Contact allergen (PPD and DNCB)-induced keratinocyte sensitization is partly mediated through a low molecular weight hyaluronan (LMWHA)/TLR4/NF-κB signaling axis

Allergic contact dermatitis (ACD) is caused by topical exposure to chemical allergens. Keratinocytes play a key role in innate immunity, as well as in ACD progression. The transmembrane Toll-like receptor 4 (TLR4), strongly implicated in skin inflammation, has the ability to bind Damage Associated Molecular Patterns (DAMPs), like Low Molecular Weight Hyaluronan (LMWHA). Previously, we had determined that p-phenylenediamine (PPD) and 2,4-dinitrochlorobenzene (DNCB) modulate keratinocyte HA deposition in a manner correlated to their sensitization. In the present study, we aimed to investigate putative co-operation of HA and TLR4 in the process of PPD and DNCB-induced keratinocyte activation. Contact sensitizers were shown to significantly increase the expression of Hyaluronan Synthases (HAS) and TLR4 in NCTC2544 human keratinocytes, as demonstrated by western blot and Real-Time PCR. These data, in correlation to earlier shown enhanced HA degradation suggest that the contact sensitizers facilitate HA turnover of keratinocytes and increase the release of pro-inflammatory, LMWHA fragments. Treatment with exogenous LMWHA enhanced TLR4, HAS levels and Nuclear factor-kappa beta (NF-\u3ba\u392) activation. PPD, DNCB and LMWHA-effects were shown to be partly executed through TLR4 downstream signaling as shown by Real-Time, western blot, siRNA and confocal microscopy approaches. Specifically, PPD and DNCB stimulated the activation of the TLR4 downstream mediator NF-\u3baB. Therefore, the shown upregulation of TLR4 expression is suggested to further facilitate the release of endogenous, bioactive HA fragments and sustain keratinocyte activation. In conclusion, keratinocyte contact allergen-dependent sensitization is partly mediated through a LMWHA/TLR4/ NF-\u3baB signaling axis