P29 Manuka honey downregulates IL-4-induced CCL26 via activation of the aryl hydrocarbon receptor

Abstract The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that regulates skin homeostasis. AHR activation by physiological ligands (e.g. indoles originated from host or microbial metabolism of tryptophan) leads to the expression of cytochrome P450 (CYP) enzymes, such as CYP1A1 and has anti-inflammatory effects in animal models and clinical samples of psoriasis and atopic dermatitis. Manuka honey (MH) is a complex nutritional material with antimicrobial, antioxidant and anti-inflammatory activity. We have previously shown that MH downregulates interleukin (IL)-4-induced CCL26 expression in immortalized keratinocytes. Recent studies suggest that MH contains potential AhR ligands, including indoles. Thus, we hypothesize that its anti-inflammatory effect may be mediated, at least partially, via AhR activation. Here, we treated HaCaT keratinocytes, either stably transfected with an empty vector (EV-HaCaT) or in which AhR had been stably silenced (AhR-silenced HaCaT), or primary normal human epithelial keratinocytes (NHEK) with 2% MH for 24 h and measured gene expression by reverse transcriptase quantitative polymerase chain reaction. We detected a 15.4-fold upregulation of CYP1A1 in EV-HaCaTs, which was significantly reduced (P ≤ 0.001) in AhR-silenced cells. Pretreatment with the AhR antagonist CH223191 significantly (P ≤ 0.0001) abrogated this effect. Similar findings were observed in NHEKs. In vivo epicutaneous treatment of Cyp1a1Cre × R26ReYFP reporter mice with pure MH significantly (P < 0.0001) induced CYP1A1 expression compared with Vaseline. Treatment of HaCaT with 2% MH significantly decreased (P < 0.0001) baseline CYP1A1 enzymatic activity at 3 and 6 h but increased it after 12 h, suggesting that MH may activate the AhR through both direct and indirect means. Importantly, MH downregulation of IL-4-induced CCL26 mRNA and protein was abrogated in AhR-silenced HaCaTs and by pretreatment with CH223191. Finally, MH significantly (P < 0.0001) upregulated filaggrin (FLG) expression in NHEKs in an AhR-dependent manner. In conclusion, MH activates AhR, both in vitro and in vivo, thereby providing a mechanism of its IL-4-induced CCL26 downregulation and upregulation of FLG expression. Taken together, our data show that the AhR-activating properties of MH could have broader clinical implications in a range of skin inflammatory conditions.