Activation of human dendritic cells : Role of the allergenic yeast Malassezia and NK cells

University dissertation from Stockholm : Karolinska Institutet, Department of Medicine

Abstract: Immature bone marrow derived dendritic cells (DCs) patrol the periphery of our body, where they encounter and take up foreign antigens. Once activated, DCs migrate to regional lymph nodes to present peptides derived from these antigens to naive T cells, and elicit different T helper (Th) cell responses. One of the keys to understanding DC function is the investigation of their activation and maturation, and the process by which activated DCs can instruct the adaptive immune system to use appropriate defense mechanisms. Few studies have so far focused on the effect of yeast on DCs. The yeast Malassezia is part of the normal flora of our skin, but can act as an allergen and elicit specific IgE antibodies and T cell reactivity in patients with atopic eczema/dermatitis syndrome (AEDS). AEDS is a chronic inflammatory skin disease that is increasing in prevalence worldwide. Though, the mechanisms that determine whether an exposed individual becomes sensitized are still poorly understood. The interaction between genetic predisposition and environmental factors seems to be crucial. Little is also known about the modulation of DC function by other cells. Only recently it became evident that NK cells might play a role in affecting DC maturation and function. It is not known, however, if this interaction takes place in vivo, and if NK-DC interaction is affected by allergen exposure. We showed, by flow cytometry and confocal microscopy, that immature human DCs bind and internalize Malassezia yeast cells and allergenic components from the yeast. This ability of the DCs is lost upon maturation. Furthermore, we showed a mannose receptor-mediated uptake of some yeast components. Using time-lapse photography we followed the kinetics of the uptake of Malassezia, and found that the yeast is taken up within 1 hour. Malassezia induces DC maturation (expression of CD80, CD83 and CD86), and production of cytokines like TNF-alpha and IL1beta, but not production of the Th1-inducing cytokine IL-12. Production of IL-18 increases after interaction with Malassezia, a phenomenon that might contribute to Th2 responses seen in AEDS patients. Finally, functional studies demonstrated that DCs pre-incubated with Malassezia induce proliferation in autologous peripheral blood mononuclear cells. Next we studied the distribution of NK cells in the skin of AEDS patients with particular emphasis on possible NK cell-DC interactions. Confocal microscopy revealed a few scattered NK (CD56+/CD3-) cells in the dermis of healthy individuals and in non-lesional skin from AEDS patients. However, NK cells were found to be differently distributed in lesional and Malassezia atopy patch test-positive skin, and for the first time, NK cell-DC contact in vivo was shown. By in vitro studies, we showed that DCs pre-stimulated with Malassezia are less susceptible to NK cell-induced cell death (51Cr release assay), suggesting that Malassezia affect the interaction through induction of DC maturation and production of soluble factors. Additionally, the results imply that the yeast has a direct effect on the NK cell cytotoxicity. We also showed that NK cells induce CD48 expression on DCs. The proportion of CD48+ cells increases in immature DCs, and even more so in those stimulated with Malassezia extract. The increased expression of CD48 on the DCs might be important for their ability to stimulate T cells, Besides, after contact with NK cells, the expression of CD83 and CD86 increases significantly in the Malassezia extract-stimulated DCs. IL-8 was shown to be induced in the NK cell-DC co-cultures. Taken together, the uptake of Malassezia and allergenic components from the yeast in the absence of IgE, imply that sensitization to Malassezia can be mediated by immature DCs in the skin. After interaction with Malassezia yeast cells, DCs become functionally mature and produce cytokines that might give rise to a Th2-like immune response. Our findings indicate that NK cells and DCs can interact in the skin, suggesting that NK cells may play a role in regulating DCs in AEDS. Malassezia yeast cells affect the NK cell-DC interaction, e.g. by decreasing NK cell cytotoxicity. The increase of CD48+, CD83+ and CD86+ DCs and induction of IL-8 production imply that NK cells activate the DCs. Furthermore, the results suggest a function of NK cell-DC interaction in shaping immune responses. Better knowledge of the activation and modulation of DCs and their function might give greater understanding of the immunological phenomenon behind the onset of AEDS. Ultimately, studies on initiation of immune responses might lead to new strategies for prevention and treatment of conditions like AEDS.

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