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Fig. 1 | Journal for ImmunoTherapy of Cancer

Fig. 1

From: Tumor cell-released autophagosomes (TRAPs) promote immunosuppression through induction of M2-like macrophages with increased expression of PD-L1

Fig. 1

TRAPs polarize macrophages toward an M2-like phenotype in vitro and in vivo. a Confocal images of BMDMs treated with CFSE-labeled TRAPs (green). After incubation with TRAPs (10 μg/ml) for 0.5 h, BMDMs were stained with PE-F4/80 antibody (red) and DAPI (blue). Scale bar, 10 μm. b Expression analysis of CD206, PD-L1, CD86 and MHC II by flow cytometry. BMDMs were stimulated with LPS (100 ng/ml) + IFN-γ (20 ng/ml), IL-4 (20 ng/ml) or TRAPs (10 μg/ml) for 48, 48 or 72 h, respectively. c Flow cytometry analysis of PD-L2, B7-H2, B7-H3, B7-H4, Tim-4 and VISTA for BMDMs after incubating with TRAPs for 48 h. d Expression analysis of NOS2 and Arg1 mRNA in BMDMs treated with TRAPs (10 μg/ml) for 6 h by qRT-PCR. e ELISA detection of IL-1β, IL-6, IL-10 and IL-12p70 produced by BMDMs exposed to TRAPs (10 μg/ml) for 72 h. f-h Mice (n = 3) were intraperitoneally injected with four different doses of TRAPs (0, 10, 30 and 100 μg) at day 0 and the phenotype of peritoneal macrophages was analyzed at day 3. Expression of CD206 (f) and PD-L1 (g) on macrophages (F4/80+CD11b+) was determined by flow cytometry. h mRNA expression level of IL-10 and Arg1 in purified macrophages was detected by qRT-PCR. Results are representative of three independent experiments. *p < 0.05, **p < 0.01 and ***p < 0.001 by unpaired t test (d, f, g and h)

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