Frank C. van Reijsen

Biphasic response against aeroallergen in atopic dermatitis showing a switch from an initial TH2 response to a TH1 response in situ: An immunocytochemical study

In the pathogenesis of atopic dermatitis (AD), IgE plays an important role; and TH2 cells, producing IL-4, have been ascribed a key role in allergic diseases such as AD. To investigate the role of TH subpopulations in the onset and continuation of AD, we performed atopy patch tests (APTs) with house dust mite allergen in patients with AD. Punch biopsy specimens were taken from the APT site, and sections were immunocytochemically double-stained for IL-4 and interferon-gamma together with different membrane markers. This provides a unique model for studying the kinetics of the TH0, TH1, and TH2 responses in situ. The results show that in lesional skin interferon-gamma-positive cells predominate over IL-4-positive cells. The interferon-gamma-positive cells are mainly CD3+ and, in particular, CD4+ cells; the remainder are CD8+, RFD-1+, and RFD-7+ cells. The IL-4-positive cells are exclusively CD4+ T cells; no eosinophils or mast cells were found to stain for IL-4. With regard to the TH cell response, a clear dichotomy of the eczematous response to allergen in skin was observed. In the initiation phase IL-4 production by TH2 and TH0 cells is predominant over interferon-gamma production by TH1 and TH0 cells. In the late and chronic phases the situation is reversed and interferon-gamma production by TH1 and TH0 cells predominates over IL-4 production by TH2 and TH0 cells. Understanding the relationship between the observed biphasic response and clinical manifestation of AD is important for the development of therapeutic strategies.

T-cell reactivity for a peanut-derived epitope in the skin of a young infant with atopic dermatitis ☆ ☆☆ ★ ★★

A role for food allergen–specific T cells in the pathogenesis of atopic dermatitis (AD) in infants with food allergy is indicated from studies performed with peripheral blood lymphocytes.1 However, no reports exist that describe the isolation of food allergen–specific T cells from the skin of infants with AD. We investigated the presence of cow’s milk–and peanut-specific T cells in the skin of the described subject.

Evaluation of the atopy patch test and the cutaneous late-phase reaction as relevant models for the study of allergic inflammation in patients with atopic eczema.

OBJECTIVE The study was designed to evaluate the atopy patch test (APT) and the late-phase reaction (LPR) after intracutaneous allergen injection as models for the study of allergic inflammation in atopic eczema. METHODS Immunocytochemistry was used to analyze skin biopsy specimens from sites of APTs and LPRs at 2 and 24 hours and to compare these with lesional and nonlesional skin of patients with atopic eczema. RESULTS A lack of neutrophil infiltration in specimens from both the APT and lesional skin sites was observed, whereas neutrophils were abundantly present in the specimens from LPR sites. With double-staining techniques it was demonstrated that the few neutrophils present in specimens from APT sites and in lesional skin were mostly located in intravascular areas, whereas in the LPR specimens they were located predominantly in extravascular areas. Eosinophils infiltrated at an earlier time point in the LPR as compared with the APT. Furthermore, there was a decrease of intact mast cells in the LPR site compared with the APT sites and lesional skin. No significant difference in T-cell number was observed between the two tests. Upregulation of E-selectin expression on endothelial cells occurred at an earlier time point in the LPR as compared with the APT. CONCLUSION There are important differences in cellular infiltrate between the APT and the LPR. The close macroscopic and microscopic similarities between the specimens from APT sites and lesional skin of patients with atopic eczema support the argument that the APT is a more valid in vivo model with which to study allergic inflammation in atopic eczema than the LPR.

Adhesion molecule expression on skin endothelia in atopic dermatitis: Effects of TNF-α and IL-4

Abstract Background: Atopic dermatitis (AD) is characterized by skin infiltrates of leukocytes, such as lymphocytes and eosinophils. Objective: To describe the mechanisms determining this inflammatory process, we have analyzed expression of adhesion molecules and their regulation on skin endothelial cells (ECs). Methods: Expression of adhesion molecules on ECs was analyzed by immunohistochemistry by using Ulex europaeus agglutin 1 as a pan-endothelial marker. Results: Vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and P-selectin were not found in skin of nonatopic individuals, whereas expression of these surface molecules was observed in nonlesional skin of patients with AD and was even more pronounced in lesional skin or after epicutaneous application of aeroallergen. Induction of adhesion molecule expression was examined on both macrovascular ECs from human umbilical cord vein (HUVECs) and human microvascular ECs (HMEC-1) from skin. TNF-α very potently upregulated adhesion molecule expression in vitro on both EC cell types. To verify the in vivo relevance of TNF-α, we performed TNF-α staining in the skin. TNF-α was observed in the dermis of nonatopic skin, both in chymase-containing mast cells and CD68 + macrophages. The increase in the number of TNF-α–containing cells was concomitant with the increase in adhesion molecule expression in the skin of patients with AD. IL-4 is supposed to be important in atopic diseases because of its IgE- and VCAM-1–inducing properties. However, IL-4 addition failed to induce VCAM-1 expression on HMEC-1, although in the same set of experiments, a clear induction of VCAM-1 expression by IL-4 on HUVECs was demonstrated. Flow cytometry revealed the absence of IL-4 receptor α-chains on HMEC-1 and their presence on HUVECs. Immunohistochemistry examination on skin sections showed no binding of the IL-4R α-chain antibodies to ECs. Conclusion: We conclude that adhesion molecule expression is increased in the skin of patients with AD. Most probably, this increased expression is not a (direct) effect of IL-4 on skin endothelium, but other cytokines, such as TNF-α, might be responsible for this increased adhesion molecule expression. Continuous adhesion molecule expression may facilitate T-cell extravasation in a nonantigen–specific manner, thus explaining the presence of increased T-cell numbers in nonlesional skin of patients with AD. (J Allergy Clin Immunol 1998;102:461-8.)

Antigen focusing by specific monomeric immunoglobulin E bound to CD23 on Epstein-Barr virus-transformed B cells

Monomeric IgE bound to the low-affinity receptor for IgE (FcERII-CD23) on EBV-transformed human B cells selectively enhances binding of antigen and therefore presentation to specific T-cell clones. To demonstrate the role of monomeric IgE in antigen focusing, we have made use of a system consisting of human T-cell clones specific for Der-P1 (major allergen of the Dermathophagoides pteronyssinus), Der-P1 coupled to NIP (Der-P1-NIP), and the commercially available chimeric (human-murine) monoclonal IgE antibodies with specificity for the hapten NIP. We have found that monomeric IgE binds to CD23 and remains detectable on the surface of the B cells for a period of at least 16 hours at 37 degrees C. Pulsing of these IgE-anti-NIP (1 microgram/ml) treated B cells for 1 hour at 37 degrees C with low amounts (10 ng/ml) of Der-P1-NIP antigen allows the B cells to stimulate Der-P1-specific T cells. Even with IgE concentrations as low as 20 ng/ml, which were not detectable by immunofluorescence, we were able to induce a significant T-cell response. Furthermore, ongoing specific T-cell-B-cell interactions were not inhibited by the presence of high concentrations of nonspecific IgE molecules (incubated with up to 25 micrograms/ml) on the surface of the B cells. Our data confirm the hypothesis that IgE, bound by either CD23 or the high-affinity receptor for IgE, potentiates the immune response. Therefore, IgE may be seen as the fourth general mechanism for antigen capture by (nonspecific) antigen-presenting cells.

Nonspecific T-cell homing during inflammation in atopic dermatitis: Expression of cutaneous lymphocyte-associated antigen and integrin αEβ7 on skin-infiltrating T cells

Atopic dermatitis (AD) is a chronic skin disorder, characterized by infiltration of activated memory CD4+ T cells into skin. A model to study the onset of allergic inflammation in a patient with AD is the atopy patch test (APT), in which, by epicutaneous application of aeroallergen, an eczematous reaction is induced in 50% of sensitized patients with AD. Extravasation of T cells into skin is thought to be critically dependent on expression of the surface molecule cutaneous lymphocyte-associated antigen (CLA), which recognizes and binds its ligand E-selectin on endothelium. We studied the dynamics of expression of CLA and the gut homing receptor alphaE beta7 (HML-1) on T cells in the skin of patients with AD and in APT reactions and nickel and sodium lauryl sulfate patch test reactions by means of immunohistochemical double staining of skin biopsy specimens. The results show an increase in the number of CD3+ T cells in the lesional skin of patients with AD, APT reactions, and nickel and sodium lauryl sulfate patch test reactions as compared with nonlesional skin of the same patients and nonatopic individuals. In contrast, the percentages of CLA+ T cells in the lesional skin of patients with AD, in the APT reactions, and in sodium lauryl sulfate and nickel patch test reactions were decreased. In addition, we found a marked expression of alphaE beta7 by T cells present in skin, indicating a nonspecific influx of T cells during allergic skin inflammation. We propose that during allergic skin inflammation CLA expression is not a prerequisite for cutaneous T-cell infiltration. CLA expression may be important for T cells to extravasate from blood into skin during immune surveillance or for retention of allergen-specific T cells in skin.

IgE production after antigen-specific and cognate activation of HLA-DPw4-restricted T-cell clones, by 78% of randomly selected B-cell donors.

The frequency of expression of the MHC class II antigen, HLA-DPw4, in the caucasoid population is approximately 78%, and is unmatched by phenotypic frequencies of other HLA class II molecules. Here we describe three human Der-P1-specific T-cell clones (TCC), restricted by the HLA-DPw4-variant HLA-DPB1*0401, of which two TCC also responded to antigen, presented on HLA-DPB1*0402. Thus, randomly selected caucasoid donors present a 78% chance for a correct match with these HLA-DPw4-restricted TCC. This allows comparative in vitro antigen presentation studies with various antigen presenting cells (APC) from different (healthy or diseased) donors without the variable influence of responding T cells. It was subsequently demonstrated that the TCC can be used to study antigen-induced IgE production in randomly selected primary B cells. Cognate HLA-DPw4-restricted antigen presentation caused enhanced immunoglobulin production of IgE, IgG1, IgA and IgM, of which only IgE induction was reversed by addition of anti-IL-4 antibodies.