Analysis of immunoglobulin E VH transcripts in a bronchial biopsy of an asthmatic patient confirms bias towards VH5, and indicates local clonal expansion, somatic mutation and isotype switch events - PubMed (original) (raw)

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Analysis of immunoglobulin E VH transcripts in a bronchial biopsy of an asthmatic patient confirms bias towards VH5, and indicates local clonal expansion, somatic mutation and isotype switch events

R E Snow et al. Immunology. 1999 Dec.

Abstract

Immunoglobulin E (IgE)-dependent mechanisms play a pivotal role in mediating allergic disease. Previously, VH-Cepsilon transcripts from blood or spleen of atopic asthmatics have been analysed for VH gene usage and patterns of somatic mutation. An over-representation of the minor VH5 family has been observed, consistent with a superantigen drive. As local mucosal events in IgE production may be more significant in the disease process, we have analysed VH-Cepsilon transcripts from a bronchial biopsy of a patient with severe asthma. VH5 predominance was confirmed with 10 of 30 unique clones derived from this family. Repeated sequences, some with intraclonal variation, revealed clonal expansion and continuing mutational activity at the site. Unexpectedly, three unmutated VH-Cepsilon sequences were found, indicating that isotype switching to IgE can occur without mutation. Detection of a sister clone with extensive mutations was again consistent with local mutational activity. Evidence for local isotype switching was obtained by identification of clonally related immunoglobulin M (IgM), immunoglobulin G (IgG) and immunoglobulin E (IgE) sequences. However, in contrast to findings in blood, no IgG4 transcripts clonally related to IgE were detected, suggesting that the balance between synthesis of IgG4 and IgE may differ between systemic and local sites. These data confirm a VH5 bias in IgE, and support the concept that IgE-synthesizing B cells arise via local differentiation.

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Figures

Figure 1

Figure 1

Graph showing a significant over‐representation of the VH5 family by immunoglobulin E (IgE) in the bronchial biopsy when compared with the germline repertoire, a polymerasec chain reaction (PCR) control for immunoglobulin M (IgM) and the productive repertoire from normal B cells.

Figure 2

Figure 2

Immunoglobulin M (IgM)‐, immunoglobulin G (IgG)‐ and immunoglobulin E (IgE)‐related clones with a common CDR3/FWR4 sequence. The nucleotide sequences of CDR3/FWR4/CH transcripts are aligned to D‐segment and JH germline genes. Primer sequences are underlined, the Cε primer is 3′ to the sequence shown.

Figure 3

Figure 3

VH5‐immunoglobulin E (IgE) amino acid sequences from bronchial biopsy aligned to closest germline genes. Clones 1a, b, and 2a, b, c are sets of related clones. Identical repeated sequences were found for clones 2b, 4 and 10, with number of repeats shown in brackets. Upper case: replacement mutation; lower case: silent substitution. *Indicates sites that have previously been reported as hot‐spots of mutation. ↓Indicates frequently mutated sites not previously reported.

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