Fig 1.
Effect of pH on the motility and velocity of sperm.
Purified human sperm were incubated in pH 5.2, 6.2, 7.2 and 8.2 nutrient solutions for 15, 30, 60, 90 and 120 min, respectively. (A) Sperm mobility (PR + NP): a,b,e-g p<0.05, statistically significant different from pH 7.2 group at 15, 30, 60, 90, and 120 min, respectively; c p<0.05, statistically significant different from pH 6.2 group at 15 min; dp<0.05, statistically significant different from pH 5.2 group at 15 min. (B) PR: h,i,l-n p<0.05, statistically significant different from pH 7.2 group at 15, 30, 60, 90, and 120 min, respectively; jp<0.05, statistically significant different from pH 6.2 group at 15 min; kp<0.05, statistically significant different from pH 5.2 at 15 min. (C) mean straight line velocity (VSL): o-rp<0.05, statistically significant different from pH 7.2 group at 30, 60, 90 and 120 min, respectively; t p<0.05, statistically significant different from pH 6.2 group at 15 min; sp<0.05, statistically significant different from pH 5.2 group at 15 min. (D) mean curvilinear velocity (VCL): u-w p<0.05, statistically significant different from pH 7.2 group at 60, 90 and 120 min, respectively; y p<0.05, statistically significant different from pH 6.2 group at 15 min; xp<0.05, statistically significant different from pH 5.2 group at 15 min. and; (E) mean average path velocity (VAP): α, β, δ, ε, η p<0.05, statistically significant different from pH 7.2 group at 15, 30, 60, 90 and 120 min, respectively; θ p<0.05, statistically significant different from pH 6.2 group at 15 min; λp<0.05, statistically significant different from pH 5.2 group at 15 min (N = 3).
Fig 2.
The viability of sperm in nutrition solutions with different pH.
(A), the images of HOS spermatozoa in different pH nutrition solutions were obtained by CASA. (B), HOS rate markedly declined in pH 5.2 and 6.2 nutrition solutions in comparison with pH 7.2 solution, * mean p<0.01.
Fig 3.
Linear regression between HOS rate and sperm movement.
(A), the correlation between HOS rate and sperm mobility, r = 0.98, p<0.05. (B), the correlation between HOS rate and sperm PR, r = 0.98, p<0.05.
Fig 4.
Sperm penetration decreased in acidic environments.
Compared with pH 7.2 group, the sperm ascending altitude in pH 5.2 and 6.2 groups was significantly lower * p<0.01 (N = 9).
Fig 5.
Sperm Na+/K+-ATPase activity decreased in acidic environments.
Compared with pH 7.2, sperm Na+/K+-ATPase activity decreased evidently at pH 5.2 or 6.2, while there were no significant differences with that at pH 8.2. * p<0.05 (N = 4).
Fig 6.
Measurement of intracellular Ca2+ during human sperm capacitation by FCM.
(A), the FCM results of spermatozoa in sperm nutrition solutions at pH 5.2, 6.2, 7.2 and 8.2, respectively. (B), compared with pH 7.2 group, the mean fluorescent intensity of intracellular Ca2+ in pH 5.2 and 6.2 groups significantly declined, but the pH 8.2 group did not show significant differences. * p<0.05 (N = 6)
Fig 7.
Measurement of intracellular Ca2+ during human sperm capacitation by LSCM.
(A), the LSCM results of spermatozoa in sperm nutrition solutions at pH 5.2, 6.2, 7.2 and 8.2, respectively. (B), compared with pH 7.2 group, the mean fluorescent intensity of intracellular Ca2+ in pH 5.2 and 6.2 groups significantly declined, but the pH 8.2 group did not show significant differences. * p<0.05 (N = 10)