Dendritic Epidermal T Cells Research Articles

Skin-resident T cells limit UV damage through sensing the danger signal ATP (P3246)

Abstract Exposure to ultraviolet radiation (UVR) is a major risk factor for skin cancers and skin inflammation. However, molecular and cellular mechanisms to minimize UV damage in the skin are incompletely understood. Based on their sentinel function, we hypothesized that resident T cells in the skin, murine dendritic epidermal T cells (DETC) and human skin-resident T cells, sense UVR-induced danger signals such as extracellular adenosine triphosphate (ATP) to initiate protection and repair. Supporting this hypothesis, we find that epidermal keratinocytes release ATP upon UVR to activate DETC and human skin-resident T cells in a direct and IL-1β-mediated manner and this response could be inhibited by ATPases. Using mice deficient in DETC, we show a previously unrecognized role for DETC in repairing keratinocyte DNA damage upon UVR. Functionally, ATP enhanced TCR-mediated DETC proliferation and IL-17A production. IL-17A induced molecules relevant to DNA repair and apoptosis in keratinocytes, including TNF related weak inducer of apoptosis (TWEAK) and Growth arrest and DNA damage gene (GADD45), providing a possible link to the observed UV protective mechanisms exerted by DETC. Ongoing studies quantifying UVR-induced DNA damage and apoptosis in wildtype, TCRδ-/-, and IL-17A-/- skin cells and in human skin cultures will provide novel information on skin-resident T cell function, and may identify new targets to treat and prevent sunburn and skin cancer in humans.

CCR6 Is Required for Epidermal Trafficking of γδ-T Cells in an IL-23-Induced Model of Psoriasiform Dermatitis

A subset of CCR6+, γδ-low (GDL) T cells that express Th17 cytokines in mouse skin participates in IL-23-induced psoriasisform dermatitis. We use CCR6-deficient (KO) and wildtype (WT) mice to analyze skin trafficking patterns of GDL T cells and function-blocking mAbs to determine the role of CCR6 in IL-23-mediated dermatitis. Herein, CCL20 was highly upregulated in IL-23-injected WT mouse ear skin as early as 24 hours after initial treatment, and large numbers of CCR6+ cells were observed in the epidermis of IL-23-injected WT mice. Anti-CCL20 mAbs reduced psoriasiform dermatitis and blocked recruitment of GDL T cells to the epidermis. In CCR6 KO mice, GDL T cells failed to accumulate in the epidermis after IL-23 treatment, but total numbers of GDL T cells in the dermis of WT and CCR6 KO mice were equivalent. There was a ~70% reduction in the proportion of IL-22+ GDL T cells in the dermis of CCR6 KO mice (vs. WT mice), suggesting that effector function as well as epidermal recruitment of GDL T cells are impaired in CCR6-deficient mice. Thus, these data show CCR6 regulates epidermal trafficking of γδ T cell subsets in skin and suggest the potential of CCR6 as a therapeutic target for psoriasis.

Migratory properties of skin-resident γδ T cell populations (49.20)

Abstract γδ T cells preferentially reside in epithelial tissues like skin. Dendritic epidermal T cells (DETC) in epidermis have been studied extensively. Recently, a novel IL-17-producing γδ T cell population in dermis was identified and demonstrated to have pivotal role in skin inflammation. Nevertheless, their migration patterns are largely undefined. In this study, we found that almost 30% γδ T cells in dermis, lymph nodes (LNs) and spleen are Vγ2+. This proportional distribution led us to hypothesize that γδ T cells may re-circulate between dermis and secondary lymphoid tissues. To test this hypothesis, we created GFP+:GFP- parabiotic mice, and demonstrated that dermal γδ T cells are constantly circulating, whereas DETC are stationary and non-recirculating. We next focused on skin homing of dermal γδ T cells and found that these γδ T cells were not diminished in lymphotoxin α-/- mice (lacking LNs), suggesting that dermal γδ T cells can be stably maintained after seeding from perinatal thymus. Further study revealed that dermal γδ T cells express high levels of CD69, CD103, and E- and P-selectin ligands, all of which are required for mature T cell migration to skin. However, in E- and P-selectin ligand-deficient (Fuc T IV/VII-/-) mice, dermal γδ T cells are paradoxically increased, while DETC are remarkably diminished. Taken together, these findings may help better define the role of γδ T cells in peripheral epithelial tissues.

Skin-derived γδ T cells as a source of IL-17 in the lymph nodes and a possible initiator in contact hypersensitivity (121.22)

Abstract In the murine skin, there exist two subsets of γδ T cells, Vγ5+ dendritic epidermal T cells (DETCs) and Vγ4+ dermal γδ T cells. It remains unclarified about the role of skin γδ T cells in contact hypersensitivity and their homeostasis. Initially, to evaluate the mobility of cutaneous γδ T cells, we used kaede-transgenic mice which express photoconvertible fluorescence protein “kaede”. By exposing the skin to violet light, cutaneous cells changes its fluorescence from green to red. Using this labeling system, we demonstrated that γδ T cells migrated from the skin to draining lymph nodes (DLNs) even in the steady state and such transmigration was enhanced after hapten-application. Skin-derived γδ T cells were mainly Vγ4+ cells, suggesting that they were dermal origin. In addition, this subset produced a higher amount of IL-17 than either LN-resident γδ T cells or αβ T cells migrating from the skin. In addition, we found that γδ T cells assisted αβ T cells proliferation and production of IFN-γ and IL-17 in vitro. These data suggest that the main source of IL-17 in the DLN is γδ T cells from the dermis, and this population may play an important role to establish contact hypersensitivity response through modulating αβ T cell functions.

Cutting Edge: Dendritic Epidermal γδ T Cell Ligands Are Rapidly and Locally Expressed by Keratinocytes following Cutaneous Wounding

TCR-specific activation is pivotal to dendritic epidermal T cell (DETC) function during cutaneous wound repair. However, DETC TCR ligands are uncharacterized, and little is known about their expression patterns and kinetics. Using soluble DETC TCR tetramers, we demonstrate that DETC TCR ligands are not constitutively expressed in healthy tissue but are rapidly upregulated following wounding on keratinocytes bordering wound edges. Ligand expression is tightly regulated, with downmodulation following DETC activation. Early inhibition of TCR-ligand interactions using DETC TCR tetramers delays wound repair in vivo, highlighting DETC as rapid responders to injury. To our knowledge, this is the first visualization of DETC TCR ligand expression, which provides novel information about how ligand expression impacts early stages of DETC activation and wound repair.

Rank Signaling Links the Development of Invariant γδ T Cell Progenitors and Aire+ Medullary Epithelium

SummaryThe thymic medulla provides a specialized microenvironment for the negative selection of T cells, with the presence of autoimmune regulator (Aire)-expressing medullary thymic epithelial cells (mTECs) during the embryonic-neonatal period being both necessary and sufficient to establish long-lasting tolerance. Here we showed that emergence of the first cohorts of Aire+ mTECs at this key developmental stage, prior to αβ T cell repertoire selection, was jointly directed by Rankl+ lymphoid tissue inducer cells and invariant Vγ5+ dendritic epidermal T cell (DETC) progenitors that are the first thymocytes to express the products of gene rearrangement. In turn, generation of Aire+ mTECs then fostered Skint-1-dependent, but Aire-independent, DETC progenitor maturation and the emergence of an invariant DETC repertoire. Hence, our data attributed a functional importance to the temporal development of Vγ5+ γδ T cells during thymus medulla formation for αβ T cell tolerance induction and demonstrated a Rank-mediated reciprocal link between DETC and Aire+ mTEC maturation.

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.

Cutting edge: Identification of a motile IL-17-producing gammadelta T cell population in the dermis.

Dendritic epidermal T cells (DETCs) are a well-studied population of γδ T cells that play important roles in wound repair. In this study, we characterize a second major population of γδ T cells in the skin that is present in the dermis. In contrast to DETCs, these Vγ5-negative cells are IL-7R(hi)CCR6(hi) retinoic acid-related orphan receptor γt(+) and are precommitted to IL-17 production. Dermal γδ T cells fail to reconstitute following irradiation and bone marrow transplantation unless the mice also receive a transfer of neonatal thymocytes. Real-time intravital imaging of CXCR6(GFP/+) mouse skin reveals dermal γδ T cells migrate at ∼4 μm/min, whereas DETCs are immobile. Like their counterparts in peripheral lymph nodes, dermal γδ T cells rapidly produce IL-17 following exposure to IL-1β plus IL-23. We have characterized a major population of skin γδ T cells and propose that these cells are a key source of IL-17 in the early hours after skin infection.

Functions of skin-resident γδ T cells

The murine epidermis contains resident T cells that express a canonical γδ TCR and arise from fetal thymic precursors. These cells are termed dendritic epidermal T cells (DETC) and use a TCR that is restricted to the skin in adult animals. DETC produce low levels of cytokines and growth factors that contribute to epidermal homeostasis. Upon activation, DETC can secrete large amounts of inflammatory molecules which participate in the communication between DETC, neighboring keratinocytes and langerhans cells. Chemokines produced by DETC may recruit inflammatory cells to the epidermis. In addition, cell-cell mediated immune responses also appear important for epidermal-T cell communication. Information is provided which supports a crucial role for DETC in inflammation, wound healing, and tumor surveillance.

Id3 is a critical regulator of γδ-T cell development (153.41)

Abstract αβ and γδ T cells are thought to arise from a common precursor during development in the thymus. Previously, we have shown that αβ/γδ lineage choice is dictated by differences in T cell receptor (TCR) signal strength defined by differential activation of the ERK-Egr-Id3 pathway. In particular, we found that Id3 is required for strong TCR signals to both promote adoption of the γδ-fate and oppose the αβ-fate outcome. Importantly, the effects of Id3-deficiency on γδ T cell development differ depending on the γδ subset. While Id3-deficiency impairs the development of the Vγ2-expressing and Vγ3 dendritic epidermal T cell (DETC) subsets, it results in the dramatic expansion of Vγ1.1Vδ6.3 T cells which are thought to be innate type NK γδ T cells. Fetal thymic organ culture analysis indicates that Id3-deficiency blocks DETC development by interfering with selection in the thymus, not by blocking expression of the canonical DETC TCR complex. Conversely, single-cell PCR analysis has revealed that the junctional diversity of CDR3s of the Vγ1.1Vδ6.3 expressing cells that are expanded in Id3-deficient mice exhibit evidence of ligand-mediated selection. These results are consistent with the notion that development of these cells is normally restricted by ligand-engagement, but this restriction is disabled in the absence of Id3. Altogether, these results indicate that Id3 plays a critical role not only in regulating fate selection, but also in shaping the γδ TCR repertoire.

Cutaneous immunosurveillance by self-renewing dermal γδ T cells

The presence of γδ T cell receptor (TCR)–expressing cells in the epidermis of mice, termed dendritic epidermal T cells (DETCs), is well established. Because of their strict epidermal localization, it is likely that DETCs primarily respond to epithelial stress, such as infections or the presence of transformed cells, whereas they may not participate directly in dermal immune responses. In this study, we describe a prominent population of resident dermal γδ T cells, which differ from DETCs in TCR usage, phenotype, and migratory behavior. Dermal γδ T cells are radioresistant, cycle in situ, and are partially depend on interleukin (IL)-7, but not IL-15, for their development and survival. During mycobacterial infection, dermal γδ T cells are the predominant dermal cells that produce IL-17. Absence of dermal γδ T cells is associated with decreased expansion in skin draining lymph nodes of CD4+ T cells specific for an immunodominant Mycobacterium tuberculosis epitope. Decreased CD4+ T cell expansion is related to a reduction in neutrophil recruitment to the skin and decreased BCG shuttling to draining lymph nodes. Thus, dermal γδ T cells are an important part of the resident cutaneous immunosurveillance program. Our data demonstrate functional specialization of T cells in distinct microcompartments of the skin.

Skint-1 is a highly specific, unique selecting component for epidermal T cells.

αβ T-cell repertoire selection is mediated by peptide-MHC complexes presented by thymic epithelial or myeloid cells, and by lipid-CD1 complexes expressed by thymocytes. γδ T-cell repertoire selection, by contrast, is largely unresolved. Mice mutant for Skint-1, a unique Ig superfamily gene, do not develop canonical Vγ5Vδ1(+) dendritic epidermal T cells. This study shows that transgenic Skint-1, across a broad range of expression levels, precisely and selectively determines the Vγ5Vδ1(+) dendritic epidermal T-cell compartment. Skint-1 is expressed by medullary thymic epithelial cells, and unlike lipid-CD1 complexes, must be expressed by stromal cells to function efficiently. Its unusual transmembrane-cytoplasmic regions severely limit cell surface expression, yet increasing this or, conversely, retaining Skint1 intracellularly markedly compromises function. Each Skint1 domain appears nonredundant, including a unique decamer specifying IgV-domain processing. This investigation of Skint-1 biology points to complex events underpinning the positive selection of an intraepithelial γδ repertoire.

Neutrophils and natural killer T cells as negative regulators of wound healing

Cutaneous wound healing is a dynamic process requiring coordination of immune cell infiltration, keratinocyte proliferation, angiogenesis, extracellular matrix deposition and remodeling to ensure adequate wound closure [1–3]. Multiple pathologic conditions are known to perturb efficient cutaneous repair by impacting one or more of the three interdependent phases of wound healing: the early inflammatory, proliferative, or late remodeling phase [1,2,4,5]. Innate immune cells are critical during wound repair, as innate immune cell dysfunction has been demonstrated to augment normal wound healing, leading to poor wound closure or hypertrophic scar formation. While several immune cells, in particular macrophages, dendritic epidermal T cells (DETCs) and mast cells, have all been demonstrated to enhance aseptic wound healing [6–15], two members of the innate immune population, neutrophils and natural killer T (NKT) cells, act to negatively regulate the pace of wound closure. Herein, we focus our discussion on the negative regulatory role of these neutrophils and NKT cells, and consider potential therapeutic interventions that may enhance wound repair by alterations in the number or function of these populations.

Embryonic trafficking of gammadelta T cells to skin is dependent on E/P selectin ligands and CCR4.

Dendritic epidermal T cells (DETC) express an invariant Vgamma3/Vdelta1 T-cell receptor, appear in fetal epidermis, and form a population of resident epidermal T cells. Their temporal development in the thymus has been studied extensively. However, little is known about the mechanisms involved in the embryonic trafficking of DETC from thymus to epidermis. We demonstrate that DETC in adult skin, as well as the DETC precursors in fetal thymus, express E and P selectin ligands (E- and P-lig). Mice deficient in alpha1,3 fucosyltransferases IV and VII (FTIV/VII) cannot synthesize the carbohydrate motifs that form key elements of these selectin ligands. The numbers of DETC in the epidermis of FTIV/VII(-/-) mice were dramatically reduced compared with normal mice. However, the development of DETC precursors in fetal thymus was identical in normal and FTIV/VII(-/-) mice, suggesting that the DETC precursors produced in FTIV/VII(-/-) mice could not traffic effectively to skin because they lack E- and P-lig. We tested this hypothesis by daily injection of blocking antibodies against E and P selectin into pregnant mice. Mice born from dams treated with anti-selectin antibodies, but not those born from dams treated with isotype control, had significantly diminished numbers of DETC. To test the role of chemokine receptors in DETC skin homing, we examined skin from CCR4(-/-) and CCR10(-/-) mice, respectively. DETC were significantly reduced in CCR4(-/-) mice but were present at normal levels in CCR10(-/-) mice. Our results present evidence for the crucial role of trafficking molecules in embryonic migration of DETC precursors to skin.

Id3 is a critical mediator of lineage commitment and functional maturation of γδ-T cells (36.63)

Abstract αβ and γδ T cells are thought to arise from a common precursor during development in the thymus. However, the basis for separation of these lineages is poorly understood. Previously, we showed that strong TCR signals promote γδ lineage choice, while weaker signals favor adoption of the αβ lineage. Here we show that the HLH factor Id3 plays a defining role in this process. We found that Id3 is required for strong TCR signals to both promote adoption of the γδ-fate and oppose the αβ-fate outcome. Importantly, the effects of Id3-deficiency on γδ lineage cells appear to differ depending on the γδ subset. In Id3-/- mice, development of the Vγ2 subset and Vγ3 dendritic epidermal T cell (DETC) subset are both reduced, while innate type NK γδ T cells bearing a Vγ1.1/Vδ6.3 TCR are dramatically expanded. Our analysis suggests that the γδ subsets whose development is impaired by Id3-deficiency are dependent on TCR signals of intermediate intensity. In contrast, those which are expanded (Vγ1.1Vδ6.3) in Id3-deficient mice may normally be deleted, perhaps by high-affinity ligand. In support this view, KN6 γδ TCR transgenic thymocytes normally deleted by high affinity ligand (T-10/22b) are rescued by Id3-defciency. Likewise, the junctional sequences of the TCR subunits of Vγ1.1Vδ6.3 T cells in Id3-/- exhibited almost no variation, suggesting strong selective pressure. These results indicate that Id3 not only in regulates fate selection, but also in shapes the γδ TCR repertoire.

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.

Integrin αE(CD103)β7 influences cellular shape and motility in a ligand-dependent fashion

AbstractWhile the extravasation cascade of lymphocytes is well characterized, data on their intraepithelial positioning and morphology are scant. However, the latter process is presumably crucial for many immune functions. Integrin αE(CD103)β7 has previously been implicated in epithelial retention of some T cells through binding to E-cadherin. Our current data suggest that αE(CD103)β7 also determines shape and motility of some lymphocytes. Time-lapse microscopy showed that wild-type αE(CD103)β7 conferred the ability to form cell protrusions/filopodia and to move in an amoeboid fashion on E-cadherin, an activity that was abrogated by αE(CD103)β7-directed antibodies or cytochalasin D. The αE-dependent motility was further increased (P < .001) when point-mutated αE(CD103) locked in a constitutively active conformation was expressed. Moreover, different yellow fluorescent protein–coupled αE(CD103) species demonstrated that the number and length of filopodia extended toward purified E-cadherin, cocultured keratinocytes, cryostat-cut skin sections, or epidermal sheets depended on functional αE(CD103). The in vivo relevance of these findings was demonstrated by wild-type dendritic epidermal T cells (DETCs), which showed significantly more dendrites and spanned larger epidermal areas as compared with DETCs of αE(CD103)-deficient mice (P < .001). Thus, integrin αE(CD103)β7 is not only involved in epithelial retention, but also in shaping and proper intraepithelial morphogenesis of some leukocytes.

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 Cells Promote Wound Healing by Production of Vascular Endothelial Growth Factor Mediated by HIF‐1α Signaling

Local hypoxia is general consequence of an acute skin wound due to vascular disruption and high oxygen consumption by cells at the edge of the wound. The resident γδ T cells in the skin, Dendritic Epidermal T Cells (DETC), are known to support local tissue homeostasis and wound repair by local production of growth factors, such as keratinocyte growth factors (KGFs) and insulin‐like growth factor (IGF)‐1. We have analyzed the functional role of DETC in response to local tissue hypoxia generated by an acute skin wound. The hypoxic status of the skin wound induced HIF‐1α expression in DETC. HIF‐1α signalling during hypoxia induced expression of vascular endothelial growth factor (VEGF) by DETC. Wound healing was impaired after HIF‐1α gene‐silencing in DETC and this impairment was rescued by VEGF administration. Together this data demonstrates that acute skin wound induces HIF‐1α signaling in DETC, which promote VEGF production with the result of enhancing skin wound healing.Grant: NIH‐AI‐36964