Lab 6 Immune System 1
Objectives:
Immune system
Be able to identify (based on morphology and organ location) the various cells of the immune system:
B-lymphocytes (plasma cells)
T-lymphocytes (T-helper; T-cytotoxic; Natural Killer cells)
Macrophages
Others (Dendritic cells, Neutrophil, Mast cell
Thymus
Be able to recognize the anatomical features of the thymus:
Describe pattern of blood flow within the thymus from the corticomedullary arteries through the cortex and medulla and back to the corticomedullary veins.
Define the relationship of the thymus to lymphocytes that originate in bone marrow and to lymphocytes which populate lymph nodes and unencapsulated lymphatic tissue.
GALT and MALT
Identify non-encapsulated aggregates of lymphatic tissues (e.g., Peyer's patches, palatine and nasopharyngeal tonsils) present in the mammalian body.
Correlate the function of tonsils with their location in the body.
Lymph node
Describe the architecture of a lymph node in the following terms: cortex, medulla, capsule, trabeculae, and reticular connective tissue.
Describe the flow of lymph through a lymph node from afferent lymph vessels to the efferent lymph vessel incorporating structures such as the subcapsular sinus, cortical lymph sinusoid, and medullary lymph sinusoids.
Define the lymph (lymphatic) nodule and germinal center (identify these structures on your slides).
Describe the route of recirculation of lymphocytes from blood to lymph within the lymph node.
Spleen
Describe the cellular and fibrous connective tissue elements present in the capsule and trabeculae of the spleen.
Describe the morphologic and functional differences between red and white pulp.
Identify the following vascular components of the spleen and explain the route of blood flow through the spleen:
Splenic artery
Trabecular artery
Artery (arteriole) of the white pulp
Artery of the red pulp (a.k.a. pulp arteriole)
Sheathed capillaries
Splenic (venous) sinuses or venules
Trabecular vein
Splenic vein
Slides:
#73 Bovine hyoid bone
#79 Thymus
#76 Palatine tonsil
#80 Nasopharyngeal tonsil
#74 Lymph node
#29 lymph Node - Silver stain
#75 Mesenteric node
#77 Spleen
Bovine Hyoid Bone - decalcified - Glass slide #73
Bone marrow, in the aggregate, is the second largest organ of the mammalian body. Marrow is the reticular connective tissue that fills the central cavity of bone perfused by blood from the nutrient artery. Blood is distributed radially by thin-walled arterioles to the blood sinusoids and is returned by small venules to a central vein. Review the compact bone of the cortex and the spicules representing spongy (cancellous) bone, which project into the central cavity of the bone. Marrow in cancellous bone is the soft material found in the spaces between the bone spicules. It consists of reticular connective tissue that forms a delicate meshwork within the marrow cavity, and it is permeated by numerous thin-walled blood vessels. Within the spaces of this tissue, the developing red and white blood cells are segregated into developing cell nests. Recall that while all cells of the blood originate from pluripotent stem cells in the bone marrow, the bone marrow also serves as the major site of maturation of B lymphocytes in most mammals studied to date. Bone marrow also contains blood, adipocytes and megakaryocytes (large basophilic cells that give rise to platelets).
Erythroid nest:
The interstices of the marrow reticular connective tissue are filled with developing blood cells. Because the cells divide through the early stages of differentiation, they form clonal groups, or 'nests'. In this example the cell morphology is slightly compromised by the bone decalcification process. Nonetheless, erythroid nests are most distinguishable because they typically contain reticulocytes - enucleate red blood cells with a basophilic staining tint due to residual cytoplasmic RER.
Recall from the lecture on blood that developing blood cells lose volume as they differentiate. Thus, many large cells are present in bone marrow.
Topic: Erythroid nest
Blue arrows = erythrocytes and reticulocytes.
Granulocytic nest:
Granulocyte nests are best distinguished using the oil immersion objective on the microscope to resolve the cytoplasmic granules. Granulocytic nests typically contain larger but fewer cells. [Look for band-form granulocytes if the granulocyte nuclear profiles are evident.]
Cellular detail has been diminished by decalcification of surrounding bone, but it appears that the small nest just below the erythrocytes contains some basophilic granules.
Topic: Granulocytic nest
Blue arrows indicate granulocytic precursors. (Specific granules cannot be resolved in every cell.)
Megakaryocytes:
Megakaryocytes are the largest hematopoietic cells in the bone marrow (40-100 µm in diameter). Recall that platelets are produced by fragmentation of the megakaryocyte cytoplasm. These large cells are typically located adjacent to marrow sinusoids where it is possible for the platelets to be shed directly into the blood.
Topic: Megakaryocytes
Blue arrows = megakaryocyte
Erythrocytic precursor cell nest - note purplish enucleate reticulocytes, late-stage erythroblasts, and ejected erythroblast nuclei.
Many areas of this marrow slide have a mix of cell lineages - here a combination of erythrocytic and granulocytic precursors.
In a few of these eosinophilic myelocytes, prominent eosinophilic granules are resolved.
In this image the central clonal group is a relatively rare basophilic granulocyte cell nest.
Thymus - Glass slide #79
Intro:
The thymus is a bilobed organ that extends from the thoracic inlet to the pericardium in the prepubertal feline and canine. Recall the thymus is the site where T–lymphocytes develop and mature. The insubstantial capsule may be associated with adipose tissue, and fine connective tissue trabeculae extend from it into the parenchyma. These collagen-rich trabeculae branch off from the capsule and extend into the parenchyma dividing the organ into incomplete lobules. Each thymic lobule contains darker cortical and lighter medullary regions.
Topic: Intro
C = cortex; M = medulla. Blue line is through the corticomedullary junction.
Blood supply:
The blood supply to the thymus is derived from arteries that penetrate the parenchyma at the corticomedullary junction by way of the connective tissue septa. The arteries divide into arterioles that course along the junction and give rise to a capillary network that forms arcades in the cortex. Cortical capillaries have a continuous endothelium, relatively thick basal lamina, and a sheath of epithelial cell processes that collectively form a blood-thymus barrier. These capillaries drain into postcapillary venules located in the medulla or corticomedullary junction. Note: Immigrant hemopoietic cells enter the thymus through large venules at the corticomedullary junction and undergo a program of proliferation, lineage commitment and MHC-restricted selection of T-cell receptor (TCR, positive selection) under the influence of cortical epithelial reticular cells. Although negative selection caused by unsuccessful MHC–TCR interactions occurs in the cortex, it is the medulla that is the main site for deletion of autoreactive thymocytes. Selected single-positive CD4+ or CD8+ thymocytes undergo final maturation in the medulla and are then exported to the peripheral T-cell pool as fully functional self-tolerant T cells.
Topic: Blood supply
Blue arrows = thymic capillaries.
Medulla:
The medulla has a distinct structural pattern. It harbors a highly heterogeneous population of thymic reticular epithelial cells that have a similar structure to those in the cortex but they are often much larger and their epithelial nature is more obvious. In addition to epithelial reticular cells the thymic medulla contains epithelial cysts and neuroendocrine-like cells.
Thymic corpuscles:
Thymic corpuscles are stable groups of cells displaying the stratified organization and cellular morphology (such as intracellular keratin filaments) of other epithelial tissues. Formerly known as Hassal's corpuscles, they are thought to produce interleukins that assist with the differentiation and education of T cells. Thymic corpuscle morphology can vary considerably - in size, cellular morphology, and relative isolation from other cells. Representative detailed images are on the next page.
Topic: Thymic corpuscles
Blue arrows = thymic corpuscles.
Trabeculum:
Collagen-rich trabeculae branch off from the outer capsule and extend into the parenchyma, dividing the organ into incomplete lobules.
Topic: Trabeculum
Blue arrows = a trabeculum separating two lobules.
Thymic corpuscles - Glass slide #79
These corpuscles are in various stages of development. Note the significant variation in epithelial reticular cell morphology with respect to shape, size, aggregation or layering, and the presence or absence of recognizable keratin.
Porcine Ileum - Glass slide #161
Mesentery:
Peyer’s patches (glass Slide #161, ileum of a mature pig) are large aggregates of unencapsulated lymphatic nodules in the mucosa of the small intestine. In young ruminants, pigs and carnivores, a single large aggregate of lymphatic nodules (the ileal Peyer’s patch) is present in the distal jejunum/ileum. In sheep, cattle and rabbits this patch may represent the mammalian equivalent of the avian Cloacal Bursa and is a site where processes of negative and positive selection of B-lymphocytes occur. For this reason, this early Peyer’s patch region is often considered a primary lymphatic/immune organ. However, this area shows atrophy at a young age and thus is distinct from the numerous distinct small intestinal Peyer patches and scattered nodules of the colon that persist into adulthood (these are considered secondary lymphatic tissue).
Topic: Mesentery
Green outline = areas containing GALT in this particular field of view; Blue arrows = mesentery.
Dome epith.:
A typical area of Peyer’s patches is illustrated below. Peyer's patches are characterized by large lymphatic nodules and the infiltration of lymphoid cells into the overlying “dome” epithelium. The epithelium of the Peyer's patch “dome” region overlying the lymphatic nodules appears low columnar and contains specialized cells with deeply invaginated basal surfaces known as microfold cells or M-cells. M-cells have been shown to be the principal sites for the sampling of gut luminal antigens and are important in the transfer of antigen from the gut lumen to Peyer's Patches. Thus, Peyer’s Patches have long been considered the gatekeepers of the mucosal immune system. Peyer's Patches facilitate the generation of an immune response within the mucosa. B cell precursors and memory cells are stimulated by antigen in Peyer's Patches. Bacteria and other antigens pass to the mesenteric lymph nodes where the immune response is amplified, and activated lymphocytes pass into the blood stream via the thoracic duct. These cells then home in on the gut and carry out their final effector functions.
Topic: Dome epith.
Blue arrows = dome epithelium
The black arrow indicates typical dome epithelium. Note the dense population of lymphocytes beneath the epithelium
Absorptive epith.:
Similar to the GI system, absorptive epithelium is simple columnar (or occasionally pseudostratified) epithelium with microvilli. Lymphocytes may normally be found beneath gut absorptive epithelia but not in large numbers. Contrast the appearance of these surfaces that are specialized for nutrient absorption with the epithelial surfaces that overly Peyer's patches.
Topic: Absorptive epith.
Blue arrows = absorptive epithelium.
Peyer's patch nodule:
Each Peyer's patch contains many lymphatic nodules. In this example, the cytoplasm of central cells are not enlarged, indicating the nodule is mostly quiescent.
Topic: Peyer's patch nodule
Green asterisks indicate one immune nodule.
Many GALT nodules do not have a specialized dome for interacting with enteric antigens. This nodule is an example.
M-cells can be difficult to recognize on glass slide #161 because of the section thickness. This image of probable M-cells is digitally synthesized from four different focal planes taken with a 100X objective. Note a small lymphocyte within the recess created by each cell. The apical surface of M-cells typically has low microvilli and is slightly recessed, forming a small indentation. [In this example the detailed surface morphology is blurred due to the imaging technique.] M-cells are often associated with subepithelial macrophages when they are actively phagocytizing potential antigens.
Palatine Tonsil - Glass slide #76
Invaginations:
Mucous membranes are faced with the problem of maintaining a moist, often absorptive surface in the presence of large numbers of infectious organisms and other irritants. Thus, it is not surprising that lymphoid cells underly significant stretches of the mucosa of the digestive and respiratory systems. An important accumulation of lymphoid tissue is found in the tonsils. This is essentially lymphoid tissue that is embedded in the wall of the oropharynx (palate) and nasopharynx. The overlying epithelium is typical of the original region so one can identify the tonsil by its epithelium (best seen at higher magnification). Note that lymphatic nodules in the tonsil typically form a single layer close to the epithelial surface. The mucosa may form invaginations, as seen in this example.
Topic: Invaginations
Blue arrows = mucosal invaginations
Submucosal gland
The nodules are more superficial than these glands located in the submucosa of the pharynx.
Epithelium:
This unencapsulated aggregate of secondary lymphatic tissue lies under the epithelium of the oropharynx. A portion of the mucosal invagination reveals the stratified squamous non-keratinized epithelium, which in this case is variably infiltrated with lymphocytes. Note the juxtaposition of the dense lymphatic tissue to the epithelium in many areas.
Topic: Epithelium
Yellow arrow = submucosal gland; Green asterisk = nodular germinal center; Blue arrow = nodular corona or mantle; Red arrow = mucosal epithelium.
Germinal center:
Identify the light staining germinal center of the tonsillar secondary lymphoid nodules. These contain medium to large B lymphocytes whose cytoplasm occupy more volume than the small lymphocytes elsewhere in the nodule. The peripherally located darker staining mantle or corona consists of small lymphocytes. Note that the mantle layer is best developed near the portion of the nodule that faces the overlying epithelium. The large size of the germinal centers containing many mitotic figures is suggestive of a high level of immunologic activity in the tonsil.
Topic: Germinal center
Yellow arrow = submucosal gland; Green asterisk = nodular germinal center; Blue arrow = nodular corona or mantle; Red arrow = mucosal epithelium.
Mantle/corona:
Identify the light staining germinal center of the tonsillar secondary lymphoid nodules. These contain medium to large B lymphocytes whose cytoplasm occupy more volume than the small lymphocytes elsewhere in the nodule. The peripherally located darker staining mantle or corona consists of small lymphocytes. Note that the mantle layer is best developed near the portion of the nodule that faces the overlying epithelium. The large size of the germinal centers containing many mitotic figures is suggestive of a high level of immunologic activity in the tonsil.
Topic: Mantle/corona
Yellow arrow = submucosal gland; Green asterisk = nodular germinal center; Blue arrow = nodular corona or mantle; Red arrow = mucosal epithelium.
Mucous acini:
Recall from the previous page that these mucous acini are located in the submucosa of the pharynx.
Topic: Mucous acini
Yellow arrow = submucosal gland; Green asterisk = nodular germinal center; Blue arrow = nodular corona or mantle; Red arrow = mucosal epithelium.
Nasopharyngeal Tonsil - Glass slide #80
Epithelium:
These images will orient you to the section of nasophyringeal tonsil from the ewe. Under the microscope, compare the morphology of the nasopharyngeal tonsil with that of the palatine (oropharyngeal) tonsil (glass slide #76). Note the different epithelial types. What other differences do you find? This example is from a ewe.
Topic: Epithelium
Red arrows = lymphatic nodules, blue arrows = pharyngeal mucosa, Green asterisks = submucosal C.T.
Submucosal C.T.:
Under the microscope, compare the morphology of the nasopharyngeal tonsil with that of the palatine (oropharyngeal) tonsil. Note the different epithelial types. What other differences do you find? This example is from a ewe.
Nodules:
Under the microscope, compare the morphology of the nasopharyngeal tonsil with that of the palatine (oropharyngeal) tonsil. Note the different epithelial types. What other differences do you find? This example is from a ewe.
Epithelium:
As in the previous oropharyngeal (palatine) tonsil, note the juxtaposition of the dense lymphatic tissue to the epithelium in many areas. Identify the epithelium. Identify the light staining germinal center of the tonsillar secondary lymphoid nodules. The peripherally located darker staining mantle or corona is more prominent in this specimen. Again, the mantle layer is best developed near the portion of the nodule that faces the overlying epithelium. The fundamental similarity between the two tonsils is that their nodules are not encapsulated, as are lymph nodes.
Germinal center
As in the previous oropharyngeal (palatine) tonsil, note the juxtaposition of the dense lymphatic tissue to the epithelium in many areas. Identify the epithelium. Identify the light staining germinal center of the tonsillar secondary lymphoid nodules. The peripherally located darker staining mantle or corona is more prominent in this specimen. Again, the mantle layer is best developed near the portion of the nodule that faces the overlying epithelium. The fundamental similarity between the two tonsils is that their nodules are not encapsulated, as are lymph nodes.
Mantle/corona
As in the previous oropharyngeal (palatine) tonsil, note the juxtaposition of the dense lymphatic tissue to the epithelium in many areas. Identify the epithelium. Identify the light staining germinal center of the tonsillar secondary lymphoid nodules. The peripherally located darker staining mantle or corona is more prominent in this specimen. Again, the mantle layer is best developed near the portion of the nodule that faces the overlying epithelium. The fundamental similarity between the two tonsils is that their nodules are not encapsulated, as are lymph nodes.
Lymph Node - orientation to glass slide #74
Afferent lymphatic:
Lymph nodes, situated along the extensive drainage system of lymph vessels, filter the lymph before returning it to the bloodstream. Lymph nodes are the only lymphatic organs with both afferent (incoming) and efferent (outgoing) lymph vessels and sinuses. Study the general morphologic features of the lymph node on glass Slide 74 with a low power objective. One afferent lymphatic duct is found entering the capsule from the right.
Topic: Afferent lymphatic
Blue arrow = afferent lymphatic duct; Red arrows = efferent lymphatic duct; Yellow asterisks = paracortex; Green asterisks = cortex.
Efferent lymphatic:
The node drains via an efferent lymphatic vessel that exits the hilum. This vessel may contain valves and the lymph may exhibit pale homogeneous staining.
Topic: Efferent lymphatic
Blue arrow = afferent lymphatic duct; Red arrows = efferent lymphatic duct; Yellow asterisks = paracortex; Green asterisks = cortex.
Paracortex:
Surrounded by the capsular sinus and capsule, the outer cortex contains the lymphatic nodules. Adjacent to them are areas of paracortex (or deep cortex).
Topic: Paracortex
Blue arrow = afferent lymphatic duct; Red arrows = efferent lymphatic duct; Yellow asterisks = paracortex; Green asterisks = cortex.
Cortex:
Surrounded by the capsular sinus and capsule, the outer cortex contains the lymphatic nodules. Adjacent to them are areas of paracortex (or deep cortex).
Topic: Cortex
Blue arrow = afferent lymphatic duct; Red arrows = efferent lymphatic duct; Yellow asterisks = paracortex; Green asterisks = cortex.
Medulla:
Most of the middle of this section consists of medulla, relatively open areas in which nodal cells and lymph intermix. The medulla, consisting of medullary sinuses adjacent to stromal medullary cords, funnels lymph toward the hilus.
Topic: Medulla
Green asterisks indicate areas of medulla; Blue arrows = a trabeculum; Red asterisk = region of the hilus.
Trabeculum:
Trabeculae of reticular connective tissue divide the cortex and carry with them extensions of the subcapsular sinus toward the medulla or hilus. Trabecular sinuses appear as spaces bordering the invaginating CT stroma.
Topic: Trabeculum
Green asterisks indicate areas of medulla; Blue arrows = a trabeculum; Red asterisk = region of the hilus.
Hilus:
Locate the hilus of the node and observe any blood vessels, which both enter and leave the node in this region. The hilus contains a single efferent lymph vessel.
Topic: Hilus
Green asterisks indicate areas of medulla; Blue arrows = a trabeculum; Red asterisk = region of the hilus.
Subcapsular sinus:
The subcapsular sinus may contain various lymphatic cells in addition to open space, occupied by lymph fluid. Examine its relationship to the node's capsule and cortical lymphatic nodules.
Topic: Subcapsular sinus
Green arrows = subcapsular sinus; Yellow asterisks = cortex; Red arrows = paracortex.
Cortex:
The cortex, containing the lymphatic nodules, is divided into two areas: the outer cortex containing lymph nodules with large germinal centers and the inner paracortex, such as the areas found between deeper nodules.
Topic: Cortex
Green arrows = subcapsular sinus; Yellow asterisks = cortex; Red arrows = paracortex.
Paracortex:
The cortex, containing the lymphatic nodules, is divided into two areas: the outer cortex containing lymph nodules with large germinal centers and the inner paracortex, such as the areas found between deeper nodules.
Topic: Paracortex
Green arrows = subcapsular sinus; Yellow asterisks = cortex; Red arrows = paracortex.
Germinal center:
Within the germinal center of the cortical nodules, note the presence of mitotic lymphocytes - particularly the condensed and aligned chromatin of metaphase or anaphase cells.
Reticular cells:
In the subcapsular and medullary sinuses, note the presence of various immune cells plus reticular cells, which may be identified by an oval nucleus and delicate, elongated cytoplasm (arrows). Reticular cells can also be observed in the paracortex. The reticular cells, with their pale staining, exilic branches form a meshwork or reticulum through which lymphocytes wind their way toward the hilus. Cytoplasmic processes of the reticular cells enwrap the reticular fibers that are the primary c.t. stroma of the lymph node. Compare these areas to glass slide #29 (and next page), in which reticular fibers (collagen type III) are specifically stained.
Macrophages
The paracortex is a T-cell rich area composed of diffuse lymphatic tissue and a mix of cell types. Identify active macrophages based on vacuoles in their cytoplasm (arrows in the center of this field).
Trabeculae:
Collagenous trabeculae from the outer capsule extend inward through the cortex. The thin blue collagenous trabeculae visible in this image are presumably attached to the capsule in a different plane of section.
Medullary cords:
The medulla contains many medullary cords that provide stromal reinforcement and an attachment site for cells. The medullary cords are surrounded by pale staining medullary sinuses, which are channels for the lymph fluid. Note: Lymph flows from the afferent lymphatics→subcapsular sinus→trabecular sinus→medullary sinus→efferent lymphatics.
Topic: Medullary cords
Blue arrows are located in medullary sinuses and point to medullary cords.
Lymph node silver stain - Glass slide #29
Nodule
This lymph node was stained with a special silver stain that reveals reticular fibers. Cells also absorb some stain, which allows the regions of the organ to be discerned even though little cellular detail is evident. Note that the nodules have few reticular fibers, located around their margins. The paracortex and medulla have a significant fibrous framework that is not evident with routine staining. Medullary cords and adjacent medullary sinuses are clearly evident in the stromal patterns.
Paracortex:
This lymph node was stained with a special silver stain that reveals reticular fibers. Cells also absorb some stain, which allows the regions of the organ to be discerned even though little cellular detail is evident. Note that the nodules have few reticular fibers, located around their margins. The paracortex and medulla have a significant fibrous framework that is not evident with routine staining. Medullary cords and adjacent medullary sinuses are clearly evident in the stromal patterns.
Medulla
This lymph node was stained with a special silver stain that reveals reticular fibers. Cells also absorb some stain, which allows the regions of the organ to be discerned even though little cellular detail is evident. Note that the nodules have few reticular fibers, located around their margins. The paracortex and medulla have a significant fibrous framework that is not evident with routine staining. Medullary cords and adjacent medullary sinuses are clearly evident in the stromal patterns.
Mesenteric Node - orientation to glass slide #75
Cortex:
Glass Slide 75 is a section of a mesenteric lymph node from a pig. Identify the cortex and medulla of the pig lymph node. All of the functional components of a lymph node are present but the node appears inverted compared to the usual mammalian lymph node such as the dog (glass slide #74). Lymph enters these nodes at the hilus and leaves at the periphery of the node. This is one of the unusual aspects of the lymphatic system in the pig.
Medulla:
Glass Slide 75 is a section of a mesenteric lymph node from a pig. Identify the cortex and medulla of the pig lymph node. All of the functional components of a lymph node are present but the node appears inverted compared to the usual mammalian lymph node such as the dog (glass slide #74). Lymph enters these nodes at the hilus and leaves at the periphery of the node. This is one of the unusual aspects of the lymphatic system in the pig.
The Spleen - glass slide #77
Capsule:
The spleen filters blood and is involved in immunologic responses to blood borne antigens. Use low power to survey the general morphologic features of the spleen on glass Slide 77. Note the capsule and trabeculae consist of a mixture of collagenous connective tissue and smooth muscle. The splenic artery and vein are visible at the top of the section.
Topic: Capsule
Blue arrows = capsule; Violet arrows = trabeculae; Red arrows = red pulp; Green asterisks = nodules (white pulp).
Trabeculum:
Trabeculae distribute the largest of the organ's substantial arteries. The amount of smooth muscle in the capsule and trabeculae varies by species. The cat spleen contains less smooth muscle and less prominent vessels than many larger species.
Topic: Trabeculum
Blue arrows = capsule; Violet arrows = trabeculae; Red arrows = red pulp; Green asterisks = nodules (white pulp).
Nodule:
Note the area of white pulp, consisting of nodules in which basophilic lymphocytes aggregate, is much smaller than the area of red pulp.
Topic: Nodule
Blue arrows = capsule; Violet arrows = trabeculae; Red arrows = red pulp; Green asterisks = nodules (white pulp).
Red pulp:
Red pulp contains a rich vascular network in which erythrocytes pool, imparting the obvious color. Erythrocytes in this section may appear either orange or red, the significance of which is unclear.
Topic: Red pulp
Blue arrows = capsule; Violet arrows = trabeculae; Red arrows = red pulp; Green asterisks = nodules (white pulp).
Trabecular artery:
To understand the organization of the red pulp and white pulp it is necessary to understand the vasculature of the spleen. The trabeculae within the spleen distribute blood to the pulp. Recall from lecture that blood flows through the vasculature of the spleen in the following order: Splenic artery→trabecular artery→artery of the white pulp→artery of the red pulp (pulp arteriole)→sheathed capillary or terminal capillary→splenic sinuses or venules→trabecular vein.
Also remember that reticular fibers do not stain in this triple-stained preparation. However, they are thought to play a role in directing red cell 'open' circulation in red pulp.
Topic: Trabecular artery
Red arrows = trabecular artery
Artery white pulp:
The artery of the white pulp may branch significantly, and in larger species may also give off small nodular arterioles and capillaries within the white pulp that eventually drain into surrounding marginal sinuses. Note that in this specimen many of the 'arteries of the white' pulp are categorized as arterioles. [Splenic artery→trabecular artery→artery of the white pulp→artery of the red pulp (pulp arteriole)→sheathed capillary or terminal capillary→splenic sinuses or venules→trabecular vein.]
Topic: Artery white pulp
Blue arrows = artery in longitudinal section.
Topic: Artery white pulp
Blue arrows = artery in cross section.
PALS:
White pulp is the aggregated lymphoid tissue of the spleen and is surrounded by the marginal sinuses, which are the border between the red and white pulp. Typical lymphoid nodules with many lymphocytes are located eccentrically in areas of white pulp. The lymphoid tissue associated with arteries in the white pulp is called the periarterial lymphatic sheath (PALS). The sheath may be incomplete and not fully surround the artery. Arteries of the white pulp almost all divide into smaller vessels in the marginal zone. [Splenic artery→trabecular artery→artery of the white pulp→artery of the red pulp (pulp arteriole)→sheathed capillary or terminal capillary→splenic sinuses or venules→trabecular vein.]
Topic: PALS
Green asterisks = PALS; Red arrows = artery of white pulp (longitudinal).
Marginal sinuses:
[Splenic artery→trabecular artery→artery of the white pulp→artery of the red pulp (pulp arteriole)→sheathed capillary or terminal capillary→splenic sinuses or venules→trabecular vein.]
Topic: Marginal sinuses
Blue arrows = sinuses containing red blood cells.
Marginal sinuses:
Because the arteries of the red pulp are a continuation of the artery of the white pulp, they are smaller in caliber and may be no larger than a terminal arteriole. Most vasculature of the red pulp consists of capillaries and sinuses that are variable in size and empty directly in to the trabecular veins. The red pulp contains many capillaries and blood sinusoids that are bordered by cells aggregated around reticular connective tissue. These cell-lined spaces are the sites where blood cells are sufficiently retarded to allow the removal of old worn-out red cells and any foreign material present in the blood. Cells of the macrophage system are present throughout the red pulp to serve that function. [Splenic artery→trabecular artery→artery of the white pulp→artery of the red pulp (pulp arteriole)→sheathed capillary or terminal capillary→splenic sinuses or venules→trabecular vein.]
Topic: Artery red pulp
Sheathed capillary:
The artery of the white pulp may branch significantly, and in larger species may also give off small nodular arterioles and capillaries within the white pulp that eventually drain into surrounding marginal sinuses. Note that in this specimen many of the 'arteries of the white' pulp are categorized as arterioles. [Splenic artery→trabecular artery→artery of the white pulp→artery of the red pulp (pulp arteriole)→sheathed capillary or terminal capillary→splenic sinuses or venules→trabecular vein.]
Topic: Sheathed capillary
Blue arrows = capillary in cross section; Green circle = approximate edge of sheath surrounding the capillary.
Trabecular vein:
The trabecular veins are easily distinguished from trabecular arteries by the orientation of the smooth muscle. Veins have little smooth muscle, hence the smooth muscle visible is oriented longitudinal to the vessel - whereas arterial smooth muscle will be circumferentially-oriented around the vessel lumen. [Splenic artery→trabecular artery→artery of the white pulp→artery of the red pulp (pulp arteriole)→sheathed capillary or terminal capillary→splenic sinuses or venules→trabecular vein.]
Topic: Trabecular vein
Blue arrows = trabeculum; Red arrows = trabecular vein.