a, b, Representative images of CH25H immunostaining (a) and safranin-O staining (b) in mice that underwent intra-articular injection of Ad-C or Ad-CH25H once weekly for 3 weeks (n = 12 mice per group). c, Representative immunostaining of CH25H in cartilage sections of 20-week-old wild-type and Ch25h−/− (KO) mice (n = 12 per group). d, Staining of whole skeleton, hindlimbs and forelimbs, and the length of the indicated bones, in wild-type (n = 6) and Ch25h−/− (n = 12) mice. e, Staining of growth plates in metatarsal bones, along with the lengths of the indicated zones, from Ch25h−/− mice and wild-type littermates (n = 20 mice per group). f, Representative immunostaining images of CH25H, DIPEN and NITEGE in Ch25h−/− and wild-type littermates subjected to sham or DMM surgery (n = 8 mice per group). g, OA parameters quantified 8 and 12 weeks after Ch25h−/− and wild-type littermates were subjected to sham operation (n = 8 mice) or DMM surgery (n = 12 mice). h, Wild-type and Ch25h−/− mice fed regular diet or HCD were subjected to sham or DMM operations, with OA parameters quantified at 4 (n = 8 mice for sham and 6 mice for DMM) and 6 weeks (n = 9 mice for sham and 9 mice for DMM). i, von Frey (n = 10 mice per group) and hot-plate (n = 15 mice per group) pain assays in DMM-operated Ch25h−/− and wild-type mice at the indicated times after operation. Means ± s.e.m. with two-tailed t-test (d, e). Means ± 95% CI with Mann–Whitney U test for OARSI grade, osteophyte maturity and von Frey assay; means ± s.e.m. with two-tailed t-test for SBP thickness and hot-plate assay (g−i). Exact P values (for P < 0.001) can be found in the accompanying Source Data. Scale bars, 25 μm.