Figure 4. Role of integrin α_vβ₈ in RORγt⁺ antigen presenting cell-dependent iTreg cell differentiation.

a, Frequency of iTreg cells among proliferating donor-derived Hh-specific cells in the MLN at 3 days after transfer of naïve CFSE-labeled Hh7–2 T cells into mice treated with 200μg of ADWA11 blocking antibody (n=8) or left untreated (n=8), on the day of adoptive transfer. Data summarize three independent experiments. b, Hh7–2 T cell proliferation and differentiation in the MLN of Itgav^ΔCD11c (n=5) and littermate controls (n=15) at 3 days after adoptive transfer. Data summarize three independent experiments. c, Proliferation and differentiation of Hh-specific iTreg cells in the MLN of Itgav^ΔRORγt (n=6) and littermate control mice (n=8). CFSE-labeled Hh7–2 T cells were analyzed at 3 days following their adoptive transfer into Hh-colonized mice. Representative flow cytometry profiles (left) and aggregate data (right). Data summarize three independent experiments. d, Transcription factor expression in Hh7–2 T cells (left panels) and in endogenous CD4⁺ T cells (right panels) from colon lamina propria (LP) at 10 days after adoptive transfer into Itgav^ΔRORγt mice (n=3) and control littermates (n=6). Data summarize two independent experiments. Representative dot plots and aggregate data are shown (right panels). e, Scheme for mixed bone marrow chimeric mouse experiment, with control, Itgav^ΔRORγt or MHCII^ΔCD11c cells administered to irradiated host mice (created with BioRender.com). f, Bar graphs showing iTreg frequency among Hh7–2 T cells in the colon LP at 10 days after their transfer into the bone marrow chimeric mice, reconstituted with different combinations of donor cells as indicated. Control mice (n=3), MHCII^ΔCD11c (n=3), MHCII^ΔCD11c and WT (n=4), Itgav^ΔRORγt and WT (n=4) and MHCII^ΔCD11c and Itgav^ΔRORγt (n=4). All statistics were calculated by unpaired two-sided Welch’s t-test. Error bars denote mean ± s.d. p-values are indicated in the figure.