Fig. 3: SELP inhibits the microglia pro-inflammatory phenotype and promotes their immunosuppressive activity.

A ELISA assay showing higher secretion of SELP by human microglia treated with PD-GB CM compared to naïve DMEM. Data represent mean ± s.d. Each dot represents a triplicate. The graph shows the average of three independent repeats. Statistical significance was determined using an unpaired, two-sided Student’s t test. B Real-time PCR showing higher expression SELP mRNA by human microglia treated with PD-GB4 CM compared to naïve DMEM. Data represent mean ± s.d. Each dot represents a triplicate. The graph shows the average of three independent repeats. Statistical significance was determined using an unpaired, two-sided Student’s t test. Representative image (C) and quantification (D) of flow cytometry analysis of PSGL-1 expression by human microglia, showing higher expression when treated with PD-GB4 GB CM compared to naïve DMEM. Data represent mean ± s.d. The graph shows the average of five independent experiments. Statistical significance was determined using an unpaired, two-sided Student’s t test. E tSNE plot of single-cell RNA-seq analysis of microglia and macrophages isolated from patients′ GB tumors, showing the expression of PSGL-1 by microglia compared to macrophages. Data obtained from Darmanis et al.⁵⁶. Treatment with rSELP resulted in elevated mRNA expression of ARG1 (F), reduced expression of iNOS (G) and elevated expression of IL-10 (H) and TGF-β (I) by human microglia. SELPi or anti-PSGL-1 neutralizing antibody rescued rSELP effects while anti-CD44 or CD24 neutralizing antibody did not affect gene expression. Data represent mean ± s.d. Each dot represents a triplicate. The graphs show the average of three independent studies. Statistical significance was determined using one-way ANOVA test with multiple comparisons adjustment. Phagocytic activity (J) and NO release (K) by human microglia were reduced when treated with rSELP, and were restored when inhibiting SELP or PSGL-1 but not when neutralizing CD44 or CD24. Data represent mean ± s.d. of four wells per group. The graphs are representative of three independent repeats. Scale bars represent 200 µm. Statistical significance was determined using one-way ANOVA test with multiple comparisons adjustment. Source data are provided as a Source Data file.