The regeneration of hyaline articular cartilage by cell-mediated gene therapy using transforming growth factor beta(1) (TGF-beta(1))-producing fibroblasts (NIH 3T3-TGF-beta(1)) has been reported previously. In this study, we investigated whether TGF-beta(1)-producing fibroblasts irradiated with a lethal dose of radiation are still capable of inducing the regeneration of hyaline articular cartilage. NIH 3T3TGF-beta(1) fibroblasts were exposed to doses of 20, 40, or 80 Gy, using a irradiator, and then injected into artificially made partial defects on the femoral condyle of rabbit knee joints. The rabbits were killed 3 or 6 weeks postinjection and hyaline articular cartilage regeneration was evaluated by histological and immunohistochemical staining (n = 5 per each group). Irradiated NIH 3T3-TGFbeta(1) fibroblasts started to die rapidly 3 days after irradiation; moreover, the kinetics of their viability were similar regardless of the radiation intensity. TGF-beta1 expression, measured by ELISA, showed that the TGF-beta(1) protein produced from the irradiated cells peaked 5 days after irradiation and thereafter declined rapidly. Complete filling of the defect with reparative tissue occurred in all the groups, although variations were observed in terms of the nature of the repair tissue. Histological and immunohistochemical staining of the repair tissue showed that the tissue newly formed by irradiated NIH 3T3-TGF-beta(1) fibroblasts after exposure to 20 Gy had hyaline cartilage-like characteristics, as was observed in the nonirradiated controls. On the other hand, the repair tissue formed by NIH 3T3-TGF-beta(1) fibroblasts irradiated with 40 or 80 Gy showed more fibrous cartilage-like tissue. These results suggest that TGF-beta(1)-producing fibroblasts irradiated up to a certain level of lethal dose (i.e., 20 Gy) are able to induce normal-appearing articular cartilage in vivo. Therefore, irradiated heterologous cell-mediated TGF-beta(1) gene therapy may be clinically useful and an efficient method of regenerating hyaline articular cartilage.