No abstract

Atopic dermatitis (AD) is an itchy, chronic form of eczema with multiple pathogenic factors, including skin barrier dysfunction and immune response dysregulation. Type 2 inflammation, characterized by the accumulation of Th2 cells, ILC2, and type 2 cytokines (interleukin (IL)-4, IL-5, and IL-13), plays a crucial role in the pathophysiology of AD, although other cytokines derived from Th1, Th17 and Th22 are also involved in the formation of AD skin lesions. 1,2 Skin barrier dysfunction is also linked to inflammation of the skin and vice versa. Skin barrier dysfunction enhances transepidermal allergen exposure and triggers type 2 inflammation. The type 2 cytokines IL-4, IL-13, IL-31, and IL-22, which are produced by Th22, decrease the production of filaggrin (FLG), loricrin, and involucrin in epidermal keratinocytes, resulting in skin barrier dysfunction.

Staphylococcus aureus (S. aureus) is frequently detected in patients with atopic dermatitis (AD). Decreased diversity of bacterial flora on the skin surface (dysbiosis) has been considered as an important pathogenic factor of AD. We previously demonstrated that the skin immune responses induced by AD skin-derived strains of S. aureus (AD strain) was altered, compared to those by standard strains of S. aureus. However, detailed mechanisms have not been elucidated.To investigate how the colonization of S. aureus is controlled by the skin barrier and cytokine milieu, we quantitated the number of AD strain internalized into keratinocyte (KC, HaCaT). Fluorescent labeled S. aureus was detected using a cell imaging system (Opera Phenix TM ), KC was stimulated by heat-inactivated AD strain or standard strain, with or without Th1 cytokine (IFN-γ, 100ng/ml) and Th2 cytokines (IL-4 and IL-13, 10ng/ml respectively) for 24hr.Time course study revealed that only AD strain could adhere to KC within 15 minutes, and was rapidly internalized into KC within 3 hr. The number of AD strain internalized to KC was significantly higher than that of standard strain. The treatment with Th1 cytokine significantly decreased the number of AD strain internalized into KC, while Th2 cytokines exerted no effect on the internalization. When filaggrin (FLG) in KC was knocked down by using specific siRNA, AD strain was significantly accumulated in KC. These characters of AD strain may promote the long-term retention of S. aureus in AD skin.

Atopic dermatitis (AD) is a common and multifactorial skin disease. Genetically determined skin barrier defects and a dysregulated immune system lead to chronic inflammation. It is known that atopic and healthy skin differ in bacterial colonization. Atopic skin is associated with a lower microbiota diversity and higher amounts of potentially pathogenic bacteria like Staphylococcus aureus. However, it is still subject of debate whether this different microbiota composition is mainly a result or a cause of the inflammatory state in AD. To investigate this question further, we started to analyze the influence of the skin microbiota of healthy individuals and AD patients on a 3D skin equivalent representing healthy skin. To this end, defined skin areas were rinsed with a saline/detergent solution to harvest the microbiota. Subsequently, the rinsing solution was sequentially centrifuged and stored at-80 C in a glycerol-pellet. These microbiota samples were then applied to the surface of a 3D skin equivalent. After stimulation, gene expression of inflammatory mediators, barrier and defense molecules was analyzed by real-time PCR. The microbiota of AD patients was compared with localization-, gender-and age-matched microbiota samples of healthy control persons. As a general result, we observed that the microbiota-treatment of the 3D skin equivalents induced the expression of several inflammatory and defense mediators (e.g. cytokines, antimicrobial peptides). The microbiota derived from lesional AD-skin induced higher levels of several inflammatory mediators as compared to the microbiota of healthy skin or non-lesional AD skin. Especially the expression of the AD-associated inflammatory mediator thymic stromal lymphopoietin (TSLP) was markedly increased by the microbiota derived from lesional AD skin. These findings indicate that the altered skin microbiota in AD may amplify the inflammation process through the induction of host inflammatory mediators such as TSLP.