T-lymphocytes (T cells)
Ontogeny
The process of development and maturation of the T Cells in mammals begins with the haematopoietic stem cells (HSC) in the fetal liver and later in the bone marrow where HSC differentiate into multipotent progenitors. A subset of multipotent progenitors initiates the transcription of recombination activating gene 1 and 2 (RAG 1 and RAG2) and become lymphoid-primed multipotent progenitors and then common lymphoid progenitors (CLP). Only a small subset of pluripotent cells migrates to the thymus and differentiates into early thymic progenitors (ETP). The thymus does not contain self-renewing progenitors; and therefore, long-term thymopoiesis depends on the recruitment of thymus-settling progenitors throughout the life of the individual (1). These progenitors must enter the thymus to become gradually reprogrammed into fully mature and functional T Cells. The T Cell’s distinct developmental steps, as illustrated in Figure 1, are coordinated with the migration of the developing thymocytes towards specific niches in the thymus that provide the necessary stage-specific factors that are needed for further differentiation.
Figure 1
Overview of T Cell development and maturation. Adapted from Rothenberg et al. (4). Abbreviations. HSC: Haematopoietic stem cells, CLP: Common lymphoid progenitors, ETP: Early thymic progenitors, DN: Double negative; DP: Double positive, SP: Single positive, (more...)
The ETP are multipotent and can generate T Cells, B Cells, Natural killer cells (NK), myeloid cells, and dendritic cells (DC). ETP represent a small and heterogenous subset, have the ability to proliferate massively, and can be identified by the phenotype Linlow, CD25−, Kithigh as well as by their expression of Flt3, CD24, and CCR9 (1). These cells, which are attracted by the chemokines CCL19 and CCL21, enter the thymus via the corticomedullar junction. In the stroma of the thymus, the ETP encounter a large number of ligands for the Notch receptors as well as growth factors such as Kit-ligand and IL-7 which trigger and support the differentiation and proliferation of these cells in the initial stages of T Cell development (2). Moreover, the expression of Notch-1 receptors and their interaction with Delta-like ligands is essential for the differentiation of the T Cells in the thymus and for the inhibition of the non-T Cell lineage development (3).
Within the thymic cortex, ETP differentiate into double negative (DN) cells that do not express either CD4 or CD8 (i.e., CD4− and CD8−). Some authors consider the ETP a DN1 cell that later differentiates into DN2 when it acquires the CD25+ and CD44+ receptors. At this stage of development, the cells lose the B potential and begin to express proteins that are critical for the subsequent T Cell receptor (TCR) gene rearrangement such as RAG1 and RAG2. They also begin to express proteins necessary for TCR assembly and signaling as CD3 chains, kinases, and phosphatases such as LCK, ZAP70, and LAT (4). DN3 cells can take two divergent routes of differentiation. A cell can either express the αβ chains of the TCR and follow the process of selection to generate CD4+ or CD8+ T Cells or express the γδ chains to generate a subpopulation of γδ lymphocytes with special functional characteristics (5,6) (Table 1).
Table 1
Characteristics of αβ T cells and γδ T cells.
The expression of the β chain of TCR, at the DN3 stage, cascades the simultaneous expression of the CD4 and CD8 molecules and thus, the cells convert into double positives (DP), which constitutes the largest population of cells in the thymus (4,7). At this stage of maturation, the DP cells enter a control point known as positive selection to select the cells with functional TCRs that bind to self-peptides with intermediate affinity and avidity. For this, the epithelial cells of the thymic cortex “put the DP cells to the test” by presenting their own peptides in the context of the class I (HLA-I) and class II (HLA-II) HLA molecules. Only a fraction (1%-5%) of the DP cells, that express a TCR with intermediate affinity for these Ags persists by survival signals. DP cells incapable of binding HLA-I or HLA-II undergo apoptosis. Positive selection allows the differentiation of the DP thymocytes towards a single positive (SP) population that is restricted to HLA (i.e., DP cells that recognize HLA-I differentiate into CD4−CD8+ and those that recognize HLA-II differentiate into CD4+CD8−) (8, 9). Subsequently, SP cells enter the medulla of the thymus where a second control point known as negative selection takes place. At the medulla, positively selected thymocytes are exposed to a diverse set of self-antigens presented by medullary thymic epithelial cells (mTEC) and DC. mTECs use a special epigenetic mechanism to give rise to what is often referred to as promiscuous gene expression which contributes to the low expression of many genes including tissue-restricted self-antigens. SP cells with a high affinity or avidity for binding self peptides presented on HLA-I or HLA-II are eliminated by apoptosis, thus assuring the destruction of potentially autoreactive cells (9). Cells that survive negative selection mature and become naïve T Cells given the fact that they have not been primed by Ag for which they express a specific TCR. Naïve T Cells leave the thymus and migrate continuously to the secondary lymphoid organs to be primed and differentiate into effector cells with specialized phenotypes.
T cell receptor (TCR) complex
During the maturation process, T Cells acquire a receptor called TCR that recognizes a specific Ag. TCR is a multiprotein complex composed of two variable antigen-binding chains, αβ or γδ, which are associated with invariant accessory proteins (CD3γε, CD3δε, and CD247 ζζ chains) that are required for initiating signaling when TCR binds to an Ag (10).
The αβ-TCR does not recognize Ag in its natural form but recognizes linear peptides which have been processed and presented in the HLA-I or HLA-II context. The peptides presented by HLA-I molecules are small (8–10 aminoacids) and have an intracellular origin while those presented by HLA-II molecules are longer (13–25 aminoacids) and are generally of extracellular origin. Nevertheless, the αβ-TCR of NKT cells and the γδ-TCR can recognize glycolipids and phospholipids presented by CD1 molecules.
TCR α and β chains are very polymorphic, which favors the recognition of a great diversity of peptides. Each chain has a variable (V) and a constant domain (C) with a joining segment (J) that lies between them. The β chain also has an additional diversity segment (D). Each (V) domain has three hypervariable sectors known as CDR-1, -2, and -3 (complementarity-determining regions) and is capable of generating an inmense pool of combinations to produce different TCR specific for an Ag. CDR3α and β regions bind to the central region of the peptide presented. This region represents the most diverse region of the TCR and is considered to be the main determinant of specificity in Ag recognition. CDR1α and β also contribute to peptide recognition and bind to it through the amino and carboxy-terminal motifs respectively. TCR regions that come into contact with HLA mainly correspond to CDR-1 and CDR-2 (10). TCR associates with a molecule called CD3, which is composed of three different chains: gamma, delta, and epsilon γδε. These chains are associated as heterodimers γε and δε. TCR is also associated with a hom*odimer of δε chains (CD247) that has a long intracytoplasmic portion and participates in the downstream transductional activation signals. Both the CD3 chains and the δε chains that associate with TCR possess tyrosine-based activation motifs (ITAMs) in their intracytoplasmic moeities, which are phosphorylated to initiate T Cell activation (11).
TCR gene rearrangement is essential during T Cell development. Multiple gene segments dispersed in the genomic DNA must bind and transcribe to produce a functional TCR. This process occurs independently for each chain beginning with the recombination of genes for the β chain (12). Genes that code for the TCR chains in humans map to four loci: TCRA and TCRD on chromosome 14 and TCRB and TCRG on chromosome 7. The locus for the β chain has 42 gene segments for the region (V), 2 for (D), 12 for (J), and 2 for (C) while the locus for the α chain has 43 gene segments for the region (V) and 58 for (J) (13) (Figure 2). Somatic recombination of these gene segments occurs at the DN2 and DN3 stages of T Cell development and is mediated by the gene products RAG-1 and RAG-2. Nuclease and ligase activity, as well as the addition or elimination of nucleotides, generates the great variety of TCR present in our organism at the moment of birth. It is estimated that the diversity of TCR in humans may reach 2×107 (13).
Figure 2
TCR generation by somatic recombination. Adapted from Turner et al. (13). Abbreviations. TCR: T Cell receptor; C: constant gene segment, V: variable gene segment, D: diversity gene segment, J: junctional gene segment, N: addition of non-template-encoded (more...)
Activation of the naïve T cells
T Cell activation and differentiation will only be sucessfull if three signals are present: i) interaction of the TCR with the peptide presented by the HLA molecule, ii) signaling through co-stimulatory molecules, and iii) participation of cytokines that initiate clonal expansion (14).
Additionally, the cytokine microenvironment that accompanies the activation defines the type of response that will be generated later.
Ag recognition and signal transduction pathways in T cells
Constant migration of the naïve T Cells towards the secondary lymphoid organs is essential in order for each one to encounter its specific Ag presented by an antigen-presenting cell (APC) (15). For this to occur, the naïve T Cells constitutively express L-selectin, an adhesion molecule which acts on the initial binding of T Cells to the high endothelial postcapillary venules located in the lymph nodes, tonsils, and aggregated lymphatic follicles. Only the specialized endothelial cells in the post-capillary veins allow constant passage of the T Cells from the blood towards the lymph nodes or Peyer’s patches given that the latter two constitutively express the addressins PNAd (peripheral node addressin) or MAdCAM-1 (mucosal addressin cell adhesion molecule-1) respectively. Both interact with the L-selectin of the lymphocytes. The endothelial cells of the rest of the vasculature restrict or impede binding of lymphocytes unless their receptors are induced by inflammation mediators (16).
In the lymph nodes, T Cells establish temporary contact with a great number of dendritic cells (DC) but only halt and bind to those which present an Ag which is compatible and specific to their receptor (15).
T Cells within lymph nodes migrate at high speeds of about 11–14 μ per minute. This is in contrast to DCs which transit through lymph nodes at speeds of about 3–6 μ per minute and then stop. This allows DCs to constantly establish new contacts with T Cells. In the absence of Ag, T Cells do not stop, but in the presence of an Ag, the duration of the interaction with the DC may be transitory (3 - 11 min) or stable (several hours) depending on the affinity for the Ag (15). Stable unions are favored by the high presence of peptides in the DC, highly antigenic ligands, mature DC, and expression of molecules such as ICAM-1 (15).
Antigen recognition by TCR induces the formation of several “TCR microclusters” that accompany the reorganization and approach of other membrane molecules and signaling proteins towards the contact zone with the DC. This contact zone between the T Cell and DC membranes is known as an immunological synapse and consists of a highly organized and dynamic molecular complex divided into three concentric zones known as the central, peripheral, and distal supramolecular activation clusters. The central region is composed of the TCR complex, co-stimulatory and co-inhibitory molecules, and co-receptors. These co-receptors are known as primary and secondary activation signals. The peripheral zone is mainly made up of the adhesion molecules LFA-1-ICAM-1 and CD2-LFA-3 that, due to their affinity, maintain and stabilize binding between the cells. The distal zone consists of F-actin and phosphatase CD45 (17).
After Ag recognition, a complex signaling process is initiated on the internal side of the membrane and in the cytoplasm for the subsequent activation of three essential transcription factors: NFAT, AP-1, and NF-κB. These signaling pathways are shown in a simplified diagram in Figure 3 and start when phosphatase CD45 activates the tyrosine-kinases, Fyn and Lck, which are associated with the ε chains of the CD3 and the co-receptors CD4/CD8 respectively (18). Once activated, these kinases autophosphorylate and phosphorylate the ITAM moieties of the δε chains and CD3. Phosphorylated ITAMs attract the ZAP-70 molecule. Then, the binding of ZAP-70 to phospholipase C γ1 (PLC γ1) or LAT initiates two different cascades.
Figure 3
Overview of TCR signalling pathways. Adapted from Brownlie et al. (18). Abbreviations. LCK: lymphocyte-specific protein tyrosine kinase, FYN: a member of Src tyrosine kinases, ZAP70: ζ-chain associated protein kinase of 70 kDa, LAT: Linker for (more...)
A first cascade is initiated when PLC- γ1 converts the phosphatidylinositol biphosphate (PIP2) into inositol triphosphate (IP3) and diacylglycerol (DAG). The IP3 diffuses into the cytoplasm and binds to the receptors of the endoplasmic reticulum, where it induces the release of Ca2+ deposits to the cytosol. The intracellular increase of Ca2+ stimulates the enzyme calmodulin, which is a serine/threonine-kinase. The activated calmodulin, in turn, activates calcineurin, a phosphatase that catalyzes the desphosphorylation of the nuclear transcription factor NF-AT to allow entry into the nucleus and activate the expression of several genes (e.g., IL-2, etc.) (19).
A second signaling cascade is initiated when ZAP-70 phosphorylates an adaptor protein known as LAT. LAT recruits several proteins that allow transference of guanine nucleotides from GDP to GTP for the activation of some proteins called Ras. These initiate a cascade of phosphorylations resulting in the activation of mitogen activated protein kinases (MAPK). These MAPK are tasked with activating the transcription factor AP-1 which is composed of the proteins c-fos and c-jun. MAPKs allow dimerization of those proteins to initiate the transcription of genes (18).
The third signaling pathway is initiated with the production of DAG, which activates protein kinase C (PKC). Later, it gives rise to recruitment of the IKK complex which requires the proteins Carma1, Bcl10, and MALT1 for its activation. Activation of the IKK kinases permits the phosphorylation of the IκB inhibitors, which are then ubiquitinated and degraded. This releases NF-κB dimers that translocate to the nucleus and activate transcription of their target genes (20).
The transcription factors NF-AT, AP-1, and NF-κB enter the nucleus and induce the transcription of genes to initiate the secretion of IL-2; the expression of its high affinity alpha receptor (IL-2Rα); the expression of integrins that promote cellular adhesion; the expression of costimulatory molecules such as CD40L; and the expression of anti-apoptotic proteins (19, 20).
Co-stimulation
A co-stimulatory molecule is defined as a surface molecule that is not itself able to activate T Cells but which can significantly amplify or reduce the signaling induced by the TCR complex (21,22). The main T Cell co-stimulatory molecules and their respective ligands for the profesional and non-professional APC are shown in Table 2.
Table 2
T Cell co-stimulatory molecules and their ligands.
Positive co-stimulatory signals are known as the second activation signal and are indispensable for potentiating the production of IL-2 due to the induction of a sustained activation of the nuclear transcription factor NF-κB. Furthermore, interaction between these molecules initiates antiapoptotic signals that prolong T Cell life span and initiate the expression of adherence molecules as well as the production of growth factors and cytokines that promote their proliferation and differentiation.
Only CD28, CD27, and HVEM are expressed constitutively while the remaining co-stimulatory molecules are inducible and expressed only after activation. Constitutive co-stimulatory molecules have a positive regulatory effect (21, 22).
Although most of the co-stimulatory molecules have a monotypic binding (one ligand), some of them, e.g., CD28 and PD-1, interact with more than one ligand. Moreover, other molecules such as those of the SLAM family interact hom*otypically with identical molecules. Almost all of the T Cell co-stimulatory molecules belong to the CD28/B7 superfamily or the TNF/TNFR family (21, 22).
There is a hierarchy in the downstream activation of these co-stimulatory molecules. For example, it has been observed that co-stimulation with CD28 significantly increases the induction of ICOS and OX-40 on the surface of the T Cell (22).
Clonal expansion
In response to antigen recognition and co-stimulatory signals, T Cells initiate the synthesis of IL-2 and express the high affinity receptor for it (IL2Rα or CD25) transitorily. CD25 binds to the other chains of the IL2R which are the β chain (CD122) and common γ chain (CD132). However, it does not participate in the signaling, but increases the affinity for IL-2 from 10 to 100 times (23).
IL-2 acts as an autocrine and paracrine growth factor. IL-2 activates blastogenesis or clonal expansion which gives rise to large numbers of T Cells with receptors identical to the original, able to recognize only the Ag that initiated its activation. IL-15 and IL-21 also participate in this process of clonal expansion (23).
T Cell activation and clonal expansion is followed by a death phase during which 90% of the effector cells are eliminated by apoptosis. The mechanisms which induce this phase of contraction or death include interactions Fas-FasL, TNF, and TNFR I and II as well as CD40-CD40L. In addition, molecules such as perforins, IFN- γ, and IL-2 regulate the contraction phase of the T Cells (24).
CD4+ T cell subsets
The differentiation of a CD4+ T Cell into distinct subpopulations or cell phenotypes is determined by the nature and concentration of the Ag, the type of APC and its activation state, the cytokine microenvironment that accompanies the antigenic presentation, and the presence and quantity of co-stimulatory molecules, along with other variables.
If the T Cell expresses CD4, it is converted into a T-helper cell (Th) which has a double function: to produce cytokines and to stimulate B Cells to generate Abs. Until recently, only four distinct phenotypes had been identified: Th1, Th2, Th17, and T-regulatory cells (Treg) each of which secretes a different cytokine profile. However, in the last few years, new T-helper subsets such as Th9, Th22, and follicular helpers (Tfh) have been identified. Figure 4 summarizes the main characteristics of these T Cell subsets, the factors that induce them, and the cytokines they produce.
Figure 4
CD4 T Cell subsets. Adapted from Lloyd et al. (30). Abbreviations. TH: T-helper, IFNγ: interferon gamma, DTH: delayed type hypersensitivity, TNF: tumor necrosis factor, FGF: fibroblast growth factor, AHR: airway hyperresponsiveness.
Th1. The differentiation of the Th1 cells is induced by IL-12, IL-18, and type 1 IFNs (IFN-α and IFN-β) secreted by DC and macrophages after being activated by intracellular pathogens. IL-18 potentiates the action of IL-12 on the development of the Th1 phenotype. In general, the response mediated by Th1 depends on the T-bet transcription factor and the STAT4 molecule. These cells produce IFN- γ, IFN-α, IFN-β, and IL-2 and express CXCR3 and CD161 (25). They stimulate strong cell immunity to intracellular pathogens as well as participate in the pathogenesis of the autoinmune diseases and in the development of delayed type hyper-sensitivity. In Th1 cells, the IL-2 increases the expression of T-bet and IL-12Rβ2 which then promotes the sustainability of this phenotype (23).
Th2. A Th2 response is induced by extracellular pathogens and allergens. It is generated by the effect of the IL-4, IL-25, IL-33, and IL-11 secreted by mast cells, eosinophils, and NKT cells. These cytokines induce the intracellular activation of STAT-6 and GATA-3, which initiates the secretion of cytokines of the Th2 phenotype such as IL-4, IL-5, IL-9, IL-13, IL-10, and IL-25, as well as, the expression of CCR4 and ICOS (26, 27). Th2 cells induce immunoglobulin class switching to IgE, through a mechanism mediated by IL-4. The IgE, in turn, activates cells of the innate immune system such as basophils and mast cells and induces their degranulation and the liberation of histamin, heparin, proteases, serotonin, cytokines, and chemokines. These molecules generate contraction of the smooth muscle, increase vascular permeability, and recruit more inflammatory cells. Th2 cells also migrate to the lung and intestinal tissue where they recruit eosinophils (through the secretion of IL-5) and mast cells (through IL-9). This leads to tissue eosinophilia and hyperplasia of mast cells. When acting upon epithelial cells and the smooth muscle (through IL-4 and IL-13), the Th2 cells induce production of mucus, metaplasia of the Goblet cells, and airway hyper-responsiveness as observed in allergic diseases (26). In the Th2 cells, IL-2 induces the expression of IL-4Rα and keeps the loci of the IL-4 and IL-13 genes in an accessible configuration during the final stages of the differentiation of these cells, which helps to conserve this phenotype (23).
Th9. This subset of T-helper cells arises through the effect of TGF-β and IL-4. Th9 cells produce IL-9 and IL-10 and do not express cytokines or transcription factors of the Th1, Th2, or Th17 subsets (28). IL-9 promotes the growth of mast cells and the secretion of IL-1β, IL-6, IL-13, and TGF-β. Nevertheless, IL-9 is not exclusive to this cell subpopulation. It is also produced by Th2, Th17, Treg, mast cells, and NKT cells (29). In allergic processes and infections by helminthes, the IL-9 stimulates the liberation of mast cell products and, through IL-13 and IL-5, indirectly induces the production of mucus, eosinophilia, hyperplasia of the epithelium, and muscular contraction (30).
Th17. These cells are induced by the combined action of IL-6, IL-21, IL-23, and TGF-β. The IL-6 activates the naïve T Cell resulting in the autocrine production of IL-21 which in synergy with TGF-β induces the nuclear transcription factor (ROR)c and the production of IL-17A and IL-17F. IL-23 is essential for the survival and activation of Th17 after its differentiation and selectively regulates the expression of IL-17 (31).
The Th17 cells are mainly located in the pulmonary and digestive mucosa. They produce IL-17A, IL-17F, IL-6, IL-9, IL-21, IL-22, TNF-α, and CCL20. IL-17, in synergy with TNF-α, promotes the expression of genes that amplify the inflammatory process. IL-17 binds to its receptor in mesenchymatous cells such as fibroblasts, epithelial cells, and endothelial cells to promote the liberation of chemokines and inflammation mediators such as IL-8, MCP-1, G-CSF, and GM-CSF (31). IL-17 and IL-22 also induce the production of defensins. The inflammatory environment generated by Th17 cells is associated with diseases that have an important inflammatory component such as rheumatoid arthritis, systemic lupus erythematosus (SLE), bronchial asthma, and transplant rejection (32).
Th22.This T Cell subset is generated by the combined action of the IL-6 and TNF-α with the participation of plasmacytoid DC. Th22 cells are characterized by the secretion of IL-22 and TNF-α. The transcripcional profile of these cells also includes genes that encode for FGF (fibroblast growth factor), IL-13, and chemokines implicated in angiogenesis and fibrosis. The main transcription factor associated with this phenotype is AHR. In the skin, IL-22 induces antimicrobial peptides, promotes the proliferation of keratinocytes, and inhibits their differentiation which suggests a role in the scarring of wounds and in natural defence mechanisms (33). The Th22 cells express CCR4, CCR6, and CCR10 which allows them to infiltrate the epidermis in individuals with inflammatory skin disorders. They participate in Crohn’s disease, psoriasis, and the scarring of wounds (34).
Follicular helper T Cells (Tfh). These cells were discovered just over a decade ago as germinal center T Cells that help B Cells to produce antibodies. The development of these cells depends on IL-6, IL-12, and IL-21. They are characterized by the sustained expression of CXCR5 and the loss of CCR7, which allows Tfh cells to relocate from the T Cell zone to the B Cell follicles that express CXCL13. There, they induce the formation of germinal centers, the transformation of B Cells into plasma cells, the production of antibodies with different isotypes, and the production of memory B Cells (35).
Among all the T-helper cell subsets, the Tfh express the TCR with the highest affinity for Ag and the greatest quantity of costimulatory molecules such as ICOS and CD40L. Furthermore, they express the transcription factor BCL-6 and cytokines such as IL-21, IL-4, and IL-10 which induce the differentiation of B Cells and the production of Ab (35).
Regulatory T Cells (Treg). Regulatory T Cells represent 5% to 10% of CD4+ T Cells in healthy adults. They constitutively express markers of activation such as CTLA-4 (CD152), α receptor of IL-2 (CD25), OX-40, and L-selectin (36). These are considered anergic in the absense of IL-2 which makes them dependent on the IL-2 secreted by other cells. By their mechanism of action and origin, they represent a heterogenous population of cells that can be divided into two: natural Treg cells of thymic origin and induced Treg cells differentiated on the periphery (37).
The natural Treg cells are CD4+CD25high and constitutively express the transcription factor FOXP3+ which is essential for their development. The CD4+CD25−FOXP3− cells can differentiate into Treg cells in the presence of IL-10 and TGF-β and for interaction with immature DC. In contrast, the differentiation of Treg cells is inhibited when mature DC produces IL-6.
The production of Treg cells is essential in preventing autoimmune diseases and avoiding prolonged immunopathological processes and allergies. They are also essential for inducing tolerance to allogenic transplants as well as tolerance of the foetus during pregnancy. They supress the activation, proliferation, and effector function of a wide range of immune cells including autoreactive CD4 or CD8 T Cells which escape negative selection in the thymus, NK cells, NKT, LB, and APC. Like a double-edged sword, Treg cells also supress antitumoral responses, which favors tumor development (37).
Action mechanisms of Treg cells are depicted in Figure 5. These mechanisms can be broadly divided into those that target T Cells (regulatory cytokines, IL-2 consumption, and cytolysis) and those that primarily target APCs (decreased costimulation or decreased antigen presentation). Major mechanisms by which Treg cells exert their functions include (36, 38):
Figure 5
Mechanisms of action of T regulatory cells. Abbreviations. TGFβ: Transforming growth factor beta, CTLA4: cytotoxic T-lymphocyte antigen 4.
Liberation of regulatory molecules such as IL-10, TGF-β, IL-35, and adenosine which inhibit the secretion of specific cytokines of Th1, Th2, and Th17 phenotypes.
Liberation of granzymes and perforins that induce apoptosis of the effector cell.
Competition for IL-2. Treg cells do not produce IL-2, but they express high CD25, the IL-2 receptor α chain, and have the ability to compete with effector T Cells for IL-2. This results in a state of privation of proliferation signals of the effector cells and apoptosis.
Downregulation of APC maduration and co-stimulatory function. CTLA-4 on the surface of Treg cells downregulates or prevents the upregulation of CD80 and CD86, the major costimulatory molecules on antigen-presenting cells.
See Also15.4C: B Cells and T Cells
Additionally, Treg cells induce the production of an enzyme in the DC called IDO (indolamine deoxygenase) that degrades tryptophan, transforming it into proapoptotic metabolites called kynurinines.
Transference of cAMP by gap junctions, which exert immonosuppressive functions.
The clinical importance of these cells is shown in patients with mutations of the FOXP3 gene who develop an immunodeficiency linked to chromosome X which presents with pluriglandular and intestinal manifestations known as IPEX. This autoimmune multiorgan disease requires a bone marrow transplant in early infancy (37).
CD8+ T cells
When a CD8+ T Cell develops its effector functions, it is converted into a cytotoxic T Cell able to attack cells directly and destroy those that are malignant or infected with virus (39). In order to exert this function, a cytotoxic T Cell induces apoptosis in its target cells by the liberation of cytolytic granules or by the expression of ligands for death receptors such as FasL (CD95) (40).
The cytolytic granules contain pore-forming proteins called perforins or cytolysins, proteases known as granzymes or fragmentins, granulolysins which participate in the degradation of membrane lipids, inhibitors of perforins that protect the cytotoxic T Cell from autolysis (calreticulin, catepsin G), and FasL.
Once the immunological synapse between the cytotoxic T Cell and the target cell has been established, the content of these granules is liberated. Perforins polymerize in the plasma membrane and produce pores, which act as channels that allow water entry and generate an osmotic disequilibrium in the cell. Furthermore, they facilitate the passage of granzymes to the cytosol and to the nucleus of the target cell which favors their proteolytic action on the mitochondria and fragmentation of the DNA (41).
Cytotoxic T Cells also liberate IFN- γ and TNF-α, which are important in the defence against viral infections and in controlling the proliferation of tumoral cells (39).
B lymphocytes (B cells)
Ontogeny
The first stages of B Cell development take place in complex microenvironments created by the stromal cells of the bone marrow known as “niches” from which come the stimuli and factors required to initiate a series of cell signals. These, in turn, activate transcription factors that induce, or repress, the expression of different target genes that modulate cell survival, proliferation, and differentiation. IL-7 is critical to the development of the B Cells and is produced by the cells of the stroma.
Thus, and as shown in Figure 6, the development of the B Cells initiates from a haematopoietic stem cell (HSC). This transforms into an early lymphoid progenitor (ELP) and, then, becomes a common lymphoid progenitor (CLP) from which is derived, on one hand, natural killer cells (NK) and dendritic cells (DC) and, on the other, common lymphoid progenitor-2 (CLP-2), which is responsible for the B Cell lineage. This is considered the first stage of the immature B Cells (42-44). A prerrequisite for the development of the B Cells in bone marrow is the absence or supression of protein Notch-1 (N1) signaling, which is necessary for T Cell development (45).
Figure 6
B Cell development and B Cell subsets (see text for details). Abbreviations. HSC: haematopoietic stem cell, ELP: early lymphoid progenitor CLP: common lymphoid progenitor, CLP-2: common lymphoid progenitor-2, NK: Natural Killer cell, DC: Dendritic Cell, (more...)
During the differentiation of the B Cells, a process of gene recombination is structured initially that codes for segments V (Variable), D (Diversity), and J (Joining) of the heavy chain (chain H) together with that of the genes for segments V and J of the light chain (chain L) of the membrane-bound immunoglobulin (mIg). This recombination process is initiated by the complex of proteins RAG1- RAG2 that generate the rupture of the double chain of DNA between segments of genes and specific recognition sites that are also known as “recombination signal sequences.” This process leads to the generation of B Cells that express a wide repertoire of mIg. This will form the B Cell receptor (BCR) which is able to recognize more than 5×1013 different Ags (44,46).
Allelic Exclusion: During its development, the B Cell generates a wide diversity of BCR for the gene recombination process mentioned above. Although each cell has many allelic loci for the different BCR chains (two loci for the heavy chain and multiple loci for the light chain), each mature cell eventually expresses only a single type of receptor. This is achieved by restricting the gene expression of the BCR of a single allele in a process known as allelic exclusion, which involves a monoallelic activation and feedback inhibition (46).
The expression of pre-BCR is an important control point for the recombination of the heavy chain. Its product Igμ associates with a surrogate light chain (SLC), a heterodimer composed of two germline-encoded invariant proteins (VpreB and λ5) to thus produce the molecular complex known as pre-BCR in the B Cell precursors (Figure 7 Panel A) (44,46). In the absence of Ag, signaling through the pre-BCR occurs by the cross-reaction between the positively charged arginine residues present in the λ5-region of the SLC and negatively charged molecules in the stromal cells (47). Once this pre-BCR is expressed on the cell surface, it generates a signal that induces proliferation of the pre-B cells and, as a result, significantly increases their number and guarantees that they successfully carry out the heavy chain recombination. Furthermore, signaling through the pre-BCR is implicated in activation of the gene recombination for the light chain and is thus required for B Cell differentiation to continue (46). With respect to this, it is known that pre-BCR plays an essential role in controlling the development of determined secretory cells of auto-Abs and may represent an important factor in multifactorial autoimmune diseases (47, 48).
Figure 7
A) pre-B Cell receptor. B) B cell receptor. Abbreviations. Igα: Accesory Immunoglobulin alpha (CD79a), Igβ: Accesory Immunoglobulin beta (CD79b).
Based on the differential expression of a complex of surface markers, five stages (A, B, C, D and E) have been described for the development of immature B Cells that occur inside bone marrow (Figure 6). These stages are as follows: CLP-2 corresponds to stage A with expression of B220+, KIT−, CD19−, FLT3+, CD24low/−; CD43+, IgM−; stage B corresponds to early Pro-B cell with expression of B220+, KIT+, CD10+, CD19+, CD24+, CD43+, FLT3−, IgM−; stage C to late Pro-B cell which, furthermore, expresses BP1+. During stage D, the immature B Cell expresses a pre-receptor B (Igμ+SLC λ5 and VpreB) which converts the cell into Pre-B with expression of B220+, CD19+, CD24+, CD25+, and CD43; and finally, stage E corresponds to the B immature cells with expression of B220+, CD19+, CD24+, CD43−, and IgM+, which emerge from bone marrow and are guided towards the secondary lymphoid organs (spleen, lymphonodules, Peyer’s patches, tonsils, and mucosal tissue) to continue their differentiation into transitional B Cells (type 1 and type 2). These, through the effect of molecules such as BAFF and APRIL, differentiate into marginal zone B Cells (MZB) or enter the germinal centers and transform into follicular B Cells (FoB). Later, depending on the Ag stimuli they receive and the microenvironment of cytokines that surrounds them, each of these cells transforms into a plasma Ab-secretory cell or memory B Cells (42,44,49).
Formation of the germinal centers
Homing of B Cells in the spleen is regulated by expression of chemokines such as CCL21, CCL19, and CXCL13 produced by follicular dendritic cells (FDCs). These facilitate the movement of B Cells to the marginal zone or the follicules thereby giving rise to the formation of the germinal centers (GC). A GC is considered to be a specialized microenvironment of lymphoid tissue where an intense cell proliferation, somatic hypermutation, and selection by antigenic affinity occur. Here, the early development of B Cell differentiation is completed and cells undergo apoptosis (50).
During this process, Tfh cells activate the B Cells, which proliferate and create the first part of a germinal center within the follicle. At this stage, somatic hypermutation occurs (this is dependent on proliferation and the microenvironment of the germinal center although the exact factors that induce it are unknown). This process generates a progeny of B Cells with distinct receptors (almost identical but mutated in the variable zones). Some of these receptors do not recognize the presented Ag while others do with greater avidity. Where this maturation is happening, there are Ag-presenting DCs, and those B Cells that have increased their affinity for the Ag recognize it avidly and remain bound. This interaction is known as “maturation by affinity.”
While the former happens, there is a change of BCR isotype through a process known as class-switch recombination (CSR), for which an intrachromosomal deletional rearrangement produces a change from the Cμ chain (which codes the constant region of IgM) to C γ, Cα or Cε, encoding the constant regions of IgG, IgA, and IgE respectively (51). Finally, different B Cells are generated with a BCR that has a specific isotype and a modified affinity for and exit to peripheral circulation. Some of these cells convert into plasma cells and go to the bone marrow (increasing the Igs) while others remain within the same lymphoid organ. Many other B Cells become memory B Cells. The germinal center forms at least one week after contact with the Ag (50).
B cell receptor (BCR)
The BCR is a macromolecular complex that is built in the membrane by IgM/ IgD with two additional Ig accessories denominated Igα (CD79a) and Igβ (CD79b) (Figure 7, Panel B). The membrane-bound immunoglobulins (mIgs) are glycoproteins with a basic monomer. Each of these is made up of four polypeptidic chains of which two are heavy or H chains with a molecular weight of approximately 65 kDa, and the others are light or L chains with a molecular weight of 25 kDa. Each L chain is linked to an H chain by a disulphur bridge. The H chains are linked to each other by at least one other disulphur bridge. These IgM or IgD monomers correspond to the extracellular segments of the BCR. However, the mIg also have another two segments, the transmembranal and the intracytoplasmic, which result from an extension of the carboxy-terminal portion of the two H chains. The two V domains, which form the active sites allowing each BCR to bind specifically to an antigenic determinant, are found in the amino-terminal (H and L) portions of each peptide chain of the BCR. The antigen-BCR interaction is a non-covalent reaction (52). The intracytoplasmic region of mIg, which presents only 3 aminoacids (lysine-valine-lysine), is very small and does not permit mlg to carry out the signaling process per se. The transmembranal segment of the C-terminal portion of the H chain of mIg consists of 25 aminoacids found close to the tyrosine kinase (PTK) enzymes which, in turn, are close to the heterodimers (Igα and Igβ). The latter are responsible for the signaling process since they can provide the substrate for the tyrosinases in their ITAM regions (52).
Signaling mechanisms
Stimulation of the B Cells via antigenic BCR begins with the recognition and capture of the Ag through BCR molecules. This induces their aggregation and triggers the signaling process by activating the SRC family kinase (SFK) which then phosphorylates the ITAM moieties of the accessory chains Igα and Igβ. These carry out the same function as the δε chain (CD247) to activate the TCR and produce lipid-raft-associated calcium-signaling module forms (Figure 8). This complex contains 3 classes of activated protein tyrosine kinases (PTKs): i) Lyn, Fyn, and Blk of the Src family; ii) Syk/ZAP70; and iii) of Bruton (Btk) of the Tec family
Figure 8
B Cell receptor signaling mechanisms. Abbreviations. Lyn: Tyrosine-protein kinase Lyn, SHIP: 5’ phospholipid phosphatase SHIP-2, Syk: Spleen tyrosine kinase, BLNK: B Cell linker protein, BTK: Bruton kinase of Tec family, PLCγ2: phospholipase (more...)
This initiates the formation of a macromolecular complex known as ‘signalosome’ composed of the BCR, the tyrosine kinases already mentioned, some adaptor proteins such as CD19 and BLNK (B Cell linker protein), signaling enzymes such as the phospholipase C gamma - 2 (PLC γ2), the phosphatidylinositol 3-kinase (PI3K), and molecules of the Vav family.
The signaling produced by the signalosome activates multiple signaling cascades which implicate other kinases, GTPases, and transcription factors such as NF-kB, Bcl6, NF-AT, FoxO, Jun, and ATF-2, etc. In order to study these signaling routes in greater detail, our recommendation is to consult the following web page: http://www.cellsignal.com/reference/pathway/B_Cell_Antigen.html
The activation of all these mechanisms gives rise to changes in cell metabolism, gene expression, and the organization of the cell cytoskeleton. The result of the response is determined by different factors or conditions such as: the state of maturation of the B Cells, the nature of the Ag, the magnitude and duration of signaling through the BCR, and the signals of other receptors such as CD40, the receptor of IL-21, and BAFF-R. Thus, many other transmembranal proteins such as CD45, CD19, CD22, PIR-B, and Fc γRIIB1 (CD32) modulate specific elements of the signaling BCR. During the in vivo processes, the B Cells are also activated by APCs, which capture and present the antigenic fractions on their cell surfaces. This type of B Cell activation by such cell membrane–associated Ags also requires a reorganization of the B Cell cytoskeleton. Thus, it is to be expected that the complexity of the signaling mediated by the BCR allows diverse biological effects to occur, including cell survival, tolerance, or apoptosis as well as proliferation and differentiation in Ab-producing cells or memory B Cells. (http://www.cellsignal.com/reference/pathway/B_Cell_Antigen.html) (53, 54).
Co-receptors of the BCR
During B Cell activation, another series of molecules participates to build a molecular complex that acts as a BCR co-receptor. These molecules can significantly increase the signaling Induced initially by the BCR and include: CR2 (CD21), CD19, CD45, CD38, and CD81 (Figure 9) (44, 55). The binding of CD21 with the Ags which are found to be opsonized with the complement fraction C3d facilitates grouping of the co-receptor with the BCR. The position of the CD21 allows the associated kinases to phosphorylate the tyrosine residues in the cytoplasmic domain of the CD19, and this binds to the tyrosine-kinases of the Src and the PI3-kinase family.
Figure 9
B Cell co-receptors. Abbreviations. CD: Cluster of differentiation, CR2: Complement receptor-2.
Co-stimulatory molecules
Another important aspect in the activation of the B Cells is the presence of molecules which positively or negatively regulate the process. Together these are known as co-stimulatory molecules and some of them are described as follows.
B cell activating factor (BAFF) is a cytokine and member of the TNF family. It is produced by a wide variety of cells (neutrophils, monocytes, macrophages, DCs, and T Cells) (49). It is essential for the maturation and survival of the B Cells since it participates in the processes of differentiation and proliferation. To date, three BAFF receptors have been identified: i) BAFF-R, ii) TACI (transmembrane activator, calcium modulator, and cyclophilin ligand interactor), and iii) BCMA (B Cell maturation factor). Interaction with the BAFF-R is the most critical to making maturation possible and causing BAFF-R to act in combination with the BCR (44, 49). Elevated levels of BAFF have been observed in the sera of patients with SLE, Sjögren’s Syndrome (SS), and rheumatoid arthritis (RA) (49).
APRIL (A proliferation-inducing ligand) is a BAFF hom*ologue that binds to TACI and BCMA but does not interact with BAFF-R. In addition to its co-stimulatory function, APRIL improves the ability of B Cells to present Ag and to increase their survival time and also regulates tolerance. On the other hand, it promotes the proliferation and survival of malignant B Cells and other tumoral cells. As for BAFF, elevated levels of APRIL have been observed in the sera of patients with SLE (56).
CD40, a transmembranal glycoprotein type I receptor, belongs to the TNF receptor superfamily. It is expressed on a great variety of cells (e.g., monocytes, Ag-presenting cells, endothelial cells, smooth muscle, fibroblasts, keratinocytes, and platelets) and in all the stages of the B Cells. This receptor N-terminal extracellular domain includes several cysteines, has four subregions, and binds to its ligand (CD40L or CD154) through the 2nd and 3rd subregion. The interaction between the CD40 and its ligand, which is present in the T Cells, increases the expression of cytokines (IL-2, IL-6, IL-10, TNF-α, Lymphotoxin-α, and TGF-β), chemokines, metalloproteinases of the matrix, growth factors, and adhesion molecules. This allows signaling processes to occur through activation of several protein tyrosine kinases (PTK) such as ERK-1, ERK-2, p38, and JNK. This, in turn, permits the activation of several transcription factors such as NF-kB, AP-1, and NF-AT. These processes lead to maturation, differentiation, and cell proliferation of the B Cells with the subsequent production of Abs and, finally, the production of memory B Cells. Additionally, mutations in this molecule, or in its ligand, are responsible for the syndrome of hyperIgM linked to chromosome X. Although this CD40-CD154 interaction mediates many mechanisms of the humoral and cell immune responses, it is also implicated in a wide spectrum of chronic inflammatory and autoimmune diseases. The blockage of this signaling route is therefore considered to be a potential therapeutic mechanism for these pathologies (57).
Other co-stimulatory molecules: Additionally, and as is detailed in Table 2, the B Cell expresses another series of molecules from the superfamilies CD28/B7 and TNF/TNFR which interact directly with the T Cells as co-stimulatory molecules. Included among these are B7-1 (CD80), B7-2 (CD86), CD70, the ICOS ligand (ICOS-L), the CD30 ligand (CD30-L or CD153), the 4-1BB ligand (4-1BBL), SLAM (CD150), etc. (21).
Molecules of adhesion
B-cell mobilization requires adhesion mechanisms in which several types of molecules participate such as chemokines, their receptors, the selectins, and integrins.
Lymphoid chemokines are a group of chemokines/receptors that are expressed constitutively in the lymphoid tissue cells and aid in the recirculation of the lymphocytes. They include CXCL12, CXCL13, CCL19, CCL21, CXCR5, CCR7, and CXCR4 (16). The differential positioning of the B Cells in the GC or in the external part of the follicules is regulated by the EBI2 receptor coupled to protein G (also known as GPR183) which directs the B Cells to the perifollicular and interfollicular areas. This location of the B Cells mediated by EBI2 is important for the first stages of the Ab response (16). Molecules such as S1P (Sphingosine 1 Phosphate) and its S1PR1 receptor, in turn, control the exit of the B Cells from the lymphonodules (16, 58).
The Ab-secretory cells express high levels of integrins such as α4β1 and LFA-1 as well as ICAM-1, α5β1, and α6β1. The plasma cells in the intestine and the mammary glands express α4β7 (59, 60).
The selectins are another family of molecules that contribute to the adhesion and mobilization of the B Cells. The plasmocytes have a high expression of PSGL-1 (P selectin glycol-protein ligand 1) which recognizes the P and the E selectin (59). Another molecule that participates in the B Cell homing processes is the CD22, a member of the Siglec (sialic acid-binding immunoglobulin-like lectin) family which preferentially binds sugars with α2,6 -sialic acid radicals and is mainly present in mature B Cells (59).
B cell subsets
Mature B Cells can be divided into several subsets based on their location, cell surface phenotype, Ag specificity, and activation routes.
The transitional B Cells are considered to be the first stages of development of the B Cells once they leave the bone marrow to migrate to the secondary lymphoid organs. The lymphocytes CD20+, CD21±, CD23±, IgM++, and IgD± CD38++ are designated B Cell transitional type-1 (T1) and differentiate from type 2 (CD20+, CD21++, CD23±, IgM++, and IgD++CD38±). The transitional B Cells T2 can evolve intomarginal zone B Cells or GC (44,61,62).
The Follicular B Cells (FoB) or B-2. These are generated directly in the bone marrow and reach the follicules of secondary lymphoid organs and the circulation. They are considered to be resting (naïve) cells and constitute the largest subpopulation of B Cells. Their differentiation is influenced by a great variety of factors including chemokines, BCR signaling, and some Ags. They participate in T-dependent (TD) immune responses since they can use the BCR to engulf the Ag, process it, and present it to the Ag-specific T Cells (63).
The Marginal zone B Cells (MZB). This type of cell is located as a sentinel in the marginal zones of the spleen which correspond to the interphase between the circulation and the splenic lymphoid tissue. These B Cells also inhabit the inner wall of the subcapsular sinus of the lymph nodes, the epithelium of tonsillar crypts, and the subepithelial dome of intestinal Peyer’s patches. In humans, they present the following phenotype: IgMhigh IgDlow CD1c+ CD21high CD23−CD27+. MZB express high levels of TLR (similar to macrophages, DCs, and granulocytes) phenomena that allow them to play a role of a bridge between innate and adaptive immune responses. MZB have the ability to rapidly respond to an Ag-specific stimulus by using both T-independent (TI) and dependent (TD) mechanisms and to transform into plasma cells that secrete IgM, IgG1, IgG2 (for both, TD and TI pathways), IgA1 (on the TD pathway), and IgA2 (on the TI pathway) low affinity Abs (51).
B1 B Cells. These are the first B Cells to form in the fetal liver. They subdivide into B1a and B1b with the former expressing the glycoprotein of membrane CD5, which is absent in the latter. Both express CD9 and CD45RA markers, are involved in type TI immune responses, are found in the peritoneal and pleural cavities, and are the main source of circulating Abs. As with the MZB, the B-1 responds rapidly to Ag-specific stimuli and transforms into plasma cells. Their numbers have been observed to increase in experimental studies and in humans with autoimmune diseases.
Seven sub-populations of mature peripheral B Cells have been identified in human tonsils based on the expression of two surface markers (CD38 and IgD). This has made it possible for a TD model for the differentiation of mature B Cells to be proposed. The subpopulations suggested are: i) B- cell mature naïve (Bm); ii) B- cell mature 1, Bm1 (CD38- IgD+); iii) Bm2 (CD38+ IgD+). - These three would be activated by their specific Ag in the extra-follicular areas through interaction with interdigital DC and Ag-specific T Cells. Once activated, the three can be transformed into Bm2’ founder cells of GC (CD38++ IgD+), and then differentiate into Bm3 centroblasts (CD38++ IgD−). These Bm3 cells are selected during their differentiation into Bm4 centrocytes (CD38++ IgD−) as a function of their BCR affinity. Finally, these cells differentiate into either memory B Cells (CD38+ IgD−), Bm5 cells (CD38−IgD−), or high affinity plasma cells (Figure 10) (62).
Figure 10
Sub-sets of peripheral mature B Cells. Adapted from Cornec et al (62). Abbreviations. Bm: B Cells mature.
Plasmocyte or Ab-secretory cells. These differentiate from an activated B Cell which, in the presence of IL-2 and IL-10, stops expressing surface molecules such as CD19, CD20, CD22, HLA class II molecules, and their BCR. These cells also lose the ability to divide. At the same time, they undergo a series of cellular modifications, e.g., an increase in their cytoplasm due to enhanced growth of the endoplasmic reticulum that is required to harbor the high number of ribosomes for robust production of Abs. They also stop expressing CXCR5 and CXCR7 and increase CXCR4 which causes them to lose contact with the DC and forces the TFH cells to migrate from the GC to the medullar cords of the ganglia (44).
Two classes of plasma cells are known: Short-lived cells, which are located in the medulla of the ganglia and, later, quickly exit to the circulation and seek the site where the Ag enter to initiate, in situ, the production of specific IgM type Abs. Long-lived cells migrate to a special niche in the bone marrow following the expression of SDF-1 by the stromal cells. Within this niche, an extended production of IgG type Abs, which can be used to mount a prolonged or permanent defence against the Ag that originally activated the B Cells, is initiated. The prolonged survival of these latter cells is due to the effect of IL-16. Development of short- or long-lived plasma cells depends on the expression of the transcription factor Blimp-1 (B lymphocyte-induced maturation protein-1) (44).
Memory B Cells. There are several subsets of memory B Cells that are classified based on their origin, the differential expression of CD27, and the isotype of the mIg being expressed. Three different origins for the cells have been described: i) the spleen, ii) the germinal center, and iii) the intestine lamina propria outside the GC. In the spleen, they present CD27−IgG+ markers. At the GC, they are CD27+IgM+IgD−and change from mIg to CD27+IgG/IgA+. Last of all, those generated in the intestine express CD27−IgA+ (44, 64).
Regulatory B Cells (Bregs). B Cells also liberate a wide variety of cytokines and, as with the T Cells, can be classified according to the profile of cytokines that they produce. Thus, the Bregs are a functional sub-set of B Cells, and they contribute to the maintenance of the fine equilibrium required to guarantee tolerance. Bregs restrict the excessive inflammatory responses that are produced during autoimmune diseases or that can be caused by unresolved infections. IL-10 is fundamental to the function of Bregs since they inhibit pro-inflammatory cytokines (IFN- γ, IL-17), reduce the expression of the MHC clase II molecules and support the differentiation of Tregs (65). It has also been reported that CD40-CD154 interaction is an essential activation pathway for the Bregs. With regard to the Breg surface markers, there is considerable controversy and the consensus is that there is no single marker, or even set of markers that make identification of this type of regulatory cell possible. Among the markers reported in these types of cells are: CD1dhigh, CD5+, CD19+, CD24high, CD27variable, CD38 variable, CD138 variable, IgMhigh, and IL-10+ (65).
Innate B cell helpers
It has also been demonstrated that B Cells receive additional help from other cells besides the T-helper cells. These include: the iNKT, DCs, epithelial cells, macrophages, and diverse granulocytes, including neutrophils, eosinophils, basophils, and mastocytes (51, 66-68).
The iNKT cells express a TCR invariant Vα14+ that recognizes soluble glycolipids, e.g., α-galactosylceramide, presented by DCs or subcapsular macrophages in the CD1d context. Soluble glycolipids improve the expression of CD40L and IFN-α, which stimulates the maturation of DCs in the efficient antigen-presenting cells. These interact with the TFH cells to form active CGs with the consequent generation of long-living plasma cells that produce IgG (66, 67).
The B Cell helper neutrophils (NBH) occupy the perimarginal zone of the spleen in the absence of inflammation or infection. They interact with perifollicular B Cells and MZB through the liberation of APRIL, BAFF, CD40L, IL-6, and IL-21 in response to stimuli by cytokines and microbial products. This interaction results in CSR processes by which the plasma cells generated stop expressing IgM and start to produce IgG and IgA (68).
In general, we can say that these innate immune cells can stimulate and help the response of Abs to both TD and TI processes. For the former, these cells make use of helper signals for B Cells in the GC and in the central lymphoid sites such as the bone marrow. On top of this, theTI type responses take place on the surface of the mucosae and in the marginal zone of the spleen to give rise to a rapid response from natural Abs (66, 67).
Types of immune responses mediated by Abs
Traditionally, the humoral immune response mediated by Abs is classified based on whether or not the B Cells receive help from the T Cells, i.e., if they are TD or TI responses of the thymus (69).
One charactistic of the TD response is the induction of follicular GCs in which the Tfh cells select B Cells with high affinity BCRs by somatic mutation and cause them to differentiate into memory B – cells. In contrast, the TI response may be provoked by microbial ligands, which are classified as type TI -1 or by extensive crosslinking of the BCR with the Ag, which is known as type TI -2 (69).
Recent studies describe the existence of two new mechanisms that participate in the B- cell response, e.g., the B Cells can also receive TD type help but from cells of innate immune system. Some examples are those induced from iNKT cells which are classified as TD - 2. Furthermore, an innate TI-3 type response has been described that involves myeloid cells such as B Cell helper neutrophils (NBH) (68), monocytes, eosinophils, mastocytes, and basophils (70). Much remains to be discovered with regard to the steps in the human B - cell response, including which cells and molecules are involved in the new mechanisms described above. The authors of the present chapter, therefore, recommend that those readers who are interested in a deeper understanding of this subject keep abreast of the reports that confirm and clarify such mechanisms.
