Genetic enhancement of matrix synthesis by articular chondrocytes: Comparison of different growth factor genes in the presence and absence of interleukin-1

Abstract

Objective. To determine whether articular chon-drocytes express growth factor genes delivered by ad-enoviral vectors and whether expression of these genes influences matrix synthesis in the presence and absence of interleukin-1 (IL-1). Methods. Monolayer cultures of rabbit articular chondrocytes were infected with recombinant adenovi-rus carrying genes encoding the following growth factors: insulin-like growth factor 1 (IGF-1), transforming growth factor 1 (TGF1), and bone morphogenetic protein 2 (BMP-2). As a control, cells were transduced with the lac Z gene. Cultures were also treated with each growth factor supplied as a protein. Levels of gene expression were noted, and the synthesis of proteogly-can, collagen, and noncollagenous proteins was measured by radiolabeling. Collagen was typed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. The effects of growth factor gene transfer on proteoglycan synthesis in the presence of IL-1 were also measured. Results. The expression of all transgenes was high following adenoviral transduction. Proteoglycan synthesis was stimulated 8-fold by the BMP-2 gene and 2-3-fold by the IGF-1 gene. The effects of BMP-2 and IGF-1 genes were additive upon cotransduction. Synthesis of collagen and noncollagenous proteins, in contrast, was most strongly stimulated by the IGF-1 gene. In each case, collagen typing confirmed the synthesis of type II collagen. IL-1 suppressed proteoglycan synthesis by 50-60%. IGF-1 and TGF genes restored proteoglycan synthesis to control levels in the presence of IL-1. The BMP-2 gene, in contrast, elevated proteoglycan synthesis beyond control levels in the presence of IL-1. Conclusion. Transfer of growth factor genes to articular chondrocytes can greatly increase matrix synthesis in vitro, even in the presence of the inflammatory cytokine IL-1. This result encourages the further development of gene therapy for the repair of damaged cartilage.

Age-related Changes in the Response of Human Articular Cartilage to IL-1α and Transforming Growth Factor-β (TGF-β)

“…Furthermore, this is supported by studies in which decreased levels of PG synthesis could be restored using adenoviral delivery of TGF-␤1 to monolayer cultures of rabbit articular chondrocytes (37). Although our studies do not provide a direct mechanistic link between the effects of TGF-␤1 on UDPGD activity and GAG production, they indicate that the actions of IL-1␣ on GAG synthesis do not appear to involve modification of chondrocyte UDPGD activity, whereas the response to TGF-␤1 is closely correlated with changes in UDPGD activity.…”

Benefits of Recombinant Adeno-Associated Virus (rAAV)-Mediated Insulinlike Growth Factor I (IGF-I) Overexpression for the Long-Term Reconstruction of Human Osteoarthritic Cartilage by Modulation of the IGF-I Axis

“…Even though application of these stimuli was capable of containing cartilage degradation, it was not sufficient to compensate for the loss of matrix elements and cells and to reestablish an original cartilage surface. Growth, transcription and enzymatic factors are potent candidates to achieve these goals because of their anabolic or mitogenic properties such as fibroblast growth factor 2 (FGF-2) (20,21), bone morphogenetic proteins 2 and 4 (BMP-2 and BMP-4) (22,23), transforming growth factor β (TGF-β) (22,24,25), transcription factor sex-determining region Y-type high mobility group box 9 (SOX9) (20,26,27), glucuronosyltransferase-I (28), bcl-2 (29) and telomerase (30). A critical agent that may have the strongest value to readjust the disturbed homeostasis in OA cartilage is insulinlike growth factor (IGF)-I, since it has the ability to influence concomitantly metabolic and proliferative processes, affording protection against extracellular matrix degradation in horse and rabbit articular cartilage explant cultures experimentally treated with proinflammatory cytokines (13,14,22).…”

“…In contrast with vectors derived from adenoviruses (6,10,13,14,(16)(17)(18)(19)21,22,25) and retroviruses (7,11,12,23,26,30) or with nonviral compounds (8,15,28,29), systems based on the replication-defective, nonpathogenic human adeno-associated virus (AAV) may provide better tools for OA, since recombinant AAV (rAAV) can deliver genes in nondividing cells such as chondrocytes both in vitro and in situ in their dense extracellular matrix at high efficiencies and for extended periods of time (20,24,27,31). Also, removal of the viral protein coding sequences in rAAV make them less immunogenic than adenoviral vectors characterized by shortterm transgene expression levels (32).…”

Gene-based approaches for the repair of articular cartilage

“…For example, transfer of cDNAs encoding certain bioactive factors can be used to stimulate anabolic pathways. Delivery by recombinant adenovirus of the cDNAs for TGF-b, 56,57 IGF-1, 11,57 BMP-2 57 or BMP-7 58 to monolayer cultures of chondrocytes isolated from experimental animals has been shown to stimulate expression of cartilage matrix genes, resulting in increased synthesis of proteoglycan and collagen type II. TGF-b1, IGF-1 and BMP-2 overexpression was also found to rescue proteoglycan synthesis following pretreatment of the chondrocytes with IL-1, a potent inhibitor of matrix synthesis.…”

“…TGF-b1, IGF-1 and BMP-2 overexpression was also found to rescue proteoglycan synthesis following pretreatment of the chondrocytes with IL-1, a potent inhibitor of matrix synthesis. 57 The delivery of genes whose products enable maintenance of the phenotype of hyaline cartilage without hypertrophy of the chondrocytes would be highly desirable. In this regard, IGF-1 gene transfer was also found to render chondrocyte cultures resistant to dedifferentiation in monolayer, enabling maintenance of the chondrocytic phenotype for at least 28 days in culture.…”