eMedicine Specialties > Pediatrics: General Medicine > Hematology

White Blood Cell Function

Author: Tiffany Chang, MD, Staff Physician, Tri-Campus Pediatric Residency Program, Mattel Children's Hospital, Cedars Sinai Medical Center, Olive View Medical Center
Coauthor(s): Pamela H Kempert, MD, Assistant Clinical Professor, Division of Hematology/Oncology, David Geffen School of Medicine at University of California at Los Angeles, Mattel Children's Hospital and UCLA Medical Center; Kathleen Sakamoto, MD, Professor, Department of Pediatrics, Division of Hematology-Oncology and Pathology and Laboratory Medicine, Mattel Children's Hospital, David Geffen School of Medicine, University of California at Los Angeles; David Buchbinder, MD, Fellow, Department of Pediatrics, Division of Hematology-Oncology, Mattel Children's Hospital, David Geffen School of Medicine, University of California at Los Angeles; Anne E Hagey, MD, Global Project Head, Department of Oncology Cytotoxics, Abbott Laboratories
Contributor Information and Disclosures

Updated: Apr 11, 2008

Overview of the Immune System

The immunopathogenesis of many human diseases is characterized at the molecular level. Therefore, a basic understanding of immune function is often useful. Specific manipulation of the immune system for therapeutic purposes is now possible.

Types of immunity

The 2 recognized types of immunity are innate and adaptive.

Innate immunity is relatively nonspecific. It is the body's first-line defense against many bacterial pathogens. Innate immunity resides in the skin, mucous membranes, polymorphonuclear (PMN) cells, complement system, and a select group of cells that possess cytotoxic capabilities.

The skin and mucous membranes act as physical barriers to invading microorganisms. PMN cells (ie, granulocytes, monocytes, macrophages) primarily have a phagocytic function. Granulocytes are mobile phagocytes that travel to areas of inflammation to engulf and destroy invading microorganisms. They are relatively indiscriminate in their function. Monocytes circulate, whereas macrophages are fixed in lymphoid and mucosal tissues. They can also phagocytose foreign microorganisms. Binding of complement to a foreign substance, or antigen, amplifies and augments the body's innate immune system by means of its role as an opsonin (a factor that enhances phagocytosis of unwanted particles) and as a chemoattractant (a factor that recruits cells to areas of inflammation). Natural-killer (NK) cells are specialized lymphocytes that have cytotoxic properties in addition to their ability to produce cytokines that assist in the orchestration of adaptive immunity.

In contrast to basic innate immunity, adaptive immunity is specific and depends on antigenic stimulation. Antigens are foreign substances that evoke an immune response. They can take on many different forms, including proteins, lipids, or carbohydrates. The generation of receptors specific for antigens is a unique and complex process that generates 1012 specific receptors for each cell type of the adaptive immune system, including T and B cells.

After a complex process of education and maturation, a circulating lymphocyte can bind to an antigen. Various cell types can process and present these antigens to T cells, or antigens may be soluble and bound to B-cell receptors. Cell-to-cell interactions set off a cascade of events that may result in T- or B-cell activation and, ultimately, host defense. The adaptive immune system consists of 2 types of lymphocytes: T cells (70-75% of the adaptive immune force) and B cells (10-20% of the adaptive immune force). NK cells are specialized lymphocytes and are considered in the context of the innate immune system.

Immunodeficiency diseases

The table below provides a brief overview of several immunodeficiency diseases. The normal functions of the immune system and the disorders that affect it are considered in detail below.

Innate Immune System

Key components

Granulocytes are a key component of the innate immune system (ie, nonspecific immune defense system). The granulocyte network includes 3 main components: neutrophils, eosinophils, and basophils. This network makes up 50% of the body's circulating WBCs. Other cellular components of the innate immune system include mononuclear phagocytes, dendritic cells, and NK cells.

Neutrophils are the first-line defense the body has upon invasion by a foreign microorganism. Neutrophils move to the site of invasion by means of chemotaxis, which occurs in response to microbial products, activated complement proteins, and cytokines. Chemotaxis of neutrophils involves movement of pseudopodia and polymerization of cytoskeletal proteins or actin. The cell may then ingest the foreign invader. PMNs, monocytes, and eosinophils can participate in phagocytosis. Opsonins are often antibodies or components of the complement pathway that bind to the surface of target organisms to facilitate this phagocytosis. 

Myelopoiesis

The neutrophil maturation and differentiation pathway termed myelopoiesis takes approximately 12 days. Neutrophils derive from a pluripotent myeloblast, or stem cell, which expresses the cell markers CD34 and human leukocyte antigen (HLA)-DR. This stem cell can give rise to colony-forming unit (CFU)–granulocyte, erythrocyte, megakaryocyte, and macrophage (CFU-GEMM) cells that express CD33, CD34, and HLA-DR. Some progeny of these cells become CFU–granulocyte-macrophage (CFU-GM) cells that express CD13, CD33, CD34, and HLA-DR. These cells give rise to CFU–granulocyte (CFU-G) cells that express CD13, CD33, CD15, and HLA-DR. Myeloblast formation marks the next stage in myelopoiesis. Myeloblasts express the cell surface markers CD13, CD33, and CD38 and have few or no granules. The myeloblast gives rise to the promyelocyte.

The promyelocytic stage is characterized by the appearance of azurophilic (bluish) granules called primary granules and by fewer mitochondria than observed in its myeloblast precursor. This change indicates a heavily anaerobic metabolism. Primary granules contain myeloperoxidase, arginine-rich (cationic) proteins, lysozymes with a bactericidal function, sulfated mucopolysaccharides, acid phosphatases, proteases, and hydrolases, among other contents. Microperoxisomes that contain catalase are also present at the promyelocytic stage of development.

The myelocyte represents the next stage. This cell is smaller and rounder than its predecessor, with a smaller Golgi apparatus and endoplasmic reticulum. Glycogen appears at this stage and serves as a glucose store that the hexose monophosphate shunt can directly oxidize. These changes indicate an increase in the amount of cellular anaerobic metabolism. Secondary granules appear at this stage of development. Contents of secondary granules include collagenase and lysozyme. The secondary granules do not contain peroxidase. The production of primary granules ceases during the myelocyte stage of development.

Appearance of the metamyelocyte marks the next step in myelopoiesis. The metamyelocyte is a nonproliferative cell. It has many more secondary granules than primary granules have. At the final stages of neutrophil development, the metamyelocyte becomes a band neutrophil and then a mature neutrophil.

Types of granules

The stages of neutrophil differentiation may be characterized by the type of granules present and their composition. Primary granules appear at the transition between the myeloblast stage of development and the promyelocyte stage of development. The appearance of secondary granules occurs at the transition from the promyelocyte to the myelocyte stage of development. This is associated with the commitment of the cell to a neutrophil lineage, as opposed to an eosinophil or a basophil lineage. The contents of the secondary granules also change during development. Myelocyte secondary granules have lactoferrin and no gelatinases. Metamyelocytes have both lactoferrin and gelatinases. Bands and mature neutrophils have gelatinases and no lactoferrin.

Mature neutrophils, eosinophils, basophils, mononuclear phagocytes, and natural killer cells

When the neutrophils reach a mature stage, they are released into the circulation under the influence of granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF). Once in the circulation, neutrophils can migrate from the circulating pool of neutrophils into the marginating pool of neutrophils. The circulating pool captures approximately 5% of the body's neutrophils (reflected by the peripheral WBC count), although 10-15% remain adherent to the endothelium as the marginating pool. The remaining 85% reside in the bone marrow. Neutrophils circulate for 6-12 hours in the bloodstream before irreversibly infiltrating tissues and completing their life cycle 24 hours later.

Eosinophils also develop from the promyelocyte. They contain approximately 1012 nonazurophilic (pinkish) granules, which are primarily composed of peroxidase and acid phosphatases. They have a developmental period of 3-6 days and a half-life of 30 minutes after they arrive in the circulation. In tissues, they are present for about 8-12 days. Eosinophils participate in less phagocytosis than neutrophils do but can promote inflammation and assist in host defense.

Basophils evolve in the bone marrow from a common progenitor to eosinophils and basophils. Basophils have granules which stain deep blue or purple with basic dyes. They may resemble mast cells, although mast cells evolve and mature in the tissue and not the bone marrow and arise from a different progenitor. Basophils infiltrate tissues, secrete cytokines, and have roles in immunoglobulin (Ig) (specifically IgE) synthesis and in B-cell signaling.

The mononuclear phagocytes serve a host of functions. They can phagocytose, secrete mediators of inflammation, process antigens and interact with lymphocytes, perform cytotoxic activities, and perform specialized functions dictated by the tissue location of the mononuclear phagocyte. Dendritic cells arise from the same bone marrow progenitor as the mononuclear phagocytes, although they have poor phagocytic activity and primarily serve as antigen-presenting cells to lymphocytes.

NK cells are specialized lymphocytes derived from the bone marrow that possess cytotoxic capabilities. They secrete cytokines and can interface with the adaptive immune system. These cells contain cytoplasmic perforin and granzymes granules, which mediate the killing of target cells. NK cells may also recognize targets in the context of decreased or absent expression of major histocompatability complex (MHC) I molecules. This is a common tool some viruses and tumor cells use to evade detection. NK cells express the cell-surface markers CD56 and CD16. CD16 is a low affinity IgG receptor that may be used in antibody-dependent cell-mediated cytotoxicity (ADCC). ADCC is a process in which CD16 is triggered by IgG bound to a target cell.

Molecular and genetic factors

Transcription factors drive patterns of hematopoietic progenitor cell genetic expression and direct lineage commitment. In particular, PU.1 and CCAAT/enhancer-binding protein alpha (C/EBP alpha) play key roles in the regulation of normal and abnormal myeloid gene expression. PU.1, a member of the Ets family of proteins, is responsible for myeloid-specific gene expression. PU.1 expression markedly increases during myeloid differentiation, and most myeloid-specific promoters possess binding sites for PU.1.

Murine models that lack PU.1 expression have demonstrated PU.1–dependent expression of genes that encode select cytokine receptors, integrins, and granule components. Murine models have confirmed that loss of PU.1 disrupts myeloid development. C/EBP alpha is a leucine-zipper transcription factor that is important in neutrophil development. Expression of C/EBP alpha increases during early myeloid development; C/EBP alpha also activates critical myeloid genes. In murine models that lack C/EBP alpha expression, GCSF levels, interleukin (IL)-6 receptor levels, and some cytokine levels are decreased. Murine models with a loss of C/EBP alpha lack mature granulocytes.

Transcription factors play a critical role in normal and aberrant myelopoiesis. One well-characterized example is the role of retinoic acid receptors in the development of acute promyelocytic leukemia. PU.1 and C/EBP alpha have also been implicated in leukemogenesis. Mutations in PU.1 were found in 9 of 126 patients with acute myeloid leukemia (AML).¹ The frequency of C/EBP alpha mutations is 7-11% in patients with AML.

AML1 protein, which has an established role in the expression of early myeloid genes and neutrophil differentiation, is involved in the t(8;21) translocation observed in AML. This translocation fuses AML1 to the ETO gene. Normal AML1 cooperates with both PU.1 and C/EBP alpha to regulate the expression of myeloid specific genes. This fusion product disrupts the function of PU.1 and C/EBP alpha.

Functional capabilities of neutrophils

Consideration of neutrophil development from a functional viewpoint highlights the work of Glasser and Fiederlein (1987).² They described a distinct order to the functional capabilities that neutrophils acquire. The functional differentiation of the human neutrophil is characterized by the initial appearance of Fc receptors, followed by the ability to perform phagocytosis and respond to complement by acquiring and displaying appropriate receptors. Fc receptors and complement receptors (CRs) promote neutrophilic phagocytic function, which is why their appearance coincides with the ability to phagocytose. CR1 and CR3 react with C3 opsonins. CR1 and CR3 appear later than the Fc receptors, which are expressed at the myelocyte-metamyelocyte stage of development. CR1 and CR3 then work synergistically with Fc receptors to promote phagocytosis.

The capacity for oxygen-independent microbial killing appears next and is followed by the ability of the neutrophil to engage in oxygen-dependent microbial killing and, finally, by its ability to take part in chemotaxis. Promyelocytes have no oxidative killing ability, and their oxygen-independent bactericidal mechanisms are inoperative despite the possession of primary granules. Oxygen-dependent bacterial killing is a late manifestation of functional differentiation, as demonstrated by the fact that only band and segmented neutrophils have a substantial respiratory burst.

Chemotaxis is a late manifestation of functional differentiation. Random motility is expressed in the absence of chemotaxis in immature myeloid cells, but further maturation of the cell is necessary for directed chemotaxis. Chemoattractant cytokines called chemokines assist in the orchestration of chemotaxis of mature neutrophils.

Neutrophilic response

A neutrophilic response is initiated when circulating neutrophils flowing through the postcapillary venule experience low chemokine levels. They also detect other factors released at a site of infection. This milieu triggers changes in the surface molecules on endothelial cells and neutrophils, which cause these cells to associate. The initial associations have low affinity, are reversible, and are mediated by L selectins on neutrophils and are mediated by E and P selectins on endothelial cells. These loose associations are responsible for the process known as leukocyte rolling, in which connections are made and then repeatedly broken, causing the neutrophil to roll along the surface of the endothelium. This process exposes the neutrophil to many activating factors.

Activating factors lead to the induction of quantitative and qualitative changes in integrins, which mediate neutrophil adhesion. Activated integrin complexes mediate adhesion between neutrophils and endothelial cells by using intercellular adhesion molecule-1 (ICAM-1), which is located on endothelial cells. Neutrophils also adhere to other neutrophils. Neutrophil-to-neutrophil adhesion and neutrophil-to-platelet aggregates begin to occlude the venule and reduce blood flow.

The next phase involves loosening of the integrin-mediated adhesion. This accompanies further conformational changes that allow migration of the neutrophil between endothelial cell junctions and into the extravascular tissue. This process is called diapedesis. Once through the endothelium, neutrophils sense the chemotactic gradient and migrate to the site of infection. The migration of neutrophils involves a complex signal-transduction cascade, ending in remodeling of the actin-based cytoskeleton in these cells. Granule secretion occurs at this time, releasing heparinase, gelatinase, and other enzymes that aid neutrophil transit through the basement membrane and connective tissue.

IgG and cleaved forms of C3 act as opsonins. At the site of infection, neutrophils adhere to pathogens by means of their Fc receptor and CR. Internalization or endocytosis of the microorganism by the neutrophil generates a phagosome or phagocytic vesicle. Membranes of the neutrophil granule fuse with the phagosome membrane, delivering a wide variety of potent antimicrobial proteins to the phagosome that contains the target microorganism. Neutrophils contain azurophilic or primary granules and specific basophilic or secondary granules. Contents of both granules are secreted into the phagosome.

The assembly and activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex occurs at the phagosome membrane. This enzyme complex generates large amounts of superoxide, which combines with water to produce hydrogen peroxide. Other oxidants are also formed, including hydroxyl radical and peroxynitrite. Inducible nitric oxide synthase manufactures the nitric oxide precursor to the formation of peroxynitrite. In the presence of myeloperoxidase from the azurophilic granules, a reaction is catalyzed that uses hydrogen peroxide and chloride anion to create hypochlorous acid in the phagosome. These oxidants denature proteins, enabling proteolysis of the foreign particles.

The oxidants also activate proteases in the neutrophil granules. The reactive oxygen species formed may also act as signaling molecules that eventually lead to apoptosis of target cells. These events enhance breakdown and clearance of pathogens from the site of infection. Oxidants may also help terminate neutrophil influx by inactivating inflammatory mediators.

Toll-like receptors

Toll-like receptors (TLR) play a critical role as pattern recognition receptors.³ They bind molecules derived from infectious and noninfectious agents. They are expressed on antigen-presenting cells, such as macrophages and dendritic cells. Their signal activates antigen-presenting cells to provoke innate immunity and to establish adaptive immunity. Each TLR has common effects, such as production of inflammatory cytokines or upregulation of costimulatory molecule expression, but each also has specific functions, such as inducing type 1 interferon The TLR signal activates antigen-presenting cells to support T helper 1 (TH1) cell differentiation. The main function of TLR is induction of inflammation and the establishment of adaptive immunity. TLR-activated antigen-presenting cells stimulate T cells. TH1 cells produce interferon-g and mediate antiviral and antibacterial immunity. 

Complement system

The complement system is composed of more than 30 proteins encoded throughout the genome with important clusters of genes on chromosome arm 1q and in the MHC region on 6p. These proteins make up approximately 4% of the serum proteins of the body. These serum proteins amplify the innate immune response and interface with the adaptive immune system by initiating the complement cascade. A host of complement pathway regulators control the pathways of complement activation.

The activation of the complement system may result in the lysis of targets by the creation of an active membrane-lysing complex. Activation of the terminal components of the complement cascade generates a membrane-attack complex that creates a pore in the target cell membrane. This pore allows for the exchange of cellular contents intended for direct killing of selected pathogens. The complement system may also result in the release of inflammatory mediators and chemoattractants, which bring other neutrophils to the site of invasion. It is also involved in the coating of pathogens by generating opsonins. Opsonization involves the binding and activation of C3. When activated, C3 acts as the bridge between the microbe and any phagocytic cell that encounters it. Complement is also involved in other activities that assist in maintaining immunologic homeostasis and host defense. In addition to its role in host defense, the complement system can orchestrate anaphylactoid-type reactions.

A multitude of pathways may be invoked to activate of the complement system. The classic pathway requires the adaptive immune system because it requires antibody. The alternative pathway is activated in multiple ways but does not require antibody for activation. A third pathway is called the mannan-binding lectin pathway. This last pathway requires the binding of mannan-binding lectin to specific polysaccharides, which subsequently activates proteases and the complement system.

Adaptive Immune System

Key components and antigen-receptor interaction

The adaptive immune system is composed of T and B lymphocytes. Lymphocytes constitute 40% of circulating WBCs. B cells make up 10-20% of lymphocytes, T cells account for 70-75%, and NK cells comprise 10-15%. They are considered in the context of the innate immune system above.

The cornerstone of adaptive immunity is antigen-receptor interaction. The B cell expresses surface antibody or Ig, which serves as an antigen receptor. T cells express antigen receptors. Genes for Ig and T-cell receptor (TCR) are rearranged in developing B and T cells, respectively. The variable (V), diversity (D), and junctional (J) gene recombinations are responsible for the nearly 10¹² specific receptors generated for each cell type. Recombination events of gene segments and other processes, such as the addition of nucleotides at splicing junctions and somatic hypermutation (in B-cell development), aid in diversification. The receptors created are specific for particular antigens.

B cells and their products constitute one arm of the adaptive immune system. B cells arise from hematopoietic stem-cell precursors in the bone marrow and must undergo 2 phases of maturation: an antigen-independent phase and an antigen-dependent phase. A common lymphoid progenitor gives rise to progenitor B cells (proB cells), which are the earliest identifiable cells committed to development in the B-cell lineage. These proB cells rearrange their Ig heavy-chain genetic segments to create a functional IgH gene that is then expressed as a pre–B-cell receptor.

Once a functional pre–B-cell receptor is present, the Ig light-chain genetic segments can rearrange and be expressed with the already rearranged heavy chain. This immature B cell may then populate secondary lymphoid organs and undergo antigen-dependent maturation, which requires antigen encounter and T-cell support. The result is the creation of Ig-secreting plasma cells or memory B cells. B cells express CD19, CD20, and CD21. Of note, CD21 is the receptor for the Epstein-Barr virus, which enhances B-cell proliferation in vivo and immortalization in vitro.

An environmental milieu composed of activated CD4+ T cells and cytokines regulates class-switch recombination and isotype switching. The serum Igs produced make up 30% of serum proteins. The antibodies that B cells manufacture function as opsonins and antitoxins. They also neutralize viral pathogens, agglutinate and lyse bacteria, and participate in ADCC.

Types and subtypes of Ig

Five types of Igs are recognized: IgG, IgM, IgA, IgD, and IgE. IgG is the most abundant antibody class. It has antitoxin, antiviral, and antibacterial functions. It makes up 80% of the total Ig concentration and is evenly divided between extravascular tissues and the circulation. IgG has a half-life of 20-25 days. It is the primary component of intravenous (IV) Ig (IVIG) used clinically. IgG crosses the placenta and has 4 subclasses: IgG1, IgG2, IgG3, and IgG4. IgG is involved in little direct killing of targets but does activate the complement system and is involved in opsonization and ADCC. The presence of IgG after an initial response to antigen is associated with immunologic memory.

IgM is the first antibody formed after antigen stimulation. It is present as a pentamer with a joining, or J, chain. It makes up 5% of the total Ig and is mostly found in the circulation. It functions as an antipolysaccharide, an opsonin, which is an activator of complement and rheumatoid factor. IgM plays a key role in host defense demonstrated by the fact that deficiency may result in the rapid development of sepsis.

IgA has 2 subclasses (IgA1 and IgA2) and makes up 15% of the serum Ig, with 40% of the total amount of IgA present in the circulation. Secretory IgA is dimeric and is joined by a J chain. It is predominantly IgA2, whereas serum IgA is monomeric and predominantly IgA1. The major sites of production are the GI, respiratory, and genitourinary tracts. Secretory IgA can resist digestion. It is the key immunologic component of the mucosal immunity and guards against sinopulmonary, GI, and genitourinary infections. A lack of IgA is the most common antibody deficiency and occurs in about 1 per 400 individuals. Anaphylaxis can occur when a person with IgA deficiency is given exogenous Ig. This occurs because of antiIgA antibodies, which a person with an absence of IgA can manufacture. Low-IgA Ig products are available for use in patients with IgA deficiency.

IgD is present in trace quantities in the serum. It is present in large quantities bound to the B cells of a fetus or newborn. This finding is consistent with its role as an antigen receptor in B-cell development.

IgE triggers immediate hypersensitivity reactions by binding to basophils and mast cells. Allergens may also combine with IgE to activate cells. It is also believed to play a protective role in the defense against intestinal parasites.

Neonatal antibody system

The neonatal antibody system is composed mainly of B cells. Plasma cells are rare, and antibody responses are not well developed. Neonatal Igs largely consist of maternal IgG acquired by their transfer across the placenta during the third trimester by means of Fc receptors on placental cells. These maternal antibodies disappear in the first few months of life due to normal catabolism, resulting in a physiologic hypogammaglobulinemia by age 3-5 months. This passive transfer of immunity is helpful for protection against viral pathogens and bacterial pathogens that infants encounter due to deficient opsonic antibody.

Secretory IgA received in breast milk is also protective for neonates. In addition to their protective role, maternal antibodies can also cause disease, such as in neonatal lupus, isoimmune thrombocytopenic purpura, and hemolytic disease of the newborn. Neonates do not receive maternal IgM, IgA, or IgE because they do not cross the placenta; therefore, these antibodies are relatively absent until the babies' own production of antibodies becomes adequate. Active fetal antibody production begins shortly before birth with low levels of IgM followed by IgG and IgA production. As a result of the sequence and timing of the development of the antibody milieu, children have a poor response to polysaccharide antigens until age 2 years.

B cells

B-cell development occurs in the bone marrow and in the peripheral lymphoid organs. B-cell commitment and differentiation is a complex process that requires the careful regulation of gene expression and transcriptional activity. One protein deemed critical in the early stages lymphoid development is the zinc finger protein Ikaros. Murine models in which Ikaros is disrupted result in blockage of B-cell development.

Murine models lacking E2A, another transcription factor, have impaired lymphoid development and lack B-lineage cells. E2A is also implicated in leukemogenesis; for example, E2A is sometimes found as part of a fusion protein coupled to the homeobox transcription factor PBX in preB-cell leukemia.

Intracellular signaling is also required for normal B-cell development. For example, absence of the intracellular protein Bruton tyrosine kinase (BTK) results in the phenotype of X-linked agammaglobulinemia, or Bruton agammaglobulinemia. B-cell differentiation in the bone marrow is blocked in individuals with this defect and results in a lack of circulating B cells and antibody production. Another X-linked primary immunodeficiency, X-linked hyper-IgM syndrome, results from a lack of the T-cell surface molecule CD40 ligand, which normally binds CD40 on B cells to induce their class-switch recombination. The absence of CD40 ligand results in excess IgM and a failure to produce IgG, IgA, and IgE.

T cells

Fetal lymphoid progenitors migrate to the thymus, where they undergo unique and necessary interactions that are critical to their development and maturation. T cells reside in the thymic microarchitecture, in which only about 20% survive as mature functional T cells. The other 80% die in situ during their maturation. During T-cell differentiation, segments of the TCR gene undergo recombination to allow for necessary diversification. Prothymocytes (CD3^-, CD4^-, and CD8^-) begin at the subcapsular domain and become double-positive thymocytes (CD4⁺ and CD8⁺) and then single-positive thymocytes (CD4⁺ or CD8⁺).

During this process, they also undergo positive selection for thymocytes with TCRs that can interact with self MHC molecules and negative selection, which induces apoptosis in thymocytes that are strongly self-reactive. In the medulla of the thymus, mature T cells emerge and then populate peripheral lymphoid organs.

One factor embedded in the thymic microarchitecture and critical to the induction of T-cell development is the Notch system. Stromal cells express the Notch ligand (specifically the delta-like family of proteins), and thymocytes express Notch receptors. Data from murine models confirmed the necessity of Notch signaling in the development of T lymphocytes. Mice with constitutively active Notch have impaired B-cell development. Notch-deficient mice have impaired T-cell development. Activation of the Notch system results in cleavage of intracellular Notch, which then translocates to the nucleus to induce the transcription of genes specific to T cells. Notch is critical for the cellular decision of development in the B- versus T-cell lineage.

All T cells express CD3, the TCR. About 60% of mature T cells express CD4 and are T-cell helpers-inducers, whereas 40% express CD8 and are suppressor-cytotoxic cells. The proper ratio of CD4⁺ to CD8⁺ cells in the body is more than 1. A ratio of less than 1 is abnormal. Numbers of mature T cells peak at age 6-12 months and then decline to adult levels. CD4⁺ levels are high in neonates but then decrease over time. Levels of CD8⁺ T cells are low in neonates but then increase over time.

T cells can also assist in the clearance of intracellular organisms, virally infected cells, and tumor cells. T cells are implicated in the rejection of foreign tissues and in the pathogenesis of graft versus host disease (GVHD). They are also involved in the regulation of other immune system cells. T cells perform many of these functions by means of their production and response to cytokines.

Cytokines

Cytokines orchestrate growth, differentiation, and activation of the immune system. Cytokines have a molecular weight of 10-30 kD and typically have a short half-life. They are secreted by many different types of cells and have different functions; they are immune regulators, lymphocyte growth factors, mediators of inflammation, and stimulators of the bone marrow.

ILs have proinflammatory functions in the regulation of the immune response. Some ILs promote the proliferation, differentiation, activation and apoptosis of the cells of the immune system. Some ILs, including IL2, also have antiviral and antitumoral functions.

Interferons (-a, -b, -g) are involved in the regulation of immune responses and promote the activation and differentiation of the cells of the immune system. Interferons also possess antiviral and antitumoral properties.

Tumor necrosis factor (TNF)-a and TNF-b have proinflammatory properties as well as antitumoral functions.

Hematopoietic growth and survival factors are other types of cytokines and include G-CSF, GM-CSF, and macrophage colony-stimulating factor (M-CSF). They are primarily stimulators of myelopoiesis.

Some of these cytokines have been clinically used, with various degrees of success, to manipulate the immune system. Interferon-γ has been used successfully in the treatment of chronic granulomatous disease (CGD). Interferon-α has been successfully used in the treatment of some childhood vascular tumors. Administration of G-CSF to promote neutrophil development has been successful in some individuals with congenital or acquired neutropenia.

T cells respond to endogenously or exogenously derived peptide fragments. These antigenic fragments include viral proteins and antigenic fragments derived from invading organisms by means of the phagocytic process. The antigenic fragment is displayed by an antigen-presenting cell in the context of MHC molecules. MHC class I–restricted HLA molecules include HLA-A, HLA-B, and HLA-C and are found on nearly all nucleated cells. MHC class II–restricted HLA molecules include HLA-DR, HLA-DP, and HLA-DQ and are also expressed on antigen-presenting cells, including monocytes, dendritic cells, and B cells. TCRs subsequently bind to the complex of the antigenic fragment and the HLA molecule. CD4⁺ T cells recognize exogenously derived antigens in the context of MHC class II molecules. CD8⁺ T cells recognize endogenously derived antigens in the context of MHC class I molecules.

CD4⁺ T cells generate memory cells after the initial encounter with the antigen. In addition to memory cells, 3 types of CD4⁺ T-helper cells (TH0, TH1, and TH2) aid in orchestrating the immune response. TH0 cells are the source of TH1 and TH2 cells. TH1 cells produce and are stimulated by interferon-γ, IL-12, and other cytokines. TH1 cells contribute to delayed-type hypersensitivity reactions.

In contrast, TH2 cells produce and are stimulated by IL-4, IL-5, IL-6, and IL-10. TH2 cells aid in the humoral or innate immune response by helping B-cell differentiation. TH1 and TH2 cellular responses are believed to operate in opposition. For example, an active TH1 response inhibits TH2 activity and an active TH2 response inhibits TH1 activity. TH1 and TH2 cellular responses are also affected by cytokines in a similar fashion. For example, IL-12 stimulates a TH1 response and inhibits a TH2 response, whereas IL-4 and IL-10 stimulate a TH2 response and inhibit a TH1 response.

Most cytotoxic T lymphocytes express CD8 and recognize endogenously derived antigens in the context of MHC class I molecules. They are critical in mediating allograft rejection, tumor surveillance, and destruction of intracellular pathogens. They may also induce apoptosis by means of the Fas/Fas-ligand system or cytolytic granules. CD8⁺ T cell cytokine-secretion patterns may play a role in suppressing immune responses.

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Immunodeficiency

Primary and secondary immunodeficiency

Causes of immunodeficiency are primary and secondary. Approximately 70 primary immunodeficiencies have been described. About 70% of these are antibody deficiencies. The overall incidence of primary immunodeficiencies is approximately 1 case per 10,000 individuals. This rate excludes IgA deficiency, which has an incidence of 1 case per 400 individuals. Approximately 70% of secondary immunodeficiencies are associated with T-cell defects. About half of these cases occur in patients who are hospitalized. Secondary immunodeficiencies in this population are caused by immunosuppressive therapy, splenic hypofunction due to surgery or disease states, or nutritional deficiency.

Clinical details

Immunodeficient states can manifest as an increased frequency or severity of infections, as an unusually prolonged or complicated infection, or as an infection with a nonvirulent or unusual organism. A thorough medical history is essential to diagnose an immunodeficiency disorder. A healthcare professional should elicit details about previous infections, previous medical conditions (including allergic or autoimmune disorders), birth history, family history, drug or toxin-exposure history, travel history, and immunization history. A thorough history also includes a comprehensive growth and development history and a review of systems to assess for signs and symptoms of infection and inflammation.

Careful physical examination may demonstrate an absence of lymph nodes in a patient with infection. Other physical findings that may suggest an immunodeficiency include oral ulceration, gingivitis, or an autoimmune phenomenon. Eczema is common in patients with WAS, and oculocutaneous albinism is characteristic of Chediak-Higashi syndrome.

Roughly half of patients referred to a pediatric immunologist because of concerns of immunodeficiency are confirmed to be healthy. They have normal growth and development and a history of typical childhood infections separated by healthy periods. Healthy children have an average of 6-8 respiratory tract infections each year through the first decade of life. For the first 2-3 years of life, a typical child can have roughly 6 episodes of otitis media and roughly 2 episodes of gastroenteritis.

About a third of patients who are referred to the immunologist have allergic disorders. They have normal growth and development and afebrile illnesses that respond poorly to antibiotic therapy. These children may have chronic cough, atopic skin changes, and other physical changes suggestive of an allergic disorder.

Laboratory testing

Initial screening tests should include a CBC count and differential. Lymphopenia suggests immunodeficiency, especially in newborns. Abnormal granules are characteristic of Chediak-Higashi syndrome. A peripheral blood smear may also allow the clinician to identify small platelets characteristic of WAS.

In general, children with B-cell or antibody deficiencies, phagocytic deficiencies, or complement deficiencies have recurrent infections with encapsulated bacteria. Quantitation of IgG, IgM, IgA, and IgE levels is a useful screening test. Levels more than 2 standard deviations below the levels for age-matched control subjects are abnormal. If IgG levels are normal but functional deficits are present, subclass studies may be useful. If Ig levels are low, B-cell enumeration may be useful.

One can assess numbers of B cells using flow cytometry. An absence of B cells suggests a diagnosis of Bruton (X-linked) agammaglobulinemia. When interpreting laboratory results, the clinician should remember that some laboratories fail to use appropriate age-adjusted normal ranges. In assessing B-cell function, determination of antibody titers to proteins (eg, tetanus and diphtheria toxins) and to polysaccharides antigens (eg, pneumococcal antigens) are useful following immunization. Measurement of isohemagglutinins, which are IgM antibodies to the polysaccharide antigens that define the ABO blood system, are another method to assess B-cell function.

Children with complement deficiencies may present with autoimmune disorders. They often have severe recurrent infections with encapsulated bacteria. They may also present with disseminated neisserial infection or meningococcal meningitis. Complement function can be assessed by measuring levels of C3 and C4. A CH50 titer is a useful screening test for deficiencies in the complement cascade. The CH50 titer is used to measure the ability of a dilution of the patient's serum to lyse antibody-coated sheep RBC. With a blockage in the complement cascade, the titer is zero.

Patients with T-cell defects may have infections with opportunistic organisms. The most simple and effective screening test for assessing T-cell function is an intradermal skin test, which provides a qualitative assessment of the T-cell response to antigens to which a person has been exposed. This skin test may involve use of a Candida albicans extract, although other intradermal agents, including purified protein derivative and tetanus toxoid, may also be used to assess cellular immunity. T-cell enumeration using cell surface markers may be useful in the workup of a patient with suspected cellular immunodeficiency. T-cell activation may be assessed in vitro by mitogen-response testing. T-cell activation and proliferation in response to various mitogens may provide useful information with respect to T-cell function.

Phagocytes may be enumerated with a WBC count and differential. Phagocytic activity can be evaluated using a respiratory-burst assay with fluorescent dyes or the older NBT test (see Chronic granulomatous disease). For advanced testing of phagocytic activity, phagocytic and bactericidal assays are available.

In addition to the laboratory tests noted above, various research laboratory tests can be performed to further characterize a primary immunodeficiency disorder. These can be accessed by a pediatric immunologist at a tertiary-care or academic center.

Treatment options for immunodeficiency

Treatment options for immunodeficiency disorders may be disease-specific, whereas others may help regardless of the particular diagnosis. Patients should adhere to particular vaccination instructions. In some cases, prophylactic use of antimicrobial agents provides protection against bacterial, viral, and fungal pathogens. IVIG may also be used therapeutically.

Allogeneic hematopoietic stem-cell transplantation (HSCT) from HLA-identical siblings, matched unrelated donors, or haploidentical donors are sometimes used. Sources of hematopoietic stem cells are numerous and include bone marrow, peripheral blood stem cells, and umbilical-cord blood. HSCT has been used with various degrees of success. The results of transplantation depend on the timing of the procedure in relation to the patient's disease, the type of transplant, the particular immunodeficiency, and the general health of the recipient.

Gene therapy has been used, although results are limited. Understanding and characterizing immunodeficiency at a molecular level has been extremely helpful and is likely to become even more important as clinicians attempt to tailor specific therapies for particular immunodeficiencies.

Antibody Deficiencies

Selective IgA deficiency is the most common primary immune deficiency. The incidence of this disorder is approximately 1 case per 400 individuals, and it arises sporadically in most instances. This condition may be transient, acquired, or partial. Laboratory evaluation reveals an IgA level of less than 5 mg/dL without other major findings. Normal serum IgG and IgM levels are noted. Most cases involve normal numbers of IgA-bearing B-cell precursors, but those IgA-bearing B cells fail to mature into IgA-secreting plasma cells.

Most patients remain asymptomatic and healthy. Upper respiratory infections typically involve encapsulated bacteria, such as Streptococcus pneumoniae or Haemophilus influenzae. Lower respiratory tract infections are rare. GI infections with parasites, such as Giardia organisms, are also common. In addition to the infectious complications, patients may develop allergic disorders, autoimmune disorders, GI disorders, CNS disorders, and malignancy. Approximately one fourth of patients have allergies, one fourth have recurrent respiratory infections, one fourth have other autoimmune disorders, and one fourth are largely asymptomatic. Close to 20% of those with IgA deficiency also have an IgG2-subclass deficiency.

Treatment consists of antibiotic therapy and management of specific autoimmune conditions and associated disorders. IVIG therapy is generally avoided because of the risk of anaphylaxis to IgA-containing substances, although low-IgA formulations of IVIG are available.

Transient hypogammaglobulinemia of infancy

The genetics and pathogenesis of transient hypogammaglobulinemia of infancy are not well characterized. The postnatal decrease in serum IgG levels is accentuated, and endogenous Ig synthesis occurs more slowly in children with this disease than in others. Patients typically present at age 4 months to 2 years with recurrent respiratory tract infections, bacterial infections, or chronic diarrhea. Laboratory test findings reveal low quantities of Ig, specifically IgG (at least 2 standard deviations below levels for age-matched control subjects) with or without depressed levels of other Ig isotypes. B cells are present in normal numbers. Patients are usually asymptomatic. IgG levels commonly normalize by age 2-3 years. IVIG treatment is usually reserved for symptomatic infants with severe infections.

IgG-subclass deficiencies

IgG has 4 subclasses (IgG1, IgG2, IgG3, and IgG4). Each subclass has a unique biologic and functional characteristic. The basic genetic defects responsible for immunodeficiencies involving these subclasses are not well characterized. Clinical findings include recurrent respiratory infections with encapsulated bacteria.

IgG1 makes up the bulk of IgG; therefore, a deficiency of IgG1 is classified as panhypogammaglobulinemia, as opposed to an IgG-subclass deficiency. IgG2-subclass deficiency is the most common subclass deficiency after IgG4-subclass deficiency. It is associated with impaired polysaccharide responsiveness because the IgG2 subclass contains the antibodies to polysaccharide antigens. Therefore, individuals with an isolated IgG2-subclass deficiency may have infections with encapsulated bacterial, such as S pneumoniae and H influenzae. Patients with IgG4-subclass deficiency are usually asymptomatic.

Laboratory evaluation findings demonstrate normal absolute levels of IgG, but a subclass is deficient. Levels less than 2 standard deviations below age-matched control data are of concern but are clinically significant only if the response to protein or polysaccharide antigens is deficient. Minimal normal levels of IgG subclasses are 280 mg/dL for IgG1, 50 mg/dL for IgG2, and 25 mg/dL for IgG3. About 25% of healthy persons do not have any detectable IgG4; therefore, no minimal level of this subclass appears to be necessary. IgG-subclass deficiency is a common part of other immunodeficiencies. Typical patterns of Ig deficiency include paired deficiencies of IgG2-IgG3, IgG2-IgA, and IgG1-IgG3. In specific cases, treatment of these disorders may include IVIG or antibiotics.

Impaired polysaccharide responsiveness

Impaired polysaccharide responsiveness is a heterogenous disorder with multiple causes. It commonly affects children aged 2-10 years. In this disorder, normal numbers of B cells and Igs are present but do not respond well to polysaccharide antigens; this leads to infections with H influenzae, Streptococcus, Pneumococcus, Staphylococcus, and Neisseria meningitides. Patients have normal antibody responses to protein antigens.

Children with impaired polysaccharide responsiveness typically have multiple sinus, ear, or pulmonary infections. This disorder may be transient. The diagnosis cannot be established before age 2 years because the immunologic response to polysaccharide antigens may not be mature until this age. Patients with this disorder can be treated with IVIG or antibiotics.

X-linked hyper-IgM syndrome

This is an X-linked recessive disorder caused by one of 3 different genetic defects. It is characterized by the absence of CD40 ligand, which normally appears on activated T cells. CD40, which is constitutively expressed on B cells, requires binding of CD40 ligand for B-cell proliferation and differentiation. Without CD40 ligand, B cells cannot undergo class-switch recombination. Blockage of this normal process leads to antibody diversification. IgM accumulates, and IgG and IgA are scarce. Although patients have normal numbers of B cells, they may have various degrees of T-cell immunodeficiency and neutropenia. Autosomal recessive forms of hyper-IgM syndrome are recognized, though the X-linked recessive form is most common.

Clinical presentations of this disease typically occur after age 6 months and consist of pneumonia, sepsis, meningitis, osteomyelitis, sinusitis, vaccine-related polio infection, echoviral encephalitis, or conjunctivitis. Affected children typically present with upper and lower respiratory tract infections and are susceptible to infection with P jiroveci.

They may also present with diarrhea and are notably susceptible to cryptosporidium with subsequent sclerosing cholangitis. Other manifestations include a host of autoimmune phenomena and hepatic malignancies. Neutropenia (cyclic or chronic) is commonly seen. The basis of the neutropenia is unknown, but it may respond to G-CSF. 

The diagnosis is confirmed by demonstrating the absence of CD40 ligand on activated T cells. Monitoring for liver and biliary disease is important. Untreated patients can develop infections, chronic liver disease, and other sequelae, including malignancy. Treatment consists of prophylactic IVIG, P jiroveci prophylaxis, and HSCT.

Duplantier et al (2001) described their experience with HSCT in X-linked hyper-IgM syndrome.⁴ They also briefly review HSCT and its success in treating this disorder. Data from small case series suggest that transplants from HLA-identical siblings provide the best outcome. Survival was 20% at 25 years without bone marrow transplantation (BMT). Eleven patients have been transplanted for this disorder; 5 received transplants of bone marrow from a fully matched sibling donor, and all patients are alive and have engrafted. Only one of the 3 with matched unrelated donor transplants is alive. One died of complications, and one did not engraft. Only one of 3 haploidentical transplants engrafted.

This is an X-linked recessive disorder caused by a defect in the BTK gene, which encodes BTK, which is a tyrosine kinase essential for B-cell proliferation, differentiation, and survival. BTK is expressed at all stages of B-cell development. Blockage at the pre–B-cell stage of development results in a lack of mature B cells. As a result, numbers of B cells and levels of Igs are decreased.

Children with this disorder present after age 6 months as maternal antibody levels fall. Bacterial infections, such as sinusitis, conjunctivitis, pneumonia, osteomyelitis, meningitis, and sepsis, are common. Patients can also have arthritis, bronchiectasis, echoviral encephalitis, vaccine-related polio infection, and malignancy. Infectious complications typically involve encapsulated bacteria because antibodies are involved in opsonization of these bacteria. Patients are also at increased risk of enteroviral infections.

Physical examination reveals absent lymphatic tissue. Laboratory test findings reveal an Ig level of less than 400 mg/dL, no inducible antibody production, and complete or partial absence of B cells. Cellular immunity remains normal. Treatment consists of IVIG, antibiotic therapy, and, potentially, antiviral therapy. More recently, murine models studied by Yu et al (2004) demonstrated that gene transfer into HSCs can reconstitute BTK-dependent B-cell development and function in vivo, supporting the feasibility of pursuing BTK gene transfer for X-linked agammaglobulinemia.

CVID is also known as acquired agammaglobulinemia. This condition is most commonly diagnosed after puberty but is difficult to diagnose because of variable phenotypes that may markedly change over time. IgG levels must be below 500 mg/dL, and IgA and IgM levels must be 2 standard deviations below age-matched control data. However, the diagnosis remains one of exclusion.

Other primary antibody deficiencies or other causes of hypogammaglobulinemia must be ruled out. Numbers of B cells are usually normal, but they may be reduced or absent. Serum IgG and IgA values are most commonly low, although IgM levels may also be reduced. Numbers of IgA-, IgG-, and IgM-bearing B-cell precursors are normal, but numbers of IgA-, IgG-, and IgM-producing plasma cells are reduced. This finding suggests that patients have B cells that do not differentiate into antibody-producing plasma cells.

The inability to make specific antibody responses remains a consistent hallmark of the disease. Molecular defects in the inducible costimulatory (ICOS), CD19, the transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI), and the B-cell activating factor that belongs to the TNF family (BAFF-R) have been identified. Mutations in the TNF-receptor family member TACI, which mediates isotype switching in B cells, were found in 10-20% of patients with CVID. 

TACI is expressed on B cells and interacts with BAFF and APRIL, which are expressed on macrophages and dendritic cells. These proteins all have roles in B-cell activation and class switching. TACI and BAFF-R are critical to the maintenance of B-cell homeostasis and have important roles in isotype class switching. CD19 deficiency highlights the importance of antigen receptor signaling, whereas ICOS deficiency illustrates the absolute need for T:B-cell interaction in coordinating an effective secondary humoral response. 

Children with CVID may present with recurrent sinopulmonary infections, including bacterial pneumonia, sinusitis, otitis, and bronchiectasis. They may develop GI inflammation, chronic diarrhea, hepatosplenomegaly, lymphadenopathy, benign lymphoproliferative disorders, autoimmune phenomena, GI malignancies, and hematologic malignancies (usually B-cell lymphomas).

Ig levels are low. The ratio of CD4⁺ to CD8⁺ T cells can be abnormal, with a decreased number of CD4⁺ T cells relative to CD8⁺ T cells. The treatment of CVID may include prophylactic antibiotic therapy and the use of IVIG.

T-Cell Deficiencies

DiGeorge syndrome

DiGeorge syndrome is caused by the failure of development of the third and fourth pharyngeal pouches. A spectrum of findings includes characteristic facies with hypertelorism, ear malformations, palatal defects, and micrognathia. Other findings include hypocalcemia manifested as neonatal tetany secondary to parathyroid hypoplasia or aplasia. Thymic hypoplasia or aplasia with subsequent cellular immunodeficiency may occur. Congenital heart disease, including anomalies of the aortic arch, tetralogy of Fallot, and truncus arteriosus, is often associated with this disorder. Patients are at increased risk of autoimmune phenomenon, malignancy, and language delay.

Secondary to palatal defects and defects in cellular immunity, patients are predisposed to developing upper respiratory infections. As a result of deficient cell-mediated immunity, patients have an increased susceptibility to viral infections in particular. The deficit in cell-mediated immunity may improve over time and is variable with respect to the number and function of T cells. Some patients may have impaired humoral immunity as well.

Fluorescent in situ hybridization (FISH) reveals a deletion at chromosomal band 22q11.2 in 90% of patients. Laboratory evaluation may reveal hypocalcemia, lymphopenia with normal Ig levels, an attenuated antigen response, or an absent thymic silhouette on chest radiographs.

Prophylaxis against P jiroveci infection is often indicated. BMT, peripheral-blood lymphocyte transfusion, and thymic transplantation have been used, with some success in patients with severe T-cell defects, although most cases involve a mild-to-moderate defect in cell-mediated immunity.

Markert et al (2007) reviewed 54 patients with complete DiGeorge anomaly.⁵ Of the 44 who underwent thymus transplantation, 33 (75%) survived; median follow-up posttransplantation was 3 years and 10 months, and the range was 7 months to 13 years. Transplantation was well tolerated and resulted in stable immunoreconstitution in these infants.

Individuals with CMC typically have chronic superficial fungal infections with Candida or other fungal pathogens. These infections may involve the skin, nails, or mucous membranes without evidence of sepsis.

CMC is a heterogenous disorder divided into 7 clinical subgroups based on the extent, location, underlying molecular defects, and associated complications. Patients may have endocrine abnormalities, including hypothyroidism, hypoparathyroidism, adrenal insufficiency, or diabetes. These endocrine abnormalities are least common in patients who present in adolescence.

CMC has been described as part of the autosomal recessive polyendocrinopathy-candidiasis-ectodermal dysplasia syndrome. The underlying genetic defect associated with this syndrome is a mutation in the autoimmune regulator (AIRE) gene. Other subgroups of CMC include chronic oral candidiasis, familial chronic mucocutaneous candidiasis, CMC with thymoma, localized candidiasis, candidiasis with keratitis, and candidiasis with hyper-IgE syndrome. In most children with CMC, the underlying genetic defect is unknown.

CMC is a disorder of selective T-cell unresponsiveness. The range of immunologic abnormalities is wide, although the most common abnormality is a selective defect of cell-mediated immunity against Candida species. This feature is best demonstrated by the finding of cutaneous anergy to Candida antigen or decreased lymphocyte proliferation in response to candidal antigens. Numbers and function of T cells remain normal. Humoral deficiency is not uncommon in CMC. The most common humoral deficiency associated with CMC is IgG-subclass deficiency, but IgA deficiency is possible.

The mainstay treatment consists of local and systemic antifungal agents. Infusions of peripheral-blood leukocytes and transplantation of thymic tissue have been used. Prior to effective antifungal therapy, thymus transplants were performed (as early as 1968 through 1989) with limited success. Two patients received peripheral blood leukocytes from donors, but both later lost their hypersensitivity response to Candida.^6,7

Combined Immunodeficiencies

The ZAP-70 protein is a tyrosine kinase that plays a role in the selection and maturation of T cells in the thymus. A deficiency of this protein results in a paucity of CD8⁺ T cells.

The ZAP-70 defect is due to mutations of the gene on chromosome 2q12 that encodes ZAP 70. The ZAP-70 tyrosine kinase has a critical role in signals that are transmitted via the TCR. Patients may have a classical SCID phenotype; they may have elevated total lymphocyte counts with higher CD4 T cell levels and almost complete absence of CD8 T cells. Although the CD4 cells appear normal, they fail to respond to signals via the TCR. ZAP 70 is critical for normal thymic development of CD8 cells. Mature CD4 T cells depend on ZAP 70 for activations. 

Patients may have humoral immunodeficiency that manifests as decreased serum Ig levels and impaired B-cell function. This spectrum of immunodeficiency leads to bacterial, viral, and fungal infections similar to those seen in patients with SCID. One key difference of ZAP-70 defects compared with SCID is the presence of lymph nodes and a normal thymus in patients with ZAP-70 defects because they still have normal CD4 counts. Patients can present with a moderate or severe phenotype and may appear clinically similar to patients with SCID. Numbers of NK and B cells are normal. B-cell activity may be diminished, and serum Ig levels may be abnormal, although humoral immunity is usually spared.

Use of antimicrobial prophylaxis, IVIG, and HSCT has been described. Four patients have received a transplant, and 3 have been cured.^8,9,10

SCID, or acquired agammaglobulinemia, is characterized by lymphopenia and a complete absence of normal B- and T-cell function. Some types of SCID decrease numbers of NK cells and decrease activity of NK cells.

SCID is usually identified in the first 2-3 months of life. Clinical manifestations can be severe and may include candidiasis, P jiroveci infection, GI infections, and failure to thrive. Rashes, malabsorption, chronic cough, and absent lymphatic tissue are also characteristic. Maternal lymphocytes may engraft in patients and cause GVHD. At least 9 genetic mutations have been identified as causes of SCID, and each is associated with a lymphocyte phenotype.

The X-linked form of SCID (XSCID) is the most common form of SCID and represents 30-40% of all cases. It is thought to occur in approximately 1 per 50,000 births. In patients with XSCID, the lymphocyte phenotype includes T cells, which are decreased or absent in number. B cells are present in normal numbers, but their function is abnormal. Ig levels are low, and specific antibody responses are diminished or absent. NK cells are decreased or absent in number. Numerous mutations have been demonstrated in patients with XSCID. Identified gene defects include defects in IL-2RG (the gene that encodes the gamma chain of the IL-2 receptor). Common gamma-chain deficiency results in faulty signaling through a cytokine receptor with subsequent effects on T, B, and NK cells.

Autosomal recessive forms of SCID share a lymphocyte phenotype similar to that described for XSCID. One of these forms results from mutations in the Jak3 protein, a signaling molecule associated with the common gamma chain. A second autosomal recessive form of SCID involves a deficiency in CD45, a membrane-associated tyrosine phosphatase that regulates Src kinases needed for T- and B-cell–receptor signal transduction.

Another lymphocyte phenotype includes decreased or absent numbers and function of T cells with relatively normal proportions of B and NK cells. This phenotype is the result of mutations in the receptor for IL-7 (IL-7R alpha). Mutations in the CD3 delta chain can mimic the lymphocyte phenotype described for mutations in IL-7R alpha. The CD3 delta chain functions as a T-cell antigen receptor that is essential for T-cell development.

A third lymphocyte phenotype includes absent or decreased numbers of T and B cells with normal numbers of NK cells. This autosomal recessive form of SCID involves mutations in the recombinase activating genes RAG1 and RAG2, the gene products of which are essential for the rearrangement of antigen receptor genes. Artemis deficiency is a deficiency of a variable diversity joining (VDJ) recombination and DNA repair factor. As a result of Artemis deficiency, cells cannot repair DNA after RAG1 and RAG2 products make double-stranded cuts. The lymphocyte phenotype is similar to that described for RAG1 and RAG2 mutations. Partial deficiency of RAG is associated with autoimmune phenomenon caused by oligoclonal lymphocytes. This has been described as Omenn syndrome, which is characterized by erythroderma, adenopathy,hepatosplenomegaly, edema secondary to protein loss, marked hypereosinophilia, and an elevated IgE level.

Laboratory findings in SCID include lymphopenia. B- and T-cell counts are typically decreased, although numbers of B and NK cells may vary, as demonstrated in the various lymphocyte phenotypes described. Other laboratory findings include cutaneous anergy, absent in vitro mitogen responses, and decreased Ig levels. Physical examination may demonstrate decreased lymphoid tissue, and radiographic examination may demonstrate the absence of a thymic silhouette.

HSCT is the only curative treatment for SCID. T-cell depletion of the graft before transplantation may decrease the likelihood of GVHD. Many patients with SCID cannot reject allografts because of the absence of T-cell function; therefore, they typically do not require substantial immunosuppression during or after transplantation.

Buckley reviewed an experience with 132 patients with SCID who underwent HSCT.¹¹ Approximately three fourths of the patients were alive at a median follow-up of 5.4 years. Grunebaum et al (2006) reviewed 94 infants with SCID who underwent BMT. The survival rate was 92.3% in patients who received HLA-identical donor BMT, 80.5% in patients who received HLA-matched unrelated donors BMT, and 52.5% in patients who received HLA-mismatched related donors BMT. Survival was significantly higher with HLA identical donor BMT or with HLA-matched unrelated donor BMT, suggesting that, in the absence of a relative with identical HLA, HLA-matched unrelated donor BMT may provide better engraftment, immune reconstitution, and survival for patients with SCID than HLA-mismatched related donor BMT.

SCID associated with the purine enzyme defects of ADA deficiency or PNP deficiency results in a combined B- and T-cell deficiency. Patients with SCID associated with purine-enzyme defects can present with skin, respiratory, and GI infections. P jiroveci and Candida species are frequently isolated from affected children. Patients may present with evidence of failure to thrive.

ADA and PNP deficiency are both autosomal recessive. Only ADA deficiency is associated with skeletal abnormalities. ADA deficiency is associated with decreased or absent T-, B-, and NK cell immunity. PNP deficiency is associated with neurologic impairment (usually motor dysfunction) and immunodeficiency characterized by normal B-cell immunity, which may decrease over time, and severely depressed or absent T-cell immunity. PNP deficiency may be associated with autoimmune phenomenon.

In ADA deficiency, intracellular accumulation of toxic levels of purine intermediates, such as deoxyadenosine and deoxyguanosine occurs. These are converted to 5 prime triphosphates, which inhibit ribonucleotide reductase and prevent de novo synthesis of deoxynucleotides. These cells cease to divide and undergo apoptosis. The T-cell precursors in the thymus are especially sensitive to apoptosis. This excess impairs normal cellular function and directly or indirectly leads to lymphocyte apoptosis. Partial deficiency of ADA is described.

Mutations that preserve some enzyme activity may lead to mild forms of combined immunodeficiency, which occur in adulthood as numbers of T cells decline. Many distinct mutations in the ADA gene have been described. The clinical spectrum of disease is associated with the underlying mutation. In PNP deficiency, the accumulation of guanosine and deoxyguanosine appears to be toxic to T cells and the CNS.

Laboratory assessment of PNP or ADA enzyme activity demonstrates low or absent activity in RBCs and WBCs. T-cell number and phytohemagglutinin (PHA) response is decreased. High levels of circulating deoxyadenosine are observed in ADA enzyme deficiency. Skeletal dysplasia and absence of a thymic silhouette may be observed during radiographic evaluation of patients with ADA deficiency. Patients with PNP deficiency typically have a low uric acid level because PNP is needed for purine degradation.

Treatments for ADA deficiency include HSCT, which is the treatment of choice for those patients with an HLA-identical sibling. The use of polyethylene glycol–modified ADA (PEG-ADA) has largely replaced RBC transfusion to provide the deficient enzyme. PEG-ADA is administered intramuscularly once or twice per week.

Retrovirally mediated gene therapy based on autologous cord blood, marrow, or lymphocytes into which the normal gene is transduced has been used with some success in patients with ADA deficiency. Although patients may typically still require PEG-ADA administration, their levels of ADA are higher than those of patients not undergoing gene therapy. Transduced cells express normal levels of ADA and showed normal function in vitro. These cells also function normally in vivo. However, they did not provide a sufficiently diverse population of T cells to reconstitute protective immunity in all patients. Retroviral transduction is only 5-25% efficient. 

Transduction of mature T cells appears superior to attempts to reconstitute hematopoietic stem cells. Although improvements in T-cell function have occurred, complete immune reconstitution has not been achieved.¹²  The SCID trials carried out in Paris and London have yielded therapeutic benefit in most patients. Therapy-related leukemia has occurred in 3 patients. Genomic insertions of retroviruses in mouse bone marrow has led to leukemogenesis; only one case has been reported in primates (a monkey) and no cases have been reported in patients with ADA deficiency. In PNP deficiency, enzyme replacement has not been effective in the long term. HSCT has been used with some success.

Buckley reviewed an experience with 132 patients with SCID who required HSCT.¹¹ Approximately three quarters of the patients were alive at a median follow-up of 5.4 years. Approximately three quarters of the patients with SCID secondary to ADA deficiency survived after HSCT.

Reticular dysgenesis is an autosomal recessive or X-linked recessive disorder that leads to congenital failure of stem cells committed to myeloid and lymphoid development. Patients typically present in the first few days of life with failure to thrive, emesis, diarrhea, and infectious complications.

The condition is thought to be an early defect in hematopoiesis because all leukocytes fail to develop, although the molecular basis of this disorder is unclear. Reticular dysgenesis is characterized by lymphopenia and subsequent B- and T-cell immunodeficiency and agranulocytosis. Lymphatic and thymic tissue is hypoplastic. Patients with reticular dysgenesis have low serum concentrations of IgG and IgM. Thrombopoiesis and erythropoiesis remain normal, although anemia and thrombocytopenia may be present. Bacterial and viral infections are severe and begin early in life.

G-CSF therapy has not been effective, but HSCT can be used as a treatment modality. Bertrand et al (2002) reported their experience with haploidentical HSCT for reticular dysgenesis.¹³ Three of 10 patients were still alive after transplantation.

Other Primary Immunodeficiencies and Phagocyte Deficiencies

Other Primary Immunodeficiencies

The gene for WAS has been identified on chromosome X and is expressed in all hematopoietic stem-cell lineages. The gene product of WAS serves as an intracellular signaling molecule that plays a critical role in cytoskeletal organization, lymphoid development, phagocytosis, and apoptosis.

The classic presentation of WAS includes eczema, thrombocytopenia (particularly with microthrombocytes), and severe bacterial infections. This X-linked recessive disorder is often associated with decreased T cell-mediated immunity, resulting in opportunistic and viral infections. Numbers of B and NK cells are normal, although their function may be diminished.

Other manifestations include lymphadenopathy, hepatosplenomegaly, otitis with chronic otorrhea, food intolerance, autoimmune phenomenon (including hemolytic anemia glomerulonephritis), inflammatory bowel disease, and vasculitis, as well as malignancy (including leukemia and lymphoma). Malignancies are present in 13% of patients by age 9.5 years and are most commonly associated with Epstein-Barr virus. 

Lymphopenia and decreased cellular and humoral immunity may evolve over time. IgM and IgG levels are typically low, and IgA and IgE levels are typically high. Other laboratory findings include eosinophilia, small platelets, low isohemagglutinins, decreased number of T cells, cutaneous anergy, and decreased T-cell responses to mitogens (polysaccharide antigens). Monocytes, macrophages and dendritic cells have defective adhesion and motility. Screening for defects in WASP expression can be performed using flow cytometry with a suitable anti-WASP antibody; the diagnosis is ultimately confirmed by mutation analysis of the WASP gene.

Available treatments include antibiotics, splenectomy (in some cases followed by lifelong antibiotic prophylaxis), immune modulators (eg, anti-CD20, steroids), and platelet transfusions as needed. IVIG may also be used in the supportive care of some patients. At present, the only curative therapy is HSCT. Killed vaccinations can be given, but the response rate may be poor and should be measured.

Filipovich et al (2001) reviewed the outcomes of 170 patients with WAS from the International Bone Marrow Transplant Registry.¹⁴ The 5-year survival probability for all subjects was 70% (with a range of 63-77%). The 5-year survival probability was 87% with HLA-matched sibling donors, 52% with other related donors, and 71% with matched unrelated donors.  Many deaths were due to Epstein-Barr virus lymphoproliferative disorders.^15,16,17 HLA-identical sibling transplantation provided the best outcome, with an 80% probability of survival at 5 years.

Reports of successful gene therapy of SCID and CGD have encouraged the development of similar strategies for WAS. Gene transfer has demonstrated corrections in multiple cellular defects of murine and human cells in vitro . Clinical studies for treatment of WAS using gene transfer to HSCs are currently in preparation.

Ataxia telangiectasia is an autosomal recessive disorder caused by abnormalities of the ATM gene on chromosome band 11q22-23. The gene product is a protein kinase that is involved in regulating the cell cycle, cellular responses to DNA damage, and DNA recombination. Cells that harbor the mutant protein may have chromosomal instability, accelerated telomere shortening, sensitivity to radiation, and dysregulation of the cell cycle.

Affected patients typically present with progressive ataxia and motor development may be delayed. These neurologic findings are associated with a loss of cerebellar Purkinje cells. Patients can develop oculocutaneous telangiectasias (small dilated blood vessels) involving the conjunctivae, eyelids, ears, and skin. Patients also develop cellular and humoral immunodeficiency with progressive lymphopenia and Ig deficiencies. This spectrum of immunodeficiency may lead to recurrent sinopulmonary infections caused by bacterial and viral pathogens. These recurrent infections may lead to chronic pulmonary insufficiency.

Ectodermal changes give rise to an appearance of premature aging and endocrinopathies, including growth retardation, pubertal delay, and hypogonadism. Cancer is a common complication of ataxia telangiectasia. The risk of cancer is approximately 1% per year after age 10 years. Leukemia and lymphoma are common during childhood years in these patients, whereas epithelial cancers predominate in adulthood. This predisposition to the development of T- and B-cell malignancies may be associated with their increased sensitivity to ionizing radiation and genetic instability.

Laboratory findings reveal an IgA deficiency in 75% of patients and an IgE deficiency in 25%. Other findings include cutaneous anergy, decreased numbers of T cells, a decreased response to mitogens, and marked chromosomal fragility. Levels of carcinoembryonic antigen (CEA) and alpha-fetoprotein are markedly elevated and may aid in the diagnosis. No effective treatment for ataxia telangiectasia is available. IVIG can be used in cases of antibody deficiency, and antibiotics should be used when appropriate.

Hyper-IgE syndrome, also known as Job syndrome, is a rare and complex disorder characterized by high levels of serum IgE and chronic dermatitis. Children with this disorder may have recurrent staphylococcal infections of the joints, lungs, and skin. They may also have asthma, allergies, coarse facial features, and bony abnormalities (eg, scoliosis or osteopenia) with subsequent fractures.

Sporadic cases are the most common, although autosomal dominant and autosomal recessive forms have been described. The autosomal dominant form is characterized by immune dysregulation (eczema, elevated IgE levels, eosinophilia, recurrent staphylococcal infections, pneumatoceles), characteristic facies, skeletal abnormalities (hyperextensibility, scoliosis), and abnormal dentinogenesis. The autosomal recessive form is characterized by a different spectrum of immune dysregulation (recurrent viral and fungal infections), neurologic complications (facial paralysis, hemiplegia), and an absence of skeletal and dental abnormalities.

FHL is an autosomal recessive disorder that is often fatal. It typically occurs in early childhood and may have an onset temporally associated with an infection. Patients have an increased inflammatory response that involves hypersecretion of proinflammatory cytokines. All known genetic defects that result in FHL also result in impaired NK cell function and impaired cytotoxic T-lymphocyte function.  Previously healthy children may present with a family history of the disorder, fever, organomegaly, pancytopenia, disorders of coagulation, hypofibrinogenemia, hypertriglyceridemia, and hemophagocytosis. Hemophagocytosis refers to activated macrophages or histiocytes that ingest RBCs, WBCs, and platelets.

The disease is characterized by nonmalignant accumulation and visceral infiltration of activated T lymphocytes and macrophages. Cytotoxic T-cell and NK cell activity is markedly reduced or absent. The disease seems to be characterized by uncontrolled activation of T cells and macrophages with an associated overproduction of inflammatory cytokines. High levels of interferon-g and TNF-a are found in the serum.

This disease has been associated with defects in perforin, a cytotoxic pore-forming protein and a lytic granule constituent. Perforin is manufactured as an inactive precursor that is subsequently modified to create its active form that may then bind to the phospholipid bilayer of target cells. The pores created by perforin may act in concert with granzymes to induce caspase or caspase-independent apoptosis.

Perforin-deficient mice have normal numbers of NK and CD8⁺ T cells. They are healthy in the absence of infection. After viral infection, some perforin-deficient mice develop a dysregulated immune response. These and other findings suggest that, given this blocked pathway and persistently infected cells, antigen-presenting cells lead to persistent T-cell stimulation and production of proinflammatory cytokines.

The association of perforin, which has been mapped to chromosome band 10q22, and FHL has been further delineated as FHL2. Three other loci have been linked to FHL. The gene for FHL1 is not known, although it has been linked to chromosome band 9q21. The third locus is unidentified. FHL3 has been linked to chromosome band 17q25 and involves the gene MUNC13-4, which is instrumental in the fusion of the granule membrane. Cultured lymphocytes from selected patients with FHL showed defective perforin dependent cytotoxic activity and decreased or absent perforin in their granules.

Mutations in the soluble N -ethylmaleimide–sensitive factor attachment-receptor (SNARE) protein (syntaxin 11) have been shown to be responsible for a specific subtype of FHL4. However, syntaxin 11 is not known to be involved in the secretary machinery of the cell or the cytotoxic arsenal of the NK cells and cytotoxic T lymphocytes. Arneson et al suggest that syntaxin 11 in NK cells and activated cytotoxic T lymphocytes may regulate the events that control vesicle movement from the interior of the cell during the development of the cell-mediated cytotoxicity, rather than the terminal steps that influence granule fusion with the plasma membrane.¹⁹  Pathologies involved in FHL4 are likely due to alterations in syntaxin 11–dependent processes directly within cytotoxic lymphocytes. 

Treatment for FHL involves induction chemotherapy with etoposide and corticosteroids, followed by maintenance therapy with cyclosporine and alternating cycles of the same agents, and followed by HSCT in cases with primary hemophagocytic lymphohistiocytosis or recurring noninherited forms.

Ouachee-Chardin et al (2006) reviewed the outcomes of HSCT in patients with FHL.²⁰ The overall survival was 58.5%, with a median followup of 5.8 years (extending to 20 y). Long-term sequelae were limited, with only 7% of patients experiencing a mild neurologic disorder.

Complement deficiencies

Complement proteins are inert precursors that are activated by various different enzymatic cleavages. Activation of the complement system may result in direct lysis of target microorganisms. Complement also aids in the inflammatory process and in opsonization. It also interfaces with the adaptive immune system.

The classic pathway requires antibody for activation. The alternative pathway acts through a series of proteins analogous to the classic pathway to form a C3 convertase that then continues along the classic pathway.

In general, heterozygous deficiency of complement is asymptomatic because half the amount of protein is sufficient for adequate functioning of the complement system. Most complement deficiencies are autosomal recessive, although some are sex linked and others autosomal dominant.

Redundant pathways can overcome deficiency of early components of the classic pathway (upstream of C3). If they are deficient, associated autoimmunity occurs. C3 deficiency is a defect characterized by autoimmunity and infection with Pneumococcus and Neisseria species. Defects of complement factors C5-C9 are autosomal recessive and generally do not manifest until late adolescence. Patients with C5-C8 deficiency have increased susceptibility to neisserial infection. C9 deficiency results in slow lysis of targets and an increased susceptibility to neisserial infection.

Deficiencies in alternative pathways are relatively uncommon and result in an increased propensity to develop Neisserial infections. The gene that encodes properdin is located on the X chromosome. Various types of properdin deficiency are known. Type 1 and type 2 properdin deficiency are quantitative defects in properdin, whereas type 3 properdin deficiency is a qualitative defect. These defects lead to an increased propensity to develop Neisserial infections and autoimmune phenomenon.

Patients with mannan-binding lectin deficiency may have severe infections early in life, although the consequences of deficiencies in this pathway are not completely clear. Serine proteases are activated by pathogens independent of the antibodies that activate the lectin pathway. Deficiency of the mannan-binding lectin is the most common and is associated with susceptibility to infections and autoimmune diseases, especially in infants and children whose immunity is already decreased.²¹ Immunodeficiency laboratory assessment usually reveals a decrease in the CH50 titer, except in cases of isolated C9 deficiency. The CH50 titer is based on the ability of a dilution of the patient's serum to lyse antibody-coated sheep RBCs. Assays for specific complement proteins are also available.

Prophylactic antibiotics may have a role in treatment. Vaccination with polysaccharide vaccines is also recommended.

Phagocyte Deficiencies

In patients with CGD, white cells can move and phagocytose normally but cannot kill intracellular bacteria. This impairment is due to an inability of phagocytes to form reactive oxygen intermediates. The impaired formation of reactive oxygen species results from functional defects in the NADPH oxidase enzyme complex. NADPH oxidase normally produces reactive oxygen intermediates, which then serve as signaling molecules to activate granule proteins with microbicidal activity. The NADPH oxidase complex consists of 2 membrane-bound components designated gp91 and p22 and 4 cytosolic components designated p47, p67, p40, and Rac.

Four genetic variants of CGD are described. Two thirds of cases are X-linked and are characterized by abnormal gp91, which is encoded by a gene on chromosome band Xp21. Roughly one third of cases are transmitted in autosomal recessive fashion.

Patients with CGD have recurrent skin, lung, bone, and soft tissue infections. Other common complications include pneumonitis, conjunctivitis, adenitis, rhinitis, sepsis, cellulitis, and meningitis. Recovered organisms include Staphylococcus aureus (liver abscesses with staphylococci are nearly pathognomonic for CGD), Serratia marcescens, Burkholderia species, Nocardia species, and Aspergillus fumigatus. Physical examination may reveal hepatosplenomegaly, evidence of intestinal or genitourinary obstruction due to exuberant granuloma formation, and short stature.

Laboratory evaluation reveals hypergammaglobulinemia because of an ineffective humoral response. Cellular immunity is normal. NBT is a yellow dye that is reduced to an insoluble blue-purple form in normal cells. In phagocytes affected by CGD, no reduction occurs, and the yellow color remains. An accurate quantitative respiratory-burst assay for superoxide production, which relies on flow cytometry, has largely replaced the NBT test.

Available treatments include subcutaneous interferon-γ, which attenuates the severity of infections. The use of prophylactic antimicrobial agents, including trimethoprim-sulfamethoxazole, is recommended. The effect of Aspergillus pathogen has led to the use of prophylactic antifungals, such as itraconazole. Steroids can be used to shrink granulomas that cause luminal obstruction but may increase the risk of fungal infections.

HSCT has also been used. Seger et al (2002) reviewed their experience with HSCT for the treatment of CGD.²² An HLA-identical sibling donor was used in 27 individuals who underwent HSCT for CGD. More than 80% of patients survived and were cured. Leung et al reported that 9 of 13 patients who received a transplant were alive with follow-up at 14 months and 6 years.²³  One patient died of sepsis, and one died of complications of BMT. Two were alive and experienced a relapse of their disease. Of 24 patients who received an HLA-identical sibling transplant, 20  were alive upon follow up at 1-7 years.²⁴

Leukocyte involvement in host defense is a complex process that involves rolling, adhesion, and diapedesis. Many molecules mediate these processes; selectins and integrins are critical. Many defects in these pathways result in attenuated neutrophil recruitment and poor inflammation collectively known as LAD. Several forms of LAD are recognized.

LAD1 is an autosomal recessive disorder characterized by deficiency of CD18, which is encoded by a gene on chromosome band 21q22.3 and is the beta chain common to several complexes of leukocyte surface integrin. The integrin complex also includes CD11. Decreased expression of CD18 leads to low expression of CD11a, CD11b, and CD11c. Deficiency of CD18 renders the cell membrane unable to adhere normally. As a result of defective adhesion, neutrophils fail to mobilize and migrate to sites of tissue injury. These adhesion defects also decrease granulocyte adherence to each other, they decrease complement mediated phagocytosis, and they decrease antibody-mediated cytotoxicity.

The classic clinical presentation is an infant whose detachment of the umbilical cord is delayed. Severe infections occur and may include soft tissue, sinopulmonary, and perirectal infections. Dental disease, including periodontal and gingival disease, may also occur. Each form of LAD has characteristic phenotypes including findings of chronic neutropenia and bleeding.

Diagnosis is based on flow cytometry to assess for CD18 expression. However, CD18 expression may be normal with abnormal CD18 function. The complications of LAD can be reduced with adequate antibiotic therapy. Transplantation is also a consideration.

LAD includes LAD2, which is an autosomal recessive disorder of fucose metabolism. Mutations in the guanosine diphosphate-fucose transporter gene decreases expression of fucosylated proteins that function as selectin ligands. In addition to immune dysregulation, patients have severe mental retardation, short stature, characteristic facies, and the Bombay blood phenotype. The diagnosis is based on flow cytometry. The use of oral fucose replacement has been attempted.

Other forms of LAD include defective selectin expression on endothelial cells and mutations involving the guanosine 5'-triphosphatase (GTPase) Rac2, which is involved in actin cytoskeletal regulation and NADPH oxidase function.

Shwachman-Diamond syndrome is a rare autosomal recessive syndrome, with over 95% of cases having alterations in the 7q11 chromosome in the SBDS gene. Most mutations are due to gene conversion between the SBDS gene and an adjacent pseudogene. All known patients are heterozygous because some SBDS protein is required. SBDS functions in ribosome formation (60S subunit). The reason for bone marrow failure and neutropenia remains unclear. Alter (2007) and Ganapathi et al (2007) reported that levels of unbalanced production of ribosome precursors might contribute to increased apoptosis.^26,27

The most common clinical findings are recurrent infections, failure to thrive, pancreatic exocrine insufficiency, and skeletal abnormalities secondary to skeletal dysplasia. The skin, liver, heart, and kidney can also be affected. Shwachman-Diamond syndrome is the second most common cause of exocrine pancreatic insufficiency after cystic fibrosis. A spectrum of bone marrow failure (most commonly neutropenia with no associated monocytosis) has been associated with the syndrome.²⁸ Defects in myeloid progenitors with subsequent alterations in granulocyte mobility, migration, and chemotaxis have been described. Associated defects in lymphoid progenitors with subsequent T-cell and B-cell dysfunction are also described. NK cell function may be affected.

Patients with Shwachman-Diamond syndrome have a 10-25% incidence rate of pancytopenia, a 20% incidence rate of aplastic anemia, a 20-33% incidence rate of myelodysplasia, and 12-25% incidence rate of acute leukemia.²⁹  Since 1964, 300 cases of Shwachman-Diamond syndrome have been reported Exocrine pancreatic insufficiency is characterized by pancreatic fluid without trypsin, lipase, or amylase activity.

Treatment for this disorder consists of pancreatic-enzyme replacement therapy and antibiotic therapy. G-CSF is only used in patients with persistent severe neutropenia and recurrent serious infections. Although patients do respond to GCSF, the risk of malignant transformation and the functional defects of neutrophils in Shwachman-Diamond syndrome make it less than ideal.²⁸ Projected median survival is 35 years but is only 14 years with the development of aplastic anemia and 9 years if leukemia develops.

As of 2002, BMT had been used in only 15 patients.²⁹ Most of these patients had already progressed to myelodysplasia or marrow aplasia; 9 patients died after BMT (3 relapsed). Transplant-related mortality was 40%. The range of follow-up was 9 months to 3 years. No increase in morbidity when compared with other patients receiving BMT. Patients who developed myelodysplasia or leukemia had a significantly worse outcome than those with just bone marrow aplasia.

Chediak-Higashi syndrome is a rare autosomal recessive syndrome. Chediak-Higashi syndrome is caused by mutations of the CHS1 gene , also known as the lysosomal trafficking regulator, which is involved in the regulation of packaging and sorting of intracellular proteins. CHS1 is located on chromosome 1q42-44. Disruption of this gene results in abnormal fusion of intracellular granules.

In affected individuals, neutrophils, eosinophils, basophils, and platelets are characterized by large blue-gray granules in their cytoplasm. In affected neutrophils, this change leads to a deficiency or absolute lack of some granule proteins. This lack may be partly responsible for the abnormalities of neutrophil chemotaxis and neutrophil microbicidal activity observed in individuals with this disorder. Patients have deficiencies in T-cell cytotoxicity and NK cell activity, as well as decreased bacterial killing by monocytes and polymorphonuclear leukocytes.

Other cell types may also be affected. Dense bodies may be reduced in the platelets, a finding which may explain the easy bruising and prolonged bleeding time that is common in affected patients. A lack of NK cell function and lymphocyte function may be observed. Schwann cells, melanocytes, thyroid follicular cells, and renal tubular cells may also be affected.

Other pathognomonic findings associated with this disorder include oculocutaneous albinism due to abnormal melanin granules transport. Patients may also have cranial neuropathies, peripheral neuropathies, and hepatosplenomegaly. Patients with Chediak-Higashi syndrome have frequent bacterial infections, usually involving the skin or respiratory tract. Patients may also develop a lymphoproliferative phase, termed an accelerated phase, which is a common cause of death. This is typically associated with Epstein-Barr virus that causes hemophagocytosis. This form of hemophagocytic lymphohistiocytosis is best treated with regimens noted above.

Aggressive treatment of infections is critical, although the use of antibiotic prophylaxis is unclear. HSCT has been used to treat patients with this disorder in both the chronic and accelerated phases. Haddad et al (1995) summarized their experience with HSCT for Chediak-Higashi syndrome.³⁰ Six of seven patients who received HLA-identical transplants were alive and well at median follow-up of 6.5 years. Eapen et al (2007) reviewed outcomes after allogeneic hematopoietic cell transplantation in 35 children with Chediak–Higashi syndrome.³¹ With a median follow-up of 6.5 years, the 5-year probability of overall survival is 62%. Mortality was highest in those with accelerated phase disease at transplantation and after alternative-related donor hematopoietic cell transplantation. Thus, early transplantation in remission after accelerated phase disease appears to improve disease-free survival.

Griselli Syndrome

Griselli syndrome is an autosomal recessive syndrome that involves partial albinism, silver hair, frequent infections, cellular immunodeficiency, and neurologic abnormalities, with early demise due to uncontrolled activation of lymphocytes and macrophages.

Defects of MYO5A or RAB27A genes on chromosome 15q21 are responsible. MYO5A and RAB27A lead to defective transport of melanosomes to surrounding keratinocytes. Only MYO5A is present in brain tissue. RAB27A is associated with abnormal lymphocyte cytotoxic activity; defective NK cell cytotoxicity and defective T-cell cytotoxicity are seen. Progressive neurological signs also occur. 

Treatment of the accelerated phase includes steroids, antithymocyte globulin, cyclosporin, etoposide, and, possibly, intrathecal methotrexate. Other chemotherapy agents have not been helpful. Allogeneic BMT is only curative for a RAB27A defect and prevents recurrence of the accelerated phase. No treatment is available for the MYO5A defect.^32,33,34

Neutropenia

Neutropenia is defined as a neutrophil count of less than 1500 cells/mcL, but this varies with age and ethnicity. The lower limit of normal for neonates and infants aged 2 weeks to 1 year is 1000 cells/mcL. African Americans have neutrophil counts lower than those of other ethnic groups, with a lower limit 100-200 cells/mcL below the reference ranges described above.

The absolute neutrophil count (ANC) is calculated by multiplying the WBC count by the percentage of neutrophils (segmented neutrophils plus band neutrophils). An ANC of less than 500 cells/mcL is considered severe neutropenia. An ANC of 500-1000 cells/mcL is considered moderate neutropenia, and an ANC of 1000-1500 cells/mcL is considered mild neutropenia. The risk of infection increases as the ANC decreases. Severe infections are common when the ANC is less than 500 cells/mcL. Other factors associated with an increased risk of infection may include the cause and duration of neutropenia.

Mechanisms involved in neutropenia may include decreased production of neutrophils, increased consumption of neutrophils as a result of destruction or sequestration, dilution of neutrophils, or any combination of these mechanisms.

Neutropenia may also be considered primary or secondary in origin. Secondary causes are most common and are related to cytotoxicity, immune-mediated destruction, or infection. Drugs, including chemotherapeutic agents, decrease neutrophil counts. Infections with various viral pathogens may directly impair myelopoiesis or lead to sequestration of neutrophils. These pathogens may also be instrumental in causing immune-mediated destruction. Examples include parvovirus, Epstein-Barr virus, and human immunodeficiency virus (HIV). Autoimmune neutropenia can be seen in various disorders that are autoimmune in nature, including celiac disease. Overwhelming infection and sepsis are also secondary causes of neutropenia.

Cyclic neutropenia

Cyclic neutropenia is characterized by a history of recurrent infections and neutropenia approximately every 21 days. Cyclic neutropenia has a genetic basis in 25% of cases and is commonly autosomal dominant in inheritance. Cyclic neutropenia is associated with the ELA2 gene, which encodes neutrophil elastase, a glycoprotein manufactured and stored in promyelocytes and myelocytes. Both the sporadic and autosomal dominant forms have coding sequence mutations of neutrophil elastase (ELA-2) a serine protease synthesized during the promyelocyte stage. 

The link between ELA2 mutations and cyclic neutropenia is unclear; however, ELA2 mutations and abnormal localization of neutrophil elastase may be linked to apoptosis of differentiating cells, which has been documented.³⁵ Neutrophil elastase is synthesized early in neutrophil development by CD34+ cells. The neutrophil elastase is packaged in the azurophilic granules and cycling is the result of the interrupted flow of cells down the pathway of neutrophil differentiation. Mutations in the gene for neutrophil elastase are present in more than 90% of patients, and this condition shows accelerated apoptosis. The neutrophil elastase may leak out of the reticuloendothelium system and kill the cells.

The severity varies, but typical complications during the neutropenic period include fever, gingivitis, stomatitis, cellulitis, and perirectal abscess formation. Death from overwhelming infection, sepsis, and peritonitis, is unusual but does occur in approximately 10% of patients.

An interval of about 21 days between the occurrence of signs and symptoms of infection suggests cyclic neutropenia. Cycles can vary in periodicity from 14-36 days; approximately 70% of patients have a 21-day cycle. Associated neutropenia can persist 3-10 days before recovery.

Cyclic neutropenia is confirmed by performing serial CBC counts 2-3 times per week for approximately 2 months while other causes of neutropenia are excluded. The diagnosis is confirmed by identification by ELA2 mutations. Bone marrow biopsy findings may reveal hypoplasia or arrested maturation at the myelocyte stage of development. Findings in the recovery phase commonly reveal granulocyte hyperplasia. The administration of G-CSF has clearly benefitted patients with cyclic neutropenia. Management of symptomatic patients include use of antibiotics and G-CSF.

Severe congenital agranulocytosis, also known as Kostmann syndrome, is characterized by the early onset of severe neutropenia. Life-threatening pyogenic infections often complicate severe congenital agranulocytosis. Various types of inheritance, including autosomal recessive or autosomal dominant inheritance, have been reported, although most cases are sporadic. The hematopoietic system has reduced bone marrow progenitor colony-forming potential, bone marrow stromal abnormalities, increased bone marrow apoptosis, and defects of neutrophil chemotaxis.²⁸

This disorder is associated with mutations that involve the ELA2 gene, occurring in approximately 60% of cases.³⁶ Mutations in ELA2 with abnormal intracellular targeting of neutrophil elastase occur in the most severely affected patients with lower neutrophil counts and more vulnerability to myelodysplasia and acute myeloid leukemia.³⁷ The mutations are more diverse than the mutations in cyclic neutropenia. ELA2 mutations may cause incorrect folding of nonfunctional neutrophil elastase proteins, which accumulate in the cytoplasm and induce apoptosis. 

In other patients, mutations of the HAX1 gene in chromosome 1 have been reported.³⁸ The protein produced is a member of the BCL2 family of apoptosis-regulating proteins. HAX1 is critical for maintenance of the inner mitochondrial membrane potential and protects myeloid cells from apoptosis. Other gene defects (GFI1, AP3B1, WAS, TAZ1, and MAPBPIP have been discussed, but the mechanism for these neutropenias is unknown.  G-CSF abrogates mitochondrial-dependent apoptosis of bone marrow cells.³⁸  Mutations in GFII and WASP have also been associated with severe congenital agranulocytosis. These mutations may lead to aberrant transcription of ELA2 and abnormal packaging of neutrophil elastase, respectively. 

Although both severe congenital neutropenia and cyclic neutropenia both have mutations in the ELA2 gene and despite the fact that patients with cyclic neutropenia do not have an excess of acute myeloid leukemia conversion, the pathogenic mechanisms still need to be elucidated. 

Life-threatening infections occur in the first year of life and may include cellulitis, perirectal abscess formation, meningitis, peritonitis, and sepsis. Isolated organisms may include Staphylococcus and Pseudomonas species and Escherichia coli. Oral signs are common and may include aphthous ulcers, gingivitis and stomatitis. Many patients may become edentulous. Patients with congenital agranulocytosis are at increased risk for myelodysplastic syndrome or acute leukemia.

Point mutations in the gene for the G-CSF receptor CSF3R have been implicated in the progression of severe congenital neutropenia to leukemia. Germeshausen et al reported that, of 23 patients with congenital neutropenia with signs of malignant transformation, 18 (78%) were shown to harbor a CSF3R mutation.¹² More than 90% of these patients respond to G-CSF. However, during long-term treatment, an increasing number of patients acquire mutations in the G-CSF receptor and develop myelodysplasia with monosomy 7 that progresses to myeloid leukemia. The cumulative incidence of malignant transformation is 21% after 10 years of treatment. The cumulative risk after 8 years of treatment was 13%.³⁶  The group aged 10-19 years had a greater proportion of conversions than any other age group. 

Clinically, patients are shorter than their peers, and, by age 10 years, 50% have heights lower than the 10th percentile.³⁹ Patients also have osteopenia and osteoporosis, as well as an increased frequency of autoimmune complex disease.

Laboratory studies demonstrate severe neutropenia at birth, with neutrophil counts of less than 100-300 cells/mcL. Peripheral monocytosis and eosinophilia are common. Anemia may develop secondary to chronic inflammation and illness. The platelet count is usually normal or increased. Examination of the bone marrow reveals myeloid hyperplasia with maturational arrest at the promyelocyte stage of development. Treatment may consist of G-CSF, prednisone, or HSCT. Treatment with GCSF may lead to splenomegaly.

The International Neutropenia Registry has reported that 95% of patients have an increase in the ANC to more than 1.0 x 10⁹/L. Complications of G-CSF include splenomegaly, hepatomegaly (20% of patients), osteoporosis, and vasculitis (≤4%). As of 2003, 57 of 348 patients died; the median age of death was 12 years, and the leading cause of death was leukemia. 

Although glucocorticoids may transiently elevate the WBC count, they are not suitable for long-term use due to side effects. HSCT has been used with success.⁴⁰  Of the 101 patients in the International Neutropenia Registry, 9 have received BMT. Of these 9 patients, 8 engrafted, and 3 died. Six patients are alive and in complete remission. Only 2 received stem cells form related donors. Infections, including GVHD, are the main causes of death. 

However, the results are superior if the patients receive transplantation before they develop myelodysplasia or acute myeloid leukemia; 17 of the 24 patients in the registry who underwent transplantation after developing these conditions died. In contrast, 10 of the 12 patients transplanted for failure to respond to G-CSF before they developed myelodysplasia survived. Eight of the 10 that survived had an HLA-identical sibling. Mino reported a successful cord-blood transplant for severe congenital neutropenia.⁴¹  

Incidental neutropenia

Incidental neutropenia refers to a healthy child with neutropenia on a CBC count obtained for reasons other than an evaluation for neutropenia. This finding requires thorough history taking and physical examination. A complete history should focus on present or past illnesses or infections, along with a thorough birth and family history. Focus should also be placed on any past or current medications. Careful physical examination should also be completed, with documentation of the presence or absence of gingivitis, mouth ulcers, cellulitis or perirectal abscesses.

In a study of 387 healthy children, Moyer and Grimes found that nearly 75% had CBC counts outside of the reference range with no demonstrable abnormalities on repeat examinations.⁴² About 10% of the children had laboratory evidence of neutropenia. When interpreting the laboratory data for neutropenia, clinicians should consider the patient's ethnicity and other relevant clinical information. Neutropenia is common and is frequently of little or no consequence.

Laboratory evaluation should not reveal any evidence of anemia or thrombocytopenia. Involvement of other cell lines should suggest failure or infiltration of the bone marrow. Minor infectious illnesses may cause thrombocytopenia and neutropenia because the circulating half-lives of platelets and neutrophils are short. Transient marrow suppression can cause clinically significant deficiencies of platelets and neutrophils.

Incidental neutropenia in a newborn may reflect alloimmune sensitization in which maternal antibodies to fetal neutrophil antigens cross the placenta and lead to the destruction of fetal neutrophils. This disorder can be diagnosed by identifying granulocyte-specific alloantibodies in the patient or maternal serum. Affected infants may require antibiotic therapy in the presence of infection. Severe infections or sepsis may require the administration of G-CSF.  

Chronic neutropenia

Chronic neutropenia is defined as neutropenia that lasts longer than 6 months. The entity of chronic benign neutropenia of childhood is a heterogenous disorder. Rajantie et al estimated that chronic benign neutropenia of childhood occurs with an incidence of about 4 cases per 100,000 children, with an equal distribution between boys and girls.⁴³ The course of disease varies among individuals. The median duration of chronic benign neutropenia of childhood is estimated at 20 months, with most patients recovering by age 4 years. The likelihood of spontaneous remission may decrease with increasing age.

One common cause of chronic neutropenia during infancy and early childhood is primary autoimmune neutropenia (AIN). The etiology is unclear, and no specific hereditary mechanism has been identified. The female-to-male ratio is 3:2, and the incidence is 1 case per 100,000 population. Median age of diagnosis is 12 months. The diagnosis of primary AIN is made when circulating anti-NA1 and anti-NA2 granulocyte antibodies are detected. Antineutrophil antibodies may interfere with the bactericidal capabilities of the neutrophil, including phagocytosis, adhesion, opsonization, and microbicidal activity. Antibodies directed against granulocyte precursors instead of mature neutrophils may be associated with a severe clinical course.

Bux and colleagues found a close relationship between the development of NA1 antibodies and HLA-DR2 in infants with AIN.⁴⁴ Periodic measurement of antineutrophil antibody titers may help in predicting the course of AIN, but the clinical usefulness of such information is not confirmed. AIN is typically benign and self-limited. The mean duration of the disorder is 20 months from diagnosis, and 95% of patients recover by age 4 years, with recovery defined as the disappearance of the antineutrophil antibodies, combined with at least 3 consecutive CBC counts in which the ANC is more than 1500/mcL.

Repeated infections can be treated with G-CSF and continuing routine supportive care and treatment. The use of oral corticosteroids and IVIG has been described, but the results are not consistently good. The best treatment is the use of prophylactic antibiotics (eg, Bactrim Septra) and appropriate antibiotics for each infection. In cases with severe infections or repeated low-dose G-CSF may be helpful. No patient with this diagnosis  has been reported to develop myelodysplasia or acute myeloid leukemia.  

Keywords

primary immunodeficiency diseases, immunology, neutrophil function, white blood cell disorders, white blood cell function, neutropenia, T cells, B cells, innate immunity, adaptive immunity, WBC, selective immunoglobulin A deficiency, transient hypogammaglobulinemia of infancy, immunoglobulinemia subclass deficiencies, impaired polysaccharide responsiveness, X-linked hyperimmunoglobulin M, X-linked agammaglobulinemia, Bruton agammaglobulinemia, Bruton agammaglobulinemia, DiGeorge syndrome, DiGeorge's syndrome, mucocutaneous candidiasis, hyperimmunoglobulin E, Job syndrome

Job syndrome, chronic granulomatous disease, reticular dysgenesis, Chediak-Higashi syndrome, Shwachman-Diamond syndrome, congenital agranulocytosis, Kostmann syndrome, Kostmann syndrome, neutropenia, complement defect, leukocyte adhesion defect, LAD, common variable immunodeficiency, CVID, Wiskott-Aldrich syndrome, WAS, severe combined immunodeficiency, SCID, adenosine deaminase deficiency, ADA, nucleoside phosphorylase deficiency, PNP, acquired agammaglobulinemia, ataxia telangiectasia, IgA deficiency, chronic mucocutaneous candidiasis, CMC, hyper-IgE syndrome, ZAP-70 defect, familial hemophagocytic lymphohistiocytosis, FHL, severe congenital agranulocytosis

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