Knee joint distraction (KJD) has been associated with clinical and structural improvement and synovial fluid (SF) marker changes. However, structural changes have not yet been shown satisfactorily in regular care, since radiographic acquisition was not fully standardized. AI-based modules have shown great potential to reduce reading time, increase inter-reader agreement and therefore function as a tool for treatment outcome assessment. To 1) analyze structural changes after KJD in patients using this AI-based measurement method, and relate these changes to 2) clinical outcome and 3) SF markers. 20 knee OA patients (<65 years old) were included in this study. KJD treatment was performed using an external fixation device, providing 5 mm distraction for 6-7 weeks. SF was aspirated before, during and immediately after treatment. Weight-bearing antero-posterior knee radiographs and WOMAC questionnaires were collected before and ∼one year after treatment. Radiographs were analyzed with the Knee Osteoarthritis Labelling Assistant (KOALA, IB Lab GmbH, Vienna, Austria) to obtain whole-joint KL grade, medial and lateral joint space narrowing (JSN) scores, compartmental continuous minimum and standardized JSW, and osteophytes and sclerosis scores of the medial and lateral tibia and femur. 10 Pre-defined biomarker levels in SF were measured by immunoassay. Radiographic one-year changes were analyzed using Wilcoxon Signed Rank tests for categorical and paired t-test for continuous variables. Linear regression was used to calculate associations between changes in standardized joint space width (JSW) and WOMAC, and changes in JSW and SF markers. After treatment, radiographs showed an improvement in Kellgren-Lawrence grade in 7 of 16 patients that could be evaluated; 3 showed a worsening. Joint space narrowing scores and continuous JSW measures improved especially medially. Radiographic changes were not statistically significant (p>0.1), except for the medial JSW (p<0.05). A greater improvement in JSW was significantly associated with a greater improvement in WOMAC pain (β=0.64; p=0.020). A greater increase in MCP1 (β=0.67; p=0.033) and lower increase in TGFβ1 (β=-0.787; p=0.007) were associated with JSW improvement. Despite the small number of patients, also in regular care KJD treatment shows joint repair as measured automatically on radiographs, significantly associated with certain SF marker change and even with clinical outcome. Versus Arthritis project grant (20783), Centre for OA Pathogenesis Versus Arthritis (grants 20205 and 21621), the Kennedy Trust for Rheumatology Research, and ReumaNederland (ISP14-3-301/16-1-404). CS, EB, MD: ImageBiopsyLab GmbH. NA CORRESPONDENCE ADDRESS: m.p.jansen-36@umcutrecht.nl