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Y.J. Gordon, E.G. Romanowski, S.A. Harvey; Adenovirus Infection of Human Conjunctival Cells: Insights into the Innate Immune Response by Microarray Analysis . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4186.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Adenovirus (Ad) is the major cause of viral conjunctivitis worldwide and innate immunity serves an immediate role to prevent and eradicate infection. The defensins and defensin-like molecules are key components of innate immunity, small cationic peptides that may play a direct antiviral role in the eye (Romanowski et al. 2003). We characterized their expression changes in target conjunctival cells responding to Ad infection. Methods: Cultured primary human conjunctival epithelial cells were infected with Ad serotype 5 at multiplicities of infection (MOI) of 0, 0.1 or 1.0 and harvested at different times post-infection (1.5h, 6h and 16h). Total cell RNA was isolated, processed and analyzed using the Affymetrix HU133A GeneChip System (≈ 22,000 transcripts). We focused on transcripts which were present in control cells and were highly expressed (> 50th percentile). Results: At 16h post-infection, the 6 host genes up-regulated by ≥ 2-fold in highly infected cells (MOI=1.0) vs. uninfected cells (MOI=0) included those plausibly required for viral replication, assembly and packaging: ribonucleotide reductase, thymidylate synthetase, DEK oncogene, and the Hsp70 heat shock proteins HSPA1A and HSPA1B. At this time, human defensin ß2 (hDEF B2) was one of 54 genes down-regulated by ≥ 2-fold. We reasoned that during sparse infection (M=0.1) signaling from infected cells would up-regulate antiviral genes in the majority of uninfected cells, providing a more focused comparison. At 6h post-infection 46 transcripts were up-regulated by ≥ 2-fold in sparsely infected (MOI=0.1) vs. highly infected (MOI=1.0) cells; these transcripts included human defensin ß1 (hDEF B1). At 16h post-infection the same criteria yielded 18 genes, 14 of which are known to be interferon-induced. These 14 included chemokines CXCL 10 and CXCL 11, specific ligands for a receptor (CXCR3) absent from conjunctival epithelial cells but highly co-expressed with IFN-γ in CD8+ T cells. As well as their ability to recruit T-cells with antiviral properties, these chemokines are reported to have direct defensin-like activity against bacteria. Conclusion: The ocular innate immune response to physiologically relevant, sparse (MOI=0.1) Ad infection includes up-regulation of chemokines CXCL 10 and CXCL 11. Both this up-regulated expression and the constitutive expression of hDEF B1 and hDEF B2 are ablated by a greater multiplicity of Ad infection (MOI=1.0). Future studies will determine whether CXCL 10 and CXCL 11 share the direct antiviral effects of hDEF B1 and hDEF B2.
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