Multivariate Analysis of Inflammatory and Regenerative Mediators in an In Vitro Canine Osteoarthritis System Treated...

Osteoarthritis (OA) is characterized by interactions among inflammatory and extracellular matrix mediators that may not be fully captured through individual evaluation. This study investigated the multivariate organization and temporal dynamics of selected mediators in a lipopolysaccharide-challenged in vitro canine OA cartilage-synovium explant coculture system treated with platelet-rich gel supernatant (PRGS) and platelet-poor gel supernatant (PPGS). Tissue explants obtained from six dogs were exposed to PRGS or PPGS at 25% and 50% concentrations. Interleukin-1β (IL-1β), IL-10, transforming growth factor beta 1, platelet-derived growth factor BB, and hyaluronic acid were quantified at 1 and 48 h. Principal component analyses (PCA) were performed using both individual mediator concentrations and biologically relevant mediator ratios, which were intended to capture the balance among inflammatory, regulatory, GF-related, and matrix-associated responses beyond absolute mediator concentrations alone, and principal component scores were evaluated using linear mixed-effects models. Mediator-based PCA revealed that variability was distributed across multiple dimensions, with significant treatment, time, and treatment × time effects detected in different principal components. Ratio-based PCA concentrated a larger proportion of total variance within the first two components (81.1%) than mediator-based PCA (61.4%) and identified treatment- and time-related effects involving regulatory and matrix-associated mediator balances. Ratio-derived PCA showed greater structural stability across resampling analyses than the mediator-based approach. PRGS 50% generated the most pronounced multivariate separation, particularly within components associated with GF- and matrix-related mediator profiles. These findings indicate that biologically relevant mediator ratios provide complementary information regarding the organization of inflammatory and regenerative responses in this experimental system.