Said Tamer M, Grunewald Sonja, Paasch Uwe, Glander Hans-Juergen, Baumann Thomas, Kriegel Christian, Li Liang, Agarwal Ashok
Center for Advanced Research in Human Reproduction, Infertility and Sexual Function, Glickman Urological Institute and Department of Obstetrics-Gynecology, Cleveland Clinic Foundation, 9500 Euclid Avenue, Desk A19.1, Cleveland, OH 44195, USA.
Reprod Biomed Online. 2005 Jun;10(6):740-6. doi: 10.1016/s1472-6483(10)61118-2.
The selection of vital, non-apoptotic spermatozoa is a prerequisite for achieving optimal conception rates in assisted reproductive techniques. Magnetic cell sorting using annexin-V microbeads can effectively separate apoptotic and non-apoptotic spermatozoa. The objective of the present study was to optimize the integration of magnetic cell sorting in standard sperm preparations and to correlate the effect of different sperm preparation procedures on apoptotic markers. Semen specimens collected from 15 healthy donors were prepared by either density gradient centrifugation or by one-step sperm wash technique separately and in combination with magnetic cell sorting. The preparation methods were evaluated by assessment of semen parameters (motility, viability and morphology) as well as markers of apoptosis (levels of active caspase-3, integrity of membrane mitochondrial potential and externalization of phosphatidylserine). The apoptotic markers were measured using fluorochrome dyes coupled with flow cytometry. The results showed that the combination of density gradient centrifugation and annexin-V magnetic cell sorting was superior to all other sperm preparation methods in terms of providing motile, viable and non-apoptotic spermatozoa. This study clearly shows the advantage of integrating magnetic cell sorting as a part of sperm preparation, which in turn may positively affect the success rates of assisted reproductive techniques.
选择有活力、非凋亡的精子是在辅助生殖技术中获得最佳受孕率的前提条件。使用膜联蛋白-V微珠进行磁性细胞分选能够有效分离凋亡精子和非凋亡精子。本研究的目的是优化磁性细胞分选在标准精子制备中的整合，并关联不同精子制备程序对凋亡标志物的影响。从15名健康供体收集的精液标本分别通过密度梯度离心或一步法精子洗涤技术单独制备，并与磁性细胞分选联合使用。通过评估精液参数（活力、存活率和形态）以及凋亡标志物（活性半胱天冬酶-3水平、线粒体膜电位完整性和磷脂酰丝氨酸外化）来评价制备方法。使用荧光染料结合流式细胞术测量凋亡标志物。结果表明，在提供有活力、存活且非凋亡的精子方面，密度梯度离心与膜联蛋白-V磁性细胞分选相结合的方法优于所有其他精子制备方法。本研究清楚地表明了将磁性细胞分选作为精子制备一部分的优势，这反过来可能对辅助生殖技术的成功率产生积极影响。
