Figure 2.

Chitosan transforms tumor immune microenvironment and increases infiltration of neutrophils and monocytes. (A-L) Chitosan transforms tumor immune microenvironment in CT26 peritoneal tumor model. CT26 cells (2 × 10⁵) were transplanted into the peritoneal cavity of mice to establish CT26 peritoneal tumor model. Mice were intraperitoneally administrated with 0 or 100 mg/kg chitosan every three days and sacrificed on day 15 to collect the tumors. Then the single-cell suspension of tumors was prepared and subjected to FCM analysis. (A-B) Representative scatterplots of the gated neutrophils (CD45⁺ CD11b⁺ LY6G⁺) are shown in (A) and quantified in (B) (n = 4 mice). (C-D) Representative scatterplots of the gated monocytes (CD45⁺ CD11b⁺ LY6G^- LY6C⁺) are shown in (C) and quantified in (D) (n = 4 mice). (E-F) Representative scatterplots of the gated N2 neutrophils (NEU2, CD45⁺ CD11b⁺ LY6G⁺ Arg-1⁺) are shown in (E) and quantified in (F) (n = 4 mice). (G-H) Representative scatterplots of the gated N1 neutrophils (NEU1, CD45⁺ CD11b⁺ LY6G⁺ TNF-α⁺) are shown in (G) and quantified in (H) (n = 4 mice). (I-L) The percentages of M1 macrophages (M1 Mac, CD45⁺ CD11b⁺ F4/80⁺ MHCII⁺ CD206^-), M2 Macrophages (M2 Mac, CD45⁺ CD11b⁺ F4/80⁺ Arg-1⁺ CD206⁺), activated CD4⁺ T cells (CD3⁺ CD4⁺ CD69⁺), and activated CTLs (CD3⁺ CD8⁺ CD69⁺), are quantified. (M-P) Chitosan transforms tumor immune microenvironment in B16-F10 experimental pulmonary metastasis models. B16-F10 cells (2×10⁵) were intravenously injected into mice to establish experimental pulmonary metastasis models. Mice were administrated with 0 or 100 mg/kg chitosan and sacrificed on day 15 to collect the lungs. Then the single-cell suspension of lungs and metastatic tumors was prepared and subjected to FCM analysis. The percentages of neutrophils (CD45⁺ CD11b⁺ LY6G⁺), monocytes (CD45⁺ CD11b⁺ LY6G^- LY6C⁺), activated CTLs (CD3⁺ CD8⁺ CD69⁺), and M2 Macrophages (CD45⁺ CD11b⁺ F4/80⁺ CD206⁺) are quantified (n = 4 mice). Data are represented as mean±SD. Statistical significance in (B, D, F, H, I-P) was determined by a two-sided unpaired t-test. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.