Japanese encephalitis is an acute zoonotic, mosquito-borne disease caused by Japanese encephalitis virus (JEV). Japanese encephalitis is characterized by extensive inflammation in the central nervous system (CNS) and disruption of the blood-brain barrier (BBB). However, the pathogenic mechanisms contributing to the BBB disruption are not known. Here, using a mouse model of intravenous JEV infection, we show that virus titers increased exponentially in the brain from 2 to 5 days postinfection. This was accompanied by an early, dramatic increase in the level of inflammatory cytokines and chemokines in the brain. Enhancement of BBB permeability, however, was not observed until day 4, suggesting that viral entry and the onset of inflammation in the CNS occurred prior to BBB damage.In vitrostudies revealed that direct infection with JEV could not induce changes in the permeability of brain microvascular endothelial cell monolayers. However, brain extracts derived from symptomatic JEV-infected mice, but not from mock-infected mice, induced significant permeability of the endothelial monolayer. Consistent with a role for inflammatory mediators in BBB disruption, the administration of gamma interferon-neutralizing antibody ameliorated the enhancement of BBB permeability in JEV-infected mice. Taken together, our data suggest that JEV enters the CNS, propagates in neurons, and induces the production of inflammatory cytokines and chemokines, which result in the disruption of the BBB.IMPORTANCEJapanese encephalitis (JE) is the leading cause of viral encephalitis in Asia, resulting in 70,000 cases each year, in which approximately 20 to 30% of cases are fatal, and a high proportion of patients survive with serious neurological and psychiatric sequelae. Pathologically, JEV infection causes an acute encephalopathy accompanied by BBB dysfunction; however, the mechanism is not clear. Thus, understanding the mechanisms of BBB disruption in JEV infection is important. Our data demonstrate that JEV gains entry into the CNS prior to BBB disruption. Furthermore, it is not JEV infectionper se, but the inflammatory cytokines/chemokines induced by JEV infection that inhibit the expression of TJ proteins and ultimately result in the enhancement of BBB permeability. Neutralization of gamma interferon (IFN-γ) ameliorated the enhancement of BBB permeability in JEV-infected mice, suggesting that IFN-γ could be a potential therapeutic target. This study would lead to identification of potential therapeutic avenues for the treatment of JEV infection.

Japanese encephalitis is a neuropathological disorder caused by Japanese encephalitis virus (JEV), which is characterized by severe pathological neuroinflammation and damage to the blood–brain barrier (BBB). Inflammatory cytokines/chemokines can regulate the expression of tight junction (TJ) proteins and are believed to be a leading cause of BBB disruption, but the specific mechanisms remain unclear. IP-10 is the most abundant chemokine produced in the early stage of JEV infection, but its role in BBB disruption is unknown. The administration of IP-10-neutralizing antibody ameliorated the decrease in TJ proteins and restored BBB integrity in JEV-infected mice. In vitro study showed IP-10 and JEV treatment did not directly alter the permeability of the monolayers of endothelial cells. However, IP-10 treatment promoted tumor necrosis factor alpha (TNF-α) production and IP-10-neutralizing antibody significantly reduced the production of TNF-α. Thus, TNF-α could be a downstream cytokine of IP-10, which decreased TJ proteins and damaged BBB integrity. Further study indicated that JEV infection can stimulate upregulation of the IP-10 receptor CXCR3 on astrocytes, resulting in TNF-α production through the JNK-c-Jun signaling pathway. Consequently, TNF-α affected the expression and cellular distribution of TJs in brain microvascular endothelial cells and led to BBB damage during JEV infection. Regarding regulation of the BBB, the IP-10/TNF-α cytokine axis could be considered a potential target for the development of novel therapeutics in BBB-related neurological diseases.

Japanese encephalitis virus (JEV) is a highly fatal pathogen to human beings. Toll like receptor 7 (TLR7) plays a role as first host defense against most ssRNA flaviviruses. This study aims to investigate the role of TLR7 in inducing adaptive immune response in mice against Japanese encephalitis virus. In vitro and in vivo study was conducted to examine the expression of TLRs in mice. After JEV infection, physical parameters of mice (Survival rate and body weight) were evaluated and organs or cell were collected for further analysis. TLR7 expression was increased significantly as compare to other TLRs post JEV infection. CD80, CD86 and CD273 cells expression on bmDC cells was increased significantly in TLR7-/- mice. Furthermore viral load was also increased significantly in TLR7-/- mice as compare to C57BL/6 mice. But there was no significant difference among survival rate and body weight in TLR7-/- mice as compare to C57BL/6. Interestingly, we also found that TLR8 was up-regulated in TLR7-/- mice. The study concluded that TLR8 was up-regulated in TLR7 deficient mice and it might play a compensatory role in the immune response in TLR7-/- mice.