Analysis of Human Peripheral Blood Samples from Fatal and Nonfatal Cases of Ebola (Sudan) Hemorrhagic Fever: Cellular Responses, Virus Load, and Nitric Oxide Levels

Abstract

Peripheral blood samples obtained from patients during an outbreak of Ebola virus (Sudan species) disease in Uganda in 2000 were used to phenotype peripheral blood mononuclear cells (PBMC), quantitate gene expression, measure antigenemia, and determine nitric oxide levels. It was determined that as the severity of disease increased in infected patients, there was a corresponding increase in antigenemia and leukopenia. Blood smears revealed thrombocytopenia, a left shift in neutrophils (in some cases degenerating), and atypical lymphocytes. Infected patients who died had reduced numbers of T cells, CD8؉ T cells, and activated (HLA-DR ؉ ) CD8 ؉ T cells, while the opposite was noted for patients who survived the disease. Expression levels of cytokines, Fas antigen, and Fas ligand (TaqMan quantitation) in PBMC from infected patients were not significantly different from those in uninfected patients (treated in the same isolation wards), nor was there a significant increase in expression compared to healthy volunteers (United States). This unresponsive state of PBMC from infected patients despite high levels of circulating antigen and virus replication suggests that some form of immunosuppression had developed. Ebola virus RNA levels (virus load) in PBMC specimens were found to be much higher in infected patients who died than patients who survived the disease. Similarly, blood levels of nitric oxide were much higher in fatal cases (increasing with disease severity), and extremely elevated levels (>150 M) would have negatively affected vascular tone and contributed to virus-induced shock.

Post-exposure treatments for Ebola and Marburg virus infections

“…Systematic studies in experimentally infected NHPs suggest that monocytes, macrophages and dendritic cells are the early replication sites for EBOV and MARV 41,48,51 . Filovirus infection of mononuclear phagocytes is thought to trigger a series of events that includes the production and release of the procoagulant protein tissue factor 54,55 and an assortment of pro-inflammatory cytokines, chemokines and free radical species in NHPs and humans 48,51,[55][56][57][58][59][60][61][62][63][64][65][66][67][68] . This dysregulated host response likely plays a greater role in the development of the observed pathology than any structural damage caused by viral replication in host cells and/or tissues.…”

Ebola virus: The role of macrophages and dendritic cells in the pathogenesis of Ebola hemorrhagic fever

“…By contrast, some 10-20% of ZEBOV-infected humans in African outbreaks begin to show clinical improvement during the second week of illness and recover from their infection. Survival appears to correlate with the development of an antigen-specific immune response, usually marked by the appearance of ZEBOV-specific IgG (Baize et al, 1999;Sanchez et al, 2004).…”

Comprehensive Panel of Real-Time TaqMan™ Polymerase Chain Reaction Assays for Detection and Absolute Quantification of Filoviruses, Arenaviruses, and New World Hantaviruses

“…TaqMan™-based, real-time reverse transcriptionpolymerase chain reaction (RT-PCR) can rapidly detect and identify the selected gene targets found within the genomes of these RNA viruses. Although many research groups have developed and tested standard and real-time RT-PCR assays for detection of various species and corresponding strains of Ebola, [3][4][5][6][7][8] Marburg, 4,8,9 Lassa 8, 10-12 Junin, 13 and hantavirus, [14][15][16][17][18] a comprehensive panel of TaqMan™-based real-time PCR assays using identical cycling conditions is not currently available.…”