21 Immune system

Lothar Thomas

21.1 Immune response

The immune system is in a constant defense against a multitude of invading pathogens e.g., bacteria, viruses, fungi, parasites and a pattern of non-living foreign substances. The pathogen initiates complex interactions between the pathogen-derived molecules and host sensors. The immune response is categorized in two components, innate immunity and adaptive immunity. The innate immune response, which is the first line and most effective defense plays a crucial role in defense against a majority of infections. The adaptive immune response generates a nearly unlimited number of antigen receptor specificities by random gene rearrangement that can detect extracellular and intracellular antigens using B cell antigen receptors and T cell antigen receptors, respectively /1, 2/.

21.1.1 Innate immune response

The innate immune response consists of epithelial barriers, a family of soluble antimicrobial peptides, danger and pathogen associated molecular pattern-recognizing molecules and receptors of various innate immune cells. The cells, receptors and molecules, such as pentraxins, complement, innate antibodies, nucleotide binding oligomerization domain (NOD)-like and Toll-like receptors, mast cells, monocytes/macrophages, granulocytes, natural killer cells, myeloid dendritic cells, Langerhans cells and antigen presenting cells bind to the pathogen and initiate its clearance through transcription independent immunological processes (e.g., phagocytosis, degranulation, and complement fixation) /1/.

The innate immune cells include various tissue macrophages, dendritic cells and neutrophils that express a family of innate receptors and sensors known as pattern recognition receptors (PRRs). The receptors are evolutionary conserved germ-line encoded receptors that sense pathogen associated molecular patterns (PAMPs). The PAMPs are not only essential for pathogenicity to establish infection in the host but also essential for the survival of the pathogen. PRRs include several families of receptors /2/:

• Toll-like receptors (sensing bacteria)
• NOD-like receptors (sensing bacteria)
• RiG-I-like receptors (sensing viruses)
• C-type lectin receptors (sensing fungi and mycobacteria)
• DNA-sensing molecules (sensing viruses)

This highly orchestrated defense mechanism against infectious and inflammatory insults initiates the formation of acute phase proteins driven by the endogenous cytokines interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α The cytokines are produced by macrophages and other leukocytes induced by pathogens binding to PRRs /3/.

Collectively, the responses of the innate immunity initiate either direct killing or inhibition of pathogen replication. Additionally, the responses induced by the innate immune response initiate pathogen-specific adaptive immunity through B- and T cells /2/.

Natural immune barriers

A number of mechanical factors protect the organism against pathogens. These are the skin, mucus layers, saliva and tears, in addition to a large number of chemical mechanisms such as the low pH of gastric juice.

Intestinal mucosal immunity

Mucus layers cover a surface area of several hundred square meters, which is 200 times larger than the surface area of the skin /4/. They represent the most important portals of entry for microbial pathogens that kill some 10 million children below the age of 5 years worldwide every year. Most of these deaths are due to diarrheal disease.

The mucus layers of the body defend against pathogens mainly by producing secretory IgA (sIgA). There are about 10¹⁰ IgA producing plasma cells per meter of bowel whereas the total number in all of the lymph nodes, spleen, and bone marrow together only amounts to 2.5 × 10¹⁰ plasma cells. Approximately 80% of the body’s immunoglobulin (Ig) producing plasma cells are located in the intestine and produce Ig locally in the lamina propria. In an adult, approximately 3 g of sIgA is secreted into the intestinal lumen each day.

The intestinal mucosal immune system represents the most robust Ig producing mechanism in the body and the front-line defense system against the antigens and pathogens in the more than 1,000 kg of food that passes through the intestine each year. In the intestine, the induction and regulation of mucosal immunity takes place primarily in the Peyer’s patches and mesenteric lymph nodes (gut-associated lymphoid tissue, GALT).

The mucosal immune system ensures the homeostasis of the defense system via two mechanisms:

• By limiting epithelial contact and mucosal invasion by antigens and pathogens by binding them into immune complexes
• By immunosuppression, also known as “oral tolerance” when induced via the intestine. Regulatory T cells (Treg) are located in the mesenteric lymph nodes. Treg are triggered by dendritic cells, which present them with dietary antigens and pathogens. The Treg induce mucosal tolerance to these substances by down regulating the immune system. The neonatal period is critical, both with regard to infections and to food allergies, because the mucosa and the immuno regulatory system is poorly developed.

Mucosal immune system of the respiratory tract

Particles with a diameter of less than 5 μm can reach the lower airways of the respiratory tract. There, they encounter the natural resistance of surfactant proteins (SP) and soluble components of the innate immune response such as lysozyme, lipopolysaccharide (LPS)-binding protein, fibronectin, lactoferrin, defensins, complement, and secretory IgA /5/.

The SPs are members of the collectin family and contain lectin-rich and collagen-rich domains. SPs protect against microorganisms by acting as opsonins and stimulating phagocytosis by alveolar macrophages. Low concentrations of SP aggregates stimulate lymphocyte proliferation whereas higher concentrations inhibit it. The SPs (SP-A and SP-D) stimulate cytokine secretion by macrophages. However, SPs do not protect against microbial pathogens only; they also bind pollen and mite allergens.

β defensins are low molecular weight cationic peptides with broad antimicrobial activity against bacteria, fungi, chlamydia, and viruses. They are produced by epithelial cells following the stimulation of toll-like receptors by microbial antigens.

21.1.2 Immune recognition

21.1.2.1 Danger-associated molecular patterns (DAMPs)

The immune system was developed before the separation of vertebrates and invertebrates. Therefore, it has highly conserved structures that can recognize the molecular antigen patterns of pathogens but are not differentiated enough to detect individual antigens. Many microorganisms express standard antigen molecular patterns (PAMPs) that are recognized by Pattern recognition receptors (PRRs) of immune cells. PRRs are localized on macrophages, dendritic cells and granulocytes /1/. Refer to:

• Tab. 21.1-1 – Danger associated molecular patterns (DAMPs)
• Tab. 21-1-2 – Pattern recognition receptors that mediate the intracellular signal transduction required to initiate inflammation.

21.1.2.2 Pathogen associated molecular patterns (PAMPs)

PAMPs include structures such as fungal β1,3-glucans, bacterial lipopolysaccharide (LPS), peptidoglycans, phosphoglycan, lipoteichoic acid, mannan, double-stranded RNA, and bacterial DNA. Certain pathogen groups are characterized by a particular type of PAMP and so are recognized globally, for example /5/:

• Gram negative bacteria contain LPS in their cell wall. The binding of LPS to the corresponding CD14 receptor and toll-like receptor 4 (TLR 4) on a macrophage trigger a signaling cascade that causes the macrophage to secrete cytokines, which in turn stimulate the immune system.
• Gram positive bacteria contain lipoteichoic acid
• Fungi contain mannan.

Alarmins

Alarmins are danger signals that are produced during inflammation, infection, and stress (Tab. 21.1-1 – Danger-associated molecular patterns (DAMPs)).

Opsonins

These group of proteins attract and bind microbial pathogens. Mannose-binding protein (MBP), for example, is produced as an acute phase protein by the liver. MBP is a Ca²⁺-dependent lectin receptor that binds mannose-rich carbohydrates on bacteria, fungi, parasites, and, occasionally, viruses. MBP acts as an opsonin in human blood. It accelerates the phagocytosis of mannose rich proteins and activates the classical and alternative pathways of the complement system /5/.

21.1.2.3 Pattern recognition receptors (PRRs)

These proteins are expressed as receptors on dendritic cells, macrophages, and B cells or are secreted. PRR binding sites include Ca²⁺-dependent lectin, leucine-rich peptides, and cystine-rich peptides. PRRs stimulate endocytosis in macrophages and trigger signaling cascades that lead to inflammation. The overall result is that the immune system can capture and attack entire classes of microorganisms with a relatively small number of different receptors.

Endocytosis receptor

In the absence of opsonins, this receptor, which is a mannose binding protein, facilitates the uptake of microorganisms into macrophages, dendritic cells, and occasionally endothelial cells. This is how Pneumocystis carinii is incorporated into alveolar macrophages. It is thought that the susceptibility of AIDS patients to Pneumocystis carinii infection is due to a modification in the mannose binding receptor caused by HIV /5/.

Toll-like receptor (TLR)

TLRs are found on the cell membrane of immune cells. They recognize microbial components, activate signal transmission in the nucleus, and trigger the expression of genes involved in the inflammatory response. TLRs were discovered at the end of the twentieth century in Drosophila, where they are essential receptors for host defense against fungal infection. The mammalian TLR receptor group has 11 members; in humans, TLR 10 is functionally the most important. Like cytokine receptors, TLRs have an extracellular and a cytoplasmic domain. The cytoplasmic domain is comparable to that of the IL-1 family. The extracellular domain varies, however. In IL-1R, it is immunoglobulin-like whereas in the TLRs it consists of an accumulation of leucine residues (leucine rich repeats, LRRs) /6/.

TLRs recognize conserved pathogenic structures, so they recognize many microbial components such as lipoproteins, peptidoglycans, lipoteichoic acid in Gram-positive bacteria, lipopolysaccharide (LPS) in Gram-negative bacteria, the glycosylphosphatidylinositol anchor of Trypanosoma species, and phenol-soluble modulin in Staphylococcus epidermidis. Following contact with a microbial component, TLRs located on macrophages induce the expression of genes, which results in the production of inflammatory cytokines (IL-1, IL-8) and co stimulatory molecules. Under the influence of inflammatory cytokines and co stimulatory molecules activated by TLRs, macrophages present antigens to the T-helper cells, and in this way, couple the innate immune defense to the adaptive immune system (Fig. 21.1-1 – Pathogen recognition by dendritic cells and macrophages by means of toll-like receptor (TLR)).

TLRs do not always bind antigens directly. When TLRs are activated by LPS, the LPS must be coupled to an LPS binding protein (an acute phase protein).

Co stimulatory molecules

Macrophages and dendritic cells are stimulated by TLRs to express B7 co stimulatory molecules (CD80 and CD86) on the cell surface. Co stimulatory molecules provide additional signals that, along with the TLR signals, are necessary for T-cell activation /7/.

21.1.3 Cells expressing receptors of innate immune system

21.1.3.1 Granulocytes and macrophages

The cells of the innate immune system first travel to the area of tissue damage. The PO₂ in this area is usually reduced. Polymorphonuclear neutrophils (PMNs) are the first to arrive at the diseased or injured site, followed by monocytes. Since PMNs have few mitochondria and obtain most of their energy from anaerobic glycolysis, they can perform their immune recognition and defense functions in spite of the hypoxic conditions /8/. For granulocyte functions in the innate immune response, refer to Section 19.7 – Polymorphonuclear neutrophil function.

More than 95% of macrophages in the tissues are derived from monocytes of the hematopoietic system and the remainder are derived from local tissue-resident macrophages. Circulating monocytes normally have a half-life of up to 3 days before extravasation into the tissues occurs. During inflammatory response with increased monocyte production in the bone marrow, the half-life of the circulating monocytes decreases and macrophages accumulate in the affected tissue. Macrophages are found in almost all tissues, where they often exist in tissue-specific forms (e.g., as histiocytes in connective tissue, Kupffer cells in the liver, alveolar macrophages in the lung, microglia in the central nervous system, and osteoclasts in bone).

In addition to their phagocytic capabilities, macrophages have a broad spectrum of other functions, ranging from antigen presentation, through antibacterial and antitumor activity, to secretion of regulatory substances such as enzymes, prostanoids, and cytokines.

Macrophages express a range of receptors:

• To recognize carbohydrates such as mannose. Because mannose is not normally found on the surface of vertebrate cells but is expressed on microbial cell surfaces, recognition of these sugar enables the macrophages to differentiate between self and non self antigens.
• To recognize phosphatidylserine. Cells that are subject to programmed cell death (apoptosis) express phosphatidylserine and are cleared by macrophages. Cells in necrotic tissue, on the other hand, release substances such as heat shock proteins, which involve macrophages in host defense as part of an inflammatory response (refer to Section 19.1 – Inflammatory response).
• For complement factors and immunoglobulins. Complement factors and antibodies opsonize microbes by coating them and accelerating their phagocytosis. After complement activation (see Chapter 24 – The complement system), the component C3b binds to microbial cell surfaces. Macrophages have receptors for C3b and are activated by C3b binding. In the absence of C3b, macrophages are activated by microorganisms coated with IgG, IgA, or IgM. Bacteria coated with C3b or antibacterial antibodies are surrounded in the sense of a zipper mechanism through mediation by Fc receptors on the surface of the macrophage (Fig. 21.1-2 – Nonspecific phagocytosis of a microorganism by a macrophage). Within the macrophage, pathogens are attacked by a range of mechanisms including reactive oxygen species (hydroxyl anion, super oxide anion), hypochlorous acid, NO, and antimicrobial substances such as lysozyme and cationic proteins.

In simple terms, the function of macrophages is to acquire a pathogen, processes it into smaller antigenic substances, and present them to the T cells of the adaptive immune system in order to initiate an immune response (Fig. 21.1-3 – Antigen presentation to T cells). Recognition of these antigens by T cells triggers the immune response. Macrophage function is then under the control of T cells. Interferon-γ (IFN-γ) produced by T cells is the prototypical macrophage-activating cytokine.

IFN-γ-activated macrophages produce /9/:

• IL-12 and TNF-α. IL-12 is one of the most important cytokines secreted by macrophages because it regulates the Th1 immune response. The production of IFN-γ by T cells is maintained by IL-12 and IFN-γ in turn stimulates macrophages to express co stimulatory molecules of the B7 family, which have an important role in the recognition of PAMPs by macrophage pattern recognition receptors.
• IL-1 and IL-10. IL-1 influences the Th1 immune response while IL-10 influences the Th2 immune response (Fig. 20.1-4 – Development of the sub populations of T-helper cells under the influence of IL-4 and IL-12). IL-1 stimulates the immune response while IL-10 suppresses it. IL-10 therefore reduces the production of B7 co stimulatory molecules, TNF-α, and macrophage-inhibitory factor (MIF).

Macrophage function can be summarized as follows:

• Macrophages amplify T-cell responses
• The function of the macrophages themselves is then regulated by the products (cytokines) of the T-cell immune response.

Thus, macrophages play a central role in determining the extent of the immune response.

21.1.3.2 Dendritic cells (DCs)

DCs are produced in the bone marrow and are derived from myeloid or lymphoid cell lineages. They have a characteristic star-shaped structure due to their numerous cytoplasmic extensions. This means that they have a large cell surface, which enables them to establish a high degree of contact with surrounding cells. In this way, one DC can activate 100–3,000 T cells. Antigens are ingested by macro pinocytosis. Interstitial fluid is taken up into the cell and antigens are concentrated by ejecting water through special channels.

Dendritic cells occur /10/:

• As Langerhans cells in the squamous epithelia of the epidermis and supra basal layers in the skin
• As interstitial cells in the heart, lungs, liver, and other organs
• In the covering of afferent lymphatics (veiled cells)
• As inter digital cells in T cell-rich regions of lymph organs
• As follicular cells in lymph organs. These differ from the DCs mentioned previously because they are thought to act as memory B cells.
• DCs are strategically located in the organism to ensure that invading pathogens are recognized. They continuously absorb antigens from the extracellular milieu and search through them for pathogenic antigen patterns (e.g., microbial pathogens).

Dendritic cells express a whole range of antigen recognition receptors, for example /10/:

• Mannose receptors, LPS receptors, and toll-like receptors to recognize fungal mannan, LPS in Gram-negative bacteria, and lipoteichoic acid in Gram-positive bacteria
• Receptors such as FcγRII (CD32), FcγRI (CD64), FcεRI, and the C3bi complement receptor (CD11b) to make the endocytosis of immune complexes more effective.

To activate the immune response, DCs express /10/:

• High concentrations of antigen presenting molecules such as MHC class I and MHC class II molecules and CD1a molecules. Like in macrophages, antigens are processed internally before being presented with MHC molecules as short peptides on the cell surface. MHC class II molecules present the peptides to T-cell receptors on the surface of T-helper cells, while MHC class II molecules present antigens to receptors on cytotoxic T cells (Fig. 21.1-3 – Antigen presentation to T cells). DCs are particularly effective at priming i.e., the antigen-specific activation of naive T cells (Th0) that have not had previous contact with the antigen.
• Co stimulatory molecules of the B7 family and intercellular adhesion molecules: ICAM-1 (CD54), ICAM-3 (CD50), and lymphocyte function-associated antigens such as LFA-3 (CD58), B7-1 (CD80), and B7-2 (CD86).

The mechanisms of the innate and adaptive immune response are activated as a result of this concerted activity. DCs can also migrate into local lymph nodes and trigger an adaptive immune response there. Overall, DCs represent an important link between innate and adaptive immunity.

21.1.3.3 Natural killer (NK) cells

NK cells are an important subpopulation of lymphocytes that play a role in the innate immune response to infection and malignancy. They comprise 10–20% of circulating lymphocytes and have the morphology of large granular lymphocytes (LGL cells) in the peripheral blood smear. They are called natural killer cells because of their ability to lyse target cells without prior sensitization and without the need for MHC antigen expression by the target cell /11/.

NK cells can be distinguished from T and B lymphocytes by the lack of T-cell and B-cell receptors on their surface. They are phenotypically classified as CD56⁺CD3^–. CD56 is a neuronal cell adhesion molecule (NCAM). Other surface receptors present on subsets of NK cells include FcRγIII (CD16), IL-2 receptor, c-kit receptor, CD7, CD2, and CD8.

NK cell function (target cell recognition, killer activity) is regulated by a complex interplay of activating and inhibitory cell surface receptors. Activating receptors include β₂-integrins, CD2, and receptors belonging to the immunoglobulin super family that are defined by their molecular weight (e.g., NKp46 or NKp30). The inhibitory MHC class I receptors have a molecular structure that is either lectin-like (killer cell lectin-like receptor, KLR) or immunoglobulin-like (killer cell immunoglobulin-like receptor, KIR) (Fig. 21.1-4 – Structure of inhibitory NK-cell receptors). KLR and KIR play a crucial role in determining what happens to target cells /11/.

Theories about how NK cells operate suggest that KLR and/or KIR can bind to any cell in the organism and come into play once the NK cell has bound to the target cell. If they find specific MHC structures on the target cell to which they can bind and the target cell emits an inhibitory signal, the target cell escapes lysis. If not, inhibition does not occur and the target cell is lysed (Fig. 21.1-5 – Activating and inhibitory NK cell receptors and their interaction with target cells). In virus-infected cells and tumor cells, MHC molecules are down regulated, making these cells more susceptible to lysis by NK cells.

NK cells destroy infected and malignant cells via the following mechanisms /11/:

• Cells coated with IgG antibodies are recognized via IgG receptors (FcγR) and destroyed by the mechanism of antibody-dependent cytotoxicity
• Killer cell activating receptors recognize structures on the target cell. In the absence of a signal from the inhibitory receptors, the contents of its cytotoxic granules in the NK cells are released. Perforin performs a hole in the target cell membrane and cytotoxic enzymes from the granules are then injected into the interior of the cell, whereby the cell contents are dissolved.

21.1.4 Complement system

The innate immune response frequently involves one of the three complement systems at an early stage (see also Chapter 24 – The complement system). The following pathways are activated:

• The classical pathway (activated by immune complexes)
• The alternative pathway (activated by the microbial cell wall)
• The lectin pathway (activated by the interaction between mannose binding proteins and microbial carbohydrates).

Activation of these pathways leads to production of the following:

• Fragment C3b of complement component C3. C3b is deposited on the surface of the pathogen, where complement activation takes place. This initiates phagocytosis of the pathogen by macrophages or dendritic cells, which have C3b receptors.
• Fragments C3a, C4a, and C5a, which release mediators of inflammation from mast cells. C5a also acts as a chemoattractant for polymorphonuclear neutrophils.
• The membrane attack complex, which is comprised of C5b, C6, C7, C8, and C9. This perforates the target cell membrane, leading to cell destruction.

21.1.5 Cytokines

The highly orchestrated defense mechanism against infectious and inflammatory insults initiates inflammatory cells (macrophages and other leukocytes) to produce inflammatory cytokines e.g., interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α and function as an integrated network to regulate the components of the immune system (see to Chapter 20 – Cytokines and cytokine receptors).

Innate immune response to microbial pathogens is shown in Fig. 21.1-6 – Innate immunity: responses following initial contact with microbes and microbial products.

21.1.6 Thymus

The Thymus is critical for normal development of the immune system. Infants who undergo thymectomy have reduced T-cell counts that do not recover to normal levels, even after several years of follow-up /32/.The thymus atrophies with age, the involution begins in infancy, and is accelerated in puberty. With declining central T-cell formation, peripheral clonal proliferation of T-cells the body compensates the number of T-cells that have an increasingly limited number of T-cell receptors. Age related reduction of T-cell receptors hinder immune surveillance, increasing the risk of infection and cancer. In a study about health consequences of thymus removal in adults /32/ the authors draw the conclusion: thymus contributes to new T-cell production in adulthood and to the maintenance of adult human health. The disruption of homeostasis caused by thymectomy is sufficient to adversely affect critical health outcomes, which argues strongly that the adult thymus remains functionally important.

21.1.7 Adaptive immune response

The fundamental features of the adaptive immune system are the ability for learning, gaining memory, and obtaining antigen specificity. The system provides effective response against a spectrum of environmental antigens and has the following characteristics:

• Large variety of somatic antigen receptors
• High degree of antigen specificity
• The ability to generate a rapid immune response to a previously encountered antigen based on immunological memory.

In the adaptive immune response, pathogens and their products are processed primarily by components of innate immunity. Peptides are produced of a size that allows them to associate with MHC molecules in the cytoplasm of the antigen presenting cells to form a peptide MHC complex (pMHC) /12/.

The pMHC is transported to the cytoplasmic membrane, where it is recognized by T lymphocytes (T cells) of the adaptive immune system via their T-cell receptors (TCRs). Because the TCR avidity is rather low, the concentrations of TCRs and pMHCs in the micro environment must be high enough to allow T-cell activation. Interaction between the APC and T cell is facilitated by specialized structures on their membranes called micro domains, which bring the receptors and ligands together, as well as by co receptors.

The following co receptors and ligands play a supporting role:

• CD4/CD8 molecules on the T-cell membrane, which interact with molecules of MHC II and MHC I on APCs
• CD28 molecules on T cells, which react with CD80/CD86 ligands on APCs
• CD40L (CD154) on activated T cells, which react with CD40 molecules on APCs.

Supported by the co receptors, TCR and pMHC interact in their micro-domains and activate the immune system.

The TCR repertoire consists of 10¹²–10¹⁵ possible receptors. These include receptors that bind self-peptide/MHC complexes and could theoretically destroy the body’s own tissues. In an attempt to prevent this anti-self reactivity (autoimmune reaction), bone marrow-derived lymphocytes are educated and selected in the thymus so that, in principle, autoimmune reactions cannot occur. Because this selection is not infallible, small numbers of self-reactive T cells escape and reach the peripheral tissues. In most people, however, they are harmless because the self-peptide/MHC complexes do not reach the critical concentration required to trigger an autoimmune reaction. Nevertheless, some auto aggressive T cells (known as driver cells) may trigger autoimmune reactions. Regulatory T cells (Treg) and natural killer cells (NK cells) counteract the activity of driver cells.

Antigen-mediated activation of T cells and B cells and subsequent stimulation by cytokines secreted by the T cells lead to differentiation:

• Of naive T cells (Th0) into T helper cells (Th1), Th2 cells that suppress immune reactions, and into regulatory Treg. To ensure an effective immune response, these cells are stimulated to proliferate clonally (clonal selection theory) /7/.
• Antibody production by B cells
• Macrophage activation.

Activated T cells, B cells, and macrophages collaborate with the effectors of innate immunity to eliminate the pathogen.

T and B cells are derived from primordial stem cells in the fetal liver and bone marrow under the control of interactions with stromal cells, stem cell factors, and colony stimulating factors. The initial stage of lymphocyte development is not antigen dependent.

However, once the lymphocytes express a mature antigen receptor, their survival and differentiation require the presence of an antigen /7/.

T cells, B cells, Treg and NK cells are derived from a common precursor cell. From this precursor cell, the following develop:

• Pre B cells (immature B cells)
• Pre T cells (immature T cells).

21.1.7.1 T-cell system

T cells regulate adaptive immune responses to pathogens and tumor cells and detect antigens presented by self MHC molecules. The T cell antigen receptor on conventional αβ-T cells recognizes peptide fragments bound to MHC-I or MHC-II molecules. Each developing T cell expresses a unique T cell receptor and generation of self-MHC restricted and self-tolerant T cell repertoire results from a multistep selection process in the thymus. Naive T cells develop from stem cells in the bone marrow, migrate to the thymus and become thymocytes.

The thymic selection occurs in the following steps (Fig. 21.1-7 – Selection of T cells that migrate into the thymus from the bone marrow) /13/:

• The first stage of selection is mediated by specialized thymic cortical epithelial cells, which present self-peptides (hormones of the neurohypophyseal family, tachykinin family, and insulin super family) together with MHC proteins. The thymocyte T-cell receptor (TCR) recognizes amino acids from these self peptides and MHC antigens. Thymocytes expressing a TCR weakly reactive to the host’s self antigens receive a maturation signal to generate the functional T cell repertoire in the periphery (positive selection). In contrast, thymocytes with strongly self-reactive TCRs receive a death signal (negative selection). A failure to prevent strongly self reactive T cells from entering the peripheral T cell pool is one of the main causes of autoimmune diseases. To initiate signaling an antigen activated TCR scans multiple MHC-I and MHC-II co receptors to find one that is associated with the signal initiating kinase Lck. The kinase phosphorylates immunoreceptor tyrosine based activation motifs (ITAMs) and activating tyrosines on ZAP 70 protein. MHC-II restricted TCRs require a shorter antigen dwell time (0.2 sec.) to initiate negative selection compared to MHC-I restricted TCRs (0.9 sec.) because more CD4 coreceptors are LcK loaded compared to CD8.
• T cells positively selected on the cortical epithelial cells undergo further selection. In this step specificity testing for antigens is carried out by dendritic cells and macrophages in the thymus medulla. In order to pass this selection step, the TCR must have a corresponding antigen specificity. However, because TCR genes rearrange haphazardly, the probability of a T cell having a corresponding TCR is low. If it does have a matching TCR, the signal that triggers automatic apoptosis of the T cell is switched off and the T cell is released into the circulation as functional CD3⁺CD4⁺T cell or CD3⁺CD8⁺T cell and usually migrates to the lymph nodes. More than 95% of T cells are not selected at this stage and therefore die in the thymus and fail to reach functionality.

A small subset of T cells that pass through the thymus possess a γ/δ receptor. These cells remain in the thymus for a short time only and develop in many locations outside the thymus (e.g., in the intestine-associated immune system). Like NK cells, they have cytotoxic activity and can lyse target cells by releasing perforin and lytic enzymes.

The ontogenetic development of the T-cell system is as follows: CD3⁺T cells are detectable from the 10th week of gestation while CD4⁺T cells and CD8⁺T cells are present from the 14th week of gestation.

Naive T cells

Until puberty, the thymus supplies the organism with naive T cells (Th0 cells) /14/. Even after this ceases, the size of the naive T-cell pool remains stable due to post thymic expansion of naive T cells. Naive T cells can secrete IL-2 but lack the ability to express classic effector cytokines such as IFN-γ and IL-4. Because they have not undergone clonal selection during activation with a foreign antigen, they have a highly diverse T-cell receptor repertoire.

Two populations of naive T cells exist in adults: one dormant subset from the thymus and a second subset comprising naive T cells that have proliferated in the periphery. The surface molecule CD31 (PECAM-1) can be used to distinguish CD31^+thymic naive T cells from CD31^–central naive CD4⁺ T-cells.

Individuals with reduced numbers of CD31^+thymic naive T cells are potentially low responders with respect to the primary immune response. Individuals with increased numbers of CD31^–central naive CD4⁺ T cells may be more predisposed to autoimmunity.

21.1.7.2 B cell system

B cells (B lymphocytes) are produced by hematopoietic stem cells throughout life. Mature B cells recognize pathogens and contribute to their elimination. They secrete immunoglobulins (Ig), present antigens, up regulate co stimulatory molecules, produce reactive oxygen species and cytokines, and express toll-like receptors /15, 16/.

B cells are produced in the bone marrow. Transcription factors such as PU.1, E2A, and paired box protein 5 (PAX5) are necessary for B-cell development. Successful rearrangement of heavy chain immunoglobulin gene segments in pro-B cells leads to their differentiation into pre-B cells that express μH, the IgM heavy chain (Fig. 21.1-8 – Antigen-independent development of B cells and their receptors).

As of this stage, clonal expansion of the B cells and rearrangement of the Ig light chain gene segments are possible. Auto reactive B cells that express IgM on their surface are selected and deleted. The surviving naive B220⁺IgM⁺ B cells leave the bone marrow and migrate to the spleen, where they undergo further maturation via transitional stages.

All B cells undergo transitional stage T1 (Fig. 21.1-8):

• A large proportion of the T1 B cells migrate to the periarterial lymphoid follicles of the spleen, where they acquire CD23 and IgD molecules and differentiate into T2 B cells. They become long living follicular B cells that recirculate between the spleen and peripheral lymph nodes until they die (half life 4.5 months) or encounter an antigen and undergo further differentiation. These B cells, also known as F0 B cells or B2 cells, express the surface molecules IgM^low, IgD^high, CD21^high, and CD23^high. B2 cells are antibody producing cells that constitute 80–90% of the cells in the spleen.
• A small proportion of the T1 B cells migrate to the marginal zone (MZ) of the spleen and remain there as MZ B cells. They express the surface molecules IgM^high, IgD^low, CD21^high, and CD23^low. These cells, also known as B1 cells, react quickly to antigens and involve macrophages and dendritic cells in an immune response. They express the activation markers CD80, CD86, CD40, and CD44 as well as the co stimulatory molecules B7-1 and B7-2 much more strongly than B2 cells. Although B1 cells do not recirculate, they migrate to the peripheral lymph nodes following contact with pathogens, where they activate immune defense. B1 cells have a half life of more than 54 weeks. They secrete IgM antibodies and express CD5 and CD11. B1 cells exhibit significantly less receptor selectivity than B2 cells and mainly produce the so called “natural antibodies”. Natural antibodies are poly reactive IgM antibodies that recognize a wide range of antigens and have a high complement binding capacity but low antigen affinity and selectivity. The natural blood group antibodies anti A and anti B are produced by B1 cells.

B cells undergo programmed development with the following stages (Fig. 21.1-9 – Class switch recombination following contact between B2 cell and antigen):

• Rearrangement of Ig heavy chain genes at the pro B cell stage
• Clonal expansion at the pre B cell phase
• Arrangement of light chain genes with production of IgM at the immature B cell stage
• Antibody production and immunoglobulin (Ig) switching following contact with an antigen.

Three mechanisms contribute to the diversification of the repertoire of the B cell system pool:

• VDJ recombination (also known as somatic recombination), a mechanism in which variable (V), diversity (D), and joining (J) gene segments are combined in the B cell to form an antigen receptor
• Somatic hyper mutation. This involves mutations in the VDJ sequence of the variable domains of B cell antigen receptors that lead to increased antigen specificity
• Class switch recombination. This step enables the variable domain of the heavy chain (V_H) to be expressed at the antigen binding site in association with a different constant region of the heavy chain (C_H). This enables the production of different Ig isotypes (IgG, IgA, IgE) (Fig. 21.1-10 – Structure of the mature and immature B cell receptor). As a result, the innate immune system can eliminate an Ig bound antigen in a variety of ways without altering the antigen specificity.

Whereas VDJ recombination in B and T cells takes place in the thymus and bone marrow, further development of the B cell occurs in the germinal centers of the secondary lymph organs (spleen, lymph nodes).Following secondary antigen contact in the lymph nodes, mutations in the VDJ sequence occur, resulting in improved antigen specificity. This process is called somatic hyper mutation.

In addition the differentiation of subsets of activated B cells into memory cells take place in the secondary lymph organs such as Peyer’s patches. Most plasma cells have a life span of only a few days but some can survive for longer in the bone marrow.

21.1.7.3 Antigens

Antigens are recognized based on their structure. Small antigens (haptens) do not elicit an immune response. Carbohydrates are usually poorly immunogenic and have to be coupled to a carrier to elicit an immune response.

Cell membrane receptors on T cells and B cells have binding sites with a size of 600–1700 square Angstroms, which can only bind small parts of a complex antigen. These small parts are called epitopes. Complex molecules therefore have a characteristic epitope pattern.

The antigen binding site of an antibody or the peptide MHC complex of a T cell receptor is complementary to the antigen structure. The complementary antibody and receptor structure bind non covalently. Antigen recognition only occurs if the complementary molecular structures are in relatively close proximity. For small antigens, the binding site may be a pocket or cleft, but in most cases, it is an undulating surface /7/.

The α/β T cell receptors only recognizes a linear peptide. These are formed only after processing by antigen presenting cells such as macrophages and dendritic cells.

Antibodies, whether free or bound to B cells, recognize only a small part of complex antigens, referred to as the antigenic epitopes such as those present in native protein structures. Epitopes that easily fit into a B cell receptor antigen pocket or a corresponding receptor binding site are those that dominate the polyclonal immune response.

Cryptic epitopes that are normally not recognized efficiently are more easily recognized following antigen processing by macrophages or dendritic cells. The mode of antigen presentation is also important. After processing, dendritic cells present the antigen in a form such that only few epitopes are offered to the T cell, while antigen presentation by B cells results in a greater diversity in the T cell immune response /17/.

21.1.7.4 Antigen receptors

Antigen recognition in adaptive immunity involves specific interactions between antigen epitopes and receptors on B cells and T cells. B cells recognize soluble protein and non protein (bacterial polysaccharide) antigens, with and without the help of T cells. T cells only recognize antigens that are presented in combination with MHC molecules (i.e., by cells). The difference in antigen recognition is important since it ensures that both soluble and cell bound antigens are eliminated /18/.

Each T-cell and B cell receptor has an antigen recognition unit and a signaling unit. The antigen recognition unit has 10¹²–10¹⁵ variable regions. This remarkable diversity of the immune repertoire is achieved by random rearrangement of barely 400 genes in the early stages of lymphocyte development /7/.

B cell receptors

The genes that encode the B cell receptors (BCRs) are located on three chromosomes /7/:

• The IGH cluster for the heavy chain is located on chromosome 3. This cluster contains gene segments for the variable (V), constant (C ), diversity (D), and joining (J) regions of the immunoglobulin.
• The IgK cluster for the kappa light chain is located on chromosome 2
• The IgL cluster for the lambda light chain is located on chromosome 22.

The structure of the B cell receptor is shown in Fig. 21.1-10 – Structure of the mature and immature B cell receptor.

Antigen recognition and presentation by B cells play a fundamental role in the immune response. Following B cell receptor activation by antigen in combination with co- stimulatory molecules such as CD40, B cells become potent antigen presenting cells. They process incorporated antigens in the same way as dendritic cells and macrophages and present the antigen together with an MHCI-protein. They activate CD4⁺ T cells and CD8⁺ T cells and produce cytokines.

T-cell receptors

T-cell receptor (TCR) genes are organized in a similar way to B cell receptor genes and also contain V, C, D, and J gene segments. The genes that encode the T cell recognition unit are located on three chromosomes /4/:

• TCRA/D is located on chromosome 14 and encodes the α chain
• TCRB is located on chromosome 7 and encodes the δ chain
• TCRG is located on chromosome 7 and encodes the β chain.

Each locus contains multiple V, D, and J genes, but none for D segments. Each lymphocyte uses a different combination of these gene segments to create the genetic code of its antigen receptors, which results in a high degree of diversity. The structure of the T-cell receptor is shown in Fig. 21.1-11 – Structure of the mature T-cell receptor (TCR) and pre T cell TCR.

21.1.7.5 Regulatory T cells and effector T cells

Naive T cells encounter pathogen derived antigens in lymphoid tissues and differentiate into regulatory T cells and effector Tcells. Quiescent naive T cells generate ATP through fatty acid β-oxidation (FAO)-driven oxidative phosphorylation (OXPHOS). On activation, T cells switch to anaerobic glycolysis, a process that better accomodates the energetic demands of rapid cell division /31/.

Immunoreaction is dependent on the interaction of the effector T cells and the regulatory T cells /31/:

• Multiple types of effector cells eradicate different pathogens through distinct mechanisms. Transcriptional programming governed by master transcription factors largely determines the functional phenotype of the T cell. Effector T cells typically detect pathogens and execute their function within tissue. Some effector T cells remain in the tissue and become tissue resident memory T cells to provide rapid protection against pathogen reinfection.
• Regulatory T cells play a critical role in resolving the inflammatory response and are defective in many autoimmune diseases. Metabolic programming is complex in regulatory T cells because the cells can switch metabolic states at different stages in their lives and in different microenvironments. Fox P3, the master transcription factor that directs regulatory T cell function, suppresses glycolysis-related genes and induces FAO-driven OXPHOS-related genes that are required for maximum suppression of effector T cells. The ability of regulatory T cells to switch between different metabolic programs may enhance their ability to preserve suppressive function while adapting to the different metabolic substrates found in tissue microenvironments /31/.

21.1.7.6 Antigen-dependent T-cell recognition

Antigen-dependent T cell receptors (TCRs) on the surface of the cell are associated with the CD3 complex of molecules that transmit signals into the cell when the TCR binds antigen /19/. This complex consists of a CD3γ molecule, a CD3δ molecule, and two molecules of CD3ε as well as a disulfide linked τ-chain homodimer. Cross-linking within the TCR when it binds to the antigen MHC complex initiates signal transmission. Aggregation of the receptor leads to phosphorylation of tyrosine residues in the cytoplasmic portion of the CD3 complex. The signal that is thus triggered initiates the transcription of various gene sequences in the cell nucleus. This results in cell proliferation and cytokine production. Refer to Fig. 21.1-12 – Activation of the T cell receptor of CD4⁺ T helper cells.

The B cell antigen recognition unit also associates with two signaling molecules, Igα (CD79a) and Igβ (CD79b), which transmit the activation signal into the cell in the event of antigen binding.

Co stimulatory signals

Antigen recognition by TCRs is associated with a high level of promiscuity. A second activation signal is therefore required to prevent inappropriate lymphocyte responses. This is achieved by co stimulatory signals, which are produced by contact between TCRs and ligands on the surface of neighboring cells or by TCR stimulation by cytokines. The following lymphocyte molecules react with co stimulatory ligands /20/:

• CD28 with the antigen-presenting dendritic cell B7, or CD28 with molecule 4 (CTLA-4) that is associated with the cytotoxic T lymphocytes. The proliferation and differentiation of T cells and the synthesis of IL-2 are all activated.
• CD154 ligands CD40 (expressed by B cells). The ligation of CD40 by CD154 of antigen stimulated CD4⁺T cells, stimulates B cell protein kinases that initiate antibody class switching. The switch fails when defects in the gene encoding CD154 occur. This is the case in X-linked hyper IgM syndrome with IgG, IgA, and IgE markedly decreased, but normal or elevated IgM.

Co stimulatory signals are also provided by cytokines such as TNF-α, IL-1, and IL-6. In the absence of co stimulation, antigen binding does not activate the T cell but leads instead to T cell anergy and apoptosis.

Refer to Fig. 21.1-12 – Activation of the T cell receptor of CD4⁺ T helper cells.

Inhibitory signals

IL-10 and TGF-β produce signals that down regulate the immune response. Binding of CTLA-4 to B7 or of IgG to B cell Fcγ receptors also exerts an inhibitory effect.

21.1.7.7 T cell independent immune recognition

Some antigens are recognized directly by B cells without the involvement of T cells. These include polysaccharides, polymerized flagellin, and microbial DNA, especially cytosine guanine dinucleotide sequences that are flanked by 5’ purines and 3’ pyrimidines. When they bind to B cell receptors, these antigens are taken up intra cellularly and processed into short peptides. The peptides, together with MHCII molecules, are expressed on the cell surface and recognized by adjacent Th2 helper cells. The helper cells become activated and express co stimulatory molecules such as the CD40 ligand (CD154). When CD154 on the helper T cell binds to CD40 on the B cell, a signal is generated that triggers an immune response in the form of antibody production.

Clonal selection

Activation of B cells, CD4⁺T cells, and CD8⁺T cells by antigens results in clonal selection. Each antigen can be recognized by only a few thousand lymphocytes. Following activation of B cells by CD4⁺T cells a signal is generated that prompts the B cell to begin the process of somatic hyper mutation and immunoglobulin class switching. A large number of antibodies with different specificities are produced from the lymphocytes, but every B cell expresses antibodies with only one of the many potential specificities /19/. B cells of this type are selected to participate in the immune response and proliferate to generate a family of cells (clone) that all produce the same antibody. Because a number of different clones are produced in most immune responses, microbial infections always elicits a polyclonal immune response.

Memory T and B cells

When a lymphocyte that has never been activated by an antigen (naive lymphocyte) encounters an antigen for the first time, the resulting immune response includes the production of memory T cells and B cells in addition to effector T cells and B cells. If the same antigen is encountered again, the resulting secondary immune response is faster and more effective. More lymphocytes, higher antibody concentrations and antibodies with higher avidity and specificity are produced than in the primary immune response.

21.1.7.8 Course of the immune response

The adaptive immune response is a complex process that is triggered by lymphocytes that circulate continuously through the body to detect antigens. T cells and B cells require approximately 30 minutes for each circulation. Antigen recognition and immune responses take place at various locations /19/:

• In the spleen, for antigens circulating in the blood
• In the local lymph nodes or the bronchial lymphatic tissues, when the antigens enter the respiratory tract or mucous membranes
• Responses to intranasal antigens and inhaled pathogens occur in the adenoids and palatine tonsils
• Antigens from the gut are taken up by specialized epithelial cells that transport the antigen across the epithelium to Peyer’s patches.

Registration of antigens entering the body through mucous surfaces activates lymphocytes in the mucosal associated tissues. Mainly lymphocytes in the mucous surfaces are CD8⁺α/β T cells with the appearance of large granular lymphocytes. An established function of these cells is to support the production of secretory IgA, while T cells with γ/δ receptors have a direct role in host defense. If, for example, an immune response is induced in the Peyer’s patches, then sensitized lymphocytes enter the blood and travel to the lamina propria of the intestinal mucous layer, where large amounts of secretory IgA are produced. However, responses induced in one mucous location (i.e., intra nasal), can also induce increased production of secretory IgA in the mucous tissues of other organs that are not exposed to the pathogen.

Lymphocytes from the blood enter the lymph nodes via specialized post capillary venules. The passage is mediated by adhesion molecules, for example the constitutively expressed selectin on lymphocytes. L-selectin binds to endothelial adhesion molecules of the venules. This interaction induces the lymphocytes to express lymphocyte function associated antigen (LFA-1), which facilitates the adhesion of the cells. In the next step lymphocytes migrate across the endothelium into lymphoid tissue. The spleen lacks these specialized venules.

The immune response is induced in the germinal centers of secondary lymph organs such as the spleen, lymph nodes, and Peyer’s patches. These germinal centers consist of a mesh of follicular dendritic cells in which CD4⁺T cells present antigen, B cells proliferate, plasma cell precursors are produced, immunoglobulin class switching occurs, and memory cells are separated. The germinal center provides an environment that optimizes the antibody response by bringing all of the relevant cellular components into contact /19/.

21.1.7.9 Antigen processing and presentation

T cells recognize peptides, which are presented by antigen presenting cells by way of MHC-I and MHC-II molecules (Fig. 21.1-13 – Simplified model of antigen presentation):

• CD8⁺T cells, also known as cytotoxic or killer cells, recognize peptides that are bound to MHC-I molecules. The molecules present peptides that are synthesized in the cytoplasm and are present in nearly all nucleated cells. Such antigens are mainly self peptides or peptides of viral origin in infected cells. CD8⁺T cells produce cytotoxic molecules such as Fas ligand, perforin, and serine esterases to destroy target cells. Because CD8⁺T cells are specialized in eliminating antigens produced in the cytoplasm, they are particularly efficient in attacking virus infected cells.
• CD4⁺T cells, also referred as T helper cells, recognize peptides that are bound to MHC-II molecules. These molecules present peptides that are released by cellular vesicles such as endosomes. They contain exogenous peptides that are taken up from the environment and ingested by polymorphonuclear neutrophils and macrophages. Antigens are cleaved into peptides by acid hydrolases such as nucleases, proteases, lipases, and glucosidases within endocytic vesicles. In the endosomes, the peptides are bound to MHC II molecules and trans located to the cell surface.

T-helper (Th) cell paradigm

Cytokines exert an important influence on the type of immune reaction that optimally eliminates a pathogen. CD4⁺T cells are cytokine secreting helper cells and are differentiated into Th1 and Th2 cells. According to the T helper cell paradigm /21/:

• Th1 cells activate the cell mediated immune response through cytotoxic CD8⁺T cells and macrophages. They secrete IL-2, IFN-γ and TNF-β. By producing IFN-γ and IL-2, they stimulate cytotoxic CD8⁺T cells to kill virus-infected cells (Fig. 21.1-14 – Immune response of the Th1 cell), activate macrophages to kill intracellular pathogens (Fig. 21.1-15 – Cell mediated immune response of the Th1 cell), and stimulate B cells to produce complement binding antibodies. IL-12, produced by macrophages, is the main stimulator of the Th1 cell response.
• Th2 cells stimulate B cells to produce a cytokine pattern that is dominated by IL-4, IL-5, IL-6, and IL-13. Antigen is presented to the Th2 cell by dendritic cells (Fig. 21.1-16 – Cell mediated immune response by the Th2 cell). Th2 cells promote the production of IgG antibodies, which bind hardly complement, and the production IgE. In an allergic reaction, the production of IgE antibodies is promoted by an IL-4 induced shift in the Th1/Th2 equilibrium in favor of Th2 cells.
• Cytokines secreted by Th cells modulate the immune response. For example, IFN-γ secretion by Th1 cells inhibits the immune response of Th2 cells and IL-10 secretion by Th2 cells inhibits the Th1 response by reducing macrophage function. The Th cell immune response is shown in Fig. 21.1-17 – Development and function of Th1 and Th2 cells.

Regulatory T cells (Treg)

Treg cells are CD4⁺CD25⁺T cells that are produced in the thymus and peripheral lymph organs and represent 5–10% of CD4⁺T cells in the peripheral blood and up to 20% in the bone marrow /22/. Treg cells have a normal α/β T cell receptor pattern and express the α-chain of the IL-2 receptor (CD25), cytotoxic T lymphocyte associated antigen 4 (CTLA-4), glucocorticoid-induced TNF receptor family related gene (GITR), and the transcriptional regulator Foxp3. This regulator acts as a master switch gene for Treg development and function. Treg cells have a high affinity for self peptides.

Treg functions are /22/:

• Maintenance of self tolerance by suppressing auto reactive T cells that are normally present in the periphery. Treg cells can also develop from normal CD4⁺T cells following exposure to an antigen.
• Down regulation of the immune response (refer to Fig. 20.2-2 – Cytokine activated transformation of naive T helper cells into different CD4⁺T-cell subtypes). Treg production is stimulated by IL-2.

21.1.8 Immune response in infections

Entry of a pathogen into the host initiates a multitude of interactions between soluble molecules (e. g, complement, C-reactive protein and antimicrobial peptides) and host sensors with pathogen derived molecules. The innate immune cells (e.g., neutrophils, macrophages, dendritic cells) express sensors and innate receptors known as pattern recognition receptors (PRRs). The PRRs are evolutionary conserved germ-line encoded receptors that sense pathogen derived signature molecules known as pathogen associated molecular patterns (PAMPs).

The PRRs include the following families /2/:

• Toll-like receptors (TLRs); essential in sensing bacteria
• NOD-like receptors (NLRs); important role in sensing bacteria
• RIG-I-like receptors (RLRs); important role in sensing viruses
• C-type lectin receptors (CLRs); essential for sensing mycobacteria and fungi
• DNA-sensing molecules; important role in sensing viruses.

The responses initiated by the innate immunity result in:

• Killing the pathogens or inhibition of their replication
• Initiation of the pathogen specific adaptive immunity through activation of B cells and T cells.

PRRs sense PAMPs in various compartments of the cells such as cytoplasm, cell surface and endocytotic vesicles.

21.1.8.1 Immune recognition of bacterial infection

According to the cell wall structure and composition bacteria are classified as Gram-positive and Gram-negative. The cell wall of Gram-positive bacteria are characterized by a thick peptidoglycan layer, the wall of Gram-negative bacteria contains a lipopolysaccharide (LPS) also known as endotoxin.

Innate immune defense

The innate immune system is well equipped to recognize and destroy bacteria through specialized defense cells (e.g., polymorphonuclear granulocytes, monocytes/macrophages and dendritic cells). These cells express genetically inherited receptors, called pattern recognition receptors (PRRs) for recognition of conserved pathogen associated molecular patterns (PAMPs). Signalling downstream from PRRs activates cellular responses, and killing mechanisms, and the expression of cytokines. The cytokines initiate inflammation and shape the adaptive immune responses /23/.

The peptidoglycan layer of bacteria is sensed by Toll-like receptor 2 (TLR2) of the defense cells. LPS, an immunopotent PAMP and virulence factor is sensed by TLR4. Both gram positive and gram negative bacteria contain a common ligand, the flagellin protein which is recognized by TLR5 /2/. Under normal conditions TLR2 is the major PRR involved in bacterial sensing and enhances the inflammatory response. However, this is not the case at low multiplicity of infection (MOI). At low MOI, TLR9 recognizes the bacteria by sensing bacterial DNA.

Immature dendritic cells (DCs) screen for bacteria entry using conserved PRRs and enhance the inflammatory response, which recognize PAMPs in microbial cell-wall components. These PRRs include the TLRs and C-type lectins for recognition. The TLRs relay infomation about the interacting bacteria to DCs through intracellular signaling cascades, thereby eliciting appropriate cellular processes that lead to DC maturation and the induction of inflammatory cytokines. The recognition of bacteria by C-type lectins leads to internalization of bacteria in DCs. Within the phagolysosome of DCs the bacteria target the C-type lectin DC-sign (DC-specific intercellular adhesion molecule-grabbing non integrin) and the processing for presentation by MHC class I and II molecules to T cells occurs. However, misuse DC-Sign by distinct mechanisms that either circumvent antigen processing or altered TLR-mediated signalling, skewing T-cell responses (e.g., mycobacterium, legionella, toxoplasma). This implies that adaption of bacteria to target DC-signal might support survival /23/.

Adaptive immune defense

The adaptive immune defense starts after processing of the bacterial components for presentation by MHC class I and II molecules to T cells.

21.1.8.2 Immune recognition of mycobacterium infection

M. tuberculosis infects immunocompromised individuals and children. The cell wall of M. tuberculosis consists of a mixture of polysaccharides and lipids with a high content of mycolic acid. M. tuberculosis is normally controlled, yet complete eradication of the mycobacterium does not occur. When the immune response is impaired, active disease can develop, normally through reactivation of quiescent mycobacteria or in some cases through re-infection. Refer to Section 42.12 – Infection with mycobacteria.

Innate immune defense

The adaptive immune response is as follows /23/:

• Mycobacteria are inducers of TH1-cell responses and the mycobacterial components stimulate the expression of co-stimulatory molecules and the production of IL-12 by dendritic cells (DCs) through Toll-like receptors 2 and 4 (TLR2 and TLR4). Invading M. tuberculosis is captured by macrophages/DCs and the mannose capped cell wall component lipoarabinomannan (ManLAM) is bound to the C-type lectin DC-Sign (DC-specific intercellular adhesion molecule-grabbing non integrin).
• Recognition of M. tuberculosis by TLRs expressed by DCs results in the activation of nuclear factor kappa B leading to the activation/maturation of DCs. Activation of DCs leads to the production of inflammatory cytokines.
• Increased secretion of ManLAM by infected macrophages/DCs targets DC-Sign and results in inhibitory signals that interfere with the TLR activating stimuli that lead to DC maturation. The ManLAM-DC- Sign interaction results in inhibition of DC maturation and induction of the immunosuppressive IL-10, thereby preventing an efficient cellular immune response against M. tuberculosis infection.

Adaptive immune defense

The adaptive immune defense starts after processing of of the bacterial components for presentation by MHC class I and II molecules to T-cells.

21.1.8.3 Immune recognition of viral infection

Viral molecules such as genomic DNA and RNA or double stranded RNA produced in viral infected cells are recognized by pattern recognition receptors (PRRs) expressed in innate immune cells of the host such as dendritic cells (DCs). PRR recognition depends on the detection viral envelope proteins and nucleic acid motifs within the DNA or RNA genomes of the virus (refer to Section 21.1.7.1 – Immune recognition of bacterial infection). Type I interferons (IFN-α and IFN-β) are under tight transcriptional regulation and are induced after recognition of viral components by various host PRRs /24, 25/.

21.1.8.4 Recognition of helminth infection

The surfaces of helminths as well as there excretory/secretory products are rich in glycoproteins /26/.

Innate immune defense

Recognition of these carbohydrate domains is mediated by the carbohydrate binding protein family of receptors (C-type lectins) that are expressed by cells of the innate immune system (macrophages, DCs, epithelial cells). Helminth-matured DCs have relatively immature status; they often express low levels of co-stimulatory molecules and pro-inflammatory cytokines.

Adaptive immune defense

C-type lectins, alarmins and interleukins initiate CD4⁺Th2 cell cytokine response. The patients have a high Th2 cell count, low Th1 cell count, and eosinophilia. The Th2 associated cytokines IL-4, IL-9, IL-13, IL-25 and IL-33 play important roles in mediating the effector mechanisms that contribute to worm expulsion such as golet cell hyperplasia and mucin production. IL-10 and IgG₄ concentrations are high whereas the increase in IgE is relatively slight /27/. Formation of granulomas occurs in patients with uncontrolled inflammatory responses, for example in schistosomiasis, where there is a strong immune response to the pathogen’s eggs that are embedded in the tissues. In these cases, there is a Th1 immune response with hepato­spleno­megaly and inflammation of the lymph nodes. The IgG₄ concentration is normal, while that of IgE is substantially increased. Resistant persons who remain free from helminth infections have a natural balance in the Th1/Th2 immune responses that kill invading helminths. In these cases, the ratio of IgG₄/IgE elevations is not shifted to the same extent described for the previous in favor of IgE

21.1.8.5 Recognition of fungal infection

The fungal cell wall is composed of carbohydrate polymers interspersed with glycoproteins. The three major components are polymers of glucose (β-glucans), polymer of N-acetylglucosamine (chitin), and mannans. The three components are intermingled throughout the cell wall, chitin tends to predominate near the plasma membrane, whereas the mannans have a propensity for the outer cell wall /28/. β-1,3 glucan forms the main structural scaffold of the cell wall. Many receptors in the tissues recognize β-glucans and three members of the scavenger receptor family, CD36, CD5, and SCARF1. The transmembrane receptor dentin of neutrophil granulocytes has a specificity for β-1,3 glucans. Engagement of PRRs leads to activation of signaling cascades which results in phagocytosis, respiratory burst and cytokine/chemokine gene induction. Fungi are activators of the complement system, resulting in opsonization due to deposition of C3b and iC3b on the fungal surface and activation of inflammatory cells as a result of C3a and C5a generation /29/.

21.1.9 Graft-versus-host disease

In allogenic transplantation of hematopoietic cells the most common life-threatening complication is graft-versus-host disease (GVHD) which occurs when immunocompetent T cells in the donated tissue (the graft) recognize the recipient (the host) as foreign. The resulting immune response activates donor T cells to gain cytolytic capacity and to then attack the recipient to eliminate foreign antigen-bearing cells. The two main clinical presentations are acute GVHD and chronic GVHD /30/.

In the early stage of HSCT, damaged donor tissue releases cytokines, leading to the development of a cytokine storm with increased release of adhesion molecules, co stimulatory molecules, MHC antigens, and chemokines. These danger signals activate the target tissues, including the antigen-presenting cells (APCs).

In the next step, T-cell receptors and co stimulatory molecules are activated and contact with APCs occurs. This results in allo reactive T-cell proliferation and differentiation. Activated T cells migrate to the GVHD target tissues (stomach, liver, skin, lung), where they recruit effector leukocytes (monocytes/macrophages, cytotoxic T cells, NK cells, granulocytes).

This results in destruction of the target tissue. In this effector stage, the T cell induced tissue destruction triggers an intensification of existing inflammation with further tissue injury.

References

1. Medzhitov R. Review article. Recognition of microorganisms and activation of the immune response. Nature 2007; 449: 819–26.

2. Kumar S, Ingle H, Prasad DV, Kumar H. Recognition of bacterial infection by innate immune sensors. Crit Rev Immunol 2013; 39. 229–46.

3. De Buck M, Gouwy M, Wang JM, Van Snick J, Opdenakker G, Struyf S, van Damme J. Structure and expression of different serum amyloid A (SAA) variants and their concentration-dependent functions during host insults. Current Medical Chemistry 2016; 23. 1725–55.

4. Brandzaeg P. Mucosal immunity: induction, dissemination, and effector functions. Scand J Immunol 2009; 70: 505–15.

5. Tosi MF. Innate immune responses to infection. J Allergy Clin Immunol 2005; 116: 241–9.

6. Takeda K, Akira S. Toll-like receptors in innate immunity. International Immunology 2005; 17: 1–14.

7. Delves PJ, Roitt IM. The immune system. N Engl J Med 2000; 343: 37–49.

8. Lewis JS, Lee JA, Underwood JCE, Harris AL, Lewis CE. Macrophage response to hypoxia: relevance to disease mechanisms. J Leukoc Biol 1999; 66: 889–900.

9. Robles DT, Eisenbarth GS. Immunology primer. Endocrinol Metab North Am 2002; 31: 261–82.

10. Stockwin LH, McGonagle D, Martin IG, Blair GE. Dendritic cells: Immunologic sentinels with a central role in health and disease. Immunology and Cell Biology 2000; 78: 91–102.

11. Nakamura MC. Natural killer cells and their role in disease. Laboratory Medicine 2002; 33: 278–82.

12. Rubin B. Natural immunity has significant impact on immune responses against cancer. Scand J Immunol 2009; 69: 275–90.

13. Stepanek O, Prabhakar AS, Osswald C, King CG, Bulek A, Naeher D, et al. Coreceptor scanning by T cell receptor provides a mechanism for T cell tolerance. Cell 2014; 159: 333–45.

14. Kohler S, Thiel A. Life after the thymus: CD31⁺ and CD31^{–^} human naive CD4⁺T-cell subsets. Blood 2009; 113: 769–74.

15. Jacobs R. Nicht einfach nur Killer: NK-Zellen sind auch wichtige Regulatorzellen. Immunologie Aktuell 2002; 2: 114–5.

16. Viau M, Zouali M. B-lymphocytes, innate immunity, and autoimmunity. Clin Immunol 2005; 114: 17–26.

17. Shapiro-Shelef M, Calame K. Regulation of plasma-cell development. Nature Rev Immunol 2005; 5: 230–42.

18. Doherty TM. T-cell regulation of macrophage function. Current Opinion in Immunology 1995; 7: 400–4.

19. McKay I, Rosen FS. The immune system. Second of two parts. N Engl J Med 2000; 343: 108–17.

20. Peggs KS, Allison JP. Co-stimulatory pathways in lymphocyte regulation: the immunoglobulin superfamily. Brit J Haematol 2005; 130: 809–24.

21. Mosmann TR, Sad S. The expanding universe of T-cell subsets: Th1, Th2 and more. Immunol Today 1996; 17: 138–46.

22. Chatila TA. Role of regulatory T cells in human diseases. J Allergy Clin Immunol 2005; 116; 949–60.

23. van Kooyk Y, Geitjenbeek TBH. DC-sign: escape mechanism for pathogens. Nature Reviews Immunol 2003; 3: 697–709.

24. Chen IY, Ichinohe T. Response of host inflammasomes to viral infection. Trends in Microbiology 2015; 23: 55–63.

25. Yoo JK, Kim TS, Hufford MM, Braciale JT. Viral infection of the lung: Host response and sequelae. J allergy Clin Immunol 2013; 132 (6). doi: 10.1016/j.jaci.2013.06.006.

26. Perrigoue JG, Marshall F, Artis D. On the hunt for helminths: innate immune cells in the recognition and response to helminth parasites. Cell Microbiol 2008; 10: 1757–64.

27. Maizels RM, Yazdanbakhsh M. Immune regulation by helminth parasites: cellular and molecular mechanisms. Nature Reviews Immunol 2003; 3: 733–44.

28. Levitz Sm. Innate recognition of fungal cell walls. PLOS pathogens 2010; 6: e1000758

29. Brown GD. Innate antifungal immunity: the key role of phagocytes. Annu Rev Immunol 2011; 29: 1–21.

30. Zeiser R, Blazar BR. Acute graft-versus-host disease – biologic process, prevention, and therapy. N Engl J Med 2017; 377: 2167–79.

31. Goverman JM. Regulatory T cells in multiple sclerosis. N Engl J Med 2021; 384 (6): 578–80.

32. Kooshesh KA, Foy BH, Sykes DB, Gustafsson K, Scadden DT. Health consequences of thymus removal in adults. N Engl J Med 2023; 389 (5): 406–17.

21.2 Immunodeficiency

Immunodeficiencies can be described as primary or secondary. While primary immune deficiencies are due to inherent dysfunction of the immune system and are chiefly genetic in etiology, secondary immune deficiencies are consequent to other underlying causes /1/.The number of patients with immunodeficiency is constantly increasing.

Reasons for this include:

• Demographic shift toward an aging population
• Improved tumor survival rates
• Expanding indications for and increased use of bone marrow transplantation
• Improved survival following organ transplantation (5-year survival following transplantation of solid abdominal organ is greater than 80%)
• Improved anti retroviral therapy for patients infected with HIV and greatly extended survival.

The result of all of this is an increasing number of infections with pathogens and/or courses (Tab. 21.2-1 – Typical pulmonary pathogens associated with different types of immunodeficiency).

21.2.1 Primary immunodeficiencies

Primary immunodeficiencies (PIDs) are caused by hereditary or genetic factors and more than 300 different defects have been described. The International Union of Immunological Societies Expert Committee for Primary Immunodeficiency /2/ has classified the PIDs into nine groups (Tab. 21.2-2 – Major primary immunodeficiency groups).These disorders may affect one or multiple components of the immune response, including T cells, B cells, natural killer (NK) cells, macrophages, dendritic cells, immunoglobulins and complement proteins.

PIDs can be categorized into T cell deficiencies, B cell deficiencies, phagocytosis defects, and complement deficiencies, based on the four compartments of the immune response. Most PIDs are monogenic and exhibit Mendelian inheritance. For example, Bruton’s disease affects only men since it is an X-linked disease. X-linked PID affects all men with the gene, and autosomal dominant or codominant primary immunodeficiency affects all of the progeny. However, the majority of primary immunodeficiencies are autosomal recessive and thus have a much lower penetrance /3/.

PIDs usually present in early childhood but may manifest not before adolescence and adulthood with an immune disorder that manifests as recurrent or persistent infections. The primary deficiency is often associated with autoimmune disease or the presence of autoantibodies /3/. An estimated 0.2% of the population have primary immunodeficiency and 3–5% have an autoimmune disease. The gap between the two may be explained by an underlying complete or incomplete primary or secondary immunodeficiency. PIDs with autoimmune predisposition are listed in Tab. 21.2-3 – Primary immunodeficiencies associated with a predisposition to autoimmunity and the prevalence of autoimmune diseases in selected PIDs is shown in Tab. 21.2-4 – Symptoms and clinical findings suggestive of immunodeficiency.

The overall incidence of PIDs (with the exception of secretory IgA deficiency) is estimated to be 1 in 10,000 individuals. Of the patients with PID /4/:

• Half have an antibody deficiency
• Some 20% have a combined T cell and B cell deficiency
• 10% have an isolated T cell deficiency
• 18% have phagocytosis defects
• Some 2% have a complement deficiency.

The individual incidences vary significantly, from 1 in 330 to 1 in 700 for secretory IgA deficiency to 1 in 500,000 for severe combined immunodeficiency. In children, males are affected 5 times more as females in the same age group, while the corresponding male to female ratio in adults is 1: 1.4.

21.2.1.1 T cell deficiency

Approximately 70–90% of peripheral lymphocytes are T cells, 5–10% are B cells, and 1–10% are natural killer cells (NK cells). Because the majority of peripheral lymphocytes are T cells, lymphopenia is most commonly due to a reduced number of T cells and is likely to be associated with immunodeficiency.

The antigen CD3 is carried by all T cells and is associated with the antigen receptor. Refer to Fig. 21.1-12 – Activation of the T cell receptor of CD4⁺ T helper cells.

Functional CD3 cells are:

• CD3⁺CD4⁺T cells, which produce cytokines following contact with an antigen. The cytokines are important for macrophage and B cell activation and antibody production.
• CD3⁺CD8⁺T cells, which are responsible for killing abnormal host cells (virus infected cells, malignant cells, cell mediated allogeneic graft).

T cell deficiencies are genetically heterogeneous, affect various components of adaptive immunity, and are due to a disorder in the development of the T cell repertoire.

These disorders can include:

• Lack of development of thymocytes or the environment required for their activation
• Abnormal peripheral T cells
• Disorder of signaling between T cells or between T cells and their environment
• Lack of co stimulation.

Approximately 10% of primary immunodeficiencies are caused by specific T cell immunodeficiencies, which are associated with increased susceptibility to infection by intracellular microorganisms such as Mycobacterium, Salmonella, Listeria, Toxoplasma, and viruses as well as fungal and protozoal infections /5/. On the other hand, microorganisms that are not usually pathogenic, such as the Mycobacterium vaccine strain (BCG) and infection by opportunistic pathogens such as Pneumocystis jirovecii can trigger a severe T cell immunodeficiency.

Patients with T cell deficiencies have an increased incidence of malignant tumors. This is due to several factors /6/:

• Reduced detection and destruction of free DNA due to a compromised immune response. The latter also promotes the development of malignant lymphoproliferative diseases.
• Reduced clearance of viruses such as the Epstein-Barr virus, Hepatitis B virus, Hepatitis C virus, Human T cell lymphotropic virus, Kaposi sarcoma associated virus, and Human papilloma virus. These viruses contribute to the immortalization and transformation of infected lymphocytes and are responsible for 10–15% of cancers worldwide.
• The inability to eliminate viruses also causes chronic inflammation with increased cell proliferation. This results in an increased risk that rapidly dividing cells will sustain oncogenic mutations.

A spectrum of mainly primary T cell deficiencies is shown in Tab. 21.2-5 – Primary (mainly) T cell immunodeficiencies.

21.2.1.2 B cell deficiency

Plasma cells that are capable of producing immunoglobulin develop during the final stage of B cell differentiation. They develop from B lymphocytes, which are derived from hematopoietic stem cells. The B cells undergo a series of differentiations with reassignment of the B cell receptor genes. This results in expression of the μ chain and the light chain (kappa or lambda) on the B cell surface to produce a naive B cell. The naive B cell leaves the bone marrow and migrates to the B cell pool. Contact with an antigen triggers further differentiation and leads ultimately to the secretion of immunoglobulins /7/. Following interaction with antigen-specific T cells, the B cells initially secrete IgM and then undergo a class switch to produce high-affinity IgG, IgA, or IgE antibodies (Fig. 21.1-9 – Class switch recombination following contact between B2 cell and antigen). The role of the initially produced IgM antibody in the circulation is to bind to invading pathogens and activate complement. High-affinity IgG, IgA, and IgE antibodies are produced through class switch. These protect the organism from further spread and reinfection by the pathogen.

21.2.2 Immunodeficiency categories

Some of the primary immunodeficiencies fit the criteria for more than one category.

21.2.2.1 Primary antibody deficiency

Primary antibody deficiency includes deficiencies in the production and function of individual Ig classes, Ig subclasses, and antibody specificities. They occur either in isolation or in combination. A deficiency is diagnosed based on the respective age specific reference intervals /9/. Primary antibody deficiency accounts for approximately 55% of primary immunodeficiencies. Although it can occur at any age, it is most prevalent in childhood and in the third decade of life.

Primary antibody deficiencies are a heterogeneous group of diseases with heterogeneous etiologies. They are classified in Tab. 21.2-3 – Primary immunodeficiencies associated with a predisposition to autoimmunity.

Patients with humoral immune deficiencies have increased susceptibility to infections with encapsulated bacteria such as Hemophilus influenzae type B and Streptococcus pneumoniae. Patients with B cell deficiencies usually begin having infections at the age of 7–9 months, when placental antibodies no longer provide immune protection. Viral and fungal infections are not usually a significant problem in patients with antibody deficiency, apart from patients with X-linked agammaglobulinemia (XLA), who are susceptible to infection with the Enterovirus, which can cause chronic encephalomyelitis. In addition, if antibody deficiency is diagnosed before the occurrence of organ injury (bronchiectasis, pneumonic foci), developmental disorders are not a feature /9/. Immunoglobulin substitution enables many patients to lead normal lives.

There are four distinct clinical entities that account for the majority of primary immunodeficiencies and three rare entities /10, 11/:

• Selective IgA deficiency
• IgG subclass deficiency
• Transient hypogammaglobulinemia in infancy
• Specific polysaccharide antibody deficiency
• Common variable immunodeficiency (CVID)
• X-linked agammaglobulinemia
• Hyper-IgM syndrome

Refer to Tab. 21.2-6 – Clinics and laboratory findings in primary antibody deficiency.

Algorithms for the differentiation of antibody deficiencies are shown in:

• Fig. 21.2-1 – Differentiation of antibody deficiency at normal immunoglobulin concentration
• Fig. 21.2-2 – Differentiation of antibody deficiency at low immunoglobulin concentration
• Fig. 21.2-3 – Differentiation of antibody deficiency with selective reduction of immunoglobulin classes
• Fig. 21.2-4 – Differentiation of antibody deficiency with selective increase in immunoglobulin classes.

Primary antibody disorders usually present 3–4 months after birth, once maternal immunoglobulin from placental transfer is gone. The babies present with recurrent or severe bacterial infections with encapsulated bacteria e.g., Streptococcus pneumoniae, and Haemophilus influenzae.

Antibody disorders are characterized by the presence or absence of B cells. When B cells are present, disorders are further characterized by whether B cells are of normal quality or quantity /11/. Refer to Tab. 21.2-7– B cell phenotyping in the diagnosis of B cell deficiency.

Agammaglobulinemia accounts for 13% of antibody disorders and for 84% with Bruton’s disease. Infants who have agammaglobulinemia are born with a complete absence of B cells and may have no tonsils or lymph nodes /11/.

Hypogammaglobulinemia is characterized of by low or deficient serum concentrations of any of the Immunoglobulin classes and subclasses. Common variable immunodeficiency (CVID) accounts for 46% of hypogammaglobulinemias and 82% of cases of primary antibody disorders involve a hypogammaglobulinemia /11/. IgA deficiency, IgG subclass deficiency, transient hypogammaglobulinemia and CVID are the most common types of primary hypogammaglobulinemia.

21.2.2.2 Combined immunodeficieny

Immunodeficiencies that affect both the T-cell and B cell systems are known as combined immunodeficiencies. They can occur as mild or as severe combined immunodeficiency syndrome (SCID). SCID is a heterogeneous group of congenital disorders associated with markedly reduction of T cells and variable amounts of B cells. Typically, patients presents early in life with failure to thrive, recurrent diarrhea, rashes. and serious bacterial, fungal and viral infections /1, 12/. Newborns are screened using an assay for determination of direct T cell receptor excisions circles (TRECs). TRECs are small circles of DNA created in T cells during their passage through the thymus as they rearrange their TCR genes.Their presence indicates maturation of T cells; TRECs are reduced in SCID /13/. Refer to Tab. 21.2-8– Frequency and inheritance of SCID.

21.2.2.3 Defects of the phagocytic system

Defects of the phagocytic system may be due to a defect in the function of macrophages and dendritic cells (e.g., defective migration and adhesion, or a lack of antimicrobial activity) /14/. Chronic granulomatous disease is the most common disorder /11/. It is characterized by pneumonia, abscesses, suppurative adnexitis, and gastrointestinal infections. Infections are related to the inability of the phagocytic system to kill catalase positive organisms including S. aureus, Burkholderia cepacia, Nocardia, Aspergillus, Serratia and Candida species. Patients are usually young and in some cases, the pathogen indicates the cause of the disease.

Refer to Tab. 21.2-9 – Primary phagocyte disorders.

21.2.2.4 Complement deficiencies

The actions of the complement system (opsonization, chemotaxis, destruction of bacteria via the classical, alternative, and lectin pathways) are mediated by the products of sequential complement activation and complement proteins. Complement deficiencies may be associated with severe bacterial infections. For more about complement deficiencies, refer to Chapter 24 – The complement system.

21.2.2.5 Diseases of immune deregulation

Primary immune deficiencies are often associated with autoimmune disease due to the deregulation of the immune system as a whole. In many immune deficiencies, lymphocytes may be present but dysfunctional, allowing for the development of excessive auto reactivity. Immune deregulation is commonly manifested as autoimmunity, cytopenias and inflammatory bowel disease. Clinical features not directly associated with immunodeficiency are prominent /15/. A wide range of organs may be affected. Autoimmunity is a hallmark of the clinical disease presentation. Cytokine/interleukin pathway defects involve mutations that can result in gain or loss of function.

Primary immune deficiencies with immune deregulation and/or autoimmunity /1, 16/:

• Autoimmune poly endocrinopathy candidiasis and ectodermal dystrophy (APECED)
• Autoimmune lyphoproliferative syndrome (ALPS)
• Familial hemophagocytic lymphohistiocytosis (FHL)
• Lymphoproliferative disorders associated with EBV /17/
• Immunodysregulation, polyendocrinopathy, enteropathy, and X-linked (IPEX)
• IPEX-like diseases
• IL-10/L-10 receptor deficiencies
• PLCG2-associted antibody deficiency and immune dysregulation

21.2.2.6 Defects in innate immunity

Diseases of innate immunity are disorders with a predispositin to viral and fungal infections. Included are Toll like receptor defects and natural killer cell defects. Disorders of innate immunity are /1/:

• NF-kappa B pathway defects, X-linked NEMO (online Mendelian inheritance in man; OMIM 300248 and autosomal gain of function mutations)
• Toll-like receptor signalling pathway deficiency
• Nk cell deficiency

Refer to Section 21.1.6.4– Antigen receptors.

21.2.2.7 Auto inflammatory disorders

Refer to Section 19.1.10 – Auto inflammation.

21.2.3 Secondary immunodeficiency

Secondary (acquired) immunodeficiencies occur when a healthy immune system of an individual is compromised by harmful influences. Acquired immunodeficiencies are secondary to diseases or therapy and play a far greater role than primary immunodeficiencies in numeral terms /18/. Viral infections (especially HIV), immunosuppressive therapies, malignancies, metabolic disorders, protein loss syndromes and poly trauma are the most prominent.

In transplant recipients receiving immunosuppressive medications, Cytomegalovirus infection or lymphoproliferative disease caused by the Epstein-Barr virus are indicators of severe immunosuppression. Fungal infections are associated with high morbidity and mortality in immunocompromised patients /19/.

Infections in association with low immunoglobulin levels in serum are relatively uncommon in secondary immunodeficiencies, with the exception of hypogammaglobulinemias, which occur in malignant disease, rarely due to medication, or in nephrotic syndrome. Reduced IgG levels occur in lymphoproliferative diseases and deficiencies of IgA and IgG occur in treatment with immunosuppressive, antirheumatic, or anticonvulsive medications. The dose and duration of treatment are important influence factors in treatment related antibody deficiencies /8/.

Refer to Tab. 21.2-10 – Secondary immunodeficiencies.

21.2.4 Evaluation of immunodeficiency

A congenital immunodeficiency is suggested by the symptoms listed in Tab. 21.2-4 – Symptoms and clinical findings of immunodeficiency /14/. The symptoms should prompt a thorough medical history and a step-by-step diagnostic strategy (Fig. 21.2-5 – Stepwise procedure for diagnosis of immunodeficiency).

21.2.4.1 Medical history

Regardless of the age of the patient, the medical history and clinical findings indicate possible immunodeficiency (Tab. 21.2-11 –Tests for suspected immunodeficiency) /12/:

• Increased susceptibility to infection or failure to thrive in neonates and infants. The spectrum of pathogens involved provides important clues. Infection with agents that are normally apathogenic or repeated severe infections with certain pathogens are particularly significant. Frequent viral and fungal infections may be due to an isolated T cell deficiency or combined T cell and B cell deficiency. Recurrent infections with bacteria may be due to a B cell, granulocyte, or complement deficiency or to common variable immunodeficiency syndrome (CVID).
• Signs of dysregulation of the immune system (e.g., granuloma, autoimmunity, recurring fever, lymphoproliferation, inflammatory bowel disease, unusual eczema). Refer to Tab. 21.2-12 – Prevalence of autoimmunity in primary autoimmune disorders.
• Conspicuous family history (e.g., infectious disease susceptibility, immunodeficiency, atopic disease, unknown causes of death)
• Malformations of the urinary and respiratory tracts, cystic fibrosis, ciliary dyskinesia, cerebrospinal fluid fistula, neurospora infection /18/
• HIV infection
• Malignant disease, particularly in older individuals. The B cell system is involved. Noticeably low immunoglobulin values are observed as a result of chronic lymphocytic leukemia, Hodgkin’s disease, and multiple myeloma.

21.2.4.2 Laboratory findings

Diagnosis is based on /20, 21/:

• Basic screening investigations
• Immunophenotyping of peripheral blood cells to prove the integrity of the immune system
• Functional in vitro assays to test the functionality
• Immunization with recall antigens to test the functionality of the immune system
• Molecular analysis.

Refer to Fig. 21.2-5 – Stepwise procedure for diagnosis of Immunodeficiency.

Basic screening analysis

• Complete blood cell count and differential
• Quantitative determination of IgG, IgA, IgM, IgE
• Possibly, determination of IgG subclasses
• Immunofixation electrophoresis in cases of suspected plasma cell dyscrasia in adults.

Immunophenotyping analysis

To discriminate among major lymphocyte populations, T cells, B cells, NK cells, and for the evaluation of the activation status of the immune system CD4⁺T cells and CD8⁺T cells the following cell surface markers are used /20/: CD45, CD3, CD4, CD8, CD19, CD16, CD56, and HLADR.

B cell deficiencies are more common than T cell deficiencies.

In decrease of the B cell count the differentiation of B cells is recommended Tab. 21.2-7 – B cell phenotyping in the diagnosis of B cell deficiency is recommended for further differentiation. The profile captures important B cell sub populations.

Immunophenotyping analysis assesses the count and percentage distribution of lymphocytes.

Functional in-vitro assays with antigens

After basic screening assays further workup in the diagnosis of primary immunodeficiency may need functional in-vivo assays or for testing the T cell dependent antibody response. Immunization with the recall antigen tetanus toxoid is used to test antibody response against proteins and pneumococcal vaccine is used to test the antibody response against polysaccarides (T cell independent antigen).

Genetic defects and presumed pathogenesis

For primary immune deficiency diseases refer to Reference /2/.

References

1. Raje N, Dinakar C. Overview of immunodeficiency disorders. Immunol Allergy Clin North Am 2015; 35: 599–623

2. Al-Herz W, Bousfiha A, Casanova JL, Chatila T, Conley ME, Cunningham-Rundles C, et al. Primary immunodeficiency diseases. An update on the classification from the international union of immunological societies expert committee for primary immunodeficiency. Front Immunol 2014; 5: article 162: 1–32.

3. Arason GJ, Jorgensen GH, Ludviksson BR. Primary immunodeficiency and autoimmunity: lessons from human diseases. Scand J Immunol 2010; 71: 317–28.

4. Noroski LM, Shearer WT. Screening for primary immunodeficiencies in the clinical laboratory. Clin Immunol Immunpathol 1998; 86: 237–45.

5. Edgar JDM. T cell immunodeficieny. J Clin Pathol 1998; 86: 988–93.

6. Rezaei N, Hedayat M, Aghamoammadi A, Nichols KE. Primary immunodeficiency diseases associated with increased susceptibility to viral infections and malignancies. J Allergy Immunol 2011; 127: 1329–41.

7. Wood P. Primary antibody deficiency syndromes. Ann Clin Biochem 2009; 46: 99–108.

8. Herriot R, Sewell WAC. Antibody deficiency. J Clin Pathol 2008; 61: 994–1000.

9. Dosanjh A. Chronic pediatric pulmonary disease and primary humoral antibody based immune disease. Respiratory Medicine 2011; 105: 511–4.

10. Lademenou F, Gaspar B. How to use immunoglobulin levels in ivestigating immune deficiencies. Arch Dis Child Educ Pract Ed 2016; 101: 129--35.

11. Reust CE, Evaluation of primary immunodeficiency disease in children. Am Fam Physician 2013; 87: 773–8.

12. Gaspar HB, Gilmour KC, Jones AM. Severe combined immunodeficiency – molecular pathogenesis and diagnosis. Arch Dis Child 2001; 84: 169–73.

13. Kwan A, Abraham RS, Cuurrier R, Brower A, Andruszewski K, Abbott JK, et al. Newborn screening for severe combined immunodeficiency in 11 screening programs in the United States. JAMA 2014; 312: 729–38.

14. Lekstrom-Himes JA, Gallin JI. Immunodeficiency diseases caused by defects in phagocytes. N Engl J Med 2000; 343: 1703–14.

15. Womg GK, Heather JM, Barmettler S, Cobbold M. Immune dysregulation in immunodeficiency disorders: The role of T cell receptor sequencing. Autoimmun 2017; 80: 1–9.

16. Lehman HK. Autoimmunity and immune dysregulation in primary immune deficiency disorders. Curr Allergy Asthma Rep 2015; 53. doi: 10.1007/s11882-015-0553-x.

17. Worth AJJ, Houldcroft C, Booth C. Severe Epstain-Barr virus infection in primary immunodeficieny and the normal host. Be J Haematol 2016; 175: 559–76.

18. Lehmann I, Borte M, Sack U. Diagnosis of immunodeficiencies. J Lab Med 2001; 25: 495–511.

19. Wahn V, von Bernuth H. Diagnostisches Vorgehen beim Verdacht auf einen primären Immundefekt (PID). J Lab Med 2009; 33: 179–84.

20. Hauck F, Bangol B, Rakhmanov M, Klein HG, Klein C. Rational laboratory diagnostics of primary immunodeficiency disorders. J Lab Med 2015; 39: 343–54.

21. Noroski LM, Shearer WT. Screening for primary immunodeficiencies in the clinical laboratory. Clin Immunol Immunpathol 1998; 86: 237–45.

22. Ballow M. Primary immunodeficiency disorders: antibody deficiency. J Allergy Clin Immunol 2002; 109: 581–91.

23. Hong R. The DiGeorge anomaly (CATCH 22, DiGeorge/velocardiofacial syndrome). Semin Hematol 1998; 35: 282–90

24. Conley ME, Notarangelo LD, Etzioni A. Diagnostic criteria for primary immunodeficiencies. Clinical Immunology 1999; 93: 190–7.

25. Fischer A, Notarangelo LD, Neven B, Cavazzana M, Puck JM. Severe combined immunodeficiencies and related disorders. Nature Rev Disease Primers 2015; article number 15061. doi: 10.1038/nrdp.2015.61.

26. di Santo JP. Severe combined immunodeficiency caused by defects in common cytokine receptor γc signaling pathways. Immunol Allerg N Am 2000; 20: 19–38.

27. Roifman CM, Dadi HD. Human interleukin-2 receptor α deficiency. Immunol Allerg Clin N Am 2000; 20; 39–50.

28. Zapata DA, Pacheco-Castro A, Torres PS, Millan R, Regueiro JR. CD3 immunodeficiencies. Immunol Allerg N Am 2000; 20: 1–8.

29. Picard C, Baud O, Fieschi C, Casanova JL. Diagnosis and management of heritable disorders of interferon-γ-mediated immunity. Immun Allerg N Am 2000; 20: 65–76.

30. Notarangelo LD, Candotti F. JAK3-deficient severe combined immunodeficiency. Immunol Allerg N Am 2000; 20: 97–111.

31. Elder ME. ZAP-70 and defects of T-cell receptor signaling. Semin Hematol 1998; 35: 310–20.

32. Sharfe N, Arpaia E, Roifman CM. CD8 lymphopenia caused by ZAP-70 deficiency. Immunol Allerg N Am 2000; 20: 77–95.

33. Bosticardo M, Marangoni F, Aiuti A, Villa A, Roncarola MG. Recent advances in understanding the pathophysiology of Wiskott-Aldrich syndrome. Blood 2009; 113: 6288–95.

34. Hershfield MS. Immunodeficiency caused by adenosine deaminase deficiency. Immunol Allerg N Am 2000; 20: 161–75.

35. Regueiro JR, Porras O, Lavin M, Gatti RA. Ataxia-telangiectasia. Immunol Allerg N Am 2000; 20: 177–206.

36. Ming JE, Stiehm ER, Graham JM. Genetic syndromes associated with immunodeficiency. Immunol Allerg N Am 2002; 22: 261–80.

37. Rezaei N, Hedayat M, Aghamoammadi A, Nichols KE. Primary immunodeficiency diseases associated with increased susceptibility to viral infections and malignancies. J Allergy Clin Immunol 2011; 127: 1329–41.

38. Gaspar HB, Gilmour KC, Jones AM. Severe combined immunodeficiency – molecular pathogenesis and diagnosis. Arch Dis Child 2001; 84: 169–73.

39. Gaspar HB, Kinnon C. X-linked agammaglobulinemia. Immunol Allerg N Am 2001; 21: 23–43.

40. Castigli E, Geha RS. Molecular basis of common variable immunodeficiency. J Allergy Clin Immunol 2006; 117: 740–6.

41. Urschel S, Kaykci L, Wintergerst U, Notheis G, Jansson M, Belohradsky B. Common variable immunodefi-ciency disorders in children: delayed diagnosis despite clinical presentation. J Pediatr 2009; 154: 888–94.

42. Geha RS, Rosen FS. The genetic basis of immunoglobulin class-switching. N Engl J Med 1994; 330: 1008–12.

43. Bonilla FA, Geha RS. CD154 deficiency and related syndromes. Immunol Allerg N Am 2001; 21: 65–89.

44. Rangel-Santos A, Wakim VL, Jacob CM, Pastorino AC, Cunha JM, Collanieri AC, et al. Molecular characterization of patients with X-linked Hyper-IgM syndrome: description of two novel CD40L mutations. Scand J Immunol 2009; 69: 169–73.

45. Perez E, Bonilla FA, Orange JS, Ballow M. Specific antibody deficiency: controversies in diagnosis and management. Frontiers in Immunology 2017; 8: article 586.

46. Geha RS, Rosen FS. The genetic basis of immunoglobulin class-switching. N Engl J Med 1994; 330: 1008–12.

47. Wehr C, Kivioja T, Schmitt C, Ferry B, Witte T, Eren E, et al. The EUROclass trial: defining subgroups in common variable immunodeficiency. Blood 2008; 111: 77–85.

48. Lekstrom-Himes JA, Gallin JI. Immunodeficiency diseases caused by defects in phagocytes. N Engl J Med 2000; 343: 1703–14.

49. Lanza F. Clinical manifestation of myeloperoxidase deficiency. J Mol Med 1998; 76: 676–81.

50. Planella T, Cortes M, Martinez-Bru C, Barrio J, Sambeat M, Gonzalez-Sastre F. The predictive value of several markers in the progression to acquired immunodeficiency syndrome. Clin Chem Lab Med 1998; 36: 169–73.

51. Jaffe EF, Lejtenyi C, Noya FJD, Macer BD. Secondary hypogammaglobulinemia. Immunol Allerg N Am 2001; 21: 141–63.

52. Barnes PJ. Corticosteroids. In: Samson AP, Church MK, eds. Anti-inflammatory drugs in asthma. Basel; Birkhäuser 1999: 35–85.

53. Posey WC, Nelson H, Pearlman DS. The effect of acute corticosteroid therapy for asthma on serum immunoglobulin levels. J Allergy Clin Immunol 1978; 62: 640–5.

54. Hamilos DL, Young RM, Peter JR, et al. Hypogammaglobulinemia in asthmatic patients. Annals of Allergy 1992; 68: 472–6.

55. Farr M, Kitas GD, Tunn EJ, et al. Immunodeficiencies associated with sulfasalazine therapy in inflammatory arthritis. Br J Rheumatol 1991; 30: 413–7.

56. Matthias R, Stecklein H, Guay-Wooford L, et al. Gamma globulin deficiency in newborns with congenital nephrotic syndrome. N Engl J Med 1989; 320: 389–91.

57. Ogi M, Yogoyama H, Tomosugi N, et al. Risk factors for infection and immunoglobulin replacement therapy in adult nephrotic syndrome. Am J Kidney Dis 1994; 24: 427–31.

58. Boughton BJ, Jackson N, Lim S, et al. Randomized trial of intravenous immunoglobulin prophylaxis for patients with chronic lymphocytic leukemia and secondary hypogammaglobulinemia. Clin Lab Haematol 1995; 17: 75–80.

59. Walchner M, Wick M. Elevation of CD8⁺ CD11b⁺ Leu8-T-cells is associated with the humoral immunodeficiency in myeloma patients. Clin Exp Immunol 1997; 109: 310–5.

60. Geaghan SM. Hematologic values and appearances in the healthy fetus, neonate, and child. Clin Lab Med 1999; 19: 1–37.

61. Gossage DL, Buckley RH. Prevalence of lymphocytopenia in severe combined immunodeficiency. N Engl J Med 1990; 323: 1422–3.

62. Sorensen RU, Moore C. Immunology in the pediatrician’s office. Pediatr Clin North Am 1994; 41: 691–714.

63. Maguire GA, Kumaratatne DS, Joyce HJ. Are there any clinical indications for measuring IgG subclasses? Ann Clin Biochem 2002; 39: 374–7.

64. Pacheo SE, Shearer WT. Laboratory aspects of immunology. Pediatr Clin North Am 1994; 41: 623–55.

65. Shearer WT, Paul ME, Smith CW, Huston DP. Laboratory assessment of immune deficiency disorders. Immunol Allerg Clin North Am 1994; 14: 265–99.

66. Cohen A, Grunebaum E, Arpaia E, Roifman CM. Immunodeficiency caused by purine nucleoside phosphorylase deficiency. Immunol Allerg N Am 2000; 20: 143–59.