Fig. 3

RT plus YM101 induced DCs maturation and activation. (a-c) CD14⁺ monocytes were cultured for 6 days with GM-CSF and IL-4. ImDCs were irradiated with 0 Gy, 2 Gy, 5 Gy, 10–15 Gy and cultured for 24 h. ImDCs were stimulated with LPS (1 µg/mL) as a positive control and cultured for 24 h. Representative flow cytometry histograms of CD80, CD86, and CD40 expression on DCs (n = 5 independent experiments). (d-e) ImDCs were irradiated with 0 Gy, 2 Gy, 5–10 Gy and cultured for 24 h. ImDCs were stimulated with LPS (1 µg/mL) as a positive control and cultured for 24 h. The levels of TNF-α and IL-6 were measured by ELISA (n = 5 independent experiments). (f) ImDCs were irradiated with 0–10 Gy and cultured for 24 h. ImDCs were stimulated with LPS (1 µg/mL) as a positive control and cultured for 24 h. Chemokines secretion was analyzed by ELISA (n = 3 independent experiments). Representative data in a-f were repeated at least three times with similar results. (g-i) In the 4T1 model, mice were sacrificed 5 days post RT (for DC maturation analysis) (n = 5 mice/group). In the FACS assays, quantification of total DCs (g), CD80⁺ DCs (h) and CD86⁺ DCs (i). The P values were determined by one-way ANOVA with Dunnett’s multiple comparisons test. (j-k) Representative IF staining images of CD11c (green) and CD86 (red). Scale bars represent 50 μm. All experiments were repeated 2 or 3 times yielding similar results. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001