Distinct Dictation of Japanese Encephalitis Virus-Induced Neuroinflammation and Lethality via Triggering TLR3 and TLR4 Signal Pathways

Abstract

Japanese encephalitis (JE) is major emerging neurologic disease caused by JE virus. To date, the impact of TLR molecules on JE progression has not been addressed. Here, we determined whether each TLR modulates JE, using several TLR-deficient mouse strains (TLR2, TLR3, TLR4, TLR7, TLR9). Surprisingly, among the tested TLR-deficient mice there were contrasting results in TLR3−/− and TLR4−/− mice, i.e. TLR3−/− mice were highly susceptible to JE, whereas TLR4−/− mice showed enhanced resistance to JE. TLR3 ablation induced severe CNS inflammation characterized by early infiltration of inflammatory CD11b+Ly-6Chigh monocytes along with profoundly increased viral burden, proinflammatory cytokine/chemokine expression as well as BBB permeability. In contrast, TLR4−/− mice showed mild CNS inflammation manifested by reduced viral burden, leukocyte infiltration and proinflammatory cytokine expression. Interestingly, TLR4 ablation provided potent in vivo systemic type I IFN innate response, as well as ex vivo type I IFN production associated with strong induction of antiviral PRRs (RIG-I, MDA5), transcription factors (IRF-3, IRF-7), and IFN-dependent (PKR, Oas1, Mx) and independent ISGs (ISG49, ISG54, ISG56) by alternative activation of IRF3 and NF-κB in myeloid-derived DCs and macrophages, as compared to TLR3−/− myeloid-derived cells which were more permissive to viral replication through impaired type I IFN innate response. TLR4 ablation also appeared to mount an enhanced type I IFN innate and humoral, CD4+ and CD8+ T cell responses, which were mediated by altered immune cell populations (increased number of plasmacytoid DCs and NK cells, reduced CD11b+Ly-6Chigh monocytes) and CD4+Foxp3+ Treg number in lymphoid tissue. Thus, potent type I IFN innate and adaptive immune responses in the absence of TLR4 were closely coupled with reduced JE lethality. Collectively, these results suggest that a balanced triggering of TLR signal array by viral components during JE progression could be responsible for determining disease outcome through regulating negative and positive factors.

A Japanese Encephalitis Virus Genotype 5 Molecular Clone Is Highly Neuropathogenic in a Mouse Model: Impact of the Structural Protein Region on Virulence

“…While circulation of cytokines and chemokines is known to facilitate BBB breaking and viral neuroinvasion (48,(52)(53)(54), we found that proinflammatory cytokines or immune response-related genes were induced only once the virus could be detected in the mouse brain (Fig. 6).…”

Encephalitic Arboviruses: Emergence, Clinical Presentation, and Neuropathogenesis

“…Likewise, lack of TLR3 in mice infected with JEV was associated with higher frequency of inflammatory monocytes in the CNS, enhanced number of activated microglia, and elevated levels of IL-6 and TNF-α in the CNS, along with increased BBB permeability [212]. However, ablation of TLR3 did not alter virus titers in the CNS after intracranial inoculation of mice with JEV [212]. Conversely, in another study there was no significant difference in survival between wild-type and TLR3-deficient mice after intracranial infection with WNV [117].…”

“…TLR3-deficient mice displayed increased viral load in the brain following intracranial challenge with WNV [116]. Likewise, lack of TLR3 in mice infected with JEV was associated with higher frequency of inflammatory monocytes in the CNS, enhanced number of activated microglia, and elevated levels of IL-6 and TNF-α in the CNS, along with increased BBB permeability [212]. However, ablation of TLR3 did not alter virus titers in the CNS after intracranial inoculation of mice with JEV [212].…”

Toll-like receptor-3 gene polymorphism in patients with Japanese encephalitis

“…To support this hypothesis, recent experimental study done on mice with JE showed that JEV triggered inflammatory response through TLR3 and there was an increase in viral proliferation and reduced inflammatory response in TLR3 knockout mice. Reduced inflammatory response promotes viral proliferation (Lindenbach et al, 2007;Jiang et al, 2014;Han et al, 2014). It has been demonstrated that out of 136 single nucleotide polymorphisms (SNPs) described within the human TLR3 gene, only two SNPs [A851T (rs5743316), C1234T (rs3775291)] actually affect the TLR3 function.…”