Figure 3.

Effect of metformin on tumors of the mouse colon cancer model and immunohistochemistry analysis of MDSCs and M1/M2 macrophages. (A) In vivo tumorigenesis model experimental schedule, using Apc^Min/+ treated with 3% DSS, and metformin treatment schedule. (B) After spraying methylene blue, the number and size of polyps in the colon were measured. Data are expressed as the means ± standard errors; ** p < 0.005, *** p < 0.001 (n = 3 per group). (C) Representative H&E staining of the rectum of mice (scale bar = 50 μm). (D) Immunohistochemistry (IHC) of the colon sections was performed on paraffin-embedded sections using MDSC (CD11b) and M2 macrophage (CD206) markers (scale bar = 50 μm). CD11b- and CD206-stained cells were counted in ten different fields under 200× magnification. Data are expressed as the means ± standard errors; * p < 0.05, ** p < 0.005 (n = 3 per group). (E) Mice colon tissues were homogenized and examined by real-time qPCR for CD11b (ITGAM) and CD206 (Mrc1), and normalized to β-actin expression. Data are expressed as the means ± standard errors of the three different experiments; ** p < 0.005, *** p < 0.001. n = 3 per group.