Selecting spermatozoa with intact chromatin may reduce embryo aneuploidy

Abstract

We tested a novel approach for treating couples with complete and persistent embryo aneuploidy. Using a microfluidic device, we selected spermatozoa with the highest progressive motility and genomic integrity, capable of generating euploid embryos. From October 2016 to April 2019, 13 consenting couples with male partners with high sperm chromatin fragmentation (SCF) in their ejaculate and a history of embryo aneuploidy and/or recurring implantation failure underwent a new ICSI cycle in which semen specimens were processed by microfluidics sperm selection (MFSS) and density gradient centrifugation (DGC). Consenting men had their ejaculates screened by standard semen analysis according to WHO 2010 criteria. Specimens were processed by DGC and MFSS. SCF was measured by TUNEL utilizing a commercial kit (In Situ Cell Death Detection Kit, Roche). At least 500 spermatozoa were counted under fluorescent microscopy, with an established threshold of 15%. Fertilization and clinical pregnancy rates were assessed and compared between the two preparation methods. Preimplantation genetic testing for aneuploidy (PGT-A) was performed on the resulting embryos. Embryo implantation and pregnancies after replacement of thawed euploid blastocysts were recorded. A total of 13 men with an average age of 41.5±10 years had the following average semen parameters: concentration of 40.5±44 x106/mL, 26±19 motility, and 2.2±1% morphology. After DGC and MFSS, the sperm concentration was 27±41 and 4.4±7 x106/mL, with 47.7±43% and 97.1±4% motility, respectively (P<0.0001). The average SCF decreased from 29% in the raw samples to 20% following DGC (P=NS), and dropped to 2.2% after MFSS processing (P<0.0001). These couples (female partner, 39±6 years) underwent 15 ICSI cycles with DGC-selected spermatozoa and achieved a fertilization rate of 71.3% (97/136), which generated 64.5% (31/48) morphologically good-quality embryos; of these, 12.5% (6/48) were determined by PGT-A to be euploid. Two of these euploid embryos were transferred and did not yield a pregnancy. In a subsequent ICSI cycle with MFSS processing, a fertilization rate of 73.2% (104/142) resulted in 61.8% (34/55) good-quality embryos. Of these, 36.3% (20/55) were identified as euploid and were cryopreserved (P<0.05). Seven couples received a thawed single euploid blastocyst and all 7 became pregnant (P<0.0001), resulting in the delivery of two healthy babies, with 71.4% (5/7) still ongoing. Dysfunction of the male genital tract increases both single-strand (ss) and double-strand (ds) DNA nicks and breaks, resulting in spermatozoa that impair embryonic development. Because dsDNA breaks in the male gamete can be responsible for embryo aneuploidy, the use of MFSS processing to select spermatozoa with the highest motility and genomic integrity may enhance the chances of obtaining a euploid conceptus for transfer.