Cellular Oxidative Stress Response Controls the Antiviral and Apoptotic Programs in Dengue Virus-Infected Dendritic Cells

Abstract

Dengue virus (DENV) is a re-emerging arthropod borne flavivirus that infects more than 300 million people worldwide, leading to 50,000 deaths annually. Because dendritic cells (DC) in the skin and blood are the first target cells for DENV, we sought to investigate the early molecular events involved in the host response to the virus in primary human monocyte-derived dendritic cells (Mo-DC). Using a genome-wide transcriptome analysis of DENV2-infected human Mo-DC, three major responses were identified within hours of infection - the activation of IRF3/7/STAT1 and NF-κB-driven antiviral and inflammatory networks, as well as the stimulation of an oxidative stress response that included the stimulation of an Nrf2-dependent antioxidant gene transcriptional program. DENV2 infection resulted in the intracellular accumulation of reactive oxygen species (ROS) that was dependent on NADPH-oxidase (NOX). A decrease in ROS levels through chemical or genetic inhibition of the NOX-complex dampened the innate immune responses to DENV infection and facilitated DENV replication; ROS were also essential in driving mitochondrial apoptosis in infected Mo-DC. In addition to stimulating innate immune responses to DENV, increased ROS led to the activation of bystander Mo-DC which up-regulated maturation/activation markers and were less susceptible to viral replication. We have identified a critical role for the transcription factor Nrf2 in limiting both antiviral and cell death responses to the virus by feedback modulation of oxidative stress. Silencing of Nrf2 by RNA interference increased DENV-associated immune and apoptotic responses. Taken together, these data demonstrate that the level of oxidative stress is critical to the control of both antiviral and apoptotic programs in DENV-infected human Mo-DC and highlight the importance of redox homeostasis in the outcome of DENV infection.

Author Summary

Dengue virus (DENV), the leading arthropod-borne viral infection in the world, represents a major human health concern with a global at risk population of over 3 billion people. Currently, there are no antivirals or vaccines available to treat patients with dengue fever, nor is it possible to predict which patients will progress to life-threatening severe dengue fever. Markers associated with oxidative stress responses have been reported in patients with severe DENV infection, suggesting a relationship between oxidative stress and viral pathogenesis. In order to uncover biological processes that determine the outcome of disease in patients, we utilized human dendritic cells, the primary target of DENV infection, in an in vitro model. Transcriptional analysis of pathways activated upon de novo DENV infection revealed a major role for cellular oxidative stress in the induction of antiviral, inflammatory, and cell death responses. We also demonstrated that antioxidant mechanisms play a critical role in controlling antiviral and cell death responses to the virus, acting as feedback regulators of the oxidative stress response. This report highlights the importance of oxidative stress responses in the outcome of DENV infection, and identifies this pathway as a potential new entry-point for treating dengue-associated diseases.