Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of cynomolgus macaques sequential blood samples infected with Ebola virus

ABSTRACT:

Title: Ebola-infected non-human primates - PBMCs

Summary: Infection with Ebola virus (EBOV) causes a fulminant and often fatal hemorrhagic fever. In order to improve our understanding of EBOV pathogenesis and EBOV-host interactions we have examined the molecular features of EBOV infection in vivo. Using self-spotted cDNA microarrays, we analyzed genome-wide host expression patterns in sequential blood samples from nonhuman primates (NHP) infected with EBOV.

Overall Design: Ebola infection of non-human primates (cynomolgus macaques). We took sequential samples of peripheral blood mononuclear cells (PBMC) from 15 cynomolgus macaques infected intramuscularly with 1000 plaque forming units (PFUs) of EBOV, Zaire strain. Peripheral blood samples (2.5mL) were collected on days 1, 4, or 6 prior to infection, in order to define a robust baseline, and then on successive days after infection, until death (immediately prior to euthanasia). Animals were euthanized at days 1, 2, 3, 4, 5, and 6 postinfection. PBMC's were collected at days post-infection as noted, preserved in Trizol, Trizol-extracted total RNA, 1 round amplified (Ambion MessageAmp I), directly labeled by Cy5 incorporation during reverse transcription of amplified RNA. Each blood sample was hybridized versus a common reference. The same reference is used for all samples - Stratagene Universal Human Reference RNA, 1 round of amplification (Ambion MessageAmp I), directly labeled by Cy3 incorporation during reverse transcription of amplified reference RNA. Two to three biological replicates of pre-infection samples were taken for each animal, as well as two to nine biological replicates (from different animals) at each day post-infection. A total of 50 arrays, representing 15 animals is included in this dataset.

ORGANISM(S): Macaca fascicularis

SUBMITTER: Janos Demeter

PROVIDER: E-SMDB-4098 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Non-human primates are the animals closest to humans for use in influenza A virus challenge studies, in terms of their phylogenetic relatedness, physiology and immune systems. Previous studies have shown that cynomolgus macaques (Macaca fascicularis) are permissive for infection with H1N1pdm influenza virus. These studies have typically used combined challenge routes, with the majority being intra-tracheal delivery, and high doses of virus (> 107 infectious units). This paper describes the outcome of novel challenge routes (inhaled aerosol, intra-nasal instillation) and low to moderate doses (103 to 106 plaque forming units) of H1N1pdm virus in cynomolgus macaques. All 4 challenge groups showed sero-conversion and evidence of virus replication, although the disease was sub-clinical. Intra-nasal challenge led to an infection confined to the nasal cavity. A low dose (103 plaque forming units) did not lead to detectable infectious virus shedding, but a 1000-fold higher dose led to virus shedding in all intra-nasal challenged animals. In contrast, aerosol and intra-tracheal challenge routes led to infections throughout the respiratory tract, although shedding from the nasal cavity was less reproducible between animals compared to the high-dose intra-nasal challenge group. Intra-tracheal and aerosol challenges induced a transient lymphopaenia, similar to that observed in influenza-infected humans, and greater virus-specific cellular immune responses in the blood were observed in these groups in comparison to the intra-nasal challenge groups. Activation of lung macrophages and innate immune response genes were detected at days 5 to 7 post-challenge. The kinetics of infection, both virological and immunological, were broadly in line with human influenza A virus infections. These more authentic infection models should be valuable in the determination of anti-influenza efficacy of novel entities against less severe (and thus more common) influenza infections.

Project description:Ebola (EBOV) virus causes severe and often lethal hemorrhagic fever in humans and nonhuman primates (NHP), and has been classified as a Category A bioweapon agent. There are currently no approved preventive vaccines or postexposure treatments for EBOV hemorrhagic fever. The mechanisms of EBOV pathogenesis are only partially understood, but the dysregulation of normal host immune responses (including destruction of lymphocytes, increases in levels of circulating proinflammatory cytokines, and development of coagulation abnormalities) is thought to play a major role. Accumulating evidence suggests that much of the observed pathology is not the direct result of virus-induced structural damage but rather is due to the release of soluble immune mediators from EBOV-infected cells. It is therefore essential to understand how the candidate therapeutic may be interrupting the disease process and/or targeting the infectious agent. Identification of effective treatment strategies may greatly benefit based on identification of molecular features of the host response to infection and treatment. In order to identify these gene signatures related to correlates of protection, we used a DNA microarray-based approach to compare the host genome-wide responses of EBOV-infected NHP responding to candidate therapeutics. With this approach, we have identified genes that appear to correlate with survival, including chemokine ligand 8 (CCL8/MCP-2), and revealed a subset of distinctly differently expressed genes that may provide possible targets for future diagnostics or therapeutics. These analyses will assist us in understanding the pathogenic mechanisms of EBOV infection as well as identify improved therapeutic strategies. Transcriptional analysis of global gene expression changes in Zaire Ebola Virus (ZEBOV)-infected rhesus macaques that were treated with either recombinant nematode anticoagulant protein c2 (rNAPc2) or recombinant human activated protein C (rhAPC). Animals were infected with 1000pfu ZEBOV, then subsequently treated with rNAPc2 or rhAPC. Four animals were left untreated for controls. Blood samples were taken at specified days post-infection and PBMCs were isolated from the samples and inactivated in TRIzol reagent. Total RNA was isolated from the samples, then linearly amplified and hybridized to a whole genome long-oligonucleotide microarray in a two color comparative format with a commercially available human reference RNA from Stratagene as a consistent control in dataset comparisons.

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Transcription profiling of cynomolgus macaques sequential blood samples infected with Ebola virus

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