Immune Functions of Epidermal Langerhans Cells in vivo

Epidermal langerhans cells (LC) have long been regarded as prototypic antigen presenting cells (APC) for cutaneous immunity. However, recent studies suggest a predominant role of dermal dendritic cells (dDC), whereas LC seem to exert immunosuppressive functions. However, much of these work is based on LC ablation models, and conclusions drawn were, therefore, necessarily indirect in nature. The proposed project aims at investigating LC functions under conditions that all APC subsets are present in their natural physiological state. To this end, we will generate mouse models for the inducible expression of antigens exclusively in LC. We will take advantage of a recently developed LangCreER T2 transgenic mouse strain. These mice will be crossed with floxed-stop reporter mice that have an open reading frame for a model antigen separated from a constitutively active promoter by a loxP site-flanked stop cassette. Administration of tamoxifen (TAM) will cause nuclear translocation of Cre that is expressed selectively in LC and, hence, turn on antigen expression in these cells. Transgenic mice will be examined for antigen-specific T cell and antibody responses and/or resistance to subsequent antigen challenge. TAM will be administrated solely or in combination with molecular danger signals, e.g. toll-like receptor ligands or skin irritants, to examine antigen presentation by LC under steady state or inflammatory conditions, respectively. TAM treatment, around birth or in adult mice will allow for analyzing the same proteins, either, as "self" antigens present throughout life or as "non-self" antigens expressed later when the immune system has been fully developed.

Here we demonstrate that the activation state of epidermal Langerhans cells (LCs) determines the fate of cytotoxic T cells. Langerhans cells are the only antigen-presenting cells (APC) in the epidermis. However, the presence of multiple dendritic cells (DCs) with APC properties in the subjacent dermis has complicated the analysis of these cells functions. In order to assess LC function in vivo, we have generated novel transgenic mouse models in which antigen presentation is limited to LCs. In these mice, different model- antigens, including hen egg Ovalbumin (OVA) can be inducibly expressed in LCs but no other DC subsets. When OVA, as a LC-borne neo-antigen, was presented by LCs in the absence of danger signals, transgenic mice were resistant to cutaneous immunization with OVA, suggesting the induction of tolerance. This tolerance was confirmed to be antigen- specific and dependent on regulatory T cells (Tregs), as Treg depletion reversed tolerance, allowing activation and expansion of antigen-specific cytotoxic T cells (CTLs). By contrast, when OVA was presented by LCs in the presence of microbial danger signals, OVA-specific CTLs were highly activated and developed a memory phenotype which was recallable by a cutaneous OVA challenge. Langerhans cells are located in a very prominent place, the outermost skin layer and are in permanent contact with an enormous number of environmental pathogens but also harmless microbial skin inhabitants. Thus, LCs are in the first place to either activate the immune system to fight against dangerous pathogens or to inhibit/suppress immune responses against commensals or harmless skin neo-proteins. With these novel transgenic mice we were able, for the first time, to imitate both situations to switch on a neo-antigen without danger signal (mimicry of an innocuous skin-borne protein) or with danger signals (mimicry of an infection). Importantly, as in this mouse neo-antigen expression was confined to LCs, it provides a valuable genetic tool to decipher LC functions in the presence of all other DC subsets in the skin. This knowledge has profound implication for the design of new vaccines and vaccination strategies, because the skin is also an attractive tissue for drug and vaccine delivery.