Principal Investigator: Ana Fernandez-Sesma
Abstract: Dengue virus (DenV) is a devastating human pathogen that causes disease and viremia in millions of people worldwide every year. But very little is known about the initiation of immunity by DenV. In this proposal we will study the following aims: Aim 1. What is the role of different populations of human DCs in the initiation of immunity by DenV? We hypothesize that there is a coordinated contribution of different cells in tissue and blood for the initiation/evasion of immunity and the establishment of infection by DenV. And while mosquito derived DenV may be able to inhibit important elements of innate immunity, such as type I IFN, subsequent human immune cell derived DenV may not be able to do so. This could explain the failure of DenV to transmit from human to human even though it causes viremia. We will analyze the pattern of activation and susceptibility to infection by DenV of different populations of APCs from human blood using both mosquito and primary human immune cell derived DenV. We will infect whole PBMCs to provide bystander effects and crosstalk between different cells which may resemble better the initial steps of initiation/evasion of immunity by DenV in vivo. Aim 2. What is the contribution of individual DenV proteins to the maturational profile of DCs? We hypothesize that individual DenV proteins may be responsible for the induction/evasion of immunity in vivo. We will generate recombinant Newcastle disease virus (NDV) expressing individual DenV proteins and will infect DCs and other blood APCs. We will analyze the activation and cytokine/chemokines profile in those infected cells. Since NDV induces high levels of type I IFN production in those cells we may be able to detect still unknown IFN antagonists in DenV with this system. Additionally, these recombinant viruses will be used to generate monoclonal antibodies specific for non structural proteins of DenV. Aim 3. What is the effect of DV infection or expression of individual DenV proteins on the T cell priming ability of different DCs towards Th1 immunity? DCs are the main APCs involved in priming of T cells in vivo after interaction with pathogens. Monocyte derived DCs infected with DenV have been shown to induce mixed Th0 responses in naive T cells. We hypothesize that the lack of type I IFN released by DenV infected APCs results in the generation of weak Th0 immunity in the host. We will evaluate the ability of different APCs from blood to secrete type I IFN and to prime Th1 or Th2 T cell responses after infection with DenV or with recombinant NDV expressing DV proteins. We will use qRT-PCR and multiplex ELISA for Th1/Th2 genes and cytokines/chemokines on T cells and APCs, with special emphasis on the levels of type I IFN released APCs. Dengue virus (DenV) can cause devastating disease in humans and there is no vaccine or treatment available to date. We are using a human system to study DenV initiation and evasion of immunity by analyzing the pattern of gene expression and cytokine and chemokine secretion profile of human dendritic cells (DCs)and other antigen presenting cells (APCs) and T cells after exposure to DenV. We will generate recombinant viruses expressing individual DenV proteins to analyze their role in the initiation/evasion of immunity by DenV. Understanding the modulation of cytokine responses by DenV, such as type I interferon (IFN), can help in the design of therapeutic targets to fight DenV.
Funding Period: 2008-08-15 - 2013-07-31
more information: NIH RePORT