Fig. 1

Characteristics of the cytotoxic dynamics of primary NK cells against five different epithelial target cell lines. a Cumulative survival curves of target cells, including LO2 (denoted in black), HeLa (blue), U-2 OS (green), SMMC-7721 (magenta), and MCF7 (red), in co-culture with primary human NK cells. Time 0 is when NK cells were added to the target cells and the imaging experiment was started. b Left panel: fluorescent images of the granzyme B FRET (GrzmB-FRET) reporter and the corresponding mitochondria reporter, IMS-RP, from SMMC-7721, U-2 OS, and MCF7, respectively. The GrzmB-FRET images are an overlay of the CFP (denoted by blue) and YFP (green) channels. Time (unit: hours:minutes) is indicated at the top left corner of each GrzmB-FRET image. The white scale bar is 20 μm, and the white arrows point to the specific target cells, for which we quantified the CFP and YFP signals shown in the right panels. Right panels: single-cell trajectories of CFP and YFP signals quantified from the time-lapse movies. The time of MOMP (scored by the IMS-RP signal change) and the time of death (scored morphologically by cell blebbing and lysis) are indicated by the vertical dotted line. c Distributions of the live and dead target cells killed by the three distinct cytotoxic modes mediated by granzyme B (GrzmB active), death ligand (GrzmB inactive), and necrosis after 12 h of co-culture with primary NK cells. Data were averaged from 3 independent imaging experiments, and each experiment used NK cells from a different healthy donor (this applied to all other experimental repeats). Data acquired with primary NK cells from the same donor are denoted with the same color symbols. The number of cells analyzed ranges from 40 to 223, varied between experiments and target cell lines. The error bars are standard deviations