Daniel H. Kaplan

Skin Immunity to Candida albicans.

Candida albicans is a dimorphic commensal fungus that colonizes healthy human skin, mucosa, and the reproductive tract. C. albicans is also a predominantly opportunistic fungal pathogen, leading to disease manifestations such as disseminated candidiasis and chronic mucocutaneous candidiasis (CMC). The differing host susceptibilities for the sites of C. albicans infection have revealed tissue compartmentalization with tailoring of immune responses based on the site of infection. Furthermore, extensive studies of host genetics in rare cases of CMC have identified conserved genetic pathways involved in immune recognition and the response to the extracellular pathogen. We focus here on human and mouse skin as a site of C. albicans infection, and we review established and newly discovered insights into the cellular pathways that promote cutaneous antifungal immunity.

CD301b+ dendritic cells suppress T follicular helper cells and antibody responses to protein antigens

Strong antibody response is considered a hallmark of a successful vaccine. While dendritic cells (DCs) are important for T follicular helper (Tfh) cell priming, how this process is regulated in vivo is unclear. We show here that the depletion of CD301b+ DCs specifically enhanced the development of Tfh cells, germinal center B cells and antibody responses against protein antigens. Exaggerated antibody responses in mice depleted of CD301b+ DCs occurred in the absence of any adjuvants, and resulting antibodies had broader specificity and higher affinity to the immunogen. CD301b+ DCs express high levels of PD-1 ligands, PD-L1 and PD-L2. Blocking PD-1 or PD-L1 during priming in wild-type mice partially mimicked the phenotype of CD301b+ DC-depleted animals, suggesting their role in Tfh suppression. Transient depletion of CD301b+ DC results in the generation of autoreactive IgG responses. These results revealed a novel regulatory mechanism and a key role of CD301b+ DCs in blocking autoantibody generation. DOI: http://dx.doi.org/10.7554/eLife.17979.001

Neoantigen Expression in Steady-State Langerhans Cells Induces CTL Tolerance.

The skin hosts a variety of dendritic cells (DCs), which act as professional APC to control cutaneous immunity. Langerhans cells (LCs) are the only DC subset in the healthy epidermis. However, due to the complexity of the skin DC network, their relative contribution to either immune activation or immune tolerance is still not entirely understood. To specifically study the function of LCs in vivo, without altering the DC subset composition in the skin, we have generated transgenic mouse models for tamoxifen-inducible de novo expression of Ags in LCs but no other langerin+ DCs. Therefore, this system allows for LC-restricted Ag presentation to T cells. Presentation of nonsecreted OVA (GFPOVA) by steady-state LCs resulted in transient activation of endogenous CTL in transgenic mice. However, when these mice were challenged with OVA by gene gun immunization in the contraction phase of the primary CTL response they did not respond with a recall of CTL memory but, instead, with robust Ag-specific CTL tolerance. We found regulatory T cells (Tregs) enriched in the skin of tolerized mice, and depletion of Tregs or adoptive experiments revealed that Tregs were critically involved in CTL tolerance. By contrast, when OVA was presented by activated LCs, a recallable CTL memory response developed in transgenic mice. Thus, neoantigen presentation by epidermal LCs results in either robust CTL tolerance or CTL memory, and this decision-making depends on the activation state of the presenting LCs.

A protective Langerhans cell–keratinocyte axis that is dysfunctional in photosensitivity

Langerhans cells limit ultraviolet radiation–induced keratinocyte apoptosis and skin injury, and this axis is dysfunctional in lupus photosensitivity. Abrogating keratinocyte apoptosis in lupus Lupus patients can experience photosensitivity as manifested by joint pain, fatigue, and skin rashes. To better understand the mechanisms behind this photosensitivity, Shipman et al. examined patient skin samples and mouse models of lupus. They saw that, in healthy mice and human skin exposed to ultraviolet radiation, Langerhans cells protected keratinocytes from apoptosis through a mechanism involving the metalloproteinase ADAM17 and EGFR ligands. In the lupus mice and patient samples, the Langerhans cells were unable to do this. Stimulating EGFR in the skin may bring relief to photosensitive lupus patients. Photosensitivity, or skin sensitivity to ultraviolet radiation (UVR), is a feature of lupus erythematosus and other autoimmune and dermatologic conditions, but the mechanistic underpinnings are poorly understood. We identify a Langerhans cell (LC)–keratinocyte axis that limits UVR-induced keratinocyte apoptosis and skin injury via keratinocyte epidermal growth factor receptor (EGFR) stimulation. We show that the absence of LCs in Langerin–diphtheria toxin subunit A (DTA) mice leads to photosensitivity and that, in vitro, mouse and human LCs can directly protect keratinocytes from UVR-induced apoptosis. LCs express EGFR ligands and a disintegrin and metalloprotease 17 (ADAM17), the metalloprotease that activates EGFR ligands. Deletion of ADAM17 from LCs leads to photosensitivity, and UVR induces LC ADAM17 activation and generation of soluble active EGFR ligands, suggesting that LCs protect by providing activated EGFR ligands to keratinocytes. Photosensitive systemic lupus erythematosus (SLE) models and human SLE skin show reduced epidermal EGFR phosphorylation and LC defects, and a topical EGFR ligand reduces photosensitivity. Together, our data establish a direct tissue-protective function for LCs, reveal a mechanistic basis for photosensitivity, and suggest EGFR stimulation as a treatment for photosensitivity in lupus erythematosus and potentially other autoimmune and dermatologic conditions.

Langerhans Cells Prevent Autoimmunity via Expansion of Keratinocyte Antigen-Specific Regulatory T Cells

Langerhans cells (LCs) are antigen-presenting cells in the epidermis whose roles in antigen-specific immune regulation remain incompletely understood. Desmoglein 3 (Dsg3) is a keratinocyte cell-cell adhesion molecule critical for epidermal integrity and an autoantigen in the autoimmune blistering disease pemphigus. Although antibody-mediated disease mechanisms in pemphigus are extensively characterized, the T cell aspect of this autoimmune disease still remains poorly understood. Herein, we utilized a mouse model of CD4+ T cell-mediated autoimmunity against Dsg3 to show that acquisition of Dsg3 and subsequent presentation to T cells by LCs depended on the C-type lectin langerin. The lack of LCs led to enhanced autoimmunity with impaired Dsg3-specific regulatory T cell expansion. LCs expressed the IL-2 receptor complex and the disruption of IL-2 signaling in LCs attenuated LC-mediated regulatory T cell expansion in vitro, demonstrating that direct IL-2 signaling shapes LC function. These data establish that LCs mediate peripheral tolerance against an epidermal autoantigen and point to langerin and IL-2 signaling pathways as attractive targets for achieving tolerogenic responses particularly in autoimmune blistering diseases such as pemphigus.

Isolation of Murine Skin Resident and Migratory Dendritic Cells via Enzymatic Digestion

Dendritic cells (DCs) are a highly specialized subset of professional antigen‐presenting cells (APCs) that reside in peripheral and lymphoid tissues. DCs capture antigen in the periphery and migrate to the lymph node where they prime naïve T cells. In addition, DCs have been recently appreciated to have function in innate immunity within tissues. In the skin, heterogeneous populations of DCs reside within the epidermis and the dermis. Analysis of the cutaneous DC subsets is complicated by requirements of distinct enzymatic digestion protocols for isolation of APCs from distinct anatomical compartments of the skin. Here, specific approaches for isolation of DCs from the epidermis, dermis, and the skin‐draining lymph nodes of mice are described.

Wall teichoic acid is a pathogen-associated molecular pattern of Staphylococcus aureus that is recognized by langerin (CD207) on skin Langerhans cells

Staphylococcus aureus is a major cause of skin and soft tissue infections and aggravator of the inflammatory skin disease atopic dermatitis (AD). Langerhans cells (LCs) initiate a Th17 response upon exposure to S. aureus, which contributes to host defense but also to AD pathogenesis. However, the molecular mechanisms underlying the unique pro-inflammatory capacities of S. aureus remain unclear. We demonstrate that human LCs directly interact with S. aureus through the pattern-recognition receptor langerin (CD207), which specifically recognizes the conserved β-N-acetylglucosamine (GlcNAc) modifications of wall teichoic acid (WTA) that are not expressed by other staphylococcal species. The WTA glycoprofile strongly influences the production of Th1- and Th17-polarizing cytokines by LCs. Specifically, β-GlcNAc activates LCs, whereas co-decoration of WTA with α-GlcNAc through the enzyme TarM, uniformly present in the AD-associated CC1 lineage, attenuates LC immune activation. Our findings provide important mechanistic insights into the role of S. aureus in inflammatory skin disease.Abstract Staphylococcus aureus is a major cause of skin and soft tissue infections and aggravator of the inflammatory skin disease atopic dermatitis (AD). Epicutaneous exposure to S. aureus induces Th17 responses through skin Langerhans cells (LCs), which paradoxically contribute to host defense but also to AD pathogenesis. The underlying molecular mechanisms of the association between S. aureus and skin inflammation are poorly understood. Here, we demonstrate that human LCs directly interact with S. aureus through the pattern-recognition receptor langerin (CD207). Human, but not mouse, langerin interacts with S. aureus through the conserved β-N-acetylglucosamine (GlcNAc) modifications on wall teichoic acid (WTA), thereby discriminating S. aureus from other staphylococcal species. Importantly, the specific S. aureus WTA glycoprofile strongly influences the level of Th1-and Th17-polarizing cytokines that are produced by in vitro generated LCs. Finally, in a murine epicutaneous infection model, S. aureus induced a more pronounced influx of inflammatory cells and pro-inflammatory cytokine transcripts in skin of human langerin transgenic mice compared to wild-type mice. Our findings provide molecular insight into the unique pro-inflammatory capacities of S. aureus in relation to inflammatory skin disease.

Cutaneous Dendritic Cells in Health and Disease

Dendritic cells (DCs) are a heterogeneous cell type found in lymphatic and peripheral tissues. They function as professional antigen-presenting cells (APCs), specialized to acquire antigen from their environment that they process and present to T cells. In addition, DC participate in the generation of local inflammation and suppression of inappropriate immune responses. Thus, they play a critical role in the initiation, propagation and suppression of immune responses that promote health and autoimmune disease. The skin of humans and mice contain several distinct subsets of DC (epidermal Langerhans cells are the most well-known) that are speculated to have varied and versatile functions. In this chapter, we discuss dendritic cells as important players of the innate and adaptive immune system. We will cover their roles in acquiring antigen, inducing T cell responses, and characterizing the functional differences between distinct skin DC subsets. Finally, we will relate DC in relation to human health including their contribution to diseases such as allergic contact dermatitis and psoriasis and their ability to serve as therapeutic targets in vaccinations.

Generation of a NK1R-CreER knockin mouse strain to study cells involved in Neurokinin 1 Receptor signaling.

The Neurokinin 1 Receptor (NK1R), which binds Substance P, is expressed in discrete populations of neurons throughout the nervous system, where it has numerous roles including the modulation of pain and affective behaviors. Here, we report the generation of a NK1R‐CreER knockin allele, in which CreERT2 replaces the coding sequence of the TACR1 gene (encoding NK1R) in order to gain genetic access to these cells. We find that the NK1R‐CreER allele mediates recombination in many regions of the nervous system that are important in pain and anxiety including the amygdala, hypothalamus, frontal cortex, raphe nucleus, and dorsal horn of the spinal cord. Other cell types that are labeled by this allele include amacrine cells in the retina and fibroblasts in the skin. Thus, the NK1R‐CreER mouse line is a valuable new tool for conditional gene manipulation enabling the visualization and manipulation of cells that express NK1R.