Extraplatelet Molecule Enrichment Enhances the Ability of Platelet-Rich Plasma Releasate to Buffer IL-1β-Induced Inflammation and Oxidative Stress in a Primary Human Chondrocyte–Synoviocyte Co-Culture In Vitro System of Osteoarthritis

Abstract

To evaluate the anti-inflammatory and antioxidant effects of standard platelet-rich plasma (sPRP) and balanced protein-concentrate plasma (BPCP), which is a next-generation formulation enriched in platelet-derived and extraplatelet bioactive molecules, in an indirect co-culture model of primary human chondrocytes (hCh) and synoviocytes (hSv) under IL-1β-induced inflammatory conditions. hCh and hSv were co-cultured in a transwell system (indirect co-culture) and stimulated with IL-1β (10 ng/mL) for 48 h in the presence of 10% sPRP, 10% BPCP, or serum-free medium (SF). Gene expression in hCh was analyzed by RT-qPCR for inflammatory mediators, NF-κB pathway genes, matrix-degrading enzymes and apoptosis markers. Cytokine and MMP secretion in supernatants were quantified by Luminex assay, and intracellular ROS levels were measured in hSv and hCh direct co-culture by DCFDA fluorescence after 4 h of incubation with PRP previously exposed for 24 h to IL-1β. Both sPRP and BPCP significantly reduced MCP-1 (p = 0.02) and BCL-2 expression (p < 0.0001) versus SF. Both formulations also decreased secretion of IL-1β, TNF-α, IL-6, IL-8, MCP-1, MMP-1, MMP-3, and MMP-13 compared to SF. BPCP showed greater reductions in IL-6 and MCP-1 compared with sPRP. MMP-7 and MMP-9 secretion increased under both PRP conditions, consistent with tissue remodeling activity. Both formulations significantly reduced intracellular ROS levels measured by DCFDA fluorescence, with a stronger effect observed for BPCP. sPRP and BPCP exerted consistent anti-inflammatory and antioxidant effects in a multicellular simulated osteoarthritic in vitro model. BPCP demonstrated enhanced protein-level regulation in certain markers, suggesting that enrichment in extraplatelet molecules may improve PRP efficacy.