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Guiding immune cells to the center
Germinal centers (GCs) are the site of antibody affinity maturation. The GC response fundamentally depends on contact-dependent signal exchange between antigen-specific T and B lymphocytes. Lu et al. uncovered a repulsive guidance system that inhibits GC recruitment and retention of T follicular helper (TFH) cells while simultaneously promoting their helper activities locally (see the Perspective by Moschovakis and Forster). This system comprises the GC-specific transmembrane ephrin B1 (EFNB1) molecule and two EFNB1 receptors—EPHB4 and EPHB6—expressed by activated T cells, including TFH cells. In the absence of EFNB1 on GC B cells or when EPHB6 was suppressed on T cells, inappropriately large numbers of T cells were recruited to and retained in the GC as a result of relaxed repulsion of TFH cells.
Development of long-term antibody-mediated immunity relies on the germinal center (GC) reaction, in which CD4+ T cells help B cells to proliferate and differentiate into long-lived antibody-producing plasma cells or memory B cells. The GC reaction is orchestrated in a specialized microenvironment and critically depends on guided migratory dynamics of T and B cells. Follicular T helper (TFH) cells, a subset of effector CD4+ helper T cells specialized in supporting the GC reaction, deliver contact-dependent help signals, such as the CD40 ligand, and cytokines, such as interleukin-21 (IL-21), to promote GC B cell proliferation and differentiation. However, exaggerated TFH accumulation and function can cause abnormal GC responses and potentiate autoantibody-mediated inflammatory diseases or even lymphomas. Understanding of how TFH cells are recruited, retained, and functionally regulated in the GC microenvironment is needed for mechanistic insights into long-lived antibody-mediated immunity. Such understanding will inform better design of antibody-based vaccines for microbial infections and therapies for autoimmune diseases.
Local mechanisms that regulate TFH cell dynamics and helper functions in GCs are not well defined. Multiple soluble guidance cues sensed by heterotrimeric guanine nucleotide–binding protein (G protein)–coupled receptors promote TFH confinement toward the center of the B cell follicle of the secondary lymphoid organ, where the GC is typically formed. Contact-dependent guidance mechanisms are also thought to help regulate recruitment and retention of TFH cells in GCs. To search for such mechanisms, we examined the erythropoietin-producing hepatocellular (EPH) receptors and EPH-interacting protein (ephrin, EFN) family members. These molecules are known to guide cell migration during tissue morphogenesis in a contact-dependent manner.
We found that ephrin-B1 (EFNB1), a class B ephrin, was highly expressed by GC B cells and demarcated the GC tissue domain boundary. Although EFNB1 was not required for initial GC formation or development of TFH cells, its ablation from B cells led to exaggerated local accumulation of TFH cells inside the GC. Dynamic imaging by intravital microscopy revealed that EFNB1 repulsively inhibited GC recruitment and retention of TFH cells, likely as a result of EFNB1-mediated suppression of antigen-specific GC B cell–TFH adhesion. This contact-dependent repulsion required forward signaling through TFH-expressed EPHB6 but not EPHB4 receptor. Unexpectedly, despite the exaggerated GC TFH accumulation, the magnitude of GC responses was largely unchanged, whereas development of plasma cells and affinities of antigen-specific antibodies were actually reduced. By contrast, in a GC mixed with EFNB1-sufficient and -deficient B cells, the latter cells had a competitive advantage in contributing to the plasma cell compartment. These apparently contradicting phenotypes revealed that, by EPHB4-dependent forward signaling, EFNB1 also promotes GC TFH production of IL-21, cytokine that is needed to drive plasma cell formation. Consequently, at the tissue level, EFNB1-deficient GCs lack sufficient IL-21 to support normal plasma cell formation and affinity maturation; in a competitive GC reaction where the same cohort of TFH cells were shared by EFNB1-deficient and -sufficient GC B cells, the role of EFNB1 in suppressing antigen-specific interactions between individual TFH and GC B cells dominates, affording the EFNB1-deficient GC B cells with more contact-dependent help and a competitive advantage.
By revealing a dual role for the GC-expressed EFNB1 molecule, our results uncover a regulatory system that controls GC TFH dynamics and function in the local tissue microenvironment. EFNB1 suppresses T-B adhesion and repulsively inhibits dynamic TFH recruitment and retention in the GC; yet, it also positively promotes IL-21 production by TFH cells as they dwell in the GC at any moment. This counterintuitive combination of negative regulation of residence but positive promotion of effector functions controlled by the same EFNB1 molecule likely helps to ensure that the potentially dangerous GC reaction is productive yet self-limiting. That helper T cell cytokine production is critically modulated in a tissue microenvironment–specific manner by dedicated receptor-ligand systems, the EFNB1-EPHB4 pair in the case of GCs, suggests strategies and targets of manipulation for the development of improved vaccines or therapies.
Follicular T helper (TFH) cells orchestrate the germinal center (GC) reaction locally. Local mechanisms regulating their dynamics and helper functions are not well defined. Here we found that GC-expressed ephrin B1 (EFNB1) repulsively inhibited T cell to B cell adhesion and GC TFH retention by signaling through TFH-expressed EPHB6 receptor. At the same time, EFNB1 promoted interleukin-21 production from GC TFH cells by signaling predominantly through EPHB4. Consequently, EFNB1-null GCs were associated with defective production of plasma cells despite harboring excessive TFH cells. In a competitive GC reaction, EFNB1-deficient B cells more efficiently interacted with TFH cells and produced more bone-marrow plasma cells, likely as a result of gaining more contact-dependent help. Our results reveal a contact-dependent repulsive guidance system that controls GC TFH dynamics and effector functions locally.