Fig. 4. Heat-iMVA promotes OT-I cell activation and proliferation mediated by OVA cross-presentation by dendritic cells in vitro.

a–d Proliferation of CTV-labeled OT-Ι T cells after incubation with GM-CSF-cultured BMDCs (a, b) or FLT3L-cultured dendritic cells (c, d) pulsed with OVA in the presence or absence of MVA or heat-iMVA. BMDCs were incubated with or without MVA or heat-iMVA, and then co-cultured with CTV-labeled OT-Ι cells for 3 days. e IFN-γ secretion from OT-Ι T cells after incubation with GM-CSF-cultured WT or STING^Gt/Gt BMDCs pulsed with OVA in the presence or absence of live MVA or heat-iMVA. f IFN-γ secretion from OT-Ι T cells after incubation with sorted CD103⁺ DCs from WT or STING^Gt/Gt Flt3L-cultured BMDCs pulsed with OVA in the presence or absence of live MVA or heat-iMVA. g A heat map of a one-way hierarchical clustering analysis of the differentially expressed genes between WT or STING^Gt/Gt BMDCs treated with Heat-iMVA over time (2, 4, and 6 h). h Gene set enrichment analyses (GSEA) showing differences of gene expression in several pathways including IFN-γ, IFN-α, inflammatory responses, and IL-6_JAK_STAT3 signaling in WT and STING^Gt/Gt BMDCs infected with heat-iMVA. Data are represented as mean ± SEM (n = 3-5; ^**P < 0.01 and ^***P < 0.001; unpaired multiple t test). Data are representative of three independent experiments.