Relationship between sperm-hyaluronic acid (HA) binding assay scores and HA-mediated increase in maintenance of sperm motility and velocity in IVF conditions

Abstract

OBJECTIVE: Hyaluronic acid (HA) substantially increases maintenance of human sperm motility and velocity. Also, HA presence improved IVF pregnancy rates. We studied sperm motility and velocity in Vitrolife G-IVF media with and without added HA.DESIGN: Washed sperm fractions of the same semen samples were incubated with or without added 0.1 mg/mL HA. Sperm motility and velocity were assayed by CASA (Hamilton–Thorne, Beverly, MA) a time-related fashion.MATERIALS AND METHODS: Semen was purified with 45% Isolate gradients (600xg, 20 min). The sperm were washed, and re-suspended in Vitrolife G-IVF media or in G-IVF containing 0.1 mg/mL HA in duplicates. Sperm attributes were assayed by CASA (two different drops, four fields each), after 0, 30, 90, 150 minutes, and overnight incubation, including sperm motility, curvilinear velocity(VCL), pathway velocity(VAP), progressive velocity(VSL), cross-beat frequency(BCF), amplitude head displacement(AHL), and proportions of rapid and medium velocity sperm. The sperm HA-binding scores (proportion of motile sperm bound to HA) were determined in chambers of the sperm-HA binding assay (HBA, MidAtlantic Diagnostics, NJ), in the initial semen, and after 90 minutes and overnight incubation. Statistical analyses were performed by paired-t test and Pearson's correlation using the Sigma-Stat 3.0 software.RESULTS: We studied 30 semen samples (concentration: 116±10.9x106 sperm; motility: 59.9±2.5%, mean±SEM). The HA-mediated improvements in G-IVF-HA vs. G-IVF were as follows: At 90 minutes, increases in VCL (p<0.001), VAP (p<0.001), and in proportions of rapid (p<0.001) and medium (p<0.001) velocity sperm. After the overnight incubation there were respective increases in sperm motility (p<0.05), VCL (p<0.05), VAP (p<0.05) and in rapid velocity sperm (p<0.05). In line with our hypothesis of a potential relationship between sperm HA-receptors (degree of maturity, as it measured by the HA-binding score) and HA-mediated increase in sperm velocity: HA-score vs. VSL (r=0,54, p<0.01), vs. BCF (r=0.47, p<0.05), and vs. rapid/medium velocity sperm (r=0.49 p<0.01).CONCLUSIONS: 1) HA improves the retention of sperm motility and velocity even after 24 hours in IVF conditions. 2) As hypothesized, the HA-related improvement was related to sperm maturity and consequential higher density of HA receptors. 3)s. 4) The HBA-slide test could facilitate selection of husbands who may benefit from addition of HA during IVF. OBJECTIVE: Hyaluronic acid (HA) substantially increases maintenance of human sperm motility and velocity. Also, HA presence improved IVF pregnancy rates. We studied sperm motility and velocity in Vitrolife G-IVF media with and without added HA. DESIGN: Washed sperm fractions of the same semen samples were incubated with or without added 0.1 mg/mL HA. Sperm motility and velocity were assayed by CASA (Hamilton–Thorne, Beverly, MA) a time-related fashion. MATERIALS AND METHODS: Semen was purified with 45% Isolate gradients (600xg, 20 min). The sperm were washed, and re-suspended in Vitrolife G-IVF media or in G-IVF containing 0.1 mg/mL HA in duplicates. Sperm attributes were assayed by CASA (two different drops, four fields each), after 0, 30, 90, 150 minutes, and overnight incubation, including sperm motility, curvilinear velocity(VCL), pathway velocity(VAP), progressive velocity(VSL), cross-beat frequency(BCF), amplitude head displacement(AHL), and proportions of rapid and medium velocity sperm. The sperm HA-binding scores (proportion of motile sperm bound to HA) were determined in chambers of the sperm-HA binding assay (HBA, MidAtlantic Diagnostics, NJ), in the initial semen, and after 90 minutes and overnight incubation. Statistical analyses were performed by paired-t test and Pearson's correlation using the Sigma-Stat 3.0 software. RESULTS: We studied 30 semen samples (concentration: 116±10.9x106 sperm; motility: 59.9±2.5%, mean±SEM). The HA-mediated improvements in G-IVF-HA vs. G-IVF were as follows: At 90 minutes, increases in VCL (p<0.001), VAP (p<0.001), and in proportions of rapid (p<0.001) and medium (p<0.001) velocity sperm. After the overnight incubation there were respective increases in sperm motility (p<0.05), VCL (p<0.05), VAP (p<0.05) and in rapid velocity sperm (p<0.05). In line with our hypothesis of a potential relationship between sperm HA-receptors (degree of maturity, as it measured by the HA-binding score) and HA-mediated increase in sperm velocity: HA-score vs. VSL (r=0,54, p<0.01), vs. BCF (r=0.47, p<0.05), and vs. rapid/medium velocity sperm (r=0.49 p<0.01). CONCLUSIONS: 1) HA improves the retention of sperm motility and velocity even after 24 hours in IVF conditions. 2) As hypothesized, the HA-related improvement was related to sperm maturity and consequential higher density of HA receptors. 3)s. 4) The HBA-slide test could facilitate selection of husbands who may benefit from addition of HA during IVF.