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. 2022 May 14;12(10):3952–3971. doi: 10.1016/j.apsb.2022.05.006

Figure 7.

Figure 7

MΦ(VNP-PD1nb) cells accelerate tumor cell necrosis and activate TAMs and CD8+ T cells. (A, B) Detection of alive cells in the tumor on Days 1,3 and 5 by flow cytometry in (A) (n = 4 mice per group), and flow cytometry plots of representative data are shown in (B). (C, D) Comparison of the tumor necrosis proportion in different treatment groups by H&E staining on Day 3 in (C), and representative H&E images are shown in (D). Scale bars, 1 mm (complete tumor sections) and 200 μm (enlarged tumor sections). (E) The proliferation of tumor cells (CD45-) was quantified by Ki67 staining and flow cytometry. (F, G) Changes in the tumor-infiltrating macrophage population on Days 1, 3 and 5 in (F) and phenotype (M1-like and M2-like) at Day 3 in (G) after treatment with PBS, PEMΦ(VNP-NC) or PEMΦ(VNP-PD1nb) cells in the B16F10 tumor model (n = 4 mice per group). (H) Representative immunofluorescence staining of B16F10 tumor cross sections showing cell nuclei (DAPI, blue), CD206 (marker of M2-like macrophages, red) and iNOS (marker of M1-like macrophages, green) of (G) (Scale bar = 50 μm). (I, J) Profile of tumor-infiltrating CD8+ T cell and cytokine-producing CD8+ T cell subsets on Day 3 in the B16F10 tumor model (n = 4 mice per group) (I), and representative flow cytometry plots show the method of acquisition of frequency values for each T cell subset (J). (K, L) Detection of IFNγ and TNFα in peripheral blood 3 days after treatment in the B16F10 tumor model by CBA assay in (K), and flow cytometry plots of representative CBA data are shown in (L). Boxplot representations of the spot counts, with the median, interquartile range, and minimum and maximum identifiers, are shown. All data are representative of two independent experiments. Statistics were calculated using the two-tailed, unpaired Student's t test with Welch's correction.