The generation of human natural killer cells from CD34+/DR- primitive progenitors in long-term bone marrow culture.
The generation of human natural killer cells from CD34+/DR- primitive progenitors in long-term bone marrow culture.
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DOI：
10.1182/blood.v80.9.2182.bloodjournal8092182
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发表时间：
1992-11
期刊：
影响因子：
20.3
通讯作者：
Jeffrey S. Miller;C. Verfaillie;P. Mcglave
中科院分区：
文献类型：
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作者：
Jeffrey S. Miller;C. Verfaillie;P. Mcglave
我们采用基质依赖型长期骨髓培养(LTBMC)系统来研究CD34+/HLA-DR-(CD34+/DR-)骨髓单个核细胞(BMMNC)群体中人自然杀伤细胞(NK)的发育。CD34+/DR-群体不表达任何已知的与髓系或淋巴系相关的抗原，我们和其他人已经证明CD34+/DR-群体含有原始造血祖细胞，具有自我更新和向髓系分化的能力。用荧光激活细胞分选法从正常人骨髓中获得CD34+/DR-细胞，并将其接种在照射后的同种异体骨髓基质层上。在含有重组白介素2和人血清的培养液中培养5周后，细胞扩增147+/-21倍，形态为大颗粒淋巴细胞。培养细胞(84.8%+/-1.5%)表达NK细胞特有的CD56+/CD3-表型。部分CD56+/CD3-细胞表达与成熟NK相关的其他淋巴系标志，包括CD2(7.8%+/-1.2%)、CD7(19.5%+/-2.8%)、CD8(3.1%+/-1.0%)和CD16(4.5%+/-1.3%)。培养的细胞不表达其他与T淋巴细胞(CD3、CD5、T细胞受体[TCR]α/β和TCRγ/Delta)、B淋巴细胞(CD19)、髓系(MY8、CD33和CD71)或单核细胞(CD14和CD15)相关的抗原，也不表达起始群体中与造血祖细胞相关的CD34抗原。该NK细胞对K562(E：T20：1；79%+/-1.9%)和Raji(E：T20：1；38%+/-5.7%)靶细胞均有杀伤作用。CD34+/DR-细胞的NK祖细胞频率为1：169+/-50 CD34+/DR-细胞。这些数据表明，用于研究髓系分化的人LTBMC可以被修改以研究NK以及可能的其他淋巴系的起源和发展。改良培养表明，具有NK形态、表型和功能特征的细胞可以从具有原始造血祖细胞表型且没有表型证据的淋巴系或髓系系的BMMNC群体中分化出来。进一步的研究将涉及人类NK和其他淋巴系的细胞起源和个体发育。
We have adapted the stroma-dependent long-term bone marrow culture (LTBMC) system to study the development of human natural killer cells (NK) from the CD34+/HLA-DR- (CD34+/DR-) BM mononuclear cell (BMMNC) population. The CD34+/DR- population does not express any known antigens associated with myeloid or lymphoid lineage and has been shown by us and others to contain primitive hematopoietic progenitors capable of both self-renewal and differentiation to myeloid lineage. CD34+/DR- cells obtained from normal human BM by fluorescence-activated cell sorting were plated on allogeneic, irradiated BM stromal layers. After 5 weeks of culture in the presence of media containing recombinant interleukin-2 and human serum, 147- +/- 21-fold expansion of cells with the morphologic appearance of large granular lymphocytes was observed. Cultured cells (84.8% +/- 1.5%) expressed the characteristic CD56+/CD3- phenotype of NK. A proportion of CD56+/CD3- cells expressed other markers of lymphoid lineage that have been associated with mature NK, including CD2 (7.8% +/- 1.2%), CD7 (19.5% +/- 2.8), CD8 (3.1% +/- 1.0%), and CD16 (4.5% +/- 1.3%). The cultured cells did not express other antigens associated with T-lymphocyte (CD3, CD5, T-cell receptor [TCR] alpha/beta and TCR gamma/delta), B-lymphocyte (CD19), myeloid (MY8, CD33, and CD71), or monocytoid (CD14 and CD15) lineage and did not express the CD34 antigen associated with hematopoietic progenitors present on the starting population. This NK population was cytotoxic against both K562 (E:T 20:1; 79% +/- 1.9%) and Raji (E:T 20:1; 38% +/- 5.7%) target cell lines. The NK progenitor frequency in the CD34+/DR- cell population determined by limiting dilution of CD34+DR- on stromal layers followed by a functional chromium release assay against K562 targets was 1:169 +/- 50 CD34+/DR- cells. The data suggest that human LTBMC developed to study myeloid differentiation can be modified to study the origin and development of the NK and possibly other lymphoid lineages. Modified cultures show that cells with morphologic, phenotypic, and functional characteristics of NK can be derived from a population of BMMNC with the phenotype of primitive hematopoietic progenitors and without phenotypic evidence of lymphoid- or myeloid-lineage commitment. Further studies will address the cell of origin and the ontogeny of human NK and other lymphoid lineages.