The Influence of Sperm DNA Fragmentation on Blastulation and Implantation Rates in Different Sources of Sperms.

how sperm DNA damage affects the patient reproduction ability. This suggests different outcomes, and the subject remains contentious. We investigated differences in sperm parameters and embryonic developmental steps in our study, and the observations compare the results of ICSI using sperm extracted from testicular on patients with high sperm injury to the results of patients with sperm DNA fragmentation on ejaculate samples. Materials and Methods: From March 2017 to December 2019, 418 couples were enrolled in our study at the Algazeera Centre for IVF Giza, Egypt. The parameters of the sperm were evaluated. The value sperm DNA fragmentation index (DFI) was calculated desponded on the sperm chromatin dispersion (SCD) diagnostic test, which was applied to teams in the regulation (sperm DNA fragmentation index < 30%) or abnormal (sperm DNA fragmentation index > 30%) groups. Results: decrease in fertilization and blastulation rate in a group of patients suffering high sperm damage on ejaculate samples and our results show no difference between normal ejaculate samples and testicular sperm extrication ( SDF) groups. Conclusion: no impact was detected on ICSI outcomes (pregnancy rates) between the three groups but a negative effect for sperm DNA fragmentation on fertilization and blastulation rate and blastulation rate depended on the age of women and the number of embryos transferred.


INTRODUCTION
For 44 years of IVF, all studies focus on how can increase the implantation rate. This recently improved it by obtaining blastocysts on day 5. But some factors inhabit embryos to form a blastocyst, the important factor for that male factor, which can investigate by the cornerstone of screening infertility males is semen analysis. Indicators of seminal vesicle and prostate function, respectively, are semen volume and pH level. Sperm concentration, motility, and morphology are mostly determined by testicular function and, to a lesser extent, by post-testicular (epididymal) genital tract function.
Although men who are fertile have greater mean sperm parameters than men who are infertile (concentration, motility, and morphology), these groups overlap significantly (Evenson, DP et al., 1999). Additionally, these factors typically offer poor predictions of reproductive outcomes. To differentiate between productive and unproductive men, identify pregnancy outcomes, and reduce the risk of unfavorable reproductive events, new markers are required. Standard sperm parameters are insufficient. Some data suggest that sperm DNA integrity larger studies are required to determine the clinical significance of evaluating sperm DNA integrity, but markers may be more accurate indicators of male reproductive capacity than standard assessments. (Evenson, D.P. et al., 1999, Spano M, et al., 2000, Zini A, et al., 2001, Guzick DS, el al., 2001. Research on sperm in an era where cutting-edge kinds of assisted reproductive technology are frequently used, DNA damage is particularly important. Direct contact between the sperm and the oocyte, a fusion of the cellular membranes, and the fusion of the male and female gamete genomes are all components of fertilizing. (Primakoff P 2002). The inherent quality of the sperm DNA is important in both the success of this procedure and the following embryogenesis (Ahmadi A, Ng SC 1999). Because sperm DNA integrity is not taken into account in sperm analysis, it has limitations in diagnosing male infertility (De Jonge, 2012). According to research, the sperm analysis results for 15% of evaluated infertile men are within normal parameters (Omran, Bakhiet, & Dashti, 2013;Schulte, Ohl, Sigman, & Smith, 2010). As a result, sperm DNA fragmentation and its connection to pregnancy outcomes have received a lot of interest. incorrect chromatin structure during spermatogenesis DNA damage in sperm is brought on by insufficient apoptosis just before ejaculation or by an excessive amount of reactive oxygen species (ROS). However, the exact mechanism causing this circumstance is yet unclear (Henkel R. et al., 1998, Sanocka-Maciejewska D. et al., 2005and Schuppe HC, et al., 2008. The sperm chromatin structure assay (SCSA), terminal deoxynucleotidyl transfer as mediated deoxyuridine triphosphates nick end labelling TUNEl, and other tests to assess sperm DNA integrity have all been developed since the original hypothesis-made more than 30 years ago that changes in the chromatin structure of sperm may be related to decreased fertility rates (Evenson, Darzynkiewicz, & Melamed, 1980). (Chohan et al., 2006). The WHO acknowledges the use of several of these assays for "checking the regularity of sperm chromatin and DNA," but not endorsing any of them.
In contrast, with such a diverse range of tests available, it has become increasingly clear that test standardization and consistency in clinical interpretation are essential (Shamsi et al., 2011Until there is standardization in the execution, analysis, and clinical interpretation of DNA fragmentation assays, it is doubtful that DNA fragmentation testing will be officially approved as a part of routine diagnostic testing in andrology laboratories around the world (Fernández et al., 2005).
There are several approaches to evaluating the integrity of sperm DNA, and each has benefits and drawbacks. But the only straightforward, inexpensive, and easily accessible kits are those that are sold commercially and are based on sperm. The chromatin dispersion test really does seem to satisfy these requirements. Other types of DNA fragmentation Assays necessitate specialized equipment, are employment, expensive, and time-consuming (Chohan et al., 2006). As a result, the impact of altered sperm chromatin integrity on post-embryonic development remains unknown (Larson et al., 2000). But its effect on fertilization, blastocyst formation and pregnancy rates is demonstrated in this study. Male infertile with high sperm DFI might think about getting ICSI as a treatment. However, studies have found that ICSI treatment for infertile couples with high sperm DFI results in adverse clinical outcomes, such as high rates of unsuccessful fertilization and clinical pregnancy (Lopes, Sun, Jurisicova, Meriano, & Casper, 1998;Gardner, Schoolcraft, Moffatt, & Sakkas, 2004). The sperm source, on the other hand, may have an impact on the degree of sperm DNA impairment during sperm preparation operations in an ART cycle, according to the available data. Spermatozoa from the testicles are believed to have more intact DNA signals than spermatozoa from the ejaculate, leading us to believe that using testicular spermatozoa rather than ejaculated spermatozoa would result in a more favourable reproductive outcome in an ICSI protocol. The goal of this study was to make a comparison of the clinical ICSI outcomes of using testicular spermatozoa versus ejaculated spermatozoa in infertile males with high SDF, with a focus on blastulation rates and clinical pregnancy the primary outcome and semen parameters, embryo fertilization and division as the secondary outcome, to help physicians determine the best treatment strategies for infertile males with high sperm DND Fragmentation Level.

Patients:
The prospective study concerned all the cycles of ICSI performed during a given period in Algazeera Hospital, Giza, Egypt: 418 cycles of ICSI. ART procedures involving ejaculated sperm samples or testicular extracted sperm were included in this study. The couples were divided into three groups: A Control Group: within normal sperm parameters of the patient`s (sperm concentration 20X10 6 /ml, motility 50% [types a and b, according to WHO criteria], and morphology 30% normal forms B Group: patients have high SDF and use ejaculate samples in ICSI C Group: patients use testicular biopsy sperm extraction because their sperm DNA fragmentation is high.

Semen Analysis:
After 3-5 days of sexual abstinence, each participant gave a sample of semen, and routine semen analyses were performed by one doctor in accordance with the 5th edition of the World Health Organization (WHO) laboratory manual for the examination and processing of human semen. Sperm parameters such as concentration, progressive motility (PR%), and normal sperm morphology were collected for more study.

Sperm DNA Fragmentation Assessment:
Halosperm® was used to analyze all sperm samples for fragmented DNA. Halosperm® is based on the sperm chromatin dispersion (SCD) technique, It involves a carefully controlled DNA denaturation procedure that makes it easier to remove the spermatozoon proteins later. This is how typical spermatozoa produce halos. by sperm DNA loops, which are missing in those with damaged DNA, at the sperm's head. Sperm was diluted in culture media with 20. million spermatozoa per milliliter concentration to get the optimum results 0.2 mL of fresh sample semen was diluted in medium sperm concentrations of 5 to 10 million/mL in order to get. The sperm sample was spread out across the sleeve in an agarose micro gel. After being polymerized, the agarose was chilled and then polymerized.
The slide was exposed to a denaturation solution after the cover had been removed. For 7 min with the answer, The acid denaturation solution (labeled AD tube) was combined with 10 mL of distilled water to create AD (denaturant solution), which was then added to an incubation tray. The material was submerged in the lysis solution for 25 minutes. The slide was then air-dried after being dehydrated in ethanol baths of 70%, 90%, and 100% for a total of three minutes. The sperm were then washed in water and stained with Giemsa. Each slide was air-dried before being viewed at 400x magnification using a Carl Zeiss Primo Starlight microscope to score 500 sperm. Halosperm® creates highly contrasted images of halos that may be carefully assessed utilizing Primo Star microscopy (Carl Zeiss). shows each of the five potential SCD patterns.
Sperm cells with prominent halos (thickness equal to or greater than the length of the core's Sperm cells with medium halos (thickness less than the length of the core's (minor diameter. Sperm cells with small. minor diameter but greater than 1/3 of the core's minor diameter) thickness equal to or less than 1/3 the diameter of the core's minor diameter.
A total of 500 spermatozoa were counted, and those with Degradation of sperm cells fragmented DNA was identified using the manufacturer's instructions below.

TESA Procedures:
As previously mentioned, a single doctor performed all TESA operations while under local anesthetic (Esteves, Miyaoka, & Agarwal, 2011) Briefly said, the spermatic cord was injected with about 10 ml of topical anesthetic to induce anesthesia. The spermatic cord, epididymis, and testicle fingers were interconnected during TESA treatments to immobilize the testicle and maintain skin tautness around the scrotum. The process was repeated until we had clear tissue or a spermatozoon in the biopsy needle (Argon Medical Device Inc., Gainesville, USA) that was put into the testicle's inferior pole while controlling the depth of the needle excursion. Under a microscope, one medical professional looked.

Ovarian Stimulation and ICSI Procedure:
Following three weeks of desensitization with GnRH analogues (Decapeptylâ; Ipsen), ovarian stimulation with recombinant FSH (Gonal-Fâ; Serono, or Puregonâ; Organon) was achieved and monitored. Endovaginal echography and plasma estradiol were used. When the follicles reached the desired diameter, 10 000 IU of hCG (Organon) has been administered 36 hours before oocyte retrieval. Under general anesthesia, the oocytes were extracted using a vaginal ultrasonography-guided aspiration.

Procedures for ICSI:
Denudation of oocytes was also collected, as was the on the same day as oocyte denudation, the male partner was asked to produce a semen sample, either by masturbation or TESA, which was then processed for ICSI. Fertilization was also checked, and the presence of two pronuclei and two polar bodies was considered normal fertilization; otherwise, it was considered abnormal. Embryo transfer was completed on the fifth day after the oocyte retrieval. Furthermore, clinical pregnancy was defined as the detection of a quantitative chemical pregnancy test and a fetal heartbeat via ultrasound 6 weeks after embryo transfer.

Assessment of Fertilization and Embryo Quality:
18 hours after fertilization, the resulting fertilized oocytes were placed in global media, they stayed until the fifth day. Embryos cultured after day 3 were transferred to an extended culture medium on day 3. Decide Culture for 3-5 days, depending on development speed and the Morphological quality of the cohort. Selected embryos Transfer to the new droplet on day 3 Selection of embryos for transfer Cleavage/morula stage or immature/Mature blastocyst stage. In blastocyst, in most cases, Maturity (size and distinguishable inner cell mass [ICM] and Trophectoderm) and morphology is the main determinant chosen. Embryo transfer these days Assisted hatching is not performed. this blastocyst classification according to dived Gardner. Blastocyst classification according to its size from 3:6, ICM classification from A: C and Trophectoderm classification from A: C Embryo Transfer: Embryo transfer takes place 5 days after oocyte injection and lasts approximately 115 hours. ETs must be taken carefully after counting blastocysts and transferred at 37 °C using a dewormed hot plate under a Zeiss stereomicroscope and a sterilized Falcon 1006 Petri dish. For transfer, incubate a 1 ml series filled with 20% Serum Supplement Solution (SSS) for 17 hours and remove air bubbles, connect the filled syringe to the transfer catheter, and after flushing the catheter, aspirate the selected blastocysts into the catheter, via The cervical canal inserts the catheter into the uterine cavity, slowly injects the catheter, and then gradually withdraws the catheter, and ET occurs under ultrasound in the operating room. The advantage of transferring blastocysts on day 5 is better synchronization between the embryo and the endometrium. Transferring one or two Blastocysts is the optimal number for transfer to avoid multiple pregnancies but our protocol in ET number of Embryos depends on t h e woman's age and/or the number of available blastocysts. According to our protocol.

Statistical Analysis:
Data were analyzed using IBM© SPSS© Statistics version 23 (IBM© Corp., Armonk, NY) and MedCalc© version 18.2.1 (MedCalc© Software bvba, Ostend, Belgium Numerical variables were presented as mean and SD and inter-group differences were compared using one-way analysis of variance with an application of the Tukey test for post hoc comparison if needed Categorical variables were presented as numbers and percentages and differences were compared using Fisher's exact test Correlations between numerical variables were tested using Pearson correlation. Multivariable binary logistic regression analysis was used to examine determinants of biochemical pregnancy. To examine determinants of fertilization, division, and blastulation rates, multivariable linear regression was used Two-sided p-values <0.05 were considered statistically significant.

RESULTS
The biochemical pregnancy results in the three groups were the following and no difference between the three groups. Table 1 the results success rate in the study groups.

Fisher's Exact Test:
In Table 1. Pregnancy tests amongst the three groups did not differ substantially (P > 0.05, p = 0.391). Whatever the circumstances, it was cancelled.
The comparison between three groups showing decrease on fertilization and blastocyst formation was as the following: Fertilization Rate: Control group = 83.4 ± 17 %, ICSI with ejaculate samples has SDF group = 75.0 ± 18.5 %, ICSI with TESE group=81.8±17.1 %, where, p value was = 0.003 so there is an affect by SDF.

Blastocyst Formation Rate:
Control group = 47.7 ± 18.1 % ICSI with ejaculate samples has SDF group = 38.3 ± 20.8 % ICSI with TESE = 48.1± 20.7 % where p value was = 0.002 so there is an affect by SDF. Therefore, the results had to be confirmed using linear regression, which it has more accurate where: Independent variable and a corresponding dependent variable. The independent variable is the input, and the corresponding dependent variable is the output. Table 2. shows the outcomes of a multivariable binary logistic regression analysis for factors that influence biochemical pregnancy. The odds ratio for an increase in the number of embryos transferred was 1.384, and the chance of pregnancy increased with that factor. The odds of a decrease in chemical pregnancy with an increase in the age of the wife factor were less than 1, indicating a negative effect.

Table 2. Effect of age variables and an increase in transplanted embryos on biochemical
pregnancy. †Descried to DNA fragmentation ≤30% (=0). ‡ Descried to sperm collection by ejaculation (=0). Table 3 shows the results of multivariable linear regression analysis for determinants of fertilization rate.

Table 3. Multivariable linear regression analysis for determinants of fertilization rate
This table demonstrates the effect of testicular sperm extracted effect in increase fertilization rate more than ejaculated sperm with high DNA sperm. B = regression coefficient, SE = standard error, rpartial = partial correlation coefficient (correlation between independent variable and outcome variable adjusted for the effect of the other variables in the model), rsemipartial = semi partial correlation coefficient (correlation between independent variable and outcome variable adjusted for other independent variables only.
†Descried to DNA fragmentation ≤30% (=0). ‡ Descried to sperm collection by ejaculation (=0). Table 4. shows the results of multivariable linear regression analysis for determinants of division rate; none of the included variables is a determinant of division rate (all P-values > 0.05) from the motion, not a significant (P ≤ 0.05). Table 4. Multivariable linear regression analysis for determinants of the division rate: B = regression coefficient, SE = standard error, rpartial = partial correlation coefficient (correlation between independent variable and outcome variable adjusted for the effect of the other variables in the model), rsemipartial = semi-partial correlation coefficient (correlation between independent variable and outcome variable adjusted for other independent variables only. †Descried to DNA fragmentation ≤30% (=0). ‡ Descried to sperm collection by ejaculation (=0). Table 5. shows the results of multivariable linear regression analysis for determinants of total blastulation rate.
Both sperm collection by FNA (B = -9.942, SE = 3.628, P-value = 0.006) and patient's age (B = 0.356, SE = 0.168, P-value = 0.035) are independent determinants of total blastulation rate. Where inject oocytes with testicular sperm extracted increases blastulation rate regard to oocytes injected with sperm with high sperm DNA fragmentation. And age factor was considered and the increase in the wife's age increased the blastulation rate it is according to our data analysis regardless of cancellation cases. Table 5. Multivariable linear regression analysis for determinants of total blastulation rate. B = regression coefficient, SE = standard error, rpartial = partial correlation coefficient (correlation between independent variable and outcome variable adjusted for the effect of the other variables in the model), rsemipartial = semi-partial correlation coefficient (correlation between independent variable and outcome variable adjusted for other independent variables only. †Descried to DNA fragmentation ≤30% (=0). ‡ Descried to sperm collection by ejaculation (=0). Table 6. shows the results of multivariable linear regression analysis for determinants of late blastulation rate, SDF, and wife age were showed a significant effect in blastocyst formation (P-value≤ 0.05). Where increased SDF decrease late blastulation rate, and increase wife age decrease late blastocyst formation.
Generally, there was no appreciable difference between the groups with an SDF of 30% and those with an SDF of 30% in ICSI in terms of achieving clinical or biochemical pregnancy. However, there are important differences between group B and the control in early and late blastulation, where excessive sperm DNA formation reduces blastocyst development. Additionally, there was no discernible difference between group C and the control group in any of the outcomes. Table 6. Multivariable linear regression analysis for determinants of late blastulation rate B = regression coefficient, SE = standard error, rpartial = partial correlation coefficient (correlation between independent variable and outcome variable adjusted for the effect of the other variables in the model), rsemipartial = semi-partial correlation coefficient (correlation between independent variable and outcome variable adjusted for other independent variables only. Referenced to DNA fragmentation ≤30% (=0). Referenced to sperm collection by ejaculation (=0).

Demographics:
In this part, the figures in the research are explained that the results from the statistical calculations and plenty of illustration to see the results represented in those illustrations. Figure 1 explains the relationship between the three groups in pregnancy results, as there is no difference in the results resulting from ICSI with the difference in the sperm source used that likely to be the cause of blastocyst trans fear benefits even transfer single blastocyst. While, the figure 2 explains the effect of ICSI by highly sperm DNA fragmentation, where decreased fertilization is less than the control group, which are the normal group and the group TESE group. Figure 3 explains the effect of sperm DNA fragmentation on division, no effect for high sperm DNA fragmentation on division where the results appear in this figure no different between three groups. Figure 4 descried the resulting of total blastulation rate where decreased blastulation on day fife by high sperm DNA fragmentation more than other two groups this likely to be the cause effect of paternal genome on day three. Also, The Figure 5 descried the resulting of late blastulation rate where increased late blastulation by high sperm DNA fragmentation more than other two groups this explains that the high sperm DNA fragmentation not arrest blastulation rate and delays it.  It is previously mentioned sperm DNA fragmentation showed significance in column group B. Graphical presentation of division rate in different groups A, B, and C, not significant effect (p-value > 0.05) between different columns. Graphical presentation of fertilization rate in different groups A, B, and C, between blastocyst formations in different three groups. Columns carrying different letters are significantly different at p < 0.05.

Fig. 5. Mean late blastulation rate in the three study groups. Error bars represent the standard error (SE). Dots represent individual observations.
Graphical presentation of the relationship between late blastulation in different three groups. Columns carrying different letters are significantly different at p < 0.05.

DISCUSSION
The effects of SDF on semen parameters and the requirement of sperm chromatin analyses in normal laboratory exams have conflicting conclusions. Studies have linked sperm DNA fragmentation to fertility and semen quality standards, In normal laboratory studies of semen, chromatin condensation (SCD) has been proposed as a useful metric to assess male fertility ( Hammadeh ME, et al., 2001). There are other ways to identify sperm DNA fragmentation, such as Halosperm, an enhanced SCD test created by Férnandez et al., in 2005, However, some researchers found a negative link in some (Khalili MA, et al., 2006), (Sills et al., 2004) and every parameter were to choose sperm that are normally mobile and do not affect an oocyte's potential to get fertilized (Muratori M., 2008) and (Erenpreiss J, et al.,2008) However, our results show host R et al., 2000 found that total and progressive sperm motility increased more in LFG than HFG after semen preparation, showing that lower DNA fragmentation levels benefit more from semen preparation through sperm motility. the head, acrosome, and intermediate portion of sperm, as well as concentration, do not yet correlate with sperm DNA fragmentation, but Cohen-Bacrie et al., 2009 showed a negative relationship between quick motility rate and provides a straightforward method for DNA fragmentation (Cohen-Bacrie P, 2009) That is consistent with our findings, where p =0.049 (previous manuscript khallaf et al., 2022) was found for decreased progression and quick motility. These findings may imply that the embryo's fertilization is influenced by the sperm's DNA fragmentation level but is unaffected by division and developmental potential until day 3 of development (Braude P, et al., 1988) that sperm can be repaired by oocytes and embryo DNA damage, (Ahmadi A, Ng SC et al., 1999), and that the patient's condition may have contributed to these findings. That agrees with my finding in tables 3 and 4 where the due to and the decrease of fertilization is likely to be the cause of a large amount of SDF and the ability to repair oocytes with this defect with a small amount of SDF. Sperm from the testicular exhibited less DNA damage than sperm from the ejaculate in infertile males with a high DNA fragmentation rate (Mehta, A., et al. (2015), (Moskovtsev, S. I. et al., 2010). Which can improve the embryonic potential from oocyte fertilization. (Weissman et al., 2008). discovered that the passage of spermatozoon through the epididymis was associated with loss of sperm integrity and fertilizing ability in a series of only four couples using ICSI cycles using ejaculated spermatozoon (O'Connell, McClure, and Lewis 2002). (Suganuma, Yanagimachi &Meistrich 2005). that reflect my findings, which appeared in table 3. One explanation for this finding is that the mature spermatozoa are crammed with immature spermatozoa that produce ROS as they go through the seminiferous tubule and epididymis, resulting in DNA damage to the mature sperm. (Aitken, R. J. et al., 1997, Ollero, M. et al., 2001. Because spermatozoon DNA is more susceptible to oxidation before going through chromatin condensation in the epididymis, this explanation makes sense. My article's findings imply that oxidative stress experienced during transit through the epididymis may be a greater danger to the success of ICSI than testicular sperm maturation. then use of testicular sperm in these patients to achieve better ICSI outcomes.
A Low and significantly lower cleavage and fertilization rates Note embryo quality and blastocyst formation, especially when Testicular sperm from men with OA and NOA is utilized for ICSI, which is caused by reduced testicular sperm quality, subsequent embryo quality and blastocyst growth are directly impacted. less fruitful than the previous ejaculation Epididymal maturation of sperm Lin M.H., et al.,2008. Additionally, research indicates that testes from NOA patients are more likely to have chromosomal aneuploidy than OA patients do. That may, at least in part, account for the reduced embryonic development rates seen in this study's group of patients. This contract with us There was no difference between the control group and the control group A set of testicular sperm extraction as a table (3,4,5) in the study that was presented. DFI has no impact on clinical outcomes following blastocyst transfer. because the pregnancy rate for blastocyst transfer is higher than that of early embryo transfer (day 2 or 3) There have been reports of high levels of in vitro-developed blastocysts (Zorn B et al., 2012, Goker, ENT et al., 2002 In blastocysts table, we saw adverse sperm damage effects. Form Table (5), which resulted in paternal genome effects Flow after 6-8 cell division, reducing blastocyst formation oppositely related to the number of oocytes extracted. The embryonic cell block is one of the most frequent causes of a decreased blastocyst development rate. Development blockade is made by Apoptosis that begins after the second or third division of the embryo. basic indications include apoptosis and nuclear fragmentation. Abnormalities in the development of microtubule spindles. Balaban B, et al., 2001Kolibianakis E, et al., 2002 The testicular sperm extraction group increased the blastulation rate of more Ejaculate samples which SDF, Table (5 ), because Reducing sperm damage is in this group equal to the properties of a typical ejaculated sample. according to 26 clinical outcomes of ICSI following single blastocyst transfer that have been documented Use of TESE similar to standard ICSI Table with normal ejaculation samples (Rossi-Ferragut LM et al., 2003-Arafa M, et al., 2017, Nagata S, et al., 2003, Nilsson J, et al., 2004. From table 6 we observed an increased number of late blastocysts compared with another group we believe that high SDF does not make an arrest only but can cause a delay in blastulation rate may be because of the action of oocytes in repairing itself and repairing the DNA damage take time, female aging and oocyte quality plays role .that agree with that who said ought to be taken under consideration is deoxyribonucleic acid fragmentation (Carrell and Liu 2003) report a drop in high-quality embryos and an increase in low-quality embryos on day two when gamete deoxyribonucleic acid fragmentation is substantial. It has been widely documented that the absence of the epididymal tract in spermatozoa prevents SDF in male reproductive gland gametes (Agarwal, Majzoub, Esteves, Ko, Ramasamy, & Zini, 2016;Agarwal Sharma, Samanta, Durairajanayagam, & Sabanegh, 2016;El Greco Scarselli, & Iacobelli, 2005). We incline to speculate that a higher level of deoxyribonucleic acid fragmentation in ejaculated thawed gametes may confirm a delay in embryo cleavage activation and, consequently, a delay seen in early cellular divisions.
Finally, after ICSI, we found no variations in the pregnancy rates between the 3 groups. highest-scoring embryos for transfer may be the reason why there is no correlation between sperm DNA damage and pregnancy rates. Because the pregnancy rate is affected by age of the female and the number of embryos in the blastocysts stage which is transferred to Table 2. Gerris M.R et al., 2009 where increased pregnancy chance with the increase in the number of blastocysts in a direct relationship with age and vice versa Table  1 and 2.

Conclusion
The benefit of sperm extraction from the testicles in cases of high sperm DNA. Fragmentation (SDF) gives great results in obtaining the highest fertilization equations and blastocyst formation. Similar to the injection of samples with normal parameters, which are in the results Superior to samples injected with higher SDF.