 Before studying this lesson, you should have an understanding of the formation of exudates and the role of leukocytes in phagocytosis of bacteria. This discussion will cover various chemical mediators which play a role in acute inflammation. This is a complex subject because a large number of mediators have been identified and many interacting physiological systems are involved. It is worth remembering that although the activity of many mediators has been demonstrated in experimental situations, their precise role in specific examples of inflammation is not clear. Two types of mediators are recognized, exogenous and endogenous. The exogenous mediators are primarily chemotactic factors from bacteria. Some injurious agents may also enhance vascular permeability by direct injury to vessel walls. The endogenous mediators are much more numerous and important. They may be derived from cells or from plasma. Histamine, which is derived from mast cells, is the most important vasoactive amine. It rapidly causes vasodilation and increased vasopermeability, but it has a short half-life. Two other important mediators of inflammation are derived from cell membrane lipids by oxidation. These are leukotrains and prostaglandins. The first step in formation of these substances is the release of arachidonic acid from membrane lipids by phospholipase. Phospholipase is present in the lysosomes of polymorphonuclear neutrophils and these cells are, of course, present in large numbers in inflammatory exudates. The arachidonic acid is oxidized in two pathways. The cyclooxygenase pathway leads to production of prostaglandins. The lipoxygenase pathway leads to production of leukotrains. Here are some of the important prostaglandins. Except for thromboxane, which causes vasoconstriction, as well as being a potent agent for platelet aggregation, all the prostaglandins cause vasodilation and potentiate vascular permeability. It is worth noting at this point that aspirin and the non-steroidal anti-inflammatory agents, such as endomethacin and isopropan, inhibit cyclooxygenase and therefore inhibit prostaglandin production. Lipoxygenase enzymes are also derived from neutrophils and act on arachidonic acid to produce leukotrains. Each leukotrine is derived from the preceding one by hydrolysis or addition of glutathione. Leukotrine B4 is strongly chemotactic and leukotrains C4 and D4 together are also known as the slow reacting substance of anaphylaxis. They cause bronchospasm by smooth muscle constriction as well as a profound increase of vasopermeability. Prostaglandins and leukotrains cause a delayed onset of vasodilation and prolonged increase in vascular permeability, thus resulting in formation of inflammatory exudates several hours after certain types of injury. In this way, their effects are quite different from the rapid and short-term action of histamine. Substances derived from plasma that are important mediators of inflammation include the kinins. The most important one is bradykinin. It is a small polypeptide which is released from a large precursor molecule called a kininogen. Kininogens are normal constituents of blood. The enzyme which releases bradykinin from kininogen is called calicrine. It in turn is formed from pre-calicrine by activated Higaman factor. Higaman factor is one component of the intrinsic blood coagulation pathway. It is a normal serum protein which is activated by contact with collagen and by enzymes such as plasma. Bradykinin acts very much like histamine. It causes vasodilation and separation of endothelial cells, thus increasing vascular permeability. It acts rapidly and also has a short half-life. It also stimulates nerve endings and is responsible for much of the pain associated with inflammation. The plasma system also generates mediators of inflammation primarily by the release of split products from the degradation of fibrin. Serum plasminogen is converted to the active enzyme plasmin by activated Higaman factor. The split products of fibrin are chemotactic and can also enhance vascular permeability. Activation of complement can occur during inflammation because both plasmin and calicrine are capable of splitting the third component of complement into its active fragments C3A and C3B. The latter activates C5. The small polypeptide fragments C3A and C5A cause degranulation of mast cells and histamine release. C5A is also chemotactic for leukocytes. Note that Higaman factor, when activated by contact with collagen, is capable of activating the coagulation cascade, the plasmin system, and the generation of kinins. Indirectly, the complement system will also be activated through the action of plasmin or calicrine. The complex interaction between these primary physiological systems may be accurately called the tangled web. Probably only a few mediators are important in any one specific example of inflammation. The long list of mediators of inflammation would be incomplete without mention of some polymorphonuclear neutrophil derived substances that are chemotactic or facilitate exudate formation. Likewise, the products of tissue degradation include substances which provoke inflammatory responses. Now for an in-depth examination of the systemic effects of inflammation. Five recognizable systemic effects will be discussed. Fever, leukocytosis, C-reactive protein, sedimentation rate, and coagulability of the blood. Fever is due to the action of substances called pyrogens on the temperature-regulating centers of the hypothalamus. Pyrogens are low molecular weight substances derived from exogenous sources such as bacteria or endogenously from necrotic tissue or leukocytes. An important endogenous pyrogen is interleukin-1. It is a low molecular weight protein released from activated macrophages. Leucocytosis can occur very rapidly reaching concentrations of 20 to 30,000 neutrophils per milliliter of blood. Two processes are involved. One is the rapid release of mature neutrophils normally sequestered in lungs and bone marrow. Agents such as adrenaline, prostaglandins, and activated complement components will initiate this type of leukocytosis. Direct stimulation of bone marrow to produce additional neutrophils can also result in leukocytosis. The agent involved is called colony-stimulating factor. It is produced in inflammatory exudates and can be isolated from blood or urine of patients suffering from severe infections. It is a glycoprotein derived from macrophages and T lymphocytes. Leucocytosis is usually associated with the release of younger or even immature neutrophils from bone marrow. In blood films, this is referred to as a shift to the left in the neutrophil population. Younger neutrophils have fewer nuclear segments or may even have non-segmented nuclei, in which case they are called band forms. In severe infections, metamilocytes or myocytes will occasionally appear. The appearance in the blood of acute phase reactants such as fibrinogen and C-reactive protein is a poorly understood phenomenon. C-reactive protein is not present in normal plasma, but it appears during inflammation. It is derived from the liver and one stimulus for its release is interleukin-1, which, as has been noted, is derived from activated macrophages. It also acts as a pyrogen. Elevation of the sedimentation rate of the blood is a non-specific phenomenon often observed in inflammation and in other disease processes. It is due to a change in the surface charges of red cells brought about in large part by an increase in plasma fibrinogen. Finally, increased coagulability of blood occurs during inflammation. It is due to enhanced stickiness of platelets as well as increased blood levels of coagulation factors such as fibrinogen. Acute inflammation has many different causes and can occur in almost any tissue. The specific type of injury and the type of tissue influence the character of the exudate which develops. The various kinds of exudates will now be described. The important ones are cirrus, fibrinous, membranous, and purulent. In early stages of inflammation or even in later stages if the inflammation is very mild, fluid components of the exudate predominate. And it is therefore called a cirrus exudate. Few, if any, polymorphonuclear neutrophils are present. In this photomicrograph a cirrus exudate is accumulating between heart muscle fibers due to a recent nearby infarction. Note that there are only a few neutrophils and the space between the muscle fibers seems to be empty or contains only protein preserved by the fixative. Viral infections may cause a cirrus pericarditis or a cirrus pleuritis. The fluid which accumulates in the pericardial sac or pleural spaces in such infections contains abundant protein but few inflammatory cells. Another form of inflammation characterized by a cirrus type of exudate is cellulitis. This is a particularly dangerous form of inflammation caused by streptococci which secrete hyaluronidase, an enzyme which breaks down the hyaluronic acid matrix of connective tissue. Once such a degradation occurs organisms can spread rapidly through the tissue. Beginning with a local infection an entire limb can rapidly become swollen, red, and tender. If not treated properly such infections will end an overwhelming systemic infection and death. These photographs demonstrate the changes seen in cellulitis. In addition to vascular dilation there is abundant fluid separating the muscle fibers with a few neutrophils. There is a cirrus exudate in the affected tissues and special stains will usually reveal the invading streptococci. Recall now that the fluid component of an inflammatory exudate contains all the proteins normally found in the blood including those of the coagulation cascade. If a large amount of fibrin accumulates in the exudate as a result of activation of the coagulation cascade then the exudate is called fibrinous. Fibrinous exudates are seen most often on the surface of inflamed organs such as heart, lungs, or peritoneum. Here we see a heart covered by a shaggy exudate which hides the pericardial surface. Microscopic examination shows the typical appearance of a fibrinous pericarditis. Note the large masses of bright red fibrin on the surface of the heart surrounded by various inflammatory cells. The pericardial tissues are inflamed. Note also the engorged dilated blood vessels, the evidence of fluid accumulation, and the many inflammatory cells including typical macrophages. The term serofibrinous exudate is sometimes used to describe a cirrus exudate mixed with abundant fibrin. Membranous exudates are seen in the gut and the tracheobronchial tree. The example shown here is due to diphtheria which caused formation of a loose white or grey membrane lining the surface of the trachea. Diphtheria is caused by corinabacterium diphtheriae which establish themselves on the surface of the tracheobronchial mucosa. They secrete a powerful exotoxin which kills the mucosal epithelial cells. An acute inflammatory exudate forms on the mucosal surface. This constitutes the membrane which is seen in these photomicrographs. At higher magnifications the membrane is seen to be made up of abundant fibrin which has trapped large numbers of polymorphonuclear leukocytes. Necrotic mucosal epithelial cells can be seen occasionally and many of the leukocytes are also undergoing degeneration and necrosis. The bacteria cannot be seen without special stains. The diphtheric membrane is only loosely attached to the bronchial or tracheal wall. It can easily be detached and cause asphyxiation especially in children. Another example of a membranous exudate can be seen in the gut infected with claustridia difficile. This sometimes occurs when antibiotics are given to reduce intestinal bacterial flora before surgery. Once again the bacteria secrete a powerful toxin which kills the mucosal epithelial cells and a pale membrane develops over the mucosal surface as seen in this photograph. The condition is called membranous enteritis or membranous enterocolitis and is very frequently fatal unless promptly treated with antibiotics effective against claustridia difficile. With many bacterial infections there is an outpouring of neutrophils and these cells become the most conspicuous feature of the acute exudate. If the cause of the injury is not quickly removed the neutrophils accumulate in massive numbers and a purulent exudate develops. The process is then referred to as a sub-puritive infection. Both of these terms refer to the presence of pus. Pus consists of living and dead neutrophils usually mixed with bacteria and tissue debris. This is an example of liquefactive necrosis. When pus forms it will soon be surrounded by proliferating fibroblasts and blood vessels infiltrated with neutrophils and macrophages. This is called a pyogenic membrane and it walls off the pocket of pus. Such a pocket of pus is called an abscess. Once an abscess forms it becomes very difficult for the inflammatory process to eradicate the infection. Organisms in the pocket of pus are inaccessible to circulating chemotherapeutic agents. Although they may not grow rapidly because of the acid pH and low oxygen tension of pus they persist and the abscess tends to grow larger by progressive necrosis affecting the inside of the pyogenic membrane. The tissue and cell breakdown involved in pus formation also increases the osmotic pressure so that water is drawn in further increasing the size of the abscess. This process will continue until the abscess ruptures through a surface such as skin, bronchus, pleura or peritoneum or until it is artificially drained. Once pus is expelled the abscess will usually heal with formation of scar tissue. Abscess formation means that the inflammatory process has not been successful in eradicating the bacteria but it has been successful in walling off the infection and preventing it from spreading. Look at an example of experimentally induced inflammation in the skin of a rabbit. Inflammation was produced by injecting a small amount of antibody from a rat previously immunized by rabbit tissue antigens. The antigen antibody reaction which occurs in the rabbit skin is an immunological form of tissue injury and results in acute inflammation. The first picture is rabbit skin at low magnification. Note the epidermis, the dermis containing densely packed collagen and hair follicles and beneath the dermis a layer of skeletal muscle. Underneath the muscle is loose subdermal connective tissue. The inflammatory reaction occurs in the dermis just above the muscle which was the site of injection. In low power observation of a section taken from the biopsy performed six hours after injection many dilated vessels are apparent in the deep layers of the dermis. In the surrounding tissue an early inflammatory exudate is forming. The collagen bundles of the dermis are separated by clear spaces of pale pink material. This is the fluid component of the inflammatory exudate. It is accumulating outside the blood vessels and causing the tissue to swell. This exudate contains an abundant amount of protein derived from blood. It is a serous exudate. Cellular components of the inflammatory exudate are recognizable as a mixture of polymorphonuclear and mononuclear cells. Notice the typical clover leaf of the multi lobe nucleus and the pink cytoplasm of the neutrophil. The lymphocytes present have round, dense nuclei with little or no visible cytoplasm. These cells are often clustered around the dilated small blood vessels and probably emigrated shortly before the biopsy was made. In some small vanuals, margination of polymorphonuclear neutrophils can be seen. These observations are typical of early acute inflammation. The next biopsy was taken 18 hours later. At low power, notice that the dilated blood vessels are still present and the inflammatory infiltrate is more prominent. Higher magnification of this 24-hour sample reveals that although many polymorphonuclear cells are still present, mononuclear types are becoming more numerous. In addition to the lymphocytes, typical macrophages with pale nuclei and abundant cytoplasm can be seen. These pictures demonstrate how quickly the character of an inflammatory reaction can change, at least in such a mild form of injury. The final biopsy was taken after 48 hours. It shows the inflammation undergoing resolution. A few dilated vessels are still present, but there is little edema left. The inflammatory cells are now mostly macrophages. Few polymorphs remain. The macrophages will gradually disappear over a period of several days while the tissue returns to normal. Histological signs of acute inflammation can be observed particularly well in lung tissue. An inflammatory exudate readily forms under conditions such as those resulting from bacterial pneumonia. In early stages, an exudate develops quickly and replaces air in the alveoli. At low magnification, alveolar spaces are filled with fluid, which stains pink because of its protein content. At higher magnification, the alveolar walls appear intensely hyperemic. The capillaries are widely dilated and filled with red cells. Notice that the fluid in the alveolar spaces contains scattered polymorphonuclear neutrophils and red cells. Also, there are a few macrophages to be seen. Macrophages are normal constituents of lung tissue. In many alveoli, strands of fibrin are present. This fibrin results from activation of the coagulation cascade in the exudate. Sometimes bacteria can be recognized. As time passes, more and more neutrophils accumulate in the alveoli as the cellular events of the inflammatory process become fully established. The influx of neutrophils gradually obscures all other structures. Blood is squeezed out of the capillaries so that the alveolar walls become difficult to see. The lung tissue at this stage will assume a grayish-white hue due to the large number of leukocytes and the diminished blood flow. As more time passes, the neutrophils die and the process of resolution begins as macrophages become more and more prominent. The arrow is pointing to a typical macrophage. Macrophages actively phagocetize and remove the debris from the area, carrying it to regional lymph nodes or to the bronchi where it may be expelled by coughing. Of course, the proteolytic enzymes released from dead leukocytes also assist in dissolving tissue debris and fibrin. Eventually, all components of the inflammatory exudate can be reabsorbed and the lung restored to normal. Resolution of bacterial pneumonia will be facilitated by appropriate antibiotics and by the presence of specific antibodies. Acute appendicitis is another disease caused by bacterial invasion. Here is a low-power magnification of a stained section of inflamed appendix. The lumen is lined with glandular epithelium. Note the prominent lymphoid follicles in the mucosa. Beneath the mucosa is a thick submucosa, next a layer of smooth muscle, and finally on the outside a thin serosal surface. Acute inflammation is apparent in all parts of the appendix. Here at the mucosa, the epithelium is interrupted. That is a small ulcer. The base is composed of densely packed polymorphonuclear leukocytes. This would be called a small focus of supuration. Note the typical dark, multi-lobed nuclei and pale pink cytoplasm of these cells. Moving into the submucosal tissue, note that polymorphonuclear neutrophils are numerous. The blood vessels are dilated. The edema is present, but difficult to distinguish because of the large number of inflammatory cells. In the muscular wall of the appendix, there are also numerous neutrophils, which tend to lie in rows between the elongated smooth muscle fibers. Out at the serosal surface, a typical acute inflammatory exudate is seen. Numerous neutrophils are embedded in coagulated protein. This coagulum is on the surface of the serosa, which itself is inflamed with many dilated vessels and infiltrating polymorphonuclear neutrophils. A discussion of acute inflammation would be incomplete without a brief look at a lung abscess. Such abscesses form during severe bacterial pneumonias and can also be caused by aspiration of infected material. In this low-power view of lung tissue, aerated lung can be seen at the upper right and part of an abscess lies at the lower left. The abscess was about 5 centimeters across. Scanning towards the abscess, notice the inflamed pulmonary tissue just outside the actual abscess. Alveolar walls are thickened. They contain polymorphonuclear neutrophils and macrophages. The capillaries are dilated and the alveoli nearest the abscess are compressed and airless. The wall of the abscess is called the pyogenic membrane. It consists of dilated, blood-filled capillaries surrounded by infiltrating neutrophils and macrophages. Many fibroblasts are present and can be recognized by their elongated shape. Collagen is also present. The amount of collagen seen in such cases will depend upon the severity and duration of the process. It is difficult to recognize all of the cell types in paraffin sections. Scanning further towards the center of the abscess, masses of fibrin can be seen just underneath the pyogenic membrane. Further on, there is amorphous granular material, which is the result of degeneration of leukocytes and liquefaction necrosis of pulmonary tissue. Still further toward the center of the abscess, more intact dead and dying neutrophils are recognizable. Sometimes fragments of elastic fibers can be observed in such abscesses, since these tissue elements are resistant to enzymatic digestion. This is the same histologic picture of pus, which was seen earlier. Acute inflammation, then, can occur throughout the body, and wherever it occurs, it will have similar characteristics which are modified only slightly by the type of tissue involved. The underlying processes are always the same, and the sequence of events is carefully orchestrated to result in characteristic and typical histological pictures at every stage.