Fig. 4

C87 and GSK872 pretreatment alleviated glia activation, monocyte infiltration, endothelial cell MLKL activity and downregulated KIR4.1 expression. Mice were treated with two doses of C87, a TNFα receptor inhibitor, or vehicle at 24 and 1 h before 4 mg/kg LPS injection (i.p.), or with GSK872, a RIP3 inhibitor, at 1 h before LPS was administrated, and then killed 72 h after LPS administration. Hippocampal tissues of these mice were prepared for immunostaining (n = 3 per group). A Representative images of microglia stained with Iba-1 and monocytes stained with CD68 in the CA3 region of the hippocampus. Increased cell size and irregular shape were identified as the activated microglia. B The proportions of activated microglia and C the number/mm² of CD68-positive cells in the CA3 of the hippocampus were estimated. D Representative images of astrocytes stained with GFAP and Kir4.1. Hypertrophic morphology identified as the activated astrocytes. E The proportions of activated GFAP-positive astrocytes and F the densitometric analysis of Kir4.1- and GFAP-positive protein levels in CA3 hippocampus. G Representative images of brain endothelial cells stained with CD31 and p-MLKL in the CA3 regions and H densitometric analysis of co-localized CD31- and p-MLKL-positive protein levels. Densitometry was semi-quantified using ImageJ software. Data represent the mean ± SEM of values from three animals per treatment group. One-way ANOVA; Bonferroni post hoc test vs. vehicle-treated mouse group; *p < 0.05, **p < 0.01, ***p < 0.001