Figure 1 (Case et al, 2011)
Even though they bolt their food, little chewing does happen in order for digestion to begin. Since Canis lupus familiaris (dogs) are carnivores, their teeth have adapted to enable them to chew and tear meat and they have 42 different structured teeth, shown in figure 2, which allow this function to happen (Jacobs, 2005). The incisors and canines, which are located at the front of the mouth, allow the food to be broken down into smaller molecules before entering the stomach, due to the sharp points (Aspinall, 2014: page 95). The tongue is also able to aid with this since it is composed of bundles of striated muscle, making it strong, and the cranial end is very mobile, so it can shape the food into a bolus making it easier to be chewed and travel down the oesophagus (Aspinall, 2014: page 95). Even though food is not in the dogs mouth a long time, so saliva cannot have much effect, it does help with this process as it lubricates the food aiding with swallowing (Michell et al, 1989: page 90). Paired salivary glands secrete saliva but since this contains no enzymes, rather than breaking down food in the mouth, the main role of saliva in dogs is to help kill bacteria minimising the amount that gets into the stomach (Cowell, 2017).
Figure 2 (Aspinall, 2014)
Once the food has passed through the pharynx and oesophagus, it then enters the stomach via the cardiac sphincter. In canines, digestion begins in the stomach where food is broken down into smaller molecules and passed through the GI tract. This is a large muscular bag which stores food temporarily and is the site at which food is converted into a semi – liquid; chyme (Grabowski et al, 2000: page 833). It is also lined with gastric folds and is where acid and enzymes are produced to aid with further digestion.
Figure 3 (Washington State University College of Veterinary Medicine, 2019)
The stomach consists of four regions which are shown on figure 3. The upper cardiac region is at the top of the stomach, surrounding the cardiac sphincter, the fundus is an air-filled section, the corpus is the body of the stomach where the food is stored, and the pyloric region is at the bottom of the stomach connecting to the duodenum. The pyloric region consists of two parts: the pyloric antrum and the pyloric canal (Grabowski et al, 2000: page 833). Pyloric antrum, which is connected to the body of the stomach, contains G cells which secretes the hormone gastrin. The stomach is stimulated by this hormone to release gastric acid. This hormone has two main function: breaking down proteins into amino acids and disinfecting, by killing most of the bacteria which is entered with food (Society for Endocrinology, 2011). After this process, the partially digested food enters the duodenum via the pyloric canal. Dogs eat much quicker than the food can be digested, since liquids can pass through the stomach in half an hour and solids or fatty foods take over four hours (Aspinall, 2014: page 96). As the stomach is a larger storage area at the upper part of the GI tract, it means the intestines can be supplied with partially digested food at an optimised rate whilst the rest is still being stored (Hove et al, 2010: page 578).
The mucosa, a layer of the GI tract wall, lies in large folds when the stomach is empty (Grabowski et al, 2000: page 820). This allows a lot of food to be stored at one time before being slowly released into the intestines, as they expand to accommodate large meals. Due to these folds, which can be seen in figure 3, there is space for more food than can be digested to be stored at one time, giving time for the acid to break down proteins and kill the bacteria before being digested further. This is important in dogs as they eat their food quicker than it leaves the stomach, aiding with the function to serve as a holding reservoir (Grabowski et al, 2000: page 833) (Aspinall, 2014: page 96).
Gastric pits which open into the lumen of gastric glands and secrete gastric juice (a mixture of hydrochloric acid, pepsin, mucus and water) are small apertures which the mucosa is indented by. There are four types of cells in gastric glands: peptic cells, oxyntic cells, goblet cells and argentaffine cells (Grabowski et al, 2000: page 833). Gastrin is the hormone that is secreted by the stomach wall in response to food passing through the cardiac sphincter. Then mucus, hydrochloric acid (HCI) and pepsinogen are secreted by gastric pits which are stimulated by gastrin (Aspinall, 2014: page 97).
Pepsinogen is secreted by peptic cells but into order to have enzyme energy, on contact with HCI this is converted into pepsin. The role of this enzyme is to break down proteins in the stomach. HCI is produced by oxyntic cells giving gastric juice a pH 2.0. As the stomach is very acidic, growth of bacteria is prevented from being further digested due to HCI killing the microbes. Pepsinogen requires acidic conditions to become pepsin, as well as the function of pepsin. Due to the amount of acid, the stomach can digest food even when they are in chunks which allows food to get broken down without the process getting slowed down, as they do not chew much. If this enzyme was not secreted as an inactive form, then the stomach would get digested as the tissues would get attacked before being released. Once secreted, the mucus lining of the stomach wall prevents the active enzyme from attacking any tissues. This protective barrier is produced by goblet cells. Gastric juice is formed from the secretion of this cells (Grabowski et al, 2000: page 833 – 837). This structure allows pepsin to be most efficient breaking down the large complex molecules without digesting the stomach wall (Michell et al, 1989: page 94).
Mixing waves, which is gastric rippling, peristaltic movements, pass over the stomach several minutes after food enters, approximately five waves per minute in dogs (Fulton et al, 2009: page 17). Since the fundus is a storage site, few waves occur here. Therefore, food may remain in the fundus for up to an hour or more, without becoming mixed with gastric juice (Grabowski et al, 2000: page 836). The bolus is softened during this process and mixed with secretions of gastric glands resulting in chyme (Grabowski et al, 2000: page 836). More vigorous mixing waves begin at the corpus of the stomach and intensify as they reach the lower region (Grabowski et al, 2000: page 836). Each of these waves forces several millilitres of gastric content into the duodenum, through the pyloric sphincter, as food reaches the pylorus. However, solids will not be sent through so this will only happen if it has been converted into liquids (Grabowski et al, 2000: page 836). The thick outer muscle of the stomach allows for this vigorous churning to occur without any damage. Due to the structure of the stomach it means there is space for the food to be forced back into the corpus, where further mixing will continue, and the next wave will push it forward again. These forward and backward movements are responsible for most of the mixing in the stomach, to mix the gastric content with enzymes (Grabowski et al, 2000: page 836).
The secretion of gastric juice is controlled by neural and hormonal mechanisms. The cephalic, gastric and intestinal phase are all stages where gastric digestion happens. The cephalic phase all happens in the head. This is sight, smell, taste and expectation of food as they all initiate a reflex. The medulla, which receives information from the cerebal cortex and feeding centre in the hypothalamus, transmits impulses along the vagus nerve (Grabowski et al, 2000: page 837). The gastric glands are stimulated by these impulses to secrete pepsinogen, hydrochloric acid and mucus into stomach chyme and gastrin into the blood (Grabowski et al, 2000: page 837). These are not being produced at all times, but only when the animal sees, smells, tastes and expects food. If it was to happen constantly then that would be a waste of water which could lead to dehydration in the dog. Neural and hormonal mechanisms are initiated by the sensory receptors in the stomach once food reaches it, to ensure motility and gastric secretion continues. This is the gastric phase of gastric digestion (Grabowski et al, 2000: page 837).
Distention to the stomach is caused by food as it stimulates the stretch receptors in the wall. PH of the stomach chyme is monitored by chemoreceptors. Stretch receptors and chemoreceptors are activated when stretching occurs in the stomach walls or when pH rises due to food entering the stomach. The more food that is added, the more dilute the acid becomes meaning pepsin is not very active, so cannot break down proteins since the stomach will not be at its optimum pH (Grabowski et al, 2000: page 837).
Nerve impulses travel from the receptors to the submucosa resulting in waves of peristalsis and stimulation to the flow of gastric juice, due to impulses. These waves mixes gastric juice with food (Grabowski et al, 2000: page 837). This negative feedback cycle turns down gastric juice secretion as the pH of the stomach chyme returns back to being acidic and the walls are not as stretched due to the chyme being passed into the duodenum (Grabowski et al, 2000: page 837).
G cells are stimulated to secrete gastrin, a hormone, by mechanical and chemical stimulation of the stomach lining by food. This hormone reaches the target cells (the gastric glands) by entering the bloodstream and circulating around the body until it gets there. When the pH of gastric juice drops below 2.0 then gastric secretion is stopped. Food dilutes the acid, therefore pH rises, so gastrin is now produced. This negative feedback mechanism helps provide a low pH for pepsin functioning and killing microbes (Grabowski et al, 2000: page 839).
Gastric inhibitory peptide (GIP), secretion and cholecystokinin (CCK) are the digestive hormones where are released when chyme enters the small intestine. They all have effects on the stomach, but secretin and CCK effects the pancreas, liver and gladder bladder more (Grabowski et al, 2000: page 844). When the stomach is stretched, GIP is released which promotes gastric juice secretion and growth of gastric mucosa as well as increases gastric motility. Pancreatic juice and bile are secreted by secretin which also increases gastric motility. CCK is also the hormone that secretes pancreatic juice and inhibits gastric emptying. Further growth and maintenance of the pancreas is promotes by CCK and secretin (Grabowski et al, 2000: page 847). When there are partially digested proteins present, gastrin is secreted. This hormone promotes motility of the stomach as well as relaxation of the pyloric sphincter. GIP, CCK, and enterogastric (a neural reflex) inhibit stomach emptying. Fats are the slowest to empty due to fatty acids releasing CCK and GIP which slow stomach emptying (Grabowski et al, 2000: page 852).
The movement of chyme from the stomach to the small intestine is dependent on the pancreas, liver and gallbladder (Grabowski et al, 2000: page 840). As shown in figure 1, these accessory glands are all located between the oesophagus and intestines.
With the dogs liver having six lobes it is the largest internal organ in the body, representing 3% of the animals body weight (Hove et al, 2010: page 592). The liver produces bile, which does not contain any enzymes, whereas the gallbladder stores this fluid (Grabowski et al, 2000: page 843). Since the gallbladder is located within the liver lobes, the bile is collected and passed into the duodenum by the gallbladder (Aspinall, 2014: page 97). As chyme is travelling to the duodenum, bile is forced along the bile duct into the duodenal lumen as the gall bladder contracts. The structure of the gallbladder aids with the function as it means a lot can be stored ready for when it is needed for activating enzymes which are required for further digestion. So fats can be digested, they are emulsified by bile which also increases the surface area for enzymic action (Aspinall, 2014: page 97). The structure of the liver helps with the functions as the thick outside layer is protection from any damage, allowing bile to still be produced and fats to be broken down. Also, there is a large storage space, because of the size, allowing a lot of bile to be produced (Agar, 2001: page 106). Due to the structure of the gallbladder, even when no bile is needed as food is not being digested, the extra bile that is still being produced by the liver can be stored in the gallbladder and is ready to be used when needed (Spielman, 2015).
The pancreas is situated below the stomach, opening into the duodenal lumen, and is the site where additional digestive enzymes are produced (Aspinall, 2014: page 96). Even though it is common for dogs to have two pancreatic ducts, a ventral or accessory and a dorsal pancreatic duct, it is known for few to have one, an accessory duct, or three functional openings into the intestines (Washabau, 2012) (Jubb et al, 2016). Since the pancreas and small intestine are connected, this allows the chyme to mix with the enzymes when they are released (Jacobs, 2005).
The pancreas opens into the duodenal lumen via the pancreatic duct through which pancreatic juice is released (Aspinall, 2014: page 96). In dogs, the duodenal papilla is where the bile ducts enter the duodenum (Kararli, 1995: page 361). The pancreas is spilt up into 3 regions: head, body, and tail and has the functions of endocrine and exocrine secretions. Endocrine produces hormones that regulate sugar whereas exocrine produces enzymes that help digest food (Agar, 2001: page 102). Pancreatic juice is released from the pancreas when the exocrine has been stimulated by hormones (Aspinall, 2014: page 96). The enzymes that are produced, for example amylase that break down carbohydrates into glucose, have an outer protective layer whilst they are in the pancreas. This is because as they travel through the pancreatic ducts to reach the GI tract they would be active in the pancreas and break down tissue therefore this structure aids them to move without causing any harm.
Cells within acini, a cluster of glandular epithelial cells which make up the pancreas, secrete pancreatic juice, a mixture of fluid and enzymes (Grabowski et al, 2000: page 840). Due to the sodium bicarbonate, which makes up the pancreatic juice, it is given an alkaline pH. This buffers acidic gastric juice in chyme, stops the action of pepsin from the stomach and creates pH for the digestive enzymes in the small intestines (Grabowski et al, 2000: page 840).
Pancreatic amylase converts starch to maltose and pancreatic lipase break down fat into fatty acids and glycerol (Grabowski et al, 2000: page 841). Since food remains in the duodenum longer than in the mouth, this gives more opportunity for starch hydrolyses. Inactive enzyme precursors in the pancreas are trypsinogen and chymotrypsinogen. Once these enzymes have travelled through the pancreatic duct, another enzyme enterokinase, which is produced in the glands of the small intestine walls converts trypsinogen into trypsin. This active enzyme then activates the conversion of chymotrypsinogen into chymisetrypsin. Proteins are then broken down into polyepeptides by these enzymes that are now active (Grabowski et al, 2000: page 841).
The total length of the intestines is approximately four times the length of the dogs body (Michell et al, 1989: page 95). Canines have a relatively short small intestine as they do not eat foods that take a long time to digest in the gut (Hume et al, 1995: page 20). A beagle has a small intestine of 225 – 290cm long with the firs 25 cm being the duodenum and the ileum being the last 15cm (Kararli, 1995: page 354). This is the site at which chemical digestion is completed by mucus and enzymes which are produced from the secretory cells (Grabowski et al, 2000: page 840) (Case et al, 2011: page 50).
Once chyme has passed through the pyloric sphincter and into the small intestine, further digestion can continue with aid from the accessory glands. The duodenum, jejunum and ileum are the three sections of the small intestine and is where carbohydrates and proteins are digested by the enzymes which are produced in the pancreas (Hove et al, 2010: page 596). The exit of chyme from the stomach is slowed down, which prevents overloading the duodenum, by neural and hormonal reflexes that are initiated in the small intestine. The reflexes ensure the stomach does not empty more chyme than the small intestine can process (Grabowski et al, 2000: page 846).
The structure of the small intestines means there is a large surface area, because of lots of fold, villi and microvilli. This helps with digesting and absorbing nutrients because it means there is more areas for this to take place and more digested nutrients can diffuse into absorptive cells in less time (Grabowski et al, 2000: page 850). Also, chyme is forced to spiral, by the folds, as it passes through the small intestine which enhances absorption furthermore (Grabowski et al, 2000: page 849). Each villus contains blood and lymph capillaries.
Due to the liver and pancreas connecting to the small intestine, the enzymes required for digestion can be passed through the pancreatic and bile ducts. Also, since the gall bladder connects to the duodenum via bile ducts, bile only has a short distance to go so is pour into the small intestine (Aspinall, 2014: page 96). As bile neutralises the acids and breaks down fats which helps the small intestine with its function as alkaline conditions are created as well as a larger surface area.
The large intestine extends from the ileum to the anus and contains three sections: caecum, colon and rectum. The colon is divided into further portions: ascending (which is attached to the ileum), transverse and descending (Aspinall, 2014: page 97). This is where the final stage of digestion takes place, by bacteria which chyme for elimination by fermenting any carbohydrates that are remaining and releasing hydrogen, carbon dioxide, and methane gases (Grabowski et al, 2000: page 859). The caecum is a sac which is attached to the colon, close to the ileum (Michell et al, 1989: page 98). Water, electrolytes and some vitamins get absorbed by the colon. Any carbohydrates and proteins that have not been absorbed in the small intestines get broken down further in the large intestines (Hove et al, 2010: page 597). Glands within the mucosa of the colon secrete mucus which acts as a lubricant aiding with the passage of the faeces (Michell et al, 1989: page 98).
In canines, it takes 22 hours for food to reach the colon from the mouth (Jacobs, 2005). Throughout this process the organs work together for the bolus to travel through the GI tract breaking down and absorbing nutrients along the way, with each organ having a different structure to aid with its function.
Aspinall, V. (2014) Anatomy and Physiology of the Dog and Cat, The Digestive System. Veterinary Nursing Journal, 19 (3).
Case, L., Daristotle, L., Hayek, M.
Laboratory Investigation of Gastrointestinal Stromal Tumour
Title: Describe the laboratory investigation of GIST.
Gastrointestinal Stromal Tumour (GIST) is one of the most commonly seen mesenchymal neoplasms, a soft tissue sarcoma, present in any area of the gastrointestinal tract (GI tract) and can have extra-gastrointestinal involvement as well. The stomach is the most common site (about 70%). GIST mainly affects middle aged to elderly adults, typically in their 60s with no clear gender preference although some studies demonstrated a slightly greater number of males affected. GISTs are tumours that begin in the smooth muscle cells of the GIT tract wall known as the pace maker interstitial cells of Cajal (ICCs). ICCs are cells that are part of the autonomic nervous system (Joensuu et al., 2002).
Classification of GISTs
Before the late 1990s, any non-epithelial tumours found in the GI tract were classified as GIST. However, with the development of molecular techniques, the molecular basis of GISTs was identified. Histopathologists were now able to distinguish between different types of tumours of the GI tract as either a GIST tumour or another type of tumour based on molecular differences. CD34 and CD117 are identified as markers that can help distinguish between these different types.
Signs and Symptoms of GIST
Symptoms of GIST can include GI tract bleeding, difficulty swallowing or metastasis, particularly the liver. Rarely is intestinal obstruction seen as a result of the outward growth of the tumour. Generally there is a history of vague abdominal pain or discomfort and the tumour is often large when diagnosed.
Pathophysiology of GIST
Unlike other tumours of the GI tract, GIST tumours are non-epithelial tumours, they are connective tissue tumours such as sarcomas. Most GIST tumours occur in the stomach, then small intestine and rarely in the esophagus. Small tumours are generally benign particularly when the mitotic rate is slow, whereas large tumours can metastasize to the liver, omentum or peritoneum. Other abdominal organs, they rarely occur in.
Almost all GISTs are sporadic, meaning that the mutations are random occurrences affecting single individuals. However, there are rare examples of GIST running in families or due to idiopathic multitumour syndromes. Inheritable germ line mutations, where these tumours are driven by mutations in the genes such as KIT (85%), PDGFRA (10%) or BRAF kinase (rarely). These patients usually have multiple GISTs. GIST rarely occurs along side other syndromes such as neurofibromatosis type I. However, individuals with this syndrome contain a higher chance of GIST development (Steigen and Eide, 2009).
An abnormal c-KIT pathway is associated with about 85% of GISTs. A transmembrane receptor for a growth factor called stem cell factor (scf) is what this c-KIT gene encodes for. Mutation of the gene itself is what causes most of the abnormal c-KIT pathway, however, a small number are due to the constitutive activity of the KIT enzymatic pathway. The product of the c-KIT gene is called CD117, which is found on ICCs, and mainly mast cells, melanocytes and bone marrow cells. However, in the GI tract, a tumour staining positive for CD117 is generally a GIST, originating from ICCs. CD117 is a vital diagnostic marker in diagnosing GIST. The mutations in the c-KIT gene, allow it to function in the absence of the scf, resulting in a high mitotic rate. Other mutations are required for the cell with the c-KIT mutation are required for the cell to progress into a GIST. This mutation is more than likely the first step.
A few GISTs with no mutation in the c-KIT gene, show mutations in the gene for a closely related tyrosine kinase receptor termed Platelet Derived Growth Factor Receptor Alpha (PDGFRA). C-KIT and PDGFRA mutations are mutually exclusive. GISTs with no mutation in either genes, is referred to as wild type tumours (Quek and George, 2009).
Diagnosis of GIST
Often CT scanning is undertaken. However, the definitive diagnosis is made using a biopsy, which is taken by endoscopy, percutaneously using ultrasound or CT or taken during surgery.
Microscopic morphology of GIST
In the histopathology laboratory diagnosis of GIST, microscopically, GISTS have a broad morphological spectrum (Joensuu et al., 2002).
GISTs may be malignant or benign. However, it is now considered that all GIST tumours have malignant potential and that GIST tumours cannot be definitively classified as benign. The benign features are small size, encapsulation (can remove easily with surgery), very low mitotic activity and absence of necrosis. The malignant features are presence of metastases at the time of surgery and may have an invasive margin and high mitotic activity (Steigen and Eide, 2009).
There are three primary histological sub-types of GIST which are spindle cell type (70%), epithelioid type (about 25%) and mixed spindle -epithelioid cell type. Generally, GISTs have a broad variation ranging from hypocellular to hypercellular with higher mitotic activity. Nuclear pleomorphism is uncommon and is found more often in epithelioid type (Katz and DeMatteo, 2008).
Figure 1: Common Haematoxylin and Eosin staining histological features of GIST.
(A) spindle cells with short fascicles and whorls.
(B) ) spindle cells with longer fascicles in bundles.
(C) spindle cells with extensive perinuclear vacuolization.
(D) spindle cells with prominent nuclear palisading.
(E) epithelioid cells with pleomorphic nuclei and vacuolated cytoplasm.
(F) epithelioid cells with rhabdoid features (Quek and George, 2009).
The GIST spindle cell type is comprised of cells arranged short fascicles and whorls.They exhibit pale eosinophilic fibrillary cytoplasm, oval nuclei and undefined cell borders. Gastric spindle cells generally exhibit vast perinuclear vacuolization. The distinctive histological patterns found in spindle cells are sclerosing type and palisading vacuolated type. The sclerosing spindle contain slender spindle cells with no nuclear atypia and low mitotic rate and are normally pauicellular with vast extracellular collagen. The sclerosing spindle cells are generally small and have calcifications. The palisading vacuolated spindle cell type is one of the most occurring gastric GISTs and normally cellular with uniform and plump spindle cells. Nuclear palisading spindle cells with perinuclear vacuolization is typical. There is normally limited atypia with mitotic rate rarely greater than 10/15 high power fields (Heinrich, 2003).
GIST epithelioid cells are typically round cells organised in nests or sheets and have eosinophilic to clear cytoplasm. They also contain spectrums from sclerosing and pauicellular to sarcomatous and mitotically inactive to mitotically highly active. Although the epithelioid cells that have atypia, even with pleomorphism are sometimes found to be benign (Hirota et al., 2003).
Immunohistochemistry of GIST
In the immunohistochemistry aspect of the laboratory, most GISTs (95%) are strongly and diffusely KIT (CD117) positive, which allows KIT to be a very specific and a sensitive immunohistochemistry marker in differentiating GIST diagnostically from other mesenchyma tumours in the GI tract. KIT is a cell membrane spanning signalling molecule (receptor for tyrosine kinase) that triggers cell growth when activated by a specific stem cell ligand. The stain is visible as cytoplasmic, membrane associated or as perinuclear dots. Even though KIT positivity has important treatment implications, the intensity and extent of KIT staining neither correlates with the type mutation in KIT nor the therapeutic importance. A negative result for KIT does not rule out the patient from treatment with tyrosine kinase inhibitor (TKI) (sunitinib or imatinib) as a few wild type GISTs for both PDGFRA and KIT genes respond to TKI treatment (Joensuu et al., 2002).
CD34 is another common immunohistochemistry marker for GIST, but is not as specific or sensitive. CD34 is positive in approximately 80% of gastric GISTs, 50% of small bowel GISTs and in 95% of colorectal and esophageal GISTs (Steigen and Eide, 2009).
Other immunohistochemistry markers which may be expressed by GISTs are SMA, h-caldesmon, desmin, S100, cytokeratins 8 and 18 and Vimentin. Recently other CD markers for GISTs are reported including CD10, CD133 and CD44 (Katz and DeMatteo, 2008).
Figure 2: Immunohistochemical staining features of GIST.
(A) Spindle cell with strong and diffuse cytoplasmic staining of CD117 (c-kit).
(B) Spindle cell with strong and diffuse membrane staining of CD34.
(C) Epithelioid cell with strong cytoplasmic staining of CD117.
(D) Epithelioid cell with patchy and heterogeneous staining of CD34.
(E) Epithelioid cell with punctate staining of h-Caldesmon
(F) Epithelioid cell GIST with patchy mambrane staining of h-Caldesmon (Quek and George, 2009).
A small minority of GIST (<5%) are negative for KIT, or minimally, if positive for KIT by immunohistochemistry. These tumours look to have either mutant PDGFRA or KIT wild type, have a preference to stomach or peritoneum and are usually epithelioid or mixed subtype. For this subgroup of GISTs that are negative for KIT, many new antibodies in the diagnosis of GIST have been identified. DOG1, a transmembrane protein, has been found specifically in GISTs. Studies have found that antibodies directed against DOG1 have higher specificity and sensitivity than KIT and CD34 with 75% to 100% sensitivity overall. KIT mutant GISTs highly express DOG1 and DOG1 can also detect about one third of GISTs that are negative for KIT, which generally have the PDGFRA mutation. DOG1, like KIT, is also found in interstitial cells of Cajal (Quek and George, 2009).
Differential diagnosis of GIST
In the differential diagnosis of GISTS, although GISTS are the most common mesenchymal tumour of the GI tract, a variety of other tumours should be included in the differential diagnosis. These are schwannoma, leiomyoma, leiomyosarcoma, inflammatory myofibroblastic tumor, inflammatory fibroid polyp, fibromatosis and melanoma. However, due to the identification of the molecular basis of GIST, histopathologists are able to now distinguish between different types of tumours (Steigen and Eide, 2009).
Most frequently intramural leiomyomas are present in the esophagus and are rarely in the small intestine and stomach. Leiomyomas are normally less cellular than GISTs, and in opposition to GISTs, they frequently display greater eosinophilic cytoplasm with well-delineated cell borders. Immunohistochemically, leiomyomas are diffusely positive for desmin, smooth muscle actin (SMA) and h-caldesmon and are KIT negative. Even though most GISTs are desmin negative, rare cases express desmin, which can result in misclassification as a smooth muscle tumor. Whats different to leiomyomas, desmin immunoreactivity is focal (Katz and DeMatteo, 2008).
Primary leiomyosarcomas are extremely rare of the GI tract. They generally are present in older adults and most frequently occur in the colon. In contrast to GISTs, leiomyosarcomas are comprised of spindle-shaped cells, with brightly eosinophilic cytoplasm, along with diffuse or focal nuclear pleomorphism and high mitotic rate. KIT positivity is extremely rare in leiomyosarcomas (Katz and DeMatteo, 2008).
Uncommon tumours are gastrointestinal schwannomas that are present in the stomach or colon and are rare in other parts of the GI tract. They display a dense, peripheral cuff of lymphocytes with or without germinal centers and are comprised of cells that exhibit strong immunoreactivity for glial fibrillary acid protein and S100 in the stomach. They are KIT and CD34 negative. They normally lack well-defined nuclear palisading, foamy histocytes, verocay bodies and hyalinized vessels that are characteristic of schwannomas everywhere else. Seen in up to 33% of GISTs is nuclear palisading and perivascular hyalinization. As well as morphological differences, GI schwannomas look to be genetically distinct compared with non-GI schwannomas (Quek and George, 2009).
Mesenteric fibromatosis or intra-abdominal desmoid fibromatosis because of its gross appearance as well as its location can be confused with GIST. It may occur either sporadically or in Gardner syndrome. Microscopically, it is has an infiltrative growth pattern, in contrast to rounded, expansile borders of GISTs. It is comprised of long, sweeping fascicles of spindle to stellate fibroblasts in a collagenous or keloidal stroma. The stroma in GISTs are more myxoid or hyalinized, and keloidal collagen, when present, they surround nests of cells rather than individual cells as is seen in fibromatosis. Another important feature of feature that aids to differentiate these tumours is the vascular pattern. Desmoid tumors display small, muscular arteries and dilated, thin-walled veins in the lesion. Nuclear β-catenin immunoreactivity is present in 75% to 90% of cases of fibromatosis, while it is negative in GIST (Quek and George, 2009).
Inflammatory myofibroblastic tumor are present in paediatrics and young adults and can occur as a mesenteric mass. Histologically, this tumour has greater heterogenous composition with spindle cell areas mixed with a mainly plasma cell-rich inflammatory infiltrate. These tumors are KIT negative. The marker used for diagnosing this tumour is anaplastic lymphoma kinase; but, it is found in less than 50% of cases. (Katz and DeMatteo, 2008).
A submucosa mesenchymal tumour is a inflammatory fibroid polyp that has a preference for the stomach and small intestine. In the morphology aspect, there is a proliferation of spindle and stellate cells that form whorls around blood vessels. The stroma has a granulation tissue like display and enriched with lymphocytes, eosinophils, and plasma cells. Inflammatory fibroid polyps are confused with GISTs as they are positive for CD34 too. Although they are KIT and DOG1 negative. Mutations in PDGFRA of the same type that are in GISTs have been seen in inflammatory fibroid polyps too (Quek and George, 2009).
Epithelioid GISTs have to be separated from carcinoma, melanoma, germ cell tumors, seminoma, clear cell sarcoma and glomus tumor. Melanomas display various of morphologic patterns, such as spindle and epithelioid cell types, and, so, are included in the differential diagnosis of GIST. This is even more complicated as they are KIT positive. Melanoma marker expression such as HMB-45, S100 or Melan-A, aids to solve this differential. The immunoreactivity of KIT in metastatic melanomas can be weaker than it is in primary tumors. Rare are Glomus tumors in the GI tract and are found exclusively in the stomach. They are a uniform cell population with pale eosinophilic staining cytoplasm with round nuclei that display strong immunoreactivity for smooth muscle actin and are negative for S100, desmin and KIT. Clear cell sarcomas of the GI tract are often HMB-45 negative. Apart from the characteristic morphology of large epithelioid cells that have numerous pale eosinophilic staining to nonstaining cytoplasm and stroma that is rich in lymphocytes, seminomas are able to be separated from GISTs using their distinctive immunoreactivity for transcription factor OCT4 and placental alkaline phosphatase. They can, however, exhibit diffuse immunoreactivity with KIT resulting in a potential diagnostic downfall.
Immunohistochemistry plays a vital role in the differential diagnosis. Combining the morphology and the immunohistochemistry results, in the majority of cases enables an accurate diagnosis (Quek and George, 2009).
The risk and staging of GIST
Different GISTs have different risk classifications of their tendency to reoccur or metastasize, dependent on their size, location, mitotic activity, surgical margins and the status of the rupture of the tumour. It is thought that establishing a level of risk for GIST (low, intermediate, or high) is better than calling the tumor as malignant or benign. The most indicators of aggressive behavior are tumor size of 5 cm and 5 mitoses/50 HPF. Risk classification also incorporates the primary site of the tumor as well as the mitotic count and its size. Gastric GISTs have a better prognosis as they have a malignant potential that is lower than other GISTs found in other areas of the gastrointestinal tract. Anatomic site is a parameter in risk assessment for GIST. GISTs that are smaller than 2 cm are considered benign. Additional factors such as rupture of tumour and non-radical resection, are both linked to an adverse outcome for the patient, that are not affected by any other factors. The presence of a ruptured tumor is regarded as a high risk factor irrespective of its mitotic count or size (Joensuu et al., 2002).
All GISTs, benign or malignant are eligible for staging. In the TNM staging, GIST grading is based on mitotic activity. Mitotic activity lower than 5/50 HPFs (High Power Fields) is low (grade 1) and more than 5/50 HPFs is high (grade 2). However, different staging criteria is used for small intestinal GISTs and gastric GISTs to highlight the more aggressive clinical nature of small intestinal GISTs even with similar parameters of tumor (Steigen and Eide, 2009).
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