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J.L. Stahl, E.B. Cook, F.M. Graziano, N.P. Barney; TNF–Mediated Upregulation of ICAM–1 on Conjunctival Epithelial Cells Is Modulated by the Level of Expression of the Receptor for Soluble TNF (TNFR1) . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2815.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Upregulation of ICAM–1 expression on the ocular surface is an important feature of ocular allergic inflammation. Our research has suggested that conjunctival mast cell mediators upregulate expression of TNFR1 on conjunctival epithelial cells and enhance the sensitivity of the cells to TNFα stimulation of ICAM–1 expression. We hypothesized that increased expression of TNFR1 on conjunctival epithelial cells enhances upregulation of ICAM–1 in response to TNFα. The purpose of this study was to examine regulation of TNFR1 expression on conjunctival epithelial cells and whether increased expression results in increased sensitivity to TNFα mediated ICAM–1 upregulation. Methods: Human conjunctival epithelial cells were enzymatically dispersed and purified from multiple donor pools of cadaveric conjunctival tissues. Conjunctival epithelial cells were incubated with TNFα (various concentrations) for various times and, in some cases, were pre–treated with tissue inhibitor of matrix metalloproteinase–2 (TIMP2, to inhibit shedding of TNFR1). Supernates were collected for analysis of soluble TNFR1 (sTNFR1) via ELISA and cells were harvested for analysis of ICAM–1 and TNFR1 expression using two–color flow cytometry analysis. Results: Stimulation of conjunctival epithelial cells with TNFα resulted in shedding of TNFR1 (decreased surface expression coordinated with increased release of sTNFR1) in a dose dependent manner beginning by 5 minutes post–challenge and continuing over 24 hours. Pre–treatment with TIMP2 enhanced both TNFα stimulated TNFR1 and ICAM–1 expression. Conjunctival epithelial cells expressing high levels of TNFR1 responded to low doses of TNFα with a greater increase in ICAM–1 expression than cells expressing low levels of TNFR1. Conclusions: These results demonstrate that the level of TNFR1 expression on conjunctival epithelial cells is an important factor in regulation of the ICAM–1 response and therefore may be a significant target for pharmaceutical intervention in ocular inflammation.
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