Burgess A W, Metcalf D
Int J Cancer. 1980 Nov 15;26(5):647-54. doi: 10.1002/ijc.2910260517.
Culture of WEHI-3B myelomonocytic leukemic cells in semi-solid agar medium containing serum from mice injected with endotoxin serum (ES) led to the development of maturing granulocytes and macrophages in most leukemic colonies. ES contains high levels of granulocyte-macrophage colony-stimulating factor (GM-CSF), a regulator known to stimulate differentiation of these leukemic cells, but an antiserum which neutralized greater than 85% of the GM-CSF in ES did not suppress the differentiation-inducing activity of ES on WEHI-3B cells. The active factor in endotoxin serum stimulating differentiation in WEHI-3B leukemic cells (GM-DF) was separated from most of the GM-CSF by gel filtration using Ultrogel AcA44. The residual CSF associated with the GM-DF appeared to stimulate selectively granulocytic colonies. Disproportionation of GM-DF and GM-CSF was observed in ES fractions obtained using concanavalin-A/Sepharose chromatography: none of the GM-DF bound to this matrix, whereas 40% of the GM-CSF bound and was eluted with competing alpha methylglucopyranoside. Although no separation of GM-CSF and GM-DF was obtained using DEAE-Sepharose, non-isoelectric focusing in amphoteric buffers indicated charge differences between the differentiation factor and several sub-species of GM-CSF. Sequential purification of GM-DF from ES using 40 - 70% ammonium sulfate precipitation gel filtration and phenyl-Sepharose chromatography resulted in a 25-fold purification. In all fractionation procedures used, a sub-species of GM-CSF, stimulating granulocyte colony formation, was consistently associated with partially purified GM-DF, but some subspecies of GM-CSF clearly lacked any capacity to induce differentiation in the leukemic cells. The observations suggest that the factor in post-endotoxin serum most efficient in enforcing differentiation in myelomonocytic leukemic cells may be a subset of GM-CSF molecules with a selective capacity to stimulate granulocyte colony formation by normal cells.
在内毒素血清（ES）注射小鼠的血清存在下，将WEHI-3B骨髓单核细胞白血病细胞培养于半固体琼脂培养基中，结果大多数白血病集落中出现了成熟的粒细胞和巨噬细胞。ES含有高水平的粒细胞巨噬细胞集落刺激因子（GM-CSF），这是一种已知可刺激这些白血病细胞分化的调节因子，但一种能中和ES中超过85%的GM-CSF的抗血清并未抑制ES对WEHI-3B细胞的分化诱导活性。使用Ultrogel AcA44通过凝胶过滤将刺激WEHI-3B白血病细胞分化的内毒素血清中的活性因子（GM-DF）与大部分GM-CSF分离。与GM-DF相关的残留CSF似乎选择性地刺激粒细胞集落。在用伴刀豆球蛋白A/琼脂糖凝胶层析获得的ES组分中观察到GM-DF和GM-CSF的比例失调：GM-DF均不与该基质结合，而40%的GM-CSF结合并被竞争性α-甲基吡喃葡萄糖苷洗脱。尽管使用DEAE-琼脂糖凝胶未实现GM-CSF和GM-DF的分离，但在两性缓冲液中的非等电聚焦表明分化因子与GM-CSF的几个亚类之间存在电荷差异。使用40%-70%硫酸铵沉淀、凝胶过滤和苯基琼脂糖凝胶层析从ES中依次纯化GM-DF，纯化倍数达25倍。在所有使用的分级分离程序中，刺激粒细胞集落形成的GM-CSF亚类始终与部分纯化的GM-DF相关，但GM-CSF的一些亚类显然缺乏诱导白血病细胞分化的能力。这些观察结果表明，内毒素血症后血清中在促进骨髓单核细胞白血病细胞分化方面最有效的因子可能是GM-CSF分子的一个亚群，其具有选择性刺激正常细胞形成粒细胞集落的能力。
