BACKGROUND Cryptorchidism is the most common developmental abnormality of the male reproductive system. If left untreated, it results with infertility and testicular cancer. According to current evidence, surgery is the mainstay of treatment, and hormonal therapy approaches are still under investigation. For the protection of testicular functions, antioxidants have emerged as novel options. This study aimed to evaluate the protective properties of ozone, a strong antioxidant, on testicular tissue. MATERIAL AND METHODS Thirty-five male Wistar-albino rats, 1-month-old, were used for the study. Groups were formed as follows: 1) control, 2) sham surgery (cryptorchidism), 3) cryptorchidism plus ozone, 4) cryptorchidism plus human chorionic gonadotropin (hCG), and 5) ozone plus hCG. Surgical procedures were performed on all rats except the control group. All rats except the control group were used to create an experimental cryptorchidism model, and left testes of animals were surgically placed into the abdomen. After 1 month of surgery, groups 3, 4, and 5 were given corresponding treatments intraperitoneally for 4 weeks. At the end of the study period, testicular atrophy index (TAI) and testicular sperm motility (TSM) were assessed and biochemical, histopathological, and immunohistochemical tests were performed. RESULTS TAI and TSM were higher in the ozone, hCG, and ozone plus hCG groups than in the sham surgery group (p=0.001). TSM in the ozone group was significantly higher than in the hCG and ozone plus hCG groups. In biochemical analyses, the parameters of oxidative stress (GPx1, MDA, CAT, GSH, SOD) indicated increased oxidative activity in cryptorchidism, which was resolved by applying ozone and hCG (p=0.001). In addition, apoptotic markers, Caspase 3 and bcl-2 were significantly decreased by applying ozone and hCG (p=0.001). CONCLUSIONS Results of this study suggest that ozone therapy, either as a single agent or in combination with hCG, is a promising approach for protection of testicular functions.