Melatonin-loaded nanoparticles protecting human sperm from oxidative stress during cryopreservation

During the process of sperm cryopreservation, the overproduction of reactive oxygen species (ROS) triggers oxidative stress thereby leading to a reduction in sperm motility and quality. Therefore, it is a feasible strategy to mitigate oxidative damage during cryopreservation by adding antioxidants to freezing media. In this study, we explored the potential of melatonin to protect sperm from oxidative stress-induced damage by evaluating sperm-related parameters after thawing through self-assembly with a hyaluronic acid-bilirubin conjugate into nanoparticles (M@HBn). The optimized M@HBn exhibited uniform spherical morphology with average particle size of 112.57 ± 9.8 nm, PDI of 0.22 ± 0.02, a surface potential of -0.43 ± 1.02 mV and entrapment efficiency of 85.1 ± 4.6%. The addition of 5 μM M@HBn demonstrated a notable enhancement in frozen-thawed human spermatozoa viability, motility, and DNA integrity by scavenging ROS. Additionally, the use of M@HBn supplementation in freezing medium resulted in the most mitochondrial stability and total viability as compared to the other groups. These findings suggest that M@HBn have the potential to serve as a novel drug delivery platform for protecting spermatozoa against from cryodamage while enhancing the quality of cryopreserved sperm and the bioavailability of melatonin.