| Purpose: Toll-like receptors (TLRs) are pattern recognition receptors that serve a critical function in detecting pathogens and initiating immune responses. Activation of innate immunity through TLR4 on antigen presenting cells (APC) promotes APC anti- microbial activity and induction of antigen-specific T helper-1 (Th1) responses. The purpose of this study was to examine the TLR-4 expression patterns at the level of protein in normal and inflamed murine cornea, and to explore their relation with resident corneal APC.
Methods: Normal and inflamed corneas of BALB/c and C57BL/6 were excised and immunofluorescence single- and double-staining with multiple antibodies was performed by confocal microscopy on whole-mounted corneal stromas. Additionally, Corneal buttons were placed in culture to harvest cells and evaluate for protein cell surface expression by immunocytochemistry and flow cytometry.
Results: Numerous TLR4+ cells with dendriform morphology were found in the cornea tissue, with significant concentration of these cells in the periphery as compared to the central areas of the cornea in both BALB/c and C57BL/6 mice. Dual- color immunostaining demonstrated that 100% of the CD11b+ cells (monocytic) cells in the stroma also co-expressed TLR4 in BALB/c, whereas only a negligible number of CD11b+ cells expressed TLR4 in C57BL/6. However, inflammation led to significant upregulation in expression of TLR4 in the central areas of both BALB/c and C57BL/6 mice as compared to their normal corneas. These in vivo findings were confirmed by immunocytochemistry and flow cytometric analyses of cells derived from corneal explants.
Conclusions: These preliminary data demonstrate for the first time, at the level of protein, that murine corneal monocytic cells are capable of expressing TLR4, and that the expression pattern of this critical TLR varies depending on the microvironment (e.g. inflammatory state) of the cornea as well as the host murine strain. These data suggest that alterations in TLR4 expression may be relevant in corneal immune and inflammatory responses.
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