Fig. 6: Poly(I:C) ameliorates ASD-like behaviors in Fmr1-KO mice by enhancing IL-17A release from cerebellar microglia.

a, b Relative IL-17A mRNA expression 3 or 6 h after i.p. injection of LPS (Fmr1-KO, n = 4 mice; Fmr1-KO + 3 h LPS, n = 4 mice; Fmr1-KO + 6 h LPS, n = 4 mice) or poly(I:C) (Fmr1-KO, n = 7 mice; Fmr1-KO + 3 h poly(I:C), n = 8 mice; Fmr1-KO + 6 h poly(I:C), n = 7 mice). c Immunofluorescent staining for IL-17A (green) and DAPI (blue) showing that poly(I:C) significantly increased IL-17A expression in the cerebellar Crus I (Fmr1-KO, n = 5 mice; Fmr1-KO + poly(I:C), n = 5 mice). d A schematic representation illustrating the purification of cerebellar microglia. Created with Biorender.com. e Volcano plots showing DEGs of purified samples from the cerebellar Crus I of poly(I:C)-treated Fmr1-KO mice compared to untreated Fmr1-KO mice (n = 3 mice for each group; fold change > 1.2 or fold change < 0.84, P < 0.1). f KEGG enrichment analysis of DEGs indicated that poly(I:C) activated immune response-related pathway, including IL-17 signaling pathway (P < 0.05), in cerebellar microglia. g The gating strategy and quantification of IL-17A-positive cerebellar microglia in WT, Fmr1-KO, and poly(I:C)-treated Fmr1-KO mice. Poly(I:C) increased the ratio of IL-17A-producing cerebellar microglia in Fmr1-KO mice (WT, n = 6 mice; Fmr1-KO, n = 6 mice; Fmr1-KO + poly(I:C), n = 6 mice). h Immunofluorescence staining showing that poly(I:C) treatment elevated the number of Arc-positive (green) cerebellar PCs (calbindin-positive cells, green) in Fmr1-KO mice (Fmr1-KO, n = 8 mice; Fmr1-KO + poly(I:C), n = 8 mice). i, j Pharmacological blockade of IL-17A signaling in PCs of the cerebellar Crus I by microinjection of IL-17RA Ab abolished the poly(I:C)-induced improvement of social deficits (WT, n = 7 mice; Fmr1-KO, n = 7 mice; Fmr1-KO + poly(I:C), n = 6 mice; Fmr1-KO + poly(I:C) + IL17RA Ab, n = 7 mice) and reduced repetitive stereotyped behaviors (WT, n = 7 mice; Fmr1-KO, n = 6 mice; Fmr1-KO + poly(I:C), n = 11 mice; Fmr1-KO + poly(I:C) + IL17RA Ab, n = 6 mice). Data are presented as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, n.s. indicates not significant.