The effect of sperm preparation methods on sperm chromatin condensation and DNA damage

Infertility problem is a increasing day by day, approximately 20% of infertile couples are caused only by male factor). In the evaluation of the male factor, standard semen analysis is performed according to WHO criteria. However, semen analysis does not provide information about sperm DNA damage. One of the most important mechanisms of sperm DNA damage is abnormal or irregular chromatin packaging. In the process of spermatogenesis, the spermatid chromatin undergoes a compaction process and the histone proteins carry out the condensation of sperm chromatin by replacing them with sperm specific protamine proteins. Abnormal chromatin condensation process causes developmental anomalies in sperm. Sperm DNA fragmentation index and maturation defect rates can be related with infertility. Gradient and swim-up methods are the pre-IVF-ICSI sperm preparation techniques that applied routinely in IVF laboratories. Swim-up method allows the separation of the spermatozoa from the fast ones to the upper phase while the gradient method provides separation by utilizing the density differences of abnormal and normal sperm. Both methods aimed to obtain the highest quality, high motility and normal morphology. Semen with non-sperm cell allows to eleminate dead immobilized sperm. Mostly, all procedures are performed at 37 ° C in IVF laboratories .The aim of study was to investigate the effect of different semen preparation in different temperatures on the separation of sperm, control in with maturation defect and DNA fragmentation. The study was submitted to the Biruni University Ethics Committee and was approved in 2018-12-9 and dated 29.01.2018. The study was applied to the Biruni University Hospital between January 2018 and June 2018 in patients who applied for spermiogram analysis. In study; Patients admitted to the Biruni University Hospital Urology outpatient clinic. Standard semen analysis was performed according to WHO criteria in normospermia (n: 20) and oligoastenoteratospermia (n: 20) groups, followed by sperm maturation defect with acidic aniline blue staining method and sperm DNA fragmentation damage by acridin orange staining method. After separation of the sperm by using gradient and swim-up methods, the groups were separated from them and incubated at room temperature (22 ° C) and 37 ° C for 1 hour, followed by incubation with acidic aniline blue staining and acridin orange 4 staining prior to DNA fragmentation index. Sperm maturation defect was compared with the values obtained after the application. According to this thesis, there is no significant difference between the swim-up and gradient methods in terms of motile sperm, sperm number and morphology as a sperm preparation method and it can be preferred according to the condition of motility and concentration parameters. In terms of sperm immaturation and DNA fragmentation, it has been shown to decrease with density gradient and swim up method. Especially in cases of oligoasthenoteratospermia, the incubation at 37 ° C had bad effects especially on DNA fragmentation compared to normospermia. Incubation can be carried out at room temperature to minimize sperm DNA damage.In this study it has been shown that the most functional incubation temperature at the swim-up or gradient technique are room temperature (22 °C ). Key words: DNA damage, gradient, swim-up, DNA fragmentation