Evaluating migration and cytotoxicity of tissue-resident and conventional NK cells in a 3D microphysiological system using live-cell imaging.

Natural killer (NK) cells are critical components of the immune response against cancer, recognized for their ability to target and eliminate malignant cells. Among NK cell subsets, intraepithelial ILC1 (ieILC1)-like tissue resident NK (trNK) cells exhibit distinct functional properties and enhanced cytotoxicity compared to conventional NK (cNK) cells, positioning them as promising candidates for cancer immunotherapy. However, the specific roles and mechanisms of these cytotoxic trNK cells within the tumor microenvironment (TME) remain to be further explored. In this study, we utilized a three-dimensional (3D) microphysiological system (MPS) to model the tumor-vascular interface and investigate the distinct capabilities of cytotoxic ieILC1-like trNK and cNK cells within the TME. Through the integration of live-cell imaging and cell-tracking analysis, we quantitatively assessed NK cell migration, tumor infiltration, and cytotoxic activity in real time. Our findings revealed that trNK cells demonstrate enhanced motility, sustained tumor interactions, and superior tumor-killing efficiency compared to cNK cells. This study highlights the unique properties of trNK cells, providing a robust foundation for developing next-generation cancer therapies that harness their potent cytotoxic capabilities.