Fig. 4

SRGN induces microglial proinflammatory activation and amplifies LPS-induced inflammation in vitro. A Volcano plot showing the DEGs between the control and rSRGN-treated primary microglia. B GO analysis of the DEGs between the control and rSRGN-treated primary microglia. Top 30 GO terms were listed. C KEGG enrichment of the DEGs between the control and rSRGN-treated primary microglia. Top 20 KEGG terms were listed. D The qPCR analysis of Il1b, Tnf, Nos2 and Il6 mRNA levels in primary microglia treated with rSRGN (50 ng/ mL) for 6 h. E The qPCR analysis of Cxcl1, Cxcl2, Cxcl10 and Ccl4 mRNA levels in primary microglia treated with rSRGN (50 ng/ mL) for 6 h. F Flow chart demonstrating the co-culture procedure (Created with BioRender.com). Primary microglia were treated with rSRGN (50 ng/ mL) or control solvent for 6 h. Then, these microglia were collected and resuspended with neuron culture medium. Afterwards, microglia were replated among neurons and the co-culture system were allowed to grow for another 24 h. G Representative immunofluorescence images showing the microglia (Iba1⁺, green)—neurons (MAP2⁺, red) co-culture. Scale bar, 50 μm. H The quantification of living neuronal bodies from the co-culture system. I The qPCR analysis of Il1b mRNA level in primary microglia extracted from neonatal Srgn-KO mice or their WT littermates, with or without LPS (100 ng/ml, 6 h) stimulation. Data represented as mean ± SEM, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001