Mechanisms regulating skin immunity and inflammation.

REVIEWS Mechanisms regulating skin immunity and inflammation Manolis Pasparakis1, Ingo Haase2 and Frank O. Nestle3

Abstract | Immune responses in the skin are important for host defence against pathogenic microorganisms. However, dysregulated immune reactions can cause chronic inflammatory skin diseases. Extensive crosstalk between the different cellular and microbial components of the skin regulates local immune responses to ensure efficient host defence, to maintain and restore homeostasis, and to prevent chronic disease. In this Review, we discuss recent findings that highlight the complex regulatory networks that control skin immunity, and we provide new paradigms for the mechanisms that regulate skin immune responses in host defence and in chronic inflammation. Langerhans cells A dendritic cell population named after the German anatomist Paul Langerhans. Langerhans cells are derived from monocytes and reside in the epidermis and epithelium of hair follicles, as well as in mucosal body surfaces. They are professional antigenpresenting cells and have immune surveillance functions.

Institute for Genetics, Center for Molecular Medicine, and Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Zülpicher Strasse 47a, 50674 Cologne, Germany. 2 Department of Dermatology, University of Cologne, Kerpener Strasse 62, D-50931 Cologne, Germany. 3 St John’s Institute of Dermatology and Guy’s and St Thomas’ Hospital National Institute for Health Research Biomedical Research Centre, King’s College London, SE1 9RT London, UK. e‑mails: [email protected]; [email protected]; [email protected]. doi:10.1038/nri3646 Published online 11 April 2014 1

The skin provides a life-sustaining interface between the body and the environment. In addition to its wellcharacterized mechanical barrier function — which restricts water loss and prevents the entry of potentially harmful environmental substances and microorganisms — the skin forms an active barrier that provides the first line of immunological defence against infection1. Extensive crosstalk between epithelial, stromal and immune cells regulates immune responses in the skin to ensure effective host defence and to maintain or restore tissue homeostasis. In addition, the diverse microbial communities that colonize the surface and appendages of the skin constantly interact with host epithelial and immune cells, thereby influencing local and systemic immunity. The maintenance of immune homeostasis in the skin relies on a finely tuned equilibrium of well-regulated interactions between different cellular and microbial components. Dysregulation of this equilibrium contributes to the pathogenesis of inflammatory skin diseases such as psoriasis2. The classical approach to studying immune responses has been to focus on the individual cellular and molecular components of innate and adaptive immunity, thereby largely neglecting the contribution of the stroma to tissue immunity. However, it is now clear that understanding the mechanisms that regulate immune responses requires the study of the interactions between the different immune and non-immune cellular components of specific tissues. In addition, in barrier tissues such as the intestine and the skin, recent studies have highlighted the crucial role of the communication between host (epithelial, stromal and immune) cells and the microbiota in the regulation of immune responses.

The main challenge in the study of skin immunology is to understand the mechanisms that regulate the crosstalk between the various host and microbial cellular components of the skin, and how dysregulation of this communication contributes to altered cutaneous immune responses and to the pathogenesis of inflammatory skin diseases. In this Review, we discuss recent findings that highlight the complex regulatory mechanisms that control skin immunity. Owing to space limitations, we do not aim to comprehensively cover the field of skin immunology. Instead, we focus on recent key studies that provide new paradigms for the mechanisms that regulate skin immune responses in host defence and in the context of chronic inflammation.

Overview of skin components The skin consists of two major components: the epithelium and the connective tissue. The epithelial compartment of the skin can be subdivided into the epithelium of the hair follicle and that of the interfollicular epidermis. Little is known about the functional differences between these epithelial compartments with respect to skin immunity. The most numerous cell type present in the epithelial compartment is the keratinocyte; however, all of the epithelial zones are also colonized by nonepithelial immune cells, such as Langerhans cells and dendritic epidermal T cells (DETCs). There are speciesspecific differences in the immune cell populations that colonize the epithelium in the epidermis and hair follicles (FIG. 1). In contrast to mice, humans have a larger proportion of interfollicular epidermis relative to hair follicle epithelium. In addition, γδ T cell receptorexpressing (γδTCR +) DETCs are present in mouse

NATURE REVIEWS | IMMUNOLOGY

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REVIEWS a Mouse skin Commensal microbiota

Hair shaft

b Human skin

CD8+ T cell

Langerhans cell

Langerhans cell CD8+ T cell

Spinous cell layer

Basal keratinocyte Hair follicle

CD4+ T cell

Mast cell

Dermal DC

Basement membrane

Basal layer

Rete ridge

Sebaceous gland

ILC

γδ T cell

Lymphatic vessel

Macrophage

Epidermis

DETC

Stratum corneum Stratum granulosum

Macrophage

Dermal DC

Blood vessel

Dermis

CD4+ T cell

ILC

Adipose tissue

Figure 1 | Structure and cellular components of the skin in mice and humans. Mouse skin (panel a) has densely packed hair follicles, whereas human skin (panel b) has larger areas of interfollicular skin with sparse hair follicles. Human skin Nature Reviews | Immunology has a thicker epidermis (with more cell layers) and a thicker dermis than mouse skin, and is characterized by downward projections of the epidermal rete ridges. This feature is more prominent in psoriasis and correlates with elongation of the dermal papillae, which is known as papillomatosis. It is sometimes confused with the follicular hyperplasia that is seen in mouse models of inflammatory skin disease. The most prevalent immune cell types in human epidermis are Langerhans cells and CD8+ T cells. In mouse epidermis, there is a prominent population of Vγ5+ dendritic epidermal T cells (DETCs), which are absent from human epidermis. Human and mouse dermis are populated by macrophages, mast cells, conventional αβ T cells and a small population of innate lymphoid cells (ILCs). In mouse skin, there is an important contribution from recruited γδ T cells to skin immune surveillance and interleukin‑17 production.

Dendritic epidermal T cells (DETCs). A population of T cells present in mouse skin. DETCs express CD3 and a T cell receptor, and they derive from the fetal thymus. Following activation, DETCs can secrete large amounts of pro-inflammatory mediators, which participate in the communication between DETCs, neighbouring keratinocytes and Langerhans cells.

Fibrocytes A population of mesenchymal cells that reside in connective tissues. Fibrocytes have minimal cytoplasm and lack biochemical evidence of protein synthesis. Fibrocytes can migrate from the blood into connective tissues and have roles in wound healing and fibrotic tissue repair.

skin but are absent from human skin. It is important to take these differences into account when interpreting the inflammatory skin phenotypes observed in mouse models in relation to the mechanisms that drive the pathogenesis of human inflammatory skin diseases. The dermis is rich in extracellular matrix and contains stromal cells such as fibroblasts, fibrocytes and structural cells of the blood and lymph vessels. In addition, many different populations of myeloid and lymphoid immune cells either reside in or traffic through the dermis (FIG. 1). These cell populations are dynamic and undergo marked changes during an immune response.

Myeloid cells in skin immunity and inflammation Surprisingly little is known about the development and dynamics of the myeloid cell network in the skin. The denomination ‘myeloid cells’ is not entirely accurate, as some of the cells in this population do not originate from the bone marrow. On the basis of their location and origin, it is possible to distinguish these populations as either skin-resident myeloid cells or circulating bone marrow-derived myeloid cells that enter the skin from

the blood. However, these populations are not mutually exclusive as there is evidence that tissue-resident cells can be replenished by their circulating counterparts. In this Review, we discuss recent studies that have provided new insights into the role of skin epidermal and dermal myeloid cell populations in skin immunity and inflammation in mice and humans. Dendritic cell subsets and functions in mouse skin. It was previously thought that Langerhans cells — the dendritic cell (DC) population that seeds the epidermis before birth — are exclusively of bone marrow origin3. However, protein expression analysis and lineagetracing experiments have revealed that mouse Langerhans cells derive from two different embryonic sources under conditions of tissue homeostasis: the fetal liver and, to a minor extent, the yolk sac4. Langerhans cell maintenance in the epidermis depends on signalling through colony-stimulating factor 1 receptor (CSF1R)3. Macrophage colony-stimulating factor (M-CSF) contributes to the repopulation of the epidermis with Langerhans cells under inflammatory conditions but

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REVIEWS

Activator protein‑1 (AP‑1). A heterodimeric transcription factor composed of several different subunits of the FOS, JUN, ATF and JUN-dimerization protein families. AP‑1 regulates gene expression in response to cytokines, growth factors and infectious agents, and controls basic cellular processes such as proliferation, differentiation and apoptosis.

Motheaten phenotype A mouse phenotype caused by homozygous mutation of Ptpn6, the gene encoding SH2 domain-containing protein tyrosine phosphatase 1. It is a commonly used model of autoimmune and inflammatory disease. Motheaten mice develop chronic inflammation of the skin, produce autoantibodies and eventually succumb to lethal inflammation of the lungs.

is dispensable for the seeding of Langerhans cells in the epidermis during development. Recently, it was found that epidermal keratinocytes produce an alternative ligand for CSF1R, interleukin‑34 (IL‑34), and that this ligand was required for the migration of Langerhans cells to the epidermis during development 5,6. In addition, hair follicle keratinocytes in mice were shown to regulate Langerhans cell recruitment to the epidermis in response to stress and inflammation by expressing chemokines that either induce (CC‑chemokine ligand 2 (CCL2) and CCL20) or inhibit (CCL8) the recruitment of Langerhans cells and their precursors, which express the respective receptors (CC‑chemokine receptor 2 (CCR2), CCR6 and CCR8)7. These experiments identified hair follicle keratinocytes as key mediators of stress-induced Langerhans cell recruitment and provided evidence that hair follicles are important portals for the entry of Langerhans cells into the epidermis in adult mice. In mice, there are two DC subsets that reside in the dermis (and also in other peripheral tissues): CD103 (also known as integrin-αE)-positive DCs, which are equivalent to CD8α + migratory lymph node DCs; and CD11b (also known as integrin-αM)‑positive DCs8. The activator protein‑1 (AP‑1) family transcription factor basic leucine zipper transcriptional factor ATF-like 3 (BATF3) controls the later stages of CD8α+ DC development; however, an alternative pathway driven by IL‑12 and interferon‑γ (IFNγ) that functions through BATF and BATF2 can induce the development of CD8α+ DCs in the absence of BATF3 during infection9. Genetic tracing experiments have shown that a DC precursor, which is characterized by the expression of the C‑type lectin CLEC9A, gives rise to both CD103+ and CD103−CD207− migratory dermal DCs (but not CD103−CD207+ Langerhans cells) in the skin-draining lymph nodes10. DC subsets that are present in the skin have specialized functions. In mice, Langerhans cells have been shown to promote the induction of T helper 17 (TH17) cells in response to extracellular pathogens such as Candida albicans 11, and to cross-present exogenous skin-derived antigens to CD8+ T cells ex vivo12. However, it has also been observed that presentation of antigen by Langerhans cells can result in T cell anergy or deletion, which suggests a possible immunoregulatory role13. CD103+ dermal DCs (but not Langerhans cells) are responsible for cross-priming and for the generation of T H1 cells 11. Moreover, recent studies have identified a specific population of mouse dermal DCs that express CD301b (also known as macrophage galactose-type C-type lectin 2 (MGL2)) and that drive TH2 cell responses in an interferon regulatory factor 4 (IRF4)‑dependent manner 14–16. The regulatory role for skin DCs has also been demonstrated by the study of mice with the motheaten phenotype. In these mice, cell-specific deletion of the gene encoding SH2 domain-containing protein tyrosine phosphatase 1 (SHP1; also known as PTPN6) in CD11c+ cells resulted in increased Toll-like receptor (TLR) signalling, which correlated with urticaria-like

skin inflammation, and the development of an auto immune phenotype (including the formation of anti bodies specific for self DNA) and glomerulonephritis17. In addition, experiments in mice that involved the delivery of a model autoantigen to specific DC subsets revealed that migratory skin DCs are potent activators of regulatory T (TReg) cells, providing translational opportunities for targeted immunomodulatory therapy 18. In addition to this regulatory function, myeloid DCs can have effector functions that promote skin inflammation. In inflamed mouse skin, myeloid DCs can differentiate from invading monocytoid precursors into activated DCs that produce pro-inflammatory mediators (such as tumour necrosis factor (TNF), IL‑12 and IL‑23) and stimulate TH1‑dependent and TH17‑dependent immune responses19,20. Similar pro-inflammatory DCs have been found in inflamed skin lesions in humans21,22. Dendritic cell subsets and functions in human skin. Human Langerhans cells have been shown to have varying functions, as seen for mouse Langerhans cells. On the one hand, they show potent cross-priming activity and initiate allogeneic CD8+ T cell responses in an IL‑15‑dependent manner 23,24; on the other hand, they have been shown to contribute to the expansion of TReg cells in an autologous system, which suggests a role for Langerhans cells in tissue homeostasis25. The main DC subsets that are present in the human dermis include: CD14+CD1a− DCs, CD14−CD1a+ DCs and 6‑Sulpho LacNAc+ DCs26,27. Compared with other DC subsets, CD14+ dermal DCs are weak stimulators of T cells and they instigate humoral immune mechanisms23. Expression of CD141 (also known as thrombomodulin) by CD14+ dermal DCs characterizes a subpopulation that produces the regulatory cytokine IL‑10 and has potent immunoregulatory functions in vitro and in vivo 28. In contrast, CD14−CD141+ dermal DCs have been shown to be potent activators of CD8+ T cells29. Different types of skin inflammation are characterized by the presence of different populations of myeloid DCs in the skin, which thus contribute to the specificity of the inflammatory process30. For example, in human psoriatic skin, TNF and inducible nitric oxide synthase (iNOS; also known as NOS2)-producing DCs (TIP‑DCs) are found, in addition to inflammatory DCs that produce IL‑20 and IL‑23 (REF. 21). Skin samples from patients with another type of chronic inflammatory skin disease, atopic dermatitis, show an accumulation of a distinct population of inflammatory DCs, known as inflammatory dendritic epidermal cells, which produce CCL3, IL‑1, IL‑12p70, IL‑16 and IL‑18 (REF. 31). Thus, myeloid DCs can instigate and enhance, as well as regulate, inflammatory responses in the skin. Plasmacytoid DCs (pDCs) are important mediators of antiviral immunity owing to their ability to produce large amounts of type I IFNs in response to viral infection32. They are specialized in sensing self and viral nucleic acids, in part through TLR7 and TLR9. pDCs are absent from resting skin (or present in low numbers), but are recruited to skin that is affected by inflammation



REVIEWS

M1 macrophages A pro-inflammatory or ‘classically activated’ subset of macrophages that is characterized by phagocytic activity and the expression of particular pro-inflammatory cytokines (such as tumour necrosis factor) and pro-inflammatory mediators (such as inducible nitric oxide synthase).

M2 macrophages A pro-angiogenic or ‘alternatively activated’ subset of macrophages that is characterized by the expression of particular angiogenic cytokines (such as vascular endothelial growth factor) and anti-inflammatory mediators (such as arginase and interleukin‑10).

Wound-healing macrophages A subset of macrophages that support tissue repair by enhancing granulation tissue formation and accelerating epithelialization.

Cryopyrin-associated periodic syndromes A family of autoinflammatory syndromes, including familial cold autoinflammatory syndrome, Muckle–Wells syndrome and neonatal-onset multisystem inflammatory disease. These conditions share many clinical features (such as urticarial skin rash) and are associated with mutations in the gene encoding NLRP3 (NOD-, LRR- and pyrin domain-containing 3; also known as cryopyrin), which is a component of the inflammasome that regulates interleukin‑1β production.

in conditions such as systemic lupus erythematosus and psoriasis33,34. This has recently been illustrated by experiments showing that pDCs are required for the induction and maintenance of lupus-like skin lesions in lupus-prone mice35. In this model, mechanical disruption of the epidermal barrier by tape stripping leads to the accumulation of IFNα-producing pDCs in the skin, and blockade of TLR7 and TLR9 signalling using an oligonucleotide inhibitor can attenuate this type of skin inflammation. Moreover, pDCs seem to have a role in antitumour defence. The external application of imiquimod, a TLR7 and TLR8 agonist, to skin tumours can have a wide range of effects including the activation of both innate and adaptive immunity 36. In addition, recent studies have shown that imiquimod enables the recruitment of pDCs from the blood into the skin by a CCR6‑dependent mechanism37, and that infiltrating CD8α+ pDCs can directly kill tumour cells using cytolytic mediators, such as granzyme B38. Macrophages and monocytes. The dynamics of macrophage and monocyte populations in the skin have not been thoroughly investigated and it is not clear whether the anatomical subdivision of tissue-resident and bone marrow-derived monocytes and macrophages has a functional correlate. It is known that skin-resident tissue macrophages can originate from the yolk sac and can self renew within the tissue, at least under inflammatory conditions39. Their differentiation is determined by the activity of signalling pathways that remain poorly defined and that are, in contrast to peritoneal macrophage differentiation, independent of the transcription factor CCAAT/enhancer-binding protein-β (C/EBPβ)40. The characterization of the pathways that determine tissue-specific macrophage differentiation could potentially identify new therapeutic targets for the treatment of inflammatory skin conditions. Resident skin macro phages are thought to survey the tissue and carry out the early detection of, and response to, antigen entering the body through the skin (FIG. 2). In mice infected with Leishmania major, resident skin macrophages — in addition to DCs — are important targets of infection and can initiate an inflammatory host response41. In addition to skin-resident macrophages, circulating monocytes traffic through the skin where they support DCs in surveillance functions and in the transport of antigens to draining lymph nodes42. From the analysis of mouse models of experimental colitis, it is known that circulating LY6Chi monocytes entering the inflamed colon tissue can give rise to different myeloid cell populations43. Fate mapping of such monocytes entering the skin has not been carried out systematically but would be of great value in contributing to the understanding of inflammatory skin phenotypes. On the basis of their function, monocytes and macrophages have been divided into three major populations: classically activated (pro-inflammatory) M1 macrophages, regulatory M2 macrophages and woundhealing macrophages44. There is accumulating evidence that M1 macrophages contribute to both acute and chronic inflammation in the skin. In human psoriatic

skin, a population of IFNγ-responsive, CD163+ macro phages that express pro-inflammatory cytokines has been described45. A surrogate CD163+ cell population that was generated in vitro had a protein expression profile that overlapped with the DCs found in psoriatic skin, the so‑called TIP-DCs21. However, CD163+ macrophages and TIP-DCs are distinct cell types, which suggests that individual cell populations of the myeloid inflammatory infiltrate have non-exclusive pro-inflammatory functions, and that they are therefore not suitable as therapeutic targets. Notably, CD163+ macrophages also accumulate in skin lesions from patients with atopic dermatitis and cutaneous T cell lymphoma (CTCL)46, which suggests that their function in skin defence is not specific for a particular type of inflammation but that they have a broader role in skin immunity. Further support for the important pro-inflammatory functions of monocytes and macrophages in skin inflammation comes from two different mouse models of psoriasis-like disease: first, from mice with an epidermis-specific deletion of the gene encoding inhibitor of nuclear factor-κB (NF-κB) kinase subunit β (IKKβ; also known as IKK2), and second, from mice engineered to express a defective hypomorphic version of CD18 (also known as integrin‑β2). The pathogenic mechanisms of psoriasis-like skin inflammation in CD18 hypomorphic mice are not known. However, in both of these mouse models, depletion of CD11b+F4‑80+ skin monocytes and macrophages resulted in the resolution of skin inflammation47,48, which suggests a key pathogenic role of these cells in psoriasiform dermatitis. Recently, investigation into the pathogenesis of cryopyrin-associated periodic syndromes has revealed additional pro-inflammatory functions of skin macrophages. Mice harbouring a mutation in the gene encoding the intracellular receptor NLRP3 (NOD-, LRR- and pyrin domain-containing 3; also known as NALP3 or cryopyrin), which causes Muckle–Wells syndrome (MWS) in humans (a systemic inflammatory disease with urticarialike skin involvement), show excessive secretion of IL‑1β and IL‑18 from macrophages, monocytes, mast cells and DCs upon lipopolysaccharide (LPS) priming, without the need for a second inflammasome-activating signal such as ATP. This correlates with the development of dermatitis that is characterized by neutrophil invasion into the skin and a predominance of TH17 cell-derived cytokines49. These results suggest that uncontrolled activation of the inflammasome in myeloid cells substantially contributes to this type of skin inflammation. In contrast to pro-inflammatory M1 macrophages, regulatory M2 macrophages secrete IL‑10 and transforming growth factor‑β (TGFβ) (FIG. 2) and have an essential role in the resolution of skin inflammation50. They contribute to the dampening of skin inflammatory responses in different ways. One proposed mechanism by which skin macrophages can suppress inflammation is through the secretion of different variants of vascular endothelial growth factor (VEGF), which then leads to lymphangiogenesis and the accelerated clearance of antigen from the skin51. M2 macrophages might also directly take up antigen, thereby decreasing its availability for

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Neutrophil

Epidermis

Langerhans cell

Uptake of antigen

To draining lymph node

Keratinocyte

IL-34, M-CSF

IL-10

M2 macrophage

Dermal DC IL-23

TReg cell M1 macrophage IL-4

Lymphatic vessel

IL-2

TH1 cell

IL-1

TNF

TH17 cell To draining lymph node

M1 macrophage

M1 macrophage

Basophil

IL-12, TNF

Dermis

IL-10, TGFβ, VEGF

IL-24, CCL3, CCL4, CCL5

CCL3, CCL4, CCL5

Mast cell

Antigens

Uptake of antigen

Monocyte

Blood vessel

Figure 2 | Myeloid cells have pro-inflammatory and anti-inflammatory activities in the skin. The skin is populated by resident and trafficking myeloid cells. Langerhans cells migrate to the epidermis in response to a gradient of macrophage colony-stimulating factor (M-CSF) and interleukin‑34 (IL‑34) produced by epidermal keratinocytes. migrate NatureMonocytes Reviews | Immunology from the blood and patrol through the dermis. Epidermis-derived IL‑24 induces chemokine production by macrophages, which can attract monocytes to the skin. Dendritic cells (DCs) and monocytes can take up antigen and transport it to lymph nodes. Basophils, mast cells and dermal DCs establish a balance of pro-inflammatory and anti-inflammatory mechanisms by producing cytokines that facilitate or dampen inflammatory responses, respectively. Basophil-derived IL‑4 stimulates the differentiation of M1 macrophages into M2 macrophages that produce anti-inflammatory cytokines such as IL‑10 and transforming growth factor‑β (TGFβ), as well as vascular endothelial growth factor (VEGF), which facilitates the growth of lymphatic vessels for rapid antigen elimination. In addition, mast cells and dermal DCs produce IL‑2 and IL- 10, respectively, which facilitates the generation of regulatory T (TReg) cells. Conversely, mast cells can release preformed tumour necrosis factor (TNF), which stimulates pro-inflammatory IL‑1 production by M1 macrophages. Dermal DCs present antigen to T cells and enable T cell polarization towards T helper 1 (TH1) or TH17 phenotypes under the influence of cytokines such as IL‑12, TNF and IL- 23. CCL, CC‑chemokine ligand.

presentation to lymphocytes52. Such a mechanism has been proposed in a model of IgE-mediated allergic skin inflammation, in which the decreased development of M2 macrophages following deletion of the gene encoding IL‑4 results in enhanced inflammation52. In this model,

antigen challenge leads to the recruitment of pro-inflammatory LY6C+ monocytes to the skin via the chemokine receptor CCR2, but once in the skin they differentiate into anti-inflammatory M2 macrophages under the influence of basophil-derived IL‑4 (REF. 52). This study



REVIEWS

Resolvins Lipid mediators that are induced during the resolution phase following acute inflammation. They are synthesized in a transcellular manner from the essential omega‑3 fatty acids eicosapentaenoic acid and docosahexaenoic acid.

Protectins A family of compounds that are derived from docosahexaenoic acid and that are characterized by a conjugated triene-containing structure. They have been shown to regulate the influx of neutrophils at inflammatory sites.

Oxazolone-induced dermatitis A model of dermatitis in which the potent chemical allergen 4-ethoxymethylene-2-phenyl2-oxazolin-5-one is used experimentally to induce a delayed-type contact hypersensitivity reaction in mice.

Xenotransplant models The current gold standard for clinically relevant psoriasis models. Inflamed or symptomless skin from patients with psoriasis is transplanted into immunosuppressed mice and the development of lesions is followed over time. Such models have the disadvantage of a low throughput owing to the limited availability of patient-derived skin and they require careful standardization.

Parabiotic mice Mice in which the blood circulation has been joined surgically. Parabiotic mice share the blood circulation and exchange blood cells, such as lymphocytes, which allows the study of the role of circulating immune cells compared with tissue-resident immune cells.

provides an example of macrophage plasticity, as illustrated by their capacity to change phenotype according to the local tissue milieu, and also emphasizes the crucial role of cellular crosstalk in the regulation of skin immune responses. Another mechanism by which macrophages can dampen skin inflammation is through upregulated phagocytosis, which can be stimulated by pro-resolving mediators such as resolvins and protectins. These derivatives of polyunsaturated fatty acids are synthesized by both resident and infiltrating cells. In a model of skin infection with Staphylococcus aureus, resolvins D1 and D5, as well as protectin D1, were able to enhance the antibiotic action of vancomycin and reduce the levels of IL‑6 and granulocyte–macrophage CSF (GM‑CSF) in the inflammatory exudate53. Mast cells. Mast cells are specialized for first-line surveillance functions in the skin owing to the presence of a plethora of preformed pro-inflammatory mediators that are stored within their cytoplasmic granules and can be rapidly released upon mast cell contact with antigens. Their key role in host defence is illustrated in dengue virus infection, in which skin mast cells contribute to an effective innate immune response and to viral clearance by recruiting natural killer (NK) cells and NKT cells to the sites of infection54. In addition, mast cells have diverse functions in skin pathology. For example, in a mouse model of dermatitis, mast cell degranulation induced by staphylococcal δ‑toxin is important for the enhancement of inflammation and the increased production of systemic IgE and IL‑4 (REF. 55). In Nlrp3‑mutant mice, which are a model for MWS, cutaneous inflammation does not develop in the absence of mast cells; however, Nlrp3‑mutant mast cells are not sufficient to trigger the disease 56. Interestingly, in this model, mast cell-derived TNF seems to promote IL‑1β release and is required for the development of skin inflammation in both newborn and adult Nlrp3‑mutant mice. These results suggest that although monocytes and macrophages are crucial effector cells in the development of skin inflammation in MWS, they act downstream of mast cells in the hierarchy of a tissue‑specific myeloid cell network. Similarly to macrophages, mast cells and basophils can have pro-inflammatory and anti-inflammatory functions in the skin. In MWS, mast cells support M1 macrophagedependent inflammation (see above), whereas in chronic oxazolone-induced dermatitis they suppress skin inflammation through the production of IL‑2 (FIG. 2). In this model, mast cell-derived IL‑2 leads to an accumulation of TReg cells in the inflamed skin57. Similarly, basophils can secrete IL‑4 in an IgE-mediated allergic inflammatory response, thereby converting pro-inflammatory M1 macrophages into M2 macrophages52.

Lymphoid cells in skin immunity and inflammation T cells. The classical concept of skin immune surveillance implies that recirculating immune cells are the main effectors of the skin immune system58,59. However, data obtained from xenotransplant models of human skin inflammation have challenged this concept,

demonstrating that T cells that reside in non-inflamed human skin are necessary and sufficient to create an inflammatory pathology in the absence of recirculating lymphocytes60–62. In addition, quantitative approaches have shown that considerable numbers of T cells reside in normal human skin (~2 × 1010 in the entire skin surface, approximately twice the number of circulating T cells)63. Studies of immune surveillance in a model of herpes simplex virus (HSV) infection showed that skin-resident memory T cells achieved superior protection compared with circulating memory CD4+ T cells, and that this was mediated by pathogen-specific CD8+CD103+ tissueresident memory T cells64. Similar findings have been obtained in studies of the gut mucosa65. Upon resolution of HSV infection, the skin contained two distinct pathogen-specific memory subsets: a population of epidermis-resident memory CD8 + T cells and a population of recirculating memory CD4+ T cells66. Interestingly, peripheral tissues can support the generation and persistence of resident memory CD8+ T cells in the absence of antigen stimulation, thereby providing effective protection from de novo infection67,68. Using a vaccinia virus infection model, it has been shown that skin-resident effector memory T cells provide complete protection against cutaneous challenge, whereas protection against lethal respiratory challenge required both respiratory mucosa-resident effector memory T cells and central memory T cells69. In a subsequent study using parabiotic mice, the authors showed that long-lived nonrecirculating skin-resident memory T cells were better than circulating central memory T cells at providing rapid protection against skin re-infection 70. Similar findings have been reported for lung tissue71. What is the translational and clinical relevance of these findings? An interesting model disease in which to investigate the role of skin-resident, compared with circulating, memory T cells is CTCL, a cancer of skinhoming T cells. The CD52‑specific monoclonal antibody alemtuzumab depletes circulating and tissue-resident central memory T cells in the blood of patients with CTCL but leaves a distinct population of skin-resident effector memory T cells intact. Patients with CTCL who have been treated with alemtuzumab show efficient protection from bacterial and viral skin infections, which suggests that the small residual population of skin-resident memory T cells is sufficient for protective immunity 72. Recent work has shown that, in addition to tissueresident effector memory T cells, there is a population of tissue-resident memory TReg cells in the skin. Using a mouse model of skin-specific autoimmunity — in which disease results from the tetracycline-inducible expression of ovalbumin in keratinocytes — it was shown that thymus-derived TReg cells mediated the resolution of organ-specific autoimmunity 73. Self-reactive TReg cells that were generated by self-antigen expression in the thymus were activated following encounter with peripheral tissue autoantigens. Activated TReg cells persisted in the target tissue and suppressed autoimmune responses upon repeated encounters with tissue autoantigen73. The authors have termed this phenomenon ‘regulatory memory’73.

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Wound healing The sequence of events initiated after tissue injury that lead to its repair. The woundhealing response consists of four phases: coagulation, inflammation, proliferation and remodelling. It leads to the formation of a fibrotic replacement scar tissue.

Imiquimod-induced model of psoriasis A mouse model of psoriasiform dermatitis caused by topical application of the Toll-like receptor 7 (TLR7) and TLR8 agonist imiquimod. The imiquimod model is relatively easy to perform and, although not a perfect mimic of the human disease, it reflects key immune pathways in psoriasis such as the involvement of the interleukin‑23–T helper 17 cell axis.

Group 1 ILCs A group of innate lymphoid cells (ILCs) — including natural killer cells and a subset of ILCs (ILC1s) — that produce type 1 cytokines such as tumour necrosis factor and interferon‑γ, and express the transcription factor T‑bet. These cells contribute to immune responses against viruses and intracellular pathogens, as well as to tumour surveillance.

Group 2 ILCs A subset of innate lymphoid cells (ILCs) that produce type 2 cytokines, such as interleukin-4 and interleukin‑13. Their development depends on the transcription factors retinoic acid receptor-related orphan receptor‑α and GATA-binding protein 3. These cells contribute to tissue repair and parasite elimination, as well as to the development of asthma and allergy.

γδ T cells. γδ T cells provide early tissue immune responses as part of the ‘lymphoid stress-surveillance’ response74. In mice, the first wave of γδ T cells that leave the thymus localize to the skin, which indicates an important role for γδ T cells in mouse skin. The thymic selection of mouse Vγ5Vδ1 + skin-homing T cells depends on the expression of the immunoglobulin superfamily member SKINT1 (selection and upkeep of intraepithelial T cells protein 1)75. γδ T cells have important roles in tumour immune surveillance, wound healing and skin inflammation. They are essential for protection against skin tumours through mechanisms involving the NK group 2, member D (NKG2D) receptor and its respective ligands, as well as through IFNγ and IL‑17 production76. A new subset of γδ T cells has been identified in the dermis. These dermal γδ T cells express the IL‑7 receptor, CCR6 and retinoic acid-related orphan receptor-γt (RORγt), and are pre-committed to IL‑17 production in response to microbial challenge 77,78. Dermal γδ T cells are distinct from the classical IFNγproducing Vγ5Vδ1+ DETCs, which supports the idea that discrete subsets of γδ T cells occupy specific niches within the skin. Mouse IL‑17‑producing dermal γδ T cells have a major role in the imiquimod-induced model of psoriasis36,79–81. Mice with a mutation in the gene encoding the transcription factor SOX13 were protected from psoriasis-like skin inflammation, which supports a role for SOX13‑expressing migratory IL‑17+ γδ T cells in this setting 82. Dermal γδ T cells were also shown to be important producers of the pathological cytokine IL‑22 in this model83. IL‑15 is responsible for the expansion of IL‑17‑producing γδ (and αβ) T cell populations, a process that is inhibited by keratinocyte-derived soluble IL‑15 receptor antagonist in a negative feedback loop84. Dermal γδ T cells depend on CCR6 expression for homing to the skin and for the promotion of skin inflammation 85. The IL‑36‑mediated crosstalk between DCs and epidermal keratinocytes regulates the recruitment and activation of CCR6+ dermal γδ T cells and the pathogenesis of psoriasiform lesions in response to the administration of imiquimod86. Further evidence for a role of dermal γδ T cells in skin inflammation comes from a spontaneously developing psoriasis-like mouse model in CD18‑hypomorphic mice87. The severity of psoriasi form inflammation in these mice correlated with the loss of skin-resident Vγ5+ T cells and concurrent skin infiltration of IL‑17- and IL‑22‑producing dermal γδ T cells, and this was preceded by an increase in the frequency of Vγ4+ T cells in local lymph nodes. An extensive study of human γδ T cells in skin homeostasis and inflammation has identified the Vγ9Vδ2+ subset as a rapidly recruited immune cell subpopulation that exerts its pro-inflammatory action through the production of mediators such as IL‑17. The frequency of these cells was also increased in psoriatic human skin and their numbers in the blood correlated with disease severity 88. Thus, there is increasing evidence for an important contribution of γδ T cells to skin inflammation in both mice and humans.

Skin γδ T cells have an important role in epithelial homeostasis and tissue repair. They contribute to epidermal homeostasis through the production of growth factors such as fibroblast growth factor 7 (FGF7), FGF9 and insulin-like growth factor 1, which promote the survival of keratinocytes89. The binding of semaphorin 4D (also known as CD100) to plexin B2 (which is expressed by keratinocytes) has recently been described as crucial for the activation of epidermal γδ T cells in response to skin perturbation, and potentially for their positioning in skin tissue90. As a further example of how the immune system affects tissue regeneration, a recent study has shown that the initial secretion of FGF9 by dermal γδ T cells amplifies WNT activation in and FGF9 secretion by fibroblasts, ultimately leading to hair follicle regeneration91. The authors of the study concluded that the absence of dermal γδ T cells in humans might explain their inability to regenerate hair after wounding. Innate lymphoid cells. Innate lymphoid cells (ILCs) are a diverse family of immune cells that produce cytokines and coordinate immunity and inflammation in body surface tissues, such as the intestine, lungs and skin. They can be divided into three subsets — group 1 ILCs (ILC1s and NK cells), group 2 ILCs (ILC2s) and group 3 ILCs (ILC3s and lymphoid tissue inducer cells) — on the basis of their expression of cytokines and transcription factors92. Although ILC1s are present in human skin, their role in skin homeostasis is less well understood than that of ILC2s and ILC3s93. A group of IL‑7‑dependent and IL‑13‑producing ILC2s has recently been identified in normal mouse dermis and these ILC2s were found to have a potential role in the induction of TH2‑type dermatitis94. In keeping with this possibility, ILC2s formed stable interactions with mast cells (lasting longer than 20 minutes)94. However, ILC2‑derived IL‑13 suppressed mast cell function in vitro, indicating a complex modulation of mast cells by ILC2s in models of TH2‑type immunity 94. Another study has shown that ILC2s promote TH2‑type atopic dermatitis-like skin inflammation in mice in a thymic stromal lymphopoietin (TSLP)-dependent but IL‑25- and IL‑33‑independent manner and that similar cells exist in human atopic dermatitis95. A recent study has shown that ILC2s infiltrate human skin after antigen challenge and that ligation of E-cadherin (also known as cadherin 1) suppresses their effector function by inhibiting the expression of IL‑5 and IL‑13 (REF. 96). Group 3 ILCs are the major population of ILCs in normal human skin. In the imiquimod-induced mouse model of psoriasis, RORγt+ ILC3s (and γδ T cells) that invade the skin are necessary and sufficient for psoriatic plaque formation36,80,81. In human psoriasis, there is an accumulation of natural cytotoxicity triggering receptor (NCR)-expressing ILC3s both in the affected skin tissue and in the circulation93. Increased numbers of NCR+ ILC3s are also found in non-lesional psoriatic skin, which suggests that these cells may be responsible for triggering psoriasis93. In addition, the number of



REVIEWS circulating ILC3s in patients with psoriasis correlates with the response to TNF-targeted monoclonal antibody therapy, which indicates a potential biomarker function for ILCs93. Thus, ILC3s seem to make an important contribution to epithelial inflammation and they could act as disease biomarkers and therapeutic targets.

Group 3 ILCs A subset of innate lymphoid cells (ILCs) that mainly reside in the intestinal tract. Their development depends on the transcription factor retinoic acid receptor-related orphan receptor‑γt. These cells are thought to regulate the balance between the microbiota and the intestinal immune system. This group of cells includes lymphoid tissue inducer cells (which are involved in the development of lymphoid tissue), natural cytotoxicity triggering receptor (NCR)-expressing ILC3s (which mainly produce IL‑22), and NCR-negative ILC3s (which mainly produce IL‑17A).

Necroptosis A programmed form of necrotic cell death mediated by receptor-interacting protein kinase 1 (RIPK1) and RIPK3. It can be induced by death receptors and by TIR-domaincontaining adaptor protein inducing interferon-β (TRIF)-dependent Toll-like receptor 3 (TLR3) and TLR4 signalling. Inhibition of caspase 8 activation sensitizes cells to necroptosis.

Linear ubiquitin assembly complex (LUBAC). A ubiquitin ligase complex composed of SHARPIN (SHANK-associated RH domain-interacting protein), HOIL1L (haem-oxidized IRP2 ubiquitin ligase 1) and HOIP (HOIL1‑interacting protein) that generates linear polyubiquitin chains. LUBAC-mediated linear ubiquitylation of NF-κB essential modulator (NEMO) and other components of the tumour necrosis factor (TNF) receptor 1 signalling pathway regulates cellular responses to TNF.

Keratinocytes as initiators of inflammation Owing to their strategic positioning at the interface between the body and the environment, epidermal keratinocytes receive signals from the environment and transmit them to immune cells in the skin. This communication is achieved through the expression of receptors that sense microorganisms and other environmental stress factors, through the production of cytokines and chemokines, including members of the IL‑1, IL‑10, IL‑20 and TNF cytokine families, and through the expression of cytokine and chemokine receptors1. Many pro-inflammatory signalling cascades are activated in the epidermis of patients with inflammatory skin disease97–99, which suggests that keratinocyte-intrinsic pathways have a key role in regulating immune homeostasis and inflammation in the skin. In this section we discuss recent studies in mouse models that provide evidence of a function for keratinocytes in the pathogenesis of inflammatory skin diseases. Epithelial cell signalling in skin inflammation. AP‑1 transcription factors — consisting primarily of JUN and FOS protein dimers — are key regulators of inflammation, and dysregulation of these factors has been proposed to contribute to the initiation of skin inflammation100. Indeed, in humans with psoriasis, epidermal keratinocytes have reduced JUNB expression compared with control keratinocytes; in line with this, mice with epidermis-specific ablation of Junb and Jun have been shown to develop psoriasis-like skin inflammation accompanied with arthritis101,102. Conversely, increased FOS-mediated gene transcription in keratinocytes has been shown to trigger skin inflammation in mice103,104. Another transcription factor, signal transducer and activator of transcription 3 (STAT3), has also been shown to be more active in psoriatic epidermis than in unaffected epidermis98. Indeed, studies in transgenic mice with keratinocyte-specific expression of constitutively active STAT3 supported a causative role for epidermal STAT3 activation in the pathogenesis of psoriasis-like skin inflammation98. In addition, epidermal keratinocyte-specific knockout of suppressor of cytokine signalling 3 (SOCS3), a negative regulator of STAT3, induced constitutive STAT3 activation and resulted in the spontaneous development of IL‑6‑driven psoriasis‑like skin inflammation105. A third signalling pathway that has a well-known role in skin inflammation is the TNF pathway, as confirmed by the efficacy of TNF-neutralizing reagents in the treatment of psoriasis106. Nevertheless, the TNF-dependent cellular and molecular mechanisms that control skin inflammation remain poorly understood. Studies in mouse models have shown

that keratinocyte-restricted inhibition of NF‑κB — which was achieved by either the ablation of Ikbkb (which encodes IKKβ) or the transgenic expression of a mutated super-repressor NF-κB inhibitor-α (also known as IκBα) — resulted in the development of TNF-dependent psoriasis-like inflammatory skin lesions 107,108 . Moreover, a recent study showed that the development of skin inflammation in mice with epithelial cell-specific knockout of Ikbkb depends on keratinocyte-intrinsic TNF receptor 1 (TNFR1)‑mediated overexpression of IL‑24, which acts in a paracrine and/or autocrine manner to activate the IL‑22 receptor subunit‑α1 (IL‑22RA1)–STAT3 signalling pathway and the expression of pro-inflammatory cytokines and chemokines in keratinocytes109 (FIG. 3a). Interestingly, the absence of IL‑22 (which also signals through IL‑22RA1 and activates STAT3) did not prevent the development of skin lesions in Ikbkb-knockout mice, which suggests that IL‑22 and IL‑24 could have distinct roles in skin inflammation109. Keratinocyte death in skin inflammation. Recent studies in mouse models have suggested that keratinocyte death is a potent trigger of skin inflammation. Keratinocyte-restricted ablation of the gene that encodes FAS-associated death domain protein (Fadd) — a protein that is essential for caspase 8 activation and for apoptosis downstream of death receptor signalling — or inducible deletion of caspase 8 triggered severe skin inflammation in mice and this was prevented by genetic ablation of receptor-interacting protein kinase 3 (Ripk3)110. These findings suggest that keratinocytes that are deficient in FADD or caspase 8 undergo RIPK3‑dependent necroptosis111–113, and this leads to the release of damage-associated molecular patterns (DAMPs) that then activate TLRs and other receptors to induce skin inflammation (FIG. 3b). Mice with epidermis-specific genetic ablation of the IKK complex regulatory subunit NF‑κB essential modulator (NEMO), the upstream kinase TGFβ-activated kinase 1 (TAK1; which is required for the activation of NF‑κB and mitogen-activated protein kinases (MAPKs)) or the caspase 8 and FADD-like apoptosis regulator (CFLAR; also known as CFLIP) showed increased apoptosis of keratinocytes and developed skin inflammation114–118. Furthermore, mutant mice with chronic proliferative dermatitis caused by a lack of expression of the linear ubiquitin assembly complex component SHANK-associated RH domain-interacting protein (SHARPIN)119–121 show extensive keratinocyte apoptosis and develop TNF-dependent skin inflammation. Importantly, sensitization of intestinal epithelial cells to either necroptotic or apoptotic cell death has also been shown to trigger inflammation in the gut 122–126. These findings in mice suggest that epithelial cell death is a potent mechanism for the triggering of inflammation in barrier tissues such as the skin and the intestine. A common factor in all of these mouse models is that the development of epithelial cell death and inflammation depends at least in part on TNF signalling via TNFR1, which suggests that dysregulation of TNFR1

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Figure 3 | Tumour necrosis factor-induced signalling and cell death in skin inflammation. a | Interleukin‑24 (IL‑24) expression by keratinocytes promotes psoriasis-like skin inflammation. Mice with epidermal keratinocyte-specific Nature human Reviewspsoriatic | Immunology inhibition of nuclear factor‑κB (NF‑κB) develop chronic inflammatory skin lesions that resemble skin inflammation. During NF‑κB inhibition, tumour necrosis factor receptor 1 (TNFR1) signalling in keratinocytes causes increased expression of IL‑24 through a signalling pathway involving reactive oxygen species (ROS), mitogen-activated protein kinase kinases (MAPKKs) and extracellular signal-regulated kinases (ERKs). IL‑24 acts in an autocrine or paracrine manner to induce the activation of signal transducer and activator of transcription 3 (STAT3) in keratinocytes by binding to its heterodimeric receptor, which is composed of IL‑22 receptor subunit α1 and IL‑20 receptor subunit-β (IL‑22RA1–IL‑20RB). This results in the increased expression of pro-inflammatory cytokines and chemokines, which leads to the recruitment of macrophages and other immune cells and to the development of chronic psoriasis-like inflammatory skin lesions. Findings from studies in genetic mouse models suggest that infection with bacteria or viruses that express NF‑κB inhibitors could have a similar effect in dysregulating TNFR1 signalling in keratinocytes, resulting in IL‑24 expression and skin inflammation. b | Keratinocyte necroptosis drives skin inflammation. Mice with epidermis-specific FAS-associated death domain protein (FADD) or caspase 8 deficiency develop severe skin inflammation that depends on receptor-interacting protein kinase 3 (RIPK3). Keratinocytes lacking FADD or caspase 8 undergo RIPK3‑dependent necroptotic cell death that partly depends on TNFR1 signalling. RIPK3‑dependent necroptosis of epidermal keratinocytes probably induces skin inflammation through the release of damage-associated molecular patterns (DAMPs) from dying necrotic cells, which activate an inflammatory response by activating Toll-like receptors (TLRs) or other receptors that are expressed by cutaneous immune and stromal cells. The findings from mutant mouse models suggest that infection with bacteria or viruses expressing caspase 8 inhibitors could similarly sensitize keratinocytes to RIPK3‑dependent necroptosis and trigger skin inflammation. CFLAR, caspase 8 and FADD-like apoptosis regulator; cIAP, baculoviral IAP repeat-containing protein; IKK, inhibitor of NF-κB kinase; JAK, Janus kinase; LUBAC, linear ubiquitin assembly complex; MLKL, mixed lineage kinase domain-like protein; NEMO, NF‑κB essential modulator; TAK1, TGFβ-activated kinase 1; TAB, TAK1‑binding protein; TNF, tumour necrosis factor; TRAF2, TNFR-associated factor 2; TRADD, TNFR1‑associated death domain protein.



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Figure 4 | Microbial regulation of skin immune responses. a | Skin commensals control skin-resident T cell function Nature Reviews | Immunology and protective immunity to cutaneous pathogens. Colonization of the skin with the commensal Staphylococcus epidermidis was shown to be required for the production of interleukin‑17A (IL‑17A) by skin-resident T cells136. During subcutaneous infection with Leishmania major, the presence of S. epidermidis was required for interferon‑γ (IFNγ) production by skin-resident T cells and for protective immunity. IL‑17A and IFNγ expression by T cells and protective immunity against L. major was dependent on IL‑1 receptor signalling through the adaptor myeloid differentiation primary response protein 88 (MYD88). S. epidermidis induced the expression of IL‑1α and at the same time inhibited the expression of IL-1 receptor antagonist (IL-1RA) in skin cells, and this had the net effect of inducing IL‑17A and IFNγ expression by skin-resident T cells either directly or indirectly by acting on myeloid or other skin cells. b | S. aureus δ‑toxin induces mast cell degranulation and allergic skin inflammation. Colonization of the skin with S. aureus, a pathogenic bacterium that is often found on the skin of patients with atopic dermatitis, triggers local allergic responses by releasing δ‑toxin, which directly induces degranulation of dermal mast cells, promoting innate and adaptive T helper 2 (TH2)-type responses.

signalling in epidermal keratinocytes is a potent trigger of skin inflammation. These findings suggest that skin epithelial cells could be important cellular targets of pathogenic TNF signalling in human chronic inflammatory skin diseases such as psoriasis. The abnormal keratinocyte response to TNF could be an evolutionarily conserved mechanism that evolved to provide an early signal that alerts the immune system to infection. Given that many bacteria and viruses express proteins that inhibit different branches of the TNFR1 signalling network — such as NF-κB127,128 and the FADD–caspase 8 apoptotic cascade129–131 — it is possible that the contact of epidermal keratinocytes with such microorganisms could trigger skin inflammation in a manner similar to that observed in mouse models in which TNFR1 signalling is genetically disrupted. In individuals with a susceptible genetic background, such stochastic events could provide the trigger for the development of chronic skin inflammation.

Control of skin immunity by resident commensals The surface of the skin is colonized by rich microbial communities that interact with local epithelial, stromal and immune cell populations to establish immune homeostasis. The human skin is home to distinct bacterial and fungal communities, the complexity and characteristics of which largely depend on the milieu provided by specific sites on the skin surface132,133. This commensal microbiota protects the skin from infection by competing with pathogens and contributes to immune homeostasis by influencing local immune responses in both beneficial and detrimental ways134. This interaction is reciprocal, with epithelial and immune cells in the skin releasing antimicrobial peptides to control the microbial colonization of the skin surface. However, studies in mice have shown that the skin microbiota is not affected by the absence of B cells and T cells, Langerhans cells or TLR and IL‑1 receptor signalling, which argues against a role for adaptive

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REVIEWS and innate immunity in shaping the skin microbiota135. Although the study of microorganism–host interactions in the skin constitutes an important area of current research, extensive discussion of this field is beyond the scope of this Review. Instead, we illustrate the important role of microbiota–host crosstalk both in establishing beneficial host defence and in triggering the pathogenesis of skin inflammation in the following discussion of two recent studies. Microbial colonization of the skin was recently shown to be essential for the elicitation of effective T cell responses and for protective immunity against infection with the parasite L. major 136. Accordingly, germ-free animals failed to develop protective T cell immunity against L. major, a defect that was restored by microbial colonization of the skin but not of the gut. Colonization of the skin of these mice with the commensal bacterium Staphylococcus epidermidis was sufficient to restore effective T cell immunity to L. major by regulating IL‑1‑dependent inflammatory responses in the skin (FIG. 4a). It remains unclear whether this function is specifically performed by S. epidermidis or whether other skin commensals have similar properties. By contrast, skin bacteria can also contribute to the pathogenesis of inflammatory skin disease. An altered composition of the skin microbiota has been associated with skin inflammation but a causative relationship and the specific pathogenic mechanisms have been difficult to establish. Staphylococcus aureus, a common cause of skin infections, is frequently found on the skin of patients with atopic dermatitis but not on the skin of healthy individuals 137. Immunity against S. aureus depends on neutrophil recruitment mediated by IL‑1 receptor 1 (IL‑1R1) and myeloid differentiation primary response protein 88 (MYD88) signalling, and by the production of IL‑17 by skin γδ T cells138–140. A recent study showed that colonization of the skin with S. aureus triggers local allergic responses by releasing δ‑toxin, which directly induces the degranulation of dermal mast cells, and in turn promotes both innate and adaptive (TH2 cell) responses55 (FIG. 4b). This study illustrates how skin microorganisms can contribute to the pathogenesis of inflammatory skin disease by directly regulating local immune cell responses. Further investigation of the crosstalk that occurs between the microbiota and epithelial, stromal and immune cells in the skin will be required to better elucidate the mechanisms that regulate skin immune homeostasis and inflammation.

Nestle, F. O., Di Meglio, P., Qin, J. Z. & Nickoloff, B. J. Skin immune sentinels in health and disease. Nature Rev. Immunol. 9, 679–691 (2009). 2. Di Meglio, P., Perera, G. K. & Nestle, F. O. The multitasking organ: recent insights into skin immune function. Immunity 35, 857–869 (2011). 3. Ginhoux, F. et al. Langerhans cells arise from monocytes in vivo. Nature Immunol. 7, 265–273 (2006). 4. Hoeffel, G. et al. Adult Langerhans cells derive predominantly from embryonic fetal liver monocytes with a minor contribution of yolk sac-derived macrophages. J. Exp. Med. 209, 1167–1181 (2012). 1.

Conclusions and perspectives Understanding the mechanisms that regulate immune homeostasis to ensure efficient host defence in the absence of pathological inflammatory responses remains the major challenge in immunology, particularly in barrier tissues (such as the skin) that are constantly exposed to potentially harmful insults threatening to throw immunoregulatory mechanisms off balance. Considerable progress has been made in the past few years in understanding the functions of the different cell types in the skin and the mechanisms that they use to communicate. The role of epidermal keratinocytes as active components of the immunoregulatory network in the skin and their role in the pathogenesis of skin inflammation is now well appreciated. The discovery of new types of myeloid and lymphoid cells with specialized functional properties has shed light on the complexity and plasticity of these populations in the skin and their role in skin immunity and disease. The identification of skin-resident memory T cell populations that have key roles in controlling cutaneous immune responses has revealed the existence of local tissue immunological memory and its importance in host defence. Finally, the realization that commensal and pathogenic bacteria that colonize the skin surface have the capacity to directly regulate the functional properties of skin immune cells has provided new paradigms that highlight the importance of host–microorganism interactions in controlling local immune responses. Despite this progress, we are still far from a comprehensive understanding of the immune regulation of the skin in health and disease. For example, the pathogenic mechanisms that trigger the initiation of chronic inflammatory skin diseases such as psoriasis are not known. A better understanding of these mechanisms may derive from future investigation of how the dysregulation of epithelial cell responses to stress, caused by injury or infection, triggers skin inflammation. More studies are needed to improve our understanding of how host skin cells and microorganisms colonizing the skin negotiate a peaceful and mutually beneficial interaction, and how dysregulation of this crosstalk leads to impaired immunity and pathology. Future studies will also combine the latest genomic technologies with computational analysis approaches for the discovery of skin-relevant immune pathways and therapeutic targets141. Finally, considering the fundamental differences between mouse and human skin, an important challenge remains to translate genetic mouse model studies into a better understanding of the mechanisms regulating immune responses in human skin.

5. Greter, M. et al. Stroma-derived interleukin‑34 controls the development and maintenance of langerhans cells and the maintenance of microglia. Immunity 37, 1050–1060 (2012). 6. Wang, Y. et al. IL‑34 is a tissue-restricted ligand of CSF1R required for the development of Langerhans cells and microglia. Nature Immunol. 13, 753–760 (2012). 7. Nagao, K. et al. Stress-induced production of chemokines by hair follicles regulates the trafficking of dendritic cells in skin. Nature Immunol. 13, 744–752 (2012).


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Acknowledgements

The authors apologize to all of the authors whose work could not be discussed and cited owing to space limitations. The authors acknowledge support from the following grant fund‑ ing bodies: M.P. and I.H. are supported by grant SFB829 from the Deutsche Forschungsgemeinschaft (DFG), Germany; M.P. is supported by the DFG (grants SFB670 and SPP1656), the European Research Council (2012‑ADG_20120314), the European Commission (FP7 grants 223404 (Masterswitch) and 223151 (InflaCare)), the Deutsche Krebshilfe Association, Germany (grant 110302), the Else Kröner-Fresenius-Stiftung Foundation, Germany, and the Helmholtz Alliance Preclinical Comprehensive Cancer Center, Germany; I.H. is supported by the Deutsche Krebshilfe Association (grant 109798); F.O.N. is supported by the European Commission (FP7 grant agreement HEALTH‑F2‑2011‑261366) and the Wellcome Trust (pro‑ gramme GR078173MA). The authors’ research is funded and supported in part by the National Institute for Health Research (NIHR) Biomedical Research Centre based at Guy’s and St Thomas’ NHS Foundation Trust and King’s College London, UK. The views expressed are those of the authors and not necessarily those of the National Health Service, the NIHR or the Department of Health.

Competing interests statement

The authors declare no competing interests.


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