Ghorbani-Sini Razieh, Izadi Tayebeh, Tavalaee Marziyeh, Azadi Leila, Hajian Mehdi, Rahimi Zamani Mahshid, Nasr-Esfahani Mohammad Hossein
ACECR Institute of Higher Education (Isfahan Branch), Isfahan, Iran.
Department of Reproductive Biotechnology, Reproductive Biomedicine Research Center, Royan Institute for Biotechnology, ACECR, Isfahan, Iran.
Int J Fertil Steril. 2020 Apr;14(1):51-56. doi: 10.22074/ijfs.2020.5981. Epub 2020 Feb 25.
Telomeres are particular sequences of DNA located at the end of the eukaryotic chromosomes that are essential for genome integrity. Telomere length in spermatozoa differs among males, as well as spermatozoa. Also, decreased telomere length in spermatozoa of infertile men is associated with the reduction of fertility potential and embryo quality. Density gradient centrifugation (DGC) and swim-up are useful techniques for separation of spermatozoa with longer telomeres. Also, the selection of sperm based on surface negative electric charge or "Zeta potential", can separate high percentage of spermatozoa with intact chromatin compared to DGC alone, and also the combination of DGC-Zeta can improve clinical outcomes of infertile men candidate for intracytoplasmic sperm injection (ICSI). Therefore, we compared sperm telomere length and DNA fragmentation between two sperm preparation procedures, namely DGC and zeta potential.
In this experimental study, we assessed sperm telomere length and DNA fragmentation by quantitative real-time polymerase chain reaction (PCR) and TUNEL assay methods, respectively. The spermatozoa were obtained from infertile men with normozoospermia between September 2017 and December 2017 and prepared either by DGC or zeta potential methods. Sperm telomere length was expressed as relative and absolute units.
Compared with washed semen samples or control, no significant (P>0.05) difference was observed in the mean relative or absolute sperm telomere length when the two methods DGC or zeta potential were compared. However, the mean percentage of DNA fragmentation was significantly (P<0.05) lower in spermatozoa prepared by DGC or zeta potential methods than spermatozoa obtained from control samples.
This is the first study that compared the effect of DGC and zeta potential as the sperm preparation methods on sperm telomere length. It seems that both methods can select sperm population with high DNA integrity and the same sperm telomeres length.
端粒是位于真核染色体末端的特定DNA序列，对基因组完整性至关重要。男性之间以及精子之间的精子端粒长度存在差异。此外，不育男性精子中端粒长度的减少与生育潜力和胚胎质量的降低有关。密度梯度离心（DGC）和上游法是分离端粒较长精子的有用技术。此外，基于表面负电荷或“zeta电位”选择精子，与单独使用DGC相比，可以分离出更高比例的染色质完整的精子，并且DGC-zeta联合使用可以改善接受卵胞浆内单精子注射（ICSI）的不育男性的临床结局。因此，我们比较了两种精子制备方法（即DGC和zeta电位）之间的精子端粒长度和DNA片段化情况。
在本实验研究中，我们分别通过定量实时聚合酶链反应（PCR）和TUNEL检测法评估精子端粒长度和DNA片段化情况。精子取自2017年9月至2017年12月间患有正常精子症的不育男性，并通过DGC或zeta电位法进行制备。精子端粒长度以相对单位和绝对单位表示。
与洗涤后的精液样本或对照组相比，比较DGC或zeta电位这两种方法时，平均相对或绝对精子端粒长度未观察到显著差异（P>0.05）。然而，通过DGC或zeta电位法制备的精子中DNA片段化的平均百分比显著低于（P<0.05）从对照样本中获得的精子。
这是第一项比较DGC和zeta电位作为精子制备方法对精子端粒长度影响的研究。似乎这两种方法都可以选择具有高DNA完整性和相同精子端粒长度的精子群体。
