Immunoglobulin E antibody reactivity to bacterial antigens in atopic dermatitis patients

Abstract

Background: Atopic dermatitis (AD) is a chronic inflammatory skin disease affecting up to 20% children and 9% adults world-wide. AD patients are often sensitized against a broad variety of allergens and more than 90% of them suffer from skin superinfections with Staphylococcus aureus.

Objective: In this study, we searched for the presence of specific IgE antibodies against S. aureus and Escherichia coli antigens in AD patients.

Methods: Sera from AD patients (n=79), patients suffering only from allergic rhinoconjunctivitis (n=41) or allergic asthma (n=37) were tested for IgE reactivity to nitrocellulose-blotted S. aureus, E. coli and gut bacterial antigens. IgE-reactive bacterial antigens were affinity purified and identified by mass spectrometry.

Results: More than 30% of AD patients but not patients suffering only from allergic rhinoconjunctivitis and asthma or non-allergic persons exhibited IgE binding to several protein antigens among them DNA-binding and ribosomal proteins and flagellin. Patients with severe skin manifestations showed more frequently IgE reactivity to S. aureus compared with AD patients with mild symptoms. Positive immediate and late skin test reactions could be induced in sensitized AD patients with S. aureus extract.

Conclusion and clinical relevance: Specific IgE reactivities against a variety of bacterial antigens were observed in a subgroup comprising a third of AD patients and may contribute to allergic inflammation.

Figure 1. IgE reactivity of AD patients to nitrocellulose-blotted S. aureus and E. coli protein extracts

Total protein extracts of S. aureus (A) and E. coli (B) were separated under reducing conditions by SDS-PAGE, blotted on to nitrocellulose membrane and probed with sera from 35 patients with AD. As a control, serum from a non-atopic individual (panel N) was used. Molecular weights (kDa) are indicated in the left margin.

Figure 2. IgE reactivity of AD patients with and without skin superinfections to nitrocellulose-blotted S. aureus and E. coli protein extracts

Total protein extracts of S. aureus (A) and E. coli (B) were separated under reducing conditions by SDS-PAGE, blotted on to nitrocellulose membrane and probed with sera from the German AD patient populations with or without skin superinfections. As a control, serum from a non-atopic individual (panel N) was used. Molecular weights (kDa) are indicated in the left margin.

Figure 3. IgE reactivity of AD patients to nitrocellulose-blotted protein extracts from gut-colonizing bacteria

Nitrocellulose-blotted protein extracts of S. salivarius, K. pneumoniae, E. faecalis and B. fragilis which are ileum-colonizing bacteria (A) and B. adolescentis, C. ramosum and L. acidophilus which are colon-colonizing bacteria (B) were probed with sera from patients with AD, with IgE to E. coli proteins (E. coli +) or without (E. coli −). Lane B denotes buffer control. Patient numbers are as in Tables 1 and 2. Molecular weights (kDa) are indicated in the left margin.

Figure 4. Low cross-reactivity between IgE-reactive antigens from S. aureus and E. coli

Nitrocellulose-blotted S. aureus extracts (A) and E. coli extracts (B) were probed with sera from selected AD patients, which were pre-adsorbed with protein extracts of either S. aureus (lanes S), E. coli (lanes E), or with bovine serum albumin (BSA; lanes -). Patient numbers are as in Tables 1 and 2. Molecular weights (kDa) are indicated in the left margin.

Figure 5. Lymphocyte proliferation and cytokine releases induced in PBMC cultures of AD patients and non-allergic individuals by stimulation with S. aureus and E. coli protein extracts

(A) PBMCs from AD or non-atopic (NA) individuals were stimulated with S. aureus extract, E. coli extract or medium (MC). Mean lymphoproliferative responses of triplicate determinations were measured by ³H-thymidine incorporation and are displayed as stimulation indices. Horizontal lines denote the median value of each group. (B) The amounts of released cytokines are presented and horizontal lines denote the median value of each group.

Figure 6. Immediate and late type skin reactions to S. aureus extract in AD patients

Twenty two patients with AD, with or without IgE-reactivity to S. aureus proteins, were injected intradermally with S. aureus extract or saline buffer. Average wheal diameter was measured 20 minutes (immediate reaction) and 24 hours (late reaction) after the challenge.