191 Hair follicle-derived keratinocytes as a useful tool to study atopic dermatitis-related TH2 cytokine responses

Abstract

In vitro models based on healthy keratinocytes stimulated with TH2 cytokines are often used as a tool to study atopic dermatitis (AD)-related inflammatory responses. It seems favorable to directly use AD-derived keratinocytes without the need of invasive methods for isolation. To this end, we established the isolation and propagation of keratinocytes derived from hair follicles. To evaluate if such AD-derived keratinocytes represent a good model system to mimic AD inflammatory responses we compared 2D cultures of keratinocytes derived from healthy donors with cells derived from AD patients. Cells were stimulated with IL-4, IL-13 and IL-4+IL-13 to simulate and evaluate a TH2 cytokine response as typically seen in AD. The expression of IL-13 is known to be much higher than IL-4 in AD skin. In this model we therefore used 10-fold higher concentrations of IL-13 as compared to IL-4 because expression analyses indicate at least a 10-fold difference in cutaneous expression levels of IL-13 as compared to IL-4. We observed a higher expression change of AD-relevant factors (e.g. TSLP, CCL26, IL-33, CA2) in the AD-derived keratinocytes in response to TH2 cytokines than in healthy keratinocytes. Stimulation of the cells with both IL-4 and IL-13 generally had the highest effect on expression changes, but importantly, stimulation with IL-13 alone was sufficient to induce significant changes of AD-specific markers. To further study AD-associated epidermal inflammatory responses we established the generation of 3D skin models using AD-derived hair keratinocytes under stimulation with TH2 cytokines. Histologically, stimulation with IL-4 and IL-13 increased epidermal spongiosis, a typical hallmark of AD skin. Moreover, IL-4 and IL-13 induced a strong expression of several AD-relevant factors in the 3D models. Taken together, our data identified AD-keratinocytes, derived from hair follicles, as a beneficial tool to study AD-related inflammatory responses in vitro.