Histologic:Chapter 11

Lip

The lips are one of the most common sites of oral cancer. They are an example of a mucocutaneous junction. Three surfaces of the lips are described: an outer surface of thin skin, a free surface or red area, and an inner mucosal surface.

Slide 129 Lip, (H&E, 5-year-old child)

Scan the slide. Identify an outer surface of thin skin and on the side opposite observe the thicker mucous membrane of the inner surface. The red area is a transitional zone between the skin covering the outer surface of the lip and the true mucous membrane lining the inner surface. With low or medium power, scan the “core” of the lip containing the orbicularis oris skeletal muscle and associated connective tissue.

With medium and high power, briefly review the structure of thin skin noting the layers of the epithelium, the dermis, the hair follicles, the sebaceous glands, and the sweat glands. The sebaceous glands are not well developed in this young child. Occasional slips of skeletal muscle may be seen inserting into the dermis. These constitute portions of the muscles of facial expression (mimetic muscle) making the skin of the lip mobile.

Trace the epidermis from the thin skin to the transitional zone of the lip. At the origin of this free margin of the lip, keratinization gradually ceases; for a short distance dead and partially keratinized cells may be seen. Follow the stratum corneum from thin skin to the free margin of the lip to note its disappearance.

The connective tissue papillae in the red area are tall and highly vascularized, but this is not readily apparent in a single section. The circulating blood in the numerous small vessels of these papillae imparts the red color of the lip through its translucent epithelium.

The oral or mucosal surface of the lip is lined by a thick noncornified, stratified squamous epithelium with nucleated surface cells that slough off into the mouth. A somewhat compact zone of fine fibrous connective tissue, the lamina propria, lies beneath the mucosal epithelium.

Deep to the submucosa is skeletal muscle of the orbicularis oris. The submucosa connects the lamina propria to the thin fascia of this muscle. Are muscle fibers cut in different planes? What is the major plane of sectioning and hence the orientation of this skeletal muscle?

Note blood vessels of various sizes and small nerves throughout the muscle and connective tissues.

Slide 128 Lip, (Masson’s; adult)

Look over this slide; note the same features as above as seen with a different stain and in an older individual.

Tongue

The tongue is divided anatomically into an anterior 2/3 or body, and a posterior 1/3 or root by a posteriorly oriented V-shaped groove, the sulcus terminalis. A pit, the foramen caecum, is located at the apex of the sulcus terminalis. It represents the embryonic point of origin of the thyroid gland.

The tongue is covered by stratified squamous epithelium. On the inferior (ventral) surface, the mucosa is smooth, whereas on the superior(dorsal) surface of the body of the tongue the mucosa forms protuberances known as papillae of which there are 3 main types: filiform, fungiform and circumvallate. A fourth rudimentary type is the foliate papillae located on the posterior lateral surfaces of the tongue.

The intrinsic muscle of the tongue consists of skeletal muscle fibers arranged in longitudinal, transverse and vertical planes. A median fibrous lingual septum incompletely divides the intrinsic musculature into right and left halves.

Taste buds are barrel-shaped collections of cells found in the epithelium, on fungiform and circumvallate papillae, being most numerous on the latter.

The lingual tonsils are located behind the circumvallate papillae. They consist of numerous lymphatic nodules that form bulges on the dorsum and lateral sides of the tongue. They are aggregated along broad deep crypts.

Slide 163, Tongue, tip (H&E)

Note the stratified squamous epithelium on the surface of the section, the lamina propria, the thick mass of muscle and the glands. The epithelium of the dorsal surface is thrown up into papillae; the ventral surface is smooth and the epithelium is thinner. Note the arrangement of the muscle of the tongue and the distribution of blood vessels and nerves within it.

Identify the numerous filiform papillae on the dorsum of the tongue. They are 2 to 3 mm tall and over their tips, the epithelium is partly cornified; some desquamating cells are present at the tips. Note the core of lamina propria in the papillae. The main function of these papillae is to increase surface friction during mastication.

Toward the apex of the tip of the tongue, find several fungiform papillae. They are less numerous and larger than the filiform ones. The covering epithelium is non-keratinized. These slightly rounded and elevated papillae are said to be shaped like a mushroom with the “cap” broader than the “stalk.” The connective tissue core also shows secondary papillae, and it is quite vascular, imparting a red coloration to the papillae. Taste buds may be present in the epithelium of the apex of the fungiform papillae; however we will study them in conjunction with the circumvallate and foliate papillae below.

Locate the anterior lingual glands embedded in the muscle, lying near the ventral surface. They are mixed glands with serous, mucous and mixed alveoli. Ducts open on the ventral surface.

Slide 160, Tongue, posterior, (H&E)

Study slide 160, Tongue, posterior, (H&E) on which a vallate (circumvallate) papilla is present. Find the trench or furrow around the papilla into which the ducts of the serous glands of von Ebner empty.

Identify the purely serous von Ebner glands. Taste buds occur in the lateral epithelium of the papilla (medial wall of the trench). If a taste bud is present on this slide, study it on high power. Identify spindle-shaped cells with thin darkly staining nuclei and other more numerous cells with lighter-staining nuclei. Both types of cells have microvilli extending from their apices and projecting into the taste pore. These microvilli intertwine and makeup the so-called “taste hairs” of light microscopy. Electron microscopists have discovered four types of cells associated with the taste buds. They enumerate them as Type I, II, III and IV with the latter being a rounded basal cell that can generate the other three types. There is indecision concerning which of the cells is the actual taste cell.

Taste buds are not found on the dorsal surface of the vallate papillae.

Some slides also have a small group of mucous posterior lingual glands located laterally and deeper in the section.

Slide 162, Tongue, posterior, (H&E)

A circumvallate papilla is not present.

Note that the epithelium of the root of the tongue has no papilla (although an occasional filiform papilla may be present) and a thin lamina propria is present.

Considerable adipose tissue is present among the muscle bundles. It is more abundant in the root of the tongue but may occur in any part. Most slides show one or more sections of a small-medium sized artery in the lower right or lower left microscopic field.

Slide 161, Tongue, posterior, with lingual tonsils (H&E)

Identify again the epithelium and lamina propria of the tongue, and the masses of connective tissue and fat in which are embedded the mucous posterior lingual glands and their ducts. Some of the mucous alveoli are inactive; the cells have liberated their secretion and therefore are smaller than when filled with mucigen, and lumens are larger. Scattered skeletal muscle fibers are present but the main mass of muscle has not been included in the section.

The lingual tonsils are small aggregations of lymphatic nodules located on the dorsum and sides of the root of the tongue. Many small individual lingual tonsils, each with its own crypt, spread out under a large area of the surface of the tongue, but this arrangement may not be apparent in a small section of tissue. Lingual tonsils are not represented by a single large encapsulated mass of lymphatic nodules as is the palatine tonsil.

Identify the lymphatic nodules, some of which may show germinal centers. Note the nodules are covered with stratified squamous epithelium and that the epithelium may be infiltrated with lymphocytes (a normal occurrence). Some of the nodules and crypts may be cut tangentially and therefore their proper relationship to the epithelium is not apparent.

The nodules surround broad deep crypts extending in from the dorsal surface of the tongue. Look for sections of crypts. Ducts of the glands usually open into the bases of the crypts.

Salivary Glands

The salivary glands are divided into two main divisions. The intramural or minor salivary glands (labial, buccal, and palatine) lie within the submucosal connective tissue of the oral cavity. These intrinsic glands secrete continuously to moisten the oral cavity. Three pairs of extrinsic or major salivary glands, the parotid, the submandibular, and the sublingual, lie outside the walls of the oral cavity and must convey their secretions to the mouth by means of excretory ducts which pass through the mucosa. The major glands secrete intermittently in response to specific stimuli. The major salivary glands are classified as compound tubuloalveolar, exocrine glands. The secretory alveoli of the parotid glands are wholly serous, producing a watery secretion; those of the submandibular glands are mixed, although preponderantly serous, and those of the sublingual glands are mixed but preponderantly mucous.

Slide 156, Parotid Gland (H&E)

Scan to observe the masses of serous alveoli that make up the gland. Fat cells are scattered among the alveoli - a normal occurrence in parotid glands. Connective tissue septa divide the gland into incomplete lobules. In the septa are interlobular excretory ducts, blood vessels, nerves and lymphatics. Ducts within the lobules, that is, among the alveoli, are intralobular ducts.

With high power, identify serous alveoli (rounded oval units made up of small pyramidal cells surrounding a very small central lumen). Note the pyramidal shape of the serous cells and the rounded nucleus located toward the base of the cells. The cytoplasm had been filled with zymogen secretory granules but many of these were removed in routine section preparation leaving the fine vacuoles where they had been. A narrow, deep basophilic zone of cytoplasm is not readily apparent in this section. Try to identify myoepithelial cells (basket cells) which lie between the basement membrane and the bases of the serous cells, but they are difficult to distinguish; usually only the flattened nucleus is seen. The cells have contractile processes which “clasp” the serous cells to aid in expelling the secretion.

Note the numerous striated ducts (salivary ducts, secretory ducts) within the lobules and dispersed among the alveoli. They are supported by a small amount of connective tissue; retraction from the surrounding connective tissue is a postmortem change. These ducts are lined with a simple columnar epithelium. Focus carefully to see vertically oriented “striations” at the bases of the cells that represent mitochondria lined up between basal infoldings of the basal cell membrane. These cells contribute to the serous secretions coming from the secretory cells of the alveoli.

Slide 158, Submandibular gland (H&E)

This is from a 46-year-old man.

Look over the slide to verify that it is much like the parotid in arrangements of lobes, lobules, septa, striated ducts, intercalated ducts, and serous alveoli. Intercalated ducts are shorter than in the parotid and not as easy to identify. Fat is generally absent (unless an individual is obese), therefore the alveoli are more compactly arranged.

The submandibular gland is primarily a serous gland but scattered groups of mucous alveoli are present. Look for groups of mucous alveoli recognized by the lightly stained cells that make up an alveolus. Some mixed alveoli may be present (mucous alveoli with demilunes). Demilunes are “caps” of serous cells grouped on the periphery of a mucous alveolus.

Compare serous and mucous alveoli and note their differences.

Serous alveoli are smaller than mucous alveoli, are rounded or oval and generally uniform in size, lumens are very small. Serous cells are pyramidal in shape; cytoplasm is granular and therefore takes up stain so that the cells stain darkly; nuclei are rounded or oval and are in the basal part of the cells.

Mucous alveoli are larger, are more irregular in size, and may branch. Their lumens are larger. Mucous cells are more columnar in shape, cell outlines are visible. Cytoplasm stains very lightly since mucigen (pre-mucin) has been removed in section preparation. If the cells are filled with mucigen, nuclei are compressed and flattened at the base of the cell.

Identify lymphocytes and some plasma cells in some of the septa.

Slide 176, Submandibular Gland (PASH), same 46 year-old man

PASH (periodic acid-Schiff reagent plus hematoxylin to stain nuclei) demonstrates the presence of polysaccharides, giving a red-purple coloration to tissues that contain these substances. The degree of coloration varies with the type and amount of polysaccharide present.

Note especially the intense positive reaction (red) of the mucous cells in the mucous alveoli.

Zymogen granules in serous alveoli also give a positive reaction. Note that not all alveoli give a similar reaction, indicating that some are storing zymogen granules, others have released all or part of their secretions.

Basement membranes are demonstrated. The reticular fiber component takes the stain.

Pancreas

The pancreas is both an exocrine and an endocrine gland. The exocrine pancreas is a compound tubuloacinar gland consisting of serous secretory cells arranged in acini. As a gland of external secretion, the serozymogenic products of the acini are conveyed to the lumen of the duodenum by means of a main excretory duct, the main pancreatic duct, and by an accessory pancreatic duct which in some individuals opens into the main duct before reaching the duodenum. The main duct extends the entire length of the gland receiving branches, interlobular ducts, from the individual lobules. Striated ducts are lacking in the pancreas. The only intralobular ducts are long narrow intercalated ducts with a low simple cuboidal epithelium, which open directly into interlobular ducts . The endocrine pancreas secretes insulin and glucagon directly into the blood stream (capillaries) from irregularly dispersed aggregates of cells forming the pancreatic islets (of Langerhans). The islets are composed of anastomosing cords of cells, numerous capillaries and a fine framework of reticular fibers. They are usually directly surrounded by pancreatic acini, but islets can occur in the interlobular connective tissue. The islets range in size from only a few cells to islets containing thousands of cells. In the latter instance they are large enough to see with the unaided eye. The islets are more numerous in the tail than in any other portion of the pancreas.

Slide 154, Pancreas, (H&E)

Scan the slide to see that delicate connective tissue septa indistinctly divide the gland into lobules. In the more dense regions of connective tissue can be seen branches of the excretory duct, the interlobular ducts. Observe that serous secreting cells of the acini appear similar to the serous cells of the parotid gland. Distinguish the lighter staining islets (of Langerhans) noting the variability in size and their distribution.

With high power study a number of acini to see that they are sectioned in various planes, some show a lumen while others appear solid when the section passes through the pyramidal serous cells.

The nuclei of the pyramidal zymogenic cells are round and basally located. Observe the remarkable difference in staining between basal and apical portions of the cells. The apically located zymogenic granules are acidophilic, whereas the rough endoplasmic reticulum of the basal region is highly basophilic.

In the center of some acini, small lightly stained nuclei are present. These nuclei belong to the centroacinar cells that form the origins of the intercalated ducts. Instead of being united to the pyramidal cells of the acini directly, the ducts enter the acini and are thus enclosed or surrounded by the cells of the acini. Thus, the small cuboidal cells of the intercalated ducts lie within the acinar lumen. Intercalated ducts lead directly to interlobular excretory ducts that are lined with a simple columnar epithelium containing occasional goblet cells. Slide 153 shows goblet cells in the larger ducts. In the pancreas, the intercalated duct is somewhat larger than the intercalated duct of the salivary glands because of the absence of striated ducts.

Intercellular canaliculi occur between the serous acinar cells, but are difficult to observe with the light microscope. The canaliculi convey the secretions from the secreting cells to the lumen of the acinus.

When the pancreas is stained with H&E, special features of the cells of the islets of Langerhans cannot usually be distinguished. On slide 154, for example, even the cell boundaries of the islet cells are indistinct, and no specifically stained granules in the cells are apparent. Remember that the islets represent an endocrine gland; thus, the cells are in close association with a rich capillary bed.

The Tubular Digestive Tract

The general structure of the digestive tube consists of 4 layers that are constantly present throughout its length. From the luminal side outwards, these layers consist of a mucosa, a submucosa, a muscularis externa and a fibrosa or serosa.

Esophagus

Slide 131, Middle Esophagus (H&E)

Examine slide 131, Middle Esophagus (H&E) to identify and study the 4 layers. Examine the slide first with low or medium power.

Mucosa. This layer is approximately 500 to 800 μm thick, and it consists of an epithelium, a lamina propria and a muscularis mucosae.

• The epithelium is non-cornified stratified squamous and relatively thick.

• Lamina propria. This layer of fine-fibered loose connective tissue lies just beneath the basement membrane of the epithelium. Within this region are found thin collagenous fibers, a few elastic fibers, fibroblasts, plasma cells and lymphocytes. The infiltration of lymphocytes may be sufficiently great to form an occasional lymphatic nodule. Note the connective tissue papillae of the lamina propria indenting the epithelium.

• Muscularis mucosae. Marking the outer border of the mucosa is the muscularis mucosae consisting of smooth muscle fibers oriented mainly in the longitudinal plane. It is always a thin layer of muscle, but it is thicker in the lower esophagus than in the upper esophagus or in any other part of the alimentary tract; here it is the thickest muscularis mucosae in the body.

Submucosa. This fibro-elastic layer of coarse connective tissue fibers lies between the muscularis mucosae and the muscularis externa. Longitudinal folds of the submucosa and of the muscularis mucosae give rise to the irregular form or stellate appearance of the esophageal lumen when it is observed in cross section. The elasticity of this layer allows the folds to smooth out when food is swallowed. Note that much larger blood vessels are present in the submucosa than in the lamina propria. Within the submucosa lie a variable number of tubuloalveolar esophageal glands. The large ducts of these mucous glands penetrate the mucosa to carry secretions from the alveoli to the surface. Here the mucus acts as a lubricant in addition to that provided by the salivary glands. Observe these glands and their ducts on your slide 131. Within the lamina propria of the lower regions of the esophagus, other mucus- producing glands known as cardiac glands may be present; these are simple branched tubular glands.

Muscularis externa. The muscle of this region consists of two layers, an inner layer with the fibers oriented circularly and an outer longitudinal layer. The muscle is wholly skeletal in the uppermost part of the esophagus, it then gradually becomes a mixture of skeletal and smooth fibers (middle 1/3), and consists wholly of smooth muscle fibers in the lower 1/3. Note on slide 131 from the middle esophagus, the mixture of smooth and skeletal muscle fibers, while in the lower esophagus you would see smooth muscle fibers only.

Between the inner and outer muscle layers lies the myenteric plexus (plexus of Auerbach). Identify groups of small parasympathetic ganglion cells associated with this nerve plexus (vagus nerve).

Fibrosa. The outer layer of the esophagus is a fibrosa (tunica adventitia) of loose irregularly arranged connective tissue that serves to bind the esophagus to surrounding tissue. On slide 131 blood vessels, nerves and adipose tissue are readily identified in this layer.

Stomach

The gross subdivisions of the stomach are the cardia, the fundus, the corpus (main body) and the pyloric region consisting of a pyloric antrum, a pyloric canal and the pylorus. Only three distinct histological regions are present since the microanatomy of the fundus and corpus is similar. In all regions of the stomach, the surface epithelium consists of tall, columnar mucus-secreting cells. The muscular coat consists of three layers of irregularly arranged smooth muscle.

Cardia of the Stomach

This small, narrow, ring-shaped region of the stomach immediately surrounds the esophagus opening. Histologically, at its juncture with esophagus, the cardia exhibits a simple columnar mucus-secreting epithelium adjacent to the stratified squamous epithelium of the esophagus.

The glands in this region of the stomach are called cardiac glands, and they are similar to mucosal cardiac glands that occur infrequently in the lamina propria of the lower esophagus. Cardiac glands are simple branched tubular, mucus-secreting glands that open into bases of the gastric pits.

Slide 174, Esophagus-Stomach Junction, (H&E)

To study this slide, use only the longitudinal section seen grossly on the upper part of the slide. This will be in the lower left microscopic field. The smaller curved section is cut in varied angles, giving a false impression of relationships.

Note the following:

• The longitudinal section through the esophagus-stomach junction of the monkey shows the abrupt change from the stratified squamous epithelium of the esophagus to the simple columnar epithelium lining the stomach and gastric pits. Each cell of the stomach lining secretes mucus so the apex of the cell appears “washed out” in our routine sections. The lamina propria contains cardiac and gastric glands and is therefore greatly increased in width over that in the esophagus.

• The muscularis mucosae, submucosa and muscularis externa are continuous from esophagus to stomach.

• The cardiac glands may be very limited in extent and not even present on some slides. Other glands in the cardiac region are similar to typical gastric glands containing chief cells and parietal cells. Note that the acid-secreting parietal cells are scarce or absent in glands near the junction, but become more numerous distally as the cardia passes into the fundus of the stomach. These large, rounded acidophilic cells are in marked contrast to the smaller, basophilic serous chief cells.

• On this slide you can see some fasciculi of skeletal muscle outside the fibrosa of the esophagus. This is skeletal muscle of the diaphragm from the region where the esophagus penetrates the diaphragm to approach the junction.

The Body of the Stomach (Corpus)

Slide 133, Stomach, Corpus, (H&E)

Scan slide 133, stomach, corpus, (H&E) to observe the mucosa, submucosa and muscularis externa. Note: (1) the rugae or longitudinal folds composed of mucosa and submucosa; (2) the prominent blood vessels scattered in the coarse fibrous connective tissue of the submucosa; (3) occasional lymphatic nodules and (4) the layers of smooth muscle of the muscularis externa. Because of the arrangement, it is difficult to distinguish the three layers of smooth muscle.

Mucosa. On slide 133 study the mucosa in detail. All the simple columnar epithelial cells lining the stomach produce mucus, but the nuclei of these cells are not flattened toward the cell base as in most mucus-producing glands. The mucin stains poorly with H&E, but on slide 135 stained with PAS, the deeply stained mucin is seen to fill the apical ends of the cells. Again, on slide 133, note how the surface epithelium invaginates into the gastric pits or foveolae. The mucigen granules of the epithelial cells become progressively less abundant towards the bottom of the gastric pits as the cells themselves become shorter. From 3 to 7 gastric glands open into the bottom of each gastric pit. These closely packed glands occupy the wide lamina propria and extend all the way to the muscularis mucosae. They are simple or branched tubular glands oriented perpendicularly to the surface of the mucosa. The glands are comprised of parietal cells, chief cells, mucous neck cells and enteroendocrine cells (argentaffin cells). The latter cells can be identified on slides 135 and 174.

Parietal cells (slide 133)

These are the large, pink stained, spheroidal or pyramidal shaped cells which are most numerous in the neck and midportion of the glands. Often they occupy the periphery of the glands and bulge into the lamina propria. A constant and important feature of the parietal cells is the intracellular canaliculi. This network of canals within a cell is formed by infoldings of the plasma membrane from which numerous microvilli project. With high power observe intracellular canaliculi on slide 135 (PASH). The linings of the canaliculi are more strongly PAS-positive than the general cytoplasm so they appear as tiny “slits” in the cytoplasm. Binucleate parietal cells are easily found on slide 135. Parietal cells secrete the precursor of hydrochloric acid and the intrinsic factor (of Castle).

Chief cells (zymogenic cells), Slides 133 (above) and 135

The cuboidal or low columnar chief cells are most numerous in the deep parts of the glands. These cells secrete mainly pepsin, but it is difficult to preserve their zymogen granules, thus the apical cytoplasm appears somewhat vacuolated in our preparations. Note that the rounded nucleus lies towards the cell base, and that the basal region of the cell stains intensely with hematoxylin. This basophilia is the result of the abundant granular endoplasmic reticulum and free ribosomes found in this region.

Mucous neck cells

As the name implies, this cell type is found only in the necks of the glands except in those gastric glands near the pyloric region where the cells may be scattered. In general, mucous neck cells tend to have basal, ovoid or flattened nuclei. The cytoplasm, unless specifically stained for the mucin, may appear similar to that of the chief cells, but it is usually paler. Although typically columnar, mucous neck cells often appear irregular in shape as if deformed by adjacent cells. On slide 135 examine the necks of the gastric glands to locate mucous neck cells. Note that cells present more deeply in the glands are also PAS positive. From their nuclear morphology some of these cells appear to be “displaced” mucous neck cells. Other cells deep in the glands that are stained with PAS probably represent “overstained” chief cells.

Enteroendocrine cells (argentaffin cells)

These cells belong to a relatively diverse system of cells called APUD cells (Amine Precursor Uptake and Decarboxylation cells). A variety of substances (perhaps up to twenty or more) have been described as being produced by enteroendocrine cells. Well-preserved sections will show cells with small uniform granules in an infranuclear position. As endocrine cells they secrete through the basal lamina into capillaries and venules, not into the lumen of the gland. Identify enteroendocrine cells on slide 135 where they appear towards the base of the glands and are somewhat peripherally located among the chief cells. They have clear cytoplasm on this slide forming a “clear halo” around the nucleus. On slide 174, some of the enteroendocrine cells possess red granules (subtle to identify so search carefully) in an infranuclear position and other of these cells have clear “halos” around the nuclei as on slide 135. Identify enteroendocrine cells in the gastric glands.

Lamina propria

The lamina propria of delicate collagenous and reticular fibers and numerous cells is best studied just beneath the surface epithelium or below the base of the glands. In addition to fibroblasts, the lamina propria contains numerous other connective tissue cells including lymphocytes, plasma cells, mast cells, and eosinophilic leukocytes. Large numbers of plasma cells and lymphocytes are seen in the lamina propria of slide 133. Occasional smooth muscle fibers arising from the muscularis mucosae are seen between the glands.

Muscularis mucosae

On slides 133 and 135 scan the muscularis mucosae with medium power to observe how occasional slips of its fibers extend into the lamina propria between the glands. High power examination of the muscularis mucosae will demonstrate that its muscle fibers are cut in two planes, i.e., an inner circular layer and an outer longitudinal layer.

Submucosa

The submucosa of the stomach consists of relatively coarse connective tissue. In addition to fibroblasts, other connective tissue cells found in this layer are macrophages, mast cells, plasma cells and lymphocytes. Rather large blood and lymphatic vessels, nerve plexuses and small parasympathetic ganglia of the submucosal plexus (Meissner’s plexus) are located in the submucosa.

Muscularis externa

Locate this layer on slides 133 and 135. In the connective tissue between the circular and longitudinal layers of the muscularis externa observe the large, grayish-stained nerve cells belonging to parasympathetic ganglia of the myenteric plexus.

Serosa (visceral peritoneum)

This is a thin layer of connective tissue covered with mesothelium, forming the outermost layer of the stomach or visceral peritoneum.

Slide 136, Pyloric Stomach

The pyloric glands occupy less area than the gastric glands since the gastric pits of the pyloric region extend about ½ the thickness of the mucosa. The simple tubular pyloric glands are either coiled, or more highly branched, and have a greater diameter than gastric glands. Only one cell type, a columnar mucous cell can be identified on our slides. Recall that pyloric glands also contain gastrin-producing enteroendocrine cells but we cannot identify them on our slides.

On slide 136 observe with medium power:

• the depth of the gastric pits or foveolae.
• the lamina propria, infiltrated with lymphocytes, between the foveolae.
• the pyloric glands in the deep lamina propria.
• the relative thickness of the muscularis mucosae, the submucosa and the muscularis externa.
• under high power compare the flattened shapes of the nuclei in cells of the glands with the nuclei of cells lining the gastric pits.

Small Imtestine

The three subdivisions of the small intestine are the duodenum, the jejunum and the ileum. The general microscopic structure is similar in all three parts. In the 21 to 23 feet of small intestine, digestion of food materials continues after leaving the stomach, and its selective absorption occurs.

A. Mucosa

1. Epithelium. The absorptive process is a function of columnar epithelial cells that possess on their surface numerous microvilli forming a striated border. The microvilli increase the surface area of the lining cells at least twenty fold. Irregularly scattered among the absorptive cells are goblet cells that provide a protective secretion of mucus. The absorptive intestinal cells and goblet cells are distinctive for the small intestine, and they serve to distinguish the small intestine from the stomach that has only mucous columnar cells lining it.

2. Plicae circulares (= valves of Kerckring). Within the small intestine are circular folds of mucosa and submucosa that can be seen grossly. These visible folds, called plicae circulares or valves of Kerckring, are most prominent in the distal duodenum and proximal portion of the jejunum. They extend to the mid-ileum.

3. Villi. The mucosa also exhibits other projections or evaginations called intestinal villi that are covered mainly by columnar intestinal cells with striated borders and goblet cells. These finger-like processes, about 0.5 to 1.5 mm in height, increase the absorptive area of the small intestine in man at least five-fold. The center of each villus contains loose connective tissue of the lamina propria, a central lacteal (a lymphatic capillary), an arteriole and two venules, capillaries and slips of smooth muscle. In general the length and surface of the villi are maximal in the proximal duodenum, and they decrease gradually towards the terminal ileum.

4. Intestinal glands (crypts of Lieberkuhn). These simple tubular glands occupy the lamina propria proper. They open between the bases of the villi and extend for 0.1 to 0.7 mm through the lamina propria to the muscularis mucosae. The epithelium of the villi is continuous with the epithelium of the intestinal glands, but the striated border is progressively reduced towards the blind ends of the glands. In addition to absorptive cells and goblet cells within the crypts, cells of Paneth at the bases of the glands and scattered enteroendocrine cells (argentaffin cells) are also found.

5. Lamina propria. The intestinal glands are not as closely packed as are the gastric glands. A highly cellular lamina propria fills in the spaces between the glands or crypts and as previously mentioned, it also extends as a core of connective tissue into each villus. The main connective tissue fibers are argyrophilic reticular fibers. Elastic and delicate collagenous fibers are also present. Located within the lamina propria are large numbers of lymphocytes and plasma cells, other white cells, mast cells and scattered smooth muscle fibers. The lymphocytes may be organized into solitary nodules scattered all along the intestine. In particular, aggregates of nodules known as aggregated nodules or Peyer’s patches are found in the mucosa and submucosa of the ileum.

6. Muscularis mucosae. This thin region of smooth muscle is organized into an inner circular layer and an outer longitudinal layer, and sends strands of fibers into the lamina propria.

B. Submucosa.

1. The submucosa is a layer of loose connective tissue between the muscularis mucosae and the muscularis externa.

2. It contains the large blood vessels, lymphatics, nerve plexus and small parasympathetic ganglia (submucosal or Meissner’s plexuses).

3. Only in the duodenum are large duodenal glands (submucosal glands; Brunner’s glands) located in the submucosa.

C. Muscularis externa.

The muscularis externa of the small intestine has smooth muscle arranged into an inner circular layer and an outer longitudinal layer. In connective tissue between the two layers can be found the myenteric plexus containing parasympathetic ganglion cells and nerve fibers.

D. Serosa.

The serosa is typical, being a layer of mesothelial cells covering a thin loose connective tissue layer.

PYLORUS-DUODENAL JUNCTION

Slide 150 Pylorus-duodenum, (H&E).

1. Find under low power the typical appearance of the pyloric stomach. Identify the simple columnar mucus-secreting epithelial cells covering the surface and the coiled pyloric glands towards the muscularis mucosae.

2. Scan the slide to identify the junction of the pyloric stomach with the duodenum. This is indicated by the appearance of mucous glands in the submucosa of the duodenum.

a. Note the somewhat abrupt change where typical stomach mucosa ends and small intestinal mucosa begins. Identify:

(1) villi and the covering epithelium of absorptive cells and goblet cells. Note that the absorptive cells of the duodenum are much taller than the mucus-secreting cells of the stomach lining. (Goblet cells are not easily identified here.)

(2) the striated border of the absorptive cells.

(3) the lamina propria forming the core of the villi and supporting the short intestinal glands (crypts of Lieberkuhn) below the villi.

(4) the muscularis mucosae of the duodenum, continuous with that of the stomach.

(5) the duodenal glands (of Brunner) lying beneath the muscularis mucosae. These highly branched tubular mucus-secreting glands reside in the submucosa, and their ducts penetrate the muscularis mucosae to empty into the bases of intestinal glands. The glands may extend for a short distance into the submucosa of the pyloric stomach.

(6) the pyloric sphincter, formed by a great thickening of the circular muscle layer and inclusion of the inner oblique muscle layer of the stomach. The outer longitudinal layer does not take part in the sphincter.

(7) the muscularis externa of the duodenum, an inner circular and an outer longitudinal layer, continuous with those of the stomach.

(8) serosa.

DUODENUM

On slide 140, Duodenum (PASH):

1. Observe the plicae circulares at lowest power.

2. Examine the slide to note the numerous villi and occasional solitary lymphatic nodules.

3. With high power, study the epithelium of the villi. Observe the PAS-positive goblet cells and the columnar intestinal absorptive cells with their striated border. The glycoproteins and mucopolysaccharides of the glycocalyx (surface coat) associated with the microvilli (striated border) accounts for the PAS-positive nature of the striated border. The villi contain a connective tissue core, the lamina propria, as described previously.

4. The intestinal glands (= crypts of Lieberkuhn) open between the bases of the villi. The glands extend to the muscularis mucosae. Only a few goblet cells are present in the lower region of the glands. Cells of Paneth are not easily identified; their secretion has been liberated and few or no granules remain. (See slide 139 below.)

5. Beneath the muscularis mucosae the submucosa contains the mucus-secreting, highly branched tubular duodenal glands (= Brunner’s glands, = submucosal glands of the duodenum). The cells of these glands are PAS-positive but the mucin stains less intensely than does the mucin of the goblet cells. The ducts of these glands empty into crypts of Lieberkuhn after they penetrate the muscularis mucosae.

Study slide 139, Duodenum (H&E). (The mucosa is disrupted from post-mortem changes, villi may not be present.)

1. Observe in favorable areas the straight tubular crypts with the large pyramidal or columnar cells of Paneth at their blind ends. Paneth cells are easily identified by the large acidophilic granules in the apical regions of the cells. The round or oval nucleus is located at the base of the cell.

2. By diligent searching, find an enteroendocrine cell (argentaffin cell) with the orange-staining granules located in an infra-nuclear position, i.e., between the nucleus and the base of the cell. These cells are smaller than Paneth cells and their granules are less coarse. A variety of enzymes are secreted into capillaries by such cells.

3. This slide is excellent for studying duodenal (Brunner’s) glands. Note especially how the ducts open into the crypts.

4. In the connective tissue between the two muscle layers of the muscularis externa find ganglion cells which belong to the myenteric plexus.

JEJUNUM.

1. The jejunum lacks the submucosal glands of Brunner.

2. On slide 142, Jejunum (PASH) identify with medium power the villi on the plicae circulares, the villi located in between the plicae circulares, the lamina propria, the intestinal glands (crypts of Lieberkuhn), the submucosa and the muscularis externa.

3. The villi of the human jejunum are said to have rounded ends, but they appear somewhat variable according to the plane of sectioning.

4. Observe the goblet cells with high power. The nuclei of these cells often stain more darkly than the nuclei of adjacent absorptive cells. They are also usually narrower or more triangular in shape than in the absorptive cells.

5. In favorable regions one can locate the PAS-stained basement membrane on which the epithelial cells rest.

6. Identify the striated border and note its reduction or absence on cells at the bases of the crypts of Lieberkuhn. Small apical granules in the gland cells may form a somewhat uniform PAS stained border, but do not confuse this with a striated border. The gland cells are shorter or less columnar than the cells lining the villi.

ILEUM

This is the longest part of the small intestine. As in the jejunum, submucosal glands of Brunner are lacking.

Aggregated nodules (Peyer’s patches) form a characteristic feature of the ileum.

1. Slide 168, Ileum (H&E). This is probably from a rhesus monkey; its structure is generally similar to human, but it is small enough in diameter so that a complete cross section can be obtained for study.

a. Observe most of the features described for duodenum and jejunum. Note especially the several plicae circulares with the core of submucosa extending into them and the muscularis mucosae forming the boundary between mucosa and submucosa as usual.

b. Paneth cells are not conspicuous (will be seen better in slide 172 below).

c. Aggregated nodules (Peyer’s patches) form a prominent feature of the ileum. These are aggregates of lymphatic nodules, present in the anti- mesenteric wall of the ileum. They originate in the lamina propria, may extend through the muscularis mucosae into the submucosa. If they reach the lumen of the ileum, they are covered by a layer of cells (called M-cells), not villi.

2. Slide 172, Ileum, human (H&E). This was removed during surgery and is therefore free of postmortem changes.

a. Identify all features of the ileum. Note, only a small part of a Peyer’s patch is present. b. Study the villi that show typical normal structure. The striated border is prominent.

c. Note especially the Paneth cells, whose granules stain a brilliant pink. Also, in the lamina propria, observe eosinophils containing reddish granules.

d. Look for submucosal plexuses (Meissner’s) as well as for myenteric plexuses.

APPENDIX

1. As a blind diverticulum of the cecum, the appendix has a small and generally irregular lumen. The appendix often exhibits inflammation accompanied by infiltration of large numbers of neutrophils. A constant feature of the appendix is the vast amount of lymphatic tissue present.

2. Scan slide 145, Appendix (H&E) to find the 4 layers: mucosa, submucosa, muscularis externa, and serosa. The tissue may show postmortem changes and artifacts of preparation.

3. Note that the appendix lacks villi and plica circulares. The crypts of Lieberkuhn appear similar to the glands of the large intestine, although they may be more variable in length and less numerous per given area. The crypts contain absorptive columnar cells, goblet cells, and occasional cells of Paneth and enteroendocrine cells. Find a favorable crypt for study to identify these cell types. The striated border of the simple columnar surface cell is thin. The enteroendocrine cells are difficult to identify on our preparations but they are perhaps more numerous in the appendix (5 to 10 per gland) than in the small intestine. Mitotic figures are easily found in the lower ends of the glands.

4. The lamina propria is completely infiltrated by lymphocytes. Note how lymphatic nodules may interrupt the somewhat poorly developed muscularis mucosae.

5. Other prominent cells present here are active macrophages. The “foamy” granulated or vacuolated cytoplasm of these cells stains a darker red than that in other cells of the lamina propria. The smaller cells with bilobed nuclei and reddish cytoplasm are eosinophils.

6. Examine slide 146, Appendix (Masson’s stain). No special features are present, but note the blue-stained connective tissue, the myenteric plexus with ganglion cells, and the lymphatic nodules. The granules in the cells of Paneth are darkly stained. Observe that the circular layer of smooth muscle is thicker and better developed than the longitudinal layer. COLON

1. In the colon, the 4 layers of the alimentary tract are present, but the longitudinal layer of the muscularis externa is modified to form 3 separate thick longitudinal bands of muscle called the taeniae coli, between which is a thin layer of muscle.

2. The straight tubular intestinal glands (crypts of Lieberkuhn) are longer and more numerous than in the small intestine, and they contain more numerous mucous goblet cells. Some glands almost appear to consist of goblet cells only. Plicae circulares, villi and Paneth cells are lacking. The mucosa of the human colon therefore has a rather smooth surface. The simple columnar epithelial cells still exhibit a thin absorptive striated border.

3. The lamina propria of the colon is similar to the lamina propria of the small intestine. Lymphocytes are abundant with scattered lymphatic nodules either limited to the lamina propria or extending through the muscularis mucosae into the submucosa. Peyer’s patches are absent. The muscularis mucosae presents two layers of smooth muscle, an inner circular layer, and an outer longitudinal layer.

4. Slide 144 Colon, (PASH, rhesus monkey, complete cross-section, shows three taeniae coli). Scan the slide to see these three taeniae. Mucigen of the goblet cells is very well demonstrated. Study this slide for the features noted above. Find an area where the glands are cut longitudinally to see that they are straight and extend almost to the muscularis mucosae. Study the surface epithelium to see that in some areas more goblet cells appear in the glands than on the luminal surface between glands. Large blood vessels occur in the submucosa. Temporary folds are present. Note that the surface of the mucosa is smooth; the colon lacks villi.

5. Slide 180 Colon, (H&E, human, surgical specimen) is part of a cross-section spread open. One taenia coli is present at one end of the section. No postmortem changes are present. Again study the glands; in longitudinal sections, note their openings into the lumen of the colon and the continuity of their epithelium with the intact surface epithelium. Study other features as above. Serosa (visceral peritoneum) covers the section.

6. Slide 173 Colon, (H&E, human), is also a cross-section of the colon spread open. Much of the surface epithelium and outer parts of glands are missing, but aside from this, identify features that are listed above. Look for cell bodies of neurons in the submucosal plexuses (of Meissner). Determine if a band of the taenia coli has been sectioned and is present on your slide. RECTUM

1. Slide 170, Upper Rectum (H&E).

a. Scan the slide to observe that the longitudinal layer of the muscularis externa forms a continuous layer. Taenia coli are no longer present.

b. Observe that the structure of the upper rectum is similar to that of the colon except for the taenia coli. Identify the layers of the wall and the structures within each layer as in the colon.

ANAL CANAL

1. No slides are available for study, but you should be aware of some modifications in structure which occur in the lower rectum (upper anal canal)

a. The outer layer of smooth muscle is replaced by skeletal muscle that serves as the external anal sphincter.

b. The circular layer of smooth muscle becomes thickened to form the involuntary internal sphincter.

c. Longitudinal (vertical) ridges appear which form the rectal columns (columns of Morgagni or anal columns). They usually overlie small veins that contribute to the formation of internal hemorrhoids. Anal valves are transverse folds of the mucosa which connect the rectal columns at their bases.

d. The intestinal glands (crypts) become shorter and disappear at the level of the anal valves. Here an abrupt transition leads into the lower anal canal. The epithelium becomes noncornified, stratified squamous. Near the anal orifice, the epithelium becomes cornified, highly pigmented and it contains hairs, sebaceous glands and sweat glands.