Establishing long-term cultures with self-renewing acute myeloid leukemia stem/progenitor cells
Establishing long-term cultures with self-renewing acute myeloid leukemia stem/progenitor cells
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DOI：
10.1016/j.exphem.2007.07.001
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发表时间：
2007-10-01
影响因子：
2.6
通讯作者：
Schuringa, Jan Jacob
中科院分区：
文献类型：
--
作者：
van Gosliga, Djoke;Schepers, Hein;Schuringa, Jan Jacob
目标。随着白血病干细胞概念的出现，对它们的研究仍然是理解（白血病）干细胞生物学的关键。我们在骨髓基质上培养了急性髓系白血病CD34（+）细胞。监测长期扩张，并通过复制白血病鹅卵石区域形成细胞（L-CAs）来解决自我更新。此外，还生成了针对l - cas的慢病毒载体。在大约75%的急性髓系白血病病例（n = 30）中观察到强烈的扩增，MS5骨髓基质细胞的长期培养可维持长达24周。能够启动白血病鹅卵石区的细胞存在于CD34（+）群体中，而不存在于CD34（-）群体中。这些L-CAs内部的自我更新是通过这些L-CAs连续传代到新的MS5间质层来确定的，这导致了第二、第三和第四个L-CAs的产生，这些L-CAs能够维持长期扩增并产生大量未成熟的未分化悬浮细胞。能够启动长期培养的CD34（+）细胞均共表达MEIS1和HOXA9，并表达升高的BMI1水平。我们提出了一种新的长期白血病干细胞/祖细胞测定方法，其中可以测试新药，并且可以使用慢病毒方法过度表达或下调基因，以便更深入地了解白血病转化和自我更新的过程。(c) 2007年iseh血液学和干细胞学会。Elsevier Inc.出版。
Objective. With the emergence of the concept of the leukemia stem cell, assays to study them remain pivotal in understanding (leukemic) stem cell biology.Methods. We have cultured acute myeloid leukemia CD34(+) cells on bone marrow stroma. Long-term expansion was monitored and self-renewal was addressed by replating of Leukemic-cobblestone area-forming cells (L-CAs). Also, lentiviral vectors were generated that could target L-CAs.Results. A strong expansion was observed in about 75% of the acute myeloid leukemia cases (n = 30) and long-term cultures could be maintained for up to 24 weeks on MS5 bone marrow stromal cells. Cells that were able to initiate leukemic cobblestone areas resided in the CD34(+) population and were absent from the CD34(-) population. Self-renewal within these L-CAs was determined by sequential passaging of these L-CAs onto new MS5 stromal layers, which resulted in the generation of second, third, and fourth L-CAs, which were able to sustain long-term expansion and generated high numbers of immature undifferentiated suspension cells. CD34(+) cells that were able to initiate long-term cultures all coexpressed MEIS1 and HOXA9, and expressed elevated BMI1 levels.Conclusion. We present a novel long-term leukemic stem/progenitor assay in which new drugs can be tested and in which genes can be overexpressed or downmodulated using a lentiviral approach in order to obtain more insight into the process of leukemic transformation and self-renewal. (c) 2007 ISEH-Society for Hematology and Stem Cells. Published by Elsevier Inc.