Fig. 5: The exacerbated memory response is dependent on absence of both VEGF-C and VEGF-D.

a Schedule of HDM-induced allergic airway inflammation in mice including treatment of all mice with tamoxifen more than 2 weeks prior to inducing allergic inflammation. Allergic inflammation was induced in wild type (‘WT’) mice and mice lacking either VEGF-C (Rosa26^Cre-ERT2;Vegfc^flox/flox, here referred to as “VC KO”), VEGF-D (Vegfd^−/−, here referred to as “VD KO”), or both (Rosa26^Cre-ERT2;Vegfc^flox/flox;Vegfd^−/−, here referred to as “double KO”) that have been previously described.^33,34 b H&E staining showing inflammation (scale bar indicates 500 µm), and c blinded inflammation scores. Flow cytometric analysis of immune cell infiltration into the lungs including d eosinophils, e CD4+ T_EM/T_CM ratios, f T_Reg cells, and CD4+ Th2 cells, and g Th2/T_Reg ratios. h–i IL-5⁺, IL-13⁺, IFNγ⁺, and IL5⁺13⁺ double producing CD4+ T cells after in vitro restimulation with PMA and ionomycin on lung single cell suspensions quantified via flow cytometry. j IL-4, IL-5, and IL-13 levels after 48 h in vitro restimulation with HDM on lung single cell suspensions to assess antigen specificity of the Th2 cells in the lungs. Boxes represent median (central bar) with range from 25th to 75th percentile, and whiskers represent min to max value. Data are representative of n = 4 mice for PBS treated and n = 6 mice for inflamed mice (n = 1 experiment for mice lacking VC/VD), and statistics (one-way ANOVA with Dunnett’s multiple comparison mice lacking VEGF-C and/or VEGF-D). Tests were performed in GraphPad Prism. *p < 0.05, **p ≤ 0.01.