天然杀伤细胞（NK细胞）是先天免疫系统的淋巴细胞，在肿瘤细胞和病毒感染细胞消除中起关键作用。与T细胞不同，NK细胞的活化受其通过细胞膜上的抑制性和激活性受体与靶细胞直接相互作用的调控。这种活化机制的简单性使得可以开发基于将NK细胞转导成CAR（嵌合抗原受体）构建物的免疫治疗，用于癌症治疗。尽管与CAR-T相比，CAR-NK疗法具有不会在异基因治疗中引发移植物抗宿主病的优点，但需要对其处理的影响进行更深入的了解，以提高其功能性和适用性。鉴此，本研究对NK细胞的分离、扩增和储存所需的步骤进行了批判性检查，并分析了NK细胞对这些操纵的响应。结果显示，传统用于NK分离的磁辅助细胞分选只有在方案中包括抗体孵育和通过分离柱的通行时，才会增加NK培养物中的CD16+细胞群。此外，基于表面电位对细胞反应的重要性，对不同净表面电荷的表面对NK细胞的影响进行了评估，结果显示相对于无电荷表面，带电荷的表面上NK细胞显示出更高的增殖率。本研究突出了NK细胞操纵对改善NK细胞治疗的应用性和有效性的重要性。版权©2023 Martin-Iglesias, Herrera, Santos, Vesga, Eguizabal, Lanceros-Mendez and Silvan.

Natural killer (NK) cells are lymphocytes of the innate immune system that play a key role in the elimination of tumor and virus-infected cells. Unlike T cells, NK cell activation is governed by their direct interaction with target cells via the inhibitory and activating receptors present on their cytoplasmic membrane. The simplicity of this activation mechanism has allowed the development of immunotherapies based on the transduction of NK cells with CAR (chimeric antigen receptor) constructs for the treatment of cancer. Despite the advantages of CAR-NK therapy over CAR-T, including their inability to cause graft-versus-host disease in allogenic therapies, a deeper understanding of the impact of their handling is needed in order to increase their functionality and applicability. With that in mind, the present work critically examines the steps required for NK cell isolation, expansion and storage, and analyze the response of the NK cells to these manipulations. The results show that magnetic-assisted cell sorting, traditionally used for NK isolation, increases the CD16+ population of NK cultures only if the protocol includes both, antibody incubation and passage through the isolation column. Furthermore, based on the importance of surface potential on cellular responses, the influence of surfaces with different net surface charge on NK cells has been evaluated, showing that NK cells displayed higher proliferation rates on charged surfaces than on non-charged ones. The present work highlights the relevance of NK cells manipulation for improving the applicability and effectiveness of NK cell-based therapies.Copyright © 2023 Martin-Iglesias, Herrera, Santos, Vesga, Eguizabal, Lanceros-Mendez and Silvan.