CLEC5A Regulates Japanese Encephalitis Virus-Induced Neuroinflammation and Lethality
Figure 5
Anti-CLEC5A mAb reduces viral load in the CNS and attenuates CNS inflammation.
(A) Viral titer was determined by plaque assay for sera and tissue homogenates from JEV infected mice with or without anti-CLEC5A mAb treatment (two-tailed Student's t-tests). (B) The extent of NS3 expression in the cortex was diminished after peripheral administration of anti-CLEC5A mAb at day 5 post JEV infection. Scale bars, 50 µm. Five random fields of views in medium power field (original magnification (OM)×200) were photographed, and the numbers of NS3+ cells in each sample were counted, summed up and represented as mean ± s.e.m. (under each picture) of four independent experiments. (C) Numbers of NS3+ leukocytes (CD45hiCD11b+ and CD45midCD11b+) in JEV-infected brain were significantly reduced by administration of anti-CLEC5A mAb at day 5 post JEV infection. (D) Quantification of viral copies in CD45hiCD11b+ (R1) and CD45lowCD11b+ (R2) cells sorted from the brain of JEV-infected mice. The average of total cell numbers in R1 and R2 were approximately 5×104 and 2×104, respectively, in the isotype treated group. Q-PCR assay was performed on total RNA from each population and normalized with its respective internal control. Data were collected from four independent experiments. Two-tailed Student's t-tests were performed. Cytokines levels in the sera (E) and CSF (F) were measured by ELISA at day 5–7 post JEV infection. The effects of CLEC5A mAb (3E3G4) and isotype control were compared using two-tailed Student's t-tests. (G) Intracellular TNF-α and IL-6 were detected in CD11b+ gated F4/80+ and CD45+ cells isolated from JEV-infected brain by flow cytometry.