Molecular Mechanism of Altered Immunity in Dengue Hemorrhagic Fever

Background and hypothesis：We have previously found that dengue hemorrhagic fever （DHF）had less T helper type 1（Th1）reaction and augmented Th2 reaction (1-3), showing elevated IL-10 but lower IFNγ levels (4). Moreover, complication of DHF was higher in the elderly with chronic illness (5). Similarly, studies from Cheng-Kung University School of Medicine showed that autoimmune and cytokine storm reactions were potentially involved in DHF (6,7). These studies suggest altered immunity involved in pathogenesis of DHF. The cell and molecular mechanisms of DHF pathogenesis remain to be determined. Since dengue only causes disease in humans via mosquito bite, the virus should infect leukocytes or endothelial cells in blood via virus receptors. Variations of viral receptor genes or host conditions may affect innate immune mediators, which subsequently affect adaptive immunity towards Th2 and/or Th17 reactions and result in production of autoantibody and/or cytokine storm for immunopathogenesis and vascular damage of DHF. Specific aims：To prove the hypothesis, we propose 4 specific aims to reach the goal. Specific aim 1 is to study the responses of leukocytes and endothelial cells to dengue viruses. Studies will clarify how innate reactions under immune genes and host conditions affect viral replication and signal transduction of immune reactions. Alternatively activated monocyte/macrophage distribution after dengue infection will be assessed by flow cytometric analysis of CD14/IL-10/IL-12 staining, signal transduction will be assessed by flow cytometry and western blot, and innate immunity-mediated pro-T helper cytokine profiles: Pro-Th1:IL-12/IFNα, Pro-Th2: MCP-1/IL-10, Pro-Treg: TGFβ and Pro-Th17:IL-6/IL-17A will be assessed by multiplex bead-array assay. Specific aim 2 is to investigate how the dengue-mediated innate immunity alteration affects adaptive immune responses in molecular levels, particular polarization of Th1, Th2, Treg and Th17. Monocyte-derived dendritic cells in response to dengue viruses skew its maturation status and polarize Th cell transcription factors: T-bet/Gata-3/Foxp3/RORrT which will be assessed by RT-PCR to explore dengue-induced adaptive immunity under different types of viruses and host conditions. Specific aim 3 will validate the cell and molecular mechanism of dengue-induced immune response using plasma and leukocytes from patients with DF or DHF. Based on the first 2 years’ results, we will validate the mechanism(s) of altered immune responses in DHF by knockdown or overexpression of immune molecules. Specific aim 4 is to set up a dengue registry and sample collection core unit in Kaohsiung area. This core unit will be suitable for prospective validation of immunopathogenesis of DHF. As the dengue patients are more than 1000 cases in Kaohsiung area in 2010, we anticipate that there will be enough patients’ immune cells and plasma samples for our validation of functional immune reactions related to presence of previous dengue antibodies, viral load, genetic variations, ages, co-morbidities and treatment guidelines of DHF. Anticipated results：This study will clarify how dengue receptors mediate innate and adaptive immune responses in patients with altered innate or adaptive immune responses in patients with and without DHF. Results from this study may provide strategies to prevent DHF. Moreover this study will collaborate with subproject-1 to clarify what immune gene(s) is(are) involved in DHF, with subproject 3 to clarify how age and/ or co-morbidity is(are) involved in DHF, and with subproject 5 to investigate what virus genotype(s) is(are) involved in DHF. Finally, all these studies will provide clues for designing clinical trials of better therapeutic guidelines for DHF.

Project ID:PC10007-0352
External Project ID:NSC100-2314-B758-001-MY3