Project description:To investigate the mechanisms used by memory CD4+ T cells to protect the skin from poxvirus infection, we infected LCMV immune mice with VacV on the right ear skin and VacV expressing the dominant MHC-II restricted epitope from LCMV (VacV-Ii-GP61) on the left ear skin. On day 3 after the co-infection, RNA from whole skin was isolated and gene expression was analyzed using RNA sequencing.
Project description:The poxviruses are a family of linear double-stranded DNA viruses about 130 to 230 kbp, that belong to the family Poxviridae. The poxviruses have an animal origin and have evolved to infect a wide host range. Variola virus (VARV), the causative agent of smallpox, is a poxvirus that infect only humans, but other poxviruses such monkey pox virus and cowpox virus have also across over from animals to infect humans. Therefor understanding the biology of poxviruses can help to devise antiviral strategies. In this study we used a system-based approach to examine the host responses to three different orthopoxviruses, CPXV, VACV and ECTV in the murine macrophage RAW 264.7 cell line.
Project description:We analyzed effects of monkeypox (MPX) and vaccinia (VAC) infection on cultured human cells. Our custom-designed arrays contain >18,000 human genes as well as genes for each open reading frame of MPX and VAC. A reference experiement design type is where all samples are compared to a common reference. Compound Based Treatment: Chemical used for treating infected cells Infection: Virus used for infection Cell Line: Cell line used for infection Time: Time cells were harvested after infection Keywords: reference_design Experiments used primary human monocytes with mock infection, VAC-Western Reserve, VAC-NYBOH, MPX-Zaire, gamma irradiated (killed) MPX-Zaire, and Ebola-Zaire (EBOV) (deposited March 2004). The next set of experiments used primary human monocytes (Mf7.29) and primary human fibroblasts (S100) with mock infection, VAC-Western Reserve, MPX-Zaire, and gamma irradiated MPX-Zaire (g-MPX) (deposited August 2004). The final set used primary human monocytes (Mph), primary human fibroblasts (NHDF), and HeLa cells with mock infection, VAC-Western Reserve, MPX-Zaire, and gamma irradiated MPX-Zaire (g-MPX). Some cells were treated with ionomycin + PMA (I+P), cycloheximide (CHX) or hydroxyurea (HU). (deposited November 2005). Total RNA from cells was harvested at each timepoint, 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 sample was hybridized versus a common reference (Stratagene Universal Human Reference RNA plus a mix of poxvirus transcripts from all timepoints). The reference was1 round of amplification (Ambion MessageAmp I), directly labeled by Cy3 incorporation during reverse transcription of amplified reference RNA. Cy3 and Cy5 samples were hybridized to a custom human-poxvirus array. Fluorescent images of hybridized microarrays were acquired using the GenePix 4000B and GenePix 4200AL microarray scanners.
