Fig. 3

Mucosal associated invariant T (MAIT) cells from human breast ducts produce a more robust interleukin (IL)-17A response than blood MAIT cells. a Isolated breast duct cells or peripheral blood mononuclear cell (PBMC) samples were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin, then stained for expression of the indicated cytokines. Boxes show positive cytokine staining based on parallel analysis of unstimulated samples. Percentages shown within the gated areas represent the percentage of the total MAIT cells expressing cytokine (top boxes) or of the total non-MAIT cells (bottom boxes), respectively. b Top graph: Quantification of the fraction of MAIT cells from PBMCs (dark circles) vs. breast ducts (light circles) that stained positively for the indicated cytokines after PMA/ionomycin stimulation. Bottom graph: Fold increase in cytokine signal intensity. The geometric mean fluorescence intensity (gMFI) of the total MAIT cell population for the indicated cytokines was normalized by that of unstimulated MAIT cells. Each symbol represents the result from a different tissue donor; PBMC and breast tissue samples were not obtained from the same donors. c Top graph: Quantification of the fraction of MAIT cells (light circles) vs. non-MAIT cells (dark circles) from the same breast tissue sample that stained positively for the indicated cytokines after PMA/ionomycin stimulation. Bottom graph: Fold increase in cytokine signal intensity (gMFI of the total MAIT cell population normalized by that of unstimulated MAIT cells). Each symbol represents the result from a different breast tissue donor. d The indicated in vitro-expanded MAIT cell lines were stimulated with PMA and ionomycin, and expression of tumor necrosis factor (TNF)-α, interferon (IFN)-γ, and IL-17A was assessed by intracellular cytokine staining. The graph shows the percentage of MAIT cells in each line showing positive staining for the indicated cytokines (see also Additional file 4: Figure S3). n.s. Not significant; 5RU 5-(2-oxoprophylideneamino)-6-d-ribitylaminouracil