Failure to detect specific gluten antigens associated with the immune aggregates in the skin in dermatitis herpetiformis

Summary The univolved skin of 10 patients with dermatitis herpetiformis (DH) was examined for the presence of gluten antigens with the immunofluorescence technique using a rabbit anti-gliadin antiserum, human antibodies to wheat and to reticulin conjugated to fluorescein-isothiocyanate (FITC) and class-specific anti-human lgA immunoglobulin.In all patients, IgA deposits were found in the tips of the dermal papillae of the uninvolved skin. With the anti-wheat and anti-reticulin conjugates, as well as with the rabbit anti-gliadin antiserum, no specific immunofluorescence was observed in any of the skin specimens. Skin biopsy sections of three DH patients were treated with an acid solution (pH 3.2) in an effort to dissociate antigen-antibody complexes that might be present. After the elution procedure the sections showed undiminished IgA fluorescence, and retesting with the anti-wheat- and antireticulin antisera again gave negative results. The skin eluates, two of which contained IgA, had no antibodies to wheat or reticulin.These findings do not give support to the hypothesis that the antigens in the suspected immune complexes in the DH skin consist of gluten.

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Zusammenfassung Die nicht angegriffene Haut von 10 Patienten mit Dermatitis Herpetiformis (DH) wurde auf die Anwesenheit von Gluten-Antigenen mit der Immunfluorescenztechnik untersucht unter Anwendung eines Antigliadin-Antiserums in einem Kaninchen erzeugt, menschliche Antikörper gegen Weizen sowie gegen Retikulin verbunden mit Fluorescein-Isothiocyanate (FITC) und mit spezifisch klassifiziertem anti-menschlichem IgA-Immunglobulin.In allen Patienten fanden sich IgA-Niederschläge in den Spitzen der Dermalpapillen der nicht angegriffenen Haut vor. Sowohl mit den Antiweizen- und Antiretikulinverbindungen als auch mit dem erwähnten Antigliadin Antiserum konnte keine spezifische Immunfluorescenz in den untersuchten Häuten festgestellt werden. Hautbiopt-Schnitten von 3 DH-Patienten wurden mit einer Säurenlösung (pH 3,2) erprobt, um zu versuchen, die Antigen-Antikörperverbindungen, die möglicherweise anwesend sein könnten, voneinander zu trennen. Nach der oben erwähnten Erprobung zeigten die Versuchsteile eine ungeänderte IgA-Fluorescenz, und nach Wiederholung der soeben genannten Probe mit Antiweizen- und Antiretikulin-Antikörper zeigten sich abermals negative Resultate.Die Hauteluaten, von denen zwei IgA enthielten, zeigten keine Antikörper gegen Weizen oder Retikulin. Diese Resultate tragen nicht zu der Hypothese bei, daß die Antigene in den vermutlichen Immunverbindungen in der DH-Haut aus Gluten bestehen.

     

Class-specific antibodies to gluten in dermatitis herpetiformis

An immune reaction to wheat protein has been previously proposed to explain the pathogenesis of dermatitis herpetiformis. In order to detect and characterize antibodies to gluten in human sera, we developed an enzyme immunoassay for class-specific antibodies. Results of this assay in 49 patients with dermatitis herpetiformis were compared with those of 38 normal control subjects, 11 patients with celiac disease, and 6 small-bowel bypass patients. IgA antibodies to gluten were significantly more frequent in dermatitis herpetiformis sera (28/49) than in normal control sera (4/38). IgG antibodies to gluten were significantly more frequent in both celiac disease (10/11) and dermatitis herpetiformis (16/49) sera than in control (5/38) sera. Dermatitis herpetiformis sera also had an increased prevalence of IgM antibodies to gluten (19/49). Small-bowel bypass patients demonstrated no antibody to gluten. Antibodies to gluten in dermatitis herpetiformis objectively mark a state of immune reactivity to wheat protein and may be involved in the genesis of the cutaneous IgA immune deposits and the skin disease.     

Immediate and delayed hypersensitivity reactions to birch pollen in patients with atopic dermatitis.

We investigated immediate and delayed hypersensitivity to birch pollen in 10 patients with atopic dermatitis (AD) who had experienced a worsening of their eczema during the birch pollen season. The patients were prick- and patch-tested and antigen-induced basophil histamine release and lymphocyte proliferation were measured. 9/10 birch pollen-allergic patients proved positive in the histamine release test and the results correlated with specific IgE levels measured by RAST. Birch pollen antigen induced lymphocyte proliferation in 6/10 patients, but a positive patch test result was obtained in only one case. Both peripheral blood monocytes and purified epidermal Langerhans' cells were able to present birch pollen antigen to T cells, although Langerhans' s cells seemed to function less efficiently in this respect.     

Serum antibodies to wheat germ agglutinin and gluten in patients with dermatitis herpetiformis

Summary It has been speculated that gluten may play a role in the pathogenesis of dermatitis herpetiformis (DH) because it can act as a lectin. The lectin activity of gluten preparations was recently identified as wheat germ agglutinin (WGA). IgG and IgA serum antibodies to WGA and gluten were therefore measured in patients with DH and coeliac disease (CD) by an enzylac-linked immunosorbent assay (ELISA). Compared with healthy controls, both patients categories had increased IgG and IgA activities to WGA and gluten, the CD group showing the highest antibody levels. DH patients with subtotal villous atrophy tended to have higher activities than those with no villous changes or only minor changes. No significant difference in the gluten-to-WGA ratio of IgA or IgG antibodies was found when DH patients were compared with CD patients. If WGA plays a pathogenetic role in DH, then DH patients must have dermal characteristics, as yet undefined, that explain the initiation of their skin disease.     

Serum IL-8 in patients with dermatitis herpetiformis is produced in response to dietary gluten

Patients with dermatitis herpetiformis (DH) have a gluten-sensitive enteropathy and while on gluten-containing diets have elevated levels of serum IL-8. We hypothesized that the mucosal immune response to gluten is responsible for the elevated serum IL-8. Six DH patients were studied while on a gluten-free diet (GFD), whereas four continued on a normal diet. Patients were followed for a mean 2.2 years and serum IL-8 was analyzed. Small bowel biopsies from five DH patients on normal diets, two DH patients on GFD, and six subjects with no small bowel abnormalities were analyzed for IL-8 mRNA. Serum IL-8 levels normalized in five of six patients on GFD and decreased in one, whereas serum IL-8 levels showed no statistically significant change in DH patients on normal diets. Small bowel biopsies from DH patients on normal diets had increased expression of IL-8 mRNA compared to normal subjects, whereas patients on a GFD showed no significant increase in small bowel mRNA. No significant IL-8 mRNA was detected in normal skin biopsies from patients with DH. These observations suggest that the IL-8 in the serum of patients with DH originates from the small bowel as a mucosal immune response to gluten ingestion.     

T lymphocytes in lesional skin of patients with dermatitis herpetiformis

Ten patients with dermatitis herpetiformis had biopsies taken from involved skin.Monoclonal antibodies and the avidin-biotin peroxidase staining technique were used to stain for T cells and Langerhans cells in skin sections. A significant increase in the number of CD3-positive T cells was observed in the upper dermis of involved compared with uninvolved skin (P<0.0005). Most of the T cells in involved skin were CD45RO-positive memory cells; CD4-positive T cells exceeded the number of CD8-positive T cells by a ratio of 4:1. In addition, CD1a-positive dendritic cells were observed within the clumps of T cells in involved dermis in nine of the 10 patients, but were absent from the dermis of uninvolved skin. Double immunofluorescent staining demonstrated that approximately 20–40% of the CD3-positive T cells were activated, and expressed the HLA-DR antigen. These findings suggest that activated T cells are involved in the pathogenesis of dermatitis herpetiformis skin lesions.     

TCR Vbeta expression in the small bowel of patients with dermatitis herpetiformis and gluten sensitive enteropathy. Limited expression in dermatitis herpetiformis and treated asymptomatic gluten sensitive enteropathy

Dermatitis herpetiformis (DH) is a blistering skin disease characterized by cutaneous deposits of IgA and an associated, most often asymptomatic, gluten sensitive enteropathy (GSE). Gluten sensitive enteropathy is also seen in patients that do not have skin disease or cutaneous IgA deposits, but do have significant gastrointestinal (GI) complaints. Patients with DH and with GSE without skin disease have similar small bowel morphologic changes and HLA associations and both the skin disease and the GI symptoms can be controlled by a gluten free diet. It is not known what factors allow almost all patients with DH to continue to eat gluten and not develop symptomatic gastrointestinal disease. We have examined the expression of the Vbeta T-cell receptor (TCR) in the small bowel of patients with DH (n=11) and of patients with both symptomatic (n=10) and asymptomatic (n=7) GSE without skin disease to determine if differences in the pattern of TCR Vbeta expression are associated with differences in the clinical manifestations of these diseases. TCR Vbeta expression was analyzed using RT-PCR from small bowel biopsies. Patients with DH and those with GSE without skin disease that were on a gluten free diet and asymptomatic were found to express 6.6 and 5.6 out of 20 Vbeta families respectively, with no single family preference. Examination of peripheral blood lymphocytes from these patients did not reveal any restriction of TCR Vbeta family expression. In contrast, patients with symptomatic GSE expressed 12.6 Vbeta families (P< 0.05), with no consistent preferential expression of any single Vbeta family between patients. Patients with DH, who are continuing to ingest wheat, show a more restricted pattern of TCR Vbeta utilization, similar to that of treated patients with GSE without skin disease, and significantly different from GSE without skin disease patients eating gluten. These findings suggest that the restricted nature of the TCR Vbeta expression may play a role in the different clinical manifestations of dermatitis herpetiformis and isolated gluten sensitive enteropathy.     

Relationship of dietary gluten intake to dapsone dose in dermatitis herpetiformis

The gluten intake was quantitated utilizing a dietary history method in 43 patients with dermatitis herpetiformis on non-restricted diet. The mean daily gluten intake was 15 g. The individual intake of gluten was related to the maintenance dose of dapsone. It was significantly higher in patients on 100-150 mg dapsone daily than in those taking 0-25 mg daily. There was a significant correlation between amount of gluten in the diet and the dapsone dose (p less than 0.01, rs = 0.43). Villous atrophy was not related to the dapsone dose. It is suggested that the gluten-sensitive enteropathy changes the intestinal permeability and thus contributes to the development of blisters.     

Lymphocyte proliferation response in patients with delayed hypersensitivity reactions to heparins

Background  Heparins can induce delayed-type hypersensitivity (DTH) reactions mediated by specific T lymphocytes. However, the interaction between heparins and lymphocytes has not been sufficiently studied.
Objectives  To analyse the lymphocyte response to heparins using different types of antigen-presenting cells in patients with DTH reactions to these drugs.
Methods  We studied seven patients with DTH reactions to heparins diagnosed by delayed reading of intradermal skin testing ( n  =   5) or drug provocation test ( n  =   2) and nine tolerant controls. Biopsies were obtained from intradermal testing or during the acute reaction. Peripheral blood mononuclear cells were used to obtain T and B lymphocytes, monocytes and monocyte-derived dendritic cells (DC). T-lymphocyte proliferation was assayed by means of ³H-thymidine incorporation.
Results  Skin testing showed a high degree of cross-reactivity within low molecular weight heparins with good tolerance to sodium heparin, fondaparinux and lepirudin in most cases. The proliferative response was positive in six patients to most of the heparins tested with both monocytes and B cells (the classical lymphocyte transformation test) or immature DC as antigen-presenting cells, giving a higher response with DC. At a second evaluation 1 year later, the proliferative response was found only with DC, and mainly to the culprit drug.
Conclusions  A model using DC in the lymphocyte proliferation test is a more appropriate way to assess the immunological response in DTH to heparins; additionally, it can detect a response over a longer time. These findings may be useful for the diagnostic evaluation of drug reactions.     

IgA anti-endomysial antibody detection in the serum of patients with dermatitis herpetiformis following gluten challenge

Summary This study reports the appearance of IgA-class anti-endomysial antibodies in the serum of 8 out of 12 patients with dermatitis herpetiformis who were challenged with gluten after a number of years of control of the rash with a strict gluten-free diet. Although there was no evidence for the antibodies having any pathogenic role in the rash of dermatitis herpetiformis, their presence may be related to the deterioration in the gluten-sensitive enteropathy.     

IgA and C3 complement in the uninvolved skin in dermatitis herpetiformis after gluten withdrawal

IgA deposits in the skin in 53 patients with dermatitis herpetiformis (DH) have been studied in relation to treatment. In 19 patients the disorder was controlled by a gluten-free diet (GFD) alone, in 13 patients by dapsone and GFD and in 18 by dapsone alone. In 3 patients the skin disorder became insignificant and required no treatment. Of the patients taking a GFD alone, six had been clear of skin lesions for 7 years, 5 for 3–5 years, and 8 for periods of 6 months-3 years. IgA deposits were found in all patients in an initial biopsy and in a second biopsy after treatment for periods varying from I to 7 years. There was no difference in the quantity of IgA, as assessed by the amount of fluorescence, whether patients were controlled with a GFD alone, GFD and dapsone, dapsone alone, or in those in clinical remission. The C₃ component of complement was present in the skin in 3 of the 19 patients (16%) controlled by a GFD alone, 6 of the 13 patients (46%) of those controlled by a GFD and dapsone, and in 12 of 18 (66%) of the patients taking dapsone alone, and in one of the patients in clinical remission.     

IgG antibodies and early intradermal reactions to hydrocortisone in patients with cutaneous delayed-type hypersensitivity to hydrocortisone

Seven of 25 patients with cutaneous delayed-type hypersensitivity to hydrocortisone had an immediate reaction following the intradermal injection of hydrocortisone sodium succinate. Using an ELISA method, we found that these patients had significantly increased levels of IgG antibodies to hydrocortisone when compared with normal blood donors (P<0.005) and nickel-allergic patients (P<0.05). We suggest that these patients are at risk of developing type III and possibly type I reactions following the systemic administration of hydrocortisone and that, if needed, an alternative systemic corticosteroid should be used, for example betamethasone or dexamethasone.     

Abnormal cutaneous reactions in dermatitis herpetiformis.

In 94% of the patients with dermatitis herpetiformis, a locally applied ointment with an ester of nicotinic acid (Trafuril) induced an abnormal reaction with erythema, edema, papules, and often vesicles. The appearance of the reaction to Trafuril is similar to DH lesions. It differs markedly from the reactions to iodide where, as a rule, only one large blister results. The reaction is most pronounced 12-24 hours after application of Trafuril. The reactivity is strongest in the predilection areas for dermatitis herpetiformis. Strongly increased reactivity to Trafuril in DH is induced by pretreatment with streptokinase, streptokinase-streptodornase, or urokinase. Diminished reactivity to Trafuril is seen after pretreatment with injections of Compound 48/80, tranexamic acid, cromoglycate, or application of fluocinolone-acetonide, as well as after stroking the skin.     

Long term follow-up of dermatitis herpetiformis with and without dietary gluten withdrawal

Seventy-eight patients with dermatitis herpetiformis have been followed up for periods ranging from 3 to 14 years (mean 7.4). Forty-two patients were treated with gluten-free diet (GFD) and thirty-six took a normal diet (ND). Thirty of the forty-two (71%) taking the GFD were able to discontinue drugs previously needed to control their rash compared with five (14%) of the thirty-six patients taking a ND. The mean time taken to reduce drug requirements for patients taking a GFD was 8 months (range 4–30), and for stopping drugs, 29 months (range 6–108). The incidence of macroscopic abnormality of the small intestine decreased from 69 to 15%, and the mean intra-epithelial lymphocyte count decreased significantly in those patients taking a GFD, whereas there was no significant change in patients taking a ND. The improvement in the skin and intestinal lesions was related to the strictness of the GFD.