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  • Title: Control of delivered gene expression in chondrocytes using heat shock protein 70B promoter.
    Author: Arai Y, Kubo T, Kobayashi K, Ikeda T, Takahashi K, Takigawa M, Imanishi J, Hirasawa Y.
    Journal: J Rheumatol; 1999 Aug; 26(8):1769-74. PubMed ID: 10451076.
    Abstract:
    OBJECTIVE: To investigate whether the expressions of delivered Escherichia coli beta-galactosidase (LacZ) gene and transforming growth factor-beta1 (TGF-beta1) 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-beta1 gene regulated by HSP70B promoter were constructed. One of the adenovirus vectors was added to the culture of HCS-2/8 and gene transduced cells were produced. We applied heat stress (43 degrees C) to the transduced cells for 2 h and examined whether the expression of transduced LacZ and TGF-beta1 genes is affected by the stress, using 5 bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal) staining, measurement of beta-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, beta-galactosidase activity increased significantly, and the band of exogenous TGF-beta1 mRNA became more apparent and the expression was maintained during the 24 h monitoring period. TGF-beta1 level in culture supernatant of TGF-beta1 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.
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