Effect of AAT on NK cells and antitumor activity. (A) Treatment scheme: injection of A20 tumor cells (A20 luc tumor) and transplantation; C57BL/6 (H-2^bK^b) donors, Balb/c (H-2^d K^d) recipients (major mismatch). (B) Either donors (D) or recipients (R) or both were treated with AAT (controls received albumin), resulting in four donor/recipient combinations: transplantation of cells from AAT-treated donors into recipients treated with albumin (AAT/albumin; red); transplantation of cells from albumin-treated donors into AAT-treated recipients (albumin/AAT; black); transplantation of cells from AAT-treated donors into AAT-treated recipients (AAT/AAT; green), and transplantation of cells from albumin-treated donors into albumin-treated recipients (albumin/albumin; purple). Photon reading of luciferase activity; higher readings indicate greater tumor volume. (C) Transplantation with either donors (D) or recipients (R) given AAT while the other partner received albumin: AAT-treated donors (D:AAT; red) into albumin-treated recipients (R:albumin), and albumin-treated donors (D:albumin) into AAT-treated recipients (R:AAT; black). Tumors were imaged 3, 7, and 14 days after donor cell transplantation (ie, 7, 11, and 18 days after tumor cell injection). Tumor presence and size were determined using the luciferase reporter signal. Tumor progression was more delayed with transplantation of cells from AAT-treated donors than with direct recipient treatment. (n = 12, each condition, results presented as the mean of transplantations). (D) Survival of Balb/c (H-2d Kd) recipients of C57BL/6 [H-2b] donors after treatment of donors and recipients with the various AAT/albumin combinations (P values displayed in the figure). (E) Proportions of spleen-derived NK cells, determined by flow cytometry (FACS) using several immunophenotypic markers, in C57BL/6 donor mice treated with albumin (left, control) or AAT (right), respectively; NK cells were increased in AAT-treated mice (right panel) (horizontal axis, CD49B; vertical axis, NKp46). (F) Cytolytic function of NK cells obtained from the spleens of AAT (AAT) or albumin treated (control) mice (n = 7, each condition), and assayed at effector/target ratios of 1:1; 5:1, 10:1, and 50:1 against A20 cells. Lytic activity (percent of CFSE⁺, PI⁺ cells) was measured after 18 hours of incubation of the effector cells. Shown are the mean ± SEM of 5 experiments.