Volume 116, Issue 1 , Pages 233-236, July 2005
Local isotype switching to IgE in airway mucosa
Article Outline
Antibodies, while maintaining their antigen-binding site, can alter their effector function by switching the constant portion of their heavy chain. This process is termed isotype switching and allows the formation of the allergy-associated antibody isotype IgE. Classically, isotype switching to IgE has been considered an event restricted to lymphoid tissue, such as the regional lymph nodes and spleen, and the IgE-positive cells found within peripheral sites of allergic inflammation, such as the respiratory mucosa, have been thought to migrate from these centers. However, IgE has been detected at sites of allergic inflammation in the absence of positive skin test responses or increased serum IgE levels.1 This indicates the possibility that B cells residing within the tissue might also participate in production of IgE by means of local isotype switching.
IgE isotype switching involves the recombination of genomic DNA. Exons encoding the constant portion of the IgE antibody heavy chain (Cε) are spliced downstream of the variable portion (VH) of the antibody, allowing for the transcription of IgE mRNA (Fig 1). Coordination of recombination is performed by specific genomic switch regions, Sμ and Sε, which are present upstream of Cμ and Cε, respectively. B cells require 2 signals to undergo isotype switching. First, the cytokines IL-4 and IL-13 initiate germline transcription of a promoter upstream of Sε, Iε RNA, composed of Iε spliced directly to Cε. This transcript is necessary but not sufficient for recombination to take place. Second, activated T cells expressing CD40 ligand activate B-cell CD40, allowing class-switch recombination to continue. Recombinase activity aligns Sμ adjacent to Sε. The DNA lying between is deleted Sμ, and Sε is removed, so that the Cε region is placed adjacent to VH, forming the template for IgE heavy chain mRNA. The ends of the deleted DNA are spliced together at the S regions, giving rise to circular DNA products called switch circles.
Fig 1.
Isotype switching involves 2 steps: germline transcription and DNA recombination. Cytokines cause the transcription of Iε RNA, which is essential for switching. CD40 ligand allows recombination to proceed. Products of isotype switching are a gene for an immunoglobulin isotype (here IgE) and a switch circle, which is composed of the intermediate region that was deleted from genomic DNA.
B cells can be identified in the nasal mucosa (Fig 2)4 and can be provided with the necessary signals for isotype switching. There are increased numbers of T cells expressing IL-4 and IL-13 mRNA. T cells, mast cells, basophils, and eosinophils express CD40 ligand, and within the mucosa of sensitized individuals, these cells are present in ample numbers (Fig 3). IgE-coated granulocytes, particularly mast cells, can release IL-4 and produce IL-13 on allergen cross-linking. This would lead to local B-cell isotype switching to IgE, in effect increasing mucosal IgE levels.
Fig 2.
B lymphocytes identified on the basis of CD20 immunoreactivity within allergic nasal mucosa cultured for 24 hours in the presence of specific allergen. B cells were observed just beneath the basement membrane (A), infiltrating the epithelial layer (B), and clustering within the submucosa in groups of 3 or 4 cells (C).4
Fig 3.
Immunofluorescence with anti-CD40 ligand antibody linked to FITC. Note the large number of positive cells in the submucosa of the upper airway.
IgE mRNA–positive B cells have been identified in nasal mucosa of patients with symptoms of allergy (Fig 4). For some time, however, it was not clear whether these were truly resident B cells or simply those that infiltrated the nasal compartment after switching to IgE in lymphoid tissue. Iε RNA–positive cells were identified in the nasal mucosa of patients with allergic rhinitis after acute allergen challenge (Fig 5)2 after seasonal exposure (Fig 6).3 Additionally, increased numbers of Iε RNA–positive cells and Cε RNA–positive cells were present in explanted nasal mucosa after allergen challenge (Fig 7, Fig 8).4 Explanted tissue is devoid of cell recruitment, confirming that initiation of local isotype switching exists. Actual DNA recombination was also shown by means of PCR, in which positive Sμ-Sε switch circle DNA was detected in explanted nasal mucosal tissue with ex vivo allergen challenge (Fig 9).5 This work indicates that in addition to the lymph nodes and spleen, isotype switching to IgE can also occur locally within the respiratory mucosa. These findings suggest an explanation for the presence of IgE within these tissues in the absence of positive skin test responses, serum IgE levels, or both.
Fig 4.
A, In situ hybridization for IgE mRNA (Cε RNA) in the nasal mucosa of an allergic patient. Black arrows point to positive cells. B, Colocalization of IgE mRNA with resident CD20-positive B cells. In situ hybridization was performed with the same probe, whereas CD20-positve cells were determined with horseradish peroxidase immunocytochemistry. White arrows point to double-positive cells.
Fig 5.
In situ hybridization–positive cells expressing Cε RNA and Iε RNA detected in patients with hay fever at baseline and 24 hours after an acute ragweed allergen challenge (P < .01, Student t test).2
Fig 6.
In situ hybridization–positive cells expressing Cε RNA and Iε RNA detected in patients with allergic rhinitis before the allergen season (BS) and during the peak season (PS; P < .001, Student t test).3
Fig 7.
Expression of ε RNA transcripts within explanted nasal mucosal tissue. Significantly higher numbers of Iε (A) and Cε (B) RNA–positive cells were observed in allergen-stimulated (Ag) compared with unstimulated (medium alone [MA]) allergic tissue but not within tissue obtained from nonallergic patients.4
Fig 8.
A, Representative example of Iε RNA in situ hybridization–positive cells found in an explant from nasal mucosa of a patient with rhinitis after ex vivo allergen challenge. B, Nasal explant model in which tissue biopsy samples taken from the inferior turbinate are cultured ex vivo and stimulated with different conditions. The tissue sits on a filter dish, which floats over the media and is exposed to air, thus simulating the in vivo nasal environment.
Fig 9.
A, SεSμ switch circle DNA found within nasal mucosal tissue cultured for 24 hours with media alone or ragweed extract. B, When isotype switching occurs, the switch circles are looped out of the genomic DNA. Switch circles can be detected by means of nested PCR with primer sets that amplify the SεSμ junction of the circles but not genomic DNA.5
We thank Drs Stephen R. Durham, Donata Vercelli and Hannah J. Gould, who have been great collaborators on the work of local isotype switching.
References
- . Local specific IgE production in nasal polyps associated with negative skin tests and serum RAST. Ann Allergy. 1985;55:736–739
- Expression of epsilon germ-line gene transcripts and mRNA for the epsilon heavy chain of IgE in nasal B cells and the effects of topical corticosteroid. Eur J Immunol. 1997;27:2899–2906
- Expression of IL-4, Cepsilon RNA, and Iepsilon RNA in the nasal mucosa of patients with seasonal rhinitis: effect of topical corticosteroids. J Allergy Clin Immunol. 1998;101:330–336
- Local synthesis of epsilon germline gene transcripts, IL-4, and IL-13 in allergic nasal mucosa after ex vivo allergen exposure. J Allergy Clin Immunol. 2000;106:46–52
- . S epsilon S mu and S epsilon S gamma switch circles in human nasal mucosa following ex vivo allergen challenge: evidence for direct as well as sequential class switch recombination. J Immunol. 2003;171:3816–3822
Editor's note: The feature, Images in allergy and immunology, is designed to highlight current concepts of the immunopathology of allergic diseases and other common immunopathologically mediated diseases. The presentation will appear as sets of images that involve cross-pathology, histopathology, and molecular pathology and will cover a range of topics of interest to allergists and immunologists.
PII: S0091-6749(05)00591-9
doi:10.1016/j.jaci.2005.03.019
© 2005 American Academy of Allergy, Asthma and Immunology. Published by Elsevier Inc. All rights reserved.
Volume 116, Issue 1 , Pages 233-236, July 2005
