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Antitumor Efficacy of Human Monocyte-Derived Dendritic Cells: Comparing Effects of two Monocyte Isolation Methods.
Biological Procedures Online ( IF 6.4 ) Pub Date : 2018-02-02 , DOI: 10.1186/s12575-018-0069-6
Graça S Marques 1 , Zélia Silva 1, 2 , Paula A Videira 1, 2, 3
Affiliation  

BACKGROUND Dendritic cells (DCs), which can be used as anti-cancer vaccines, are generally obtained in vitro from isolated CD14+ monocytes (MoDCs). This generates high cell numbers and allows instructing DCs to guarantee effective antitumor responses. However, the impact of the monocyte isolation step in the antitumor effectiveness of the generated MoDCs is still unknown. Here, we compared the most used immunomagnetic technologies for monocyte isolation: magnetic activated cell sorting (MACS) from Miltenyi Biotec and EasySep from STEM CELL. RESULTS MACS technology allowed a higher monocyte yield and purity and, by flow cytometry, monocytes displayed higher size and lower granularity. In the resting state, EasySep_MoDCs showed a higher basal expression of HLA-DR, and no significant response to stimulation by LPS and TNF-α. When stimulated with whole tumor cells lysates, both MoDCs expressed similar levels of maturation and co-stimulatory markers. However, when cultured with autologous T cells, MACS_MoDCs induced significantly higher IFN-γ secretion than EasySep_MoDCs, indicating a stronger induction of Th1 cell response profile. Concordantly, T cells induced by MACS_MoDCs also showed a higher release of cytotoxic granules when in contact with tumor cells. CONCLUSIONS Overall, both the MACS and the EasySep isolation immunomagnetic technologies provide monocytes that differentiate into viable and functional MoDCs. In our experimental settings, resting EasySep_MoDCs showed a higher basal level of maturation but show less responsivity to stimuli. On the other hand, MACS_MoDCs, when stimulated with tumor antigens, showed better ability to stimulate Th1 responses and to induce T cell cytotoxicity against tumor cells. Thus, monocyte isolation techniques crucially affect MoDCs' function and, therefore, should be carefully selected to obtain the desired functionality.

中文翻译:

人单核细胞衍生的树突状细胞的抗肿瘤功效:比较两种单核细胞分离方法的效果。

背景技术可用作抗癌疫苗的树突状细胞(DC)通常在体外从分离的CD14+单核细胞(MoDC)中获得。这会产生大量的细胞,并允许指导 DC 保证有效的抗肿瘤反应。然而,单核细胞分离步骤对生成的 MoDC 的抗肿瘤效果的影响仍然未知。在这里,我们比较了最常用的单核细胞分离免疫磁性技术:美天旎生物技术公司的磁激活细胞分选 (MACS) 和 STEM CELL 公司的 EasySep。结果 MACS 技术允许更高的单核细胞产量和纯度,并且通过流式细胞术,单核细胞显示出更高的尺寸和更低的粒度。在静息状态下,EasySep_MoDCs 表现出较高的 HLA-DR 基础表达,并且对 LPS 和 TNF-α 的刺激没有显着反应。当用全肿瘤细胞裂解物刺激时,两种 MoDC 都表达了相似水平的成熟和共刺激标志物。然而,当与自体 T 细胞一起培养时,MACS_MoDCs 诱导的 IFN-γ 分泌显着高于 EasySep_MoDCs,表明对 Th1 细胞反应谱的诱导更强。一致地,由 MACS_MoDCs 诱导的 T 细胞在与肿瘤细胞接触时也显示出更高的细胞毒性颗粒释放。结论 总体而言,MACS 和 EasySep 分离免疫磁性技术都提供了分化为可行和功能性 MoDC 的单核细胞。在我们的实验设置中,静息的 EasySep_MoDCs 表现出更高的基础成熟水平,但对刺激的反应性较低。另一方面,MACS_MoDCs 在用肿瘤抗原刺激时,显示出更好的刺激Th1反应和诱导T细胞对肿瘤细胞的细胞毒性的能力。因此,单核细胞分离技术对 MoDC 的功能产生至关重要的影响,因此,应仔细选择以获得所需的功能。
更新日期:2019-11-01
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