Interleukin-4 Promotes the Expansion of Skin-Infiltrating Lymphocytes from Atopic Dermatitis In Vitro

Abstract

Functional studies of lymphocytes in atopic dermatitis (AD) have so far focused on peripheral blood mononuclear cells (PBMC), whereas cells at the involved site, the skin, have not been examined. Accordingly, we have developed methods to generate lymphocyte cultures from biopsies of inflammatory skin areas. Skin-infiltrating lymphocytes (SIL) were isolated from skin biopsies of 6 patients with severe AD and expanded in vitro in the presence of interleukin-2 (IL-2) without additional antigens. After 6-10 d in culture, outgrowth of mononuclear cells from biopsy tissue was observed in all cases. Phenotypic analysis of skin-derived cells revealed the predominance of CD4+ T-helper/inducer phenotype in SIL populations. Parallel cultures of SIL and PBMC showed an increase and expansion of CD8+ T cells in cultured PBMC, whereas the CD4+ phenotype was predominant in SIL cultures. As indicated by their expression of HLA-DR and CD25 antigens, most of the SIL were activated and the cells mainly expressed T-cell receptors (TCR) composed of alpha and beta chains. Different strategies for expansion of SIL in vitro were examined. High levels of IL-4 (1,000 U/ml) in combination with IL-2 (50 U/ml or 1,000 U/ml) preferentially promoted growth of SIL derived from AD and were much more effective than IL-2 alone. No cells expanded in cultures with IL-4 alone. SIL grown with high concentrations of IL-4 contained a significant proportion of double-positive CD4+8+ cells. No other marked differences were observed in the distribution of T cell subsets in cultures propagated under different conditions for 21 d. Our results demonstrate the feasibility of growing infiltrating T lymphocytes from inflammatory skin of AD patients. The use of high concentrations of IL-2 in combination with high levels of IL-4 allows a large expansion of these cells and thus represents a useful strategy to expand cells for further functional and molecular biologic studies.

Different growth factor requirements for human Th2 cells may reflect in vivo induced anergy

“…In addition. IL-4 was shown lo enhance the proliferative elTect of IL-2 on primary human AD skin-derived polyclonal T cell cultures [36].…”

The Relationship Between Plasma Thrombospondin Level and the Clinical Course of Atopic Dermatitis

“…For instance, it was reported recently that T cells from patients with atopic dermatitis respond more strongly to interleukin (IL) 4 26 than do T cells from normal individuals and that the proliferation of subpopulations of T cells in lesional atopic skin depends on IL-4. 27 Moreover, IL-4 and IL-5 have been shown to enhance the development of cytotoxic effectors, that cause tissue damage. 28 • 29 Most cytokines, including IL-4 and IL-5, are potent proinflammatory mediators that prime granulocytes for an enhanced generation of leukotrienes.…”

Pathophysiology of itching in urticaria and atopic dermatitis

“…Experimental data from peripheral blood T cells suggest that an imbalance in the interleukin-4/interferon-y ratio may be of great importance in the mechanisms involved in AD (59). Techniques for culturing skin-infiltrating lymphocytes obtained via skin biopsies from lesional AD skin have recently been introduced (102)(103)(104)(105) and offer an exciting possibility of studying the functional activity and spectrum of cytokine production of lesional skin lymphocytes. Such in vitro-expanded cultures of AD-skin-derived lymphocytes, stimulated with mitogens or allergens, produce interleukin-4, granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumour necrosis factor-cx, whereas the production of interferon-?…”