Received February 02, 2020
Accepted September 22, 2020

Objective: Busulfan is one of the most common chemotherapeutic drugs and has the ability to induce apoptosis in testicular germ cells, which leads to infertility. In this study, the effects of ozone therapy and melatonin were evaluated on testicular disorders induced by busulfan. Methods: In this study, 24 male mice were divided into four groups: control group, groups treated with busulfan, busulfan/melatonin, and busulfan/ozone. At the end of a 35-day period, blood samples were taken from the mice and their testosterone levels were measured. Both of the mice’s testes were removed and weighed, after which one of them was used for evaluation of morphology by Johnson’s score, as well as for measuring the diameter and thickness of seminiferous tubules. The other testis was homogenized for measuring Malondialdehyde (MDA) and antioxidant status using Catalase (CAT), Super Oxide Dismutase (SOD), and Total Antioxidant Capacity (TAC) levels. Epididymis spermatozoa were also used to evaluate motility, morphology, and sperm count. Results: Busulfan significantly reduced the testis quality (weight, sperm parameters, testosterone, CAT, SOD, and TAC levels) and increased MDA and destruction of seminiferous tubules compared to the control group. Ozone and melatonin treatments significantly increased testis quality, sperm parameters, MDA, and antioxidant status, but they did not affect the TAC level. Conclusion: This study showed that similar to melatonin, ozone can reduce the effect of busulfan toxicity on mouse testis. However, further studies are needed to understand the precise mechanism of ozone function on testis.