Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Trancription profiling of the hepatoma cell line Huh7 infected with the Hepatitis C Virus

ABSTRACT: Hepatitis C Virus is a leading cause of chronic liver disease. The identification and characterisation of key host cellular factors that play a role in the HCV replication cycle is important for the understanding of disease pathogenesis and the identification of novel anti-viral therapeutic targets. Gene expression profiling of HCV infected Huh7 cells by microarray analysis was performed to identify host cellular genes that are transcriptionally regulated by infection. The expression of host genes involved in cellular defence mechanisms (apoptosis, proliferation and anti-oxidant responses), cellular metabolism (lipid and protein metabolism) and intracellular transport (vesicle trafficking and cytoskeleton regulation) was significantly altered by HCV infection. The gene expression patterns identified provide insight into the potential mechanisms that contribute to HCV associated pathogenesis. These include an increase in pro-inflammatory and pro-apoptotic signalling and a decrease in the anti-oxidant response pathways of the infected cell. 5x105 Huh7 cells were seeded in 25cm2 culture flasks and infected in triplicate either with the genotype 2a HCV clone, JFH-1 at a multiplicity of infection (MOI) of 3 or mock infected with an equal volume of concentrated conditioned growth medium. At 6, 12, 18, 24 and 48 hours post-infection, cellular RNA was extracted using TRIzol reagent (Invitrogen). Trizol lysates were shipped to Expression Analysis (NC, USA) where RNA was purified, quality tested using the Agilent Bioanalyser and hybridised onto Human U133 Plus 2.0 Affymetrix microarray chips for fluorescence data acquisition. In summary, a total of 30 RNA samples were analysed including 3x mock infected samples taken at 6, 12, 18, 24 and 48 hours post-treatment and 3x JFH-1 infected samples taken at 6, 12, 18, 24 and 48 hours post-infection. Two samples (Mock_6hrs_1 and JFH-1_6hrs_1) did not pass our data quality control measures and were therefore excluded from the statistical analysis.

ORGANISM(S): Homo sapiens

SUBMITTER: Samantha Blackham

PROVIDER: E-GEOD-20948 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Gene expression profiling indicates the roles of host oxidative stress, apoptosis, lipid metabolism, and intracellular transport genes in the replication of hepatitis C virus.

Blackham Samantha S Baillie Andrew A Al-Hababi Fadel F Remlinger Katja K You Shihyun S Hamatake Robert R McGarvey Michael J MJ

Journal of virology 20100303 10

Hepatitis C virus (HCV) is a leading cause of chronic liver disease. The identification and characterization of key host cellular factors that play a role in the HCV replication cycle are important for the understanding of disease pathogenesis and the identification of novel antiviral therapeutic targets. Gene expression profiling of JFH-1-infected Huh7 cells by microarray analysis was performed to identify host cellular genes that are transcriptionally regulated by infection. The expression of ...[more]

PMID: 20200238

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Project description:The main goal of this study is to characterize the transcriptional modulations induced by antiviral compounds derived from chloroquine on Huh7 cells infected with HCV. And thus to identify among the genes modulated by infection, those who are regulated by chloroquine and potentially involved in the antiviral activity of the compound.Two HCV cell models were used: a non-infectious HCV replicon and the infectious HCV cell culture (HCVcc). In addition, this study aimed to highlight the characteristics of derivatives of antiviral chloroquine, compared with chloroquine itself. Abstract from the associated publication: Autophagy is a process of self-degradation of cellular components in which double-membrane autophagosomes sequester organelles or portion of cytosol and fuse with lysosomes or vacuoles for breakdown by resident hydrolases. Autophagy is upregulated in response to extra- or intracellular stress and signals such as starvation, growth factor deprivation, ER stress, and pathogen infection. Indeed, infection with hepatitis C virus (HCV) was shown to induce autophagy through ER stress signaling and subsequent Unfolded Protein Response (UPR) activation. Moreover, a role of autophagy in promoting HCV infection has been suggested and chloroquine (CQ), a lysosomal protease inhibitor, has been seen blocking autophagy as well as inhibiting HCV replication. In the present report, mechanisms accounting for these inhibitory effects were investigated. Gene expression profiling was performed on CQ treated JFH-1-infected Huh7 cells to identify the host cellular genes that are transcriptionally regulated by infection, and silenced by CQ-based treatment. Herein, we demonstrate that CQ reduces the expression of genes induced by the viral infection such as those encoding autophagic key factors triggering the turn-off of mTOR activity as well as factors involved in p53 and NF-KappaB activities. However, we present several lines of evidence demonstrating that the CQ repressive effect observed on the HCV-induced pathways results from a decrease of ER stress due to the upstream pH-dependent inhibitory effect of CQ on viral replication. Gene expression modulations were measured in Huh7 harboring replicon cells stimulated with a 10M-BM-5M of a Chloroquine derivative compound called CQd, during 6h, 12h and 24h. Two independent experiments were performed at each time. An untreated control condition was performed in each experiment, expression was measured at the 24h time point. For the HCVcc model, gene expression analysis was analyzed in two conditions: infection and treatment of infected cells. To characterize modulations in time course of infection, Huh7 naM-CM-/ve cells were infected with JFH1/CS-N6-A4 viral stock and gene expression was measured at 6h, 24h and 48h postinfection. For each kinetic time point, an uninfected condition was performed and gene expression was measured. An additional control testing the interferon (IFN) induced modulations on infected cells was included and gene expression was measured after a 48h stimulation with 100 IU of IFN (alpha 2 b). To characterize the antiviral modulations resulting from treatment with Chloroquine (CQ) or a derivative compound (CQd), gene expression was measured in JFH1/CS-N6-A4 infected Huh7 cells treated with 40M-BM-5M of CQ or CQd during 12h or 48h post-treatment and infection. For each kinetic time point 12h and 48h, an untreated infected control was also performed and gene expression measured.

Dataset Information

Trancription profiling of the hepatoma cell line Huh7 infected with the Hepatitis C Virus

Publications

Gene expression profiling indicates the roles of host oxidative stress, apoptosis, lipid metabolism, and intracellular transport genes in the replication of hepatitis C virus.

Similar Datasets

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