Fig. 9

Early and higher infection of circulating CD11b⁺Ly-6C⁺ monocytes in the infection site of IFN-I signal-incompetent mice is associated with quick viral dissemination. a Prerequisite infection of CD11b⁺Ly-6C⁺ monocytes in primary inoculation site of JEV. Cells were prepared from peripheral tissues (footpad) for primary inoculation site of JEV (5 × 10⁶ ffu/mouse) by collagenase digestion 48 h pi. Subpopulations of JEV NS1 protein-positive leukocytes were detected using flow cytometric analysis after staining cells with myeloid and monocyte/macrophage markers (CD11b, Ly-6C, F4/80) combined with Mab against JEV NS1 protein. b Earlier, higher frequency and number of JEV Ag-positive CD11b⁺Ly-6C⁺ monocytes in primary inoculation sites of IFN-I signal-incompetent mice. JEV NS1 protein-positive CD11b⁺ subpopulations were assessed by flow cytometric analysis using footpad-derived cells stained with CD11b, Ly-6C, and F4/80 combined with JEV NS1 antibody 24 and 48 h pi. c Detection of JEV-infected CD11b⁺Ly-6C⁺ monocytes in the liver following JEV infection. Cells were prepared from liver tissues by collagenase perfusion digestion 48 h pi and used for the analysis of JEV NS1-positive cells. d Enhanced JEV infection of liver CD11b⁺Ly-6C⁺ monocytes in IFN-I signal-incompetent mice. JEV NS1 protein-positive CD11b⁺ subpopulations were assessed by flow cytometric analysis 24 and 48 h pi. e Enhanced infection of IFN-I signal-deficient CD11b⁺Ly-6C⁺ monocytes in blood. Collected PBL was stained with CD11b, Ly-6C, and F4/80 combined with JEV NS1 antibody 24 h pi. JEV NS1-positive cells were assessed by flow cytometric analysis. f Enhanced CNS infiltration of JEV Ag-positive CD11b⁺Ly-6C⁺ monocytes in IFN-I signal-incompetent mice. Infiltrated leukocytes prepared from brain of IFN-I signal-competent and -incompetent mice were used to analyze JEV Ag-positive CD11b⁺Ly-6C⁺ monocytes with flow cytometry 48 h pi. g and h Involvement of CD11b⁺Ly-6C⁺ monocytes in quick viral dissemination in IFN-I signal-incompetent mice using BM chimeric models. Infiltrated leukocytes prepared from footpad and liver of BM chimeric hosts were used for the detection of JEV Ag-positive CD11b⁺Ly-6C⁺ monocytes 48 h pi. Data in bar graphs denote the mean ± SEM. Results are representative of one out of at least two individual experiments with four to five mice per group. Statistical significance *p < 0.05; **p < 0.01; ***p < 0.001 was assessed by an unpaired two-tailed Student’s t test