Fig. 7

Temporal changes in microglial phenotypes in WNE. a tSNE plots showing the relative expression of P2RY12 on live myeloid cells from WNV-infected brains at dpi 0, 4, 5, 6 and 7. b IMC images from the frontal cortex of WNV dpi 0, 3, 5, 6 and 7 animals, showing the expression of Iba-1 (myeloid cells) (green), NS1 (WNV) (red) and NeuN (neurons) (blue). Scale bar represents 50 mm. c Histograms and statistical analysis of median fluorescence intensity (MFI) changes of selected markers/parameters on/in microglia at dpi 0, 4, 5, 6 and 7. Grey peaks/bars show unstained (uns.) cells, while pink peaks/bars show fluorescence minus one (FMO) for the respective marker. Data is representative of at least three independent experiments and presented as mean ± SEM, with 3-4 mice per group. *P < 0.0332, **P < 0.0021, Kruskal-Wallis test and Dunn’s multiple comparisons test. d Schematic showing microglia morphological and immunophenotypical changes upon WNV and monocyte infiltration during WNE. e Schematic showing the overall up- (orange arrow) or downregulation (blue arrow) of selected markers on P2RY12^hi CD86⁺ (dark purple), P2RY12^lo CD86⁻ (light green), P2RY12^lo CD86⁺ (light purple), P2RY12^hi CD86⁻ (dark green) microglial subsets from dpi 0 to dpi 7. Comparative levels of marker expression (black graded to white) between microglial subsets are shown for dpi 7. Microglial populations with a highest final expression of each marker (relative to the other 3 microglia subsets) are black. Note, this does not take account of any peak changes at dpi 5 and 6