A. Bosseloir

Follicular Dendritic Cells: Origin and Function

The human immune system is made up of one trillion lymphocytes. Although dispersed throughout the organism, these cells behave as if they belonged to a single organ. To attain this homeostasis, the lymphoid cells home to and are controlled in lymphoid tissues where, in specific microenvironments, they communicate with each other or with accessory cells and proliferate, mature, or die under stringently controlled conditions. The germinal center microenvironments are among those controlling the B cells which, during the T-dependent humoral immune responses, undergo important phases of their life cycles: activation, proliferation, the isotype switch, affinity maturation, deactivation, apoptosis, etc. The follicular dendritic cells (FDC) are major components of the germinal center microenvironments. Here, we examine their origin and their influence on B cells in the light of recent experimental data.

Human follicular dendritic cells in vitro and follicular dendritic-cell-like cells.

Abstract.Human follicular dendritic cell (FDC)-like cells (FLC) have been utilized for the in vitro analysis of germinal center reactions. However, there is no consensus whether FLC represent FDC in vitro. The purpose of the present study has therefore been to determine distinguishing features of FDC and FLC in vitro. The expression of CD40, CD54, CD49d, cytokine (γ-IFN and IL-4)-dependent MHC-class II, and CD106 was observed to be specific for the determination of FDC in long-term culture. The cytokine-dependent emperipolesis of germinal center B cells was establised as another discriminating property for FDC in vitro. In 2 out of 72 long-term cultures of FDC, we encountered dividing cells among the non-dividing population of FDC. The dividing cells expressed accessory molecules similar to those of FDC but showed emperipolesis only for the initial few days of their growth. FDC did not enhance the CD40-dependent proliferation of germinal center B cells; in contrast, FLC augumented it. Both types of cells produced a significant amount of cytokine-dependent IL-6. Further studies are needed to determine whether FLC originate from FDC in vitro.

Follicular dendritic cells: whose children?

At the recent Germinal Centre Conference in Spa*, over 200 immunologists gathered to study the fate of lymph follicles, B- and T-cell selection, cell migration, accessory cells, and pathological conditions such as lymphoproliferative diseases and AIDS. Here, Ernst Heinen and Alain Bosseloir report on this fascinating field.

Moabs MAS516 and 5B5, two fibroblast markers, recognize human follicular dendritic cells

Follicular dendritic cells (FDC) are only located within follicles of secondary lymphoid tissues. The origin of this peculiar cell type is not clearly defined. To contribute to this study, we applied two monoclonal antibodies (MAS516 and 5B5) considered as specific for fibroblasts to tonsil cryosections and to isolated follicular dendritic cells. On the basis of an enzyme cocktail digestion of human tonsils and a fractionation procedure on albumin gradients, FDC can be prepared in the form of cell aggregates with associated lymphoid cells. MAS516 reacts with surface membrane molecules expressed by human fibroblasts, tissue macrophages and peripheral blood monocytes. With immunoperoxidase assays on tonsil cryosections connective tissue cells and macrophages are stained. Inside germinal centres, heavy labelling of the light zone was found. The MAS516 staining pattern is very similar to that of specific FDC markers DRC-1 or BU10. All isolated FDC reacted with MAS516 antibody. 5B5, considered as a typical fibroblast marker, reacts with human prolyl-4-hydroxylase which is an intracellular enzyme related to collagen biosynthesis. In cryosections, interfollicular and capsular areas showed 5B5 positive connective tissue fibroblasts. In germinal centres, some cells presenting features of FDC were 5B5 positive. After cell separation, 25%-50% of the isolated FDC were labelled with this antibody. This positivity of some FDC for 5B5 antibody may support the idea of their fibroblastic origin. The combination of observations realized in situ and after cell purification ensured an unequivocal recognition and identification of FDC.(ABSTRACT TRUNCATED AT 250 WORDS)

Human germinal center CD4^+CD57^+ T cells act differently on B cells than do classical T-helper cells

We have isolated two subtypes of helper T cells from human tonsils: CD4+ CD57+ cells, mostly located in the germinal center (GC), and CD4+ CD57- cells, distributed through the interfollicular areas but also present in the GC. In a functional study, we have compared the capacities of these T-cell subtypes to stimulate B cells in cocultures. In order to block T-cell proliferation while maintaining their activation level, we pretreated isolated T cells with mitomycin C prior to culture in the presence of B cells and added polyclonal activators such as PHA and Con A, combined or not with IL-2. Contrary to CD4+ CD57- cells, CD4+ CD57+ cells did not markedly enhance B-cell proliferation. Even when sIgD-B cells typical of germinal center cells were tested, the CD4 CD57 cells had no significant effect. This is in accordance with the location of these cells: They mainly occupy the light zones of the GC where few B cells divide. Even when added to preactivated, actively proliferating cells, CD4+ CD57+ cells failed to modulate B-cell multiplication. On the supernatants of B-cell-T-cell cocultures, we examined by the ELISA technique the effect of T cells on Ig synthesis. Contrary to CD57- T cells, whose effect was strong, CD57+ T cells weakly stimulated Ig synthesis. More IgM than IgG was generally found. Because CD57 antigen is a typical marker of natural killer cells, we tested the cytolytic activity of tonsillar CD4+ CD57+ cells on K562 target cells. Unlike NK cells, neither CD4+CD57+ nor CD4+ CD57- cells exhibit any cytotoxicity. Thus, germinal center CD4+ CD57+ cells are not cytolytic and do not strongly stimulate either B-cell proliferation or Ig secretion. CD4+ CD57- cells, however, enhance B-cell proliferation and differentiation, thus acting like the classical helper cells of the T-dependent areas.

THE INFLUENCE OF FOLLICULAR DENDRITIC CELLS ON B-CELL PROLIFERATION DEPENDS ON THE ACTIVATION OF B CELLS AND THE MITOGEN USED

Follicular dendritic cells (FDC) are unique non‐lymphoid cells found only in lymph follicles. They play a part in the survival, proliferation and differentiation of B cells. To analyse the influence of FDC on B‐lymphocyte proliferation, we isolatedthem from human tonsils on albumin gradients and treated them with mitomycin C to prevent the multiplication of lymphoid cells harboured in their cytoplasmic evaginations. FDC cultured for 12–16 h remained attached to the substrate;non‐adherent cells were carefully eliminated by washing. Purified B cells cultured alone or with contaminant‐cleared FDC were maintained for 2 days in the presence or absence of various stimulants, after which tritiated thymidine uptake by these cells was measured. In the absence of activators, FDC did not induce B‐cell multiplication. B cells cultured in the presence of FDC exhibited increased 3H‐TdR uptake when activated with anti‐CD40 MoAb, anti‐immunoglobulin MoAb or transferrin, but notwhen stimulated with Staphylococcus aureus strain Cowan I (SAC) at a given concentration. In the latter case, B‐cell proliferation clearly decreased. In control cocultures where mitomycin‐C‐treated non‐adherent cells were used instead of FDC inthe presence of the different stimulants, no increase in B‐cell proliferation was observed. The results suggest that, inside the germinal centres, FDC modulation of B‐cell proliferation depends on the activation state of the B cells and on the stimulantencountered.

Follicular Dendritic Cells: Isolation Procedures, Short and Long Term Cultures

The need to isolate and cultivate follicular dentritic cells (FDC) in vitro is becoming increasingly acute in view of the physiological complexity of the germinal centers and the accumulation of data on their pathology.

Germinal Center T Cells: Analysis of Their Proliferative Capacity

Several immunohistochemical studies have revealed the existence of T cells expressing the CD57 antigen in the germinal center1. A few are also found in the interfollicular zones and the mantle zone2. Phenotypically they are CD4+, CD8- cells. They do not express Leu8, CD16 or CDllb3,4,5. These cells are not fully activated being CD25-,CD71- and HLA-DR- cells6.

Modulation of B lymphocyte proliferation inside the germinal center.

Germinal centers of stimulated lymphoid follicules comprise different microenvironments where B cells proliferate or differentiate into memory B cells or precursors of Ig-producing cells [1]. Follicular dendritic cells (FDC), unique non lymphoid cells, mainly compose these microenvironments. FDC are closely associated with B cells and can be distinguished from other accessory cells in secondary lymphoid tissues by a number of features, as lack of phagocytic activity and a characteristic set of cell surface markers.

Expression and function of DRC-1 antigen.

Immune responses are generated by a series of cellular interactions which occur in distinct microenvironments. The germinal centre (GC) plays a central role in B cell proliferation and differentiation as well as in generating the secondary immune response. The architecture of the GC is designed to support cell-to-cell interactions1,2.