DNA Fragmentation Index Values Research Articles

Correlation between standard sperm parameters and sperm DNA fragmentation from 11,339 samples

Conventional semen parameters have long been considered fundamental in male fertility analyses. However, doubts have been raised regarding the clinical utility of the assessment of spermatozoa (sperm) DNA damage. In this retrospective study, we investigated the potential correlation between conventional semen parameters and semen DNA fragmentation (SDF) assessed as sperm DNA damage, in 11,339 semen samples collected between January 2019 and June 2022. We observed significant negative correlations between the DNA fragmentation index (DFI) and sperm viability (correlation coefficient [r] = −0.514) as well as progressive sperm motility (r = −0.512, p < 0.05). Samples were categorized into three groups according to DFI levels (Groups A, B, and C: ≤15%, 15 < DFI ≤30%, and >30%, respectively). Furthermore, the percentage of semen samples with normal sperm conventional parameters in Groups A, B, and C was 76.7% (4369/5697), 61.4% (2351/3827), and 39.7% (721/1815), respectively. Moreover, according to the reference values of conventional sperm parameters, the samples were divided into Groups F, G, and H with all normal, only one abnormal, and > two abnormal parameters, respectively. In addition, the proportions of samples with abnormal DFI values (>30) in Groups F, G, and H were 9.7% (721/7441), 23.1% (618/2676), and 39.0% (476/1222), respectively. Multivariate logistic regression models demonstrated that sperm vitality, progressive sperm motility, normal sperm form, total sperm count, semen volume, age, and some sperm kinematics collectively improved the area under the receiver operating characteristic curve (AUROC) to 0.861, surpassing the predictive value of a single predictor of pathologically damaged sperm DNA. Our study suggests that samples with abnormal sperm parameters may have a higher likelihood of high DNA fragmentation. Furthermore, certain semen parameters could be potential indicators of sperm DNA fragmentation, aiding sperm selection in assisted reproductive procedures.

Comparison of Embryological Results of Microinjection in Two Groups of Men with and without Requesting Sperm DNA Fragmentation Index Measurement.

The sperm DNA fragmentation index (DFI) is considered a valuable measure to assess male fertility, but the predictive value of DFI for the outcomes in assisted reproductive technology (ART) is still controversial. Therefore, this study is aimed at investigating the effect of requesting a DFI test or performing ART without DFI on the results observed in the embryology laboratory (number of embryos, fertilization rate, and embryo quality) after intracytoplasmic sperm injection (ICSI). This retrospective study was conducted on infertile men who underwent ICSI and were referred to the Avicenna Infertility and Recurrent Abortion Treatment Center in Tehran from 2019 to 2022. The samples were categorized into two groups: a case group with DFI measurement and a control group without DFI measurement. We conducted a comparative analysis of the embryology results between the two groups, focusing on parameters such as fertilization rate, number of embryos, and embryo quality. t-tests and Mann-Whitney U tests were used to conduct single variable analysis. Potential confounding effects were adjusted to use the multivariate linear and logistic regression. Data analysis showed no significant statistical difference between the case group and the control group in terms of the number of embryos (95% confidence interval for the regression coefficient (β) = -0.257-0.123), and embryo quality (95% confidence interval for β = -0.199-0.114). There was no significant statistical difference between the two groups due to the fertilization rate (95% confidence interval for β = -3.42-3.42), except for the variables of woman's age and sperm count after ICSI, as determined by adjusted linear regression. Although DFI measurement is used to assess male infertility, its importance as a predictor for the embryology outcomes after ICSI requires further evaluation and the determination of a cut-off point for predicting results. This study was based on retrospectively collected DFI data, and prospective studies confirming the superiority of ICSI outcomes are necessary.

#353 : Microfluidic Preparation of Spermatozoa: A Potential Approach in Assisted Reproductive Technology

Background and Aims: While density gradient centrifugation (DGC) is a conventional method in sperm preparation, this method can induce reactive oxygen species (ROS) which cause sperm dysfunction, sperm DNA damage, a recent advancement in sperm processing using microfluidic system is believed to reduce damage and to select the good quality sperm. The aim of this study was to evaluate the effectiveness of sperm preparation between DGC method and microfluidic system. Method: 30 semen samples were separated into 2 equal small samples, each sample was processed by DGC methods (control) and microfluidic (intervention) respectively. Pre- and post processing parameters was evaluated according to WHO-2010 and the DNA fragmentation index (DFI) was determined by flow cytometry technique. Results: The result showed that DFI values from the microfluidic preparation (0.67 ± 0.53%) was significantly lower than that of the DCG group (1.18 ± 13.73%). In particular, for semen samples with high DFI (&gt;15%), the microfluidic system showed a noticeable reduction in DFI compared to the results from the DGC method. The progressive motile sperm recovery rate was higher in the intervention group (39.75 ± 19.20 vs 32.14 ± 18.14). Our data also showed that the vitality and mobility of sperm in the microfluidic group was higher than those of the DGC group (97.9 ± 2.21 vs 96.56 ± 4.11%, 96.36 ± 2.60 vs 95.06 ± 2.35%, respectively). However, there was no difference in sperm progressive motility and sperm morphology between two groups. Conclusions: There were significant improvement in sperm DFI and progressive motility in the microfluidic system. With this result, microfluidic system is a potential approach to reduce DNA fragmentation for ICSI procedures.

Evaluation of Human Papillomavirus in the Semen of Infertile Men and Its Relationship with Semen Quality

Background: The prevalence of human papillomavirus (HPV) infection in infertile men is significantly higher than that in the general population. Nevertheless, it is unclear whether the presence of HPV in semen samples is related to reduced semen quality. Objectives: The present article aims to elucidate the possible association between HPV infection and semen parameters in Iranian infertile men. Methods: This cross-sectional study included semen samples from 140 infertile Iranian male partners of couples seeking fertility assessment. Standard semen analysis was performed. Sperm DNA fragmentation index (DFI), sperm chromatin maturation, anti-sperm antibodies, leukocyte count, fructose concentration, and zinc concentration were also assessed. Nested polymerase chain reaction (PCR) and INNO-LiPA were used to identify HPV DNA sequences in semen samples and to determine HPV genotype, respectively. Independent samples t-tests were used to compare sperm parameters between HPV-positive and HPV-negative groups. Results: Human papillomavirus DNA was detected in 18 (12.85%) out of 140 semen samples (10% high-risk genotypes, 2.14% low-risk genotypes, and one sample was excluded). Our findings revealed that HPV18 was the most prevalent type (8.57%). According to our results, the presence of HPV DNA in semen samples was associated with higher semen anti-sperm antibodies (P = 0.001), higher white blood cell (WBC) counts (P = 0.02), and increased DFI values (P = 0.03). Conclusions: Human papillomavirus -positive patients had higher semen anti-sperm antibodies, semen WBC counts, and sperm DFI values. These results highlight the importance of careful screening for the presence of HPV in semen samples from infertile men. It should also be noted that more studies are required.

Machine learning-based clustering to identify the combined effect of the DNA fragmentation index and conventional semen parameters on in vitro fertilization outcomes

BackgroundPrevious studies have demonstrated an association between male sperm quality and assisted reproduction outcomes, focusing on the effects of individual parameters and reaching controversial conclusions. The WHO 6th edition manual highlights a new semen assay, the sperm DNA fragmentation index, for use after routine semen examination. However, the combined effect of the sperm DNA fragmentation index (DFI) and routine semen parameters remains largely unknown.MethodsWe assessed the combined effect of the sperm DFI and conventional semen parameters on single fresh conventional IVF outcomes for infertile couples from January 1, 2017, to December 31, 2020. IVF outcomes were obtained from the cohort database follow-up records of the Clinical Reproductive Medicine Management System of the Third Affiliated Hospital of Guangzhou Medical University. An unsupervised K-means clustering method was applied to classify participants into several coexposure pattern groups. A multivariate logistic regression model was used for statistical analysis.ResultsA total of 549 live births among 1258 couples occurred during the follow-up period. A linear exposure–response relationship was observed among the sperm DFI, sperm motility, and IVF outcomes. In multivariable adjustment, increased sperm DFI values and decreased sperm motility and semen concentration levels were associated with reduced odds of favourable IVF outcomes. Four coexposure patterns were generated based on the sperm DFI and the studied semen parameters, as follows: Cluster 1 (low sperm DFI values and high sperm motility and semen concentration levels), Cluster 2 (low sperm DFI values and moderate sperm motility and semen concentration levels), Cluster 3 (low sperm DFI values and low sperm motility and semen concentration levels) and Cluster 4 (high sperm DFI values and low sperm motility and semen concentration levels). Compared with those in Cluster 1, participants in Cluster 3 and Cluster 4 had lower odds of a live birth outcome, with odds ratios (95% confidence intervals [CIs]) of 0.733 (0.537, 0.998) and 0.620 (0.394, 0.967), respectively.ConclusionsWhen combined with low sperm DFI values, there was no significant difference between high or moderate sperm concentration and motility levels, and both were associated with favourable IVF outcomes. Low sperm parameter levels, even when DFI values remain low, may still lead to poor IVF outcomes. Participants with high sperm DFI values and low sperm motility and semen concentration levels had the worst outcomes. Our findings offer a novel perspective for exploring the joint effects of sperm DFI and routine semen parameter values.

Correlation analysis of sperm DNA fragmentation index with semen parameters and the effect of sperm DFI on outcomes of ART

Routine semen analysis provides limited information about a man’s male reproductive potential and can not always fully explain male infertility. Many male infertilities are caused by sperm DNA defects that routine semen quality analyses fail to detect. In this study, we analyzed the association of sperm DNA fragmentation index (DFI) with the semen routine, sperm morphology, in vitro fertilization embryo transfer (IVF-ET)/intracytoplasmic sperm injection (ICSI). Further, we explored the predictive value of sperm DFI in evaluating male fertility and the outcome of IVF-ET/ICSI. Data on sperm DFI, sperm routine, and sperm morphology were collected from 1462 males with infertility. According to DFI levels, there were 468 cases in group I (DFI ≤ 15%), 518 cases in group II (15% < DFI < 30%), and 476 cases in group III (DFI ≥ 30%). The correlations of sperm DFI with semen routine and malformation rate were analyzed. Seminal plasma malondialdehyde (MDA), and total antioxidant capacity (TAC) were assessed. Sperm DFI, semen routine, and sperm morphology were detected in male patients of 101 pairs of IVF-ET/ICSI infertile couples and subdivided into IVF-I group (DFI ≤ 15%), IVF-II group (15% < DFI < 30%), IVF-III group (DFI ≥ 30%), ICSI-I group (DFI ≤ 15%), ICSI-II group (15% < DFI < 30%) and ICSI-III group (DFI ≥ 30%) according to DFI value. The effect of sperm DFI on the outcome of IVF-ET/ICSI was analyzed. There were significant differences in sperm survival rate, sperm concentration, and PR% between groupIII and group II (P < 0.01). There were significant differences in sperm survival rate, sperm concentration and PR% between group III and group I (P < 0.01). There was no significant difference in semen volume, age, abstinence days, or percentage of normal sperm between the three groups (P > 0.05). DFI was positively correlated with MDA content ( P < 0.01) and negatively correlated with TAC (P < 0.01). Sperm DFI was negatively correlated with sperm survival rate, sperm concentration, and PR% (P < 0.01). There was no correlation with age, abstinence days, semen volume, or percentage of normal-form sperm (r = 0.16, 0.05, 0.04, -0.18, p > 0.05). Compared with IVF-I group, the sperm concentration and PR were decreased in IVF-III group. The sperm malformation rate was higher in IVF-III group than that in IVF-II group. Comparatively, the PR was decreased in ICSI-III group. The sperm malformation rate was higher in ICSI-III group than that of the ICSI-I group (P < 0.05). There were no statistically significant differences in fertilization rate, cleavage rate, embryo rate, and clinical pregnancy rate between IVF group or ICSI group, and between all subgroups (P > 0.05). Sperm DFI is negatively associated with sperm survival rate, sperm concentration, and PR%. Antioxidants can decrease the rate of DNA fragmentation. Sperm DFI has proven to be very valuable in the male fertility evaluation, but its significance as a predictor of pregnancy outcomes following assisted reproductive technology. (ART) requires further investigation.

The effects of hormonal changes on sperm DNA integrity in oligoasthenoteratospermia individuals: A case-control study.

Hormonal imbalance is one of the important etiological factors for Oligoasthenoteratospermias (OAT). This study aimed to evaluate the effects of hormonal changes including prolactin, TSH, testosterone, luteinizing hormone, follicle-stimulating hormone, and anti-Mullerian hormone on sperm DNA fragmentation in normal men compared with OAT to design a clinical algorithm for the comprehensive study of male factor infertilities. We consecutively selected 60 candidates referred to the infertility clinic to collect the semen and blood samples. Then, a terminal deoxynucleotidyl transferase dUTP nick end labeling test was performed to evaluate the sperm DNA fragmentation index (DFI). After semen analysis and DFI checking, they were classified into 4 groups consisting of normospermia and OAT men each with or without increased DFI. Hormone parameters were analyzed using enzyme-linked immunoassay. Follicle-stimulating hormone and luteinizing hormone levels showed positive correlations with DFI in a significant way (p 0.01), while testosterone and thyroid-stimulating hormone were associated with sperm concentration. Prolactin and anti-Mullerian hormone levels significantly correlated (p 0.01) with sperm concentration and DFI value simultaneously. Decreased and increased levels of serum hormones could adversely affect semen profile and sperm DNA integrity which lead to severe male infertility. Although we investigated the effects of the main hormones related to male infertility on DNA damage, the role of these hormones on the fertilization rate and embryo quality needs to be evaluated in further studies.

Predicting the value of sperm analysis using an electronic nose

Total motile sperm count and DNA fragmentation Index are two parameters in sperm analysis that have recently been used to determine the outcome of the management of cases of male infertility. Total Motile Sperm Count is one of the values considered better than the 2010 World Health Organization standard sperm analysis in terms of predictive value for the success of the spontaneous ongoing pregnancy rate. High DNA Fragmentation Index values were associated with lower pregnancy success and an increased risk of low fertilization rate or total fertilization failure. In this study, we developed a method to classify sperm analysis based on total motility sperm count and DNA Fragmentation Index values by using an electronic nose. In the total motility sperm count (TMSC) study, we use four algorithms with the result of accuracy values 95% and in the DNA fragmentation Index study, we get a fairly good accuracy value for two algorithms with the accuracy values 70%.

Evaluation of sperm DNA fragmentation in male infertility.

Sperm DNA integrity could be considered a biological marker of sperm quality and may affect fertilization, embryonic development, and pregnancy outcome. The study aimed to investigate the connection between semen characteristics and sperm DNA damage in infertile patients. Standard semen analysis of 536 samples was carried out following the World Health Organization (WHO) 5th edition recommendations. Sperm DNA damage was assessed by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay, after preparation by direct swim-up. The slides were evaluated using a fluorescence microscope and the percentage of TUNEL-positive spermatozoa was expressed as the DNA fragmentation index (DFI). Patients were classified according to their DFI levels: group A (DFI < 15%) and group B (DFI ≥15%). Sperm total count, concentration, total and progressive motility, vitality, and normal morphology were significantly higher in group A compared to group B (p < 0.001). The results show a significant inverse correlation between DFI and patient's age, sperm total count, concentration, total and progressive motility, vitality and normal morphology. Higher DFI values were found to be strongly associated with poor sperm quality. In conclusion, combined with conventional semen analysis, assessment of sperm DFI could improve diagnostic accuracy and treatment management for patients with male infertility.

Correlations between elevated basal sperm DNA fragmentation and the clinical outcomes in women undergoing IUI.

ObjectiveThis study aimed to explore the impact of the sperm DNA fragmentation index (DFI) on the clinical outcomes in women undergoing artificial insemination by husband intrauterine insemination (AIH-IUI).MethodsIn this retrospective study, the value of sperm DFI was detected by sperm chromatin structure assay (SCSA) in a semen analysis collected before fertility treatment (basal DFI) in 1,500 IUI cycles at the infertility clinic of Northern Jiangsu People’s Hospital Reproductive Medicine Center from Jan 2016 to April 2021. Receiver operating characteristic (ROC) curves were used to calculate the cut-off value for the clinical outcomes of IUI, including the biochemical pregnancy rate, clinical pregnancy rate, delivery rate, and live birth rate, and multivariate logistic regression was conducted to analyse the risk factors for clinical outcomes after IUI.ResultIn 1,500 IUI cycles, the results showed that there were no statistically significant differences between the normal DFI group and the abnormal DFI group in biochemical pregnancy rate (14.41% vs. 11.3%, P = 0.386), clinical pregnancy rate (12.9% vs. 10.5%, P = 0.433), delivery rate (11.0% vs. 8.9%, P = 0.456), live birth rate (10.9% vs. 8.9%, P = 0.484) or pregnancy loss rate (14.6% vs. 15.4%, P = 1.000).ConclusionSperm DFI alone may have limited predictive power for IUI clinical outcomes.

P-006 MACS Vs Microfluidics Sperm Sorting FOR RAISED SPERM DNA FRAGMENTATION INDEX – A RCT

Abstract Study question In Individuals with raised Sperm DNA Fragmentation Index (DFI), will sperm selection by magnetic activated cell sorting (MACS) or Microfluidic Sperm Sorting (MF) optimize the reproductive outcomes? Summary answer MACS or MF offer comparable outcomes. There seems no superiority between them. Individuals with raised spermDFI, MACS/MF can be considered to optimize results. What is known already It is evident that raised sperm DFI negatively affects the reproductive outcomes. Management for raised sperm DFI to optimize reproductive outcomes is still elusive. Study design, size, duration This is an ongoing Randomized Control Trial (RCT) with prior approval from institutional Ethical Committee (IEC) (ECR/1312/Inst/TG/2019). This is preliminary data of this ongoing study. Study duration 6months(July 2020–January 2021). Couples undergoing IVF stimulation with raised spermDFI were randomized into MACS(n = 75) and MF(n = 58) for sperm selection. This study also intends to look at efficiency of MACS and MF sperm sorting to reduce spermDFI post intervention. This trial is registered with Clinical Trials Registry India (CTRI) REF/2021/05/043439 Participants/materials, setting, methods Couples with one failed IVF were offered sperm DFI. Individuals with sperm DFI&amp;gt;30% were recruited. Sperm DFI testing was done with SCSA method and randomized using software. Sperm processing done with MACS or MF based on randomization on the day of Oocyte Retrieval(OPU). MACS/MF sperm processing done as per manufacturer’s instructions. Culture done till blastocyst stage and freeze all policy followed. Two Blastocysts that showed100% survival was transferred in a Frozen Embryo transfer (FET) cycle. Main results and the role of chance Implantation rates(IR) Miscarriage Rates (MR), Multiple Pregnancy Rates(MPR) Live Birth rates (LBR) were compared between groups Reduction in sperm DFI value between MACS and MF was also evaluated In MACS arm - 75 subjects recruited, 49 patients (65%) had atleast 2 blastocysts frozen and underwent FET. 26 patients (35%) had cancelled cycles due to no blastocyst formation. In MF arm - 58 subjects recruited 38 patients (65%) had atleast 2 blastocysts and underwent FET. 20 patients (35%) had cancelled cycles due to no blastocyst formation. Sperm DFI reduction with MACS Vs MF Sperm Sorting 83% of men had sperm DFI value reduced to &amp;lt; 30% post MACS intervention 87% of men had sperm DFI value reduced to &amp;lt; 30% post MF intervention Both MACS and MF seem to be efficient in sorting sperm with raised DFI and separate sperms with good DNA quality Reproductive Outcomes of MACS Vs MF Sperm Sorting were IR - 50% Vs 48% (p = 0.8540) MR - 5.3% Vs 11% (p = 0.3280) MPR - 8% Vs 18% (p = 0.1624) LBR per ET cycle - 63% Vs 63% (p = 1.0000) Reproductive Outcomes of MACS and MF sperm Sorting seems to be comparable. Data from this study doesn’t prove superiority of one processing method over another Limitations, reasons for caution This is an ongoing study and the data presented here is the preliminary data with small sample size Wider implications of the findings Optimal intervention for management of raised spermDFI needs further research. However, MACS/MF both are non invasive and seem to optimize reproductive outcomes. Nevertheless while offering these sperm sorting methods aspects like cost, time taken to process and technical challenges if any, should be considered and offered with appropriate counselling. Trial registration number REF/2021/05/043439

P-084 Novel Single-Step Microfluidic Chamber Outperforms Density Gradient Centrifugation in Post-Selection Motility And DNA Integrity

Abstract Study question Does sperm selection by a novel single-step microfluidic chamber offer effective selection of motile sperm without DNA damage compared with Density Gradient Centrifugation (DGC)? Summary answer The novel single-step microfluidic chamber is a good alternative to conventional DGC, increasing DNA integrity and progressive sperm motility in recovered sperm populations. What is known already High levels of sperm DNA fragmentation (SDF) can have negative impacts on reproductive outcomes such as clinical pregnancy and miscarriage rates. Microfluidic sperm selection technologies have been developed to select sperm with lower level of SDF and higher levels of motility to improve Assisted Reproductive technology (ART) outcomes. While these methods have achieved variable success, very few have proven clinically relevant and fail to perform selection in a robust and simple methodology that can be adopted in clinics. Study design, size, duration A prospective cohort study including 21 donated semen samples processed in 2021. Each sample was split to perform DGC and microfluidic sperm selections side-by-side. Laboratory outcomes were evaluated using semen parameters pre and post processing and included concentration, motility, and DNA fragmentation index (DFI). Participants/materials, setting, methods The study was performed using samples donated at the Department of Obstetrics and Gynaecology, Monash University, Melbourne, Australia. DGC (10 minutes at 300g and 5 minutes wash at 300g) and microfluidic sperm selection (one-step sperm selection;15-minute incubation for selection) were performed with Sydney IVF 80/40 gradients and gamete buffers, respectively. DFI was assessed by chromatin sperm dispersion assay (SCD). Results were analysed by Friedman’s test and differences were considered significant when p-value &amp;lt; 0.05. Main results and the role of chance For the unprocessed raw semen, the starting DFI values varied from 3.0 to 23.3% with an average of 11.38% ± 5.51 and motility varied between 25.3% and 86.7% with an average of 56.42% ± 18.57. The device consistently outperformed DGC in all 21 samples for DFI (0.94% ± 1.12 vs 4.87% ±3.69, P = 0.0012) and in 20 of the 21 samples for motility post-processing DGC (90.88% ± 6.8 vs 73.58% ± 14.97, P = 0.0061). The average concentrations of the raw, DGC, and microfluidic sperm samples were 94.5, 74.12, and 3.55 million sperm/mL, respectively. Thus, both DGC and the microfluidic device showed improvements in SDF, motility, and concentration over the raw sample. However, the microfluidic device significantly outperformed DGC in terms of percentage SDF and motility, indicating its potential for clinical implementation. Limitations, reasons for caution This cohort study did not include samples from infertile patients. Furthermore, the effect of each sperm selection methods on reproductive outcomes was not a part of the study. A larger number of samples across a range of clinically infertile samples is required to fully characterise the microfluidic devices efficacy. Wider implications of the findings The microfluidic device we report here has the potential to become a new tool to improve the efficacy and consistency of sperm selections. When compared to DGC it offers improved sperm motility and DNA integrity from a platform that is simple to use and less skill-based than conventional methods. Trial registration number not applicable

Technical Evaluation of Commercial Sperm DFI Quality Control Products in SCSA Testing.

With the increase in population and the advancement of medicine, people even more hope that their offspring will be healthier. DNA fragmentation rate is currently one of the more common indicators for evaluating sperm fertility and predicting the outcome of pregnancy. In order to evaluate the technical performance and application value of commercial sperm DFI (DNA fragmentation index) quality control products in the flow cytometry sperm chromatin structure analysis (SCSA), this paper uses flow cytometry to test commercial sperm DFI quality control products (Celula) and laboratory routine self-made DFI quality control products. In this paper, the quality control of new commercial sperm DFI and laboratory-made quality control were compared and tested for 30 consecutive days. And this paper monitors the response of commercial quality control products to the interference of key reagent parameters. This paper compares the stability of the test results of two quality control products and their sensitivity to the interference of key performance parameters of the detection reagent. Experimental results show that commercial sperm DFI quality control products can simulate sperm DNA damage to achieve accurate detection of DNA integrity. The stability of commercial sperm DFI quality control (CV = 2.47%) is better than that of laboratory-made quality control (CV = 11.22%). The new commercial sperm DFI quality control product can sensitively detect changes in the concentration of acidified solution and staining solution at the same time. It can effectively control the quality of detection reagents and experimental procedures. The new commercial sperm DFI quality control product can effectively control sperm DNA integrity testing. It can be used as an external quality control product for quality control of test results to ensure that more accurate test results are provided to the clinic.

Sperm DNA Fragmentation Index in Infertility Patients vs Normal Individuals: A Cross-sectional Study

Introduction: Infertility is a major problem among married couples. The male factor contributes to around 30-40% of all cases of infertility. With advancing age, reducing sperm count may magnify the problem. The awareness of qualitative change along with quantity is important for people seeking advice in infertility. Aim: To find the prevalence of sperm DNA fragmentation with Sperm Chromatin Dispersion (SCD) test. Materials and Methods: The present study was prospective cross-sectional study which carried out on semen sample given for routine checkup in department during the period from August 2017 to August 2018. A total of 180 couples, including 88 infertile couples and 92 couples with known fertility were enrolled in the study. Sperm Count and sperm DNA Fragmentation Index (DFI) were calculated and determined by SCD test by kit method. Comparison between groups was done according to distribution of data using Mann-Whitney rank sum (two tailed) statistical test. Results: Among 180 subjects, (88 cases and 92 control) no significant difference in the age was found. The sperm count was not significantly different in cases (group of men of infertile couple) and controls (group of men of proven fertility). The highest value for DFI was 27% in controls and 97% in cases. Statistically significant difference was found in DFI (p=0.008) in both groups. DFI and sperm count showed negative correlation both in cases and controls with a correlation coefficient 0.213 (p-value &lt;0.001) and 0.754 (p-value &lt;0.005) respectively. A significant difference was observed in the median value of sperm DFI of men of infertile couple as compared to men of fertile couple. Conclusion: On comparing, significant difference was observed in the median value of sperm DFI in infertile couple. Sperm DFI at cut-off value 27% has higher diagnostic significance. SCD method is simple, easiest and standard tool to assess DFI.

The effect of micronutrient supplementation on spermatozoa DNA integrity in subfertile men and subsequent pregnancy rate

Background Male infertility has been related to an increased sperm DNA fragmentation index (DFI). Nutritional factors may improve sperm nuclear DNA integrity and thus pregnancy rates. Objective: To evaluate the effect of micronutrient supplementation on sperm DNA integrity in subfertile men and subsequent pregnancy rates. Methods In this retrospective comparative study 339 subfertile males were included on whom a sperm chromatin dispersion test (SCD) was performed as a method to detect DNA fragmentation, as well as an initial semen analysis. Of all, n = 162 received a nutritional management program for three months, consisting of two daily capsules of a standardized combined micronutrient formulation together with a guidance to diet modification and to lifestyle changes (study group). Each capsule contained L-carnitine, L-arginine, coenzyme Q10, zinc, vitamin E, folic acid, glutathione, and selenium. The control group consisted of those patients who did not receive the active treatment (n = 177), yet were instructed to engage in a healthy lifestyle, including a modification of their regular diet. The SCD test was repeated for both groups after three months. As part of the routine follow up, pregnancy rate was assessed six months after the second SCD test. Males with complete follow up and healthy female partners (aged 18 to 40 years) where included. Results Data of men with an initial mean DFI of >15% were analyzed first (n = 81;46 study and 35 control patients). After three months, both groups displayed a significant decrease of mean DFI values; however, the mean percent difference was higher in the study group (10.46 ± 1.20 % vs. 5.29 ± 0.57 %; p < .001). Then, the entire population was considered (n = 339). After three months, only the study group displayed a significant decrease of mean DFI initial values (10.48 ± 7.76 % to 6.51 ± 4.61%; p < .001); and the percent difference was higher in the study group (3.97 ± 0.28 % vs. 0.91 ± 0.28 %; p < .001). At six months follow-up, pregnancy rate was higher in the study group (27.78% vs. 15.25%, p = .002). Conclusion Both regimes significantly reduced sperm DNA fragmentation among subfertile men with a DFI >15%; however, when any baseline DFI value was considered, only micronutrient supplementation achieved a better result on DFI and thus pregnancy rate was higher.

DNA Fragmentation in Human Spermatozoa and Pregnancy Rates after Intrauterine Insemination. Should the DFI Threshold Be Lowered?

Sperm DNA fragmentation index (DFI) can be analyzed by a flow cytometric assay after treatment with acid and acridine orange. In this prospective, cohort study, the value of DFI was determined in a semen analysis collected before fertility treatment (baselineDFI) in 146 couples and during 1–3 intrauterine inseminations (IUI) in 211 couples (511 cycles). The pregnancy rate (PR)/cycle was 9.9% if baselineDFI was >10 and 21.7% if baselineDFI was ≤10, (p < 0.005). The live birth rate (LBR)/cycle was 5% if baselineDFI was >10 and 14.2% if baselineDFI was ≤10 (p < 0.005). PR/patient was 23.1% if baselineDFI was >10 and 45.5% if baselineDFI was ≤10 (p < 0.005). LBR/patient was 12.4% if baselineDFI was >10 and 34% if baselineDFI was ≤10 (p < 0.005). When isolating non-stimulated IUI cycles and couples with female age < 35, a significant difference in PR and LBR between couples with high DFI and low DFI was seen. Results suggest that DFI > 10 could advice against timed coitus and non-stimulated IUI cycles. Analysis for DFI performed before treatment provides information about PR and LBR after IUI.

Semen parameters on the intracytoplasmic sperm injection day: Predictive values and cutoff thresholds of success

ObjectiveThis study was conducted to investigate the relationship of semen parameters in samples used for intracytoplasmic sperm injection (ICSI) with fertilization and pregnancy rates in infertile couples.MethodsIn this prospective study of Infertile couples with male factor infertility that had undergone ICSI, fractions of the same semen samples obtained for microinjection (to ensure the best predictability) were evaluated to determine the semen parameters and sperm DNA fragmentation index (DFI) on the day of oocyte recovery.ResultsIn total, 120 couples completed the study and were subdivided into fertilized (n=87) and non-fertilized couples (n=33). The fertilized couples were further classified into pregnant (n=48) and non-pregnant (n=39) couples. Compared to non-fertilized and non-pregnant couples, fertilized and pregnant couples showed statistically significantly higher sperm viability and percentage of normal sperm morphology, as well as significantly lower sperm DFI values. A receiver operating characteristic curve analysis of data from the 120 ICSI cycles showed that sperm viability, normal sperm morphology percentages, and sperm DFI were significant prognostic indicators of fertilization at cutoff values of 40%, 7%, and 46%, respectively. A sperm DFI of 46% showed sensitivity and specificity of 95% and 90%, respectively, for predicting fertilization, and no clinical pregnancies occurred in couples with a sperm DFI above 46%.ConclusionSemen parameters from the ICSI day sample, especially sperm viability, normal morphology, and DFI, had an impact on fertilization and pregnancy outcomes in ICSI cycles.

Correlation between the DNA fragmentation index (DFI) and sperm morphology of infertile patients

PurposeTo evaluate the correlation between the DNA Fragmentation Index (DFI) and sperm morphology in patients undergoing ICSI, as a predictive parameter in reproductive outcomes.MethodsA retrospective study was conducted on 125 infertile patients enrolled in a fertility clinic. Seminal characteristics were measured following the WHO guidelines (2010) for the examination of the seminal fluid. After collecting motile sperm population by pellet swim up, DFI was calculated and simultaneously associated with sperm morphology using in situ TUNEL assay and an image analyzer software in at least 250 spermatozoa for each patient.ResultsAll subjects were divided into two groups according to a cutoff established, by choice, of the sperm DFI (15%): group A (< 15%) consisting of 65 patients and group B (≥ 15%) of 60 patients. Data were analyzed using non-parametric statistical methods. The results demonstrate that there is no statistical difference between the two groups in seminal characteristics. The collective data show a high significant correlation, suggesting that spermatozoa with abnormal morphology are the best candidates to contain DNA damage (p < 0.001). Also, when group A is compared with group B, an increased percentage of morphologically normal spermatozoa with fragmented DNA was observed in patients, with DFI values ≥ 15% (p < 0.001).ConclusionThese results are aimed at providing an exact value of DFI in morphologically normal spermatozoa, which will be helpful to the embryologist in evaluating the risk of transferring, during the ICSI procedure, a spermatozoon whit normal morphology but fragmented DNA.

Sperm chromatin structure assay versus sperm chromatin dispersion kits: Technical repeatability and choice of assisted reproductive technology procedure.

ObjectiveThe sperm DNA fragmentation index (DFI) guides the clinician’s choice of an appropriate assisted reproductive technology (ART) procedure. The DFI can be determined using commercially available methodologies, including sperm chromatin dispersion (SCD) kits and sperm chromatin structure assay (SCSA). Currently, when DFI is evaluated using SCD kits, the result is analyzed in reference to the SCSA-derived threshold for the choice of an ART procedure. In this study, we compared DFI values obtained using SCSA with those obtained using SCD and determined whether the difference affects the choice of ART procedure.MethodsWe compared SCSA to two SCD kits, CANfrag (n=36) and Halosperm (n=31), to assess the DFI values obtained, the correlations between tests, the technical repeatability, and the impact of DFI on the choice of ART. ResultsWe obtained higher median DFI values using SCD kits than when using SCSA, and this difference was significant for the CANfrag kit (p<0.001). The SCD kits had significantly higher coefficients of variation than SCSA (p<0.001). In vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) would be chosen for a significantly higher proportion of patients if a decision were made based on DFI derived from SCD rather than DFI determined using SCSA (p=0.003). ConclusionOur results indicate that SCD kit-specific thresholds should be established in order to avoid the unnecessary use of IVF/ICSI based on sperm DNA damage for the management of infertility. Appropriate measures should be taken to mitigate the increased variability inherent to the methods used in these tests.

DNA fragmentation of sperm: a radical examination of the contribution of oxidative stress and age in 16 945 semen samples.

What is the relationship between sperm DNA fragmentation and oxidative stress (OS) with increasing male age? Sperm DNA fragmentation increases with age and is likely related to both defective spermatogenesis and increasing OS levels. Sperm quality declines with age. The presence of DNA damage in a high fraction of spermatozoa from a raw semen sample is associated with lower male fertility in natural conception and intrauterine insemination. A retrospective cohort study of 16 945 semen samples analysed at a single reference laboratory between January 2010 and December 2018. All males were undergoing an infertility evaluation. The cohort was divided into seven age categories: <30, 30-34, 35-39, 40-44, 45-49, 50 to <54 and ≥55 years. The mean age was 37.6 years (SD 6.8). Sperm DNA fragmentation index (DFI) and high DNA stainability (HDS) were calculated using flow cytometry. OS levels were measured using the oxidative stress adducts (OSA) test, by spectrophotometry. ANOVA with weighted polynomial contrast analysis was used to evaluate trends for DFI, OSA and HDS values across age categories. Mean DFI significantly increased across all age groups (Ptrend < 0.001). OSA was lowest in patients <30 years old (mean 3.6, SD 1.0) and also increased as age increased (Ptrend < 0.001). There was a statistically significant difference between age groups for each of the three parameters (P < 0.001). There was a significant linear trend for DFI, OSA and HDS across the seven age categories (P < 0.001). Among patients with high DFI, there was a decreasing age-dependent trend in the patients observed with high OSA (P < 0.001). This is a retrospective study. All males included in the study were undergoing a work-up for infertility and may not be representative of a fertile population. Additional patient demographics and clinical data were not available. DNA and/or oxidative damage in sperm may be just as important to understand as the chromosomal aberrations that are carried in the oocyte. Further studies are needed to evaluate the effect of advancing paternal age on the male genome and, ultimately, on the health of the offspring. No funding was obtained for this study. V.D. is an employee of Reprosource/Quest Diagnostics. D.S. reports he was a Scientific Advisor to Cooper Surgical. N/A.