Figure 1 from Inhibition of Extracellular Matrix Protein Fibulin-3 Reduces Immunosuppressive Signaling and Increases Macrophage Activation in Glioblastoma

Fibulin-3 expression affects the accumulation and phenotype of TAMs. A, Representative images of GSC-derived intracranial tumors (GBM08 line) expressing control or fibulin-3 short hairpin RNAs (shRNA), showing TAM density (IBA1 staining) and expression of the immunosuppression marker CD206 in TAMs; cell nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI). Scale bars, 200 μm. B, Quantification of total TAMs (IBA1⁺ area within the tumor) and IBA1/CD206 coexpression in TAMs. The results were analyzed by Welch’s corrected t test. C, Representative flow cytometry gating of TAM populations (macrophages: CD45hi CD11b⁺; microglia: CD45lo CD11b⁺) recovered from the tumors indicated in A; note that the peripheral macrophage population is small due to the tumors being resected at an early stage. D, Flow cytometry quantification of CD206⁺ in TAMs recovered from freshly resected tumors; comparison by Student t test. E, qRT-PCR for selected genes expressed in TAMs recovered from control and fibulin-3–deficient tumors; analysis by multiple paired t tests with correction for multiple comparisons. F, Representative images of CNS1-derived intracranial tumors expressing control or fibulin-3 cDNA; stains are the same as in A; scale bars, 200 μm. G, Quantitative analysis of the tumors in (F) showing total TAMs (IBA1⁺ area within the tumor) and IBA1/CD206 coexpression in TAMs. The results were analyzed by Welch’s corrected t test. ns, not significant.