Romero-Aguirregomezcorta Jon, Laguna-Barraza Ricardo, Fernández-González Raúl, Štiavnická Miriama, Ward Fabian, Cloherty Jennifer, McAuliffe Denis, Larsen Peter B, Grabrucker Andreas M, Gutiérrez-Adán Alfonso, Newport David, Fair Seán
Department of Biological Sciences, School of Natural Sciences, Biomaterials Research Cluster, Bernal Institute, Faculty of Science and Engineering, University of Limerick, Limerick, Ireland.
Department of Animal Reproduction, INIA, Madrid, Spain.
Reproduction. 2021 Mar;161(3):343-352. doi: 10.1530/REP-20-0364.
The objective of this work was to elucidate whether a sperm selection method that combines rheotaxis and microfluidics can improve the selection of spermatozoa over density gradient and swim-up. For this purpose human sperm selected by rheotaxis were compared against density gradient, swim-up and a control group of non-selected spermatozoa in split frozen-thawed (FT) and fresh (F) semen samples. Sperm quality was assessed in terms of motility, morphology, DNA fragmentation index (DFI), viability, acrosome integrity and membrane fluidity. Using a mouse model, we compared fertilisation and embryo development rates after performing ICSI with spermatozoa, sorted using rheotaxis or swim-up. Selection by rheotaxis yielded a sperm population with reduced DFI than the control (P < 0.05), improved normal morphology (P < 0.001) and higher total motility (TM; P < 0.001) than the other techniques studied in F and FT samples. Swim-up increased TM compared to density gradient and control in FT or F samples (P < 0.001), and yielded lower DFI than the control with F samples (P < 0.05). In FT samples, selection by rheotaxis yielded sperm with higher viability than control, density gradient and swim-up (P < 0.01) while acrosomal integrity and membrane fluidity were maintained. When mouse spermatozoa were selected for ICSI using rheotaxis compared to swim-up, there was an increase in fertilisation (P < 0.01), implantation (P < 0.001) and foetal development rates (P < 0.05). These results suggest that, in the absence of non-destructive DNA testing, the positive rheotaxis can be used to select a population of low DNA fragmentation spermatozoa with high motility, morphology and viability, leading to improved embryo developmental rates.
这项工作的目的是阐明一种结合趋流性和微流控技术的精子筛选方法是否能比密度梯度离心法和上游法更有效地筛选精子。为此，在冻融（FT）和新鲜（F）精液样本中，将通过趋流性筛选出的人类精子与密度梯度离心法、上游法筛选出的精子以及未筛选的精子对照组进行比较。从活力、形态、DNA碎片指数（DFI）、存活率、顶体完整性和膜流动性等方面评估精子质量。我们使用小鼠模型，比较了采用趋流性或上游法筛选的精子进行卵胞浆内单精子注射（ICSI）后的受精率和胚胎发育率。在F和FT样本中，与其他研究技术相比，通过趋流性筛选出的精子群体的DFI降低（P < 0.05），正常形态得到改善（P < 0.001），总活力（TM）更高（P < 0.001）。在上游法中，与密度梯度离心法和对照组相比，FT或F样本中的TM增加（P < 0.001），并且在F样本中DFI低于对照组（P < 0.05）。在FT样本中，通过趋流性筛选出的精子的存活率高于对照组、密度梯度离心法和上游法（P < 0.01），同时顶体完整性和膜流动性得以维持。与上游法相比，当使用趋流性筛选小鼠精子进行ICSI时，受精率（P < 0.01）、着床率（P < 0.001）和胎儿发育率增加（P < 0.05）。这些结果表明，在没有无损DNA检测的情况下，正向趋流性可用于筛选具有低DNA碎片、高活力、良好形态和存活率的精子群体，从而提高胚胎发育率。
