Immunology (Lippincott Illustrated Reviews Series) 2nd Edition

Chapter 9: Lymphocyte Development

I. OVERVIEW

Epitope-specific T-cell and B-cell receptors (TCRs and BCRs) are randomly generated within individual thymus- and bone marrow–derived lymphocytes by gene rearrangement. Not surprisingly, some lymphocytes develop receptors that react with self epitopes. However, a selection mechanism is in place that removes these cells before they become fully functional and attack the body’s own and tissues. The adaptive immune system carefully regulates the development and differentiation of lymphocytes to prevent the maturation of self-reactive T and B cells.

Arising from hematopoietic stem cells in the bone marrow, a common lymphoid precursor (CLP) of lymphocytic lineage cells either differentiates within the thymus (T-cell lineage) or remains in the bone marrow (B-cell lineage). In addition, B- and T-cell lineages have major subdivisions (Table 9.1; also see Fig. 4.2). T lineage cells differentiate within the thymus along one of three developmental pathways: those that express αβ TCRs, those that express γδ TCRs, and those that share functional properties of natural killer (NK) cells. Thymic “graduates” in each of these groups can be differentiated from one another in several ways: the extent of their thymic “education,” their arrival and departure from the thymus, the diversity or “repertoire” of their TCRs, their geographic distribution within the body, and the nature of their responses to different categories of epitopes. B-cell lineages, designated B-1 and B-2 cells, can be distinguished from one another by many of the same characteristics used for categorizing the T cell subgroups.

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II. T-CELL LINEAGE

T cell precursors, known as prothymocytes, migrate from the bone marrow to the thymus attracted by thymic molecules (e.g., lymphotactin). Prothymocytes entering at the cortical region, now properly called thymocytes, lack TCRs, CD3, CD4, and CD8 surface molecules. The newly arrived thymocytes rapidly acquire TCRs, CD3, and both CD4 and CD8 molecules. Thymocytes must then “run a gauntlet” of selective tests as they migrate from the thymic cortex to the medulla. The selection processes are so demanding that only an estimated 1% to 5% of all thymocytes “graduate” as T cells. The other 95% to 99% either leave the thymus before undergoing selection (e.g., γδ T cells) or die an apoptotic death after failing one of the selective tests.

A. Thymus structure

By the end of the third gestational month, the bilobed thymus is increasingly populated by lymphocytes, organized into a denser outer region or cortex and looser inner region or medulla (Fig. 9.1). A connective tissue capsule within ward-extending trabeculae surrounds the thymus. Additional cell types within the thymus are epithelial reticular cells, an inclusive term that includes several cells types such as dendritic cells, macrophages, and epithelial cells that serve as “instructors” for the thymocytes as they complete their education. The epithelial reticular cells, as well as those organized to form concentric rings called Hassall’s corpuscles, express major histocompatibility molecules and secrete hormones associated with thymocyte differentiation. Medullary post capillary venules are important for the egress of thymic graduates or T cells from the thymus.

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Figure 9.1

Thymocyte development: Positive and negative selection. A. The thymus is organized into outer or cortical and inner or medullary regions. Prothymocytes enter the thymus to both increase in number and under several maturational steps. Thymocytes that “pass” exit from the thymus via postcapillary venules in the medulla. Thymic graduates are known as thymus-derived lymphocytes or T cells. Those thymocytes that fail their “tests” die. B. Prothymocytes migrating from the bone marrow enter the cortical region of the thymus. As they migrate from the cortical to the medullary regions, they begin to express T-cell receptors and other necessary accessory molecules. The γδ thymocytes exit quickly from the thymus, whereas the αβ thymocytes remain. Within the cortex, αβ thymocytes undergo positive selection. The survivors of positive selection then undergo a negative selection to remove cells that are potentially autoreactive.

B. αβ T cell development

Prothymocytes enter the subcapsular region of the thymus from the circulation, where they proliferate. These newly arrived cortical thymocytes are called double negative (DN) cells because they do not express CD4 or CD8 molecules (or TCR or CD3 complex molecules). They soon begin to generate and express αβ TCRs, associated CD3 complex, and both CD4 and CD8 molecules, as well as receptors and adhesion molecules important for their interaction with other cells and their migration through the thymus. Because they express both CD4 and CD8 surface molecules, these immature thymocytes are called double positive (DP) cells. In a process known as positive selection, DP thymocytes die within 3 to 4 days unless they recognize and bind to major histocompatibility complex (MHC) or to peptide + MHC (pMHC) molecules expressed by certain epithelial reticular cells (cortical epithelial cells) in the cortex. This process eliminates thymocytes that are incapable of recognizing self MHC. Cells that pass the positive selection test, located at the corticomedullary junction, are allowed to enter the medulla; those that fail the test die. DP cells whose CD8 molecules have engaged pMHC I then cease the expression of CD4 molecules and become single positive (SP) CD8+ cells. Likewise, those that are bound to pMHC II cease expression of CD8, becoming SP CD4+ cells (Fig. 9.2).

Survivors of positive selection then run a second gauntlet called negative selection when they arrive at the corticomedullary junction. There, they meet and interact with a second set of epithelial reticular cells (antigen-presenting cells such as dendritic cells and macrophages). Those that efficiently bind to self peptides of the pMHC I or pMHC II on these APCs are potentially autoreactive and undergo apoptotic death. Thymocytes that pass both positive and negative selection tests “graduate” from the thymus, entering the circulation through the medullary postcapillary venules as T cells. Each developmental stage is closely controlled by substances secreted by epithelial reticular cells that regulate gene expression within the thymocytes. For example, secretion of the cytokine interleukin-7 (IL-7) by epithelial reticular cells activates the genes that control the early stages of thymocyte development. Failure of early thymocytes to express IL-7 receptors terminates their development.

C. γδ T cell development

The thymus is also the differentiation site for thymocytes that express γδ TCRs and CD3 complex molecules. Many of these cells fail to express CD4 and/or CD8. Consequently, they do not undergo the same positive and negative selective processes as αβ TCR-bearing thymocytes and depart from the thymus shortly after developing their TCR complexes (Tables 9.1 and 9.2, Fig. 9.1). γδ Cells are thought to be a transitional cell type that may represent a bridge between the innate and adaptive immune systems. γδ T cells develop early in embryogenesis before many αβ T cells and migrate preferentially to the respiratory organs, the skin, and the peritoneal cavity. They use a very limited set of V, D, and J genes in the generation of the variable regions of the γ and δ chains and, thus, are much more limited in their recognition repertoire than are αβ T cells. They respond more quickly than do αβ T cells, but they do so without generating memory.

D. NKT cell origin

Natural killer T (NKT) cells are a distinctive subset of T cells that share some characteristics with NK cells. They express several surface markers and receptors found on NK cells, but unlike NK cells, they undergo some development in the thymus and express TCRs generated by DNA rearrangement and junctional diversity. NKT cells express TCRs that are extremely limited in repertoire and are predominantly specific for lipids, glycolipids, and a few specialized types of peptides. Their TCRs have an unusual restriction pattern. Although they may be either CD4+ or CD4+CD8+, they specifically recognize epitopes presented by a “nonclassical” MHC class I molecule called CD1d. The nonclassical class I molecules, encoded by genes located in a chromosomal segment adjacent to the HLA complex, appear to present epitopes (often nonpeptide in nature) to T cells other than the most abundant αβ type.

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Figure 9.2

Development of CD4+ and CD8+ αβ T cells. The type of MHC molecule or peptide + MHC (pMHC) molecule to which a double-positive (DP) CD4+CD8+ binds determines its adult phenotype. Thymocytes that fail to bind to pMHCI or pMHC II die.

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III. B-CELL LINEAGE

In humans, progenitors of immunoglobulin-producing cells are found in the yolk sac by the third week, in the fetal liver by the eighth week, and in the bone marrow by approximately the twelfth week of gestation. These cells are called bone marrow–derived lymphocytes or B cells because this is where most of these cells differentiate. B cells are defined as cells that synthesize immunoglobulin and display it on their cell surfaces as their BCRs.

A. Bone marrow

The bone marrow contains connective tissue, blood vessels, fat, and cells. Among these structures are the hematopoietic stem cells capable of giving rise to the stem cells of the myeloid, granuloid, erythroid, and lymphoid cells (see Chapter 4). The vasculature provides an efficient route for cells originating in the bone marrow to move into the periphery and for the reentry of activated, matured immune cells (e.g., plasma cells) from the periphery. Unlike lymphoid cells that are destined to differentiate into T cells, those committed to the B cell lineage remain within the bone marrow for development.

B. B cell development

B cell development reflects the stages (also called bone marrow fractions) of immunoglobulin heavy and light chain rearrangement (see also Chapter 8) and surface expression (Fig. 9.3).

• Arising from a common lymphoid progenitor (CLP), the earliest identifiable cell committed to the B-cell lineage is the pre–pro-B cell (Fraction A), within which the cell begins to express Igα and Igβ BCR accessory molecules.

• Immunoglobulin DJ gene joining and cytoplasmic expression of surrogate light chain (SLC) occurs at the early pro-B cell (Fraction B) stage followed by VDJ gene joining and cytoplasmic SLC expression at the late pro-B cell (Fraction C) stage.

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Figure 9.3

B cell development.

• The early pre-B cell (Fraction C-prime or C´) stage is characterized by the surface expression of pseudo-IgM (rearranged μ heavy chains plus SLC) and is accompanied by a burst of cellular proliferation.

• In the late pre-B cell (Fraction D) stage, immunoglobulin light chain kappa (κ) or lambda (λ) genes rearrange, and their products (κ or λ light chains) replace the SLCs.

• Immature B cells (Fraction E) express μ heavy chains plus κ or λ light chains on their cell surfaces.

• Mature B cells (Fraction F) coexpress IgM and IgD on their cell surfaces. As they pass through the developmental stage, B cell progenitors, like thymocytes, express molecules and receptors necessary for migration and interaction with other cells.

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Figure 9.4

Characteristics of B-1 and B-2 B cells.

Some attributes (e.g., DNA recombinase expression) are lost by the time cells reach the immature B cell (Fraction E) stage. If the IgM on the developing cells binds to epitopes they encounter in the bone marrow, such cells undergo apoptotic death to prevent production of autoreactive B cells.

C. B-1 and B-2 B cells

Two developmentally distinct B cell pathways are currently recognized (Fig.9.4). Conventional B cells (B-2 B cells) are widely distributed throughout the body, require interaction with T cells for their activation and proliferation, and are continually replaced from the bone marrow throughout adult life. The range of epitopes that can be recognized by B-2 B cells is vast. Upon repeated antigen exposure, B-2 B cells respond quickly with increased antibody quantity and quality, often by “fine-tuning,” the affinity of the antibody produced (affinity maturation; see Chapter 8). B-2 B cell responses are often accompanied by a change in immunoglobulin isotype. All of these properties are hallmarks of immunologic memory. Typically, more IgD than IgM is expressed on the surfaces of mature B-2 (Fraction F) B cells.

Appearing early in embryogenesis, B-1 B cells, arise from the fetal liver by the eighth gestational week. They might represent a transitional type of lymphocyte that bridges the innate and adaptive immune systems. First described about two decades ago, B-1 B cells have an importance in innate-related immunity and in autoimmune disorders that has become increasingly recognized. The B-1 B cell repertoire is quite limited in comparison to that of B-2 cells. B-1 BCRs and B-1 B antibodies are often directed against conserved microbial antigens (e.g., carbohydrates). It is thought that most, if not all, natural antibodies (e.g., IgMs directed against the A and B blood groups that exist in the absence of known immunization) are of B-1 B cell origin. B-1 B cells are found predominantly in tissues that are potential portals of microbial entry (e.g., the peritoneal cavity and respiratory tract) and are a self-renewing population within these tissues. Although they show little if any immunologic memory, limited isotype switching, and limited repertoires, they contribute greatly to protective immunity. It is estimated that more than half the IgA secreted into the mucosa is of B-1 origin.

Chapter Summary

• Arising from hematopoietic stem cells in the bone marrow, lymphocytic lineage cells differentiate either within the thymus (T cells) or remain in the bone marrow (B cells).

• T cell precursors, known as prothymocytes, migrate from the bone marrow to the thymus attracted by thymic molecules (e.g., lymphotactin). The bilobed thymus is increasingly populated with lymphocytes, organized into a denser outer region or cortex and a looser inner region or medulla.

• In the thymus cortex, T cells begin to generate and express T-cell receptors (TCRs), CD3 molecules, and sets of receptors and adhesion molecules. At this time, they begin to simultaneously express both CD4 and CD8 molecules and are known as “double positive” (DP) cells. Those thymocytes that generate and express γδ TCRs also express CD3, but many of them fail to express CD4 and/or CD8. The DP αβ thymocytes undergo a set of selective processes referred to collectively as “education,” in which the immune system begins to screen them on the basis of their ability to recognize self.

• The thymocyte population that moves from the cortex into the medulla consists of a mix of CD4+ and CD8+ cells. Fewer than 5% of thymocytes originally entering the thymus survive both positive and negative selection and leave the thymus to enter the body, where they may become activated and participate in various immune responses.

• NKT cells area distinctive subset of T cells that share some characteristics with natural killer (NK) cells.

• B cells undergo their entire developmental process within the bone marrow. Arising from a common lymphoid progenitor (CLP), the earliest identifiable cell committed to the B-cell lineage is the pre–pro-B cell. Immunoglobulin DJ gene joining and cytoplasmic expression of surrogate light chain (SLC) occurs at the early pro-B cell (Fraction B) stage. The early pre-B cell (Fraction C-prime or C´) stage is characterized by the surface expression of pseudo-IgM. Immature B cells (Fraction E) express μ heavy chains plus κ or λ light chains on their cell surfaces. Mature B cells (Fraction F) coexpress IgM and IgD on their cell surfaces.

• Two developmentally distinct B cell pathways are currently recognized. Conventional B cells (B-2 B cells) are widely distributed throughout the body. B-1 B cells represent a transitional type of lymphocyte that bridges the innate and adaptive immune systems. The importance of B-1 B cells in innate-related immunity and autoimmune disorders is increasingly recognized.

Study Questions

9.1. DiGeorge syndrome is an immune deficiency disease resulting from impaired thymic development. Which of the following is/are affected in patients with DiGeorge syndrome?

A. B cell development only

B. complement only

C. NK cell function

D. T cell development only

E. T cell and B cell functions

The correct answer is E. The defective thymic environment inhibits T cell development and function. Because so much B cell activity depends on interaction with T cells, B cell responses will also be impaired. Complement would not be impaired while sparing T and B cell activity. NK cell function should not be affected.

9.2. Negative selection of T cells occurs in the

A. blood vessels.

B. bone marrow.

C. lymph node.

D. spleen.

E. thymus.

The correct answer is E. Negative selection of T cells occurs as they move from the thymic cortex into the thymic medulla. It does not occur at sites outside of the thymus.

9.3. T cell precursors, known as prothymocytes, migrate from the bone marrow to the thymus in response to

A. eotactin.

B. IL-4.

C. IL-5.

D. IL-10.

E. lymphotactin.

The correct answer is E. Lymphotactin is one of the thymic products that help to guide prothymocytes from the bone marrow to the thymus. IL-4, IL-5, and IL-10 are cytokines produced by mature, activated T cells as well as by other cell types. Eotactin guides the movement of eosinophils.

9.4. What will be the fate of an early thymocyte that fails to express IL-7 receptors?

A. apoptotic cell death

B. development as a γδ T cell

C. development as an NKT cell

D. failure to traffic to the thymus

E. maturation along the B-cell lineage

The correct answer is A. Failure to bind IL-7 dooms the developing thymocyte. It will be unable to develop into either an αβ or a γδ thymocyte. This interaction occurs after migration of the thymocytes into the thymus. Thymocytes cannot switch to the B cell developmental pathway.

9.5. γδ T cells

A. contain very extensive antigen recognition repertoires.

B. express surface markers that are also characteristic of NK cells.

C. generate memory when recognizing antigen on multiple occasions.

D. migrate preferentially to respiratory organs, skin, and peritoneal cavity.

E. respond more slowly to antigen than do αβ T cells.

The correct answer is D. γδ T cells are found predominantly in the respiratory organs, skin, and peritoneal cavity. Their recognition repertoire is far less extensive that found in αβ T cells. They do not express significant immunologic memory but do react to antigenic stimuli more rapidly than do αβ T cells.

9.6. NKT cells

A. are usually CD8 single positive cells.

B. bind epitopes presented by MHC class II molecules.

C. express TCRs generated by DNA rearrangement and junctional diversity.

D. recognize carbohydrates and complex proteins.

E. synthesize immunoglobulin and display it on their cell surfaces.

The correct answer is C. NKT cells do express TCRs generated (like those of other T cells) by DNA rearrangement and junctional diversity. They are either CD4+ or CD4+CD8+. Despite this, their TCRs recognize lipid-related molecular fragments presented by the nonclassical class I molecule CD1d. They do not synthesize or express immunoglobulins.

9.7. Pre–pro-B cells

A. contain either κ or λ light chains.

B. demonstrate surface expression of pseudo-IgM.

C. express Igα and Igβ BCR accessory molecules.

D. have VDJ joining of genes.

E. express surrogate light chains.

The correct answer is C. Pre–pro-B cells initially express Igα and Igβ molecules. The synthesis of heavy and light chains (including surrogate light chains) occurs at later stages of development.

9.8. In contrast to B-2 B cells, B-1 B cells

A. appear later in development.

B. function in innate-related immune responses.

C. express more IgD than IgM on their cell surfaces.

D. have a more extensive antigen recognition repertoire.

E. require interaction with T cells for their activation.

The correct answer is B. B-1 B cells appear to be transitional types of lymphocytes whose functions are reminiscent of the innate immune system. B-1 B cells express more surface IgM than IgD and B-2 B cells express more surface IgD than IgM. The B-1 B cell repertoire is more limited, and their need for interaction with T cells is more limited than is seen for B-2 B cells. B-1 B cells appear developmentally earlier than B-2 B cells.