We found that the higher sperm motility induced by
H2 treatment was maintained for 2.5 h, even in the absence
of H2 (Figure 4), which is clinically enough time
for IVF and ICSI. The decrease in motility after H2 treatment
is due to several reasons, including the reduction
of intracellular ATP, and oxidative stress of oxygen in
the room air [32]. Nevertheless, the motility increased
the following day after a second H2 treatment. The increase
of motility with second treatment of H2 was still
higher than that of N2, indicating that the repeated increase
was due to H2, but not mechanical stimuli. The
repeated stimulation of sperm is likely to be useful in a
clinical setting. Higher variation of the mobility in H2-
treated sperm in Figure 4 may be dependent on its
higher mobility, because the difference of coefficients of
variation between the mobility of H2-treated sperm (0.26
to 0.59%) and that of N2 (0.48 to 0.98%) was very low.