Fig. 4

Absence of TM-LeD increases the microglia phagocytic activity and capacity to reduce HSV-1 replication in neurons. A Leukocytes purified from brains of mock-infected or infected WT or TM^LeD/LeD mice at 3 and 5 dpi were assayed for markers of microglia phagocytosis (CD45^intCD11b⁺CD68⁺). The level of cells in mock-infected WT mice was set as 100%. Primary brain microglia cultured from WT or TM^LeD/LeD mice were assessed for CD68 (B) or subjected to the zymosan phagocytosis assay (C). D Primary brain microglia cultured from WT or TM^LeD/LeD mice were mock-infected or infected with HSV-1 (MOI = 0.001) for 23 h, incubated with zymosan for 1 h, and assayed for the mean fluorescence intensity (MFI). Panels A–D were assayed by flow cytometry. E Primary brain microglia cultured from WT or TM^LeD/LeD mice were incubated with HSV-1 strain DG-1 containing VP16-GFP (MOI = 10) for 30 min and assayed for GFP levels by a microplate reader. F Primary brain neurons were cultured from WT mice, incubated with or without primary brain microglia cultured from WT or TM^LeD/LeD mice, and infected with HSV-1 (MOI = 0.001). The whole infected culture with both cells and supernatant was harvested at the indicated hours for viral titration. G Samples prepared as described in F were subjected to immunofluorescence staining 72 hpi with antibodies against mouse MAP2 or cleaved caspase 3. Scale bar, 50 μm. MAP2⁺ and cleaved caspase 3⁺ cells in images (300 × 300 μm) were counted. The data represent means ± or + SEM (error bars) of 3–12 samples per data point or group. *P < 0.05; **P < 0.01; and ***P < 0.001