Osteoarthritis (OA) is a progressive and degenerative disease of the joints, characterized by inflammation and loss of cartilage. Recently, mRNA therapies have emerged as promising disease-modifying treatments for cartilage repair and regeneration. Poly(amidoamine)-based polymeric nanoparticles (PAA-based NPs) were previously developed for intracellular mRNA delivery in chondrocytes, showing high biocompatibility and transfection efficiency. In this work, we aimed to evaluate this delivery system in models simulating the complex joint environment and in vivo in rat knee joints. For this purpose, cationic uncoated NPs and neutral PEG-coated NPs were formulated to test mRNA delivery in different models: (1) a 2D culture of chondrocytes supplemented with synthetic synovial fluid, (2) a cartilage-on-chip platform, (3) an ex vivo culture of mouse knee joints, and (4) an in vivo OA rat model. In the presence of synovial fluid, the PEG-coated NPs showed favorable physicochemical properties, higher cell uptake and equivalent GFP expression as uncoated NPs in the 2D cell culture. Similar observations were made using the cartilage-on-chip platform. In contrast, both NPs appeared to display cartilage penetration and uptake by tissue-resident chondrocytes in ex vivo joint culture. Upon intra-articular administration in vivo, the PAA-based NPs did not affect cartilage integrity in healthy nor OA rat knee joints, although enhanced synovial inflammation was observed. Uncoated NPs showed prolonged retention compared to PEG-coated NPs and higher luciferase expression in OA knee joints than in healthy joints of rats, whereas no difference was found for coated NPs. These results suggest that electrostatic interactions between cationic NPs and the anionic components of the extracellular matrix play a significant role in mRNA delivery to the articular cartilage, and that disease status may affect delivery of nucleic acids dependent on NP properties. In conclusion, PAA-based NPs are a promising platform for intra-articular mRNA delivery in the joints. STATEMENT OF SIGNIFICANCE: In this study, we investigate the application of poly(amidoamine)-based polymeric nanoparticles (PAA-based NPs) for mRNA delivery in the joints, aiming for use in osteoarthritis (OA) treatment. The formulations were tested in in vitro models mimicking the joint environment, and also following intra-articular injection ex vivo and in vivo (OA-induced rats). We demonstrate for positively charged uncoated NPs higher in vivo gene expression in OA knee joints than neutral PEG-coated NPs. However, PEG-coated NPs induced more consistent gene expression in both healthy and OA knee joints. These findings highlight the potential of PAA-based NPs for osteoarthritis research and how the interplay between the NP properties, joint biology and disease state can affect mRNA delivery.
