Lack of proliferative response by gluten-specific T cells in the blood and gut of patients with dermatitis herpetiformis.

The majority of patients with Dermatitis Herpetiformis (DH) have a gluten-sensitive enteropathy which may be triggered by a T cell-mediated immune response to gluten. Using a proliferative assay, the responses to gluten fraction III, recall antigens and mitogens of peripheral blood mononuclear cells (PBMC) and gut T cell lines (TCL) isolated from patients with Dermatitis Herpetiformis (DH) and normal controls were studied. In most cases, neither PBMC nor gut T cell lines (which were predominantly CD3+, CD4+, TCR alpha beta +) from either controls or patients proliferated in response to gluten fraction III alone. However, the addition of 10 U/ml IL-2 to PBMC cultures containing gluten fraction III resulted in a marked increase in proliferation in 9/19 DH patients and 7/11 controls compared to IL-2 alone. Furthermore, gluten-induced upregulation of IL-2 receptor (CD25) expression was demonstrated on PBMC from 4/4 patients with DH and 2/3 controls after 7 days' culture with antigen. A similar effect by exogenous IL-2, or the same concentration of IL-4, was observed in 8/11 (P = 0.02) and 5/6 respectively DH, and 3/4 normal gut T cell lines. No difference was observed in the response of DH and control PBMC to Tetanus toxin, Candida albicans and PPD; both normal and DH gut T cell lines were unresponsive to these antigens. However, the addition of IL-2 increased the response to Candida albicans by DH gut T cell lines. Moreover, the response of DH gut T cell lines to PHA (P < 0.001), Concanavalin A and anti-CD3 were markedly reduced compared to PBMC from the same patients. These findings suggest that gluten-specific T cells present in the blood and gut of normal and DH individuals are activated by but do not proliferate in response to specific antigen.

Absence of gluten-specific T lymphocytes in the skin of patients with dermatitis herpetiformis.

Dermatitis Herpetiformis (DH) is an immunobullous skin disease with an associated gluten-sensitive enteropathy. Withdrawal of gluten from the diet leads to the resolution of both the skin lesions and the enteropathy. A T cell-mediated immune response to gluten has been implicated in the damage to the gut; the possible gluten specificity of the T cell infiltrate in DH skin lesions has not, however, been investigated. T cell lines (TCL) were therefore established from the skin lesions of eight patients with DH by culturing skin fragments for 11-17 days with a medium supplemented with 20 U/ml of IL-2. In three cases, gliadin (fraction of gluten toxic to the DH gut) and irradiated, autologous peripheral blood mononuclear cells were also added. The TCL were stained for CD3, CD4, CD8, TCR alpha beta and gamma delta expression by indirect immunofluorescence, and their proliferative responses to mitogens and gluten fraction III (a peptic-tryptic digest of gluten) investigated. Of the eight CD3+ TCL, four were predominantly CD4+ (82.1-98.8%), three predominantly CD8+ (92.6-98.6%) and one TCL contained both 87.6% CD4+ and 95.2% CD8+ T cells, a substantial proportion of which were presumably double-labelled CD4+, CD8+ T cells. All eight TCL, which were almost exclusively TCR alpha beta +, proliferated in response to PHA whilst six out of the eight were stimulated by Concanavalin A. None of the TCL proliferated to gluten fraction III alone; however, two TCL showed increased proliferation to the antigen in the presence of exogenous IL-2 or IL-4 (10 U/ml) compared to cytokine alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Decreased T cell reactivity to trypsinized group A, type M22 streptococci in psoriasis.

The proliferative responses of peripheral blood mononuclear cells from patients with guttate and chronic plaque psoriasis to streptococcal M protein were investigated using whole and trypsinized group A M22-positive streptococci. Peripheral blood mononuclear cell responses to whole type M22 group A streptococci were significantly increased in guttate, but not chronic plaque, psoriasis patients compared to 17 non-psoriatic controls (p < 0.05; n = 17). A significant reduction of this response was observed in both guttate (p < 0.001; n = 17) and chronic plaque (p < 0.01; n = 27) psoriatic patients, but not in the control group, after repeated trypsinization to remove M protein from the streptococci. Furthermore, the difference between the peripheral blood mononuclear cell response to untrypsinized and trypsinized streptococci was significantly greater in the guttate patients than in the controls (p < 0.02). This preliminary study has shown an increased reactivity of T lymphocytes with specificity for trypsin-sensitive protein expressed by type M22 streptococci in the peripheral blood of patients with psoriasis.

Restricted T-cell receptor V beta gene usage in the skin of patients with guttate and chronic plaque psoriasis.

A strong association between acute guttate psoriasis and group A, beta-haemolytic streptococcal infections is well established. Furthermore, streptococcal M proteins and toxins have been shown to act as superantigens, stimulating subpopulations of T lymphocytes expressing particular V beta families. We have therefore studied the possible role of streptococcal superantigens in psoriasis by staining peripheral T lymphocytes and skin sections from patients with guttate or chronic plaque psoriasis for the expression of nine TCR V beta families, using a range of monoclonal antibodies. A marked over-representation of V beta 2+ T lymphocytes was observed in the dermis and epidermis of patients in both groups, when compared with T lymphocytes in their peripheral blood. A less marked dermal increase in V beta 5.1+ T lymphocytes was also observed in these patients. These findings are consistent with the involvement of a superantigen, possibly streptococcal, in the pathogenesis of psoriasis.

Group A streptococcal antigen-specific T lymphocytes in guttate psoriatic lesions.

A strong association exists between guttate psoriasis and group A, beta-haemolytic streptococcal infections. To demonstrate the presence of streptococcal-specific T cells in psoriatic skin, T-cell lines (TLs) were established from biopsies of lesions from five patients with guttate psoriasis, and compared with TLs from five patients with eczema, five with lichen planus, two with pityriasis rosea and three with nickel contact dermatitis. TLs from purified protein derivative (PPD)-induced delayed hypersensitivity sites in three normal individuals were also studied. All five of the psoriatic TLs responded in a proliferation assay to heat-killed isolates of group A streptococci, compared with only one eczema, two lichen planus and one pityriasis rosea. The response of one nickel contact dermatitis and two PPD TLs to group A streptococci was markedly less than to nickel and PPD, respectively. One of the psoriatic TLs was cloned in the presence of type 5 streptococcal M protein. The nine clones obtained were all CD3+, CD4+, CD45RO+, TCR alpha, beta+, gamma, delta-. However, they were all unreactive with antibodies to TCR V beta 5, 6, 8 or 12. Eight of the nine clones reacted, to a varying extent, to one or two of three preparations of group A streptococci expressing different M proteins. The streptococcal response of four consistently reactive clones from this patient was HLA-DR-restricted and inhibited by anti-HLA-DR antibody in a dose-dependent manner. On stimulation these four clones secreted high levels of gamma-interferon and detectable levels of IL-2, IL-10 and granulocyte/macrophage colony stimulating factor (GM-CSF) depending upon the nature of the stimulus, but no IL-4 or TNF-alpha production was detected. This study has demonstrated, for the first time, that T lymphocytes specific for group A streptococcal antigens can be consistently isolated from guttate psoriatic lesions. The role of streptococcal-specific T cells in the pathogenesis of psoriasis remains to be determined.

Interactions between lymphocytes and dermal fibroblasts: an in vitro model of cutaneous lymphocyte trafficking.

Cultures of dermal fibroblasts were established from skin biopsies of CBA mice and used to study the interactions with murine T-lymphocytes. Electron microscopy showed that zones of contact developed between the fibroblasts and the T-cells, particularly after mitogenic activation. The adhesion of the lymphocytes was temperature-dependent, and many more lymphoblasts than resting cells attached to the fibroblast monolayers. Flow cytometry analysis of the adherent population showed that the most prominent type of resting lymphocyte was of the CD4 phenotype, which was also observed using a T-helper lymphoid cell line. However, neither the CD4 nor the CD8 (T-cytotoxic) antigens were involved in the binding process, and while the fibroblasts expressed Class I MHC molecules (but not Class II), these also had no role in mediating lymphocyte adhesion. Although the fibroblasts did not express the ligand Mala-2, the murine homologue of human ICAM-1, a monoclonal antibody against LFA-1, its cognate receptor on the lymphocytes, nevertheless effectively inhibited binding. T-cell attachment was also partially prevented by antibody against the lymphocyte CD2 antigen and by RGDS, a protein epitope known to mediate a number of receptor-integrin interactions. Moreover, this peptide also rapidly and preferentially detached T-lymphocytes which had previously adhered to the fibroblast monolayers. Lymphocyte binding was substantially elevated following treatment of the fibroblasts with cytokines such as tumor necrosis factor-alpha and interferon-gamma, but not interleukin-1 alpha. This increase in adhesiveness was, however, almost completely abolished by monoclonal antibodies specific for LFA-1 or for Mala-2. The results of this study show that while lymphocytes recognize fibroblasts normally via a number of constitutively expressed receptor-integrin interactions, their adhesion can also be modulated by cytokine-induced changes in the expression of other surface ligands.