Efficacy and Safety of Visible and Near-Infrared Photobiomodulation Therapy on Astenospermic Human Sperm: Wavelength-Dependent Regulation of Nitric Oxide Levels and Mitochondrial Energetics

: Male infertility is a growing global concern, with asthenozoospermia being an important contributing factor. Mitochondrial dysfunction and changes in the metabolism of nitric oxide (NO) are key determinants of reduced sperm motility. This study investigates the effects of photobiomodulation (PBM) with visible and near-infrared (NIR) laser light on sperm of asthenozoospermic patients, focusing on mitochondrial energetic status, oxidative stress, and NO dynamics. Semen samples were irradiated at 450 nm, 635 nm, 810 nm, 940 nm, and 1064 nm at different power levels (0.25, 0.50, 1.00, and 2.00 W) for 60 s on a spot area of 1 cm2. ATP and AMP levels, oxidative stress markers, and NO concentrations were assessed at 10 and 60 min after irradiation, with the ATP/AMP ratio calculated as an index of cellular energy balance. The results show that the PBM modulates the energetic status of spermatozoa in a way dependent on wavelength and dose. Irradiation at 810 nm produced the most marked improvement in energetic status, whereas 635 nm exposure led to a significant decrease in cellular energy levels. NO levels showed a biphasic response, correlated with the visible range and with energy metabolism at 810 nm. Irradiation with 635 nm induced higher NO production with respect to the other wavelengths. Our findings suggest that PBM mainly involves mitochondrial photoreceptors and potentially the heme and flavin groups of nitric oxide synthases, facilitating electron transitions, enhancing the effectiveness of oxidative phosphorylation, and optimizing enzymatic activity. At longer wavelengths (940 nm and 1064 nm), interactions with water and lipids may introduce additional variables that affect membrane fluidity and mitochondrial function differently from shorter wavelengths.