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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- 1调控序列启动的造血生长因子基因在辐射后表达明显增高并促进造血祖细胞增殖作用
Objective To explore the regulation of radiation-induced genes regulating the gene expression of hematopoietic growth factor and observe its effect on hematopoiesis recovery. Methods FLFC (FL) and EGFR bicistronic gene expression vector (Egr-EF) with Egr-1 promoter were used to transfect the bone marrow stromal cell line HFCL (HFCL / EF) The mRNA expression of EGFP was detected by PCR. The expression of EGFP green fluorescent protein was observed by FACS. The content of FL in culture supernatant of HFCL / EF cells was detected by ELISA. The effect of HFCL / EF culture supernatant on the expression of CD34 + Proliferation effect. Results The integration and expression of exogenous EGFP and FL were confirmed in HFCL / EF cells. The supernatant of HFCL / EF cells after 16 h irradiation showed that the content of FL was significantly increased (P <0. At the same time, it was confirmed that the effect of HFCL / EF culture supernatant on CD34 + hematopoietic progenitor cells after 10 days of irradiation had a significant amplification compared with that before radiation (P <0.01). Conclusions The hematopoietic growth factor gene activated by Egr-1 regulatory sequence is significantly increased after radiation and promotes the proliferation of hematopoietic progenitor cells