Fig 2: High expression of SOCS3 in activated A1 astrocytes inhibited oligodendrocyte formation. (A) Schematic diagram showing the collection of astrocytes treated with control siRNA or SOCS3 siRNA, followed by culturing astrocytes with OPCs in the presence or the absence of IL-17A. (B) Representative images of OPCs cultured with astrocytes treated with control siRNA or SOCS3 siRNA for 72 h, which were immunostained for MBP and GFAP. Scale bar = 75 μm. (C) Percentage of MBP+ cells to total cell numbers. n = 3, *P < 0.05, **P < 0.01, ***P < 0.001, one-way ANOVA and Tukey’s test. Data in each statistical graph are presented as the mean ± SEM.

Fig 3: CFA induces the expression of GFAP, PKM2, p-STAT3, and HMGB1 protein in the spinal cord. (A) Immunostaining shows the expression of GFAP in the spinal cord of CON and CFA rats. GFAP was enhanced at seven days after CFA. n=3 rats/group. (B) Western blot shows the increase of GFAP protein expression in the spinal cord after CFA injection. n=3 rats/group. (C) Immunostaining shows the expression of PKM2 in the spinal cord of CON and CFA rats. PKM2 was enhanced at seven days after CFA. n=3 rats/group. (D) Western blot shows the increase of PKM2 protein expression in the spinal cord after CFA injection. n=3 rats/group. (E) Immunostaining shows the expression of p-STAT3 in the spinal cord of CON and CFA rats. p-STAT3 was enhanced at seven days after CFA. n=3 rats/group. (F) Western blot shows the increase of p-STAT3 protein expression in the spinal cord after CFA injection. n=3 rats/group. (G) Immunostaining shows the expression of HMGB1 in the spinal cord of CON and CFA rats. HMGB1 was enhanced at seven days after CFA. Scale bar: 100 µm. n=3 rats/group. (H) Western blot shows the increase of HMGB1 protein expression in the spinal cord after CFA injection. n=3 rats/group. *, P<0.05 compared to CON; **, P<0.01 compared to CON. CFA, complete Freund’s adjuvant.

Fig 4: GSDMD-KO mitigated neuropathological alterations in the peri-injury cortex at 3 days after TBI. (A) Representative images of immunoblots for synaptic proteins (PSD95, SYN1, SNAP25, and VAMP1) and total protein. (B–E) Quantitative analysis of synaptic proteins (n = 6). (F) Representative immunofluorescence images of CD68, GFAP, MAP2, and NeuN in the peri-injury cortex after TBI or in the paired sham cortex. Scale bar = 50 μm. (G–J) Quantitative analysis of integrated intensity of CD68+, GFAP+, and MAP2 + cells and NeuN+ cell number/mm2 in the cortex (three slices from each brain and three brains were used for calculation). All data are presented as the mean ± SEM and were analyzed using one-way ANOVA followed by Tukey’s multiple comparisons test for significance. ns p > 0.05, *p < 0.05, **p < 0.01, and ***p < 0.001.

Fig 5: Expression of synaptic proteins associates negatively with SIV RNA and markers of neuroinflammation, and expression of the axonal protein NFL associates positively with neuroinflammation markers. (A to I) Expression of PSD-95 (A to C), synaptophysin (D to F), and Homer1 (G to I) associates negatively with SIV, IFN-γ, and GFAP RNA expression. (J) NFL expression does not associate with SIV RNA. (K and L) NFL expression associates positively with IFN-γ and GFAP RNA expression. Statistical analysis was done using Pearson’s correlation. Each dot represents the value for 1 of the 9 regions in an animal (18 animals).