P-117 A novel method to select spermatozoa with the best morphokinetic characteristics and superior genomic integrity
Authors: Leung T , Xie P , Kocur O , Sung C , Rosenwaks Z , Palermo G 
Published: June 2022

Abstract

Study question
Can an automated device based on membrane electrophoresis be used to select spermatozoa with superior morphokinetic characteristics and lower sperm chromatin fragmentation (SCF)?

Summary answer
Albeit at an expense of concentration, a membrane-electrophoretic device was able to isolate a proportion of spermatozoa with the highest motility, morphology, and genomic integrity.

What is known already
Conventional sperm selection based on density gradient centrifugation (DGC) can enhance the proportion of progressively motile spermatozoa; however, DGC methods are limited in their ability to select spermatozoa with higher genomic integrity. Membrane-based microfluidic technologies (MFSS) have already been used clinically to select for spermatozoa with a superior chromatin status; however, these methods are based on the intrinsic characteristics of motile spermatozoa capable of selecting themselves. By incorporating an extrinsic electrophoretic drive, membrane electrophoresis would overcome a limitation of progressive motility in the semen sample and be able to isolate viable gametes with better morphology and reduced SCF.

Study design, size, duration
From August 2020 to December 2021, semen samples from 68 men were evaluated by standard semen analysis and simultaneously processed by DGC or a novel membrane-electrophoretic sperm sorter (EPSS) to select for progressively motile spermatozoa. Concentration, motility, progressive motility, morphology, and SCF were measured and compared between raw, DGC-, and EPSS-processed specimens.

Participants/materials, setting, methods
Fresh ejaculates were evaluated by standard semen analysis according to WHO 2021 criteria. Following complete liquefaction, specimens were divided into two equal aliquots for DGC and EPSS sorting. SCF was assessed by terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay under fluorescent microscopy; at least 500 spermatozoa were evaluated for each specimen with a normal threshold of ≤ 15%.

Main results and the role of chance
A total of 68 men with an average age of 38.1±6 years had the following average semen parameters: volume of 3.1±1 mL, concentration of 58.2±33 x106/mL, 43.3±9% motility, 40.5±9% forward progression motility, and a normal morphology of 2.9±0.8%. When comparing the final parameters between the EPSS and DGC methods, despite a reduced sperm concentration (7.3±9 x106/mL vs 45.0±36 x106/mL, P&lt;0.0001, respectively), EPSS was highly selective for motile spermatozoa, yielding significantly higher motility (93.1±16% vs. 86.8±15% P&lt;0.0001) and forward progressive motility (92.3±17% vs 85.2±15%, P&lt;0.0001). Despite an overall lower sperm recovery rate (21.4±25% vs 75.4±17%, P&lt;0.0001), normal morphology improved to 3.4±0.8% (P&lt;0.0001) after EPSS but remained unchanged in the DGC-processed sample. Although both EPSS and DGC improved SCF from 12.2±9% in the raw specimen to 4.8±6% and 6.6±7% (P &lt; 0.0001), respectively, EPSS outperformed DGC (P &lt; 0.05). Moreover, the DGC method took up to 60 min to process, whereas the EPSS technique took a total of 6 min.

Limitations, reasons for caution
The selection of spermatozoa by EPSS is a promising technique to isolate progressively motile spermatozoa with enhanced morphology and superior chromatin integrity, albeit at a lower concentration. This is a preliminary study; the benefit and safety of this method must be further demonstrated by insemination or IVF treatment.

Wider implications of the findings
A membrane electrophoresis device may be a viable alternative method to MFSS to identify spermatozoa with superior morphokinetic characteristics and intact chromatin. Moreover, incorporating an automated device can reduce gamete processing time while minimizing labor costs and inter-operator errors.

Trial registration number
N/A