Immunoglobulins and Their Receptors on Epidermal Langerhans Cells in Atopic Dermatitis

To elucidate the etiological role of immunoglobulin molecules on Langerhans cells (LCs) in atopic dermatitis, we conducted immunohistochemical studies on the localization of immunoglobulin G1 (IgG1), IgG2, IgG3, IgG4, IgA and IgM on epidermal LCs from 30 patients with atopic dermatitis (AD) and five non-atopic healthy volunteers. We also investigated the types of receptors for the immunoglobulins (FcεRI, FcεRII, FcγRI, FcγRII, and FcγRIII) on epidermal LCs in the patients. IgE positive epidermal LCs were observed in 28 of 30 AD patients, and 46.7% of the epidermal LCs were positive for IgE. Both IgG1- and IgG2-positive epidermal LCs were obserbed in 70% of AD patients, and 21.8% and 28.7% of the total epidermal LCs were positive for IgG1 and IgG2, respectively. IgG3- or IgG4-positive LCs were present in only small proportions of AD patients. IgA-positive LCs were observed in 8 AD patients; our study suggested that the IgA bound on LCs was secretory IgA (S-IgA). These surface immunoglobulins were observed significantly more frequently on epidermal LCs in the involved skin of AD than in clinically uninvolved skin. No IgM-positive epidermal LCs were observed in the AD patients or healthy volunteers. In non-atopic healthy controls, no immunoglobulin-binding LCs were observed. In receptors for immunoglobulins, FcεRI and FcγRII were exclusively expressed on nearly all epidermal LCs from all AD patients and all non-atopic controls. These results suggested that not only IgE but also IgG and IgA may play some etiological role in the pathogenesis of AD.     

Fc epsilon receptor II/CD23 positive lymphocytes in atopic dermatitis: II. Infiltration of Fc epsilon R II(+) T cells in the skin lesion

Cells expressing Fc receptors for IgE (Fc epsilon R II) were identified in skin from patients with atopic dermatitis (AD), eczematous dermatitis (ED), and in skin from normal nonatopic subjects, with the use of monoclonal antibodies to human lymphocyte Fc epsilon R II, H107, and to lymphoid cell-surface antigens by double immunofluorescence staining. Two to four percent of infiltrating mononuclear cells expressed Fc epsilon R II, and more than half of these cells were T epsilon cells in both acute and chronic AD lesions. Fc epsilon R II(+) T cells (T epsilon cells) bearing CD8 infiltrated preferentially acute lesions, whereas chronic lesions contained either CD8(+) or CD4(+) T epsilon cells, or both. Fc epsilon R II(+) cells rarely were present in ED lesions. There was no significant correlation between % Fc epsilon R II(+) peripheral blood mononuclear cells and the proportion of lesional Fc epsilon R II(+) cells, extent of skin lesions, or serum IgE levels, implying the selective accumulation of Fc epsilon R II(+) cells in the inflammatory infiltrate of AD. These observations suggest that the increased generation of Fc epsilon R II(+) cells in skin lesions, including CD8 (+) T epsilon cells, is involved in the pathogenesis of AD.     

Interleukin-8 immunoreactivity in the skin of healthy subjects and patients with palmoplantar pustulosis and psoriasis.

Previous studies have shown that neutrophil-activating peptide 1/interleukin-8 (IL-8) is present in psoriatic scales and to a lesser extent in normal human epidermis. A panel of monoclonal antibodies and polyclonal antisera raised against IL-8 was used to localize IL-8 with immunoperoxidase techniques in non-lesional and lesional skin of patients with psoriasis and palmo-plantar pustulosis (PPP), and in corresponding sites from healthy subjects. Intracellular IL-8 immunoreactivity was found in all epidermal cell layers in biopsies of healthy subjects and in non-lesional and lesional skin in both PPP and psoriasis. The most intense immunolabeling was regularly found in the basal cell layer. Intercellular epidermal IL-8 immunolabeling was regularly detected in lesional biopsies in PPP and psoriasis, but not in healthy subjects or non-lesional skin in PPP and psoriasis. No intercellular immunolabeling was detected after successful treatment of lesional skin. The majority of cells along the eccrine sweat glands, dermal mononuclear cell infiltrates, and endothelial cells were IL-8 immunoreactive in all biopsies studied. The present study suggests that IL-8, its precursor form, or, alternatively, a degradation product is present in normal human epidermis.     

Epidermal Langerhans cells, HIV-1 infection and psoriasis

Summary Langerhans cells (LCs) subserve an important antigen-presenting function in the skin immune system. They bear CD4 receptors, which make them potential targets for infection with the human immunodeficiency virus (HIV-1).
The observation of reduced numbers of LCs in the skin of patients with the acquired immunodeficiency syndrome (AIDS), and the association of severe psoriasis with HIV-1 infection, raise interesting questions regarding the role of LCs in the skin of HIV-1-positive psoriatic patients.
In this study, LCs were quantified in the lesional and non-lesional skin of seven HIV-1-positive psoriatic patients, and the results were compared with age-, sex- and site-matched HIV-1-negative psoriatic patients. The number of LCs was determined by staining skin sections with S-100 polyclonal antibody, using the three-step avidin—biotin immunoperoxidase method. The S-100-positive cells above the basal layer were quantified in two ways: cells/mm² of epidermal area, and cells/mm of length of basement membrane.
HIV-1-positive psoriatic patients showed a reduction in the number of epidermal LCs compared with HIV-1-negative psoriatic patients using both methods of quantification, in both lesional and non- lesional skin (P <0·05, Mann-Whitney test). In addition, a reduction in the number of LCs in lesional compared with non-lesional skin was observed in both HIV-1-positive and -negative patients when LCs were quantified per mm² of epidermal area (P<0·05, Wilcoxon test). This reduction was also observed when LCs were quantified per mm length of basement membrane, but the reduction was not statistically significant in the control group of HIV-1-negative psoriatic patients. Our findings of a reduced number of LCs in the epidermis of HIV-1 -positive psoriatic patients may be associated with the clinical deterioration of psoriasis in these patients.     

Quantitative analysis of tryptase- and chymase-containing mast cells in atopic dermatitis and nummular eczema

The distribution of mast cells (MCs) containing tryptase (T) and chymase (C) was studied in the non-lesional and lesional skin of 26 patients with atopic dertnatitis (AD) and 23 patients with non-atopic nummular eczema (NE). and in the skin of eight healthy controls. T and C activities were demonstrated enzymehistochemically using 2-Gly-Pro-Arg-MNA and Suc-Val-Pro-Phe-MNA as substrates, respectively. The T- and C-containing MCs were counted separately in the epidermis, in contact with the basement membrane. In the papillary dermis and in different dermal levels (0·2 mm each). Also, the C protein was determined immunohistochemically. T-positive MCs were similarly distributed in non-lesional and lesional skin of both AD and NE. The MC number was relatively high in the upper dermis (papillary dermis and levels I and I!) of non-lesional and lesional skin of AD. In the upper dermis of non-lesional AD and NE skin and in normal skin, about 50% of T-positive MCs displayed C activity, whereas the percentage in lesional AD and NE skin was only about 30%. hi this respect, the non-lesional and lesional samples differed significantly froLu each other in both dennatoses (in AD p = 0%005, in NEP = 0·002. Students' t-test). In all samples the MC number decreased in the deeper dermal levels, although numerous T-containing MCs were still counted in the deeper dermis (dermal levels IV-VII) of lesional AD and NE skiti. differing significantly from the MC number in normal skin (In ADp = 0·005. in NE p=0·041). In the deeper dermis. the percentage of MCs containing active C was about 70% in non-lesional and lesional AD and NE. and about 90% in normal healthy skin. However, in the upper dermis of non-lesional and lesional skin of both AD and NE. about H()% of all MCs contained the C protein, which differed significantly from the value of 100% in normal skin (p<0·5). In conclusion, the increased number of T-positive MCs in the upper dermis of non-lesional and lesional AD contributes to promoting inflammation. C apparently loses its activity in the upper dermis of lesional AD and especially in NE. Thus. Ihe enzyme partially lacks its capability to suppress inflammation, such as degradation of neuropeptides and proteins. The dysregulation of these proteinases exists already in non-lesional skin of AD and NE.     

Expression, but lack of calcium mobilization by high-affinity IgE Fcε receptor I on human epidermal and dermal Langerhans cells

Abstract In atopic dermatitis (AD) patients, IgE molecules are demonstrated on the surface of Langerhans cells (LC). FcεRI molecules, which are present on the surface of LC in AD patients as well as normal individuals, are responsible for this binding. In this study, we have investigated phenotypic and functional characteristics of FcεRI on epidermal and dermal cell populations. Epidermal and dermal cell suspensions were prepared enzymatically with dispase followed by either trypsin or collagenase treatment, respectively. Peripheral blood basophils were negatively selected by excluding other leukocytes with surface marker staining. Consistent with previous reports, both peripheral blood basophils and epidermal LC were positively stained with anti FcεRI monoclonal antibody. In addition, an FcεRI positive population was demon-strated among dermal HLA-DR positive cells. These cells express significant amounts of HLA-DR molecules (DR^Hi) and co-express CD la molecules, which identifies them as LC-like dendritie APC of the dermis. No other FcεRI positive population was found in the other dermal DR^Mid or DR populations, except for a minor DR^lo population, presumably mast cells. To analyze whether these FcεRI molecules are signal transducing for LC, intracellular calcium mobilization after crosslinking of FcεRI was measured with How cytometry. Following crosslinking, peripheral blood basophils clearly increased intracellular calcium. On the other hand, neither normal epidermal LC nor dermal DR^HiCD Ia⁺ cells changed their intracellular calcium level after FcεRI crosslinking. These data indicate that normal epidermal and dermal LC, but not basophils, are resistant to calcium flux following FcεRI engagement.     

Alterations of glucosylceramide-beta-glucosidase levels in the skin of patients with psoriasis vulgaris

Hydrolysis of glucosylceramides by the enzyme glucosylceramide-beta-glucosidase (GlcCer'ase) results in ceramide, a critical component of the intercellular lamellae that mediates the epidermal permeability barrier. A disturbance of ceramide formation is supposed to influence the transepidermal water loss in common skin diseases like atopic eczema or psoriasis. The aim of this study was to investigate whether GlcCer'ase levels were altered in the skin of subjects with psoriasis vulgaris. Skin punch biopsies were taken from lesional and non-lesional psoriatic skin and GlcCer'ase was evaluated both at the RNA and at the protein level. Normal skin from surgical patients provided the baseline GlcCer'ase expression in healthy subjects. Our results show that GlcCer'ase mRNA expression was decreased in psoriatic non-lesional skin compared to normal controls in all cases. Interestingly, in lesional psoriatic skin the level of GlcCer'ase was increased compared to non-lesional skin in all cases. For the immunohistochemical analysis, we used a newly synthesized monoclonal antibody anti-human GBC (GlcCer'ase-GST fusion protein). The results confirmed that GlcCer'ase, mainly present in the upper epidermis, was decreased in psoriatic skin compared to normal control and was increased in lesional compared to non-lesional psoriatic skin. Our findings support the concept that alteration in water permeability barrier in lesional psoriatic skin can serve as a trigger for the upregulation of the expression of enzymes like GlcCer'ase with consequent stimulation of ceramide generation.     

Mast cell chymase is increased in chronic atopic dermatitis but not in psoriasis

Mast cell chymase is a chymotrypsin-like serine proteinase primarily stored in secretory mast cell granules. Mast cell chymase has various effects on angiotensin, metalloproteases, lipoproteins, procollagen, neuropeptides and cytokines. Recent studies have demonstrated that chymase inhibitors inhibit skin inflammation. In this study we sought to determine the role of mast cell chymase in atopic dermatitis (AD) in comparison with its role in psoriasis and normal skin. Skin biopsy specimens were obtained from non-lesional and lesional skin of patients with chronic AD and psoriasis and from normal skin of non-atopic and non-psoriatic controls. The number of mast cells containing chymase was determined by immunohistochemistry using a chymase-specific monoclonal antibody. A significantly (P<0.05) enhanced number of chymase-positive cells was found in lesional AD skin as compared to normal skin as well as to lesional and non-lesional skin of patients with psoriasis. A significant (P<0.05) increase in the number of chymase-positive cells was also found in non-lesional AD skin in comparison to psoriasis. An enhanced, albeit not statistically significant difference was noted in non-lesional AD skin as compared to normal skin. In conclusion, these results suggest that mast cell chymase may play an integral part in eliciting and maintaining cutaneous inflammation in AD but not in psoriasis. The increased proteinase activity of mast cell chymase may also be involved in promoting a skin barrier defect in AD, which subsequently enhances the skins permeability to allergens and microbes and thereby aggravates the eczema.     

Impaired sphingomyelinase activity and epidermal differentiation in atopic dermatitis

A defective permeability barrier leads to the penetration of environmental allergens into the skin and initiates immunological reactions and inflammation crucially involved in the pathogenesis of atopic dermatitis (AD). Decreased stratum corneum ceramide content may cause the defect in permeability barrier function consistently found in AD. Acid and neutral sphingomyelinase (A- and N-SMase) generate ceramides with structural and signal transduction functions in epidermal proliferation and differentiation. We determined epidermal SMase activities, DNA synthesis, involucrin, loricrin, filaggrin, and keratin expression in lesional and non-lesional skin of AD patients. We found decreased epidermal A-SMase activity in lesional and non-lesional skin, correlating with reduced stratum corneum ceramide content and disturbed barrier function. N-SMase activity was reduced in non-lesional skin and more significantly reduced in lesional skin, correlating with impaired expression of cornified envelope proteins and keratins, important for skin barrier function. Changes in involucrin, loricrin, filaggrin, keratin K 5 (basal) and K 16 (proliferation associated) were noticed in non-lesional and lesional skin, whereas changes in K 10 (suprabasal), K 6 (proliferation associated), and K 17 (inflammation associated) were found only in lesional skin. In summary, reduction in SMase-generating ceramides and impaired differentiation are involved in the defective barrier function found in AD.     

Perforin and granzyme B may contribute to skin inflammation in atopic dermatitis and psoriasis

BACKGROUND: Infiltration of the skin by pathogenic T cells is regarded as a key factor in the development of inflammatory skin diseases such as atopic dermatitis (AD) and psoriasis. OBJECTIVES: To investigate whether T cells containing cytotoxic proteins may contribute to the generation of skin inflammation in these skin diseases. METHODS: Skin biopsy specimens were obtained from non-lesional and lesional skin of patients with chronic AD (n = 8) and psoriasis (n = 6), and from non-atopic controls with normal skin (n = 6). Expression of perforin and granzyme B was investigated by immunohistochemistry. RESULTS: A significant enhancement of perforin and granzyme B expression was observed in lesional AD skin as compared with normal skin, non-lesional AD skin and psoriasis. Expression of these cytotoxic proteins was also increased in psoriasis as compared with normal skin and non-lesional psoriatic skin. Immunoreactivity for perforin and granzyme B was mainly found in the cytoplasm of lymphocytic cells located in the perivascular infiltrate. In AD increased numbers of positive cells were also observed focally at sites of spongiosis in the epidermis. Double immunostaining revealed that both CD4+ and CD8+ T cells are capable of expressing perforin and granzyme B. CONCLUSIONS: Our data suggest that cytotoxic CD4+ and CD8+ T cells containing perforin and granzyme B may play an integral part in eliciting cutaneous inflammation in AD.     

Neuropeptides in the skin of patients with atopic dermatitis

There is increasing evidence that neuropeptides may be involved in the pathogenesis of atopic dermatitis (AD). This study examines whether neuropeptide distribution in the skin of patients with AD differs from normal controls. The distribution and density of several neuro-peptides were examined in lesional and non-lesional skin of AD patients (n= 5) and in normal controls (n= 4) using indirect immunofluorescence and image analysis. Cholinergic innervation was studied using cholinesterase histochemistry. Staining with the general neuronal marker protein gene product 9·5 showed a subepidermal network of nerves with fibres penetrating the epidermis, and nerves around blood vessels, sweat glands and hair follicles. Image analysis of nerves around sweat glands showed a significantly higher nerve density in non-lesional compared with both normal controls and lesional skin (P < 0·05); lesional compared with control skin showed no significant difference. In the epidermis the density of nerves was not significantly greater in non-lesional compared with lesional skin and controls. Calcitonin gene-related peptide immunoreactivity was similar in all subjects except in three of the AD patients, where more nerves appeared to penetrate the epidermis. Substance P immunoreactivity in the papillary dermis was seen in all AD patients hut no controls. Vasoactive intestinal polypeptide and neuropeptide Y staining were similar in all groups. Acetyleholinesterase-positive nerves were found around sweat glands in all subjects, the staining being greatest in non-lesional and least in lesional skin. Occasional nerves were seen in the papillary dermis in lesional skin of two out of the four patients. We have demonstrated quantitative differences in nerve growth in clinically normal skin of AD patients, and altered cutaneous neuropeptide expression in these patients which may contribute to the pathogenesis of AD.     

Increased expression of adhesion receptors in both lesional and non-lesional psoriatic skin

Adhesion receptors and their ligands play a vital role in the immune system. We studied the expression of different adhesion receptors, using single- and double-staining immunohistochemical techniques, in both lesional and non-lesional skin specimens from seven psoriasis patients and in skin biopsy specimens from eight normal healthy controls. Our results showed an overall increased expression of several adhesion receptors in both lesional and non-lesional psoriatic skin. We consistently found an increased expression in particular of ICAM-1 and E-selectin on endothelial cells, and ICAM-1 on T cells and Langerhans cells. In contrast, a weak expression of VCAM-1 was found on endothelial cells and mononuclear cells in lesional psoriatic skin specimens alone. Interestingly, LFA-1 was also expressed on Langerhans cells, with a greater frequency in skin from lesional than from non-lesional sites, but was never expressed in skin from normal healthy individuals. Furthermore, significantly increased numbers of Langerhans cells and T cells with a positive reactivity for MAb HECA-452 were found in both lesional and non-lesional psoriatic skin. We hypothesize that the enhanced expression of adhesion receptors on migrating immunocompetent cells and endothelial cells of psoriatic skin in general facilitates the increased influx of activated T lymphocytes and other immunocomponent cells into the skin, and thus underscores the generalized character of the disease.     

Mast cells are one major source of interleukin-4 in atopic dermatitis

Several reports have shown the presence of T-helper lymphocytes with Th2 characteristics in the skin lesions of atopic dermatitis (AD). However. Th2 cells themselves require an exogenous pulse of IL-4 to initiate their differentiation and synthesis of IL-4. As mast cells have recently been shown to contain IL-4, this finding prompted us to investigate IL-4 in mast cells of AD lesions, to determine if they might provide the IL-4 pulse needed by the Th2 cells. In this study, we measured IL-4 immunoreactivity in mast cells of non-lesional and lesional skin sections from 20 patients with AD. Ten patients with nummular eczema (NE) without any atopic features or background, and five healthy subjects, served as the control groups. Mast cells were first identified using an enzyme-histochemical staining method for tryptase. Subsequently, the sections were photographed, the tryptase stain was removed, and IL-4 was demonstrated with a polyclonal antibody. The sections were photographed again, and the percentage of IL-4-positive mast cells was calculated. The percentage of mast cells exhibiting IL-4 immunoreactivity in the upper dermis in lesional vs, non-lesional skin was 66±18% vs. 37±18% in AD (P<0.0001, paired t-test), but only 46±19% vs. 31±22% in NE. In the skin of healthy controls, only 23±25% of the mast cells were positive for IL-4. In addition, a significant difference was found between lesional skin of AD vs. NE patients (P<0.008, unpaired t-test). Moreover, the staining intensity of IL-4 in mast cells was clearly increased in the lesional compared with the non-lesional AD skin. Mast cells appeared to be the main cell type containing IL-4 in 40% of the lesional AD skin specimens, whereas in the remaining 60% prominent IL-4-positive mononuclear cell infiltrates were also present. In the non-lesional skin, mast cells appeared to be the main cell type containing IL-4 in all specimens. These results indicate that mast cells are one major source of IL-4 in lesional and non-lesional AD skin, and they could contribute significantly to the development of AD.     

PGP 9.5 distribution patterns in biopsies from early lesions of atopic dermatitis

Peripheral nerve fibres are often increased in lesional skin of atopic dermatitis (AD) patients. We attempted to study nerve fibre profiles, using PGP 9.5 as neuronal marker, in early AD lesions in 10 patients, as compared to non-lesional skin in the same patients and skin from healthy controls. The number of PGP 9.5-positive nerve fibre profiles was not different in the biopsies taken from normal-looking AD skin and healthy controls. The total number of PGP 9.5-positive nerve fibre profiles in the whole skin sections was higher in both the epidermis and the dermis in the group of skin biopsies taken from early lesions of AD patients. Further, the number of epidermal PGP 9.5-positive dendritic cells was increased in AD skin. It seems reasonable that PGP 9.5-positive nerve fibres and PGP 9.5-positive dendritic cells have pathological roles in AD. The findings might serve as a basis for further studies in evaluating novel diagnostic and therapeutic approaches.     

T cells and mast cells as a major source of interleukin-13 in atopic dermatitis

BACKGROUND: Interleukin (IL)-13 is a T-cell-derived cytokine that shares several functions with IL-4, including the induction of immunoglobulin E synthesis. Recent studies suggest that cytokines expressed locally in the skin play several critical roles in atopic dermatitis (AD), however, little is known about the role of IL-13 in AD lesions. OBJECTIVES: The present study was designed to characterize the involvement of IL-13 in AD in the skin and peripheral blood mononuclear cells (PBMC). METHODS: Using lesional and nonlesional skin from adult AD patients and normal skin from healthy volunteers, we performed RT-PCR, in situ RT and immunostaining to determine the IL-13 expression at the mRNA and protein levels. The actual numbers of IL-13 expressing cells in biopsy specimens were counted under the microscope. IL-13 mRNA expression in PBMC from AD patients and healthy volunteers was examined by RT-PCR analysis. RESULTS: IL-13 mRNA expression was detected by RT-PCR in lesional and nonlesional skin and in PBMC from AD patients, but not in normal skin or PBMC from healthy volunteers. In AD lesional skin, numerous IL-13 mRNA-positive cells were demonstrated by in situ RT, and similar numbers of IL-13-positive cells were also detected immunohistochemically. Smaller numbers of IL-13-positive cells were observed in AD nonlesional skin and in normal skin. The differences in the numbers of IL-13-expressing cells between lesional and nonlesional skin were statistically significant. Double immunostaining revealed that IL-13 was produced in approximately 40% of T cells and 20% of mast cells in AD lesional skin, suggesting that T cells and mast cells are major sources of IL-13 in AD lesions. CONCLUSION: IL-13 may play a local as well as a systemic role in the development of AD lesions.     

Seborrhoeic dermatitis and Pityrosporum (Malassezia) folliculitis: characterization of inflammatory cells and mediators in the skin by immunohistochemistry

BACKGROUND: The fact that Pityrosporum ovale plays a part in seborrhoeic dermatitis is well established but the mechanism of this relationship has not been established. OBJECTIVES: To compare the number and type of inflammatory cells and mediators in skin biopsies from normal and lesional skin from the trunk and scalp in patients with seborrhoeic dermatitis, Pityrosporum (Malassezia) folliculitis and in normal skin from healthy controls. METHODS: The skin biopsies were stained using the labelled Streptavidin-biotin METHOD: The following markers were studied: CD4, CD8, CD68, HLA-DR, NK1, CD16, C1q, C3c, IgG, CD54 (ICAM-1), interleukin (IL) -1alpha, IL-1beta, IL-2, IL-4, IL-6, IL-10, IL-12, tumour necrosis factor-alpha and interferon-gamma. RESULTS: HLA-DR+ cells were seen in the highest number, and were higher in lesional skin compared with normal skin from both patients and healthy volunteers. ICAM-1 expression was also increased in lesional skin. C1q and the interleukins showed an increased cellular and intercellular staining in patients compared with healthy controls and the intercellular staining was often more intense in lesions compared with non-lesional skin. Staining was often more intense when Malassezia (Pityrosporum ovale) yeast cells were present. CONCLUSIONS: An increase in NK1+ and CD16+ cells in combination with complement activation indicates that an irritant non-immunogenic stimulation of the immune system is important. The result with the interleukins showed both an increase in the production of inflammatory interleukins as well as in the regulatory interleukins for both TH1 and TH2 cells. Similarities to the immune response described for Candida albicans infections indicate the role of Malassezia in the skin response in seborrhoeic dermatitis and Pityrosporum folliculitis.     

Modulation by Chinese herbal therapy of immune mechanisms in the skin of patients with atopic eczema

Ten patients with atopic eczema (AE) received treatment with Chinese herbal therapy (CHT; Zemaphyte) for 2 months. The severity ot the eczema was recorded and skin biopsies were taken from lesional (L) and non-lesional (NL) skin before and after treatment. The skin biopsies were stained to detect T-cell subsets (CD4, CDH, CD45Ro and CD25), macrophage subsets (RFD7), dendritic cells (RFD1). Langerhans cells (CD1), HLA-DR, low-affinity IgE receptors (CD23) and high-affinity IgE receptors (15A5, 22E7). A quantitative assessment of the numbers of positively stained cells was made. Monoclonal antibody binding specifically to CD23(Fc?RII) was used, in combination with cell subset monoclonal antibodies to quantify the cellular distribution of CD23 antigen in the skin. Following 2 months of treatment with CHT, erythema was reduced by 53%. There was also a significant reduction in HLA-DR expression. The numbers of RFD1+CD23+, RFD7+CD23+, CD1+CD23+ and CD25+ cells in lesional skin decreased significantly after treatment (RFD1+CD23+ from 0.39 to 0·21, RFD7+CD23+ from 0.29 to 0·16· CD1+CD23 + from 0·24 to 0·09, CD25+ from 0·84 to 0·31 in epidermis and from 1·62 to 0·94 in dermis (mean cells numbers per unit area). No significant change in cell numbers in NL skin or expression of FcERI in either L or NL samples was observed after treatment. This study confirms that CHT is clinically efficacious and that clinical improvement is associated with a significant reduction in antigenpresenting cells expressing CD23.