Is There a Typical Germinal Center? A Large-Scale Immunohistological Study on the Cellular Composition of Germinal Centers during the Hapten-Carrier–Driven Primary Immune Response in Mice

Abstract

Germinal centers (GCs) are complex, multicell-type, transient structures that form in secondary lymphatic tissues in response to T cell-dependent stimulation. This process is crucial to the adaptive immune response because it is the source of affinity maturation and long-lived B cell memory. Our previous studies showed that the growth of murine splenic GCs is nonsynchronized, involving broad-volume distributions of individual GCs at any time. This raises the question whether such a thing as a typical GC exists. To address this matter, we acquired large-scale confocal data on GCs throughout the course of the 2-phenyl-5-oxazolone chicken serum albumin-driven primary immune response in BALB/c mice. Semiautomated image analysis of 3457 GC sections revealed that, although there is no typical GC in terms of size, GCs have a typical cellular composition in that the cell ratios of resident T cells, macrophages, proliferating cells, and apoptotic nuclei are maintained during the established phase of the response. Moreover, our data provide evidence that the dark zone (DZ) and light zone (LZ) compartments of GCs are about the same size and led us to estimate that the minimal cell loss rate in GCs is 3% per hour. Furthermore, we found that the population of GC macrophages is larger and more heterogeneous than previously thought, and that despite enrichment of T cells in the LZ, the DZ of murine splenic GCs is not poor in T cells. DZ and LZ differ in the T cell-to-macrophage ratio rather than in the density of T cells.

Essential role of immobilized chemokine CXCL12 in the regulation of the humoral immune response

“…This observation cannot be explained by sectioning artifacts because, although some organized GCs could be sectioned entirely through the FDC-rich area such that they would appear as the disorganized GCs, the overall frequency of such occurrences is expected to be similar in control and mutant mice. Although, in control mice, we observed ∼24% of disorganized GCs, a fraction comparable with the previous largescale confocal imaging studies in immunized mice (30), a significantly higher proportion of disorganized GCs were observed in the spleens of CXC12 gagtm mice. The structure of these disorganized GCs resembled that observed in CXCR4 deficiency (11,25), suggesting that B-cell responsiveness to immobilized, but not free, CXCL12 contributes to efficient organization of GCs.…”

The evolution within us

“…The number of germinal centres that forms after infection is unknown but appears to be highly variable [79]. Highaffinity naive B cells enter each germinal centre and compete for antigens presented on the surface of follicular dendritic cells.…”

A Major Hindrance in Antibody Affinity Maturation Investigation: We Never Succeeded in Falsifying the Hypothesis of Single-Step Selection

“…These simulations show that the tree shapes vary most on the initial clone affinity and the selection threshold, and much less in dependence on the rates of GC B cell recycling (22), not allowing for a unique mapping from tree shapes to selection mechanisms – likely because the investigated trees were small. In addition to global measures not always being helpful in pointing to mechanisms at the micro-evolutionary scale (15, 23, 24), simulation of global measures like peak total GC B cell numbers did not lead to results that contradict the single-step hypothesis (22). …”