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Background: Osteoarthritis (OA) is a heterogeneous joint disorder traditionally considered mechanically driven; however, evidence indicates that inflammatory mechanisms contribute to symptom expression. Exploratory analyses of peripheral biomarkers may provide insights into systemic inflammation in symptomatic knee OA, but formal phenotypic validation requires dedicated clustering or longitudinal studies. Objective: To examine associations between clinical pain, functional impairment, and circulating inflammatory biomarkers in patients with knee OA compared with healthy controls. Methods: In this prospective, single-center study, patients aged 40–80 years with radiographically confirmed knee OA and chronic knee pain were compared with age- and sex-matched healthy controls. Pain intensity and functional status were assessed using the Visual Analogue Scale (VAS) and the Knee Injury and Osteoarthritis Outcome Score (KOOS). Circulating inflammatory biomarkers, including cytokines and matrix metalloproteinases, were quantified using multiplex immunoassays. Statistical analyses included adjusted linear regression models, with age and BMI as covariates, and multiple testing correction using the Benjamini–Hochberg procedure (FDR alpha error 5%). Results: OA patients exhibited higher circulating levels of TNF-α, IL-6, IL-8, MMP-1, MMP-3, TNFSF13, TNFSF13B, and pentraxin-3 compared with controls (p < 0.01). No significant sex differences were observed. KOOSs correlated with IL-6 and IL-10 levels, suggesting an association between systemic inflammatory activity and functional limitation. All findings are presented as exploratory and associative. Conclusions: Patients with knee OA display systemic inflammatory biomarker differences associated with pain and functional impairment. These results support the role of inflammation in OA symptoms within an exploratory framework. Larger, longitudinal studies are warranted to validate these observations. Full article