Fig 2: MiR-520a-3p silencing inhibited macrophage polarization to M1 subset. (A) RT-PCR analysis showed the mRNA levels of macrophage polarization related markers TNF-α; (B) IL-6; (C) MCP-1; (D) IL-1β; (E) INOS; (F) CD206; and (G) Arg1 in plaque and artery tissue in apoE-/- mice. Data are mean ± SEM; one-way ANOVA was used for the statistical analysis. n=5 mice per group; (H) Western blot analysis showed increased CD206 and decreased INOS after miR-520a-3p inhibition in plaque and artery tissue in apoE-/- mice. Data are mean ± SEM; one-way ANOVA was used for the statistical analysis. n=5 mice per group. **P<0.01, ***P<0.001.

Fig 3: Analysis of the relationship between risk score and immune cell subtypes. (A) Proportion of immune cells in each glioma sample are indicated with different colors, and lengths of the bars in the bar chart indicate the levels of the immune cell proportions. (B) Comparison of the proportion of 22 immune cell types in high and low risk groups. (C) Correlation risk score and the proportion of 22 immune cell types. (D) Survival curves of four immune cell types significantly related to the prognosis of patients with glioma. (E) Comparison of the scoring of SPON2, IFI44, CD68 and CD206 expression in low-and high-grade gliomas tissues. (F) Correlation analysis between the scoring of SPON2, IFI44, CD68 and CD206 expression. (G) Representative immunohistochemistry of low- and high-grade gliomas tissues with the scoring of SPON2, IFI44, CD68 and CD206 expression. Abbreviations: ns, P > 0.05, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Fig 4: UVRAG mediated the inhibitory effect of miR-520a-3p on macrophage autophagy and polarization. (A) and; (B) RT-PCR analysis showed the mRNA level of UVRAG in macrophages. Data are mean ± SEM; one-way ANOVA was used for the statistical analysis. n=5 independent cell cultures; (C) Luciferase assay indicated the interaction of miR-520a-3p with 3’ UTR of UVRAG. Data are mean ± SEM; Two-tailed t test was used for the statistical analysis. n=5 independent cell cultures; (D) Western blot analysis showed the protein levels of p62, LC3 and UVRAG in macrophages. Data are mean ± SEM; one-way ANOVA was used for the statistical analysis. n=5 independent cell cultures; (E) RT-PCR analysis showed the mRNA level of CD206 and Arg1 in macrophages. Data are mean ± SEM; one-way ANOVA was used for the statistical analysis. n=5 independent cell cultures; (F) Immunofluorescence images showed the LC3 level in macrophages. Scale bars represent 50 μm. **P<0.01. ***P<0.001.

Fig 5: Validation of prognostic macrophage markers. A‐C, Representative immunofluorescence staining of breast cancer TMAs with MΦ markers CD163/CD68 (green) and CD206 (yellow), together with PLAC8 (magenta), SERPINH1 (blue), MORC4 (red) (A, B), or MHCII (red) (C). Nuclei were counterstained with DAPI (white). Magnifications serve to illustrate the presence of CD206+ MORC4+ (A, upper panels), CD206+ SERPINH1+ (B), and CD206− MHCII+ macrophages (C). Furthermore, SERPINH1+ PLAC8+ fibroblasts are displayed (A, lower panels)