Dengue Viral Infection of Endothelial Cells and Dengue Hemorrhagic Fever

Thesis (Ph.D.)--University of Rochester. School of Medicine & Dentistry. Dept. of Microbiology and Immunology, 2011.Dengue virus (DENV) is the etiological agent responsible for the severe and potentially fatal illnesses known as dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS). Microvascular plasma leakage is the hallmark of DHF/DSS. The major causes of this vascular leakage that have been suggested are: 1) DENV infection of endothelial cells and consequent apoptosis, and 2) activation of immune cells or immune factors by DENV, in the presence or absence of anti-DENV antibodies, produces mediators of vascular endothelium permeability. To examine the first postulate, we characterized and compared dengue virus-2 infection of primary human umbilical vein endothelial cells (HUVEC), the HBEC-5I brain microvascular endothelial cell line, and the HMEC-1 dermal microvascular endothelial cell line. Although these cells all produced relatively high DENV titers, dengue viral infection kinetics differed by endothelial cell type, and an overall lack of apoptosis was observed. We also found that these endothelial cells cannot support antibody-dependent enhancement of DENV infection due to their lack of Fcγ type I and II receptors. Importantly, we discovered that the major heparan sulfate proteoglycans of the vasculature, the syndecans, mediate DENV entry and infection in endothelial cells. Furthermore, syndecan-4 was essential for DENV infection of primary HUVEC. To examine the second postulate, we developed an in vitro vascular endothelium permeability model to assess the disruption of the endothelial cell barrier by supernatants from peripheral blood mononuclear cells (PBMC) activated by DENV, in the presence or absence of anti-dengue pooled human sera. The results revealed that DENV in combination with anti-dengue pooled human sera can effectively activate PBMC into secreting mediators of vascular permeability, without enhancement of viral infection. These studies highlight the complexity of the direct and indirect effects of dengue viral infection on vascular endothelial cells, and imply an active role for the vascular endothelium in vascular leakage associated with DHF. Thus, vascular leakage associated with DHF/DSS may result not from one single mechanism, but from multiple interactions between DENV, immune cells, and a dynamic vascular endothelium. Elucidation and understanding of the specific interactions with the vascular endothelium that contribute to vascular leakage will aid in the development of prophylactic and therapeutic drugs against DHF/DSS