LYMPHOID SYSTEM

1. Introduction and definitions
2. Histology of limphoid system
3. Lymphatics and lymph nodes.
4. Spleen - a filter of blood
5. Unencapsulated lymphoid aggregates of the respiratory trackt and gut
6. Genesis of T and B lymphocytes within central lymphoid organs
7. TABLE II: Some examples of autoimmune diseases.

INTRODUCTION AND DEFINITIONS

A) Function is to provide surveillance - The lymphoid system is a collection of cells and tissues which provides immune defense against disease-causing bacteria, viruses, fungi, parasites, and cancer cells (Fig. 1). A substance that elicits an immune response is called an antigen.

Figure 1: Events that may occur upon antigen stimulation of the cellular and humoral immune systems.

B) Two systems of immunity in vertebrates

1) T-lymphocytes or T-cells mediate cellular immune responses. T-cells that recognize foreign substances are activated, and they differentiate into effector T-cells which bind to and help to eliminate foreign substances.

2) The other major class of lymphocytes, B-lymphocytes or B-cells, are involved in the humoral immune responses. B-cells, when activated by antigen, differentiate into plasma cells (effector B-cells) which produce antibodies. Antibodies are proteins that specifically bind to foreign substances, leading to their elimination. Certain T-cells, called Helper T-cells (T_H), can assist B-cells in their differentiation to effector cells.

3) This dual system provides overlapping protection. Most antigens elicit both humoral and cellular responses.

4) Several specialized accessory cell types assist lymphocytes in immune defense (e.g., neutrophil, basophil, mast cell, macrophage, interdigitating reticular dendritic cell, follicular dendritic cell).

C) The immune system recognizes foreign substances by their molecular features

1) Antibodies recognize antigens; the small region of the antigen to which an antibody binds is called an antigenic determinant or epitope. For proteins, epitopes formed by adjacent amino acids in the primary sequence are called continuous, whereas discontinuous or conformational epitopes are formed by amino acids that are brought into proximity by three dimensional folding of the protein in space. 2) Recognition and binding between antibodies and epitopes are based on molecular complementarity; short-range noncovalent interactions mediate the binding.

Figure 2. Schematized diagram of the binding between an epitope of a macromolecule and two different antibody molecules, one of which binds the epitope with high affinity and the other of which binds with low affinity. The strength of binding is determined by the complementarity of the binding surfaces i.e., between the epitope and the antigen (Ag) binding site provided by each antibody (Ab). L and H denote the light and heavy chain of each antibody. Reprinted with permission from Alberts et al., Molecular Biology of the Cell, 2nd Ed., Figure 18-22, pg. 1017.

2) Antibodies belong to the class of proteins called immunoglobulins (gamma globulins). These are found in blood and lymph, interstitial fluids, and secretions. Different classes of immunoglobulins exist (IgG, IgM, IgA, IgD, and IgE).

Table I: Properties of the major classes of antibody in the human. Reprinted with permission from Alberts et al., Molecular Biology of the Cell, 2nd Ed., Table 18-1, Page 1014.

3) Structure (Fig. 3) of IgG (the most prevalent antibody class) - It is a complex of four polypeptides, joined together by disulfide bonds: two identical light (L) chains and two identical heavy (H) chains. The C-terminal portions of light and heavy chains are termed constant regions because there is very limited variation among different IgG molecules. The N-terminal portions of light and heavy chains are called the variable region. These differ substantially in amino acid sequence among individual antibody molecules. The variable region forms the antigen-binding site of the antibody molecule, the Fc region of the molecule is formed by the C-terminal halves of the two H chains.

Figure 3: An IgG molecule is comprised of two identical heavy (H) and two identical light (L) chains. Note that the antigen-binding sites are formed by a complex of the amino terminal regions of both L and H chains, but the tail and hinge regions (Fc) are formed by the H chains alone. Each H chain contains one or more oligosaccharides (carbohydrate) chains of unknown function. Reprinted with permission from Alberts et al., Molecular Biology of the Cell, 2nd Ed., Figure 18-15, page 1013.

Figure 4. Formation of an antigen-binding site by folding of H and L chains. A particular epitope (determinant) is accommodated by the antigen-binding site formed by this particular pair of H and L chains. Different binding sites would be formed by different combinations of H and L chains. Note that only restricted regions of the H and L chains make actual contact with the epitope.

4) T-cells also have specific cell surface molecules (T-cell receptor) that recognize antigens (Fig. 5).

Figure 5: Structure of the T cell receptor. A receptor molecule includes an alpha chain and a beta chain; each chain has a constant and a variable domain. A transmembrane region, rich in hydrophobic amino acids, anchors the protein in the plasma membrane of the T-cell. The a/b heterodimer is noncovalently associated with an invariant set of membrane proteins called the CD3 complex. Reprinted with permission from Alberts et al., Molecular Biology of the Cell, 2nd Ed., Figure 18-46, page 1037.

D) Clonal selection theory

1) Antigen receptors. The cell surface of lymphocytes express membrane-bound antibodies (B-cells) or T-cell receptors (T-cells) that can bind specific antigens. Each lymphocyte bears one kind of specific receptor that recognizes only a few closely related antigenic determinants. Receptor diversity is generated within this population of cells by DNA rearrangements that occur during maturation of lymphocytes within the central lymphoid organs (thymus for T-cells and bone marrow for B-cells).

Variable regions of immunoglobulins (H and L chains) and T-cell receptors (a and b chains) are encoded by gene segments, each of which specifies part of the variable region. As lymphocytes mature, these gene segments are joined to form a continuous DNA segment that encodes the entire variable region. Different combinations are formed in different cells, each cell thus forming a unique gene for the variable region of each chain (see Figure 6). From a relatively small number of gene segments, many thousands of different receptor chains are possible. Further diversity occurs because each receptor is formed by two different chains i.e., H+L or a +b. This overall mechanism of combinatorial diversity generates a very large number of potential receptors, which underlies the ability of the immune system to respond to a vast array of different antigenic substances. See Alberts, pgs. 1221-1227 for further details.

Figure 6: The V-J joining process that occurs in making a light chain (mouse). V, J, and C denote gene segments encoding regions of the light chain. In 'germ-line' DNA where immunoglobulin genes are not expressed, 4 J segments are separated from one another and from the C gene segment by short introns and from the multiple V gene segments by many kilobases. When B-cells develop, a V-gene segment (V2 in the example shown) is moved next to one of the J gene segments (J3 above) with deletion of the intervening DNA. The extraneous J4 segment and introns sequences are transcribed and then removed during RNA processing. The resulting mRNA codes for the light chain. The multiple possible combinations between V and J segments provides the potential to encode many different light chains. For heavy chains, there is an additional gene segment (D), along with V, J, and C segments, providing for greater combinatorial diversity. Reprinted with permission from Alberts et al., Molecular Biology of the Cell, 1st Edition, Fig. 17-41, pg. 981.

2) Clonal selection. Lymphocytes that bind an appropriate antigenic determinant are triggered to proliferate and differentiate (because of the specific antigen receptors that they express); the activated and dividing cells are termed immunoblasts or blast cells; clones of progeny lymphocytes are formed, each cell bearing the same surface receptors as its parental type. Some of the progeny become terminally differentiated into effector cells.

Figure 7: Diagram of the clonal selection theory. An antigen activates only those T- and B-cell clones already committed to respond to it by virtue of the antigen receptors that they express. The immune system consists of millions of different lymphocyte clones, a subset of which may be activated by any particular antigen.

3) B-cells. Approximately 10⁹ different clones of immunocompetent B-cells are available in secondary lymphoid organs (see below) to respond to an enormous number of different antigens. B-cell effector cells are called plasma cells. They secrete humoral (soluble) antibodies of a single specificity identical to the antigen-recognition specificity of their cell surface receptors. The process of differentiation triggered by antigen takes approximately five days (eight cell generations). A plasma cell has an eccentric nucleus, an extensive cytoplasm with abundant RER, and a large Golgi apparatus. These cells are found in peripheral lymphoid organs (e.g., lymph nodes and spleen) and at sites where immune reactions occur (e.g. lamina propria).

4) T-cells. T-cells can only recognize peptide antigenic determinants (degraded protein antigens) that are physically associated with proteins of the major histocompatibility complex (MHC) on the surface of target cells. There are two principle classes of MHC molecules, called MHC class I and MHC class II.

Figure 8. T-cell receptor recognition of antigen in association with MHC molecules. The antigen consists of peptide fragments. T-cells recognize and respond only to peptide antigens that are present on the surfaces of other cells in association with major histocompatibility molecules (MHC). TCR denotes the two-chained T-cell receptor. Note that recognition involves a three molecule complex: TCR, antigen, and MHC.

There are several classes of T-cells:

Cytotoxic T-cells (T_C) destroy foreign cells directly. Most T_C cells express the surface protein known as CD8. These cells recognize antigen in association with MHC class I molecules.

Helper T-cells (T_H) provide stimuli for B-cell and T_C proliferation and differentiation. Most T_H cells express a surface molecule called CD4. These cells recognize antigen in association with MHC class II molecules. T_H also facilitates activation of macrophages. Activated macrophages are more efficient in phagocytosing and killing microorganisms. T_H cells are a major target of the HIV (AIDS virus). CD4 is one of the actual receptor for binding of HIV. The loss of this class of cells cripples the immune system. Very recently, another class of receptor (chemokine receptor) has been implicated in HIV binding.

Suppressor T-cells (T_S) suppress ongoing or developing immune responses.

5) Immunologic memory. Antigen-driven differentiation of lymphocytes results in formation of both terminal effector cells (short-lived) and memory cells (long-lived). Memory cells retain the ability to respond to the same antigen, by undergoing a blast transformation and eventually producing further progeny cells. As a result of memory, a secondary response to a particular antigen is greater and faster than the primary response. The figure below illustrates the phenomenon of memory in a humoral immune response. Memory also exists for T-cells.

Figure 9: Primary and secondary antibody responses (measured in the serum) induced by a first and second exposure, respectively, to antigen A. Note that the secondary response is faster and greater than the primary response and is specific for A, indicating that the immune system has specifically "remembered" encountering antigen A before. Evidence for the same type of immunological memory is obtained if T-cell mediated responses rather than B-cell antibody responses are measured. Reproduced with permission from Alberts et al., 2nd Ed., Figure 18-9, page 1009.

Figure 10. Generation of memory and effector cells following antigen stimulation.

6) Immunologic tolerance, which is explained as the absence of an immune response to "self" molecules, can be explained by the elimination or suppression of self-reactive clones. For T-cells, this process occurs primarily during their maturation within the thymus (see below).

E) Cooperation of T cells with B cells to produce a humoral immune response

1) Helper T-cells (T_H) are required for differentiation of B-cells to plasma cells for the majority of antigens. Such antigens are referred to as T-cell dependent.

2) T- and B-cells also require accessory cells (e.g., macrophages, interdigitating reticular dendritic cells acting as "antigen-presenting cells (APC)") for cooperation; APC display partially degraded antigen (peptide fragments) on their cell surface in association with MHC molecules for optimal presentation to responsive lymphocytes. In this configuration, the "processed" antigen can optimally stimulate responsive lymphocytes.

3) A model for cooperation between T-cells, B-cells, and APC involves direct intercellular associations (Fig. 11). Soluble signals (called cytokines or interleukins) that promote growth and differentiation are passed between the cells.

Figure 11. A simplified model of cooperation among T-cells, APC, and B-cells.

F) Principal cell types to distinguish in routine histological sections

1) Small lymphocyte

2) Blast cell

3) Plasma cell

4) Macrophage

HISTOLOGY OF LYMPHOID SYSTEM

A) Four principal functions of lymphoid system

1) To generate immunocompetent lymphocytes that respond to diversity of antigens.

2) To concentrate antigens that enter the body into peripheral lymphoid organs (see below) where immune responses are initiated.

3) To circulate lymphocytes through peripheral lymphoid organs, thus exposing the antigen to the body's repertoire of antigen-specific lymphocytes.

4) To deliver the products of the immune response (antibodies and effector T-cells) to the blood and tissues.

B) Central (generative) lymphoid tissues. Sites where lymphocytes arise and mature, becoming competent to respond to antigens (i.e., immunocompetent). The enormous repertoire of antigen receptors is generated by unique mechanisms of DNA rearrangements (see above).

Figure 12: Diagram of human lymphoid tissues. Lymphocytes develop in both the thymus and bone marrow (dark color), which are therefore referred to as the central lymphoid tissues. The newly formed lymphocytes migrate from these central tissues to the peripheral lymphoid tissues (light color), where they can react with antigen. Reproduced with permission from Alberts et al., 2nd Ed., Figure 18-1, Page 1002.

1) Lymphocyte precursors originate in the bone marrow from hemopoietic stem cells. Those destined to become T-cells migrate to the thymus.

2) T-cells mature in the thymus.

3) B-cells mature in bone marrow.

4) Mature lymphocytes then migrate to the peripheral lymphoid tissues where they encounter antigen.

Figure 13. The development of T and B lymphocytes. In both mammals and birds, hemopoietic stem cells migrate via the blood to the thymus, where they differentiate into thymus lymphocytes. While some of these lymphocytes die in the thymus, others migrate to the peripheral lymphoid tissues to become T cells. In birds, other hemopoietic stem cells migrate to the bursa of Fabricius, where they differentiate into bursa lymphocytes; some of these lymphocytes die, while others migrate to the peripheral tissues to become B cells. In mammals, the hemopoietic stem cells destined to become B cells differentiate into lymphocytes in the hemopoietic tissue itself and then migrate to the peripheral lymphoid tissues to become B cells. Reproduced with permission from Alberts et al, 2nd Edition, Figure 18-3, page 1004.

C) Peripheral (secondary) lymphoid tissues. Sites where antibodies and effector T- cells are produced. Mature "immunocompetent" lymphocytes encounter antigen here and are triggered to differentiate into effector cells (antigen-dependent differentiation) and memory cells.

1) Lymph nodes are interposed along lymphatic vessels and filter the lymph fluid.

2) The spleen filters the blood.

3) Unencapsulated lymphoid tissue. Lymphoid aggregates are found along the gut and respiratory tracts e.g., tonsil, adenoids, Peyer's patch and appendix, where they provide immune protection to associated mucosal surfaces. Poorly defined collections of lymphocytes are found in connective tissues and almost all organs with the exception of the central nervous system. The cutaneous immune system is made up of lymphocytes and accessory cells within the epidermis and dermis. This system is responsible for immune responses to topically applied antigens.

LYMPHATICS AND LYMPH NODES.

A) The lymphatics form an extensive network of channels that carry lymph.

1) The lymph system originates in connective tissue spaces as extensively branched thin-walled vessels, called lymphatic capillaries (up to 100 ×m in diameter). They are lined by flattened endothelial cells with a poorly developed or absent basal lamina.

2) Fluid escapes from arterial capillaries, some of which is recovered by the venous capillaries; the excess fluid that passes into lymphatics is called lymph.

3) Lymph flows from lymphatic capillaries into larger vessels (one-way valves) which eventually empty into veins in neck, returning the lymph fluid to blood.

4) Lymphatics do not form a closed circulatory system.

5) Interposed along lymphatic vessels are lymph nodes, which filter the lymph, add lymphocytes to efferent lymph, and produce antibodies. Immune responses are generated in the local nodes draining the site of antigen introduction. The products of the response pass along the system and eventually enter the blood.

Figure 14. Generation of immune responses in local lymph nodes draining site of antigen introduction to the body. In this example, the immune response being monitored is humoral (i.e., antibody production).

B) Lymph Nodes

Figure 15. Simplified Diagram of lymph node.

1) Basic structure. Refer to Figure 14-10, page 270 of Junqueira et al., Basic Histology for a more detailed diagram of a lymph node.

a) Capsule and lymphatic vessels. Lymph nodes are enclosed by a capsule composed of dense connective tissue. They are penetrated on their convex surface by afferent lymphatic vessels. At deep indentation (hilus), the efferent lymphatic leaves, and blood vessels enter and leave. Trabeculae extend from capsule into node. Beneath the capsule is the subcapsular sinus. Trabecular sinuses run along trabeculae.

b) Reticulum. The reticulum of lymph nodes is a fibrous connective tissue network that is present throughout the organ (cortex, sinuses, and medulla). It provides a meshwork that supports the lymphocytes and macrophages. The reticulum consists of large branched, fibroblast-like cells (reticular cells) and reticular fibers synthesized by the reticular cells that lie upon them.

c) Cortex. The dense peripheral layer of a node is called the cortex. Within the cortex are follicles or nodules, which are dense ovoid collections of lymphocytes primarily with macrophages and diffuse cortex or paracortical fields, which includes lymphocytes, macrophages, and a specialized population of cells called interdigitating reticular dendritic cells (capable of antigen presentation). The follicles are predominantly B-cell domains and the diffuse cortex is predominantly a T-cell domain. When a follicle is unstimulated, it is termed a primary follicle; when active immune responses are underway, it becomes a secondary follicle and contains a germinal center (lighter-staining zone), in which there are many immunoblast cells.

d) Medulla. Central to the cortex and extending to hilus is the medulla. Plasma cells and immunoblasts (see below) form the medullary cords, which are in the form of dense cord-like collections of cells. Between the cords are medullary sinuses lined with macrophages. Lymph, arriving via the afferent lymphatics, percolates from the subcapsular sinus through the cortex and medullary sinuses. These channels contain reticular cells and macrophages. Particles and antigens are efficiently filtered out by macrophages. The lymph exits the node via the efferent lymphatic.

2) Role in the immune response

a) Recirculation of lymphocytes

Arteries and veins enter and leave a node at the hilum. Arterial branches reach the cortex and break up into capillary loops. Postcapillary venules return the blood to tributaries of veins which exit at the hilum. Postcapillary venules in the diffuse cortex are characterized by a "high" endothelial lining; that is, the endothelial cells are cuboidal to columnar rather than flat as in most other blood vessels. These morphologically distinctive postcapillary venules are called high endothelial venules or HEV. Blood-borne lymphocytes (both B and T) bind specifically to HEV via specific adhesion molecules, cross the endothelial lining, and migrate to their appropriate domains in the node (B or T). (One of these adhesion molecules is called L-selectin). Unless antigenic stimulation occurs (see below), the lymphocytes traverse their respective domains within the cortex, pass into the medullary sinuses and exit the node through the efferent lymphatic whence they are carried via lymphatic vessels back to the blood. The cells are then available to enter the same or another lymphoid organ. This process by which lymphocytes move from the blood into a peripheral lymphoid organ and are returned to the blood is known as lymphocyte recirculation. This process of recirculation enables the appropriate antigen-specific cells, which may be very small in number, to be brought into contact with a given antigen, in whichever lymphoid organ of the body the antigen has become sequestered upon entering the body. When the rare responsive lymphocytes do encounter appropriate antigens, the lymphocytes are activated and a response is initiated (see below). HEV are found in other peripheral lymphoid organs (but not in spleen) where they also serve as the portal of entry of blood-borne lymphocytes. In addition, the migration of blood-borne lymphocytes into sites of inflammation (e.g., inflamed synovium in joints) may occur through HEV that are induced at these sites.

Figure 16: Diagram of lymphocyte recirculation

Figure 17: Diagram of HEV (cross section)

b) Responses to antigen

Antigen induces the proliferation and differentiation of lymphocytes (bearing appropriate antigen receptors) within lymph node or other lymphoid organs. After administration of an antigen to a node, it is taken up by macrophages in the node and processed for presentation to antigen-specific lymphocytes. Antigen presentation is carried out by macrophages or interdigitating reticular dendritic cells, which receive the processed antigen from the macrophages. When a lymphocyte, arriving at the node via lymphocyte recirculation, encounters antigen which its antigen receptors can recognize, the lymphocyte is activated and undergoes a blast transformation. In this process, small cells with dense nuclei are converted to large, biosynthetically active cells (immunoblasts). T-cells become activated in the diffuse cortex. B-cells are activated in follicles.

For a humoral immune response to a typical antigen, activated T-cells migrate to a follicle to cooperate with B-cells. The activation of B-cells leads to the formation of a germinal center four to five days after stimulation, B-cell blasts are produced in response to T-dependent antigens. The B-cell blasts migrate from the secondary follicles and end up in the medullary cords where they terminally differentiate into plasma cells. The secreted antibody is added to the lymph. B-cell blasts can also leave the node and travel via the lymph and blood to distant tissue sites (e.g., lamina propria of gut) where they terminally differentiate into plasma cells. Memory T- and B-cells, and some effector T-cells (or their immediate precursors) exit the lymph node via the lymph, spreading the immune response beyond the immediate node. A lymph node can enlarge enormously during a response. This is due to the proliferation of lymphocytes and increased retention of fluids. The colloquial term for an enlarged lymph node is "swollen gland".

Figure 18. Structural changes in lymph node after stimulation with antigen.

SPLEEN - A FILTER OF THE BLOOD

A) Two main functions

1) White pulp. Immune responses against blood-borne antigens are carried out in the white pulp.

2) Red pulp. It is responsible for monitoring blood cells, removing old or damaged cells from the circulation, and it is site for the differentiation of monocytes into macrophages. It also serve as a reservoir for platelets.

B) Histology of spleen

Figure 19. Simplified Diagram of segment of spleen (after original by L. Weiss).

1) Overview. The spleen is enclosed by a thick capsule of dense connective tissue. A branching network of trabeculae divides the organ into communicating regions. The hilus is a deep indentation where the capsule is penetrated by blood vessels, lymphatics, etc. Throughout the spleen is a connective tissue reticulum (analogous to that in lymph nodes) consisting of reticular cells and reticular fibers. Lymphoid tissue, which is greyish-white in life, is called the white pulp. Surrounding the white pulp is the red pulp, making up most of area. The red pulp consists of two principal structures - thin walled splenic or venous sinuses and dense collections of blood cells, which are between the sinuses and are called splenic cords or cords of Billroth. The marginal zones are loose collections of lymphoid cells which divides white pulp from red pulp regions.

2) White pulp

a) Histologic organization. The splenic artery branches into trabecular arteries, which branch into the white pulp as central arteries. Lymphoid tissue (lymphocytes, macrophages, etc.) surrounds each central artery in a cylindrical form as the periarterial lymphatic sheath (PALS). Within a PALS, there are circumferential layers of reticular cells and fibers that support lymphocytes which are predominantly T-cells. Here and there along the sheaths are included follicles, analagous to those in lymph nodes, i.e., spherical clusters of predominantly B-cells. Each central artery gives off many branches at right angles; many of these empty into the marginal zone dividing the white pulp from the red pulp; some of the branches reach the red pulp.

b) Movement of lymphocytes. Circulating T- and B-cells exit the branches of the central arteries and enter the spleen, primarily in the marginal zone regions. HEV are not used for entry of lymphocytes into the spleen. The lymphocytes migrate to their appropriate domains (follicles for B-cells and PALS for T-cells) and slowly traverse these regions unless they encounter appropriate antigen (see below). Lymphocytes leave the spleen by crossing into red pulp, where they enter veins that leave spleen. Some lymphocytes may exit via efferent lymphatics. The lymphocytes that leave the spleen join the population of recirculating cells which are continuously passing in and out of the peripheral lymphoid organs.

c) Immune response. The marginal zones are major sites for the deposition of blood-borne antigens. The antigens are processed and presented to responsive lymphocytes by macrophages. Following a response, memory cells and effector T-cells pass out of spleen. The plasma cells that are generated accumulate in the marginal zone and splenic cords. The secreted antibodies are carried away by blood leaving the spleen.

3) Red pulp

a) Blood supply. The branches of the central arteries run into the red pulp where they are drained by splenic sinuses. Arterial vessels open directly into the splenic cords, adjacent to the splenic sinuses (open circulation). To complete the circulation, the blood must pass into the splenic sinuses. Some arterial vessels may drain directly into splenic sinuses (closed circulation), although this possibility is controversial. Keep in mind that the splenic sinuses carry blood whereas the sinuses of lymph nodes (e.g., subcapsular and medullary sinuses) carry lymph.

b) Splenic cords. These structures are organized around a reticular meshwork that is packed with all of the formed elements of the blood. The splenic cords should be considered as part of the vascular pathway between the blind-ending branches of the central arteries and the splenic sinuses. Only the most supple and mechanically-resilient blood cells survive this tortuous pathway. Old, damaged, or diseased cells are destroyed and engulfed by macrophages. Platelets are held in reserve in the cords. Monocytes that are removed from the circulation differentiate to macrophages while in the cords.

c) Splenic sinuses. See Figure 19. These are long vascular channels with an unusual endothelium and basement membrane. The endothelial cells are elongated with tapered ends. They lie parallel to the long axis of vessel and have nuclei that bulge into the lumen. The blood cells passing into these vessels from the cords must cross the sinus walls through thin slits between endothelial cells. The basement membrane of the splenic sinuses is fenestrated, formed by orthogonal arrays of heavy transverse ring components and lighter longitudinal strands connecting the rings; reticular fibers of the cords are continuous with the basement membrane. For blood cells migrating from the cords into the splenic sinuses, the fenestra pose no barrier. However, the inter-endothelial slits are a major mechanical barrier that the cells must traverse. This is a major site for the removal of old and damaged cells from the circulation. From the sinuses, the blood passes to pulp veins, and finally to trabecular veins.

Figure 20. Diagram of splenic sinus. Reproduced with permission from Rockefeller University Press. Modified from Drenckhahn and Wagner (1986). J. Cell Biol. 102:1738-1749. The blood cells entering from the splenic cords must squeeze through the narrow inter-endothelial slits of the sinus. Note the "barrel-hoop" basement membrane rings surrounding the "rod-shaped" endothelial cells.

Figure 21. Highly schematic diagram of relationship of white pulp to red pulp. Shown at the top is a cross-section through a periarterial lymphatic sheath (PALS) that is running longitudinally in-and-out of the page. Emerging from the PALS are branches of the central artery, some of which terminate in the marginal zone (MZ). One branch is shown ending blindly in the red pulp. It is shown in relationship to two splenic sinuses. To complete the circuit of blood flow through the spleen, the blood cells that emerge from the central artery branches must migrate across the red pulp and gain entry into the vascular lumens of the splenic sinuses. These sinuses feed pulp veins which in turn supply the trabecular veins which drain ultimately into the splenic vein. Not shown in this diagram is the fact that the PALS is packed with lymphoid cells, supported by a reticulum. Also not shown is dense packing of the splenic cords in the red pulp.by various blood cells, primarily erythrocytes.

VI. UNENCAPSULATED LYMPHOID AGGREGATES OF THE RESPIRATORY TRACT AND GUT

A) Tonsils, adenoids, Peyer's patches, and appendix are found along the respiratory or GI tract.

1) Their function is to supplement local lymph nodes in filtering out antigens and thereby to protect against antigens that penetrate the mucosal surfaces of the tracts.

2) The aggregates contain HEV, the portal of entry of B- and T- cells from the blood. These lymphoid organs, like the lymph nodes and spleen, are continuously "visited" by the population of recirculating lymphocytes.

3) The aggregates are divided into B- and T-cell domains.

4) Peyer's patches Ã0They are found in the lamina propria and submucosa of intestine; they provide precursors of plasma cells that produce IgA. Antigen-stimulated lymphocytes leave Peyer's patches, via lymphatics, and eventually reach the blood. From here, the lymphocytes selectively migrate to the lamina propria of the intestine, where they differentiate to IgA-secreting plasma cells. This class of antibody functions as an "antiseptic paint" once it is transported to the apical surface of the epithelium.

VII. GENESIS OF T AND B LYMPHOCYTES WITHIN CENTRAL LYMPHOID ORGANS

A) Stages of Maturation

1) Embryogenesis. Primordial lymphocyte precursors arise in blood islands of the yolk sac in the embryo. They move first to embryonic liver and then to bone marrow. During the second half of gestation and postnatal life, the stem cells produced in bone marrow give rise to lymphocyte precursors and other hemopoietic cells.

2) T-cell precursors enter the circulation and "home" to the thymus. Within the thymus, thymocytes (immature T cells); proliferate and differentiate and as they do so, they express characteristic differentiation markers on their cell surfaces including the antigen receptors. The immunocompetent T-cells that arise during this maturation process then migrate from the thymus and populate the T-cell domains in peripheral lymphoid tissues. This process continues throughout life.

3) B-cell precursors mature to immunocompetent B-cells in the bone marrow and then migrate to peripheral B-cell domains (continues throughout life). In birds, B-cell maturation to immunocompetent cells appears to occur in Bursa of Fabricius.

B) Histology of Thymus

Figure 22. Simplified diagram of a portion of the thymus. based on Figure 6.3 in Janeway and Travers. Immunology, 2nd Edition, Current Biology Press.

1) Embryology. The thymus develops from the endodermal and ectodermal layers of the third pharyngeal pouch and the third brachial cleft, respectively. Endodermal pouches migrate laterally into mesenchyme. The epitheliad thymic rudiments detach and migrate into the chest cavity to form the thymic lobes. Blood-borne lymphocyte precursors (originating first from yolk sac and postnatally from bone marrow) enter the thymus. Lymphocytes divide, infiltrate the epithelium, and push the epithelial cells apart although they remain attached. This process generates a cellular reticulum of epithelium (originally derived from ectodermand endoderm) with a system of spaces occupied by lymphocytes. The supporting cells are called epithelial reticular cells.

2) General organization. Each of the two lobes of the thymus is surrounded by a thin capsule of loose connective tissue. The capsule gives off septa defining incompletely separated lobules. The principal components of lobules are immature lymphocytes (thymocytes), epithelial reticular cells, dendritic cells, and macrophages. The outer zone of the lobules (cortex) contains numerous, densely packed thymocytes, whereas the central region (medulla) has fewer thymocytes and more prominent epithelial reticular cells. Epithelial reticular cells have an endodermal/ectodermal , not a mesenchymal, origin. They are large branching cells, with large nuclei, usually prominent nucleoli, acidophilic cytoplasm, and abundant tonofilaments. The tips of epithelial reticular cells are joined by desmosomes, forming a meshwork or cytoreticulum, largely free of reticular fibers.

3) Cortex. The great majority of cells are thymocytes (large, medium, and small). These are densely packed with no intervening connective tissue. Large, dividing cells are present in the most peripheral zone of the cortex. Smaller cells are evident toward the center of cortex. Macrophages are scattered through the cortex and resemble the epithelial cells in the light microscope. Frequently, they contain remnants of phagocytosed lymphocytes.

4) Medulla . This central region forms a branched mass of tissue, which is relatively rich in epithelial cells and poor in thymocytes. Thymic corpuscles or Hassall's bodies are unique to the medulla. They consist of a concentric arrangement of tightly wound epithelial cells. These corpuscles are a likely site of the destruction of thymocytes, a very extensive process within the thymus (see below).

5) Blood supply

a) Route of flow. Arteries enter the thymus through the capsule and penetrate the organ traveling with the septa. Arterioles running along junction of cortex and medulla give off capillaries that ascend into the cortex and small arteries that pass into the medulla. Blood returning from the cortex and medulla drains into venules running in the cortico-medullary junction.

b) Blood-thymic barrier. The blood-thymic barrier prevents circulating macromolecules within the capillaries of the cortex from entering the cortex, thus permitting cortical thymocytes to proliferate and differentiate in an environment free of foreign macromolecular antigens carried by the blood. The structures separating the blood from the cortex are: the capillary endothelial cells which are joined by tight junctions, the basal lamina of the capillaries, a thin collar of connective tissue, the basal lamina of the epithelial reticular cells and a continuous layer of epithelial cells. This barrier apparently does not exist in the medulla. Lymphocytes probably leave the thymus by passing into postcapillary venules of the medulla.

6) Differentiation of thymocytes. Lymphocyte precursors that are released from the marrow enter the peripheral cortex of the thymus; These are large, actively dividing cells, which give rise to small thymocytes which progressively move toward the medulla. The cells at these early stages are actively rearranging genes for their antigen receptors, producing an enormous repertoire of different specificities. The process is analogous to the rearrangement of immunoglobulin genes within B-cells as they mature in the bone marrow. As thymocytes move toward the medulla, the maturing cells are subjected to two selection processes, so that only cells with "useful" antigen receptors survive. Cells that are compatible with the individual's own MHC molecules survive ("positive selection"). Recall that T cells recognize antigen as it is presented on the surface of antigen-presenting cells in association with MHC molecules. Secondly, those thymocytes that react with self antigens are eliminated ("negative selection"). As a consequence of these positive and negative selection processes, most (>95%) of the newly formed thymocytes die within the thymus. The "useful" thymocytes that survive are exported to the periphery, departing through venules in the corticomedullary boundary. See Figure 23 for a summary of the major events in thymocyte development.

Figure 23. Major events in development of thymocytes within thymus.

7) Tolerance and autoimmunity. The elimination of auto-reactive lymphocytes in the thymus is one mechanism to ensure that the body's self components are not attacked by the immune system (tolerance). Another mechanism for producing tolerance involves the inactivation of mature lymphocytes in the periphery, so that they cannot respond to self antigens. This process is referred to as anergy. When the state of self tolerance is broken, autoimmunity results. Examples of autoimmune diseases are myasthenia gravis (autoantibodies against acetylcholine receptor on skeletal muscle cells) and multiple sclerosis (T-cells attack myelin membranes of axons in central nervous system). See Table II for further examples of autoimmune diseases.

TABLE II: Some Examples of Autoimmune Diseases

Based on L. Steinman, "Autoimmune Disease" Scientific American 269: 107-114, 1993.

8) Changes with Age. The thymus is relatively large at birth. There is rapid growth until the end of the second year, then it slows. The maximum size is achieved at puberty (30 - 40 g), and then there is a decrease in size through a process known as age involution. There is a replacement of cortical thymocytes with fat, an increase in the number and size of Hassall's bodies. Despite involution, the thymus remains functional throughout life.

9) Thymectomy. After neonatal thymectomy, the spleen, lymph nodes, and other lymphoid tissue are diminished with the loss of T-cell domains. The number of small lymphocytes in blood is reduced by 60%. There are severe immunological deficiencies including impairments in cellular immunity (e.g., graft rejection), delayed hypersensitivity, and antibody responses that require T-cell cooperation. Thymectomy in adults is much less serious, since peripheral lymphoid tissues are already stocked with T-cells.

Recommended General References

Alberts, B. et al., Molecular Biology of the Cell, 3rd Edition. 1994, Chapter 23 provides a good introduction to the principles of immunology.

Abbus, A.K., Lichtman, A.H., and Pober, J.S. Cellular and Molecular Immunology, 2nd Edition. W. B. Saunders Company, 1994 A comprehensive up-to-date textbook that covers both the basic science and clinical aspects of immunology.

Weiss, L. (Editor), Cell and Tissue Biology, Sixth Edition, 1988, Urban and Schwarzenberg, Inc. The sections on lymphoid tissues, in particular the spleen, are authoritative. Chapters 14-17.

Janeway, C. and Travers, P. The Immune System in Health and Disease, 3rd Edition 1997 , Current Biology Ltd. A superb book, very clear, comprehensive and beautifully illustrated.