Objective To evaluate the effects of systemic medical ozone (O-3) application and hyperbaric oxygen (HBO) therapy on surgically induced knee osteoarthritis (OA) in a rat model. Materials and methods We performed anterior cruciate ligament transection (ACLT) in order to create experimental OA in the right knees of 27 male rats. The left knee joints of all rats were sham-operated without ACLT as the negative control group. The rats were randomly assigned into three groups: (1) control group, which received no treatment; (2) O-3 group, which received intraperitoneal 30 mu g medical O-3; (3) HBO group, which received HBO therapy for 60 minutes twice a day. We sacrificed the rats on the tenth week after the operation. We evaluated the degree of OA using Mankin scores. Results As a result of histopathological examination, the mean Mankin scores in the right knees with ACLT were 8.17 +/- 2.12 in the control group, 6.22 +/- 1.56 in the HBO group, and 4.72 +/- 1.30 in the O-3 group. The differences between the O-3 group and the HBO group and the O-3 group and the control group were found to be statistically significant (p 0.001, p 0.039, respectively). There was no difference between the HBO group and the control group (p 0.086). Conclusions The results of the present study show that systemic medical O-3 application was more effective than HBO therapy and may reduce development of cartilage damage and prevent OA formation.