IL-4 does not affect ILC2 expansion rate, but decreases the percentage of IFN-γ–producing ILC2 cells and improves GVHD outcome. Cultured ILC2 cells were used in a xenograft GVHD model. Sublethally irradiated NSG mice received human PBMCs, with 1 group receiving donor unmatched cultured ILC2. (A) Diagram depicting PBMC isolation from donor 1, and isolation and expansion of ILC2 from donor 2, and the xenograft transplant model. (B) Kaplan-Meier plot of NSG recipient survival following transplant, 2 combined representative experiments using different donors is shown (n = 6 per experiment PBMC group and n = 4 per experiment ILC2 treated), log rank (Mantel-Cox) test. (C) Enriched human ILC2 cells were cultured in α-MEM media as described earlier with or without the addition of 50 ng/mL IL-4 and anti-IL-12 antibody for 10 days. Representative contour plots of day 10 ILC2 cultures showing IL-13 and IFN-γ expression. (D) Diagram depicting PBMC isolation from donor 1, and enrichment of human type 2 innate lymphoid cell (HuILC2) from donor 2, and expansion with or without IL-4. (E) Diagram depicting xenotransplant using the cells described in panel D. (F) Kaplan-Meier plot of NSG recipient survival following transplant, representative experiment using different donors and IL-4 treated or untreated ILC2 is shown (n = 4 per experiment), log rank (Mantel-Cox) test. (G) Clinical score of recipients from panel D, analyzed by 2-way analysis of variance, with Bonferroni correction for repeated measures of multiple comparisons between groups, ∗P < .05. BMT, bone marrow transplantation; ctrl, control; w/o, without; w/, with.