脐带血中长期培养启动细胞在转染FL和/或TPO基因骨髓基质细胞层上的生长  

Maintaining Growth of Long-Term Culture Initiating Cells from Human Cord Blood on Feeder Layers of Bone Marrow Stromal Cells Transfected with FL and/or TPO Genes

作　　者：张毅[1] 毛宁[1] 李秀森[1] 金滢[1] 张双喜[1] 吴英[1] 唐佩弦[1] 

机构地区：[1]军事医学科学院基础医学研究所,北京100850

出　　处：《中国实验血液学杂志》2001年第2期97-100,共4页Journal of Experimental Hematology

基　　金：国家 8 6 3高科技技术重点课题基金(BH 0 30 5 0 1);全军95重点课题基金资助~~

摘　　要：检测长期培养启动细胞 (LTC IC)含量可反映造血干细胞的存在和变化。用正常人骨髓基质细胞、人骨髓基质细胞系和转基因骨髓基质细胞系做培养底层 ,接种CD34+ 细胞 ,进行连续 8周的长期体外培养。研究结果表明 ,在共转染FL和TPO基因的基质细胞系HFT为培养底层所产生的LTC IC为 3.6 5± 0 .5 8 10 0 0CD34+ 细胞 ,在原代骨髓基质细胞层上所产生的LTC IC为2 .6 5± 0 .76 10 0 0CD34+ 细胞 ,二者在统计学上无显著性意义。结论 :转染FL TPO基因的HFT细胞系可作为LTCLong tem culture initiating cells(LTC IC), and in vitro assay of hematopoietic stem/progenitor cells, still represent a heterogeneous population in terms of proliferative capacity and sensitivity to different growth factors. Human umbilical cord(CB) is rich of hematopoietic progenitor cells measured by clonogenic assays and stem cells capable of reconstituting the marrow after transplantation. The influence of culture conditions on the in vitro behavior of LTC IC from CB was evaluated. First, by using IRES sequence, FL and TPO cDNA were recombined with retroviral vector pLXSN by gene recombination technology. The recombinant plasmid pLFSN, pLTSN, pLFTSN were transfected into human stromal cell line HFCL. Then, LTC IC were evaluated in long term cultures, comparing five types of stromal feeder layers: human bone marrow stromal cell, human stromal cell line HFCL, and stromal cell lines HDF tranfected with FL gene, HLT transfected with TPO gene or HFT co transfected with FL and TPO genes. The results were demonstrated that after 8 weeks of coculture, three types of stromal cell lines that supported the maintenance of CFU C for up to 3 weeks in vitro were identified. However, cocultivation of human bone marrow stromal cell and CB CD34 + cells on HFT, CFU C production continued up to 6 weeks or longer on these stroma. The absolute LTC IC frequency in CD34 + cells on human bone marrow stromal cell (2.65±0.76/1 000 cells) was no significant difference with on HFT(3.65±0.58/ 1 000 cells). Thus, HFT acts by direct contact to maintain the phenotype and function of the most primitive and quiescent human progenitors. Furthermore, HFT cell line was selected as the optimal one for supporting long term culture feeder. It was concluded that LTC IC progenitors from cord blood maintain growing upon the FL/TPO gene modified stromal feeder layers in vitro.

关 键 词：脐带血 长期培养启动细胞 骨髓基质细胞 Flt-3配体 血小板生成素基因转染 

分 类 号：R331.1[医药卫生—人体生理学]