Sperm DNA Fragmentation Risk Factors That Kill Potency
Overview
Sperm DNA fragmentation (SDF) has garnered attention as a significant factor in male infertility. Traditional semen analysis methods are insufficient for a comprehensive assessment of SDF. To address this, the authors sought to investigate the relationship between SDF and sperm parameters using computer-assisted sperm analysis (CASA), aiming to enhance treatment approaches in reproductive medicine.
In this retrospective observational study, the researchers evaluated the association between CASA-derived sperm parameters and SDF values obtained through the TUNEL assay. The study included 359 patients who sought infertility treatment at Mie University Hospital. Semen analyses encompassed concentration, motility, and morphology, with subsequent SDF measurement via flow cytometry.
Statistical analyses revealed significant correlations between SDF and various factors, including patient age, period of sexual abstinence, and specific CASA parameters. Notably, reduced sperm motility and abnormal head dimensions were linked to higher SDF levels, suggesting these parameters as predictors of SDF.
This study underscores the significance of sperm motility and head morphology as indicators of SDF, proposing their utility in evaluating male fertility. The findings advocate for detailed sperm analysis, potentially enhancing the success rates of assisted reproductive technologies by refining sperm selection criteria.
Introduction
Infertility, a condition of the reproductive system, is defined by the inability to achieve clinical pregnancy after 12 months or more of regular, unprotected intercourse. Globally, around 72.4 million couples face infertility challenges. Men are solely responsible for 20%-30% of infertility cases and contribute to 50% of all cases. Several factors, including oxidative stress, hormonal and anatomical abnormalities, and genetic, lifestyle, and environmental elements, significantly influence male infertility. Recently, sperm DNA fragmentation (SDF), which involves single- and double-strand breaks in sperm DNA, has emerged as a critical factor. High levels of SDF negatively impact male fertility and reproductive outcomes, reducing the likelihood of natural conception. Elevated SDF levels are also linked to an increased risk of recurrent pregnancy loss (RPL) and adversely affect the success rates of assisted reproductive technologies (ART) such as in vitro fertilization (IVF), intrauterine insemination (IUI), and intracytoplasmic sperm injection.
Standard semen analysis parameters defined by the World Health Organization (WHO), including sperm count, motility, and morphology, are moderate predictors of reproductive outcomes. Studies have explored the association between these parameters and SDF, revealing that men with higher SDF levels often exhibit lower semen quality. Despite normal semen analysis results, some men may still experience elevated SDF levels, indicating that traditional semen evaluations alone are insufficient to fully assess SDF. More detailed investigations are needed to understand the relationship between sperm status and SDF, with computer-assisted sperm analysis (CASA) providing a valuable tool for this purpose.
CASA offers a more reproducible and accurate evaluation of semen quality than traditional manual methods, allowing for detailed analysis of sperm motility and morphology. Research indicates that CASA-derived sperm parameters are associated with SDF, highlighting its potential utility in predicting male fertility. This study aims to elucidate the correlation between SDF and sperm parameters measured using CASA, such as motility patterns, velocity, and morphology, to better understand male fertility issues and enhance treatment strategies in ART.
Method
This retrospective study investigated the relationship between sperm parameters analyzed via computer-assisted sperm analysis (CASA) and sperm DNA fragmentation (SDF) assessed using the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay with flow cytometry (FCM). Ethical approval (approval number: H2023-129) was obtained from the Mie University Ethics Committee. The study included 359 male patients from March 2020 to December 2023, excluding those with varicocele history and insufficient sperm counts for CASA analysis. Data on BMI, sexual abstinence duration, alcohol use, and smoking were collected. Semen samples were analyzed for volume, concentration, motility, morphology, and detailed sperm measurements using the LensHooke X1 PRO CASA system. Sperm DNA fragmentation was assessed by fixing and staining sperm cells with TUNEL and analyzing them with FCM. Results were interpreted based on fluorescence intensity thresholds and sperm population characteristics, contributing insights into male fertility factors.
Statistical Analysis
In the study, data analysis was conducted using R software version 4.3.0 (The R Foundation for Statistical Computing, Vienna, Austria). Initially, a multiple regression analysis was performed to assess the relationships between various variables and Sperm DNA Fragmentation (SDF). To refine the model and adjust for potential confounders, a stepwise selection method based on the Akaike Information Criterion (AIC) was applied. This iterative process involved adding or removing variables from the initial model to optimize it by selecting the most significant predictors and achieving the lowest AIC.
The resulting model from the stepwise selection was then used for a final multiple regression analysis to explore factors associated with SDF. Similarly, for logistic regression analysis, initially all variables were included to evaluate risk factors for SDF >20%. The model was refined using stepwise selection based on AIC, ensuring inclusion of only the most significant predictors in the final logistic regression model.
To compare differences between groups, the Mann–Whitney U test examined variations in age, duration of sexual abstinence, BMI, and semen analysis parameters. Fisher’s test was employed to assess the impact of smoking and alcohol consumption on SDF. Odds ratios (OR) with 95% confidence intervals (CI) indicated increased risk of SDF >20%. Statistical significance was defined at p < 0.05 for all analyses.
Result
In a study involving 359 men seeking infertility diagnosis and treatment at Mie University Hospital, various semen parameters were analyzed. The median age of the subjects was 37 years, with a median body mass index (BMI) of 23.5 and a median sexual abstinence period of 4.5 days. The median sperm DNA fragmentation (SDF) was 13.6%.
Multiple regression analysis was conducted to explore the relationship between SDF and several variables. Age, duration of sexual abstinence, and the use of a certain sperm parameter, VSL, were found to be statistically significant. However, high variance inflation factors (VIFs) suggested potential multicollinearity issues, and the adjusted R-squared value indicated a limited ability of the model to explain SDF variation.
To address these issues, variables were selectively removed based on the Akaike information criterion (AIC), resulting in an improved model with an adjusted R-squared of 0.124. The overall regression model was statistically significant (p < 0.05), highlighting age, sexual abstinence duration, rapid sperm motility rate, and certain sperm head parameters as significant predictors of SDF. Notably, while head length and perimeter showed potential associations, multicollinearity concerns persisted.
A comparison between two groups based on SDF levels (cutoff at 20%) revealed significant differences in age, sexual abstinence duration, and various sperm motility parameters. Logistic regression further identified sexual abstinence duration, rapid sperm motility rate, and specific sperm head characteristics as factors significantly influencing the likelihood of SDF exceeding 20%. These findings underscored the complex interplay between physiological variables and SDF levels in male infertility.
Overall, the study contributes valuable insights into the factors influencing sperm DNA fragmentation, emphasizing the importance of considering multiple variables in assessing male reproductive health and infertility.
Conclusion
This study explores the impact of aging and sexual abstinence duration on sperm DNA fragmentation (SDF), emphasizing their roles in elevating oxidative stress levels, a known contributor to SDF. Advanced age correlates with increased SDF, reflecting heightened oxidative stress associated with aging. Similarly, longer periods of sexual abstinence lead to higher SDF levels, possibly due to accumulated oxidative damage in semen.
The research highlights a negative correlation between rapid sperm motility rate and SDF, suggesting that reduced motility increases the risk of significant DNA damage in sperm. Previous studies corroborate these findings, linking oxidative stress to impaired sperm motility and mitochondrial function.
Regarding sperm morphology, longer sperm heads show a lower incidence of SDF, whereas larger head perimeters are associated with higher SDF levels. This aligns with studies indicating that abnormal sperm head morphology, including head defects, correlates positively with SDF and negatively with fertility outcomes in assisted reproductive technology (ART).
The study underscores the potential of computer-assisted sperm analysis (CASA) in evaluating sperm morphology and its association with SDF risk, although its utility compared to traditional sperm parameters like concentration and motility remains debated.
Limitations include the study’s retrospective design and the absence of IVF outcome assessments, necessitating further research to clarify these relationships and minimize biases in future studies.
In conclusion, this research contributes valuable insights into male fertility assessment by identifying age, sexual abstinence duration, sperm motility, and morphology as critical factors influencing SDF. These findings may inform better criteria for sperm selection in ART, potentially improving clinical outcomes.
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