High-efficiency non-viral transfection of primary chondrocytes and perichondrial cells for ex-vivo gene therapy to repair articular cartilage defects

Background: Primary perichondrial cells and chondrocytes have been used to repair articular cartilage defects in tissue engineering studies involving various animal models. Transfection of these cells with a gene that induces chondrocytic phenotype may form an ideal method to affect tissue engineering of articular cartilage. Design: A protocol for high-efficiency transfection of primary perichondrial and cartilage cells was optimized. Plasmids carrying the marker beta -galactosidase (beta -gal), PTHrP and TGF-beta1 genes driven by a strong mammalian promoter were transfected into primary perichondrial cells and chondrocytes. A three-step method was used to achieve high efficiency of transfection: (1) permeabilization of primary cells using a mild detergent, (2) association of plasmid DNAs with a polycationic (poly-l-lysine) core covalently linked to a receptor ligand (transferrin), (3) introduction of cationic liposomes to form the quaternary complex. For in-vivo assessment, polylactic acid (PLA) scaffolds seeded with beta -gal transfected perichondrial cells were implanted into experimentally created osteochondral defects in rabbit knees for 1 week. Results: The efficiency of transfection was determined to be over 70% in vitro. The transformed cells continued to express beta -gal, in vivo for the entire test period of 7 days. Furthermore, primary perichondrial cells transfected with TGF-beta1 and PTHrP over-expressed their cognate gene products. Conclusion: The ability to transfect autologous primary perichondrial cells and chondrocytes with high efficiency using a non-viral system may form a first step towards tissue engineering with these transformed cells to repair articular cartilage defects. (C) 2001 OsteoAtthritis Research Society International.