Immunocytochemical detection of the carbohydrate antigen, Sialyl Lewisx, in normal human skin and during irritant contact dermatitis

Abstract Sialys Lewis^x (SLex) is a ligand for the E-selectin and the interaction of E-selectin on the endothelium and SLe^x on T cells may be important for T-cell migration into the skin. We investigated the expression of SLe^x on Langerhans cells (LC) in normal skin and on LC repopulating epidermis deprived of LC due to a preceding irritant contact dermatitis. SLe^x was visualized by fluorescence and light microscopic immunocytochemistry using the monoclonal antibody. CSLEX-1. The results showed that about 40% of LC in normal epidermis express SLe^x. In the repopulation phase, most of the epidermal cells were CDla⁺/SLe^x+. We suggest that SLe^x is present on epidermal LC that have recently immigrated from the dermis.     

Overexpression of protein kinase C-α and -β isozymes by stromal dendritic cells in basal and squamous cell carcinoma

Protein kinase C (PKC) isozymes transduce signals from cell surface receptors and thereby regulate important cellular functions in skin including keratinocyte growth and differentiation. Overexpression of individual PKC isozymes results in aberrant cell growth and in certain instances tumorigenicity. PKC is implicated in tumour promotion in mouse skin. Abnormal expression of PKC has been reported in several human cancers. We have, therefore, investigated expression of PKC-α and -β in basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) by immunohistochemistry. Sections were stained with specific antibodies to PKC-α, PKC-β, CDla, T cells, B cells and dermal dendritic cells (factor XIIIa), using an immunoperoxidase technique, PKC-α and PKC-β were not detected in tumour cells in BCCs or SCCs. In SCCs, PKC-β immunostaining revealed positively stained inflammatory and dendritic cells scattered through the stroma: PKC-α immunostaining was essentially negative. In contrast, in BCCs, PKC-α+ and PKC-β+ dendritic and spindle-shaped cells were observed in the stroma, immediately adjacent to the tumour islands. Double-labelling experiments showed that a proportion (approximately 20%) of PKC-β+ dendritic cells also expressed factor XIIIa. BCCs depend on stroma for growth: PKC regulates expression of type IV collagenase and stromelysis III and interactions between tumour and stroma may be important in determining tumour invasion and metastasis, Therefore, overexpression of PKC-α and -β by stromal dendritic cells in BCCs suggests that PKC activation may be involved in stromal/tumour interactions in these tumours.     

Rapid purification of human Langerhans cells using paramagnetic microbeads

Abstract Detailed studies on the biology of Langerhans cells (LC), which account for only 1-3% of all epidermal cells, require isolation from their cutaneous symbionts. Several techniques of LC isolation have been reported, including positive enrichment with mAb coupled to immuno-magnetic beads. The disadvantage of this technique is the size of the beads (= 2-5 μm), which can interfere with subsequent phenotypic and functional analyses. This limitation prompted us to test wheter paramagnetic microbeads (15 nm) employed by the MACS? system could be used to purify LC from human skin. To isolate fresh LC (fLC), epidermal cell suspensions (EC) were stained with anti-CD la mAb and with appropriate secondary reagents conjugated to microbeads and to FITC. They were then passed over a separation column and exposed to a strong magnetic field. Thereafter both CD la-depleted and CD la-enriched cells were collected. Cultured LC (cLC) were isolated by staining 72-h cultured EC with anti-HLA-DR mAb followed by the same isolation procedure. Using this technique, we could routinely isolate viable EC that were 45-88% CD la or HLA-DR⁺ as determined by FACS. Two-color FACS analysis demonstrated the majority of MACS-purified cells to be CDla⁺/HLA-DR⁺, indicating that they were indeed LC. By transmission electron microscopy (TEM), the MACS-purified CDla⁺/HLA-DR⁺ cells showed typical ultrastructural characteristics of LC. Furthermore, MACS-purified fLC or cLC were functionally intact, because they stimulated the proliferation of alloreactive T cells in a primary, one-way, mixed epidermal cell leukocyte reaction (MECLR). We conclude that MACS-separation is an efficient and rapid method to isolate human fLC and cLC of high purity and unimpaired function.     

Turnover and kinetics of epidermal langerhans cells and their dendritic precursor cells in experimental contact dermatitis

The numerical density of epidermal Langerhans cells (LCs) in contact sensitivity and toxic contact dermatitis is still a matter of controvery, mainly due to changes in the phenotypic markers of this antigen-presenting cell during the skin reactions. Since the electron microscopic detection of Birbeck granules is the most reliable marker for the identification of normal and pathologically altered LCs, we performed an ultrastructural-morphometric time-course analysis to evaluate their epidermal turnover in the earskin of BALB/c mice after painting the ears with the hapten 2,4-dinitrofluorobenzene and the irritant croton oil. The counts revealed degeneration and depletion of epidermal LCs in both allergic and toxic dermatitis. In contrast, a slightly increased number of activated epidermal LCs was found during contact sensitization. All experimental procedures resulted in an enhanced immigration of so-called indeterminate dendritic cells which also became ultrastructurally activated and often showed Birbeck granule-like formations at their cell membrane. Immunohistochemistry with the monoclonal antibody 4F7, a new marker for dendritic precursor cells of LCs, demonstrated a significant increase in these accessory cells in the epidermis. Our results indicate that contact sensitivity and toxic skin reactions are characterized by complex but distinct changes in the turnover, kinetics and cellular properties of epidermal LCs and their dendritic precursor cells. Received: 16 March 1995     

Irritant-induced migration of Langerhans cells coincides with an IL-10-dependent switch to a macrophage-like phenotype

Langerhans cells (LCs) migrate after topical exposure of the skin to irritants, despite the supposed independence of irritant contact dermatitis from adaptive immunity. Whereas allergen-activated LCs are known to migrate to the draining lymph nodes (LNs), the fate of migrated LCs upon topical irritant exposure is unknown. Here, we identified a phenotypic switch of LCs after their migration into the dermis upon irritant exposure. With the aid of ex vivo intact human skin and epidermal sheets, we show that dermal fibroblasts are necessary for an IL-10-dependent postmigrational phenotypic switch of LCs into macrophage-like cells. Exposure of ex vivo skin to a panel of seven irritants resulted in a decrease in the number of CD1a(+) cells and an increase in CD14(+)/CD68(+) cells in the dermis. Neutralizing antibodies against IL-10 totally inhibited the phenotypic LC-to-macrophage transition, but did not influence the migration of CD1a(+) cells. Exposure of epidermal sheets to irritants resulted in a fibroblast-dependent LC-to-CD14(+)/CD68(+) switch coinciding with migration, which could be totally inhibited by neutralizing antibodies against either IL-10 or CCL2/CCL5 (two chemokines responsible for epidermal-to-dermal migration). We have thus identified an IL-10-dependent phenotypic switch of LCs into macrophage-like cells upon irritant exposure and emigration from the epidermis.     

Cytochemical and Ultrastructural Studies of the Langerhans' Cells

Abstract: In the guinea pig, experimental allergic contact dermatitis (ACD) And primary irritant contact dermatitis (PICD) were induced with different concentrations of dinitrochlorobenzene (DNCB). The epidermal Langerhans' cells (LCs) were observed sequentially by both adenosine triphosphatase (ATPase) and electron microscopy. Light microscopically, in ACD, the density and dendritic processes of LC decreased markedly within 12 h after antigen challenge. Almost no recognization LCs could be seen within 2 to 5 days. Later, LCs began to repopulale in the epidermis. Within 14 days, the density and shape of the LCs returned to normal. On the contrary, LCs changed more rapidly in PICD. The dendritic processes of LC decreased within 2 h and cell density decreased dramatically within 6 h after DNCB application. LCs also repopulated more rapidly in the epidermis. Electron microscopically, in ACD, we observed that lymphocyte-like cells apposed to LCs; LCs were activated and damaged; however, in PICD, we found neither the apposition of lymphocyte-like cells to LCs, nor the activation of LCs. LCs play an important role in the convalescence phase as well as in the early and later phases of contact allergic reaction.     

Epidermal Langerhans cell apoptosis is induced in vivo by nonanoic acid but not by sodium lauryl sulphate

Exposure to irritants may cause chronic irritant contact dermatitis (ICD), characterized by irregular epidermal thickening and a predominantly dermal mononuclear cell infiltrate. The mechanisms involved, and why only certain individuals are affected, are not clearly understood. Different irritants may trigger different cellular and molecular interactions between resident skin cells and recruited inflammatory cells. In some individuals these interactions may become self-perpetuating resulting in persistent inflammation in the absence of continued exposure. This study examined Langerhans cell (LC) density in clinically normal skin of 46 patients with chronic ICD and 10 healthy individuals, and compared the action of the two irritants nonanoic acid (NA) and sodium lauryl sulphate (SLS) on the LCs and keratinocytes of clinically normal skin in patients with chronic ICD. There was a higher number of LCs/mm basement membrane in patients compared with controls, although there was no difference in the number of dendrites/LC nor in dendrite length. SLS induced keratinocyte proliferation after 48 h exposure, had no effect on LC number or distribution, and induced keratinocyte apoptosis after 24 and 48 h exposure. In contrast, NA decreased keratinocyte proliferation after 24 h exposure but this returned to basal levels after 48 h, and induced epidermal cell apoptosis after only 6 h exposure. NA dramatically decreased LC number after 24 and 48 h exposure, which was accompanied by basal redistribution and decreased dendrite length. Most significantly, NA induced apoptosis in over half of the LCs present after 24 and 48 h exposure.     

Cytochemical and Ultrastructural Studies of the Langerhans'' cells

In the guinea pig, experimental allergic contact dermatitis (ACD) and primary irritant contact dermatitis (PICD) were induced with different concentrations of dinitrochlorobenzene (DNCB). The epidermal Langerhans' cells (LCs) were observed sequentially by both adenosine triphosphatase (ATPase) and electron microscopy. Light microscopically, in ACD, the density and dendritic processes of LC decreased markedly within 12 h after antigen challenge. Almost no recognization LCs could be seen within 2 to 5 days. Later, LCs began to repopulate in the epidermis. Within 14 days, the density and shape of the LCs returned to normal. On the contrary, LCs changed more rapidly in PICD. The dendritic processes of LC decreased within 2 h and cell density decreased dramatically within 6 h after DNCB application. LCs also repopulated more rapidly in the epidermis. Electron microscopically, in ACD, we observed that lymphocyte-like cells apposed to LCs; LCs were activated and damaged; however, in PICD, we found neither the apposition of lymphocyte-like cells to LCs, nor the activation of LCs. LCs play an important role in the convalescence phase as well as in the early and later phases of contact allergic reaction.     

Effect of genetically determined immunodeficiency on epidermal dendritic cell populations in C57BL/6J mice

Summary Mice homozygous for three different recessive mutations known to cause pleiotropic defects in the immune system and in the skin were used to evaluate the relationship between the classical immune system and dendritic epidermal cell populations. Numbers of Langerhans cells (LCs) and Thy-1⁺ dendritic cells (Thy-1⁺DEC) were determined using indirect immunofluorescence microscopy of epidermal whole mounts taken from viable motheaten (me ^v ), nude (nu), and rhino (rh ^hr ) mice. All mutants were maintained on the C57BL/6J strain background and were compared with their respective littermate normal controls. Viable motheaten mice had normal numbers of LCs at 1 month of age. However, by 8 weeks of age, LC density had decreased threefold. Nude and rhino mice had normal numbers of LCs at all ages tested. There was no significant effect of the viable motheaten mutation on numbers of Thy-1⁺DEC. Although nude mice showed normal numbers of Thy-1⁺ DEC at 1 month of age, these athymic mice had a threefold decrease in numbers of such cells by 6 months. In contrast to the reduced numbers of Thy-1⁺DEC seen in nude mice, rhino mice showed a four- to fivefold increase in the numbers of these epidermal cells at all ages tested. These findings suggest new mouse models for investigating the development, regulation, and biological properties of epidermal dendritic cell populations.This work was supported in part by grants from the US-Israel Binational Foundation for Science, the United States-Israel Bi-national Agricultural Research and Development Fund (BARD), the Foundation for the Study of Cell Biology and Molecular Virology, Phoenix, Arizona and United States Public Health Service grant CA-20408. E. S. is a fellow of the Foulkes Foundation, London, UK     

Quantitative and 3-dimensional analysis of Langerhans cells in basal cell carcinoma. A comparative study using light microscopy and confocal laser scanning microscopy

Summary We have analysed Langerhans cells (LCs) in basal cell carcinoma (BCC) and in healthy skin in 15 patients, using three different techniques: light microscopic examination of horizontal sheets, and of 6-μm-thick vertical skin sections, and confocal laser scanning microscopy (CLSM) of 25-μm-thick vertical sections. The use of CLSM enables both a quantitative and a three-dimensional (3-D) analysis of the cells in the same tissue volume. A statistically significant reduction in the relative volume of epidermal CDla reactivity confined to tumour areas was found with CLSM. This difference was confirmed when the number of LCs in horizontal sheets were counted. In contrast, no significant reduction in epidermal CDla⁺ cells was found in thin vertical sections. This is probably due to the smaller tissue sample examined, and to variations in the number of CDla⁺ cells, with less cells directly overlying the tumour nests. The ratio of CDla-expressing cells in the epidermis/dermis was significantly reduced in BCCs, compared with healthy looking skin. Few LCs were observed in tumour nests, but they were numerous in the surrounding stroma of the dermis. Three-dimensional reconstructions of CDla⁺ cells in BCC revealed striking morphological changes; they had a reduced number of dendrites, and these were often short and had few branches. The results demonstrate that CLSM is a suitable technique for quantitative and morphological analysis of CDla-expressing cells in the skin. We suggest that the alterations in LC numbers, distribution and morphology in BCC most probably are secondary to changes in the local environment.     

The effect of ageing on phenotype and function of monocyte-derived Langerhans cells

Background With increasing age the immune system shows functional decline. In the skin this is associated with an increased incidence of epidermal malignancies and infections. Epidermal Langerhans cells (LCs) act as sentinels of the immune system, recognizing, processing and presenting antigen and inducing T‐cell responses. Previous investigations have demonstrated a reduction in the number of epidermal LCs in elderly subjects. Moreover, the ability of LCs to migrate in response to tumour necrosis factor (TNF)‐α, but not interleukin (IL)‐1β, is significantly impaired in the elderly. Objectives To characterize further the changes in LC function that are associated with increasing chronological age, we have evaluated age‐related changes in the response of monocyte‐derived LCs (mLCs) to IL‐1β and TNF‐α. Methods The phenotype and function of mLCs were compared in six young (≤ 30 years) and six aged (≥ 70 years) healthy individuals using a combination of flow cytometry, cytokine and chemokine array, and a Transwell migration assay. Results Monocytes from aged individuals were able to differentiate into LCs. There were no significant differences in expression of activation markers, or in baseline or inducible cytokine secretion, by mLCs derived from aged or young subjects. Furthermore, migration in response to a chemokine ligand, CCL19, was equivalent in both age groups. Conclusions These data demonstrate that changes in LC function in the elderly are not associated with changes in systemic dendritic cell phenotype and function. Conditioning of LCs in situ by the epidermal microenvironment is likely to be more important.     

Subpopulation of murine epidermal Langerhans cells identified by lectin-binding sites

Summary Lectin-binding profiles of epidermal Langerhans cells (LCs) were investigated in three strains of mice using immunofluorescence procedures. Three lectin-binding profiles were observed in each strain of mice. Most epidermal LCs reacted with concanavalin A (Con A) and Ricinus communis agglutinin 1 (RCA-1), whereas none reacted with Dolichos biflorus agglutinin (DBA). Peanut agglutinin (PNA) and wheat germ agglutinin (WGA) reacted with some of the epidermal LCs. These binding profiles were similar from site to site of the body in all strains of mice. We also investigated the lectin-binding profiles of epidermal Ia positive (Ia⁺) cells migrating into the grafted skin up to 165 days after transplantation. BALB/c (H-2^d) murine skin was grafted onto the back of (C3H/He×BALB/c)F1 (H-2^k×H-2^d) mice. The percentages of migrating I-A⁺ epidermal cells reactive with PNA and WGA were different from those of the normal epidermis soon after grafting and reached a normal level at 43 days after grafting. Our results demonstrated that there is a heterogeneous population of epidermal LCs defined by lectin-binding profiles.     

Murine epidermal Langerhans cells and keratinocytes express functional P2X7 receptors

Please cite this paper as: Murine epidermal Langerhans cells and keratinocytes express functional P2X₇ receptors. Experimental Dermatology 2010; 19 : e151–e157. Abstract: Extracellular ATP via the activation of purinergic P2 receptors has an emerging role in cutaneous biology; however, the distribution of these receptors in mouse skin is poorly defined. This study investigated whether murine epidermal cell subpopulations express functional purinergic P2X₇ receptors. P2X₇ expression was examined by immunoblotting and immunofluorescence staining of epidermal cells from C57Bl/6 mice. P2X₇ function was evaluated by nucleotide‐induced ethidium⁺ uptake measurements in epidermal cells from C57Bl/6 mice, and from P2X₇ deficient mice and wild‐type littermate controls. P2X₇ was detected in whole epidermal cell preparations, and specifically on Langerhans cells (LCs) and keratinocytes (KCs). ATP induced ethidium⁺ uptake into LCs and KCs, with EC₅₀ values of 503 and 482 μm, respectively. BzATP, and to a lesser extent ATPγS and ADP, also induced ethidium⁺ uptake; while UTP, αβ‐meth‐ATP and NAD were ineffective. ATP‐induced ethidium⁺ uptake was impaired by Na⁺ and Mg²⁺, and the P2X₇ antagonist, A‐438079 and was absent in LCs and KCs from P2X₇ deficient mice. These results demonstrate that murine LCs and KCs express functional P2X₇, and support a role for this receptor in cutaneous biology.     

Selective Expansions of T cells Expressing Vβ38 and Vβ13 in Skin Lesions of Patients with Chronic Cutaneous Lupus Erythematosus

There are several clinical types of cutaneous lupus erythematosus (LE), including acute cutaneous LE (ACLE), which occurs in 50–60% of patients with systemic LE (SLE), chronic cutaneous LE (CCLE), which is almost the same as discoid LE (DLE), and subacute cutaneous LE (SCLE). Although several important hypotheses have been proposed to explain cutaneous LE, the pathomechanisms still remain complicated and obscure. Of special interest is whether and how the T cell receptor (TCR) repertoire of infiltrating lymphocytes is involved in the development of the different types. To address this issue, we immunohistochemically examined the Vβ usage of infiltrating T cells in skin lesions, as well as in peripheral blood mononuclear cells (PBMC) of patients with cutaneous LE. The number of Vβ3.1 CD3⁺ cells in the PBMC of patients with ACLE and CCLE was significantly lower than in controls. In contrast, the number of Vβ3.1 CD3⁺ cells was elevated in the skin lesion of CCLE over that in psoriasis vulgaris or atopic dermatitis. Furthermore, skin lesions in CCLE patients showed a higher incidence of Vβ8.1 CD3⁺ and Vβ13.3 CD3⁺ cells than did those in ACLE patients. These results suggest that skin lesions of CCLE are oligoclonally associated with selective expansions of TCR Vβ chains and may be induced by antigen stimuli, including superantigens.     

Activated immunocompetent cells in human skin lymph derived from irritant contact dermatitis: an immunomorphological study

By means of microsurgical lymph cannulation, skin lymph was sampled in the course of a sodium lauryl sulphate (SLS)-induced irritant contact dermatitis in human volunteers. The lymph cells were isolated by centrifugation, and then characterized immunocytochemically using different monoclonal antibodies, and in the late phase of the skin reaction also by electron microscopy. Analyses of lymph cells before the induction of the contact dermatitis revealed median values of about 60% T cells (CD4/CD8 ratio about 2:1), 4% Langerhans cells (LCs), and 1% B cells. The remainder were varying proportions of erythrocytes and uncharacterized cells. During the skin reaction, and even after resolution of the clinical signs of dermatitis, a relative and absolute increase of T and B cells, as well as of HLA-DR positive cells, paralleled the previously reported increase of LCs; a high percentage of the T cells were CD4 and CD8 negative. In addition, surface markers such as CD11a, CD25, CD54 and CD58 were detected on lymph cells sampled during the irritant skin reaction. Cell rosettes observed in the lymph throughout the experiment were analysed in the late phase of the skin reaction, and showed a central LC with three to five peripheral, in part activated, T cells, ultrastructurally revealing gap junction-like structures between the two cell types. These data indicate that immunocompetent cells in the skin are activated by a variety of non-immunological stimuli such as operative trauma and irritant contact dermatitis.     

Antigen-presenting capacity in normal human dermis is mainly, subserved by CDla+ cells

A proposed role for antigen-presenting dermal dendrocytes in the pathogenesis of many dermal inflammatory skin diseases remains speculative. We therefore sought to determine the phenotype and functional characteristics of antigen-presenting cells isolated from normal human dermis. Normal adult human skin was incubated overnight with dispase at 4°C, the epidermis was removed, and the residual dermal preparation was then minced and digested with a mixture of hyaluronidase, collagenase, and DNAase at 37°C, prior to filtration through mesh. Dermal cell suspensions thus obtained were stained using specific monoclonal antibodies, and analysed by fluorescence micro- scopy or flow cytometry. Mean values were as follows: CD45⁺ leucocytes 39%, HLA-DR⁺ cells 39%, Ulex europaeus agglutinin I⁺ endothelial cells 26%, CD1a⁺ cells 3.9%, CD11b⁺ cells 16%, CDllc⁺ cells 6%. Mitomycin C-treated crude dermal cell suspensions induced allostimulation of peripheral blood mononuclear cells in a 7-day culture, as assessed by ³H-TdR incorporation. Depletion of CDla⁺ Langerhans-like cells from the dermal cell preparation, by 95, 74 and 90% in three separate experiments using immunomagnetic beads, reduced ³H-TdR incorporation at optimal responder-to- stimulator cell ratios by 90, 64, and 87%, respectively. Our findings suggest that, in normal human dermis, the great majority of the alloantigen-presenting capacity resides in the CDla⁺ Langerhans cell-like dendritic antigen-presenting cell population, and not to any great extent in either CDla^? macrophage-like cells, or HLA-DR⁺ endothelial cells. The relationship of the CDla⁺ dermal antigen presenting cells to the Langerhans cell lineage remains to be determined.     

CD70-CD27 interaction augments CD8+ T-cell activation by human epidermal Langerhans cells

Human cutaneous dendritic cells (DCs) from epidermal and dermal compartments exhibit functional differences in their induction of CD4+ T-cell and humoral immune responses; however, differences in the regulation of memory CD8+ T-cell responses by human skin DCs remain poorly characterized. We tested the capacity of human Langerhans cells (LCs) and dermal dendritic cells (DDCs) to induce antigen-specific cytokine production and proliferation of memory CD8+ cells. Although tumor necrosis factor-α-matured human DCs from both epidermal and dermal compartments showed efficient potential to activate CD8+ cells, LCs were constitutively more efficient than DDCs in cross-presenting CD8+ epitopes, as well as direct presentation of viral antigen to Epstein-Barr virus-specific CD8+ T cells. LCs showed greater expression of CD70, and blockade of CD70-CD27 signaling demonstrated that superiority of CD8+ activation by epidermal LC is CD70 dependent. This CD70-related activation of CD8+ cells by LCs denotes a central role of LCs in CD8+ immunity in skin, and suggests that regulation of LC CD70 expression is important in enhancing immunity against cutaneous epithelial pathogens and cancer.     

Defined in situ enumeration of T6 and HLA-DR expressing epidermal Langerhans cells: morphologic and methodologic aspects

An essential prerequisite for the in situ enumeration of epidermal Langerhans cells (LCs) is the unequivocal identification of the desired cell type. We have examined over 250 cryostat sections of normal human skin to analyze morphologic and methodologic problems underlying the quantification of epidermal LCs, defined by anti-T6 (OKT6) and anti-HLA-DR (OKIal) immunoperoxidase staining. Our findings show that OKT6 reactivity of dendritic processes in cross-sectioned epidermis yields microscopic images which are not easy to analyze objectively. The morphology that we find leads us to categorize dendritic cells into 3 arbitrary types of T6+ LC profiles. In addition we describe criteria for the assessment of OKT6 staining patterns relating to the dendritic state of epidermal LCs. Preliminary quantitative data on this issue are discussed in relation to: epidermal thickness; the thickness of skin tissue sections; and the discrepancy between the number of T6+ and HLA-DR+ LCs. We hope that the principles outlined in this report may serve to overcome potential methodologic problems with quantitation of T6+ epidermal LCs in skin sections.     

Epidermal Langerhans' cells in chronic eczematous dermatitis of the palms treated with PUVA and UVB

Epidermal Langerhans' cells (LC) were studied in patients with chronic eczematous dermatitis of the palms. The monoclonal antibodies anti-Leu 6 and anti-HLA-DR were used, and the cells visualized with an immunoperoxidase technique. Increased numbers of LC were found in allergic contact dermatitis as well as in irritant contact dermatitis and hyperkeratotic dermatitis of the palms. The increased number of epidermal LC in lesional skin may facilitate presentation of exogenous or endogenous antigens to activated T-cells. One hand was treated with PUVA or UVB and the other hand served as a non-exposed control. PUVA treatment cleared the dermatitis and the LC number decreased markedly. With UVB treatment clinical improvement was achieved, and a less pronounced decrease in epidermal LC was noticed. There seems to be a crude relationship between the extent of clinical improvement and the reduction in epidermal LC numbers. The change in LC might be a primary event or secondary to a reduction of the inflammatory process.