Han Tae Hee, Jin Ping, Ren Jiaqiang, Slezak Stefanie, Marincola Francesco M, Stroncek David F
Department of Transfusion Medicine, Clinical Center, National Institutes of Health, 10 Center Drive, Bethesda, MD 20892, USA.
J Immunother. 2009 May;32(4):399-407. doi: 10.1097/CJI.0b013e31819e1773.
Dendritic cells (DCs) are important adjuvants for cancer vaccines. Immature dendritic cells (iDCs) are often produced by the stimulation of peripheral blood monocytes with interleukin (IL)-4 and granulocyte macrophage-colony stimulating factor. For many applications iDCs are treated with cytokines or inflammatory signals to produce mature DCs (mDCs). iDCs are often treated ex vivo with lipopolysaccharide (LPS) and interferon (IFN)-gamma to produce mDCs for clinical therapy. The purpose of this study was to determine if the DC maturation cocktail LPS plus IFN-gamma could be improved by the addition of 2 other DC maturation agents IL-1beta and tumor necrosis factor (TNF)-alpha. Peripheral blood mononuclear cells were collected from 6 healthy subjects. Monocytes were isolated from the peripheral blood mononuclear cell concentrates by elutriation and were incubated for 3 days with granulocyte macrophage-colony stimulating factor and IL-4 to produce iDCs. iDCs from each subject were divided into 3 and were incubated for 24 hours with LPS plus IFN-gamma; LPS, IFN-gamma, plus IL-1beta; or LPS, IFN-gamma, IL-1beta, plus TNF-alpha to produce mDCs. The DCs were compared by measuring the expression of costimulator and antigen presenting molecules (CD80, CD83, CD86, and human leukocyte antigen-DR) by flow cytometry, cytokine production (IL-12p70 and IL-10) by enzyme-linked immunosorbent assay and global gene expression using an oligonucleotide microarray. There were no differences in the expression of costimulatory molecules, human leukocyte antigen-DR and CCR7 and production of IL-12p70 among the mDCs produced with the 3 cocktails. Global gene expression analysis found that the expression of 9576 genes differed between the iDCs and mDCs, but the expression of only 13 differed among the 3 different groups of mDCs. There was no benefit of adding IL-1beta and TNF-alpha to LPS and IFN-gamma to produce mDCs.
树突状细胞（DCs）是癌症疫苗的重要佐剂。未成熟树突状细胞（iDCs）通常通过用白细胞介素（IL）-4和粒细胞巨噬细胞集落刺激因子刺激外周血单核细胞产生。对于许多应用，iDCs用细胞因子或炎症信号处理以产生成熟树突状细胞（mDCs）。iDCs通常在体外用脂多糖（LPS）和干扰素（IFN）-γ处理以产生用于临床治疗的mDCs。本研究的目的是确定是否可以通过添加另外2种DC成熟剂IL-1β和肿瘤坏死因子（TNF）-α来改进DC成熟混合物LPS加IFN-γ。从6名健康受试者收集外周血单核细胞。通过淘析从外周血单核细胞浓缩物中分离单核细胞，并与粒细胞巨噬细胞集落刺激因子和IL-4一起孵育3天以产生iDCs。将来自每个受试者的iDCs分成3组，并与LPS加IFN-γ；LPS、IFN-γ加IL-1β；或LPS、IFN-γ、IL-1β加TNF-α一起孵育24小时以产生mDCs。通过流式细胞术测量共刺激分子和抗原呈递分子（CD80、CD83、CD86和人白细胞抗原-DR）的表达、通过酶联免疫吸附测定法测量细胞因子产生（IL-12p７0和IL-10）以及使用寡核苷酸微阵列进行全局基因表达，对DCs进行比较。在用3种混合物产生的mDCs中，共刺激分子、人白细胞抗原-DR和CCR7的表达以及IL-12p70的产生没有差异。全局基因表达分析发现，iDCs和mDCs之间9576个基因的表达不同，但3组不同的mDCs之间只有13个基因的表达不同。向LPS和IFN-γ中添加IL-1β和TNF-α来产生mDCs没有益处。
