Figure 6.

SpNK and ThNK donor cells show cytotoxicity against myeloid and mesenchymal cell targets in vitro

(A) Schematic of the effector-to-target cell cytotoxicity experiment. Effector NK cells were harvested and co-cultured with the target cell lines for 4 h (n = 3).

(B) RMA-S cells (gold-standard positive control cells sensitive to NK lysis) or Maf-DKO myeloid target cells were cultured with effector NK cells harvested from the spleen and thymus in a series of E:T ratios. RMA-S and Maf-DKO cells showed high cytotoxicity from SpNK and ThNK cells, proportional to NK cell concentration.

(C) Cytotoxicity at an E:T/ratio of 10:1. RMA-S and Maf-DKO cells show significantly higher cytotoxicity in the presence of ThNK cells compared with SpNK cells.

(D) Schematic of autologous PDGFRα⁺ cell versus NK cell (ThNK or SpNK cell) isolation and cytotoxicity assays in mutant and control mice. Fluorescence-activated cell sorting (FACS) of mesenchymal PDGFRα⁺ cells from Prf1 KO, Ncr1^gfp, Klrk1⁻, and B6 after P3 injury (9 DPA) was performed. SpNK or ThNK effector cells were isolated and matched with the original PDGFRα⁺ target (autologous) cell source.

(E) NK cell cytotoxicity toward PDGFRα⁺ cells after co-culture.

(F) Prf1 and Klrk1 KO donors (SpNK and ThNK cells) have significantly reduced cytotoxicity against PDGFRα⁺ target cells at an E:T ratio of 4:1. Ncr1 KO cytotoxicity remains similar to B following treatment with NK cells. Statistics were performed using Student’s t test (^∗p < 0.05) for two groups and one-way ANOVA and Tukey’s test ±SEM for three or more groups; n = 5. ^∗∗p < 0.01, ^∗∗∗∗p < 0.0001).