Background and objective: Cisplatin is an alkylating agent that has become a first-line therapy for some tumors. However, overproduction of reactive oxygen species (ROS) by cisplatin can cause male infertility, which can affect patients’ quality of life. Melatonin, which has the ability to provide resistance against oxidation, is a potential therapy for male infertility caused by cisplatin. Material and methods: Normal human spermatozoa samples were divided into (i) control group incubated with physiological saline solution; (ii) cisplatin group incubated with cisplatin; (iii) melatonin group incubated with melatonin; and (iv) melatonin + cisplatin group incubated with melatonin and cisplatin. Spermatozoa motility was measured using a computer-aided semen analysis system. Additionally, we determined spermatozoa apoptosis through terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling. The mitochondrial membrane potential (\mathrm{\Delta }\mathrm{\Psi }m) and levels of ATP, ROS, and malondialdehyde, as a metabolite of oxidative stress, were measured to study the mechanism by which melatonin can protect human spermatozoa. Results: Melatonin improved sperm motility, and decreased apoptosis and increased the \mathrm{\Delta }\mathrm{\Psi }m value and level of ATP in sperm. More importantly, the overproduction of ROS induced by cisplatin was reduced by melatonin. Furthermore, western blotting revealed that melatonin increased the expression of antioxidant pathway components nuclear factor erythroid 2-related factor (Nrf2) and heme oxygenase-1 (HO-1) that was reduced by cisplatin. Conclusion: Our studies suggested that melatonin might protect human spermatozoa against the effects of cisplatin through Nrf2/HO-1.