Genome-wide analysis of poly(I:C)-responsive gene expression by primary human keratinocytes with and without episomal human papillomavirus
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ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Judith Boer
PROVIDER: E-GEOD-21260 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Analysis of the effect of high-risk human papillomaviruses HPV16 and HPV18 in keratinocytes at gene expression level. The hypothesis tested in the present study was that HPVs inhibit the function of pathogen recognition receptors (PRRs), thereby evading the immune system. Results show that HPV-positive keratinocytes have a weaker response to poly(I:C), a synthetic double strand RNA agonist of viral PRRs, with IL1B as a central hub in a gene expression network of relative downregulation.
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Project description:The interaction between the transmembrane glycoprotein surface receptor CD40 expressed by skin keratinocytes (KCs) and its T cell-expressed ligand CD154 was suggested to exacerbate inflammatory skin diseases. However, the full spectrum of CD40-mediated effects by KCs underlying this observation is unknown. Therefore, changes in gene expression after CD40 ligation of KCs were studied by microarrays. CD40-mediated activation for 2 hours stimulated the expression of a coordinated network of immune-involved genes strongly interconnected by IL8 and TNF, while after 24 hours anti-proliferative and anti-apoptotic genes were upregulated. CD40 ligation stimulated the production of chemokines that attracted lymphocytes and myeloid cells from peripheral blood mononuclear cells (PBMCs). Thus, CD40-mediated activation of KCs resulted in a highly coordinated response of genes required for the local development and sustainment of adaptive immune responses. The importance of this process was confirmed by a study on the effects of human papilloma virus (HPV) infection to the KCâs response to CD40 ligation. HPV infection clearly attenuated the magnitude of the response to CD40 ligation and consequently the KCâs capacity to attract PBMCs. The fact that HPV attenuates CD40 signalling in KCs indicates the importance of the CD40-CD154 immune pathway in boosting cellular immunity within epithelia. Total RNA from eight 72 hours 50 IU/ml IFNgamma pre-stimulated primary undifferentiated keratinocyte cultures, four uninfected and four HPV-positive, that were either left unstimulated, or stimulated for 2 hours with IFNgamma and Control- or CD40L-expressing L-cells (mouse fibroblasts), or stimulated for 24 hours with IFNgamma and Control- or CD40L-expressing L-cells. The stimulated samples were generated in duplo. 72 samples were generated in total.
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Project description:Previous studies have shown that normal and HPV immortalized keratinocytes are sensitive to TNF anti-proliferative effect. Conversely, HPV18-immortalized keratinocytes are resistant to the cytostatic effect mediated by this cytokine. In this study we have compared gene expression patterns in primary cultures of human foreskin epidermal keratinocytes (PHK or NHFK) and HPV16 (HF698) and HPV18(HF18Nco)-immortalized cell lines, and profile the transcriptional changes 3 and 60 hours after TNF treatment. Keywords: global expression profile, microarray, HPV, TNF
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Project description:Previous studies have shown that normal and HPV immortalized keratinocytes are sensitive to TNF anti-proliferative effect. Conversely, HPV18-immortalized keratinocytes are resistant to the cytostatic effect mediated by this cytokine. In this study we have compared gene expression patterns in primary cultures of human foreskin epidermal keratinocytes (PHK or NHFK) and HPV16 (HF698) and HPV18(HF18Nco)-immortalized cell lines, and profile the transcriptional changes 3 and 60 hours after TNF treatment. Keywords: global expression profile, microarray, HPV, TNF In order to determine the effects of HPV infection and TNF treatment on global gene expression, were performed 2 independent experiment for each cell line, including the two periods of treatment with TNF. Furthermore, were performed 2 experiments for each control plate containing the non-treated cells.
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2014-01-14 | E-GEOD-54008 | biostudies-arrayexpress
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2014-06-26 | E-GEOD-58841 | biostudies-arrayexpress
Project description:To identify early processes in carcinogenesis, we used an in vitro model, based on the initiating event in cervical cancer, human papillomavirus (HPV) transformation of keratinocytes. We compared gene expression in primary keratinocytes (K) and HPV16-transformed keratinocytes from early (E) and late (L) passages, and from benzo[a]pyrene treated L cells (BP). The transformed cells exhibit similar transcriptional changes to clinical cervical carcinoma. We revealed a contraction in expression of the apoptotic network during HF1 cell transformation, which affected the ability of L and BP cells to execute apoptosis, but did not lead to resistance to apoptotic stimuli. The contraction in the apoptotic machinery during the process of transformation was accompanied by a switch from apoptosis to necrosis in response to CDDP. The shrinkage of the pro- and anti-apoptotic networks appears to be part of a general contraction in the number of genes transcribed in L and BP cells. We also identified a large group of genes with induced expression, which are involved in cell metabolism and cell cycle, suggesting increased investment of the transformed cell in cellular proliferation. We hypothesize that the decrease in expression of many diverse pathways, including the pro- and anti-apoptotic networks, cuts the energy requirements for cell maintenance, allowing energy to be diverted towards rapid cell proliferation. This study supports the hypothesis that the process of cancer transformation may be accompanied by a shift from apoptosis to necrosis. Experiment Overall Design: We isolated mRNA from three independent cultures each of Keratinocytes and HPV16-immortalized keratinocytes : E, L and BP cells, grown under basal conditions. cDNA was prepared and hybridized to the Human Genome U133A Array (Affymetrix).
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