Activation of monocytes via the CD14 receptor leads to the enhanced lentiviral transduction of immature dendritic cells
Activation of monocytes via the CD14 receptor leads to the enhanced lentiviral transduction of immature dendritic cells
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DOI：
10.1089/104303404323142015
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发表时间：
2004-06-01
影响因子：
4.2
通讯作者：
Thielemans, K
中科院分区：
文献类型：
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作者：
Breckpot, K;Corthals, J;Thielemans, K
在这项研究中，我们比较了树突状细胞（DC）分化的阳性选择的单核细胞（CD 14-DC）的DC分化的粘附选择的单核细胞（adh-DC），重点是慢病毒转导。使用第二代、含三螺旋的自失活慢病毒载体，在感染复数（MOI）为15时，我们观察到CD 14-DCs的转导增强（72.8 +/-5.3%，平均荧光强度[MFI] = 166 +/-76）与adh-DC（32.3 +/-13.1%，MFI = 119 +/-76，n = 5）相比。更重要的是，当单核细胞与抗CD 14抗体偶联的珠粒、抗CD 14抗体或脂多糖（LPS）孵育时，对CD 3adh-DCs的效率显著增加，分别达到高达86.6%、53.3%和80.9%的转导效率。我们发现这种增强的转导与单核细胞的活化相关，其特征在于细胞因子白细胞介素（IL）-1 β和肿瘤坏死因子（TNF）-α的上调以及IL-6和IL-10的从头合成。然而，未成熟CD 14-DCs的增强转导与细胞周期从G（0）到G（1）的进展无关。我们进一步表明，CD 14-DC在表型上与adh-DC相当。功能分析显示，通过逆转录酶-聚合酶链反应（RT-PCR）评估，同种异体刺激能力、CD 40连接上IL-12 p70的产生或IL-1 β、IL-6、IL-8、IL-10、IL-12和TNF-α的表达无差异。最后，我们发现慢病毒转导的CD 14-DCs在体外刺激MAGE-A3抗原特异性CD 4（+）和CD 8（+）T细胞方面与adh-DCs具有相同的能力。
In this study, we compared dendritic cells (DCs) differentiated from positively selected monocytes (CD14-DCs) to DCs differentiated from adherence-selected monocytes (adh-DCs) with emphasis on lentiviral transduction. Using a second-generation, triple-helix containing, self-inactivating lentiviral vector at a multiplicity of infection (MOI) of 15, we observed enhanced transduction of CD14-DCs (72.8 +/- 5.3%, mean fluorescence intensity [MFI] = 166 +/- 76) compared to adh-DCs (32.3 +/- 13.1%, MFI = 119 +/- 76, n = 5). More importantly, the efficiency to transduce adh-DCs was significantly increased when monocytes were incubated with anti-CD14 antibody coupled beads, anti-CD14 antibodies, or lipopolysaccharide (LPS), reaching transduction efficiencies up to 86.6%, 53.3%, and 80.9%, respectively. We showed that this enhanced transduction was correlated to an activation of the monocytes, characterized by the up regulation of the cytokines interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha and the de novo synthesis of IL-6 and IL-10. However, the enhanced transduction of immature CD14-DCs was not correlated with a progression in the cell cycle from G(0) to G(1). We further showed that CD14-DCs were phenotypically comparable to adh-DCs. Functional analysis revealed that there were no differences in allostimulatory capacity, production of IL-12 p70 on CD40 ligation or expression of IL-1beta, IL-6, IL-8, IL-10, IL-12, and TNF-alpha as evaluated by reverse transcriptase-polymerase chain reaction (RT-PCR). Finally, we showed that lentivirally transduced CD14-DCs were equally capable as adh-DCs in stimulating MAGE-A3 antigen-specific CD4(+) and CD8(+) T cells in vitro.