Control of delivered gene expression in chondrocytes using heat shock protein 70B promoter

Objective. To investigate whether the expressions of delivered Escherichia coli β-galactosidase (LacZ) gene and transforming growth factor- β1 (TGF-β1) gene are regulated by the stress response of human chondrocyte- like cells (HCS-2/8) when heat shock protein 70B (HSP70B) promoter is inserted into the adenovirus vector. Methods. Two adenovirus vectors that contain either LacZ gene or TGF-β1 gene regulated by HSP70B promoter were constructed. One of the adenovirus vectors was added to the culture of HCS2/8 and gene transduced cells were produced. We applied heat stress (43°C) to the transduced cells for 2 h and examined whether the expression of transduced LacZ and TGF-β1 genes is affected by the stress, using 5 bromo-4- chloro-3-indolyl-β-D-galactopyranoside (X-gal) staining, measurement of β- galactosidase activity, Northern blotting, and ELISA. Results. The percentage of X-gal positive stained cells in LacZ gene-delivered cells with heat stress was significantly higher than in controls (no heat stress). With heat stress, β-galactosidase activity increased significantly, and the band of exogenous TGF-β1 mRNA became more apparent and the expression was maintained during the 24 h monitoring period. TGF-β1 level in culture supernatant of TGF-β1 gene-delivered cells with heat stress (5477.3 ± 321.1 pg/ml) was significantly higher than in the controls (853.2 ±29.2 pg/ml). Conclusion. HSP70B promoter could regulate the expression of delivered genes according to the intensity of heat stress.