P-042 Sperm Selection Based on Speed Using Rheotaxis in a Parallel Array of Microfluidic Apertures
Authors: Yaghoobi M , Xie P , Cheung S , Kocur O , Rosenwaks Z , Abbaspourrad A , Palermo G 
Published: June 2023

Abstract

Study question
Does selecting human spermatozoa using a dynamic microfluidic system impact semen parameters and genomic characteristics?

Summary answer
Flow rate adjustment in a dynamic microfluidic device selects spermatozoa with superior DNA integrity. It also appears to select a higher proportion of Y-chromosome-bearing spermatozoa.

What is known already
Mammalian spermatozoa demonstrate rheotactic motion against the flow. This is important in order to overcome the fluid drag force in the female genital tract to reach the oocyte. Sperm processing methods have been developed based on these principles; more recently, static and dynamic microfluidic devices have been tested due to the association between high progressive motility and greater genomic integrity. These devices also appear to have an impact on sperm sex selection.

Study design, size, duration
Human ejaculates (n = 7) were evaluated and simultaneously processed by density gradient centrifugation (DGC) or a novel microfluidic sperm sorter (MFSS) with an adjustable flow rate inspired by the constriction of the uterotubal junction. Concentration, motility, morphology, and SCF were assessed and compared between raw, DGC-, and MFSS-processed specimens. To identify differences in the SCF and the proportion of X- and Y-spermatozoa, spermatozoa isolated at different flow rates (150, 250, 350, and 450 μl/h) were evaluated.

Participants/materials, setting, methods
Ejaculates were evaluated by standard semen analysis according to WHO 2021 criteria. Following complete liquefaction, 1 mL of sample was used for DGC, and 0.8 mL of semen was used for the prototype MFSS at 0.2 mL for each flow rate. SCF was assessed by TUNEL assay with a normal threshold of ≤ 15%. Fluorescent in situ hybridization (FISH) was performed using X- and Y-chromosome probes.

Main results and the role of chance
A total of 7 men (35.4±5 years) had the following semen parameters: volume of 3.3±2 mL, concentration of 76.7±33 x106/mL, 45.5±1% motility, and a normal morphology of 3.2±0%. When comparing the post-processing parameters between the two methods, DGC yielded a concentration of 49.9±25 x106/mL and MFSS yielded a reduced concentration of 4.5±5x106/mL (P&lt;0.0001). DGC yielded 91.1±1% motility and normal morphology of 3.2±0.4%. Albeit at a cost of concentration, MFSS yielded spermatozoa with significantly higher motility (96.3±3%; P&lt;0.0001) and a higher proportion with normal morphology (4.0±0.6%; P&lt;0.0001). Moreover, the SCF of the raw sample was 9.4±2.0%; although the SCF was 8.1±3.6% with DGC, it was 4.5±2% (P&lt;0.001) with MFSS.
An analysis was performed within the different flow rates of the MFSS; a 250 μl/h flow rate yielded the highest recovery rate (42.0±6%; P&lt;0.001), highest concentration (5.6±4 x106/mL; P&lt;0.01), highest motility (97.0±2.2%; P&lt;0.05), and a normal morphology rate (4.3±1%). The SCF for this flow rate was &lt;3%. Surprisingly, we noticed that the proportion of Y-chromosome–bearing spermatozoa positively correlated to the flow rate, suggesting that there may be a subtle difference in the kinetics of male and female spermatozoa.

Limitations, reasons for caution
Although our novel MFSS platform is a promising technique, it is costly and requires specific expertise and training. This is a limited observation and requires integration of clinical outcomes to prove its benefits.

Wider implications of the findings
A microfluidic device with a controlled flow rate allowed the fine-tuned selection of spermatozoa with superior hydrodynamic and kinetic characteristics and a higher genomic integrity. Once automated, this device may help to select spermatozoa with greater embryo developmental competence and be useful to isolate spermatozoa of the desired sex.

Trial registration number
N/A