摘要
目的应用小鼠骨髓微血管内皮细胞能够在体外大量扩增骨髓造血细胞。本文旨在进一步研究（１）内皮细胞分泌的造血细胞因子；（２）优化培养条件，提高骨髓细胞的扩增指数；（３）扩增细胞的长期造血重建功能。方法采用ＲＴ┐ＰＣＲ和生物活性测定技术分析内皮细胞表达的造血细胞因子；小鼠骨髓细胞经与内皮细胞共培养方法进行扩增，扩增的骨髓细胞移植给大剂量化疗后的异基因小鼠，然后检测外周血中供体细胞的比例。结果（１）内皮细胞能持续地、或经肿瘤坏死因子（ＴＮＦα）刺激后表达干细胞因子（ＳＣＦ）、血小板生成因子（ＴＰＯ）和白介素┐６（ＩＬ┐６）；（２）将Ｃ５７小鼠骨髓细胞置于ＴＮＦα刺激后的内皮细胞层上４小时，粘附的骨髓细胞在０．２５μｇ／ｍｌ重组人粒细胞集落刺激因子（ｒｈＧ┐ＣＳＦ）存在下培养１周，细胞数量增加了６０倍，其中ＣＤ３４＋细胞和ｃ┐ｋｉｔ＋细胞比例分别为５８．５％和６３．９％；（３）将扩增的Ｃ５７小鼠骨髓细胞移植给大剂量ＶＩＰ方案化疗（顺铂，１０ｍｇ／ｋｇ，腹腔注射，第一天；异环磷酰胺，５００ｍｇ／ｋｇ，腹腔注射，第２天；依托帕甙，５ｍｇ／ｋｇ，腹腔注射，第１～２天）后的ＢＡＬＢ／ｃ小鼠，８周时外周血供体细胞比例达３５％。?
bjective:Mouse bone marrow hematopoietic cells can be expanded ex vivo on the monolayers of bone marrow microvascular endothelial cells. This study investigated (1) the hematopoietic cytokines expressed by endothelial cells;(2)optimizing cultural conditions to increase the marrow cells; and (3) the longterm hematopoietic reconstitution of expanded marrow cells.Mehtods:The hematopoietic cytokines expressed by endothelial cells were evaluated by reverse transcription PCR(RTPCR) and biological assay. Mouse bone marrow cells were expanded ex vivo on the monolayers of endothelial cells, and were transplanted into allogeneic BALB/c mice after highdose chemotherapy; the donor cells were tested in the peripheral blood of recipient mice.Results:(1)The endothelial cells can express spontaneously or upon stimulation with TNFα the genes encoding for stem cell factor (SCF), thrombopoietin (TPO),and interleukin6(IL6)(2)After adherence to the TNFαstimulated endothelial monolayers the number of bone marrow cells was increased by 60fold within one week in the presence of recombinant human GCSF, among which 58.5% and 63.9% were CD34+and ckit+cells respectively.(3)After transplantation of expanded marrow cells into BALB/c mice which were treated with highdose VIP regimen (cisplatin, ifosfamide, etoposide), eight weeks later 35% of the peripheral blood nucleated cells of recipient mice were donor origin.Conclusion:Bone marrow microvascular endothelial cells can stimulate the proliferation of hematopoietic cells by producing several hematopoietic cytokines. The expansive index of hematopoietic cells can be increased markedly by pretreating endothelial cells with TNFα, depleting nonadherent marrow cells and increasing GCSF concentration. The expanded marrow cells, which contain approximately 60 % of CD34+ and ckit+ cells, have the capacity of longterm hematopoietic reconstitution after transplantation.
出处
《肿瘤》
CAS
CSCD
北大核心
1998年第2期66-70,共5页
Tumor
关键词
骨髓
造血细胞
体外扩增
移植
Hematopoietic cells Cell expansion ex vivo Transplantation Mouse
