Arai Y, Kubo T, Kobayashi K, Takeshita K, Takahashi K, Ikeda T, Imanishi J, Takigawa M, Hirasawa Y
Department of Orthopaedic Surgery, Kyoto Prefectural University, Japan.
J Rheumatol. 1997 Sep;24(9):1787-95.
To investigate the effects of adenovirus vector-mediated gene transduction of E. coli beta-galactosidase (LacZ), transforming growth factor-beta 1 (TGF-beta 1), and heat shock protein 70 (HSP 70) on human chondrocyte-like cell line (HCS-2/8).
We examined expression of transduced genes and their expression periods by 5 bromo-4-chloroindolyl-beta-D-galactoside (X-gal) staining. Northern blotting, ELISA, and Western blotting. To assess the influence of TGF-beta 1 and HSP70 gene transduction, the expression of mRNA of type II collagen, proteoglycan core protein, matrix metalloproteinase 3 (MMP3) and tissue inhibitor of matrix metalloproteinase 1 were examined by Northern blotting.
Staining with X-gal indicated that the genes were transduced into a majority of the cells. Expression of the transduced genes in the cells was continued for at least 21 days. Transduction of TGF-beta 1 gene enhanced mRNA expression of type II collagen and proteoglycan core protein, but suppressed MMP3 mRNA expression in the cells. Expression of HSP70 was also high. Enhanced expression of HSP70 elevated mRNA expression of proteoglycan core protein.
These results indicate adenovirus vector is useful in chondrocyte gene therapy, and it could be an efficient mediator of TGF-beta 1 and HSP70 gene transduction.
研究腺病毒载体介导的大肠杆菌β-半乳糖苷酶（LacZ）、转化生长因子-β1（TGF-β1）和热休克蛋白70（HSP 70）基因转导对人软骨样细胞系（HCS-2/8）的影响。
我们通过5-溴-4-氯吲哚-β-D-半乳糖苷（X-gal）染色、Northern印迹法、酶联免疫吸附测定（ELISA）和蛋白质印迹法检测转导基因的表达及其表达时期。为评估TGF-β1和HSP70基因转导的影响，通过Northern印迹法检测II型胶原、蛋白聚糖核心蛋白、基质金属蛋白酶3（MMP3）和基质金属蛋白酶组织抑制剂1的mRNA表达。
X-gal染色表明基因被转导到大多数细胞中。转导基因在细胞中的表达持续至少21天。TGF-β1基因的转导增强了II型胶原和蛋白聚糖核心蛋白的mRNA表达，但抑制了细胞中MMP3 mRNA的表达。HSP70的表达也很高。HSP70表达的增强提高了蛋白聚糖核心蛋白的mRNA表达。
这些结果表明腺病毒载体在软骨细胞基因治疗中有用，并且它可能是TGF-β1和HSP70基因转导的有效介质。
