Identification of TLR patterns and TLR-mediated epithelial protection mechanisms during localised candidosis
Final Report Abstract
The oral mucosa is an important site of microbe host interactions and the keratinocytes may participate in the host defence by expression of TLR. There are conflicting result reports in the literature for constitutive expression of the TLR family on keratinocytes. In summary recently previous studies detected transcripts for TLR1-5 and TLR9 in human keratinocytes. In the present study we were able to enlarge the epithelial TLR expression profile by demonstrating constitutive epithelial mRNA expression of TLR1-6 and TLR8-10. Confirming previously published studies we detected TLR2, TLR4, TLR6 and TLR8-10 mRNA in unstimulated human PMNs and in addition we found transcripts for TLR1, TLR5 and TLR7. These results suggest an important role of these pattern recognition receptors in infectious and inflammatory mucosal diseases. Although there is clear evidence for the expression of TLR by keratinocytes there is a lack of studies investigating TLR mRNA expression upon stimulation with C. albicans. Heat killed C. albicans cells failed to modulate epithelial TLR expression. Also in the present study viable C. albicans cells were unable to stimulate epithelial TLR expression although inducing clear signs of mucosal infection and a strong cytokine response. These results suggest that TLR expression is not required for stimulation of a C. albicans induced epithelial cytokine response. The lack of TLR expression may be explained by the fact that C. albicans is a harmless colonizer of mucosal surfaces in healthy individuals. During the period of colonization, extensive fungal growth is limited through release of antimicrobial peptides from keratinocytes, or due to existence of other bacteria of the microbial flora. In this stage of colonization without clinical symptoms and signs of inflammation neither for the facultative pathogen nor for the host a TLR mediated inflammatory cytokine response might be of advantage. In contrast, we found a strong stimulation of epithelial TLR4 mRNA expression induced by C. albicans in the presence of PMNs. Confocal laser and electron microscopy confirmed epithelial TLR4 stimulation on the protein level and demonstrated increased cytoplasmic expression only by keratinocytes directly in contact with the pathogens. Interestingly the effect was not necessarily connected to the direct interaction of PMNs with C. albicans or the infected RHE. TLR4 upregulation was associated with protection against Candida induced tissue damage and could be reversed by blocking with anti-TLR4 antibodies. PMNs are frequently found at sites of Candida infections but not during colonization, and are thought to be a hallmark of oral candidosis in humans. Therefore it is tempting to speculate that infection of the epithelium induces a not TLR mediated chemo-attractive immune response for PMNs. In the presence of PMNs an immunological cross-talk between infected epithelial cells and PMNs via e.g. cytokines stimulates a strong epithelial TLR4 immune response which may be of prime importance in initiating a protective anti-candidal effect.
