Marginal-Zone B Cells in the Human Lymph Node and Spleen Show Somatic Hypermutations and Display Clonal Expansion

Abstract

Splenic marginal-zone B cells, marginal-zone B cells of Peyer’s patches in the gut, and nodal marginal-zone B cells (also identified as monocytoid B cells) share a similar morphology and immunophenotype. These cells likely represent a distinct subset of B cells in humans and rodents, but their precise ontogenetic relationship as well as their origin from B cells of the germinal center is still debated. To study this, we performed a mutation analysis of the rearranged immunoglobulin variable genes (VH) of microdissected single nodal and splenic marginal-zone cells. In addition, we investigated the presence of proliferating cells and B-cell clones in the human splenic and nodal marginal zone as well as adjacent germinal centers. This was performed by immunohistochemical staining for the Ki-67 antigen and denaturing gradient gel analysis of amplified immunoglobulin heavy chain genes’ complementarity determining region 3 of microdissected cell clusters. A variable subset of nodal and splenic marginal-zone B cells showed somatic mutations in their rearranged VH genes, indicating that both virgin and memory B cells are present in the nodal and splenic marginal zone. Nodal and splenic marginal-zone B cells preferentially rearranged VH3 family genes such as DP47, DP49, DP54, and DP58. A preferential rearrangement of the same VH genes has been shown by others in the peripheral CD5− IgM+ B cells. These data suggest that the splenic and nodal marginal-zone B cells are closely related B-cell subsets. We also showed that marginal-zone B cells may cycle and that clones of B cells are frequently detected in the nodal as well as the splenic marginal zone. These clones are not related to those present in adjacent germinal centers. These data favor the hypothesis that clonal expansion occurs in the marginal zone. Whether the somatic hypermutation mechanism is activated during the clonal expansion in the marginal zone and which type of immune response triggers the clonal expansion need to be elucidated.

Persistent polyclonal B lymphocytosis: an expansion of cells showing IgVH gene mutations and phenotypic features of normal lymphocytes from the CD27+ marginal zone B‐cell compartment

“…The overall frequency of mutations per 100 bp was 2AE3% ± 0AE3, which is clearly higher than the figures observed for resting naive B lymphocytes (Brezinschek et al, 1995;Spencer et al, 1998;Tangye et al, 1998). The mutation frequencies detected in PPBL were in the range of those described for normal lymphocytes from the CD27 + MZ B-cell compartment (1AE6-5AE6) (Dunn-Walters et al, 1995;Paramithiotis & Cooper, 1997;Klein et al, 1998;Spencer et al, 1998;Tangye et al, 1998;Tierens et al, 1999;Dono et al, 2000). It is well established that Ag-selected IgVH mutations occurring in the germinal centres lead to the accumulation of replacement mutations in the complimentarity-determining regions (CDRs) while this kind of mutation are suppressed in the framework regions (FRs) (Chang & Casali, 1994).…”

“…Recent evidence demonstrates that B lymphocytes from normal MZ and MZ-like areas have already triggered Ig-V gene somatic mutations events (Dunn-Walters et al, 1995;Paramithiotis & Cooper, 1997;Klein et al, 1998;Tangye et al, 1998;Tierens et al, 1999;Dono et al, 2000) and, accordingly, these cells are now considered as a memory B-lymphocyte compartment. In consequence, the presence of Ig-V gene somatic mutations was examined in the B cells isolated from the three PPBL patients under study.…”

“…Nevertheless, a complete pattern of Ag-selected mutations (Chang & Casali, 1994) was only observed in a few clones (Table I). The significance of the mutational process occurring in the normal MZ-like B-cell compartment is not entirely understood, and the well-defined antigen-selected mutation distribution pattern (Chang & Casali, 1994) has not been consistently demonstrated (Dunn-Walters et al, 1995;Paramithiotis & Cooper, 1997;Klein et al, 1998;Spencer et al, 1998;Tangye et al, 1998;Tierens et al, 1999;Dono et al, 2000). Therefore, PPBL cells harbour mutational events similar to those described in the normal MZ B-cell compartment.…”

New Insights into the Cell Biology of the Marginal Zone of the Spleen

“…In addition to the presence of marginal zone B cells with their distinct features as outlined in the previous sections, especially in humans and rats, there is ample evidence for the existence of B memory cells in the marginal zone. These cells have been generated as a result of classical T-dependent responses, involving germinal center reactions, and showing somatic hypermutation and high aYnity of their BCRs (Dunn-Walters et al, 1995;Liu et al, 1991;Shih et al, 2002;Tierens et al, 1999). They may react swiftly to antigen, and as such the marginal zone can form a reservoir of memory B cells.…”

Heterogeneity of stromal cells in the human splenic white pulp. Fibroblastic reticulum cells, follicular dendritic cells and a third superficial stromal cell type

“…23 In addition, MZ B cells are regarded as pre-activated cells that are easily stimulated to differentiate into IgM-secreting plasma cells. 12,24,25 Human MZ B cells therefore function in T-cell-independent immune reactions, but may also be recruited into Tcell-dependent antibody production involving somatic hypermutation and immunoglobulin isotype switching. 12,23 It is still controversial, whether human MZ B cells represent a unique B-cell lineage with an a priori pre-diversified repertoire existing independent of germinal centre formation 24,[26][27][28][29] or whether they are a special population of post-germinal centre cells.…”