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Gene expression of a variety of Huh-7 derived hepatoma cells and their susceptibility to infection by JHF1 HCV in the HCV cell culture system.

ABSTRACT: Hepatitis C virus (HCV) is a global problem. To better understand HCV infection researchers employ in vitro HCV cell-culture (HCVcc) systems that use Huh-7 derived hepatoma cells that are particularly permissive to HCV infection. A variety of hyper-permissive cells have been subcloned for this purpose. In addition, subclones of Huh-7 which have evolved resistance to HCV are available. However, the mechanisms of susceptibility or resistance to infection among these cells have not been fully determined. In order to elucidate mechanisms by which hepatoma cells are susceptible or resistant to HCV infection we performed genome-wide expression analyses of six Huh-7 derived cell cultures (Huh-7, Huh-7.5.1, Huh-7.5.1c2, R1.09, R1.10 and R2.1) R that have different levels of permissiveness to infection. A great number of genes, representing a wide spectrum of functions are differentially expressed between cells. To focus our investigation, we identify host proteins from HCV replicase complexes, perform gene expression analysis of three HCV infected cells (infected Huh-7, Huh-7.5.1 and Huh-7.5.1c2) and conduct a detailed analysis of differentially expressed host factors by integrating a variety of data sources. Our results demonstrate that changes relating to susceptibility to HCV infection in hepatoma cells are linked to the innate immune response, secreted signal peptides and host factors that have a role in virus entry and replication. This work identifies both known and novel host factors that may influence HCV infection. Our findings build upon current knowledge of the complex interplay between HCV and the host cell, which could aid development of new antiviral strategies. Six Huh-7 derived hepatoma cell types that have different levels of susceptibility to HCV infection in cell culture are used: Huh-7, Huh-7.5.1, Huh-7.5.1c2, R1.09, R1.10 and R2.1. Of these the first three (label starting Huh are susceptible to HCV infection and the latter three (label starting R are resistant to HCV infection. All cell types are derived from Huh-7. Huh-7.5.1 is a subclone of Huh-7.5 that in turn is a subclone of Huh-7. Huh-7.5.1c2 is a subclone of Huh-7.5.1. R1.09 and R1.10 are subclones of R1 that is inturn a sublone of Huh-7.5,1. R2.1 is a subclone of Huh-7.5.1. Overall design: 39 samples are used. In every case there are 3 biological replicates, i.e., there are 13 unique conditions (39/3=13). These samples are subdivided between two studies: (1) A comparison of HCV infection resistant cells R1.09, R1.10 and R2 against HCV susceptible Huh-7.5.1. (2) A comparison of JFH1 HCV infected Huh-7, Huh-7.5.1 and Huh-7.5.1c2 cells versus their uninfected counterparts. In each case there are two uninfected counterparts, cells that were harvested after being in culture for 20 hours and cells that were harvested at the same time point as the infected cells, that is, at the peak time of infection. Peak times for infection vary between cells, depending on their susceptibility to infection: 168 hours for Huh-7, 120 hours for Huh-7.5.1 and 96 hours for Huh-7.5.1c2.

INSTRUMENT(S): [HG-U133_Plus_2] Affymetrix GeneChip Human Genome HG-U133 Plus 2 Array [Brainarray Version 12]

ORGANISM(S): Homo sapiens  

SUBMITTER: David Robertson  




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