Analysis of the human VH gene repertoire. Differential effects of selection and somatic hypermutation on human peripheral CD5(+)/IgM+ and CD5(-)/IgM+ B cells.

Abstract

To analyze the immunoglobulin repertoire of human IgM ϩ B cells and the CD5ϩ and CD5 Ϫ subsets, individual CD19 ϩ / IgM ϩ /CD5 ϩ or CD5 Ϫ B cells were sorted and non-productive as well as productive V H gene rearrangements were amplified from genomic DNA and sequenced. In both subsets, the V H 3 family was overrepresented largely as a result of preferential usage of a small number of specific individual family members. In the CD5 ϩ B cell subset, all other V H families were found at a frequency expected from random usage, whereas in the CD5 Ϫ population, V H 4 appeared to be overrepresented in the nonproductive repertoire, and also negatively selected since it was found significantly less often in the productive compared to the nonproductive repertoire; the V H 1 family was significantly diminished in the productive rearrangements of CD5 Ϫ B cells. 3-23/DP-47 was the most frequently used V H gene segment and was found significantly more often than expected from random usage in productive rearrangements of both CD5

Heterogeneity of Tonsillar Subepithelial B Lymphocytes, the Splenic Marginal Zone Equivalents

“…Unfortunately, it is impossible at present to determine the reciprocal contribution to the mutated memory cell pool by these two subsets, since clonally related sequences were not found across the different subsets. This is similar to the observations of other groups (20,40,41).…”

“…Mutated SE B cells may move again from the GCs to the SE area or migrate to the peripheral blood or to the splenic MZ or to other equivalent sites. Indeed, a minority of IgM-only cells sharing some of the phenotypic features of SE B cells have been detected in the circulation (41,49,50). Interestingly, V H and V L gene analyses have determined that these cells are almost invariably mutated, suggesting that primarily memory cells tend to recirculate.…”

Disturbed Peripheral B Lymphocyte Homeostasis in Systemic Lupus Erythematosus

“…14 and 20). In peripheral blood as well as in the bone marrow, memory B cells have been identified in populations of B cells expressing either class-switched Ig isotypes: IgM and IgD, or IgM only (21)(22)(23)(24)(25). More recently, IgD ϩ /CD27 ϩ B cells have been identified as having somatically mutated Ig genes and, therefore, being memory B cells (25).…”

The VλJλ Repertoire in Human Fetal Spleen: Evidence for Positive Selection and Extensive Receptor Editing

“…The cells were then stained with a PE-labeled anti-CD19 mAb (Becton Dickinson, Mountain View, CA) and a FITC-labeled anti-human IgM mAb (Caltag, Burlingame, CA). The CD19 ϩ /IgM ϩ B cells were sorted using a FACStar Plus flow cytometer (Becton Dickinson, San Jose, CA) outfitted with an automated single cell deposition unit, and one cell was deposited into each well of a 96-well PCR plate assembled on a microAmp base, as described previously (24,25). Each well contained 10 l of PCR buffer (50 mMKCl, 10 mM Tris-HCl, pH 9, 0.1% Triton X-100).…”