Abstract:
Japanese encephalitis virus (JEV), a mosquito-borne flavivirus, is a major cause of viral encephalitis in Asia. Even though the principle target cells for JEV in the central nervous system are neurons, the microglia is activated in response to JEV infection. This research aimed to investigate the relationship between JEV and microglial cells. The percentage of JEV infectivity in mouse microglial (BV-2) cell line at 8, 15 and 24 hr post infection was determined by flow cytometry. It was found that the percentage of infected cells were approximately 53.5, 71.3 and 83.6 respectively. The JEV binding protein (s) expressed on the surface of BV-2 cells was also identified. Using One dimensional and Two-dimensional gel electrophoresis to separate the membrane proteins, we later identified the 43 kDa laminin receptor precursor protein as a JEV binding protein by virus overlay protein binding assay (VOPBA) followed with liquid chromatography-mass spectrometry (LC/MS/MS). This newly identified JEV binding protein was further characterized by infection inhibition assay. BV-2 cells were mock-infected or infected with JEV in the presence of either 0 (control), 5,10 and 20 μg anti-laminin receptor antibody or 20 μg soluble laminin. The percentage of inhibition of JEV infection was determined by flow cytometry. Results showed a dose dependent pattern of inhibition in the presence of anti-laminin receptor antibody, determined at 15 hr post infection, compared to non-relevant antibody and control. Taken together, 43 kDa laminin receptor precursor protein is verified as JEV putative receptor on mouse microglial cell surface.
