Models to Study the Pathogenesis of Ebola Virus

In general, studies on Ebola virus require sophisticated yet inconvenient safety precautions, considerably limiting the investigation and understanding of Ebola pathogenesis. Fortunately, isolation of the viral cDNAs and the development of artificial expression strategies have allowed the study of Ebola virus gene products in vitro under less restrictive conditions. For example, an artificial replication system has been developed based on the vaccinia virus T7 expression model. Also, a reverse genetics system enables the generation of infectious Ebola virus from cloned cDNA. Using this strategy, cultured cells are transfected with plas-mids for the expression of the Ebola proteins, a plasmid for the Ebola viral RNA controlled by T7 RNA polymerase promoter, and a plasmid for T7 RNA polymerase. This system is effective for the study of mechanisms underlying the path-ogenicity of Ebola virus, as viral genes and proteins can be manipulated as desired.

Several animal models have also been used to examine Ebola virulence, the host responses to infection, and the efficacy of immunization. Although Ebola Zaire infection is traditionally examined using nonhuman primate hosts, the virus has recently been adapted to produce uniformly fatal infection in other animals including guinea pigs and mice, through serial animal-to-animal passaging. These models have provided critical insights into Ebola pathogenesis and are especially valuable for testing antiviral medications and vaccines.

More recently, a safe high-throughput avian-derived retroviral gene transfer system was engineered to specifically express candidate viral genes in mouse endothelial cells in vivo [10]. The TIE2-TVA transgenic mouse system expresses the avian leukosis virus (ALV) receptor, TVA, under the control of the endothelial cell-specific TIE2 promoter. Using this system, only mammalian cells engineered to express the TVA receptor can be transduced by infection with avian-derived viruses, enabling the somatic introduction of multiple genes in vivo, in a tissue-specific manner, using a single transgenic animal. This unique animal model safely mimics the infectious process by which Ebola targets endothelial cells in vivo. To verify viral infectivity and demonstrate the potential of this model to study HFV virulence genes, an ALV-derived vector [RCASBP(A)] expressing Polyoma Middle T Antigen (PyMT)—which induces hemorrhagic diathesis when expressed in mice—was used. Endothelial cell-specific retroviral transduction with PyMT had a dramatic effect on mouse survival. Histological examination of killed animals revealed massive hemorrhaging in the liver and spleen, very similar to that seen in human patients infected with Ebola virus. These promising preliminary results suggest that this model may be uniquely suited to safely examine the contribution of Ebola genes, individually or in combination, to VHF. This system may provide fundamental insight into the molecular pathogenesis of this lethal virus and may ultimately help identify diagnostic markers and gene-product targeted therapies for VHF.

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