C57BL/6J (WT) mice (8 weeks old, male) were purchased from Nanjing Medical university, and AMPK-KO mice were purchased from Shanghai Model Organisms. They were used to create an OA model by destabilization of the medial meniscus surgery (DMM) (n = 6 per group). Briefly, after the mice were anaesthetized, a medial articular incision was made to expose the leftjoint cavity, and then the tibial collateral ligament was transected. Finally, the articular incision was closed. In the control group, only the joint cavity was opened. One week after surgery, 1 mg/kg baicalein (MCE, HY-N0196) per knee, 1 mg/kg ML385 (MCE, HY-100523) per knee, 1 mg/kg AICAR (MCE, HY-13417) per knee or 1 mg/kg of the ferroptosis inhibitor ferrostatin-1 (Fer-1, MCE, HY-100579) was injected into the joint cavity of the mice once a week. Meanwhile, saline was injected into the control group. Mice were sacrificed after surgery 10 weeks.

C57BL/6J (WT) mice (8 weeks old, male) were purchased from Nanjing Medical university, and AMPK-KO mice were purchased from Shanghai Model Organisms. They were used to create an OA model by destabilization of the medial meniscus surgery (DMM) (n = 6 per group). Briefly, after the mice were anaesthetized, a medial articular incision was made to expose the leftjoint cavity, and then the tibial collateral ligament was transected. Finally, the articular incision was closed. In the control group, only the joint cavity was opened. One week after surgery, 1 mg/kg baicalein (MCE, HY-N0196) per knee, 1 mg/kg ML385 (MCE, HY-100523) per knee, 1 mg/kg AICAR (MCE, HY-13417) per knee or 1 mg/kg of the ferroptosis inhibitor ferrostatin-1 (Fer-1, MCE, HY-100579) was injected into the joint cavity of the mice once a week. Meanwhile, saline was injected into the control group. Mice were sacrificed after surgery 10 weeks.

C57BL/6 J mice (8 weeks old, female) were purchased from Dossy Experimental Animal Limited Company (Chengdu, China). For surgery, mice were anaesthetized with pentobarbital sodium (100 mg/kg, injected intraperitoneally) and subjected to unilateral ACLT procedures. 28 The sham group received a skin incision and suturing without patellar dislocation or ligament transection. For virus injection, mice were intraarticularly injected with 1 x 10⁹ pfu (8 ul) of mock or AdEpas1 virus after one week of surgery. For Fer1 (MCE, Monmouth Junction, HY100579) injection, mice were intraarticularly injected with 1 mg/kg Fer1 or with vehicle two weeks after surgery, the injection was repeated once a week.

Thirty-two 8 weeks old male C57BL/6 mice were used in this study. Eight mice were randomly allocated to the sham surgery group. After anesthetized by intraperitoneal injection of pentobarbital (35 mg/kg), sham surgery group were subjected to sham surgery. Then, destabilized medial meniscus (DMM) were performed to establish the OA model in twenty-four mice. DMM mice were randomly divided into three groups (N=8 for each group): the DMM group, the DMM+0.1mg/kg ferrostain-1 group and the DMM+1mg/kg ferrostain-1 group. The DMM+0.1 mg/kg ferrostain-1 group and the DMM+1mg/kg ferrostain-1 group were intraarticularly injected with 0.1 mg/kg or 1 mg/kg ferrostain-1 respectively. The sham surgery group and DMM group were intraarticularly injected with same volume of vehicle. The injection was repeated twice a week for 8 consecutive weeks.

Thirty-two 8 weeks old male C57BL/6 mice were used in this study. Eight mice were randomly allocated to the sham surgery group. After anesthetized by intraperitoneal injection of pentobarbital (35 mg/kg), sham surgery group were subjected to sham surgery. Then, destabilized medial meniscus (DMM) were performed to establish the OA model in twenty-four mice. DMM mice were randomly divided into three groups (N=8 for each group): the DMM group, the DMM+0.1mg/kg ferrostain-1 group and the DMM+1mg/kg ferrostain-1 group. The DMM+0.1 mg/kg ferrostain-1 group and the DMM+1mg/kg ferrostain-1 group were intraarticularly injected with 0.1 mg/kg or 1 mg/kg ferrostain-1 respectively. The sham surgery group and DMM group were intraarticularly injected with same volume of vehicle. The injection was repeated twice a week for 8 consecutive weeks.

A total of 50 healthy, wild-type adult male C57BL/6 mice (age, 8weeks; weight 20-25 g) were purchased from the Laboratory Animal Center of Ningxia Medical University. After mice were anesthetized with an intraperitoneal injection of 1% sodium pentobarbital (60 mg/kg), the meniscotibial ligament of the right knee was transected to free the anterior crus of the meniscus, and sham surgery was performed without transecting the meniscotibial ligament. Mice were injected intra-articularly with Erastin (15 mg/kg), and the sham and DMM groups were injected with an equal volume of saline twice a week. Moreover, Met was administered by gavage (200 mg/kg/day). Mice were killed by overdose anesthesia 8 weeks after surgery, and the right knee joint was collected for subsequent experiments.

Mice were orally administered 0.1 g/kg Co-Q10 daily for 8 weeks after the DMM model was established to investigate the therapeutic effect of Co-Q10 in GPX4-CKO mice with osteoarthritis. Mice and rats were anaesthetized with pentobarbital. Destabilization of the medial meniscus (DMM) surgery was performed under a microscope. The incision was sutured and disinfected daily until it healed.

A total of 16 Sprague-Dawley rats (6 weeks old, female) were purchased from Guangdong Medical Laboratory Animal Center (Foshan, China) and randomly divided into four groups (n = 4): Sham, Hulth, Hulth + ScpI2 (0.1 mg/kg) (Vitas-M, Apeldoorn, Netherlands), Hulth + ScpI2 (0.5 mg/kg). One week after surgery, the rats were intraarticularly injected with vehicle or ScpI2 twice a week for 4 consecutive weeks.

A rat model of OA with destabilization of the medial meniscus was established . After anesthetization with 3% pentobarbital sodium (Tocris, Avonmouth, UK), the hair on the right knee was clipped. Right knee was subsequently exposed before an incision was made in the medial aspect of the joint capsule, then the anterior cruciate ligament was transected, and the medial meniscus was completely resected in a manner that did not injure the articular cartilage. Subsequently, the joint was irrigated with normal saline, the capsule was sutured with 4-0 chromic catgut, and the skin was closed with 4-0 nylon mattress sutures. And the rats were allowed to move, eat, and drink freely after surgery. Experimental groups are as follows: sham group (A medial incision was made to expose the knee joint cavity, and sutured), OA model group (Destabilization of the medial meniscus), sh-NC group (OA rats were injected with sh-NC), and sh-SND1 group (OA rats were injected with sh-SND1).

A rat model of OA with destabilization of the medial meniscus was established . After anesthetization with 3% pentobarbital sodium (Tocris, Avonmouth, UK), the hair on the right knee was clipped. Right knee was subsequently exposed before an incision was made in the medial aspect of the joint capsule, then the anterior cruciate ligament was transected, and the medial meniscus was completely resected in a manner that did not injure the articular cartilage. Subsequently, the joint was irrigated with normal saline, the capsule was sutured with 4-0 chromic catgut, and the skin was closed with 4-0 nylon mattress sutures. And the rats were allowed to move, eat, and drink freely after surgery. Experimental groups are as follows: sham group (A medial incision was made to expose the knee joint cavity, and sutured), OA model group (Destabilization of the medial meniscus), sh-NC group (OA rats were injected with sh-NC), and sh-SND1 group (OA rats were injected with sh-SND1).

Adult male C57BL/6 mice (eight weeks of age) were used for in vivo experiments. We randomly divided mice into four groups with 9 mice per group: SHAM + AAV9-GFP, SHAM + AAV9-NCOA4, DMM + AAV9-GFP, and DMM + AAV9-NCOA4 groups. For in vivo experiment of SP600125 administration, we randomly divided mice into three groups with 6 mice per group: DMM + AAV9-GFP, DMM + AAV9-GFP + SP600125, and DMM + AAV9-NCOA4 + SP600125. 10 ul of SP600125 (1 mg/kg) or vehicle solution was injected articularly once per week for 7 weeks.