Densities,distribution and phenotypic expression of T cells in human fetal skin

T cells are present in normal adult human skin, but their occurrence in fetal skin is unknown. T cell and Langerhans cell (LC) populations were studied using single or double immunohistochemical staining on cryostat-section. Skin samples taken from different body regions of 17 fetuses ranging from 18 to 30 weeks estimated gestational-age (w-EGA), were examined. In all specimens but one, we did not find any epidermal T cell. In contrast, dermal CD3⁺ T cells occurred at all w-EGA. The density of these cells increased with increasing age. Double staining showed that CD3⁺ T cells were predominantly CD4⁺/CD45RA⁺. On the other hand, LC, as assessed by CD1a expression, was evenly distributed within the interfollicular epidermis and papillary dermis at all gestational ages. Analysis of T cell and LC density in different body regions did not show significant topographic differences. We suggest that lack of epidermal T cells, although the LC network was fully represented, might reflect the scarce opportunity of fetal LC to contact foreign antigens in utero.     

Modulation of Ia+ Langerhans cell numbers in vivo by cultured epidermis derived supernatants and by GM-CSF

Abstract This paper demonstrates that epidermal cells in culture produce an activity which can increase the frequency of Ia⁺ epidermal Langerhans cells (LC). This was achieved by treating mice topically with a mixture containing supernatant derived from primary culture of murine epidermis (ES) and a synthetic corticosteroid, triamcinolone acetonide (TAG). The presence of the supernatant in the mixture partially protected the Ia⁺ LC from depletion by the steroid. The Ia⁺ LC frequency increasing activity was measured as the difference between the Ia⁺ LC frequency due to treatment with steroid mixed with supernatant and the Ia⁺ LC frequency due to treatment with steroid mixed with negative control medium. The mean frequency of Ia⁺ LC in epidermis treated with TAC mixed with ES was 606(SD 43) cells/mm², as compared with 486 (SD 68) cells/mm² in the epidermis treated with TAC mixed with control medium. The activity appeared to be caused by (a) proteinaceous factor(s). A fraction of ES which was retained above a ≥10 KDa molecular weight cut-off membrane was capable of partially protecting Ia⁺ LC frequency from TAC depletion. Supernatants from cultured lymph nodes, dermis as well as the squamous cell carcinoma lines T7 and T79, but not the human osteosarcoma cell-line 143B, also contained similar activities. We demonstrate that GM-CSF also increased the number of Ia⁺ epidermal LC when applied topically to mouse skin in this system. Therefore, using this Ia⁺ LC frequency modulation system, we propose that GM-CSF is one example of a cytokine which may be involved in the regulation of Ia⁺ LC numbers in epidermis and that epidermal cells produce factors which can increase the number of Ia⁺ LC.     

Down-regulation of Langerhans cell protein kinase C-β isoenzyme expression in inflammatory and hyperplastic dermatoses

The family of protein kinase C (PKC) isoenzymes plays a fundamental part in signal transduction, and thereby regulates important cellular functions, including growth, differentiation, cytokine production and adhesion molecule expression. In lesional psoriatic skin. Ca²⁺-dependent PKC activity, PKC-β protein and epidermal Langerhans cell (LC) PKC-β immunostaining are significantly decreased, indicating activation and subsequent down-regulation of PKC. Whether these changes occur in other inflammatory/hyperplastic dermatoses is, however, unknown. We examined PKC-α and PKC-β expression in normal skin, psoriasis, cutaneous T-cell lymphoma (CTCL), lamellar ichthyosis, non-bullous ichthyosiform erythroderma, atopic dermatitis, urushiol-induced allergic contact dermatitis, and sodium lauryl sulphate (SLS)-induced irritant contact dermatitis. Cryostat sections were stained for PKC-α and PKC-β, and the LC marker CDla, using an immunoperoxidase technique and specific monoclonal antibodies. Double-labelling studies, in normal skin, revealed co-expression of PKC-β and CDla by epidermal LCs. Analysis of the number of PKC-^β+ and CDl^a+ epidermal LCs, in diseased compared with normal skin, revealed three categories: (i) in psoriasis and CTCL. the PKC-^β+ epidermal LC number was significantly reduced, whereas the CDl^a+ epidermal LC number was unchanged; (ii) in allergic and irritant contact dermatitis, both PKC-β+ and CDla⁺ epidermal LCs were significantly reduced in number; and (iii) in atopic dermatitis, the PKC-β⁺ epidermal LC number was normal, and CDla⁺ epidermal LCs were significantly increased in number. Moreover, the ratio of epidermal LC PKC⁺/CDla⁺ was reduced in all the dermatoses studied, suggesting activation of PKC-β, with subsequent down-regulation. Within the dermis, increased PKC-β staining of infiltrating cells was observed in all the conditions studied except lamellar ichthyosis and non-bullous ichthyosiform erythroderma. These data indicate that: (i) down-regulation of LC FKC-β occurs in a variety of inflammatory and hyperplastic skin disorders, and is not unique to psoriasis, and (iii the pattern of epidermal LC PKC-β and CDla expression varies among the diseases studied. In mice, PKC activation induces LC migration. Thus, down-regulation of epidermal LC PKC-β associated with reduced CDla⁺ epidermal LCs in allergic and irritant contact dermatitis suggests that PK.C-β may transduce the signal for migration of LCs from human epidermis.     

Evaluation of Sialyl Lewisx Antigen in the Skin and the Sera of Patients with Psoriasis Vulgaris

The roles of sialyl-Lewis^x antigen were evaluated in the pathogenesis of psoriasis. Sialyl-Lewis^xexpression was investigated immunohistochemically in the epidermis of normal human skin and erythematous lesional skin of psoriasis vulgaris by avidin-biotin-peroxidase complex procedures. A few sialyl-Lewis^x positive dendritic cells were detected in the epidermis of normal human skin. In 7 out of 9 cases of psoriasis vulgaris, the number of sialyl-Lewis^x-positive epidermal dendritic cells increased in the erythematous lesion over the adjacent normal skin; there were no marked changes in the numbers of CD1a-positive cells in the epidermis between the two skin types. In the double immunofluorescence studies, more than half of the sialyl-Lewis^x-positive epidermal cells in psoriatic erythema were stained with a monoclonal Lag antibody that specifically reacts with Birbeck granules and related structures of human Langerhans cells. Furthermore, we determined the changes in serum levels of sialyl-Lewis^x antigens in patients with psoriasis. Although levels in the sera were not significantly elevated over those of controls, the increases correlated with the degree of disease activity. These findings suggest that sialyl-Lewis^x antigen is possibly involved in the development of psoriasis.     

Effect of UVB 311 nm irradiation on normal human skin

Ultraviolet radiation B (UVB) on the skin induces erythema, inflammation and modifications of the immune system. These changes have been reported after excessive short-term or long-term exposure to broad spectrum UVB. In this study, we examined the effects of local repetitive UVB irradiation of 311 nm wavelength on the skin of seven young volunteers. Skin biopsies were taken before and after UVB irradiation, and we immunohistochemically analyzed the expression of CD1a and HLA-DR antigens of Langerhans cells (LC), the possible infiltration of dermis/epidermis by CD11b macrophages, the modifications or the induction of intercellular adhesion molecule-1 (ICAM-1), E-selectin and vascular cell adhesion molecule-1 (VCAM-1) involved in the binding of leukocytes to the endothelial surface and the development of perivascular infiltrates of LFA-1⁺ mononuclear cells. We also determined the expression of substance P receptors (SPR) using biotinylated substance P (SPB). Exposure of UVB 311 nm induced a drastic reduction of CD1a⁺ cells and a moderate increase of HLA-DR⁺ dendritic cells in the epidermis without infiltration by CD11b macrophages. An increase of the binding of SPB to upper layer epidermal cells was noted in five of seven biopsies. In the dermis, vessel-associated ICAM-1 expression increased and an induction of E-selectin occurred on nearly 20 to 40% of endothelial cells, but VCAM-1 expression remained undetectable. The percentage of LFA-1⁺ cells did not change significantly after irradiation. These observations may be compatible with a selective role of UVB 311 nm on the skin immune response.     

Presence of somatostatin in normal human epidermis

Somatostatin (SOM) is a ubiquitous peptide which is responsible for the inhibition of numerous biological functions. SOM is described as an antiproliferative molecule and an inhibitor of exocrine or endocrine secretion from a variety of tissues, including pancreas, gastrointestinal tract, central and peripheral nervous system. Mediation of SOM effects can be indirect or direct, respectively, through other molecules or receptors on target cells. We have searched for the presence of SOM in the epidermis using immunofluorescence, confocal laser scanning microscopy, radioimmunoassay, and chromatography. Immunofluorescence and confocal laser scanning microscopy studies were performed using rabbit antiserum anti-SOM and mouse monoclonal antibody directed to CD1a Langerhans cell (LC) marker disclosed with fluorescein or tetramethylrhodamine isothiocyanate conjugates. SOM was extracted from whole skin or epidermal cell suspension or LC-enriched suspensions and analysed by radioimmunoassay. We used an antiserum which was reactive for the 6–11 portion of native SOM. Chromatographic columns were performed on extracts from whole skin. The epidermis was SOM immunoreactive. LC were immunoreactive for SOM and the staining was membranous. SOM was extracted from the whole skin at about 0·13±0·02 fmol/mg of tissue (mean±SEM). The SOM concentration in epidermal cell suspensions was 1·5±0·9 fmol/10⁶ cells. Data obtained with LC-enriched suspensions showed large variations between donors. Extracts from skin showed one peak with an elution profile like that of 14 amino acid SOM. This study demonstrates that 14 amino acid SOM is expressed in normal human epidermis.     

Effect of granulocyte macrophage-colony stimulating factor on Langerhans cells in normal and healthy atopic subjects

Granulocyte macrophage-colony stimulating factor (GM-CSF) is a multipotent cytokine produced by many cutaneous cell types including keratinocytes. Langerhans cells (LC) represent the major antigen-presenting cells in skin, and in vitro studies demonstrate that GM-CSF is of pivotal importance in LC. Healthy volunteers ( n  = 3 non-atopic, n  = 3 with atopy) received recombinant human GM-CSF (0.05 μg/mL) by intradermal injection for 3 days to the same site. Diluent was injected in a similar manner as control. Biopsies were taken 24 h after the final injection and examined immunohistochemically for LC and inflammatory cell markers. Compared with control sites, intradermal GM-CSF resulted in shortening of dendritic cell processes and redistribution of LC in the epidermis; numbers of CD1a + cells in the epidermis were significantly decreased ( P  < 0.005), while those in the dermis were significantly increased ( P  < 0.05) following intradermal GM-CSF when compared with controls. Double labelling studies on epidermal CD1a + cells indicated de novo expression of intercellular adhesion molecule (ICAM)-1 and increased expression of HLA-DR following GM-CSF ( P  < 0.005, P  < 0.005, respectively). Additional findings included a marked mixed inflammatory cell infiltrate in the dermis and increased expression of the endothelial cell adhesion molecules E-selectin and ICAM-1. These data indicate that in normal human skin, GM-CSF induces changes in the phenotype and distribution of CD1a + cells consistent with LC functional maturation and exit from the epidermis to the dermis. As these events are central to the initiation of cutaneous inflammation, GM-CSF may potentially play a critical role in the pathogenesis of inflammatory dermatoses.     

IMMUNOHISTOLOGIC LOCALIZATION OF RAS P21 IN NORMAL, HYPERPLASTIC, AND NEOPLASTIC EPIDERMIS

Background. Ras p21, a ras oncogene product, plays an important role in tumorigenesis, proliferation, and differentiation in various tissues and cells. Methods. Using a monocolonal antibody raised against ras p21 (RASK 4), localization of ras p21 in normal epidermis and involved epidermis of various skin diseases was examined immunohistologically. Results. Ras p21 was not present in basal cells of normal epidermis or basaloid cells of basal cell epithelioma but was found almost evenly in the cytoplasm of squamous cells and granular cells. This suggests that ras p21 is concerned with differentiation of epidermal cells. The mode of distribution of ras p21 differed from one cell to another in epidermal cells that turned to malignancy. The distribution was uneven and irregular in the tumorous region on the whole. Conclusions. This result might possibly represent abnormal differentiation of epidermal cells that turned to malignancy, deviating from the regular mode of the distribution of ras p21, which is necessary for normal differentiation.     

Interleukin 1β and the stimulation of Langerhans cell migration: comparisons with tumour necrosis factor α

Epidermal Langerhans cells (LC) and the cells into which they mature are believed to play a pivotal role in cutaneous immune function. The induction phase of contact sensitization is associated with the migration of LC from the skin and their accumulation as dendritic cells (DC) in lymph nodes draining the site of exposure. We have demonstrated previously that tumour necrosis factor α (TNF-α), an epidermal cytokine produced by keratinocytes, provides one signal for LC migration. We describe here experiments designed to evaluate the influence of interleukin 1β (IL-1β), a product exclusively of LC in murine epidermis, on LC migration, LC morphology and DC accumulation, and to compare the effects of this cytokine with those of TNF-α. Both cytokines induced a significant reduction in the frequency of epidermal LC and the arrival of DC in draining lymph nodes. Changes in both parameters were induced more rapidly following intradermal administration of TNF-α than were observed after treatment with IL-1β. However, the reduction in LC frequency was more persistent with IL-1β. Both cytokines caused the activation of LC, characterized by the acquisition of a more dendritic morphology and the increased expression of Ia molecules. These results demonstrate that IL-1β and TNF-α can each stimulate the migration of epidermal LC, but that the changes induced by these cytokines are not identical. Received: 9 September 1996     

The tolerogenic molecule CD95-L is expressed on the plasma membrane of human activated, but not resting, Langerhans' cells

Although dendritic cells (DC) are well known for their immunogenic capacities, they may even induce peripheral T-cell tolerance, and such a tolerogenic potential can be exerted in mouse through the expression on the DC plasma membrane of the CD95-ligand (CD95-L) molecule, which is able to trigger apoptosis of CD95-expressing antigen-specific T cells. We therefore asked whether epidermal DC, namely Langerhans' cells (LC), either resting (i.e. within the epidermis, 'in situ') or activated (i.e. suspended from the epidermis) or both, could express the CD95-L molecule on the plasma membrane. For such a purpose, two colloidal gold-immunoelectron microscopy (IEM) double-step procedures were carried out: an 'in situ' method, able to investigate resting LC, was performed on ultrathin frozen sections obtained by ultracryomicrotomy (UCMT) of normal skin biopsies; a pre-embedding (P-E) method, able to investigate suspended LC, was performed on epidermal cells (EC) suspended from normal skin specimens. In UCMT/IEM sections, resting LC showed gold particles within the cytoplasm but very rarely within organelles and never along the plasma membrane: resting LC are therefore capable of synthesizing CD95-L but not of expressing it in a functional location, thus autoreactive phenomena against CD95-expressing EC being avoided in normal epidermis. On the other hand, in P-E/IEM preparations, suspended LC showed several gold particles along the plasma membrane: activated LC are therefore capable of expressing CD95-L in a functional location, thus bearing the potential to exert tolerogenic capabilities against CD95-expressing T cells, e.g. to prevent inflammatory/autoimmune cutaneous disorders and/or favor the resolution thereof.     

The steady-state turnover of murine epidermal Langerhans cells

We have investigated the steady-state turnover of murine epidermal Langerhans cells (LCs) using an X-irradiation model, 3H-thymidine autoradiography and cultured epidermal sheet explants, and by assessing the LC population in normal mice. The LC density after whole-body irradiation without any cutaneous shielding was not significantly different from that in skin shielded during whole-body irradiation (P > 0.05), indicating that the additional irradiation to the skin did not contribute to a decrease in LC density. In both instances, the LC number gradually decreased in a linear fashion. The results indicate that epidermal LCs continuously leave the epidermis and are continually replaced by circulating precursor cells from the bone marrow at a steady rate. Autoradiographic studies after a pulse injection of 3H-thymidine showed a labelling index of 0.013%, indicating that local mitosis is not an important contributor to the maintenance of the epidermal LC population. Although local X-irradiation resulted in temporary reduction of LC density, epidermal sheet explant culture obtained immediately after local X-irradiation showed no difference in LC density as compared with control unirradiated skin, indicating that the decrease in LC density was not due to significant LC destruction. From these data, we calculated that the half-life of murine LCs in the epidermis is approximately 9 days.     

Self‐renewal capacity of human epidermal Langerhans cells: observations made on a composite tissue allograft

Abstract: Epidermal Langerhans cells (LC) are dendritic, antigen‐presenting cells residing within mammalian epidermis and mucosal epithelia. When massively depleted, they are replaced by cells of bone‐marrow origin. However, their renewal within normal skin under steady‐state conditions is not precisely known. We observed that epidermal LC within a human hand allograft remain stable in the long term (10 years) and are not replaced by cells of recipient’s origin; furthermore, we observed a Langerhans cell in mitosis within the epidermis 8 years postgraft. These results show that under almost physiological conditions, human LC renew in the epidermis by local mitoses of preexisting cells.     

In situ evidence that the population of Langerhans cells in normal human epidermis may be heterogeneous

Epidermal Langerhans cells (LC) may occur in subsets with different phenotypic and functional characteristics. In this work we give further evidence that the CDla-positive LC population in the normal human epidermis may be heterogeneous. We found that one of our monoclonal antibodies (TE4B) to stratum corneum chymotryptic enzyme (SCCE) stained a population of dendritic cells in the normal epidermis, in addition to high suprabasal keratinocytes. The staining of the dendritic cells was seen only when the biopsies had been fixed with formaldehyde and when the sections had been pretreated, either with proteolytic enzymes or with Triton X-100. The blinding of the antibody was mediated through its antigen binding site, as it could be inhibited by adsorption with recombinant pro-SCCE. Experiments with double labelling showed that the TE4B-positive dendritic cells were also CDla-positive. On the other hand, not all CDla-positive cells were TE4B-positive. By means of confocal microscopy of double-labelled cells, the TE4B binding site could be localized intracellularly. SCCE-mRNA could be detected by in situ hybridization in high suprabasal keratinocytes only. A possible explanation may be that there is a subset of LC which have taken up SCCE secreted by high suprabasal keratinocytes. Alternatively. TE4B may bind to an epitope present in a subgroup of epidermal LC which cross-reacts immunologically with SCCE. It is suggested that the demonstrated heterogeneity of the population of LC in the normal epidermis should be taken into account in studies on the possible role of epidermal autoantigens in the development of immune-mediated skin diseases.     

Lymphocytes and macrophages of the epidermis and dermis in lesional psoriatic skin,but not epidermal Langerhans cells,are depleted by treatment with cyclosporin A

Summary Since cyclosporin A (CsA) is an immuno-suppressive agent, its beneficial effect in psoriasis suggests that immune cells may play a role in the pathogenesis and resolution of psoriasis. To determine early effects of CsA in psoriasis, we quantitated immune cells using double immunofluorescence microscopy on biopsy specimens obtained prior to therapy and after 3,7, and 14 days of CsA therapy. CsA therapy resulted in significant reductions in the absolute number of immune cells (including T cells, monocytes/macrophages, and antigen presenting cells) contained within psoriatic skin. The effect was rapid, with over one-half of the reduction in the density of HLe1⁺ (human leukocyte antigen-1 positive or bone marrow derived) cells, including T cells, activated T cells, monocytes, and Langerhans cells (LCs), occurring within 3 days. Despite the overall reduction in the numbers of immunocytes in the skin, the proportion of T cells, Langerhans cells, and monocytes in relation to the total number of immune cells was unchanged with therapy, reflecting equally proportional losses of each subtype. Dermal CD1⁺DR⁺ cells (putative Langerhans cells), which are not found in normal skin but are present in lesional psoriasis skin, were virtually cleared from the papillary dermis after CsA therapy. Although absolute numbers of epidermal Langerhans cells, defined as cells expressing both CD1 (T6) and DR molecules (CD1⁺DR⁺), were also reduced after CsA, epidermal non-Langerhans CD1^-DR⁺ cells (macrophages, activated T cells, DR^- keratinocytes) demonstrated a proportionally greater decrease, with the ratio of CD1⁺DR⁺ Langerhans cells/non-Langerhans CD1^-DR⁺ epidermal cells changing from a mean of 0.82 at baseline to 1.92 at day 14. Thus, early in the course of therapy, CsA appears to be effective at clearing CD1^-DR⁺ cells while leaving LC relatively intact in the epidermis.This work was supported in part by the Babcock Foundation     

UVB-Dependent Modulation of Epidermal Cytokine Network: Roles in UVB-Induced Depletion of Langerhans Cells and Dendritic Epidermal T Cells

The epidermis of mice consists of three cellular components, i.e., keratinocytes, Langerhans cells (LC), and dendritic epidermal T cells (DETC). Each epidermal subpopulation produces a different set of cytokines, thereby forming a unique cytokine milieu. These cytokines, in turn, support the survival and growth of LC and DETC and regulate their immunological functions. LC and DETC play important, but distinct, effector roles in protective immunity against antigens that are generated in or penetrate into the epidermis. Acute or chronic exposure of mice to ultraviolet B (UVB) radiation is known to impair this cutaneous immunity, as evidenced by the failure to induce T cell-mediated immune reactions, by the generation of antigen-specific immunological unresponsiveness, and by the development of skin cancers. Importantly, these changes are associated with reduced densities of LC and DETC in UVB-exposed skin, suggesting that the deficiency in these epidermal leukocytes may account for some of the deleterious influences of UVB radiation on skin. Here I will review the recent advance in our understanding of the mechanisms by which UVB radiation may deplete LC and DETC from epidermis. More specifically, I will discuss the following possibilities: a) UVB-mediated suppression of the production of relevant growth factors for LC and DETC, b) UVB-induced abrogation of surface expression of growth factor receptors, and c) UVB-triggered apoptotic cell death in epidermal leukocytes.     

Effect of orally administered aromatic retinoid on murine Langerhans cells

Summary The effect of orally administered aromatic retinoid (Ro 10-9359) on murine epidermal Langerhans cells (LC) was studied in vivo and in vitro. Daily administration of retinoid caused a transient increase in LC density, as determined by staining for Ia antigens, during the first few days of treatment and thereafter a continuing decrease that reached a maximum at 2 weeks. In addition, the morphology and location in the epidermis had been altered. When the treatment was continued to 4 weeks, the density of LC returned to normal. The Ia-antigen-presenting function of epidermal cells to an allo-Ia-reactive cloned T cell line was elevated at all stages of retinoid treatment examined. This elevation did not correlate with the density of histochemically stainable Ia⁺ LC. These findings suggest that orally administered retinoid profoundly alters the functional capacity of Ia⁺ LC.     

Distinctive dendritic cell subsets expressing factor XIIIa, CD1a, CD1b and CD1c in mycosis fungoides and psoriasis

The papillary dermis of psoriasis and mycosis fungoides (MF) lesions is characterized by prominent collections of cells with dendritic morphology. Immunophenotypically distinct populations of cutaneous dendritic cells have been identified as CD1a+, FXIIIa-Langerhans cells (LC) and CD1a-, FXIIIa+ dermal dendritic cells (DDC). In this study, antibodies against the human GDI cluster of antigens (i.e. CD1a, CD1b and CD1c) and the DDC) marker (FXIIIa) were used to further characterize the subsets of dendritic cells in normal skin as compared to neonatal foreskin, psoriasis and MF by both immunoperoxidase and double immunofluorescence techniques. Normal skin and foreskin epidermis and dermis contained few CD1b+ or CD1c+ cells along with normal numbers of CD1a+ LC and FXIIIa+ DDC. Both MF and psoriasis were characterized by CD1a+ cells in the epidermis and dermis. FXIIIa+ cells were greatly expanded in the upper dermis of MF lesions and to a lesser degree in psoriasis as has been previously described by our group. MF contained significantly increased epidermal and dermal CD1b+ (15.7/5 high power fields [HPF] and 59.7/5 HPF respectively) and CD1c+ dendritic cells (33.8/5 HPF and 95.9/5 HPF respectively), while in psoriasis these cells were not statistically different from normal skin. Double immunofluorescence studies revealed that some (<25%) FXIIIa+ cells co-expressed CD1b and CD1c in MF>psoriasis> foreskin, while FXIIIa+ DDC never co-expressed CD1a. Thus, in contrast to normal skin in which epidermal or dermal dendritic cells rarely express CD1b and CD1c antigens, these members of the CD1 family are upregulated on both LC and DDC in benign and malignant inflammatory states. Upregulation of CD1b and CD1c on MF epidermal and dermal dendritic cells, as compared to psoriasis, foreskin and normal skin, may be useful in the immunophenotypic recognition of MF, as well as in helping to understand its immunobiology.     

Tumour necrosis factor alpha is pro-inflammatory in normal human skin and modulates cutaneous adhesion molecule expression

Tumour necrosis factor a (TNF-α) is a potent immunoregulatory cytokine produced by many cutaneous cells, including kcratinocytes, mast cells and Langerhans cells. To explore its potential role in inflammatory skin disease, we have studied immunohistochemically the effects of intradermal recombinant human TNF-α (rHuTNF-α) on cutaneous inflammatory cells, adhesion molecules and Langerhans cells in normal human skin. Volunteers received rHuTNF-α 100U (group A), 5000 U (group B), or 100 U daily for 5 days (group C), and biopsies were taken at 6 h (groups A and B), or 6 h after the final injection (group C). An inflammatory cell infiltrate developed in all cases: following single injections of either 100 or 5000 U rHuTNF-α this was predominantly neutrophilic, whereas following multiple injections of 100 U few neutrophils were seen, although many lymphocytes (CD3⁺, CD4⁴) were present. In all groups there was an increase in cells of monocyte/macrophage lineage (CD36⁾⁺. TNF-α induced a dose- and time-dependent decrease in CDla⁺ epidermal Langerhans cell numbers and an increase in dermal CDla⁴ cells, suggesting migration of Langerhans cells away from the epidermis. TNF-α induced endothelial E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in all groups, and adhesion molecule expression by interstitial dermal dendritic cells (ICAM-1 and VCAM-1) and keratinocytes (ICAM-1) was observed. These findings indicate that TNF-α is a potent modulator of cutaneous immune function in vivo, and this central role in the cutaneous immune response suggests that TNF-α may be an attractive target for therapeutic inhibition.     

The ICAM-3/LFA-1 interaction is critical for epidermal Langerhans cell alloantigen presentation to CD4+ T cells

Intercellular adhesion molecule (ICAM)-3 is a recently described member of the immunoglobulin superfamily and, as such, is closely related to ICAM-1 and ICAM-2. All three ICAMS are cognate for the counter-receptor lymphocyte function associated antigen-1 (LFA-L CD11a/CD18). Unlike ICAM-1 and ICAM-2. ICAM-3 is constitutively expressed at high levels on resting leucocytes. We investigated the expression and function of ICAM-3 in normal skin (n= 5), as well as its expression in psoriasis (n= 4). atopic eczema (n= 4), allergic (rhus) contact dermatitis (n=3). and cutaneous T-cell lymphoma (CTCL. n=2). Five-micrometre cryostat sections of skin were stained using monoclonal antibodies to ICAM-3 and A well characterized immunoperoxidase technique. In normal skin. ICAM-3 was expressed by all cutaneous leucocytes hut most striking was the strong expression of ICAM-3 by Langerhans cells within both epidermis and dermis. This observation was confirmed by double-labelling with CD1a and negative staining with an IgG1 isotype control. In psoriasis, atopic eczema, allergic contact dermatitis, and CTCL. ICAM-3 was co-expressed on all CD1a⁺ cells, although, in psoriasis, the intensity of ICAM-3 expression was reduced. Functional blocking experiments were performed to determine whether the observed ICAM-3 expression on Langerhans cells was functionally important in antigen presentation. CD4⁺ T cells were prepared from peripheral blood and 10⁵ CD4⁺ T cells combined with 10⁵ epidermal cells harvested from keratome biopsies of normal skin of an individual allogeneic to the T-cell donor. Addition of 50 μg anti-ICAM-3 to the co-culture resulted in a consistent (50%) reduction in degree of alloantigen presentation by Langerhans cells to T cells. Inhibition was 77% of that produced by the addition of anti-LFA-1. These data indicate that ICAM-3 is constitutively expressed by Langerhans cells and is a major ligand for LFA-1 on CD4⁺ T cells during their response to Langerhans cells. Because fresh Langerhans ceils constitutively express little ICAM-1. whereas ICAM-3 is constitutively expressed at high levels, it would appear that 1CAM-3 is the dominant functional ICAM on in situ Langerhans cells in the normal epidermis.     

Induction by tumour necrosis factor α of dose-related changes in Langerhans cell frequency in mice

Abstract It has been demonstrated previously that tumour necrosis factor α (TNF-α) provides an important signal for the migration of epidermal Langerhans cells (LC) from the skin. Intradermal administration to mice of homologous recombinant TNF-α induces both a rapid reduction in the frequency of LC local to the site of exposure and, somewhat later, an accumulation of dendritic cells in draining lymph nodes. It has been proposed recently, however, that the influence of TNF-α on LC function may be dose-dependent in nature with lower concentrations inducing migration, but higher concentrations immobilizing LC in the epidermis. To investigate this proposal we examined the kinetics and dose-response relationships of TNF-α-induced LC migration in mice. At all concentrations tested (50, 150 or 300 ng/ear), intradermal exposure to TNF-α caused within 30 min a significant reduction in the frequency of MHC class II (Ia)⁺ LC within epidermal sheets. With the lower concentrations of TNF-α this effect was still apparent when LC were enumerated in the epidermis up to 4 h following cytokine treatment. In contrast, however, exposure of mice to 300 ng of TNF-α was consistently associated with a considerably less marked, and statistically insignificant, reduction in LC frequency by 4 h. These data indicate that at all concentrations of the cytokine examined here, TNF-α was able to stimulate a rapid (within 30 min) reduction in epidermal LC numbers, but that the rapidity with which the epidermis was repopulated following the initiation of LC migration was influenced by the concentration of TNF-α administered. It is suggested that TNF-α may influence not only the tempo of LC migration, but also the kinetics of epidermal repopulation. Received: 18 June 1998 / Received after revision: 25 February 1999 / Accepted: 19 March 1999