Cell Cooperation in the Antibody Response

Chapter 9 Cell Cooperation in the Antibody Response




Summary




The primary development of B cells is antigen independent. Pre-B cells recombine genes for immunoglobulin heavy and light chains to generate their surface receptor for antigen.


T-independent antigens activate B cells without requiring T-cell help. They can be divided into two groups. TI-1 antigens can act as polyclonal stimulators, while TI-2 antigens are polymers which activate by cross-linking the B-cell receptor.


T-dependent antigens are taken up by B cells, processed and presented to TH cells. T cells and B cells usually recognize different parts of an antigen.


B-cell activation requires signals from the B-cell receptor and costimulation. CD40 is the most important costimulatory molecule on B cells. Ligation of the B-cell–coreceptor complex can lower the threshold of antigen needed to trigger the B cell. Intracellular signaling pathways are analogous in B cells and T cells.


Activated B cells proliferate and differentiate into antibody-forming cells. Cytokines from TH cells control the process of division, differentiation and class switching.


Somatic hypermutation of immunoglobulin genes, followed by selection of high-affinity clones is the basis of affinity maturation. These processes occur in germinal centers.


Class switching is effected by somatic recombination occurring within the heavy chain genes. Somatic hypermutation and class-switching by recombination are linked processes, which require selective targeting of DNA-modification and DNA-repair enzymes to the heavy chain gene locus.


The antibody response is the culmination of a series of cellular and molecular interactions occurring in an orderly sequence between a B cell and a variety of other cells of the immune system. This chapter discusses the principles of B-cell development, activation, proliferation and differentiation leading to the generation of plasma cells and memory cells. In addition, the consequences of these interactions, including affinity maturation and class switching, are examined.


In adults, B cell development occurs in the bone marrow and does not require contact with antigen. During this time the B cells rearrange the genes for their heavy and light chains, and synthesize cell surface IgM which acts as their antigen receptor (BCR). The BCR complex includes:



Immature transitional B-cells exit the bone marrow and enter the periphery where they further mature in secondary lymphoid organs. If these cells do not encounter antigen, they soon die within a few weeks by apoptosis. If, however, these mature B cells encounter specific antigen, they undergo activation, proliferation and differentiation leading to the generation of plasma cells and memory B cells.image



Development of B cells



Primary B-cell development is antigen independent


Within the bone marrow, a sequence of immunoglobulin rearrangements and phenotypic changes takes place during B-cell ontogeny, analogous to that described for T cells in the thymus which leads to the production of the B cell’s antigen receptor (Fig. 9.w1). The molecular processes involved in immunoglobulin gene rearrangement have been described in Chapter 3, and this section relates these events to B-cell development.



The earliest stage of antigen-independent B-cell development identified is the progenitor B (pro-B) cell stage. Pro-B cells can be divided into three groups based on the expression of:



Early pro-B cells express TdT alone, intermediate pro-B cells express both TdT and B220, and late pro-B cells express B220 and have downregulated TdT. B220 remains expressed on the surface throughout the remainder of B-cell ontogeny.


As the cells progress through the pro-B cell stage, they rearrange their Ig heavy chain genes and begin to express CD43 (leukosialin), CD19, RAG (recombination-activating gene)-1 and RAG-2. As late pro-B cells pass into the pre-B-cell stage, they downregulate TdT, RAG-1, RAG-2, and CD43.



Genes for immunoglobulin H and L chains are recombined to generate the surface receptor


Pre-B cells can be divided into large mitotically active pre-B cells and small non-dividing pre-B cells. Both large and small pre-B cells express Igμ heavy chains in the cytoplasm (cμ) and the pre-B cell–receptor complex on their surface (Fig. 9.w2). Large pre-B cells have successfully rearranged their Ig heavy chain genes. As these cells pass from the large pre-B cell group into the small pre-B cell group, they begin to rearrange their Ig light chain genes and upregulate RAG-1 and RAG-2. The final stage of B-cell development is the immature B-cell stage. Immature B cells have successfully rearranged their light chain genes and express IgM. Once again, RAG-1 and RAG-2 expression has been downregulated. As immature B cells develop further into mature B cells, they begin to express both IgM and IgD on their surface. These mature B cells are then free to exit the bone marrow and migrate into the periphery.




Other phenotypic markers such as CD25 (IL-2Rα chain), CD79a (Igα), CD79b (Igβ), and c-Kit can help to identify particular populations of pro-B, pre-B or immature B cells (see Fig. 9.w1).






B cell activation



T-independent antigens do not require T cell help to stimulate B cells


The immune response to most antigens depends on both T cells and B cells recognizing the antigen in a linked fashion. This type of antigen is called a T-dependent (TD) antigen.


A small number of antigens, however, can activate B cells without MHC class II-restricted T cell help and are referred to as T-independent (TI) antigens (Fig. 9.1).



Importantly, many TI antigens are particularly resistant to degradation. TI antigens can be divided into two groups (TI-1 and TI-2) based on the manner in which they activate B cells:




Many TI-1 antigens possess the ability in high concentrations to activate B cell clones that are specific for other antigens – a phenomenon known as polyclonal B cell activation. However, in lower concentrations they only activate B cells specific for themselves. TI-1 antigens do not require a second signal.


TI-2 antigens, on the other hand, are thought to activate B cells by clustering and cross-linking immunoglobulin molecules on the B cell surface, leading to prolonged and persistent signaling. TI-2 antigens require residual non-cognate T cell help, such as cytokines.


Several signal transduction molecules are necessary for mediating TI antigen responses in B cells. These include CD19, HS1 protein and Lyn.



T-independent antigens induce poor memory


Primary antibody responses to TI antigens in vitro are generally slightly weaker than those to TD antigens. They peak fractionally earlier and both generate mainly IgM. However, the secondary responses to TD and TI antigens differ greatly. The secondary response to TI antigens resembles the primary response, whereas the secondary response to TD antigens is far stronger and has a large IgG component (Fig. 9.2). It seems, therefore, that TI antigens do not usually induce the maturation of a response leading to class switching or to an increase in antibody affinity, as seen with TD antigens. This is most likely due to the lack of CD40 activation (see below). Memory induction to TI antigens is also relatively poor.



There are potential survival advantages if the immune response to bacteria does not depend on complex cell interactions, as it could be more rapid. Many bacterial antigens bypass T-cell help because they are very effective inducers of cytokine production by macrophages – they induce IL-1, IL-6 and tumor necrosis factor-α (TNFα) from macrophages. The short-lived response and lack of IgG may also be due to lack of costimulation via CD40L and lack of IL-2, IL-4 and IL-5, which T cells produce in response to TD antigens.


It is possible to convert TI antigens into T-dependent antigens by altering their structure. For example, pneumococcal polysaccharides are TI antigens and do not induce immunological memory or antibodies in infants. However, conjugation of pneumococcal polysaccharides to a carrier protein induces polysaccharide-specific antibody in infants, and memory similar to T-dependent antigens (imagesee Method boxes 9.1 and 2.1).



Method box 9.1 Measuring antibody production – the PFC assay and ELISPOT


Various methods have been developed for assaying antibody production. Two such methods are the plaque-forming cell (PFC) assay and the enzyme-linked immunospot assay (ELISPOT).


Antibody-forming cells (AFCs) are measured by means of a quantitative PFC assay (Fig. MB9.1.5), originally developed by Niels Jerne in the 1960s. B cells (e.g. spleen cells) are plated in agar with sheep red blood cells sensitized by binding the specific antigen to their surface. Antibody produced by any B cell will coat the red blood cells. Following the addition of complement, these coated cells may be lyzed, causing the appearance of a zone of lyzed cells (plaque) in the agar. Figure MB9.1.5 shows the appearance of such a plaque, with a B cell in the center, under the microscope. The plaques are then counted to give a quantitative measure of the number of PFCs.


Another way of detecting antibody-producing cells is by means of an enzyme-linked immunospot assay (ELISPOT). An ELISPOT assay starts out by coating a plastic well with antigen and adding a known quantity of B cells. The antigen coated onto the plastic will then capture any antibody in the vicinity of the activated B cell that is producing the antibody. After a period of time, the B cells are removed, and the specific antibody can be detected by adding an enzyme-labeled anti-immunoglobulin plus chromogen. Development of the label in this assay results in a spot surrounding the active B cell. Counting each spot and knowing the quantity of B cells originally added to the well allows one to enumerate the frequency of B cells producing the specific antibody. Method Box 2.1, Fig. MB2.1.3 shows the method of detecting antibodies and the appearance of the spots on the developed plates. In addition to analyzing specific antibody-secreting B cells, the ELISPOT assay has been adapted to measure the frequency of cytokine-secreting T cells and various other cell types (right panel). With the improvement in ELISPOT assay plate design and in ELISPOT detection equipment, antibody- or cytokine-secreting cells can now be detected at the single cell level.





Activation of B cells by T-dependent antigens



T cells and B cells recognize different parts of antigens


In the late 1960s and early 1970s, studies by Mitchison and others, using chemically modified proteins, led to significant advances in understanding of the different functions of T cells and B cells. To induce an optimal secondary antibody response to a small chemical group or hapten (which is immunogenic only if bound to a protein carrier), it was found that the experimental animal must be immunized and then challenged using the same hapten–carrier conjugate – not just the same hapten. This was referred to as the carrier effect.


By manipulating the cell populations in these experiments, it was shown that:



These experiments were later reinforced by details of how:



One consequence of this system is that an individual B cell can receive help from T cells specific for different antigenic peptides provided that the B cell can present those determinants to each T cell.


In an immune response in vivo, it is believed that the interactions between T and B cells that drive B cell division and differentiation involve T cells that have already been stimulated by contact with the antigen on other antigen-presenting cells (APCs), for example dendritic cells.


This has led to the basic scheme for cell interactions in the antibody response set out in Figure 9.3. It is proposed that antigen entering the body is processed by cells that present the antigen in a highly immunogenic form to the TH and B cells. The T cells recognize determinants on the antigen that are distinct from those recognized by the B cells, which differentiate and divide into antibody-forming cells (AFCs). Therefore two processes are required to activate a B cell:




Jun 18, 2016 | Posted by in IMMUNOLOGY | Comments Off on Cell Cooperation in the Antibody Response

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