 This program examines some of the morphological expressions of cell injury. Cells and tissues may respond to environmental changes by using several adaptive mechanisms, such as atrophy, hypertrophy, hyperplasia, and metaplasia. However, there are limits within which a cell is capable of adapting to environmental stress. When such limits are exceeded, the cell can no longer adapt and it will show signs of injury. It goes almost without saying that the threshold for injury by any given stress varies from cell to cell. As shown here diagrammatically, the injured cell may still revert to normal, provided the stress has not been too great and restoration of a normal environment begins within a finite period of time. There are many diverse substances and forces within the environment which may produce cell injury. Many such environmental changes are the same ones which elicit adaptive reactions from cells. However, injury is the result if the stress is too great or lasts too long. There is considerable variability among cells in their sensitivity to changes in the environment. For example, hypoxia, for a period of only a few minutes, is sufficient to cause injury to neurons. Fibrocytes, on the other hand, may withstand hypoxia for long periods without injury. The basis for this difference lies largely in the metabolic activity of the cell. Environmental variations which may produce injury include such diverse phenomena as excessive mechanical forces, temperature extremes, electricity, ionizing radiation, biological and chemical agents, and the interaction of antibodies with various antigens. You should keep in mind that reference to the environment means external as well as internal environment. Although there are many and diverse environmental variations which may cause cell injury, cells and tissues can react in only a limited number of ways. Most cell injury involves interference with membrane functions, energy production, and or enzyme systems. After a cell has been injured, some time must elapse before the morphological changes that ensue become detectable. Morphologic indications of some types of cell injury may be detected within a few minutes by electron microscopy. Ordinarily, longer periods of time are required for such changes to become visible with a light microscope. How are cell injuries detected? Findings such as dilatation of the endoplasmic reticulum and mitochondria, loss of ribosomes, decreased intracellular glycogen, and increased calcium are common expressions of cell injury. Also, nuclear changes such as condensation of chromatin on the nuclear membrane may occur. These changes are primarily due to loss of membrane function secondary to interference with energy metabolism of the cell. Any interference with energy production results in a decrease in ATP concentration, causing the sodium pump to fail. The cell membrane then becomes leaky, permitting an influx of sodium, calcium and water, and efflux of potassium and other cellular components, including enzymes. An influx of water causes dilatation of subcellular organelles. This can be demonstrated using electron microscopy as shown in the diagram. The influx of enzymes into the interstitial fluid can be of clinical significance, since detection of elevated levels of certain enzymes in the serum may reflect cell injury. To illustrate the use of electron microscopy in the detection of cell injury, some electron micrographs will now be examined. The first electron micrograph shows a normal rat liver cell. This large oval structure at the edge of the picture is the nucleus with a normal chromatin pattern. These smaller, round to ovoid structures are mitochondria. You should take note of the appearance of normal internal membranes or Christi. These groups of parallel membranes are the rough endoplasmic reticulum. Between these cellular organelles, the cytoplasmic matrix can be seen. These small dark clusters are normal deposits of glycogen. Now you see an electron micrograph of rat liver that had been experimentally injured by acute poisoning with methylmercury. This preparation was made four hours after exposure to the poison. Cellular edema is detectable. This vertical line represents the border between two hepatocytes, and here is a bile canaliculus lying between the liver cells. Over here is the nucleus of one of the cells. Note that the cell matrix is pale, reflecting increased water content. Look at these mitochondria. They are enlarged as a result of the influx of water. They are pale and so distended that the Christi are widely separated and compressed toward the periphery. For comparison, note this more normal mitochondrion over here. To give you an opportunity to compare the electron micrographs of normal and injured rat hepatocytes, this frame shows a normal cell on the left and a cell poisoned with methylmercury on the right. The next two illustrations present electron micrographs of rat neurons that further demonstrate early changes in cell injury. This is an electron micrograph of a normal rat neuron. This round structure is the nucleus. These are nucleoli. Outside the nucleus here are the cell matrix and subcellular organelles. Mitochondria are seen here, and these parallel lines are part of the cytoplasmic membrane system. The next three slides demonstrate cells damaged by a neurotoxin trimethyltin. This swollen cell shows changes 48 hours after exposure to the toxic agent. This is the nucleus, and the dark area here is a nucleolus. Surrounding the nucleus are membranes of the endoplasmic reticulum, showing dilatation which resulted from the influx of water. For comparison, look at the previous two electron micrographs side by side, showing a normal neuron on the left and an injured one with a dilated endoplasmic reticulum on the right. Finally, this rat neuron shows a more advanced stage of cellular edema, 72 hours after exposure to trimethyltin. The nucleus is here, and it has been distorted by the accumulation of water inside the cell. This dark area is the nucleolus, and these large clear spaces represent water accumulated by the endoplasmic reticulum. Many of the membranes have coalesced into larger membrane-bound structures. These structures with dense sheaths are axons. Turning now to observations made with the light microscope shown here diagrammatically is the result of cellular imbibition of excess water. The cells become visibly enlarged and take on a slightly opaque appearance. This phenomenon is referred to as cloudy swelling or cellular edema. Not only are the individual cells distended, but the entire organ is larger due to the increased water content. Remember, this is a reflection of fluid accumulation or edema and should not be confused with hypertrophy or hyperplasia. Organs containing a large population of epithelial cells such as the liver or kidney are ordinarily used to demonstrate cell injury. In these organs, early signs of injury are more easily detected. However, such expressions of cell injury occur in all cells and organs. Changes that occur in injured cells may also be seen by the light microscope. Good examples may be found in renal tubular epithelial cells that have been injured by hypoxia, hepatocytes previously exposed to a hepatotoxic virus, liver cells injured by ethyl alcohol, and myocardium damaged by metallic phosphides. This slide shows on the left the normal epithelial cells that form a part of the proximal convoluted tubule. Note the normal pink stain cytoplasm and the blue nucleus. On the right are cells from a kidney that have been injured by hypoxia. The structure at the tip of the pointer is a section of a proximal tubule. The epithelial cells are so swollen that the tubular lumen is obliterated. If you examine the cytoplasm carefully, you should be able to detect very small clear vacuoles which are subcellular organelles distended with water. Now you can see the same alterations at higher magnification. Here is another example that demonstrates more extensive cellular edema. Note the prominent water vacuoles. When cellular edema progresses this far, the condition is referred to as hydropic or vacular degeneration. It is also worthwhile to examine nuclear changes that result from cell injury. The left side of this slide shows normal tubular epithelial cell nuclei. The chromatin material has a diffuse reticular pattern and there is a prominent nucleolus. On the right you can see the central nuclear clearing and condensation of chromatin material that often occurs in cellular edema. These nuclear changes are secondary to water accumulation and alterations in intracellular pH. All of the changes that have been demonstrated are morphological expressions of reversible cell injury. In life, these cells could have recovered and reestablished normal homeostasis. Had the environment been restored to normal within a finite time frame? Swelling can also be demonstrated in liver cells. In this case the cause of injury was a virus. On the left side of this frame are normal hepatocytes with pink cytoplasm and distinct nuclei. On the right is a group of hepatocytes that had been injured and subsequently had imbibed water. Note also in the right hand frame the slight haziness of the cytoplasm. Higher magnification of the same tissue may make the changes more obvious to you. The cell at the end of the pointer shows no signs of irreversible damage. Had this cell remained in vivo it would have had the potential to recover normal function. Other nearby cells exhibit changes which indicate that they were irreversibly damaged and are now dead. These features will be discussed when cell death is considered. Another common manifestation of cell injury is the appearance of fat within the cell. Certain cells such as lipocytes, adrenal cortical cells and cells of the corpus luteum normally contain lipids that are visible by light microscopy. Most cells however do not contain lipid in sufficient quantity to make it visible by the light microscope. Therefore the appearance of fat in some cells may be an indication of cell injury. Injured liver cells frequently accumulate fat. This diagram depicts some aspects of lipid metabolism in liver cells. Dietary lipids reach the liver in the form of chylomicrons via the portal vein. The triglycerides in the chylomicrons are broken up into glycerol and free fatty acids by hepatocyte enzymes. These products of enzymatic degradation are then reassembled back into triglycerides and bound to proteins that are synthesized by the hepatocyte. These protein bound lipids called lipoproteins are then secreted into the hepatic sinusoids and leave the liver via the hepatic veins. Common agents that damage hepatocytes are ethyl alcohol, usually consumed orally, and chlorinated hydrocarbons, usually by inhalation. These substances interfere with hepatocyte metabolism and among the enzymatic reactions affected are those involved in lipoprotein synthesis. Triglycerides cannot be secreted by the hepatocyte until they are bound to protein. Consequently, when lipoprotein synthesis is disturbed, triglycerides accumulate in the hepatocyte. If the intracellular concentration becomes high enough, the triglycerides coalesce into visible droplets. Following injury, myocardial cells often accumulate fat. Muscle, including cardiac tissue, utilizes fatty acids as a primary substrate for ATP production and is therefore actively involved in lipid metabolism. In muscle, then, as in liver, cell injury may result in intracellular fat accumulation. Although abnormal fat accumulation may occur in practically any cell which participates in lipid metabolism, historically, the liver and the heart, particularly the liver, are generally used as examples when abnormal fat accumulation is considered. This is a section taken from a normal human liver. Note the chords of hepatocytes with eosinophilic cytoplasm. Now you see the same tissue under higher magnification. Notice that the cell nuclei are mostly round and located near the centers of the hepatocytes. Examining injured hepatocytes, notice this nucleus which is deformed and displaced to the periphery of the cell. This large, clear vacuole represents a globule of lipid. Since lipid is hydrophobic, all of the hydrophilic cell components have been displaced toward the cell membrane. To give you an opportunity to compare normal and injured liver cells, this frame shows normal cells on the left and injured cells filled with lipid on the right. Note that the nuclei on the left are round and centered. On the right, they are displaced to the periphery by large fat globules. It is not difficult to imagine that a cell such as this one might have problems in carrying on normal functions. In fact, if such severe fat accumulation affected large numbers of cells, clinical signs of liver dysfunction would be present. In cells such as these, you see several smaller, clear vacuoles. Notice that the hepatocyte nucleus is still round and not yet flattened against the cell membrane. This represents an earlier stage of fatty change. With progressive fat accumulation, the smaller droplets would coalesce into larger globules and compress the intracellular contents against the cell wall. Now examine the section of normal liver on the left and compare it with the section on the right in which small fat globules are contained in the hepatocyte. The lipid first accumulates as liposomes, which are membrane-bound structures probably derived from endoplasmic reticulum. These liposomes later transform into non-membrane-bound droplets of neutral fat that coalesce into larger and larger lipid globules. Examining an electron micrograph of a rat liver cell will give further insight into this phenomenon. Here is a bile canaliculus between two hepatocytes. Over here is the edge of a nucleus. Here are a few mitochondria, and these structures are membranes of the rough endoplasmic reticulum. Notice especially these small membrane-bound structures containing electron-dense droplets. These are liposomes, and the dark droplets are lipoproteins. This represents a very early stage of abnormal lipid accumulation in cells that have been injured by methylmercury. This electron micrograph represents a slightly later stage of lipid accumulation. Here is the nucleus. These structures are neutral lipids, which are not membrane-bound. These lipid droplets are formed by coalescence of liposomes and loss of enveloping membranes. These droplets continue to coalesce until they form the large clear spaces seen in light microscopy. The H&E sections of fatty livers you have seen were processed in the routine manner, which involves exposure to various fat solvents. During such routine tissue processing, lipid was extracted from the tissue, producing the space or vacuoles where the lipid was before it was dissolved. This slide shows a section of liver prepared by frozen-section technique to avoid contact with fat solvents. A red dye called oil-red-o was used to stain fat selectively. Here you see the lipid stained bright red in the hepatocytes. Under high magnification, the red-stained lipid is even more striking. This is a section of myocardium from a young man accidentally poisoned by metallic phosphide. This tissue was stained with Sudan black, another dye which selectively binds to lipid. The black areas are collections of fat within injured myocardial cells. Under high magnification, perhaps you can better appreciate the presence of intracellular lipid.