Fig. 4: FBFO@HM@aOPN NPs significantly polarized M0- and M2-like macrophages towards the M1-like phenotype in vitro.

a Flow cytometry analysis showing the internalisation of Cy5.5-labelled FBFO by Raw264.7 cells (n = 3 independent experiments). b CLSM image of Raw264.7 cells stained with DCFH-DA to indicate NP-induced ROS generation. Scale bar, 200 μm. c Volcano plot showing the expression profile related to DEGs related to macrophage polarisation in Raw264.7 cells treated with PBS or FBFO@HM@aOPN. d GO BP and (e) GSEA enrichment analyses of the DEGs between the PBS- and FBFO@HM@aOPN-treated Raw264.7 cells. Flow cytometry analysis of the proportions of f CD80⁺CD86⁺ (n = 3 independent experiments), g MHC I⁺ (n = 4 independent experiments), h H-2Kb/SIINFEKL⁺ (n = 4 independent experiments), and i CD206⁺ (n = 3 independent experiments) among Raw264.7 cells treated with different formulations under 660 nm irradiation. j Establishment of BMDMs. Flow cytometry analysis of the proportions of k CD80⁺CD86⁺, l MHC I⁺, m H-2Kb/SIINFEKL⁺, and n CD206⁺ cells in BMDMs treated with different formulations under 660 nm irradiation (n = 3 independent experiments). o GSEA enrichment analyses of the DEGs between the PBS- and FBFO@HM@aOPN-treated Raw264.7 cells. p Western blot analysis of STING-IRF3 and NF-κB p65 phosphorylation in Raw264.7 cells and BMDMs treated with different formulations under 660 nm irradiation. q Schematic diagram of the mechanism by which FBFO@HM@aOPN reprograms macrophages to the M1-like phenotype. All the data are presented as the means ± SDs. The p values were determined via two-tailed one-way ANOVA with a Tukey post hoc test (f–i, k–n), p > 0.05, no significance (ns), *p < 0.05; **p < 0.01; ***p < 0.001; and ****p < 0.0001. Source data are provided as a Source Data file.