Effect of adenovirus-mediated TGF-β1 gene transfer on the function of rabbit articular chondrocytes.

BACKGROUND

Articular chondrocytes are important in maintaining normal cartilage tissue and preventing articular degeneration. Exogenous genes have previously been transduced into articular cells using adenoviral vectors to contribute to the maintenance of cell function. This study aimed to transfer the transforming growth factor-β1 gene (TGF-β1) into rabbit articular chondrocytes by adenovirus infection to elucidate its effects on cell function.

METHODS

Rabbit chondrocytes were isolated and cultured both as monolayers and three-dimensional culture systems. To achieve overexpression, TGF-β1 was transfected by adenovirus infection, using the LacZ gene as a control. TGF-β1 protein expression was analyzed by western blotting. Quantitative DNA fluorometric analysis evaluated cell proliferation, and quantitative reverse transcriptase PCR determined the mRNA expression of related chondrocyte marker genes. Western blotting and glycosaminoglycan quantitative testing were used to examine changes in extracellular matrix components.

RESULTS

TGF-β1 protein expression was found to increase in Adv-TGF-β1-transduced cells, reaching a maximum after chondrocytes had been cultured for 4 weeks. Adv-hTGF-β1 transduction altered chondrocyte morphology from fibrocyte-like long spindle-shaped to round or oval. TGF-β1-transduced cells showed an increase in DNA synthesis, glycosaminoglycan content, and increased aggrecan and collagen II protein expression, while collagen I was significantly decreased. Moreover, TGF-β1 overexpression significantly promoted the mRNA expression of the chondrogenic gene SOX9, and inhibited that of the hypertrophic marker COL10A1 and the mineralization marker MMP-13.

CONCLUSIONS

TGF-β1 overexpression positively improved the phenotype, function, and proliferation of chondrocytes, even after several generations.