The role of Ebola virus secreted glycoproteins in target cell activation

Ebola virus, a member of the family Filoviridae, causes one of the most severe forms of viral hemorrhagic fever with mortality rates up to 90%. In the terminal stages of disease symptoms progress to hypotension, coagulation disorders, and hemorrhages and there is prominent involvement of the mononuclear phagocytic and reticuloendothelial systems. Cells of the mononuclear phagocytic system are primary target cells and producers of inflammatory mediators and their activation is independent of virus replication. Virus-induced dysfunction of the endothelium, including endothelial cell damage and increased permeability, may occur directly through virus infection that leads to activation and lytic replication as well as indirectly by mediator-induced inflammatory responses. Ebola efficiently produces three secreted glycoproteins during infection: sGP, delta peptide, and GPI. While the presence of these glycoproteins has been confirmed in blood (sGP) and in vitro systems, it is hypothesized that they are of biological relevance in pathogenesis, particularly target cell activation. To gain insight into their function, we expressed the soluble glycoproteins in mammalian cells, characterized and subsequently purified them using immunoaffinity purification. Functional studies using these proteins were then performed on primary macrophages and human umbilical vein endothelial cells (HUVECs). Cells were treated with increasing amounts of glycoproteins and subsequently tested for activation by detection of pro-inflammatory cytokines and chemokines (macrophages) and adhesion molecules (HUVEC). Furthermore, HUVEC were monitored for changes in morphology, trans-endothelial resistance, and rearrangements of the endothelial junctions. Our data clearly demonstrate that the secreted glycoproteins do not play a role in activation of target cells.