Activation Of Dendritic Epidermal T Cells Research Articles

Dendritic Epidermal T Cells in Allergic Contact Dermatitis

Allergic contact dermatitis (ACD) is a common inflammatory skin disease with a prevalence of approximately 20% in the European population. ACD is caused by contact allergens that are reactive chemicals able to modify non-immunogenic self-proteins to become immunogenic proteins. The most frequent contact allergens are metals, fragrances, and preservatives. ACD clinically manifests as pruritic eczematous lesions, erythema, local papules, and oedema. ACD is a T cell-mediated disease, involving both CD4+ and CD8+ T cells. In addition, γδ T cells appear to play an important role in the immune response to contact allergens. However, it is debated whether γδ T cells act in a pro- or anti-inflammatory manner. A special subset of γδ T cells, named dendritic epidermal T cells (DETC), is found in the epidermis of mice and it plays an important role in immunosurveillance of the skin. DETC are essential in sensing the contact allergen-induced stressed environment. Thus, allergen-induced activation of DETC is partly mediated by numerous allergen-induced stress proteins expressed on the keratinocytes (KC). Several stress proteins, like mouse UL-16-binding protein-like transcript 1 (Mult-1), histocompatibility 60 (H60) and retinoic acid early inducible-1 (Rae-1) α-ε family in mice and major histocompatibility complex (MHC) class I—chain-related A (MICA) in humans, are upregulated on allergen-exposed KC. Allergen-induced stress proteins expressed on the KC are consequently recognized by NKG2D receptor on DETC. This review focuses on the role of γδ T cells in ACD, with DETC in the spotlight, and on the role of stress proteins in contact allergen-induced activation of DETC.

Epidermal loss of phospholipase Cδ1 attenuates irritant contact dermatitis

Irritant contact dermatitis (ICD) is one of the most common inflammatory skin diseases caused by exposure to chemical irritants. Since chemical irritants primarily damage keratinocytes, these cells play a pivotal role in ICD. One of the phosphoinositide-metabolizing enzymes, phospholipase C (PLC) δ1, is abundantly expressed in keratinocytes. However, the role of PLCδ1 in ICD remains to be clarified. Here, we found that croton oil (CrO)-induced ear swelling, a feature of ICD, was attenuated in keratinocyte-specific PLCδ1 knockout mice (PLCδ1 cKO mice). Dendritic epidermal T cells (DETCs), which have a protective role against ICD, were activated in the epidermis of the PLCδ1 cKO mice. In addition, the skin of CrO-treated PLCδ1 cKO mice showed increased infiltration of Gr1+CD11b+ myeloid cells. Of note, elimination of Gr1+CD11b+ myeloid cells restored CrO-induced ear swelling in PLCδ1 cKO mice to a similar level as that in control mice. Taken together, our results strongly suggest that epidermal loss of PLCδ1 protects mice from ICD through induction of Gr1+CD11b+ myeloid cells and activation of DETCs.

Role of TNF-α reception in epidermal γδ T cell function during homeostasis and after TCR stimulation

Abstract Inflammatory responses mediated by the cellular components of epithelial tissues are essential to the wound healing process. Epidermal γδ T cells, also known as dendritic epidermal T cells (DETC), play key roles in the wound healing process by producing growth factors, chemokines and cytokines necessary for the proliferation and migration of epithelial cells. DETC activation occurs via the γδ TCR, however, second and third signals are required to sustain the activation and production of factors needed in functions such as wound repair. While progress has been made on elucidating signal 2 costimulatory signals such as JAML and CAR, less is understood about what signals are required for signal 3 in DETC and what roles TNF-α plays in regulating and sustaining DETC function. TNF-α signals through two receptors, tumor necrosis factor receptor 1 and 2 (TNFR1 and TNFR2). These receptors elicit varied responses that include apoptosis, survival or inflammation. Our data indicate that DETC upregulate type 1 chemokines and their corresponding chemokine receptor expression during inflammation. We have also found that a subset of DETC express TNFR1 and TNFR2 on their cell surface constitutively suggesting a requirement for tonic TNFR signaling for particular DETC populations. We have quantified chemokine, cytokine and growth factor production from DETC stimulated in the presence or absence of TNF-α using Luminex technology. Additionally, we have investigated which TNFR is responsible for modulating DETC function by comparing DETC function in cells isolated from wild-type, TNFR1 and/or 2 knockout mice. Identifying which factors regulate DETC function and activation could provide potential targets for improving DETC function in wound repair.

IL-15 Enhances Activation and IGF-1 Production of Dendritic Epidermal T Cells to Promote Wound Healing in Diabetic Mice.

Altered homeostasis and dysfunction of dendritic epidermal T cells (DETCs) contribute to abnormal diabetic wound healing. IL-15 plays important roles in survival and activation of T lymphocytes. Recently, reduction of epidermal IL-15 has been reported as an important mechanism for abnormal DETC homeostasis in streptozotocin -induced diabetic animals. However, the role of IL-15 in impaired diabetic wound healing remains unknown. Here, we found that, through rescuing the insufficient activation of DETCs, IL-15 increased IGF-1 production by DETCs and thereby promoted diabetic skin wound repair. Regulation of IGF-1 in DETCs by IL-15 was partly dependent on the mTOR pathway. In addition, expression of IL-15 and IGF-1 were positively correlated in wounded epidermis. Together, our data indicated that IL-15 enhanced IGF-1 production by DETCs to promoting diabetic wound repair, suggesting IL-15 as a potential therapeutic agent for managing diabetic wound healing.

NKG2D-Dependent Activation of Dendritic Epidermal T Cells in Contact Hypersensitivity

The interaction between keratinocytes (KC) and skin-resident immune cells plays an important role in induction of contact hypersensitivity (CHS). A specific subset of γδ T cells termed dendritic epidermal T cells (DETC) are located in mouse epidermis, and we have recently shown that DETC become activated and produce IL-17 in an IL-1β-dependent manner during CHS. Various receptors on DETC, including NKG2D, are involved in DETC responses against tumors and during wound healing. The ligands for NKG2D (NKG2DL) are stress-induced proteins such as Mult-1, H60, Rae-1 in mice and MICA, MICB and ULBP in humans. Here, we show that allergens up-regulate expression of the NKG2DL Mult-1, H60 and Rae-1 in cultured mouse KC and of MICA in primary human KC. We demonstrate that Mult-1 is expressed in mouse skin exposed to allergen. Furthermore, we find that the vast majority of DETC in murine epidermis and skin-homing cutaneous lymphocyte-associated antigen (CLA) positive γδ T cells in humans express NKG2D. Finally, we demonstrate that blocking of NKG2D partially inhibits allergen-induced DETC activation. These findings demonstrate that NKG2D and NKG2DL are involved in allergen-induced activation of DETC and indicate that the NKG2D/NKG2DL pathway might be a potential target for treatment of CHS.

IL-1β–Dependent Activation of Dendritic Epidermal T Cells in Contact Hypersensitivity

Substances that penetrate the skin surface can act as allergens and induce a T cell-mediated inflammatory skin disease called contact hypersensitivity (CHS). IL-17 is a key cytokine in CHS and was originally thought to be produced solely by CD4(+) T cells. However, it is now known that several cell types, including γδ T cells, can produce IL-17. In this study, we determine the role of γδ T cells, especially dendritic epidermal T cells (DETCs), in CHS. Using a well-established model for CHS in which 2,4-dinitrofluorobenzene (DNFB) is used as allergen, we found that γδ T cells are important players in CHS. Thus, more IL-17-producing DETCs appear in the skin following exposure to DNFB in wild-type mice, and DNFB-induced ear swelling is reduced by ∼50% in TCRδ(-/-) mice compared with wild-type mice. In accordance, DNFB-induced ear swelling was reduced by ∼50% in IL-17(-/-) mice. We show that DNFB triggers DETC activation and IL-1β production in the skin and that keratinocytes produce IL-1β when stimulated with DNFB. We find that DETCs activated in vitro by incubation with anti-CD3 and IL-1β produce IL-17. Importantly, we demonstrate that the IL-1R antagonist anakinra significantly reduces CHS responses, as measured by decreased ear swelling, inhibition of local DETC activation, and a reduction in the number of IL-17(+) γδ T cells and DETCs in the draining lymph nodes. Taken together, we show that DETCs become activated and produce IL-17 in an IL-1β-dependent manner during CHS, suggesting a key role for DETCs in CHS.

Delayed Activation of Innate Immunity in Murine Model of Diabetic Wounds: Initiating An Aberrant Inflammatory Response

Introduction: Diabetic wounds are a costly and morbid condition requiring continuous wound care, frequent debridement, and in some cases, amputation. Aberrant immune responses, characterized by delayed infiltration of inflammatory cells, paucity of growth factors and persistence of the inflammatory phase of wound healing, are well-documented manifestations in chronic diabetic wounds. the relationship of these findings has not been well described. Skin resident immune cells function as a first line of defense in the activation of the inflammatory wound-healing cascade. Skin resident γδ T cells, also known as Dendritic Epidermal T cells (DETC), recognize damage-associated molecular patterns. Upon activation, DETC recruit neutrophils and monocytes to the wound bed and stimulate fibroblast and keratinocyte proliferation. Abnormal activation and signaling from these cells would result in poor clearance of pathogens and debris, faulty ECM formation, and delayed keratinocyte migration. We hypothesize that wounds in leptin receptor deficient, diabetic mice will exhibit decreased activation of DETC resulting in atypical macrophage phenotypes. Methods: An excisional wound-healing model was implemented with leptin receptor deficient (db/db) mice and their wild type (WT) littermates. on post-operative days (POD) 3 and 7, wounds were photographed, and skin was harvested. Wound samples were dispersed in an enzyme cocktail to single cell suspension. Cells were stained with two antibody panels for 12-color Flow Cytometry (FC) to identify surface markers for leukocyte subtypes, skin homing, and activation states. Results: the leptin receptor deficient model of Type II Diabetes was validated with serial blood glucose and body mass readings (p<0.05) (n>5; 2-tailed Ttest). Delayed wound healing was observed in the db/db group with widened epithelial gap on photomicroscopy (p<0.05). DETC in wounds of db/db animals had lower levels of surface activation markers CD25 (p=0.03) and CD44 (p=0.02) on POD3. Activation of these cells in skin was comparable to WT on POD7. on POD3, Ly6G+ neutrophils were increased (p=0.006) and activated macrophages (F480+, MHCII+, CD11b high) (p=0.003) were decreased in db/db wounds. Activated macrophages were also decreased in the db/db wound on POD7 (p=0.02). Total wound macrophages were equivocal in both groups on POD3 and POD7. Conclusions: Diabetic mice show an impaired ability to activate DETC upon wounding. Blunted activation of DETC likely leads to delayed recruitment of neutrophils and inflammatory monocytes in the early phases of wound healing. the result is persistent neutrophils and less activated macrophages in the wound on POD3 and 7. Ineffective macrophage activation could lead to poor clearance of wound debris and pathogens as well as altered signaling within the microenvironment of the wound. These findings could explain the improper regulation of the inflammatory wound-healing cascade and the clinical manifestations of chronic diabetic wounds.

Role for E-Cadherin as an Inhibitory Receptor on Epidermal γδ T Cells

E-cadherin is a homophilic adhesion molecule that maintains homotypic intercellular adhesion between epithelial cells such as epidermal keratinocytes. E-cadherin is also expressed on resident murine epidermal γδ T cells, known as dendritic epidermal T cells (DETCs), but they express another receptor for E-cadherin, α(E)(CD103)β(7) integrin, as well. In this study, we analyzed functional differences between E-cadherin-mediated homophilic binding and heterophilic binding of α(E)β(7) integrin to E-cadherin in heterotypic intercellular adhesion of DETCs to keratinocytes. E-cadherin, but not α(E)β(7) integrin, was downregulated on activation of DETCs in vivo and in vitro. Short-term (1-h) adhesion of DETCs to keratinocytes in vitro was primarily mediated by α(E)β(7) integrin, and blocking of the binding of α(E)β(7) integrin to E-cadherin inhibited the lysis of keratinocytes by DETCs. Stable binding of E-cadherin on DETCs to plate-bound recombinant E-cadherin was observed only after 24-h culture in vitro. Cytokine production and degranulation by DETCs in response to suboptimal TCR cross-linking and mitogen stimulation were augmented by coligation of α(E)β(7) integrin. In contrast, engagement of E-cadherin on DETCs with immobilized anti-E-cadherin Ab, plate-bound recombinant E-cadherin, and E-cadherin on keratinocytes inhibited DETC activation. Therefore, E-cadherin acts as an inhibitory receptor on DETCs, whereas α(E)β(7) integrin acts as a costimulatory receptor. Differential expression of E-cadherin and α(E)β(7) integrin on resting and activated DETCs, as well as their opposite functions in DETC activation, suggests that E-cadherin and α(E)β(7) integrin on DETCs regulate their activation threshold through binding to E-cadherin on keratinocytes.

Visualizing the rapid and localized keratinocyte expression of dendritic epidermal T cell antigens in response to wounding (93.3)

Abstract Within the skin, dendritic epidermal T cells (DETC) function at the interface of innate and adaptive immunity, playing important roles in homeostasis, tumor surveillance and wound repair. Central to their role in wound repair is stimulation of DETC through their monoclonal Vγ3Vδ1 T cell receptor (TCR) by an uncharacterized antigen expressed on stressed keratinocytes. While only DETC proximal to the wound site become activated, expression patterns and kinetics of antigens that stimulate DETC activation are uncharacterized. Using soluble DETC TCR tetramers, we show that the DETC TCR binds to a glycosylated, cell surface expressed, protein that is constitutively expressed by transformed epithelial cell lines in vitro. Additionally, in situ studies show that DETC TCR ligands are rapidly upregulated by keratinocytes proximal, but not distal to, wound edges. Following antigen expression, DETC become activated and antigen expression is subsequently down-modulated. Furthermore, inhibition of TCR-ligand interactions using DETC TCR tetramers delays wound repair in vivo, highlighting the importance of rapid, TCR mediated, DETC activation by keratinocytes for proper wound healing. This study represents the first visualization of DETC TCR ligand expression by keratinocytes in murine skin and provides a mechanism for the rapid and localized activation of DETC in response to wounding. Supported by the National Institutes of Health (AI064811 to W.L.H, A142267 to L.T.), and the National Science Foundation Graduate Research Fellowship to H.K.K.

Visualizing skin γδ T cell antigen expression during wound repair

Skin is the largest organ through which we experience the outside world and our first line of defense against environmental and pathogenic assault. Within the skin, dendritic epidermal T cells (DETC) are uniquely poised to monitor the tissue for damage and disease and play key roles in wound repair and homeostasis. Central to the role in wound repair is stimulation of the DETC through their invariant γδ TCR by an unknown antigen expressed on stressed keratinocytes. Characterizing ligands for the DETC TCR is crucial to understanding DETC activation and function. We developed soluble DETC TCR tetramers to investigate expression patterns and kinetics of putative antigens specific for the DETC TCR. In vitro tetramer studies showed stimulatory keratinocyte cell lines express DETC TCR ligands. Ex vivo tetramer staining in wounded epidermal sheets revealed rapid upregulation of antigen on keratinocytes at, but not distal to, the wound site prior to DETC activation followed by a loss of antigen after DETC activation. This study represents the first visualization of DETC antigen expression during wound repair and provides novel information about patterns of antigen expression and early stages of DETC activation during wound repair. Ongoing studies using the DETC TCR tetramer are focused upon identification of the unknown DETC antigen. This work is supported by NIH grants AI064811 to W.L.H, A142267 to L.T, and a NSF GRFP to H.K.K.

Dendritic Epidermal T-Cell Activation

Although gammadelta T cells compose a small proportion of lymphocytes in lymphoid compartments and peripheral blood, they are the major T-cell population present in epithelial tissues. However, the role played by gammadelta TCR expressing intraepithelial lymphocytes (IEL) has been enigmatic. The location of tissue-resident IEL suggests that they are important members of the first line of defense against insult for organs exposed to the environment, including the skin, gut, lungs, and reproductive system. Dendritic epidermal T cells (DETC) are the skin-resident gammadeltaIEL and serve as a model system for gammadeltaIEL in other locations. DETC have demonstrated importance in the modulation of immune responses, surveillance and repair of tissue, and resistance to infection. This work discusses recent developments in understanding DETC activation.

Activation of murine epidermal γδ T cells through surface 2B4

Dendritic epidermal T cells (DETC) are gamma delta T cells that normally reside in murine skin. They express on their surface the 2B4 molecule, a 66-kDa glycoprotein of the immunoglobulin gene superfamily thought to be associated with anti-tumor cytotoxicity by natural killer and lymphokine-activated killer cells. Here, we show that ligation of surface 2B4 transduces cell activation signals in DETC. Treatment with anti-2B4 monoclonal antibodies triggers the secretion of interferon-gamma and interleukin-2 by DETC lines, induces proliferation of resting DETC lines, amplifies anti-CD3-dependent proliferation of DETC freshly isolated from mouse skin; and up-regulates egr-1 and c-fos mRNA expression. These results indicate a unique pathway for DETC activation.

In vivo activation of mouse dendritic epidermal T cells in sites of contact dermatitis

Adult mouse epidermis contains a population of dendritic, Thy-1+, CD3+, CD4-, CD8- and T cell receptor (TcR) V gamma 3/V delta 1+ leukocytes termed dendritic epidermal T cells (DETC). DETC isolated from skin and placed into culture will proliferative vigorously in response to T cell mitogens and T cell growth factors. In the present study, we examined whether DETC can be activated in situ by modulating their epidermal environment. Ear skin of CBA mice was painted with the chemical irritant, croton oil, and the epidermal cells (EC) isolated from such sites were then tested for proliferative responses to exogenous interleukin-2 (IL-2), in the absence of added mitogens. Cells from croton oil-treated skin showed marked IL-2 responsiveness, whereas cells from phosphate-buffered saline-treated skin failed to proliferate. IL-2 responses were seen as early as 2 days after croton oil treatment and peaked between days 5 and 10. gamma delta TcR-bearing cells, most likely resident DETC, were the major population to respond to IL-2, since depletion of gamma delta TcR+ cells, but not alpha beta TcR+ cells, abolished that responsiveness, and since gamma delta TcR+ cell numbers increased markedly in the cultures that contained added IL-2. These results indicate that DETC in normal skin, which are at a state of rest, may be activated when their residential epidermal environment is disrupted externally. This process of DETC activation may be a critical step in the maturation of DETC into effector leukocytes in vivo.

Electric Shock

Substances that penetrate the skin surface can act as allergens and induce a T cell–mediated inflammatory skin disease called contact hypersensitivity (CHS). IL-17 is a key cytokine in CHS and was originally thought to be produced solely by CD4⁺ T cells. However, it is now known that several cell types, including γδ T cells, can produce IL-17. In this study, we determine the role of γδ T cells, especially dendritic epidermal T cells (DETCs), in CHS. Using a well-established model for CHS in which 2,4-dinitrofluorobenzene (DNFB) is used as allergen, we found that γδ T cells are important players in CHS. Thus, more IL-17–producing DETCs appear in the skin following exposure to DNFB in wild-type mice, and DNFB-induced ear swelling is reduced by ∼50% in TCRδ^−/− mice compared with wild-type mice. In accordance, DNFB-induced ear swelling was reduced by ∼50% in IL-17^−/− mice. We show that DNFB triggers DETC activation and IL-1β production in the skin and that keratinocytes produce IL-1β when stimulated with DNFB. We find that DETCs activated in vitro by incubation with anti-CD3 and IL-1β produce IL-17. Importantly, we demonstrate that the IL-1R antagonist anakinra significantly reduces CHS responses, as measured by decreased ear swelling, inhibition of local DETC activation, and a reduction in the number of IL-17⁺ γδ T cells and DETCs in the draining lymph nodes. Taken together, we show that DETCs become activated and produce IL-17 in an IL-1β–dependent manner during CHS, suggesting a key role for DETCs in CHS.