A NOVEL METHOD TO ATTENUATE EMBRYO ANEUPLOIDY DUE TO PATERNAL INHERITANCE

Abstract

To test the efficacy of a sperm-selection method to generate genetically healthy embryos in couples with recurrent embryo aneuploidy. Consenting men with high sperm DNA fragmentation (SCF) in their spermatozoa who were treated at our center during the last three years were included. Spermatozoa were selected by density gradient centrifugation (DGC) for ICSI cycles, which frequently yielded aneuploid embryos, resulting in no embryo transfer or failed implantation. In subsequent ICSI cycles, their spermatozoa were processed by microfluidics sperm selection (MFSS). SCF, including the assessment of double-stranded DNA breaks (DSBs), was performed before and after each selection method. SCF, embryo aneuploidy, and clinical outcomes were assessed and compared. A total of 29 men underwent semen analysis according to WHO 2010 criteria. SCF was carried out by TUNEL on at least 500 spermatozoa with an established threshold of 15%. DSBs were measured by a neutral Comet test using a modified in-house protocol. A threshold of ≤3% was considered normal, with at least 200 spermatozoa assessed per patient. Clinical outcomes were compared between ICSI cycles performed with the DGC and MFSS methods. Preimplantation genetic testing for aneuploidy (PGT-A) was carried out on the resulting blastocysts. Euploid embryos were replaced into the uterine cavity, and implantation and clinical pregnancy rates (CPR) were recorded. A total of 29 men (40±7 years) had the following semen parameters: a concentration of 38.7 ± 39 x106/mL, 31 ± 17% motility, and 2.1 ± 1% morphology. After DGC and MFSS, the sperm concentration was 18±29 and 7.3±11 x106/mL, with 55.2±30% and 98.3±2% motility, respectively (P<0.0001). The average SCF decreased from 22±9% in the raw samples to 16±3% following DGC, and dropped to 1.8±1% after MFSS processing (P<0.0001). The DSB rate was 3.8±2% in the raw samples, 2.8±1% after DGC, and only 0.3±0.1% after MFSS. These couples (female partner, 37.2±5 years) underwent 42 ICSI cycles with DGC-selected spermatozoa and achieved a fertilization rate of 61.8% (254/411) with 23.8% (26/109) of the embryos identified as euploid. The replacement of these euploid embryos yielded an implantation rate of only 4.3% (1/23) and a CPR of 8.3% (1/12), with 8.3% (1/12) resulting in a pregnancy loss. When these couples underwent subsequent ICSI cycles with MFSS, a fertilization rate of 80.2% (334/416) (P<0.00001) was achieved, with 48.9% (90/184) of the embryos identified as euploid (P<0.0001). A total of 29 embryos were replaced, resulting in an implantation rate of 65.5% (19/29; P<0.0001) and a CPR of 73% (19/26) (P<0.001), resulting in 5.2% (1/19) pregnancy loss and 69.2% (18/26) ongoing pregnancy or deliveries (P<0.0001). Spermatozoa with DNA damage, particularly double-stranded nicks/breaks, may impair the ability of the oocyte to repair the male genome. Because the male genome contributes to embryo aneuploidy, selecting spermatozoa with the highest motility and chromatin integrity may enhance the chances of obtaining a euploid embryo for transfer and boost embryo implantation.