Flagellin Induces the Expression of Thymic Stromal Lymphopoietin in Human Keratinocytes via Toll-Like Receptor 5

Abstract

Background: Thymic stromal lymphopoietin (TSLP), highly expressed by keratinocytes in skin lesions of atopic dermatitis patients and bronchial epithelial cells in asthma, plays a key role in allergic diseases. Information on triggers for the release of TSLP in keratinocytes is still limited. Keratinocytes express Toll-like receptor (TLR) 5, the ligand for which is flagellin, the major structural protein of the flagella of Gram-negative bacteria. IL-4, IL-13 and TNF-α (Th2/TNF) are associated with allergic diseases. TGF-α, one of the ligands for the epidermal growth factor receptor, is overexpressed in keratinocytes in atopic dermatitis. We investigated the induction of TSLP expression in keratinocytes stimulated with flagellin and its modulation by the Th2/TNF cytokines and TGF-α. Methods: Primary human keratinocytes were stimulated with flagellin with or without cytokines. The TSLP released was measured by ELISA. Gene expression was analyzed by quantitative real-time PCR. Results: Stimulation of keratinocytes with flagellin induced the release of TSLP protein and upregulation of the gene expression of TSLP and other pro-inflammatory molecules. The flagellin-induced release of TSLP was enhanced by the Th2/TNF cytokines or TGF-α. Small interfering RNA-mediated knockdown of TLR5 expression suppressed the flagellin-induced TSLP gene expression. Conclusions: Flagellin induces TSLP expression in keratinocytes via TLR5 and the expression can be upregulated by a cytokine milieu with Th2/TNF or TGF-α, suggesting that exposure of barrier-defective skin to Gram-negative bacteria or environmental flagellin contributes to the initiation and/or amplification of Th2-type skin inflammation including atopic dermatitis through the induction of TSLP expression in keratinocytes.

Recent Advances in Mucosal Immunology and Ocular Surface Diseases

“…TLR-mediated TSLP induction has been recognized [39,53,54]. We have demonstrated that TSLP was largely induced by specific TLR ligands in human corneal epithelial cells [40].…”

“…Substantial evidence now indicates that epithelial cells are central participants in innate and adaptive immune responses [4,5,7]. Based on the observation that we and other groups have made [40,53,54] that TSLP is mainly induced via TLR mediated innate response in epithelia exposed to microbial products, we hypothesized that pollen, such as Ambrosia artemisiifolia short ragweed (SRW), the most widespread plant in North America, may serve as a functional TLR4 agonist that induces production of a proallergic cytokine TSLP via innate immune response to trigger Th2-dominant allergic inflammation. To uncover the novel phenomenon and molecular signaling pathways involved in pollen induced allergic inflammation, a comprehensive set of experiments has been conducted using a wellcharacterized murine model of allergic conjunctivitis induced by SRW pollen in BALB/c, TLR4 deficient and MyD88 knockout mice, as well as a murine topical ocular surface challenge model and a culture model of primary human corneal epithelial cells exposed to an aqueous extract of defatted SRW pollen.…”

Flagellin suppresses experimental asthma by generating regulatory dendritic cells and T cells

“…Recent studies have shown contradictory results on the effect of flagellin, 10,33 suggesting that flagellin can promote asthma or atopic dermatitis. However, both of these studies 10,33 are based on experiments in which flagellin doses were 1/50th to 1/1000th of those used in this study. We recapitulated the asthma-promoting activity of lower doses of flagellin in our system.…”

Immunologic Targets in Atopic Dermatitis and Emerging Therapies: An Update

“…One might postulate that this aspect of an innate immune response may help prevent persistent TLR2 activation and ensuing inflammation because TJs are present in the superficial layers of the epidermis, whereas TLR2 is expressed below these layers [28]. Interestingly, TLR stimulation can also induce keratinocyte production of thymic stromal lymphopoietin (TSLP), an important cytokine in priming the Type 2 adaptive immune response [40][41][42].…”