed cake in the diet on male rabbit reproductive and some non-reproductive indexes. Male rabbits were fed either a standard diet (control group; C; n = 10) or a diet enriched with hempseed cake (experimental group E5 with 5% of a hempseed cake; n = 10, and experimental group E10 with 10% of a hempseed cake; n = 10) in 100 kg of the milled complete feed mixture. Rabbit weight gain, sperm concentration, motility, progressive motility, and sperm quality were evaluated using CASA and flow cytometry. Feeding with a hempseed cake, given at both tested concentrations, had no effect on weight gain per week and the total average weight gain compared to the control group (p > 0.05). Hempseed cake addition had no effect on sperm concentration in ejaculate, sperm motility, and progressive motility (p > 0.05). Selected haematological and biochemical indexes were examined. The E5 group showed positive tendencies in hepatic profile parameters, while in the E10 group the tendencies were opposite, though within the reference values. Based on our results, no negative effects of hempseed cake feeding on rabbit reproduction and health status were found, and we can recommend the use of hempseed cake at doses up to 10% in the nutrition and feeding of rabbits. Therefore, agro-industrial co-products can decrease the feeding cost.

Trifluoperazine effect on human sperm: The accumulation of reactive oxygen species and the decrease in the mitochondrial membrane potential.

A strong link between antipsychotic drug use and reduced human sperm quality has been reported. Trifluoperazine (TFP), a commonly used antipsychotic, is now being explored for anticancer applications. Although there are hints that TFP might affect the male reproductive system, its impact on human sperm quality remains uncertain. Using a human sperm and TFP in vitro coculture system, we examined the effect of TFP (12.5, 25, 50 and 100 µM) on human sperm function and physiological parameters. The results showed that 50 µM and 100 µM TFP induced the accumulation of reactive oxygen species (ROS) and a decrease in the mitochondrial membrane potential (MMP) of human sperm, leading to decreased sperm viability, while 25 µM TFP inhibited only the penetration ability, total sperm motility, and progressive motility. Although 12.5 µM and 25 µM TFP increased [Ca2+]i in human sperm, they did not affect capacitation or the acrosome reaction. These results may be explained by the observation that 12.5 µM and 25 µM TFP did not increase tyrosine phosphorylation in human sperm, although TFP increased [Ca2+]i in a time-course traces similar to that of progesterone. Our results indicated that TFP could cause male reproductive toxicity by inducing the accumulation of ROS and a decrease in the MMP in human sperm.

Association Between Dietary Habits and the Semen Quality of South Asian Men Attending Fertility Clinic: A Cross-sectional Study.

The purpose of the study is to explore the impact of dietary habits, including daily intake of nutrients and dietary patterns, on the semen characteristics of South-Asian men. This cross-sectional study included 162 South-Asian men aged 21-40 years who were undergoing infertility evaluation. Based on semen analysis, men were grouped into categories of normal and abnormal semen characteristics. The participants' dietary habits were captured using '24-hour dietary recall' and 'Food Frequency Questionnaire (FFQ)'. Daily dietary intake of various nutrients and adherence to prevalent dietary patterns were compared between men with normal and abnormal semen characteristics. Six dietary patterns were identified following factor analysis of FFQ data. Men with normal semen characteristics showed significantly higher consumption of "coastal diet" rich in seafood, fish, cereals, beans, and coconut oil than those with abnormal semen characteristics. Further, the multivariable-adjusted model, adjusted for clinically relevant confounders, like age, body mass index (BMI), physical activity, smoking and sexual abstinence, also showed a lower risk of abnormal semen characteristics among men consuming "coastal diet" (odds ratio [OR] = 0.56, 95% confidence interval [CI] 0.37-0.82). In terms of daily nutrient intake, men with normal semen characteristics showed significantly lower consumption of monounsaturated fatty acids (MUFA) compared to men with abnormal semen characteristics (p = 0.008). Our findings suggest that higher consumption of "coastal pattern diet" lowers the risk of abnormal semen characteristics. Additionally, a negative association of MUFA consumption with semen quality was also identified, which needs cautious interpretation until replicated in future studies with biochemical validations.

Determinant genetic markers of semen quality in livestock.

The reproductive efficiency of livestock is crucial for agricultural productivity and economic sustainability. One critical factor in successful fertilization and the viability of offspring is the quality of semen. Poor semen quality, especially in frozen-thawed semen used in artificial insemination (AI) have been shown to influence conception outcomes, resulting a negative impact on livestock production. Recent advancements in genetic research have identified specific markers linked to semen quality traits in various livestock species, such as cattle, sheep, goats, pigs, buffalo, and equines. These genetic markers are essential in screening males for breeding suitability, which in turn enhances selective breeding programs. Understanding these markers is crucial for improving reproductive performance and increasing productivity in livestock populations. This review offers a comprehensive overview of the genetic markers associated with semen quality in key livestock. It explores the underlying genetic mechanisms and their practical implications in animal breeding and management. The review underscores the importance of integrating genetic insights into breeding strategies to optimize reproductive efficiency and ensure the sustainable development of livestock industries.

The moderation effect of GSTM1/GSTT1 gene polymorphisms on the association of sperm mitochondrial DNA copy number and sperm mobility.

Oxidative stress (OS) is believed to be a significant factor in the decline of semen quality, with mitochondrial DNA copy number (mtDNAcn) serving as a sensitive biomarker for both semen quality and mitochondrial dysfunction resulting from oxidative stress. While glutathione S-transferases (GSTs) are commonly known as 'antioxidant' enzymes, there is ongoing debate regarding the relationship between GST genotypes and semen quality. In a study involving 568 male volunteers from the outpatient department of Puyang Reproductive Medicine Center, sperm mtDNAcn, semen quality, and GSTM1/GSTT1 genotypes were analyzed to investigate the potential link between GSTM1/GSTT1 gene variations and semen quality, as well as the impact of GSTs gene variations on the connection between sperm mtDNAcn and semen quality. Adjusting for variables such as age, BMI, smoking, and alcohol consumption, it was found that mtDNAcn was significantly correlated with decreased sperm concentration and total sperm count (b = - 0.109, - 0.128, respectively; P = 0.002, 0.001, respectively). GSTM1 was associated with progressive motility (OR 0.390, 95% CI 0.218, 0.697), Straight line velocity (VSL) (OR = 0.606, 95% CI 0.385, 0.953), and Straightness (STR) (OR 0.604, 95% CI 0.367, 0.994), while GSTT1 was linked to progressive motility (OR 0.554, 95% CI 0.324, 0.944) and Beat crossover frequency (OR 0.624, 95% CI 0.397, 0.982). The GSTT1 was found to moderate the relationship between mtDNAcn and sperm motility parameters linearity (LIN), STR, and Wobble (WOB), with additive interaction effects observed between GSTT1 and mtDNAcn on LIN, STR, and WOB (P for interaction = 0.008, 0.034, 0.010, respectively). Overall, this study suggests that GSTT1 and GSTM1 gene variations may play a role in sperm motility, with GSTT1 potentially influencing the impact of oxidative stress on sperm motility.

Effect of paternal overweight or obesity on semen parameters, clinical pregnancy and live birth outcomes in men treated with intrauterine insemination (IUI).

Background: Overweight and obese individuals are steadily increasing in recent years. Male overweight or obesity has adverse impact on reproductive functions. The study aimed to evaluate the potential impact of paternal overweight or obesity on sperm quality and clinical pregnancy outcome in patients undergoing intrauterine insemination (IUI) treatment. Methods: This retrospective study included 1,036 couples from our reproductive center between July 2019 and August 2022. All males were categorized into normal weight, overweight, or obese groups according to their body mass index (BMI). Baseline characteristics and reproductive hormones were analyzed. Semen parameters, clinical pregnancy and live birth outcomes were compared among the different BMI groups. Results: There were no significant differences in sperm concentration, total sperm motility, progressive sperm motility, normal sperm morphology and sperm DNA fragmentation index (DFI) among the three groups. However, the obese group exhibited a significantly decreased semen volume compared to the other two groups (P<0.01). No differences were found in clinical pregnancy rate (CPR), abortion rate (AR) and live birth rate (LBR) among the groups (P>0.05). Slight higher ARs were observed in overweight and obese groups compared to normal group (13.64%, 21.05% vs. 11.11%, P=0.49). Conclusions: These data suggest that male obesity leads to a significant decrease in semen volume. It is thus recommended that male BMI should be regarded as one of the predictors for IUI treatment to avoid a decrease in semen volume.

Exploring Myrciaria dubia liquid extract as a potential semen extender for breeding roosters.

The current investigation aimed to explore the effects of Myrciaria dubia liquid extract (MDLE) as the primary component of an extender for breeder rooster semen over different periods at room temperature. Fifteen breeder roosters (40 weeks of age, average body weight of 2.05±0.12) with confirmed fertility were used. Employing a factorial design (3x4), the treatments consisted of semen in natura and two semen extenders (an experimental based on MDLE and a commercial) subjected to four periods at room temperature post-collection (5, 10, 15 and 20 minutes) with four replicates (tubes) each. All variables evaluated in this study yielding significant results (p<0.05). Analyzed individually, the experimental extender based on MDLE exhibited a linear reduction (p<0.05) in motility and vigor results, while it caused an increase in pH values and percentages of sperm defects evaluated. When compared with semen in natura and commercial extender, the efficiency of MDLE as a semen extender was inferior to that observed with the commercial extender and similar to the results observed with semen in natura. Nonetheless, the experimental extender based on MDLE yielded satisfactory results for up to 15 minutes of storage time. In conclusion, MDLE can be considered as an alternative for composing a roosters' semen extender, maintaining sperm characteristics within acceptable limits for up to 15 minutes at room temperature. However, this experimental extender demonstrated lower efficiency than the commercial extender in maintaining the sperm quality at room temperature across all periods tested.

Exploring the association between urinary bisphenol A, S, and F levels and semen quality parameters: Findings from Led-Fertyl cross-sectional study.

Infertility is recognized as a multifaceted condition affecting approximately 15% of couples globally, influenced by various factors including genetic predisposition and environmental exposures. Among these environmental factors, bisphenol A (BPA) emerges as a prominent Endocrine-disrupting chemical (EDCs) widely distributed, leading to chronic human exposure in daily life. As regulations on BPA became more stringent, alternative substances such as bisphenol S (BPS) and bisphenol F (BPF) have emerged. Animal studies have demonstrated a dose-dependent decline in fertility and embryotoxicity following chronic exposure to BPA. However, literature data on human studies are limited and heterogeneous. Additionally, even less is known about the relationship between exposure to the BPA analogues (BPS and BPF) and sperm quality. Therefore, the present study aimed to examine the association between urinary concentrations of BPA, BPF, and BPS and semen quality parameters among 195 adult Spanish men from the Led-Fertyl study cohort using multiple linear regression models adjusted by potential confounding variables. Our results revealed an inverse association between log-transformed creatinine-adjusted concentration (ng/mg) of BPA and BPF levels and the percentage of sperm vitality (ß: 3.56 %; 95%CI: 6.48 to -0.63 and ß: 4.14 %; 95%CI: 6.97 to -1.31; respectively). Furthermore, participants in the highest quartile of BPA and BPF urinary concentration exhibited lower sperm vitality compared to those in the lowest quartile (ß: 6.90 %; 95%CI: 11.60 to -2.15 and ß: 9.68 %; 95%CI: 14.43 to -4.94; respectively). These results supply epidemiological evidence establishing a relationship between bisphenols urine exposure and sperm quality, suggesting that a re-evaluation of the overall safety of BPA alternatives is warranted.

Alginate oligosaccharide supplementation improves boar semen quality under heat stress.

Heat stress is a serious problem that affects animal husbandry by reducing growth and reproductive performance of animals. Adding plant extracts to the diet is an effective way to help overcome this problem. Alginate oligosaccharide (AOS) is a natural non-toxic antioxidant with multiple biological activities. This study analyzed the potential mechanism of AOS in alleviating heat stress and improving semen quality in boars through a combination of multiple omics tools. The results indicated that AOS could significantly increase sperm motility (P < 0.001) and sperm concentration (P < 0.05). At the same time, AOS improved the antioxidant capacity of blood and semen, and increased blood testosterone (P < 0.05) level. AOS could improve the metabolites in sperm, change the composition of gut microbiota, increase the relative abundance of beneficial bacteria such as Pseudomonas (P < 0.01), Escherichia-Shigella (P < 0.05), Bifidobacterium (P < 0.01), reduce the relative abundance of harmful bacteria such as Prevotella_9 (P < 0.05), Prevotellaceae_UCG-001 (P < 0.01), and increase the content of short chain fatty acids. Proteomic results showed that AOS increased proteins related to spermatogenesis, while decreasing heat shock protein 70 (P < 0.05) and heat shock protein 90 (P < 0.01). These results were verified using immunofluorescence staining technology. There was a good correlation among sperm quality, sperm metabolome, sperm proteome, and gut microbiota. In conclusion, AOS can be used as a feed additive to increase the semen quality of boars to enhance reproductive performance under heat stress.

Intensity and type of physical activity and semen quality in healthy young men.

OBJECTIVE: To study the relationship between intensity of physical activity (PA) and semen quality in healthy young men. DESIGN: A prospective cohort study with repeated measures for each subject. SETTING: Not applicable. PATIENT(S): Healthy high school and university students who did not regularly smoke tobacco, drink alcohol, or take drugs or medicine, with normal body mass index and abdominal circumference. INTERVENTION(S): The participants underwent urologic visit, fasting blood and semen sampling, and anthropometric measurements, and filled in the International Physical Activity Questionnaire, at enrollment and after 4 and 8 months. Duration and frequency of walking, moderate-intensity, and vigorous-intensity activities in the last week were assessed, and a score was computed for total PA. MAIN OUTCOME MEASURE(S): Semen specimens were taken at each visit through masturbation, after 3-5 days of abstinence, and analyzed by an expert urologist. Sperm concentration, total and progressive motility, and proportion of spermatozoa with normal morphology were measured. Linear and generalized linear mixed models with the Poisson family were fitted to assess the relationships between PA variables and sperm parameters, after adjusting for season, time, and study arm. The shape of the relationship was modeled through restricted cubic spline regression. RESULT(S): A total of 143 male subjects, aged 18-23 years (median, 20 years), were enrolled. They had a median PA of 1,960 (95% confidence interval, 1,055-3,182) Metabolic Equivalent of Tasks in min/wk. Statistically significant differences were found for total, progressive motility, and percent of cell with normal morphology across categories of total PA; the highest medians of total (47%) and progressive motility (34%) and of the percentage of normal morphology cells (7%) were found for medium PA. Positive associations of sperm total motility and normal morphology with medium levels of PA, and negative associations with walking and vigorous-intensity activity emerged. Spline regression analysis confirmed these findings, showing an inverse U-shape relationship, with the highest value of total motility and normal morphology for medium PA, and the lowest values for lower and higher activity. CONCLUSION(S): These findings support the present recommendations to practice moderate PA for health improvement, including semen quality.

Trends and challenges in liquid-preserved boar semen production: From boar to product.

Boar semen production plays a pivotal role in modern swine breeding programmes, influencing the genetic progress and overall efficiency of the pork industry. This review explores the current challenges and emerging trends in liquid-preserved boar semen production, addressing key issues that impact the quality and quantity of boar semen. Advances in new reproductive technologies, boar selection, housing, semen processing, storage and transport, and the need for sustainable practices including the use of artificial intelligence are discussed to provide a comprehensive overview of the field.

Temporal effect of flaxseed oil in boar's diet on semen quality, antioxidant status and in-vivo fertility under hot humid sub-tropical condition.

The present study investigates the temporal effects of flaxseed supplementation on boar semen quality, antioxidant status, and in-vivo fertility under high-temperature humidity index (THI) conditions in a sub-tropical climate. Twelve Hampshire crossbreed boars were randomly assigned to control and treatment groups, with the treatment group receiving flaxseed oil supplementation. Semen samples were collected and analyzed for semen quality parameters, sperm kinematics, and antioxidant status. Fertility outcomes were assessed through in-vivo mating trials. Flaxseed supplementation resulted in time dependent significant improvements in semen volume, sperm concentration, total and progressive sperm motility, sperm quality parameters, and antioxidant status. Fertility outcomes, including farrowing rates and litter sizes, were also enhanced in the flaxseed-supplemented group. These findings highlight the potential of flaxseed supplementation to improve boar fertility under high ambient stress conditions, with implications for optimizing reproductive performance in swine production systems.

Novel approach for oligospermia (NAPO) - Protocol for a randomized controlled trial.

Background: Infertility affects millions of couples globally, with up to 40-50 % of cases linked to impaired semen quality. Insemination or in vitro fertilization are used frequently, regardless of the cause of infertility due to the lack of specific medical interventions for male infertility. Denosumab, an antibody blocking RANKL signaling, may enhance semen quality in infertile men. This randomized controlled trial evaluates if denosumab improves spermatogenesis in men with severely impaired semen quality identified by serum AMH levels as a predictive marker. Methods: NAPO is a single-center, sponsor-investigator-initiated, placebo-controlled, double-blinded randomized trial. Subjects will be randomized in a 2:1 fashion to receive either denosumab 60 mg subcutaneously or a placebo. The study will be carried out at the Division of Translational Endocrinology, Copenhagen University Hospital, Herlev, Denmark. The primary outcome of the study is defined as the difference in sperm concentration (millions/mL) at one spermatogenesis (80 days) after inclusion. Discussion: An important step in addressing infertility is establishing a viable treatment option for male infertility. With this study, we describe the protocol for a planned RCT aimed at evaluating whether treatment with denosumab can improve sperm concentration in men with severely impaired semen quality. The results of this study will provide evidence crucial for future treatment in a patient group where treatment options are minimal at best. Trial registration: Clinical Trials: NCT06300229. Registered on March 12, 2024. Clinical Trials Information System (CTIS): 2023-508325-27-00. Approved on December 19, 2023.

Standardization and application of ARMS TaqMan real-time PCR for screening of folate metabolism genes in Han Chinese.

Folate has antioxidant properties, and low concentration in seminal plasma may be associated with increased DNA damage in sperm. Mutations of the methylenetetrahydrofolate reductase (MTHFR) and methionine synthase reductase (MTRR) genes, including MTHFR C677T (rs1801133), MTHFR A1298C (rs1801131), and MTRR A66G (rs1801394), can lead to decreased activity of the encoded folate metabolic enzymes, thereby affecting male reproduction. The current SNP detection methods commonly used in clinical practice have some shortcomings, such as long time-consuming, complex detection steps, or high cost. The purpose of this study was to establish a simple, time-saving, sensitive, accurate, and easy to clinical popularization method for folate metabolism gene detection. We combined ARMS-PCR with TaqMan fluorescent probe to establish an ARMS TaqMan real-time PCR detection method. According to the variation of rs1801131, rs1801133, and rs1801394, two specific primers (one wild type and one mutant) were designed. Mismatched nucleotides were introduced at the penultimate or third position to improve the specificity of the primer. Specific TaqMan probe was introduced to detect PCR products to improve the sensitivity of the method. The results showed that the sensitivity of ARMS TaqMan real-time PCR in SNP genotyping was 1 ng, and the accuracy was 100%. A total of 249 clinical samples were detected by the established method, and the correlation between three SNPs and semen quality was analyzed. We found that individuals carrying the AG + GG genotype of rs1801394 had a lower risk of abnormal semen quality. In conclusion, we developed a highly sensitive, accurate, rapid, and easy to be popularized method for detecting SNPs of rs1801394, rs1801131, and rs1801133. ARMS TaqMan real-time PCR is a reliable SNP genotyping method in folate metabolism genes.

Prehypertension in male affects both semen quality and pregnancy outcomes in their first single blastocyst frozen-thawed embryo transfer cycles.

OBJECTIVE: To investigate whether prehypertension (pre-HTN) in male affects semen quality and assisted reproductive technology (ART) outcomes. DESIGN: Retrospective cohort study. SETTING: University-affiliated reproductive medicine center. PATIENT(S): Clinical data were collected from 1,043 couples undergoing their first single blastocyst frozen-thawed embryo transfer cycles. INTERVENTION(S): According to the blood pressure (1 mm Hg = 0.133 kPa) in male, including systolic blood pressure (SBP) and diastolic blood pressure (DBP), the subjects were divided into the control group (90 ≤ SBP < 120 mm Hg and 60 ≤ DBP < 80 mm Hg, n = 611) and the pre-HTN group (120 ≤ SBP < 140 mm Hg and/or 80 ≤ DBP < 90 mm Hg, n = 432). The association between pre-HTN and semen quality, and ART outcomes was then evaluated. MAIN OUTCOME MEASURE(S): Primary outcome: live birth rate. SECONDARY OUTCOMES: semen quality, laboratory embryo outcomes, clinical pregnancy rate, biochemical pregnancy rate, miscarriage rate, and other pregnancy outcomes. RESULT(S): Compared with the control group, the pre-HTN group showed lower total sperm motility, total sperm count, sperm progressive motility, progressive sperm count , and higher prevalence of oligozoospermia (17.6% vs. 13.1%), and asthenozoospermia (37.7% vs. 19.8%). Meanwhile, compared with the control group, the pre-HTN group had a lower clinical pregnancy rate (42.8% vs. 57.6%) and a lower live birth rate (32.9% vs. 47.3%). There were no differences in the remaining parameters of semen quality, laboratory embryo outcomes (except for the proportion of intracytoplasmic sperm injection), and pregnancy outcomes between the two groups. Additionally, regression analysis showed that pre-HTN in male was an independent risk factor for clinical pregnancy (adjusted odds ratio, 0.57; 95% confidence interval, 0.44-0.74) and live birth (adjusted odds ratio, 0.58; 95% confidence interval, 0.44-0.75). CONCLUSION(S): Prehypertension in male not only results in lower semen quality, but also has negative consequences on the success of ART. It is an independent risk factor for clinical pregnancy and live birth in the first single blastocyst frozen-thawed embryo transfer cycles.