转hGM-CSF基因骨髓基质细胞在体外扩散盒中的表达

Expression of the bone marrow stromal cell line transfected by hGM-CSF in the diffusion chamber in vitro

目的 研究转人粒 巨噬细胞集落刺激因子 (hGM CSF)基因骨髓基质细胞系在体外扩散盒中的表达。方法 采用阳离子脂质体介导法将构建好的真核表达载体pIRESIneo hGM CSF导入人骨髓基质细胞系HFCL ,建立转基因人骨髓基质细胞系HFCL hGM CSF ,用Southernblot和Northernblot方法分别检测其基因组DNA整合和mRNA转录 ,并将其移入体外扩散盒中 ,用ELISA法检测其蛋白表达量。结果 Southernblot和Northernblot分析表明hGM CSF基因已整合到HFCL细胞基因组DNA中 ,在mRNA水平上可以检测到其表达 ,移入扩散盒的HFCL hGM CSF细胞稳定分泌hGM CSF ,量为 5 1 6± 0 5ng 10 6 细胞 天。结论 转hGM CSF基因骨髓基质细胞可以在体外扩散盒中持续稳定表达hGM CSF 。

Objective To study the expression of an bone marrow stromal cell line transfected by hGM-CSF in the diffusion chamber in vitro. Methods The bone marrow stromal cell line transfected with hGM-CSF was constructed by using liposome-mediated gene transfer method and transferred into the diffusion chamber.Integration of hGM-CSF in the genome, transcription of hGM-CSF mRNA and expression of hGM-CSF protein in the transfected HFCL cells in the diffusion chamber were assayed by Southern blot, Northern blot and ELISA. Results It showed that hGM-CSF cDNA was integrated into HFCL genome successfully,hGM-CSF mRNA was transcripted and hGM-CSF protein was expressed with(51 6±0 5)ng.10 6 cell -1 d -1 in the supernatant. Conclusion The transgene HFCL cells express hGM-CSF continuously in the diffusion chamber.