Project description:We inoculated ARPE-19 human retinal pigment epithelial cells with EBOV, and followed course of infection by immunocytochemistry and measurement of titer in culture supernatant. To interrogate transcriptional responses of infected cells, we combined RNA sequencing with in silico pathway, gene ontology, transcription factor binding site and network analyses. Human retinal pigment epithelial cells were permissive to infection with EBOV, and supported viral replication and release of virus in high titer. Unexpectedly, 28% of 560 up-regulated transcripts in EBOV-infected cells were type I IFN responsive, indicating a robust type I IFN response.
Project description:We used small RNA sequencing (RNA-Seq) to identify miRNA expression in EBOV-infected human RPE cells, and conducted in silico analyses to identify biological targets of, and molecular interactions with, these miRNAs.
Project description:Episodic Ebola virus (EBOV) outbreaks, such as the current one in West Africa, emphasize the critical need for novel antivirals against this highly pathogenic virus. Here, we demonstrate that interferon gamma (IFNγ) prevents morbidity and mortality associated with EBOV infection when administered to mice either 24 hours prior to or 2 hours following EBOV infection. Microarray studies with IFNγ-stimulated human macrophages identified novel interferon-stimulated genes (ISGs) that inhibit EBOV infection upon ectopic expression. IFNγ treatment reduced viral RNA levels in macrophages to a similar degree as cells treated with the protein synthesis inhibitor, cycloheximide, suggesting that IFNγ treatment inhibits genome replication. As IFNγ treatment robustly protects mice against EBOV infection, we propose that this FDA-approved drug may serve as a useful prophylactic or therapeutic strategy during EBOV outbreaks, contributing to the currently limited arsenal of filovirus antivirals.
Project description:Episodic Ebola virus (EBOV) outbreaks, such as the current one in West Africa, emphasize the critical need for novel antivirals against this highly pathogenic virus. Here, we demonstrate that interferon gamma (IFNγ) prevents morbidity and mortality associated with EBOV infection when administered to mice either 24 hours prior to or 2 hours following EBOV infection. Microarray studies with IFNγ-stimulated human macrophages identified novel interferon-stimulated genes (ISGs) that inhibit EBOV infection upon ectopic expression. IFNγ treatment reduced viral RNA levels in macrophages to a similar degree as cells treated with the protein synthesis inhibitor, cycloheximide, suggesting that IFNγ treatment inhibits genome replication. As IFNγ treatment robustly protects mice against EBOV infection, we propose that this FDA-approved drug may serve as a useful prophylactic or therapeutic strategy during EBOV outbreaks, contributing to the currently limited arsenal of filovirus antivirals.
Project description:<p align="left"><b>Title: </b> Ebola-infected non-human primates - PBMCs<br> <p align="left"><b>Summary: </b> 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. <br> <p align="left"><b>Overall Design: </b> 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.
Project description:We performed transcriptomic analysis of a time course of Ebola virus (EBOV) iPSC-derived hepatocytes. Mock infected and EBOV infected hepatocytes were harvested 1, 2, 3, and 7 days post-infection for analysis. Hepatocytes were infected with EBOV at a multiplicity of infection of 10.
Project description:Ebola virus can cause a severe and often fatal hemorrhagic fever in humans and other mammals, known as Ebola virus disease (EVD),the mechanism of how this pathogenesis comes about is not well understood, It is assumed that miRNA may have important roles in virus infection response. To better understand the function of miRNA in EBOV infection disease, we undertook a miRNA profiling analysis using the whole blood of EBOV infection patients.
Project description:We used microarrays to evaluate the effect of SRPIN803 on gene expression in ARPE-19 cells. ARPE-19 cells were treated with SRPIN803 (10 uM) or the negative control (0.1% DMSO) for 4 hours for Total RNA isolation and hybridization on Microarray.