摘要
目的探索辐射诱导基因调控序列启动造血生长因子基因表达及观察其对造血恢复的作用。方法本实验将带有 Egr- 1调控序列启动的 FL T3配基 ( FL )和 EGFP双顺反子基因表达载体 ( Egr- EF)转染骨髓基质细胞系 HFCL (称 HFCL /EF) ;用 RT— PCR鉴定细胞内目的基因的 m RNA表达 ;采用 FACS观察 EGFP绿色荧光表达的阳性细胞 ;用 EL ISA方法检测 HFCL / EF细胞培养上清 FL的含量 ;观察 HFCL / EF培养上清液对 CD34+细胞的增殖作用。结果在 HFCL / EF细胞中证实有外源性基因 EGFP和 FL 的整合和表达 ,在辐照 16 h后的 HFCL/ EF细胞培养上清液中表明 FL 含量较照射前明显增高( P<0 .0 1) ;同时证实辐射 10 d后 HFCL/ EF培养上清液对 CD34+造血祖细胞的作用较辐射前具有明显的扩增作用 ( P<0 .0 1)。结论 Egr-
Objective To explore the regulating effects of radiation inducible gene on the expression of hematopoiedtic growth factor genes Methods The human FLT3 Ligand (FL) cDNA and EGFP cDNA were linked together with IRES and then inserted into the eukaryotic expression vector pCIEgr, which was constructed by substituting CMV promoter in pCIneo with the Egr1 promoter(EgrEF) The expression of FL in HFCL/EF cells were confirmed with RTPCR、ELISA、FACS and cell culture Results The activity of EGFP in transfected cells increased after exposure to 25Gy The amounts of secreted FL in serumfree supernatants of HFCL/EF were significantly higher than the control group FL cDNA was successfully expressed in the cells by ELISA and RTPCR analysis At day 10 of culture the number of CD34+ cells in HFCL/EF culture supernatants was significantly higher than that of nonradiation group ConclusionThese results showed that radiation can enhance the ability of the supernatants containing FL of HFCL/EF to expand early hematopoietic progenitor cells and protect hematopoietic cells from radiationinjury effects [
出处
《免疫学杂志》
CAS
CSCD
北大核心
2000年第3期166-171,共6页
Immunological Journal
基金
This work is supported by Natural Science Foundation of China(39900040) and Natural Science Outstan- ding Youth Foundation of C
关键词
辐射诱导基因
造血生长因子
基因治疗
hematopoietic growth factor
radiation inducible gene
gene therapy
radiation
bone marrow stromal cell
