辐射诱导性启动子调控GM—CSF基因的表达

构建携带早期生长反应基因（Egr-1)启动子的GM-CSF和增强绿色荧光蛋白（EGFP）双顺反子基因表达载体（Egr-EG)。通过脂质体转染骨髓基质细胞系HFCL（称为HFCL/EG）,采用FACS分析EGFP表达的阳性细胞，用RT-PCER、ELISA及集落增殖法分析GM-SCFmRNA、GM-CSF含量的表达及对CFU-GM抚养殖作用。结果显示：HFCL/EG细胞证实有外源性基因EGFP和GM-CSF的整合和表达及对CFU-GM的增殖作用。提示Egr-1调控序列启动的GM-CSF基因修饰的基质细胞经辐射造血因子表达明显增高，并对造血细胞具有一定的保护作用。

STUDIES ON THE EXPRESSION OF GM-CSF REGULATED BY RADIO INDUCIBLE PROMOTER

The human GM CSF cDNAandEGFP cDNA were linked together with IRES and then inserted into the expression vector pCI Egr 1, which was constructed by substituting CMV promoter in pCIneo with the Egr 1 promoter(Egr EG).The vector was transfered into human bone marrow stromal cell line HFCL by Lipofectin TM . The results indicated that the activity of EGFP in transfected cells increased at 18h after exposure to 2.5 Gy. The amounts of secreted GM CSF and CFU GM in serum free supernatants of Egr EG was significantly higher than the control group. EGFP and GM CSF cDNA were successfully integrated and expressed in the cells, which were confirmed by FACS and RT PCR analysis respectively. These in vitro data provide an experimental basis for the in vivo use of gene therapy of GM CSF gene regulated by Egr 1 promoter to protect hematopoiesis from total body irradiation.