Chapter 6: The Organization and Structure of Lymphoid Tissues

The thymus is the site for development of thymic-derived lymphocytes, or T cells. In this organ, developing T cells, called thymocytes, differentiate from lymphoid stem cells derived from the marrow into functional, mature T cells.¹ It is here that T cells acquire their repertoire of specific antigen receptors to cope with the antigenic challenges received throughout one’s life span. Once they have completed their maturation, the T cells leave the thymus and circulate in the blood and through secondary lymphoid tissues.

THYMIC ANATOMY

The thymus is located in the superior mediastinum, overlying, in order, the left brachiocephalic (or innominate) vein, the innominate artery, the left common carotid artery, and the trachea. It overlaps the upper limit of the pericardial sac below and extends into the neck beneath the upper anterior ribs. It receives its blood supply from the internal thoracic arteries. Venous blood from the thymus drains into the brachiocephalic and internal thoracic veins, which communicate above with the inferior thyroid veins.

Arising from the third and fourth branchial pouches as an epithelial organ populated by lymphoid cells and endoderm-derived thymic epithelial cells, the thymus develops at about the eighth week of gestation.² The thymus increases in size through fetal and postnatal life and remains ample into puberty,³ when it weighs approximately 40 g. Thereafter, the size progressively decreases with aging as a consequence of thymic involution.⁴ The cause of thymic involution is likely in part a result of the influence of glucocorticoid hormones.⁵ Nonetheless, there is evidence that T lymphocytes continue to develop throughout life, potentially including in some extrathymic sites.⁶

The volume of the thymus can be estimated by sonography. In one study of 149 healthy term infants within 1 week of birth, there was a significant correlation between the estimated thymic volume and the weight of the infant.^3,7 However, no correlation was apparent between the estimated thymic volume and the infant’s sex, length, or gestational age. Also, there was no apparent correlation between estimated volume and the proportions of CD4+ T cells or CD8+ T cells found in the blood. The estimated thymic volume of healthy infants increases from birth to 4 and 8 months of age and then decreases.³ Most of the individual variation at 4 and 10 months of age appears to correlate with breastfeeding status, body size, and, to a lesser extent, illness. Breastfed infants at 4 months of age have significantly larger estimated thymic volumes than do age-matched formula-fed infants with similar thymic volumes at birth.⁸

THYMIC ARCHITECTURE

A longitudinal fissure divides the thymus into two asymmetrical lobes, a larger right and a smaller left, that are derived from the right and left branchial pouches, respectively. These two developmentally separate parts of the thymus are easily separated from each other by blunt dissection.

Each lobe of the thymus is divided into multiple lobules by fibrous septa that extend inward from an outer capsule. Each lobule consists of an outer cortex and an inner medulla (Fig. 6–1). The cortex contains dense collections of thymocytes (developing immature T cells) that cytologically appear as lymphocytes of slightly variable size with scattered, rare mitoses. The lighter-staining medulla is loosely arranged and more sparsely populated by mature thymocytes and characteristic tightly packed whorls of squamous-appearing epithelial cells, called thymic or Hassall corpuscles (Fig. 6–2). These appear to be remnants of degenerating cells and are rich in high-molecular-weight cytokeratins. Hassall’s corpuscles are thought to serve a critical role in the development of regulatory T cells.⁹

The thymus contains several important cell types that serve a variety of functions including supporting the maturation of thymocytes into mature T cells. There are several types of specialized epithelial cells within the thymus.¹⁰ The three main categories of thymic epithelial cells are the medullary epithelial cells, which are organized into clusters; the cortical epithelial cells, which form an epithelial network; and the epithelial cells of the outer cortex. The epithelial cells in the cortex and medulla often have a stellate shape, display desmosomal intercellular connections, and likely function as support cells to developing thymocytes by providing important growth factors such as interleukin (IL)-7.¹¹ In addition, at primarily the corticomedullary junction, the thymus contains marrow-derived antigen-presenting cells, mostly interdigitating dendritic cells and macrophages. Scattered B cells are also present in the thymus, and these interact with maturing thymocytes and potentially regulate T-cell development.^12,13

After puberty, thymic involution begins within the cortex. This region may disappear completely with aging, while medullary remnants persist throughout life. Glucocorticoids also may induce atrophy of the cortex secondary to glucocorticoid-induced apoptosis of cortical thymocytes.⁵ This also may be seen in conditions that are associated with increases in circulating glucocorticoid hormones, for example, pregnancy or stress.^14,15

THYMIC IMMUNE FUNCTION

The thymus is the site of T-cell development. The importance of the thymus is underscored by patients with DiGeorge syndrome, or chromosome 22q11.2 deletion syndrome, who lack the genes required for thymic development.¹⁶ These patients do not develop T cells and hence have profound immune deficiency.

Prothymocytes originate in the marrow and migrate to the thymus, where they mature into T cells (Chap. 76). Maturation of T cells is accompanied by the sequential acquisition of various T-cell markers including CD2, CD3, CD4 or CD8, CD5, and the T-cell receptor (TCR) (Fig. 6–3).¹⁷ Terminal deoxynucleotidyl transferase (TdT) is found in prothymocytes and immature thymocytes but is absent in mature T cells. TdT facilitates the successful rearrangement of TCR genes in immature thymocytes.¹⁸

T-cell precursors can be found in distinct microenvironments within the thymus. Marrow-derived CD34+ pre-T cells enter the cortex via small blood vessels and are double-negative for CD4 and CD8 antigens.¹ One of the earliest identifiable T-cell membrane antigens is CD2. As the thymocytes proliferate and differentiate in the cortex, they acquire CD4 and CD8 antigens. They subsequently acquire the CD3 antigen and the TCR for antigen as they migrate toward the medulla. In the cortex, the thymocytes are induced to express the chemokine receptor, CCR7, which directs their migration to CCL19- and CCL21- producing cells in the thymic medulla where they undergo further maturation.¹⁹

Positive and negative selection of maturing T cells takes place in the thymus.²⁰ Double-positive (CD4+ and CD8+) thymocytes undergo an initial positive selection step that is mediated exclusively by thymic cortical epithelium to ensure that developing T cells can recognize peptides in the context of self major histocompatibility complex (MHC) molecules.²¹ Thymocytes that have TCRs capable of interacting with self MHC molecules expressed by thymic cortical epithelial cells undergo expansion, whereas thymocytes with defective TCR undergo apoptosis.^22,23,24 As these positively selected cells migrate toward the medulla, they experience negative selection through their interaction with thymic medullary epithelial cells in order to ensure that any T cells that react too strongly to self MHC molecules are deleted. These thymic medullary epithelial cells uniquely express the autoimmune regulatory gene (AIRE). AIRE encodes a transcriptional regulator that promotes ectopic expression of a large repertoire of transcripts encoding proteins that ordinarily are restricted to differentiated organs residing in the periphery.²⁵ This allows the thymic medullary epithelial cells to express many different self-antigens, which are presented to developing thymocytes. Those thymocytes that have TCR that react too vigorously with the MHC molecules of the medullary epithelium will undergo apoptosis.²³ Most of the developing thymocytes are destroyed. In this way, only those T cells that have the appropriate level of affinity for self-MHC molecules yet are not reactive against self antigens will reach the medulla to undergo the final maturation stages and eventually exit the thymus via efferent lymphatics as functionally competent naïve CD4+ and CD8+ single-positive T cells.

Patients with the rare disease autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) or polyglandular autoimmune (PGA) syndrome type I (PGA I) underscore the importance of negative selection of thymocytes by thymic medullary epithelial cells. APECED, or PGA I, is characterized by chronic mucocutaneous candidiasis, hypoparathyroidism, and adrenal insufficiency. In addition, most patients also have a number of other autoimmune manifestations, including thyroiditis, type 1 diabetes, ovarian failure, alopecia, and/or hepatitis.²⁶ These patients have genetic defects in AIRE,²⁷ which precludes their thymic epithelial cells from expressing the large variety of tissue differentiation self-antigens required for the negative selection of self-reactive thymocytes and the generation of central T-cell tolerance.^25,28

The spleen is a specialized abdominal organ serving multiple functions in erythrocyte clearance, innate and adaptive immunity, and the regulation of blood volume. In general the spleen contains two structurally and functionally distinct components: white and red pulp. The white pulp of the spleen consists of secondary lymphoid tissue that provides an environment in which the cells of the immune system can interact with one another to mount adaptive immune responses to bloodborne antigens. The splenic red pulp contains macrophages that are responsible for clearing the blood of unwanted foreign substances and senescent erythrocytes, even in the absence of specific immunity. Thus, it acts as a filter for the blood.

SPLENIC ANATOMY

The spleen is located within the peritoneum in the left upper quadrant of the abdomen between the fundus of the stomach and the diaphragm. It receives its blood supply from the systemic circulation via the splenic artery, which branches off the celiac trunk, and the left gastroepiploic artery.²⁹ The blood returning from the spleen drains into the portal circulation via the splenic vein. Therefore, the spleen can become congested with blood and increase in size when there is portal vein hypertension (Chap. 56).

Approximately 10 percent of individuals have one or more accessory spleens. Accessory spleens are usually 1 cm in diameter and resemble lymph nodes. However, they usually are covered with peritoneum, as is the spleen itself. Accessory spleens typically lie along the course of the splenic artery or its gastroepiploic branch, but they may be elsewhere.³⁰ The commonest location is near the hilus of the spleen, but approximately 1 in 6 accessory spleens can be found embedded in the tail of the pancreas, where they may be occasionally mistaken for a pancreatic mass lesion.³¹

The average weight of the spleen in the adult human is 135 g (range: 100 to 250 g). However, when emptied of blood it weighs only approximately 80 g. On autopsy of 539 subjects with normal spleens, there was a positive correlation between the spleen weight and both the degree of acute splenic congestion and the subject’s height and weight, but not with the subject’s sex or age.³²

The splenic volume can be estimated by computed tomography (CT) of the abdomen.³³ In one study, the splenic volume was calculated from the linear and the maximal cross-sectional area measurements of the spleen, using the following formula: splenic volume = 30 cm³ + 0.58 (the product of the width, length, and thickness of the spleen measured in centimeters).³⁴ Using this formula, the mean value of the calculated splenic volume for 47 normal subjects was 214.6 cm³, with a range of 107.2 to 314.5 cm³. The calculated splenic volume did not appear to vary significantly with the subject’s age, gender, height, weight, body mass index, or the diameter of the first lumbar vertebra, the latter being considered representative of body habitus on CT.

The splenic volume also can be estimated by sonography, which provides good correlation with volumes measured by helical abdominal CT or actual volume displaced by the excised organ. In one study of 50 patients, the linear measurement by sonography that correlated most closely with CT volume was the spleen width measured on a longitudinal section with the patient in the right lateral decubitus position (correlation coefficient [r] = 0.89, p <0.001). There was also good correlation between splenic length measured in the right lateral decubitus position and CT volume (r = 0.86, p <0.001). In another postmortem analysis of 32 normal adult spleens, the ultrasonogram measurements of maximal height, width, and breadth of the spleen were compared with the actual volume displaced by the excised organ.³⁵ The mean actual splenic volume was approximately 148 cm³ (±81 cm³ SD), whereas mean splenic volume estimated from ultrasonography was 284 cm³ (±168 cm³ SD). Despite the differences between the actual and estimated volumes, these investigators did find a roughly linear correlation between actual splenic volume and the estimated splenic volume measured by ultrasound. However, there may be operator-to-operator variation in measurement of the estimated splenic volume, making the use of sonography in longitudinal studies technically demanding.

SPLENIC ARCHITECTURE

The spleen has an open circulation, which lacks endothelial continuity from artery to vein. When isolated spleens are perfused in washout studies, erythrocytes that appear in the splenic vein appear to be flushed out from three compartments. The red cells that are flushed out first come from a compartment that presumably is formed by the splenic vessels. The erythrocytes that are flushed out next come from a second compartment, where they presumably are loosely held within the filtration beds. The erythrocytes that are flushed out last presumably were adherent to cells of the filtration beds. Although 90 percent of the blood flow passes through the splenic vessels, only approximately 10 percent of the total splenic red cells are found within this first compartment. The second compartment is perfused by 9 percent of the total inflow yet contains 70 percent of the splenic red cells. The last compartment is perfused by only 1 percent of the inflow but contains 20 percent of the splenic red cells.

These compartments reflect the anatomy of the spleen and its stroma. The stroma is composed of branched, fibroblast-like cells called reticular cells. These cells produce slender collagen fibers, the reticular fibers, which are rich in type III collagen. The reticular cells and fibers form a meshwork, or reticulum, which filters the blood. Three major types of filtration beds can be distinguished by their structure and content: the white pulp, the marginal zone, and the red pulp.

White Pulp

The white pulp contains the lymphocytes and other mononuclear cells that surround the arterioles branching off the splenic artery. After the splenic artery pierces the splenic capsule at the hilum, it divides into progressively smaller branches. Each branch is called a central artery because it runs through the central longitudinal axis of a distinctive filtration bed that surrounds each central artery (Fig. 6–4). This is composed of a cuff of lymphocytes called the periarteriolar lymphoid sheath (PALS). The PALS is composed mostly of T lymphocytes, about two-thirds of which are CD4+ T cells. The PALS around white pulp arterioles of the human spleen is not continuous.³⁶ Indeed, segments of the central arterioles might not be surrounded by T cells in areas where they run through lymphoid follicles containing pale areas of activated B lymphocytes interspersed with large, pale macrophages and dendritic cells.¹ The migration of T cells to the PALS is governed by stromal cell production of chemokines, primarily CCL19 and CCL21, which interact with the chemokine receptor CCR7 that is expressed by naïve T cells.³⁷ Stromal production of these chemokines can be stimulated by certain cytokines, such as lymphotoxin.³⁸

On gross inspection of the surface of a freshly cut spleen, these follicles appear as white dots referred to as malpighian corpuscles (Fig. 6–5). These corpuscles contain a germinal center and have the same anatomic features and functions as secondary follicles in the lymph node. Branches coming off the central artery deliver disproportionate amounts of plasma and lymphocytes to the rim of the PALS (Fig. 6–6). These branches tend to run at acute angles, leading to a selective loss of plasma from the blood, a phenomenon referred to as “skimming.” After becoming relatively depleted of plasma, the arterioles then carry plasma-reduced blood into the filtration beds of the red pulp and marginal zone. As a result, the red pulp and marginal zone beds contain relatively high concentrations of red cells.

The Marginal Zone

The marginal zone surrounds the PALS and follicles. It is composed of a reticulum, which forms a finely meshed filtration bed, serving as a vestibule for much of the blood that flows through the spleen. The marginal zone surrounds the white pulp and merges with the red pulp. It contains more lymphocytes than the red pulp. These are primarily B cells and CD4+ T cells that appear especially well equipped for rapid antibody immune responses to bloodborne antigens.^39,40,41 However, like the red pulp, the marginal zone may become congested and remove damaged and senescent red cells and parasites.

The Red Pulp

The splenic red pulp is composed of a reticular meshwork, called the splenic cords of Billroth, and splenic sinuses.⁴² This region predominantly contains erythrocytes but also has large numbers of macrophages and dendritic cells as well as fewer numbers of granulocytes, cytolytic CD8+ T cells, and natural killer (NK) cells.

As the central arteries branch and decrease in size, the PALS also branches and decreases in diameter to but a few cells surrounding the arteriole. The small arteriole finally emerges from its sheath and then terminates in either the marginal zone or the red pulp. Here these vessels are suspended and anchored by adventitial reticular cells in the periarterial beds. They often terminate abruptly as arteriolar capillaries or as vessels with a trumpet-like flare with widened slits called interendothelial slits. The blood flows through these slits into filtration beds composed of large-meshed loculi that open to one another.

The blood in the red pulp and marginal zone drains into venous sinuses that form anastomosing, blind-ending vessels. These venous sinuses actually are specialized postcapillary venules. The endothelial cells are shaped as tapered rods that are stiffened by basal, longitudinal, intermediate cytoskeletal filaments and contractile filaments of actin and myosin. These intracellular contractile filaments can shorten the vein, causing the endothelium to buckle and form interendothelial gaps, favoring transmural passage.

The endothelial cells are attached to a basement membrane. Although this appears to be fashioned of fibers, the basement membrane actually is an extracellular membranous wall with large, regular defects that expose considerable basal endothelial surface. This includes the interendothelial slits through which blood may flow from the filtration bed and into the vein. Ordinarily the interendothelial slits are narrow or even closed unless forced apart by cells in transmural transit or by endothelial contraction.

Splenic arterioles terminate at varied distances from the walls of venous vessels. Blood flowing from arterioles that terminate at the venous vessel wall may flow directly into the splenic vein. However, blood flowing from arterioles that terminate at a distance from a vein must traffic through the spleen. In so doing, the blood either may pass quickly through a nonsinusoidal venous aperture or slowly through sinusoidal interendothelial slits and the fibroblast stroma.

The fibroblast stroma contains reticular cells and myofibroblast cells, which are also called barrier cells. The latter may fuse with each other to form a syncytial membrane that connects the arterial terminals with venous interendothelial slits or apertures. Like other myofibroblasts, these cells contain actin and myosin and may contract, thereby approximating splenic arterial and venous vessels with one another. Thus, the fibroblast stroma may affect the relative proportion of blood that flows through the stroma or the sinusal interendothelial slits. Such redistribution might occur during periods of acute physiologic stress, allow for increased expulsion of red cells from the spleen, and account for some of the increase in hematocrit observed during strenuous exercise.⁴³

SPLENIC FUNCTION

Red Cell Clearance

Mixed within the stroma of the red pulp and marginal zone are monocytes and macrophages. As the blood passes through the stroma, monocytes adhere to the stroma, where the microenvironment is conducive to their maturation into macrophages and large, dendritic, lysosome-rich phagocytes. These cells assist the reticular cells in mechanical filtration. More importantly, these cells have phagocytic activity that allows them to ingest imperfect erythrocytes, store platelets, and remove infectious agents, such as plasmodia, from the circulation.⁴⁴ In addition, the monocytes and macrophages have nonphagocytic functions, such as the presentation of antigens to T cells or the elaboration of immunomodulatory cytokines.

Collectively, the anatomy of the spleen allows the marginal zone and red pulp to cull defective erythrocytes. As the blood passes slowly through the sinusoidal interendothelial slits and the fibroblast stroma, the erythrocytes must undergo alterations in shape to squeeze through the mechanical barrier generated by this filtration compartment. Normal red cells that are supple may pass through readily because the interendothelial slits can open to approximately 0.5 μm. However, erythrocytes containing large, rigid inclusions, such as plasmodium-containing erythrocytes, are delayed or sequestered.⁴⁵ Antibody-coated red cells, as present in autoimmune hemolytic anemia, are also recognized and removed by macrophages in the splenic red pulp. Polymorphisms of FcγRII (CD32) or FcγRIII (CD16) that affect immunoglobulin (Ig) G binding in vitro can alter the efficiency of clearance of antibody-coated red cells in vivo.⁴⁶

When these filtration beds sequester imperfect red cells, the blood pools inside the spleen, causing stasis and congestion. This stimulates sphincter-like contraction of the distal vein, resulting in proximal plasma transudation that produces a viscous luminal mass of high-hematocrit blood. During episodes of enhanced red cell sequestration, as occurs during malarial crises or hemolytic episodes in a small proportion of patients with sickle cell disease, the splenic volume and weight may increase 10- to 20-fold (Chap. 56).⁴⁷ Although the white pulp may enlarge, particularly in germinal centers, the marginal zones and red pulp become greatly widened with pooled erythrocytes and macrophages in this setting.

Regulation of Blood Volume

The spleen also can play a role in modulating blood volume. Release of high-hematocrit blood through splenic contraction occurs in response to activation of the baroreflex, which also may be activated during conditions of decreased blood pressure and cardiac output.^48,49 On the other hand, physiologic agents such as atrial natriuretic peptide, nitric oxide, and adrenomedullin can induce fluid extravasation from the splenic circulation into lymphatic reservoirs.⁵⁰ Excessive splenic extravasation can contribute to the inability to maintain adequate intravascular volume during septic shock. There also is evidence that the splenic afferent and renal sympathetic nerves play a role in maintaining renal microvascular tone.⁵⁰ This splenorenal reflex can influence blood pressure and, during septic shock, help promote renal sodium and water reabsorption and release of the vasoconstrictor angiotensin II. On the other hand, in portal hypertension, the splenorenal reflex can promote renal sodium and water retention and possibly play a role in the hemodynamic complications of portal hypertension through neurohormonal modulation of the mesenteric vascular bed.

Splenic Immune Function

The spleen and its responses to antigens are similar to those of lymph nodes, the major difference being that the spleen is the major site of immune responses to bloodborne antigens, while lymph nodes are involved in responses to antigens in the lymph.⁴² Antigens and lymphocytes enter the spleen through the vascular sinuses, because the spleen lacks high endothelial venules. Upon entry, the lymphocytes home to the white pulp. T cells, which express the chemokine receptor CCR7, migrate to the PALS in response to CCL19 and CCL21, and B cells, which express CXCR5, migrate to the lymphoid follicles in response to CXCL13.^37,51 Dendritic cells also express CCR7 and hence migrate to the same area as do naïve T cells. T and B cells migrate within these compartments for about 5 and 7 hours, respectively. In the absence of an immune response, these cells migrate through a reticulum arranged around the circumference of the central artery.

Upon immune activation in response to antigen, the lymphocytes may remain in the spleen to sustain a primary or secondary immune response. Activation of B cells is initiated in the marginal zones that are adjacent to CD4+ T cells in the PALS. Activated B cells then migrate into germinal centers or into the red pulp.⁵² Lymphoid nodules appear and expand by recruiting lymphocytes from the blood and the peripheral zone of the follicles, termed the mantle zone. These cells then proliferate and differentiate in the center of a lymphoid nodule, forming a germinal center.⁵³ In their path from the marginal zone to the follicles, B cells pass into the PALS, where they remain in contact with T lymphocytes for a few hours, allowing ample time for T- and B-cell interaction in response to antigens. If they are not recruited in an immune response to antigen, both T and B lymphocytes exit the spleen via deep efferent lymphatics, not the splenic veins.

These efferent lymphatics are not distinguished as separate structures within the PALS, being quite thin-walled and often packed with efferent lymphocytes. However, they are important in moving nonreactive lymphocytes out of the spleen and in producing high-hematocrit pulp blood. After leaving the spleen, the efferent lymphocytes become the afferent lymphatics of the perisplenic mesenteric lymph nodes or empty into the thoracic duct. This duct empties into the left subclavian vein, thus returning the lymphocytes to the venous circulation.

The lymphoid nodes are secondary lymphoid tissues. They form part of a network that filters antigens from the interstitial tissue fluid and lymph during its passage from the periphery to the thoracic duct. Thus, the lymph nodes are the primary sites of immune response to tissue antigens.

LYMPH NODE ANATOMY

The lymph nodes are round or kidney-shaped clusters of mononuclear cells that normally are less than 1 cm in diameter (Fig. 6–7). A collagenous capsule surrounds a typical lymph node and has an indentation called the hilus where blood vessels enter and leave.

Lymph nodes typically are present at the branches of the lymphatic vessels and form part of the extensive network of lymphatic channels that extends throughout the body. Several afferent lymphatic channels that drain lymph from regional tissues into the lymph node perforate the capsule of each lymph node. The lymph draining from the node leaves through one efferent lymphatic vessel at the hilus. The lymph from the node, in turn, empties into efferent lymphatic vessels that eventually drain into larger lymphatic channels leading eventually to the thoracic duct. The thoracic duct, in turn, drains into the left subclavian vein, thus returning lymph into the systemic circulation.

Clusters of lymph nodes are located strategically in areas that drain various superficial and deep regions of the body, such as the neck, axillae, groin, mediastinum, and abdominal cavity. The lymph nodes that receive lymph that drains from the skin, termed somatic nodes, are superficial. The lymph nodes that receive their lymph from the mucosal surface of the respiratory, digestive, or genitourinary tract, termed visceral nodes, are usually deep within body cavities.

LYMPH NODE STRUCTURE

Beneath the collagenous capsule is the subcapsular sinus, into which the afferent lymphatic channels drain (Fig. 6–8). This sinus is lined with phagocytic cells. Fibrous trabeculae radiate from the medulla adjacent to the hilus of the node to the subcapsular sinus, thus breaking the node into several follicles, called cortical follicles. These trabeculae, together with the capsule and a network of reticulin fibers, support the various cellular components of the node and serve as the scaffolding for lymphatic spaces, namely, the subcapsular and cortical sinuses. These lymphatic spaces are continuous with medullary sinuses and the solitary efferent lymphatic channel exiting the hilus.

Each cortical follicle contains dense collections of small, mature, recirculating lymphocytes. These consist of a B-cell area (cortex), a T-cell–rich area (paracortex), and a central medulla with cellular cords that contain T cells, B cells, plasma cells, and macrophages.¹ Some follicles contain lightly staining areas of 1- to 2-mm in diameter, called germinal centers. Germinal centers are the specialized sites for the generation of memory B cells and antibody affinity maturation via the process of immunoglobulin variable-region somatic hypermutation.⁵⁴ Follicles without germinal centers are called primary follicles, and those with germinal centers are called secondary follicles. Primary lymphoid follicles contain nodules that consist predominantly of small, mature, recirculating B lymphocytes.

Within 1 week after antigenic stimulation, secondary follicles develop a germinal center that contains proliferating B cells and macrophages.^55,56 The small, nonreactive B cells are apparently forced to the periphery of the follicle, where they form a dense follicular mantle. The B cells within the germinal center, on the other hand, are highly activated, typically forming blasts that have abundant cytoplasm and round, cleaved, or convoluted shapes. Follicular dendritic cells also are found within the germinal centers. These cells can trap and retain antigens for months, possibly in the form of immune complexes.⁵⁷ The germinal centers of the secondary follicle may gradually regress after the antigenic stimulus is eliminated.

Surrounding the lymphoid follicles of the superficial cortex are sheets of lymphocytes that extend to the deep cortex, the paracortex, that blend into medullary cords of cells. The paracortical zones are formed mostly of T cells. The ratio of T cells to B cells in these zones is approximately 3:1. The medulla, however, contains scattered B cells, dendritic cells, macrophages, rare NK cells, and, during an immune response, plasma cells. The superficial cortex and medulla of the lymph nodes are the thymic-independent areas, while the deep cortex is particularly enriched with T cells, forming an area that sometimes is referred to as the thymic-dependent area. The major T-cell population found within the lymph node consists of CD4+ T cells. The scattering of CD4+ T cells in the follicles, and in more prominent numbers in the interfollicular zones, reveals the proximity of CD4+ T and B cells important for T-B cooperation during proliferation and maturation of antigen-stimulated B cells.⁵⁸

The T-cell–rich paracortex is also relatively enriched with specialized NK cells and other innate lymphoid cell (ILC) populations that likely have roles in shaping innate and adaptive immune responses in lymph nodes, as well as other secondary lymphoid tissues (SLT), through the production of immunomodulatory cytokines. The NK cell population present in the lymph node paracortex is predominantly composed of the CD56^bright subset that can rapidly produce cytokines, such as interferon-γ and tumor necrosis factor-α, in response to monocyte-derived cytokines (IL-12, IL-15, and IL-18).^59,60 However, these NK cells show relatively weak cytotoxicity in comparison to CD56^dim NK cells that predominate in the blood (Chap. 77).⁶¹ There is accumulating evidence that CD56^bright NK cells represent the immediate precursors to CD56^dim NK cells and likely differentiate into the latter just prior to their egress from lymph nodes.^62,63,64 Moreover, CD34+CD45RA+ hematopoietic progenitor cells capable of giving rise to CD56^bright NK cells are also enriched in the lymph node paracortex indicating that these tissues are also likely sites of NK cell development.^65,66,67

Lymphocytes primarily enter lymphatic tissues from the blood by migrating across the tall, active endothelium of specialized postcapillary venules called high endothelial venules.⁶⁸ Cellular adhesion molecules and various chemokines, including CXCL13, CCL19, and CCL21, are responsible for the pattern of lymphocyte trafficking and determine the associations among stromal (e.g., reticular and endothelial) and parenchymal (e.g., T and B lymphocytes, dendritic cells, and macrophages) cells in the lymphoid tissues.^54,69

LYMPH NODE FUNCTION

The lymph node is the site where different types of lymphocytes, macrophages, and dendritic cells can interact with one another to generate an immune response to antigens carried within the lymph. As the lymph passes across the nodes from afferent to efferent lymphatic vessels, particulate antigens are removed by the phagocytic cells and transported into the lymphoid tissue of the lymph node.¹ Abnormal cells within the lymph, such as neoplastic cells, also can be trapped within the lymph node.

Within the lymph node, antigen is presented to T cells as processed peptides by MHC molecules of antigen-presenting cells (Chap. 76). Various T-cell subsets comprise a network of interactive cells. CD4+ and CD8+ cell-mediated contacts, as well as T-cell–derived soluble factors, induce and regulate the immune response. T-cell recognition is mediated by the TCR for antigen. Which T cells are activated is determined by the specificity of the TCRs, the structure of MHC molecules, and the nature of antigen-presenting cells, including the dendritic reticular cells, macrophages, and B cells.

Along with TCR recognition of processed antigen presented in the MHC of the antigen-presenting cell, adequate T-cell activation requires secondary signals, or costimulation, delivered through accessory molecules, such as CD28.⁷⁰ Without these secondary signals, T cells may become anergic or specifically nonresponsive to antigen stimulation.⁷¹ This specific suppression is thought to play an important regulatory role in the maintenance of self-tolerance.^72,73

T-cell recognition of specific antigen may induce release of soluble factors, such as interleukins, that can activate the T cells themselves or have paracrine effects on other neighboring leukocytes.⁷⁴ Also, activated T cells express surface molecules, such as CD40-ligand (CD154), that also can activate B cells, dendritic cells, or macrophages.^75,76

The T-dependent immune response includes the formation of early germinal centers within days after antigen exposure. There is a mixture of B cells and activated CD4+ T cells in the lymphoid follicles. T-B cooperation involves the accessory B-cell antigen CD40 and the CD154 antigen expressed on activated T cells (Chap. 76). Activated B cells differentiate and take on the cytomorphologic characteristics of centroblasts and comprise the largest numbers of cells in the early germinal center.⁵⁶ Subsequently, centroblasts give rise to smaller B cells, the centrocytes. B cells undergo affinity maturation within the germinal center. During this process, the genes encoding the surface immunoglobulin of B cells undergo high rates of mutation, called somatic hypermutation.^53,77 B cells that express immunoglobulin with little or no affinity for antigen undergo apoptosis.⁷⁸ The resulting cellular debris is tingible, or capable of being stained, and is found prominently within macrophages specifically designated tingible body macrophages. On the other hand, B cells expressing surface immunoglobulin with high affinity for antigen are selected to proliferate and differentiate to memory B cells or plasma cells.⁵⁴ As well as promoting activation of B cells, CD4+ T cells, and CD8+ T cells may give rise to circulating memory T cells.^79,80

Following the release of specific antibody, antigen–antibody complexes may form and become sequestered on the surface of follicular dendritic cells within the germinal centers. These antigen–antibody complexes produce a coating of small, bead-like, immune complex-coated bodies called iccosomes. Iccosomes may be presented to CD4+ T cells by B cells and dendritic cells. Iccosomes also appear to assist in anamnestic recall following reentry of antigen in the host.⁸¹ T- and B-cell memory functions depend upon persistence of antigen.⁸²

MUCOSA-ASSOCIATED LYMPHOID TISSUES

The mucosa-associated lymphoid tissues (MALTs) are diffusely organized aggregates of lymphocytes that protect the respiratory and gastrointestinal epithelium.⁸³ The lymphoid aggregates associated with the respiratory epithelium are sometimes referred to as the bronchial-associated lymphoid tissue. The lymphoid aggregates associated with the intestinal epithelium are sometimes referred to as the gut-associated lymphoid tissue (GALT). Lymphocytes in the GALT are located in three main regions: within the epithelial layer, scattered through the lamina propria, and clustered in organized collections in the lamina propria. The latter includes the tonsils, adenoids, appendix, and specialized structures called Peyer patches found in the ileum (Fig. 6–9). Most intraepithelial lymphocytes are CD8+ T cells, 10 percent of which express the γ/δ form of the TCR (Chap. 76). On the other hand, the intestinal lamina propria contains a mixed population of cells, including activated CD4+ T cells as well as a recently described and heterogeneous population of ILC. ILCs are now known to be key players in mucosal immunity; they release immunomodulatory cytokines, and a subset also produces IL-22 which supports epithelial homeostasis.⁸⁴ Similar to the lymphoid follicles of the spleen and lymph nodes, the mucosal follicles in the lamina propria contain mostly B cells, which sometimes are organized into germinal centers.

Solitary lymph nodules with follicular and germinal center structures occur in the mucosa and submucosa of the respiratory tract, the gastrointestinal tract (particularly within the ileum), the urinary tract, and the vagina. Microfolds overlying specialized epithelial cells in the gut transport antigenic material by pinocytosis, with potential subsequent activation of the immune response. During states of chronic inflammation, lymphoid nodules may form as a localized center of lymphocytes with marked follicular activity. The Waldeyer ring of pharyngeal lymphoid tissues and Peyer patches in the ileum contain prominent aggregated nodular lymphoid tissue. No capsule or efferent or afferent lymphatic vessels are present in these accessory lymphoid tissues.

The MALT are rich in plasma cells and eosinophils. The plasma cells are a source of secretory immunoglobulin that is transferred into the lumina of the bronchi and gastrointestinal tract. The majority of plasma cells in the mucosa of the bronchi and gut contain IgA.⁸⁵ IgA is released from the plasma cell and then combines with a secretory piece synthesized within the mucosal epithelium to become secretory IgA (Chap. 75). Secretory IgA then is secreted across the microvilli of mucosal epithelium into the lumen, where it may prevent colonization of mucosal membranes by pathogens. Lymphoid nodules along mucosa-lined tracts serve as precursors of IgA-producing cells. These nodules form a barrier against many microorganisms and antigens.⁸⁶

Peyer Patches

Peyer patches are the most important and highly organized of the GALT.⁸³ They are found in the lamina propria of the ileum (near the ileocolonic junction) and consist of up to 50 or more lymphoid nodules covered by a single layer of columnar epithelium (see Fig. 6–9). They are well developed in youth and regress with age. Antigens from the intestinal epithelium are collected by specialized epithelial cells called M cells, allowing for generation of specific immune responses against intestinal pathogens.⁸⁷ Peyer patches are the sites at which B cells differentiate in response to these antigens into the plasma cells found within the intestine.⁸⁸

Tonsils

The tonsils are the major component of the Waldeyer ring of pharyngeal lymphoid tissues. They are covered by variable epithelial surfaces that have deep, branching depressions called crypts. Fused lymphatic nodules lie adjacent to the crypts, and germinal centers are prominent. A pseudocapsule of condensed connective tissue surrounds the tonsils, and septae within the structures form lobulations. Together with the other lymphoid tissues of the Waldeyer ring, the tonsils provide the initial barrier to pathogens entering the oral pharynx.