Ma F, Manabe A, Wang D, Ito M, Kikuchi A, Wada M, Ito M, Ohara A, Hosoya R, Asano S, Tsuji K
Division of Cellular Therapy, The Advanced Clinical Research Center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
Leukemia. 2002 Aug;16(8):1541-8. doi: 10.1038/sj.leu.2402547.
The in vitro proliferation of T cell acute lymphoblastic leukemia (T-ALL) cells in its entirety has not been well delineated because of a lack of an appropriate culture system that mimics the growth pattern in a living body. We applied a NOD/SCID mouse fetal thymus organ culture (FTOC) for leukemic cells from fresh (one case) and frozen (seven cases) bone marrow (BM) samples of children with T-ALL. Cell growth was observed in all seven samples in the culture, reaching a proliferational peak at 4 weeks, and it was calculated that the proliferation potential was 212-to 319-fold. The FTOC-derived T-ALL cells showed similarity to the original cells morphologically and immunophenotypically, still possessed clonalities and were able to regenerate overt leukemia in NOD/SCID mice. These FTOC-derived T-ALL cells differed from ordinary cell lines because they always need FTOC support. Thus, we established a new in vitro culture for T-ALL cells. A comparison of the original and FTOC-derived T-ALL cells revealed that the proportion of cells expressing IL-7R increased in all seven cases. Sorting and re-seeding of FTOC-derived IL-7R+ and IL-7R- cells into secondary FTOC resulted in a predominant generation of IL-7R+ cells from both fractions, while IL-7R- cells proliferated more potently than did IL-7R+ cells, suggesting that a pathway for the conversion of IL-7R- to IL-7R+ exists during the proliferation of T-ALL lymphoblasts. Addition of exogenous IL-7 or neutralization with anti-IL-7 antibody did not influence the growth pattern of T-ALL cells in FTOC. The current study provides a unique assay system for the exploration of the hierarchy within human T-lymphoid leukemic cells, and should facilitate the establishment of novel therapeutic modalities.
由于缺乏模拟体内生长模式的合适培养系统，T细胞急性淋巴细胞白血病（T-ALL）细胞的整体体外增殖情况尚未得到很好的描述。我们将NOD/SCID小鼠胎儿胸腺器官培养（FTOC）应用于来自T-ALL患儿新鲜（1例）和冷冻（7例）骨髓（BM）样本的白血病细胞。在培养的所有7个样本中均观察到细胞生长，在4周时达到增殖峰值，计算得出增殖潜能为212至319倍。FTOC衍生的T-ALL细胞在形态和免疫表型上与原始细胞相似，仍具有克隆性，并且能够在NOD/SCID小鼠中再生明显的白血病。这些FTOC衍生的T-ALL细胞与普通细胞系不同，因为它们始终需要FTOC支持。因此，我们建立了一种新的T-ALL细胞体外培养方法。对原始T-ALL细胞和FTOC衍生的T-ALL细胞的比较显示，所有7例中表达IL-7R的细胞比例均增加。将FTOC衍生的IL-7R+和IL-7R-细胞分选并重新接种到二级FTOC中，导致两个部分均主要产生IL-7R+细胞，而IL-7R-细胞比IL-7R+细胞增殖更强劲，这表明在T-ALL淋巴母细胞增殖过程中存在IL-7R-向IL-7R+转化的途径。添加外源性IL-7或用抗IL-7抗体中和对FTOC中T-ALL细胞的生长模式没有影响。本研究为探索人类T淋巴细胞白血病细胞内的层次结构提供了独特的检测系统，并应有助于建立新的治疗方法。
