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I. Maltseva, C. Basbaum, S. Fleiszig; In vitro Contact Lens Exposure Attenuates Pseudomonas aeruginosa–mediated Toll–like Receptor Gene Expression . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2106.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: We previously reported that contact lens exposure (CLE) of corneal cells in vitro inhibits the induction of human beta defensin–2 (hbd–2) by Pseudomonas aeruginosa (PA). This seemed to derive from a failure of PA to stimulate JNK in the presence of CLE. Hbd–2 expression requires both NF–kB and JNK activation, both of which are downstream effectors of Toll–like receptor (TLR) signaling. To further understand the effect of CLE on hbd–2 expression, we investigated its effect on TLR signaling induced by PA. Methods: To determine whether TLRs were involved in hbd–2 expression, we co–transfected HCE cells with a dominant negative construct for the common TLR adaptor protein MyD88 (MyD C) and an hbd–2 reporter plasmid. Cells were exposed to PA supernatant in serum–free medium for 6 hours prior to luciferase assay. HCE were grown with or without CLE for 3.5 days. CLE was terminated and cells were exposed to PA for 6 hours before Real–Time PCR analysis. Results: Cells expressing dominant negative MyD88 lost responsiveness to PA. Real–Time PCR analysis revealed that P. aeruginosa induced mRNA expression of TLR2 4.8x, TLR4 9x and TLR5 2.5x. This response to the pathogen was decreased to 1.7x, 2x and 1.3x respectively in cells pre–exposed to the contact lens. Conclusions: PA stimulation of hbd–2 promoter is TLR–mediated. In addition, PA exposure increased TLR 2, 4 and 5 gene expression. This induction of expression is reduced in the presence of CLE. These data might explain the observed decrease in JNK activity and subsequent decrease in hbd–2 expression in cells co–cultured with a contact lens. They might also at least partly explain why contact lens wearers are more susceptible to serious infection by PA.
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