T Follicular Helper Cell Overview

T follicular helper (Tfh) cells are a subset of CD4-positive T helper cells that are involved in the humoral response [1]. Tfh cells are found in secondary lymphoid tissues, including tonsils, spleen, and lymph nodes. The role of Tfh cells is to trigger germinal center (GC) B cells into antibody-secreting plasma and memory B cells [2,3]. Therefore, Tfh cells are critical for mounting immune responses against pathogens, and understanding their function is important for vaccine development.

Germinal centers (GCs) are microanatomical structures formed within secondary lymphoid organs after infection or immunization. GCs are where B cells proliferate, undergo diversification and increase their affinity to antigens. The GC is divided into two compartments—the dark zone (DZ) and the light zone (LZ)—with different functions and cell types (Table 1) that influence the role of GC B cells [4]. The DZ is occupied with highly proliferative B cells that hypermutate in order to produce diverse antibodies. The LZ is more diverse, with follicular dendritic cells (FDCs), Tfh cells, macrophages, and both naive and GC B cells. LZ cells work in concert to select B cells that (1) will exit the GC as a memory B cell or plasma cell, (2) are removed by macrophages, or (3) will be directed back into the DZ to go through the hypermutation process again [5].

Table 1. Nonexhaustive list of cells found in the germinal centers.

Tfh cells are CD4-positive cells that express high levels of CXCR5 protein and transcription factor B cell lymphoma 6 (BCL6) [6]. Other important markers include CXCR3, PD1, and ICOS. Tfh cells produce IL-21 to drive B cell differentiation and affinity maturation. Markers identifying GC Tfh cells are conserved across humans and mice (Table 2) [1].

Table 2. Nonexhaustive list of human and mouse Tfh cell markers.

Circulating Tfh (cTfh) cells are being investigated as surrogates for GC Tfh cells in order to understand vaccine response in vitro [7]. GC Tfh may not authentically differentiate in vitro and requires dissection of the GC region from secondary lymphoid tissues in order to study T cell–B cell interactions. It has been observed that co-culturing cTfh cells with autologous naive B cells generated plasmablasts [8,9]. cTfh cells can be identified with the following markers: CD4, CXCR5 with PD1, CCR7, CXCR3, CCR6, and ICOS [8,9,10]; cTfh cells do not express BCL6.

T helper cell subset differentiation is dependent on polarizing CD4⁺ T cells with specific cytokines. Robust routes for differentiation have been recognized for type 1 and type 2 Th (Th1 and Th2) cells, Th17 cells, and regulatory T (Treg) cells. Tfh cell differentiation requires multiple factors and not one single event [1]. It is accepted that Tfh differentiation requires TCR stimulation of naive CD4⁺ T cells with MHC II-specific peptides with the presence of both IL-6 and IL-21 [11, 12, 13].
 

Cytokines play an important role in Tfh cell differentiation, although there are reported differences between mice and humans regarding which cytokines play a key role. IL-6, for example, is a potent inducer of murine Tfh cell differentiation along with the costimulatory molecule ICOSL, whereas the role of IL-6 in human Tfh differentiation is far less defined. In contrast, human Tfh cell differentiation can be accomplished in vitro via coculture using a combination of IL-12 with TGF-β or activin A, conditions that do not influence murine Tfh cell differentiation. Among inhibitory cytokines, IL-2 is the most potent inhibitor of Tfh differentiation.

Key cytokines secreted by Tfh cells include IL-21 and IL-4 which in combination with the co-stimulatory molecule CD40L provide the signals that are required for B cell proliferation and differentiation. IL-21 secretion precedes the expression of IL-4, which is only acquired upon full differentiation of Tfh cells in the germinal centers. In addition to these cytokines, human Tfh cells also secrete the chemokine CXCL13 which is responsible for the recruitment of CXCR5⁺ B cells to follicles.

Table 3: Key cytokines involved in Tfh differentiation and secretion.

Profiling cytokines and chemokines involved in differentiation or those secreted by Tfh cells can be measured individually using ELISA or multiplex immunoassays such as the Invitrogen ProcartaPlex assays.