Platelet-rich plasma-loaded hydrogels: diffusivity and release of individual proteins in multicomponent complex environments.

Platelet-rich plasma (PRP) treatments have demonstrated clinical benefit for osteoarthritis (OA), although reported outcomes remain inconsistent. Hydrogel-based delivery devices can enhance therapeutic efficacy by prolonging local protein exposure, but effective design depends on characterizing PRP diffusion and the complex protein-protein and protein-matrix interactions that govern release kinetics. Conventional bulk release studies provide limited mechanistic insight into protein transport within complex, physiologically relevant microenvironments, where macromolecular crowding, protein-protein interactions, hydrogel confinement, and patient-specific synovial fluid composition collectively affect release kinetics. To capture this complexity, we used fluorescence correlation spectroscopy (FCS) to directly quantify diffusion coefficients of four representative PRP proteins with varying size and properties across a series of complex environments that represent PRP, the delivery device, and the intended host microenvironment. Diffusivity decreased with increasing protein size, solution viscosity, crowding density, micro-clot formation, and confinement from the hydrogel. Diffusion coefficients obtained via FCS correlated strongly (R2 = 0.84) with effective diffusivities calculated from bulk release experiments and enabled mathematical modeling of protein release kinetics using Fickian diffusion models from both slab hydrogels and microspheres of varying diameter. Together, this study demonstrated a framework for the utility of FCS in characterizing hindered diffusion within complex, multicomponent biological systems, thereby guiding the rational design of hydrogel-based PRP delivery platforms.