Molecular analysis of IgD-positive human germinal centres

Abstract

It is controversially discussed whether human IgM+IgD+CD27+ B cells, which carry somatically mutated Ig variable region (IgV) genes, are derived from germinal centres (GC) B cells or originate from another developmental pathway. GC composed of IgM+IgD+ B cells, which co-express the CD70 surface marker, have been described in ~10% of tonsils. As IgM+IgD+CD27+ B cells might be generated in such GC, we characterized IgD+ tonsillar GC cells. GC dominated by IgD+ B cells were present in 10 of 67 tonsils analyzed. Three GC were additionally positive for CD70. Detailed analysis of one such GC by microdissection and single-cell DNA PCR revealed IgD+ GC B cells undergoing somatic hypermutation during clonal expansion. However, further analysis of this GC as well as five additional microdissected GC by reverse transcription (RT)-PCR for clonally related Igm and Igd transcripts indicated that the B-cell clones in five of these six IgD+ GC belong to the IgD-only B cell subset, which has deleted the Cm gene, and that only one GC harboured a large IgM+IgD+ B-cell clone. Hence, most IgD+ GC consist of IgD-only B cells and fully developed IgM+IgD+(CD70+) GC are very rare. This indicates that the rare IgM+IgD+ GC B-cell clones from IgD+ GC contribute little to the large population of IgM+IgD+CD27+ B cells. Finally, an RT-PCR analysis with clone-specific primers for two IgD+ GC B-cell clones showed an absence of IgG or IgA class-switched clone members, indicating strict regulation of class switching and a selective production of IgD+ B cells from such clones. © The Japanese Society for Immunology. 2010. All rights reserved. For permissions, please e-mail: journals.permissions@oxfordjournals.org.