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Mucosal Permeability and Smooth Muscle Function in Asthma
Obstructive Lung Disease
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Mucosal Permeability and Smooth Muscle Function in Asthma James C. Hogg, MD, PhD, FRCP(C)*
The changes in airway structure that occur in asthma are characteristic of a chronic inflammatory process that involves tissue covered by a mucosal surface. 12. 13 This process produces changes in the airway epithelium and in the smooth muscle surrounding the airways that could contribute to abnormal airways function in asthma. In this article will be a brief review of the inflammatory process and a presentation of evidence that the structural changes seen in asthmatic airways are a result of this response. The contribution of the airway muscle and epithelium to the abnormal function of asthmatic airways will then be reviewed.
THE STRUCTURAL FEATURES OF THE INFLAMMATORY RESPONSE Following tissue injury, there is a rapid series of changes in the microvessels that result in vascular congestion and the escape of plasma into the tissue. 13 The balance between exudation from the vessels and fluid drainage into the lymphatic vessels determines the degree of tissue swelling. The exudation of fluid from the vascular space is followed by the appearance of platelets, polymorphonuclear leukocytes, monocytes, and lymphocytes in the inflammatory site. This greatly increases cell traffic in the damaged tissue and sets off a number of events, which occur simultaneously, but must be described separately. These events determine the structural features of the inflammatory response in all tissues but certain tissuespecific changes occur in some organs. For example, airways, like other structures covered with a mucosal surface, show two important tissuespecific features of the inflammatory response. 12 The first of these is a shedding of the epithelium, which can sometimes occur in large clumps. In asthma, these clumps are referred to as "Creola bodies" after the patient *Professor of Pathology, University of British Columbia; and Director, Pulmonary Research Laboratory, St. Paul's Hospital, Vancouver, British Columbia, Canada
Medical Clinics of North America-VD!' 74, No. 3, May 1990
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(Creola Jones) in whom they were first described. 2H The second is an excess secretion of mucus from both epithelial lining cells and submucosal glands. The combination of the exudation of fluid and cells from microvessels, shed epithelium, and mucus hypersecretion often fills the airways with tenacious material referred to as a mucus plug. In fact, the majority of the material occluding the airway is an exudate of plasma and cells that contains a very small amount of mucus. In the repair phase of the inflammatory response, epithelial cells multiply and cover the denuded surface. The rapid turnover of the epithelium probably results in the characteristic thickening of the basement membrane in this condition. Similarly, the proliferation of the microvessels, connective tissue, and muscle results in a thickening of the submucosa. Mediators of the Inflammatory Response The mediators of the various events that characterize the inflammatory response are chemical in nature. These chemicals are derived from the cells that are resident in the tissue, from the plasma exudate, and from the cells that migrate into the tissue in response to injury. The account below provides a very brief overview of the complex subject and is designed to outline a basis for this discussion of mucosal and smooth muscle function in asthma. Tissue-derived Mediators. An important group of mediators of the inflammatory reaction are derived from peroxidation of arachidonic acid, which is a major constituent of lipid in cell membranes. Arachidonic acid can be metabolized in two ways. The cyclo-oxygenase pathway involves two unstable endoperoxide intermediates that are metabolized to stable prostaglandins, such as prostaglandin E z, a potent vasoconstrictor and platelet activator thromboxane A2 , a dilator and antiaggregatory compound, prostaglandin 12 , On the other hand, the lipoxygenase pathway of arachidonic acid peroxidation produces a family of compounds known as leukotrienes. One of these compounds (leukotriene C) is now known to be identical with a slow-reacting substance of anaphylaxis, which has been implicated as a possible mediator of the prolonged smooth muscle contraction seen in asthma. The best known tissue-derived mediator is histamine, which is formed by decarboxylation of histidine in the granules of mast cells. These cells are found in high concentration around the airways particularly in the distal parts of the lung. Because these distal airways are muscular, histamine is an important candidate as a mediator of airways narrowing. A third group of tissue-derived mediators (the tachykinins) are produced by antidromic stimulation of sensory nerves and are thought by some to be important in some forms of bronchial hyperreactivity. Plasma-derived Mediators. Hageman factor is activated when plasma comes in contact with a foreign surface during the exudative process. Activated Hageman factor converts several plasma proenzymes to their active form. This in turn results in activation of the complement, coagulation, kinin, and fibrinolytic cascades, which generate a host of inflammatory mediators. Mediators derived from the complement cascade can be generated through the classical pathway that is triggered by antigen antibody complexes or by the alternate pathway by endotoxin and by plasma proteins
MUCOSAL PER:vJEABILlTY AN]) S!\100TH .MUSCLE FUNCTION I!\I ASTH~jA
733
and proteolytic enzymes contained in the plasma exudate. In either event, this cascade releases potent mediators of the inflammatory response into the tissues. Mediators Derived from Migrating Cells. The cell population migrating into an acutely inflamed area contributes a variety of inflammatory mediators. The neutrophil can release cationic proteins as well as acid and neutral proteases from its granules. Activation of the nicotinamide-adenine-dinucleotide phosphate (NADPH) oxidase in the neutrophil membrane generates a variety of toxic oxygen species including the superoxide anion, hydroxyl radical, and hydrogen peroxide. The interaction of H 20 2 with halide ions and myeloperoxidase from the neutrophil granules produces a variety of short- and long-acting oxidants that can destroy invading bacteria. Monocytes migrating into inflamed tissue transform to macrophages that are capable of processing antigen and generating many important cytokines capable of stimulating the proliferation of T- and B-lymphocytes. The cytokines produced by monocytes and lymphocytes affect antibody production as well as the function of neutrophils, eosinophils, monocytes, and lymphocytes in the inflammatory response. The major problem with understanding the function of individual mediators is that many of them function simultaneously and produce interactive results. Although individual mediators can be isolated and studied in a controlled fashion both in vitro and in vivo, less is known about their natural function in tissue where opportunities for interaction abound. This makes the casual observer (including the author) very skeptical about ever finding the mediator that is responsible for asthma. Airway Structure in Asthma The histologic appearance of an asthmatic airway (Fig. 1) provides a snapshot of the process that is responsible for the structural changes that occur in the airways of asthmatic patients. As the inflammatory process is dynamic, this appearance serves as a single frame of a long motion picture that must be interpreted in the context of our knowledge of the general features of the inflammatory response. Studies of airway tissue obtained at postmortems. 9, 10, 15, 17, 18, 25, 31, 34, 35 open lung8 and bronchial biopsy32 provide evidence that the structural changes are consistent with the general features of an inflammatory process involving tissue covered with a mucosal surface. Figure 1 shows an airway, from a patient who died of asthma, in which the lumen is filled with an exudate of fluid that contains inflammatory cells, This exudate arises primarily from the bronchial microvasculature and spills over into the airway wall and lumen. The exudate contains small amounts of mucus, plasma protein, and inflammatory cells including eosinophils. The epithelium and subepithelium contain mast cells,s, 10 and there is goblet cell metaplasia,lO basement membrane thickening;';, 25, 35 smooth muscle hypertrophy, and hyperplasia. 15 Furthermore, a recent report2 concerning biopsy and bronchoalveolar lavage 2 of airways of mild asthmatics has confirmed epithelial and submucosal changes that are consistent with a milder form of the inflammatory process that has been clearly documented at postmortem. This study2 showed an inverse correlation between bronchial hyper-reactivity and lavage epithelial cell count and evidence of deposition
734
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Figure 1. A histological section of a bronchiole from a patient who died of asthma is shown. Note that the airway wall is infiltrated by inflammatory cells and that the lumen is filled with exudate containing a small number of these cells. The basement membrane is thickened, the epithelium contains a large number of goblet cells, and there is a marked infiltration of cells surrounding the muscle.
of collagen below the epithelial basement membranes, mast cell degranulation, mucosal infiltration with eosinophils, and migration of eosinophils and monocytes from vessels into the tissue of the asthmatic airways. Since these changes serve as markers of the severity of the inflammatory process, they suggest that airways reactivity increases as the inflammatory response becomes more exuberant. Inflammatory changes are present early in the course of asthma, 2 suggesting that this condition, like many other important disease processes, has a long preclinical phase. The Role of Smooth Muscle in Asthma The popular hypothesis that abnormal behavior of the smooth muscle accounts for the abnormal behavior of the airways has been difficult to prove. Several studies 1, 7, 36 attempting to correlate in vivo airway responsiveness and in vitro smooth muscle sensitivity have found no consistent relationship. This suggests that the excessive airways narrowing that characterizes asthma is a property of intact airways and that abnormal airway smooth muscle shortening may not be required to produce the reversible reduction in airway caliber that characterizes asthma. An alternate hypothesis 3 , 14 is that the airway wall thickening caused by the chronic inflammatory process works in series with normal smooth muscle shortening to produce an exaggerated reduction in airway caliber. An analysis of this hypothesis 26 showed that changes in airway wall thickness that have little
MUCOSAL PERMEABILITI AND SMOOTH MUSCLE FUNCTION IN ASTHMA
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effect on baseline airways resistance can markedly increase airways responsiveness to inhaled agonists. Studies of human lungs 21 resected for tumor, which were inflated with Krebs' solution, bisected in the sagittal plane, and then incubated in Krebs' solution containing either supersaturated theophylline or 1O- 3M carbachol, have demonstrated the effect of muscle relaxation and contraction on airways structure. This showed that contraction of the airways muscle caused the lumen area to decrease and the mucosal surface to fold. Furthermore, a linear relationship established between wall area and the internal perimeter (PI) has provided an excellent method of categorizing airways size. Postmortem studies in which PI and wall area were measured in patients who died of asthma22 showed that the degree of muscle shortening required to produce airway closure was less in the asthmatic patients because of the increased wall area. Quantitative study of the tissue types in these airways showed that the epithelium, vasculature, interstitial tissue, and muscle were all increased in amount. These studies 22 also showed that smooth muscle shortening of 40 per cent resulted in an approximately I5-fold increase in resistance of the airway from the nonasthmatic patients, whereas the same degree of muscle shortening causes approximately a 290-fold increase in resistance in the asthmatic airway. These results support the concept that inflammatory thickening of the airway wall can cause a marked increase in airways resistance with smooth muscle shortening that remains within the normal range. It follows that relaxation of the smooth muscle would also account for a rapid lowering of airways resistance as the smooth muscle lengthens. Both the rapid increase and decrease in airways resistance may occur with changes in smooth muscle lengths that remain within the normal range, suggesting that normally functioning smooth muscle acts in series with a thickened submucosa to reversibly narrow the airways lumen in asthma. Epithelial Changes in Asthma NadeP7 was the first to suggest that airways reactivity was related to the inflammatory injury of the mucosa. Some years ago it was suggested that this might result from an increase in epithelial permeability in the bronchial mucosa that was associated with the inflammatory reaction. 16 The epithelium in the central airways30 is said to be pseudostratified because the cell types of which it is made have a variable purchase on the basement membrane. The outer margins of the cells are sealed by tight junctions that are quite impermeable to large molecules. A sensory nerve net located just below the tight junction surrounds many of the cells forming a tripwire mechanism that is important in the acute response to many irritants. 37 In animal experiments, epithelial permeability can be measured using tracer substances such as horseradish peroxidase 4, 33 or, more recently, fluorescein-Iabeled dextran. 6 Studies of this type have shown that irritants, such as cigarette smoke, cause a marked increase in epithelial permeability that falls back to control values in the 24-hour period following exposure 20 (Fig. 2). In other experiments 19 in which airways reactivity was measured following a similar exposure to cigarette smoke, an increase in airways reactivity was observed (Fig. 3) that also fell back to the control values over
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Hours Post Challenge with 100 Puffs Whole Cigarette Smoke Figure 2. The rate of appearance of the tracer horseradish peroxidase in the blood of guinea pigs at varying intervals after an acute exposure to cigarette smoke is shown. Note that the marked increase in airway permeability V2 hour after exposure fell back to control values 24 hours later. (From Salvato G: Some histological changes in chronic bronchitis and asthma. Thorax 23:168-172, 1968; with permission.)
the same time course following exposure. This temporal association between changes in epithelial permeability and airways reactivity supported the hypothesis that there is a causal relationship between changes in airways mucosal permeability and their responsiveness to inhaled agonists. Unfortunately the relationship between increased airways reactivity and epithelial permeability established in these animal experiments has not been confirmed in human studies. Although several groups23, 24 have
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Figure 3. The increase in airways resistance to inhaled histamine in guinea pigs following cigarette smoke exposure is shown. Note that the greatest increase in resistance occurred '12 hour after the animals were exposed to cigarette smoke and that this increase in resistance fell below the control values 24 hours later. This change parallels the change in airway permeability (see Fig. 2) and suggests a relationship between these two observations. (From Simani IAS, Inoue S, Hogg Je: Penetration of the respiratory epithelium of guinea pigs following exposure to cigarette smoke. Lab Invest 31:75-87, 1974; with permission.)
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measured an increase in ailway permeability to 99Tc DTPA in human smokers, other studies of chronic smokers24 showed that this increase in epithelial permeability was not related to an increase in airways reactivity. Furthermore, when asthmatics with very marked airways hyperreachvity were studied, no increase in airway epithelial permeability could be found. II These human studies show that there is no simple relationship between airways permeability and reactivity in patients with asthma. They also suggest that the increase in permeability seen in animals after acute exp~sure to cigarette smoke may be different than that seen in chronic human smokers. Recent studies 6 of guinea pigs exposed to cigarette smoke have shown that the major site of the increased airways permeability is in the respiratory bronchioles. This may also be the leakage site in human smokers because it is well established that cigarette smoking results in a respiratory bronchiolitis. 29 However, the permeability change responsible for the increase in airways reactivity in the animal studies may be more centrally located and may only develop after the heavy acute exposures these animals received. A comparable human situation might be the smoke exposures that occur in fires and industrial accidents rather than the more modest exposures associated with cigarette smoking. Unfortunately, there have been no studies of airways reactivity and permeability before and after such events that would provide the data needed to test such a hypothesis. In summary, there is ample evidence that the airways of patients suffering from asthma contain an inflammatory response that is consistent with that seen in tissue covered by a mucosal surface. This response can be demonstrated by both autopsy and biopsy studies and is present even in mild asthmatic patients. Although this inflammatory process is associated with abnormalities in the epithelium and hypertrophy of the airway muscle, neither of these provides a clear explanation for the observed changes in airways function. These latter changes are most readily explained by normal smooth muscle contraction and shortening acting in series with a thickened submucosa and luminal exudate to narrow the airway caliber.
REFERENCES 1. Armour CL, Lazar NM, Schellenberg RR, et al: A comparison of in vivo and in vitro human airway reactivity to histamine. Am Rev Respir Dis 129:907-910, 1984 2. Beasley R, Roche WR, Roberts JA, et al: Cellular events in the bronchi in mild asthma and after bronchial provocation. Am Rev Respir Dis 139:806-817, 1989 3. Benson MK: Bronchial hyperreactivity. Br J Dis Chest 69:227-239, 1975 4. Boucher HC, Johnson J, Inoue S, et al: The effect of cigarette smoke on the permeability of guinea pig airways. Lab Invest 43:94-100, 1980 5. Bullen SS: Correlation of clinical and autopsy findings in 176 cases of asthma. J Allerg Clin Immunol 23:193-203, 1952 6. Burns AH, Hosford SP, Dunn LA, et al: Hespiratory epithelial permeability after cigarette smoke exposure in guinea pigs. J Appl Physiol 66:2109-2116, 1989 7. Cerrina J, Ladurie ML, Labat C, et al: Comparison of human bronchial muscle responses to histamine in vivo with histamine and isoproteronol in vitro. Am Hev Hespir Dis 134:51-61, 1986 8. Cudz E, Levison H, Cooper DM: Ultrastructure of airways in children with asthma. Histopathology 2:407-421, 1978
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9. DUBBili ~.,.IS, Massarella GR, Anderson JA: A comparison of the quantitative anatomy of the bronchi in normal subjects, in status asthmaticus, in chronic bronchitis and in emphysema. Thorax 24: 176-179, 1969 10. Dunnill MS: The pathology of asthma with special reference to changes in the bronchial mucosa. J Clin Path 13:27-33, 1960 11. Elwood HK, Kennedy SM, Belzberg A, et al: Respiratory mucosal permeability in asthma. Am Rev Respir Dis 128:523-527, 1983 12. Florey HvV: Secretion of mucus and inflammation of mucus membranes. In General Pathology. London, Lloyd Luke (Medical Books) Ltd, 1962, pp 167-197 13. Florey HW: Inflammation. In General Pathology. London, Lloyd Luke (Medical Books) Ltd, 1962, pp 1-167 14. Freedman BJ: The functional geometry of the bronchi. Bull Physiopathol Resp 8:545551, 1972 15. Heard BE, Hossain S: Hyperplasia of bronchial muscle in asthma. J Pathol 110:319-331, 1971 16. Hogg JC: Bronchial mucosal permeability and its relationship to airways hyperreactivity. J Allergy Clin Immunol 67:421-425, 1981 17. Houston JC, De Vavasquez S, Trounce JR: A clinical and pathological study offatal cases of status asthmaticus. Thorax 8:207-213, 1953 18. Huber HL, Koessler KK: The pathology of bronchial asthma. Arch Int Med 30:689-760, 1922 19. Hulbert WC, McLean T, Hogg JC: The effect of acute airway inflammation on bronchial reactivity in guinea pigs. Am Rev Respir Dis 132:7-11, 1985 20. Hulbert WC, Walker DC, Jackson A, et al: Airway permeability to horseradish peroxidase in guinea pigs: The repair phase after injury by cigarette smoke. Am Rev Respir Dis 123:320-326, 1981 21. James AL, Hogg JC, Dunn LA, et al: The use of internal perimeter to compare airway size and calculate smooth muscle shortening. Am Rev Respir Dis 138: 136-139, 1988 22. James AL, Pare PD, Hogg JC: Mechanisms of airway narrowing in asthma. Am Hev Respir Dis 139:242-246, 1989 23. Jones JG, Lawler P, Crawley JCW, et al: Increased alveolar epithelial permeability in cigarette smokers. Lancet 1:66-68, 1980 24. Kennedy SM, Elwood RK, Wiggs BJ, et al: Increased airway mucosal permeability of smokers: Relationship to airways reactivity. Am Rev Respir Dis 129:143-148, 1984 25. Messer l, Peters GA, Bennet W A: Cause of death and pathological findings in 304 cases of bronchial asthma. Dis Chest 38:616-624, 1960 26. Moreno RH, Hogg lC, Pare PD: Mechanisms of airway narrowing. Am Rev Respir Dis 133: 1171-1180, 1986 27. Nadel lA: Structure and function relationship in the airways. Medicina Thoracalis 22:231, 1965 28. Naylor B: The shedding of the mucosa of the bronchial tree in asthma. Thorax 17:69-72, 1962 29. Neiwoehner DE, Kleinerman l, Rice DB: Pathologic changes in the peripheral airways of young cigarette smokers. N Engl 1 Med 291:755-758, 1974 30. Reid L, lones R: Bronchial mucosal cells. Fed Proc 38:191, 1979 31. Richards W, Patrick JR: Death fi'om asthma in children. Am J Dis Child 110:4-21, 1965 32. Salvato G: Some histological changes in chronic bronchitis and asthma. Thorax 23:168172, 1968 33. Simani IAS, Inoue S, Hogg lC: Penetration of the respiratory epithelium of guinea pigs following exposure to cigarette smoke. Lab Invest 31:75-87, 1974 :34. Takizawa T, Thurlbeck WM: Muscle and mucous gland size in the major bronchi of patients with chronic bronchitis, asthma and asthmatic bronchitis. Am Rev Hespir Dis 104:331-336, 1971 35. Unger L: Pathology of bronchial asthma. South Med 1 38:513-522, 1945 36. Vincenc KS, Black lL, Yan K, et al: A comparison of in vivo and in vitro responses to histamine in human airway. Am Rev Respir Dis 128:875-879, 1983
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37. Widdicombe JG: Reflex control of tracheobronchial smooth muscle in experimental and human asthma. In Austen KF, Lichtenstein (eds): AsthmaII: Pathology, immunopharmacology and treatment. Second International Symposium. New York, Academic Press, 1977, pp 225-231
Address reprint requests to James G. Hogg, MD, PhD, FRCP(C) UBC Pulmonary Research Laboratory st. Paul's Hospital 1081 Burrard Street Vancouver, British Columbia V6Z 1Y6 Canada
0025-7125/90 $0.00
+
.20
Mucosal Permeability and Smooth Muscle Function in Asthma James C. Hogg, MD, PhD, FRCP(C)*
The changes in airway structure that occur in asthma are characteristic of a chronic inflammatory process that involves tissue covered by a mucosal surface. 12. 13 This process produces changes in the airway epithelium and in the smooth muscle surrounding the airways that could contribute to abnormal airways function in asthma. In this article will be a brief review of the inflammatory process and a presentation of evidence that the structural changes seen in asthmatic airways are a result of this response. The contribution of the airway muscle and epithelium to the abnormal function of asthmatic airways will then be reviewed.
THE STRUCTURAL FEATURES OF THE INFLAMMATORY RESPONSE Following tissue injury, there is a rapid series of changes in the microvessels that result in vascular congestion and the escape of plasma into the tissue. 13 The balance between exudation from the vessels and fluid drainage into the lymphatic vessels determines the degree of tissue swelling. The exudation of fluid from the vascular space is followed by the appearance of platelets, polymorphonuclear leukocytes, monocytes, and lymphocytes in the inflammatory site. This greatly increases cell traffic in the damaged tissue and sets off a number of events, which occur simultaneously, but must be described separately. These events determine the structural features of the inflammatory response in all tissues but certain tissuespecific changes occur in some organs. For example, airways, like other structures covered with a mucosal surface, show two important tissuespecific features of the inflammatory response. 12 The first of these is a shedding of the epithelium, which can sometimes occur in large clumps. In asthma, these clumps are referred to as "Creola bodies" after the patient *Professor of Pathology, University of British Columbia; and Director, Pulmonary Research Laboratory, St. Paul's Hospital, Vancouver, British Columbia, Canada
Medical Clinics of North America-VD!' 74, No. 3, May 1990
731
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C. Haec
(Creola Jones) in whom they were first described. 2H The second is an excess secretion of mucus from both epithelial lining cells and submucosal glands. The combination of the exudation of fluid and cells from microvessels, shed epithelium, and mucus hypersecretion often fills the airways with tenacious material referred to as a mucus plug. In fact, the majority of the material occluding the airway is an exudate of plasma and cells that contains a very small amount of mucus. In the repair phase of the inflammatory response, epithelial cells multiply and cover the denuded surface. The rapid turnover of the epithelium probably results in the characteristic thickening of the basement membrane in this condition. Similarly, the proliferation of the microvessels, connective tissue, and muscle results in a thickening of the submucosa. Mediators of the Inflammatory Response The mediators of the various events that characterize the inflammatory response are chemical in nature. These chemicals are derived from the cells that are resident in the tissue, from the plasma exudate, and from the cells that migrate into the tissue in response to injury. The account below provides a very brief overview of the complex subject and is designed to outline a basis for this discussion of mucosal and smooth muscle function in asthma. Tissue-derived Mediators. An important group of mediators of the inflammatory reaction are derived from peroxidation of arachidonic acid, which is a major constituent of lipid in cell membranes. Arachidonic acid can be metabolized in two ways. The cyclo-oxygenase pathway involves two unstable endoperoxide intermediates that are metabolized to stable prostaglandins, such as prostaglandin E z, a potent vasoconstrictor and platelet activator thromboxane A2 , a dilator and antiaggregatory compound, prostaglandin 12 , On the other hand, the lipoxygenase pathway of arachidonic acid peroxidation produces a family of compounds known as leukotrienes. One of these compounds (leukotriene C) is now known to be identical with a slow-reacting substance of anaphylaxis, which has been implicated as a possible mediator of the prolonged smooth muscle contraction seen in asthma. The best known tissue-derived mediator is histamine, which is formed by decarboxylation of histidine in the granules of mast cells. These cells are found in high concentration around the airways particularly in the distal parts of the lung. Because these distal airways are muscular, histamine is an important candidate as a mediator of airways narrowing. A third group of tissue-derived mediators (the tachykinins) are produced by antidromic stimulation of sensory nerves and are thought by some to be important in some forms of bronchial hyperreactivity. Plasma-derived Mediators. Hageman factor is activated when plasma comes in contact with a foreign surface during the exudative process. Activated Hageman factor converts several plasma proenzymes to their active form. This in turn results in activation of the complement, coagulation, kinin, and fibrinolytic cascades, which generate a host of inflammatory mediators. Mediators derived from the complement cascade can be generated through the classical pathway that is triggered by antigen antibody complexes or by the alternate pathway by endotoxin and by plasma proteins
MUCOSAL PER:vJEABILlTY AN]) S!\100TH .MUSCLE FUNCTION I!\I ASTH~jA
733
and proteolytic enzymes contained in the plasma exudate. In either event, this cascade releases potent mediators of the inflammatory response into the tissues. Mediators Derived from Migrating Cells. The cell population migrating into an acutely inflamed area contributes a variety of inflammatory mediators. The neutrophil can release cationic proteins as well as acid and neutral proteases from its granules. Activation of the nicotinamide-adenine-dinucleotide phosphate (NADPH) oxidase in the neutrophil membrane generates a variety of toxic oxygen species including the superoxide anion, hydroxyl radical, and hydrogen peroxide. The interaction of H 20 2 with halide ions and myeloperoxidase from the neutrophil granules produces a variety of short- and long-acting oxidants that can destroy invading bacteria. Monocytes migrating into inflamed tissue transform to macrophages that are capable of processing antigen and generating many important cytokines capable of stimulating the proliferation of T- and B-lymphocytes. The cytokines produced by monocytes and lymphocytes affect antibody production as well as the function of neutrophils, eosinophils, monocytes, and lymphocytes in the inflammatory response. The major problem with understanding the function of individual mediators is that many of them function simultaneously and produce interactive results. Although individual mediators can be isolated and studied in a controlled fashion both in vitro and in vivo, less is known about their natural function in tissue where opportunities for interaction abound. This makes the casual observer (including the author) very skeptical about ever finding the mediator that is responsible for asthma. Airway Structure in Asthma The histologic appearance of an asthmatic airway (Fig. 1) provides a snapshot of the process that is responsible for the structural changes that occur in the airways of asthmatic patients. As the inflammatory process is dynamic, this appearance serves as a single frame of a long motion picture that must be interpreted in the context of our knowledge of the general features of the inflammatory response. Studies of airway tissue obtained at postmortems. 9, 10, 15, 17, 18, 25, 31, 34, 35 open lung8 and bronchial biopsy32 provide evidence that the structural changes are consistent with the general features of an inflammatory process involving tissue covered with a mucosal surface. Figure 1 shows an airway, from a patient who died of asthma, in which the lumen is filled with an exudate of fluid that contains inflammatory cells, This exudate arises primarily from the bronchial microvasculature and spills over into the airway wall and lumen. The exudate contains small amounts of mucus, plasma protein, and inflammatory cells including eosinophils. The epithelium and subepithelium contain mast cells,s, 10 and there is goblet cell metaplasia,lO basement membrane thickening;';, 25, 35 smooth muscle hypertrophy, and hyperplasia. 15 Furthermore, a recent report2 concerning biopsy and bronchoalveolar lavage 2 of airways of mild asthmatics has confirmed epithelial and submucosal changes that are consistent with a milder form of the inflammatory process that has been clearly documented at postmortem. This study2 showed an inverse correlation between bronchial hyper-reactivity and lavage epithelial cell count and evidence of deposition
734
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HOGG
Figure 1. A histological section of a bronchiole from a patient who died of asthma is shown. Note that the airway wall is infiltrated by inflammatory cells and that the lumen is filled with exudate containing a small number of these cells. The basement membrane is thickened, the epithelium contains a large number of goblet cells, and there is a marked infiltration of cells surrounding the muscle.
of collagen below the epithelial basement membranes, mast cell degranulation, mucosal infiltration with eosinophils, and migration of eosinophils and monocytes from vessels into the tissue of the asthmatic airways. Since these changes serve as markers of the severity of the inflammatory process, they suggest that airways reactivity increases as the inflammatory response becomes more exuberant. Inflammatory changes are present early in the course of asthma, 2 suggesting that this condition, like many other important disease processes, has a long preclinical phase. The Role of Smooth Muscle in Asthma The popular hypothesis that abnormal behavior of the smooth muscle accounts for the abnormal behavior of the airways has been difficult to prove. Several studies 1, 7, 36 attempting to correlate in vivo airway responsiveness and in vitro smooth muscle sensitivity have found no consistent relationship. This suggests that the excessive airways narrowing that characterizes asthma is a property of intact airways and that abnormal airway smooth muscle shortening may not be required to produce the reversible reduction in airway caliber that characterizes asthma. An alternate hypothesis 3 , 14 is that the airway wall thickening caused by the chronic inflammatory process works in series with normal smooth muscle shortening to produce an exaggerated reduction in airway caliber. An analysis of this hypothesis 26 showed that changes in airway wall thickness that have little
MUCOSAL PERMEABILITI AND SMOOTH MUSCLE FUNCTION IN ASTHMA
735
effect on baseline airways resistance can markedly increase airways responsiveness to inhaled agonists. Studies of human lungs 21 resected for tumor, which were inflated with Krebs' solution, bisected in the sagittal plane, and then incubated in Krebs' solution containing either supersaturated theophylline or 1O- 3M carbachol, have demonstrated the effect of muscle relaxation and contraction on airways structure. This showed that contraction of the airways muscle caused the lumen area to decrease and the mucosal surface to fold. Furthermore, a linear relationship established between wall area and the internal perimeter (PI) has provided an excellent method of categorizing airways size. Postmortem studies in which PI and wall area were measured in patients who died of asthma22 showed that the degree of muscle shortening required to produce airway closure was less in the asthmatic patients because of the increased wall area. Quantitative study of the tissue types in these airways showed that the epithelium, vasculature, interstitial tissue, and muscle were all increased in amount. These studies 22 also showed that smooth muscle shortening of 40 per cent resulted in an approximately I5-fold increase in resistance of the airway from the nonasthmatic patients, whereas the same degree of muscle shortening causes approximately a 290-fold increase in resistance in the asthmatic airway. These results support the concept that inflammatory thickening of the airway wall can cause a marked increase in airways resistance with smooth muscle shortening that remains within the normal range. It follows that relaxation of the smooth muscle would also account for a rapid lowering of airways resistance as the smooth muscle lengthens. Both the rapid increase and decrease in airways resistance may occur with changes in smooth muscle lengths that remain within the normal range, suggesting that normally functioning smooth muscle acts in series with a thickened submucosa to reversibly narrow the airways lumen in asthma. Epithelial Changes in Asthma NadeP7 was the first to suggest that airways reactivity was related to the inflammatory injury of the mucosa. Some years ago it was suggested that this might result from an increase in epithelial permeability in the bronchial mucosa that was associated with the inflammatory reaction. 16 The epithelium in the central airways30 is said to be pseudostratified because the cell types of which it is made have a variable purchase on the basement membrane. The outer margins of the cells are sealed by tight junctions that are quite impermeable to large molecules. A sensory nerve net located just below the tight junction surrounds many of the cells forming a tripwire mechanism that is important in the acute response to many irritants. 37 In animal experiments, epithelial permeability can be measured using tracer substances such as horseradish peroxidase 4, 33 or, more recently, fluorescein-Iabeled dextran. 6 Studies of this type have shown that irritants, such as cigarette smoke, cause a marked increase in epithelial permeability that falls back to control values in the 24-hour period following exposure 20 (Fig. 2). In other experiments 19 in which airways reactivity was measured following a similar exposure to cigarette smoke, an increase in airways reactivity was observed (Fig. 3) that also fell back to the control values over
736
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the same time course following exposure. This temporal association between changes in epithelial permeability and airways reactivity supported the hypothesis that there is a causal relationship between changes in airways mucosal permeability and their responsiveness to inhaled agonists. Unfortunately the relationship between increased airways reactivity and epithelial permeability established in these animal experiments has not been confirmed in human studies. Although several groups23, 24 have
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Figure 3. The increase in airways resistance to inhaled histamine in guinea pigs following cigarette smoke exposure is shown. Note that the greatest increase in resistance occurred '12 hour after the animals were exposed to cigarette smoke and that this increase in resistance fell below the control values 24 hours later. This change parallels the change in airway permeability (see Fig. 2) and suggests a relationship between these two observations. (From Simani IAS, Inoue S, Hogg Je: Penetration of the respiratory epithelium of guinea pigs following exposure to cigarette smoke. Lab Invest 31:75-87, 1974; with permission.)
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measured an increase in ailway permeability to 99Tc DTPA in human smokers, other studies of chronic smokers24 showed that this increase in epithelial permeability was not related to an increase in airways reactivity. Furthermore, when asthmatics with very marked airways hyperreachvity were studied, no increase in airway epithelial permeability could be found. II These human studies show that there is no simple relationship between airways permeability and reactivity in patients with asthma. They also suggest that the increase in permeability seen in animals after acute exp~sure to cigarette smoke may be different than that seen in chronic human smokers. Recent studies 6 of guinea pigs exposed to cigarette smoke have shown that the major site of the increased airways permeability is in the respiratory bronchioles. This may also be the leakage site in human smokers because it is well established that cigarette smoking results in a respiratory bronchiolitis. 29 However, the permeability change responsible for the increase in airways reactivity in the animal studies may be more centrally located and may only develop after the heavy acute exposures these animals received. A comparable human situation might be the smoke exposures that occur in fires and industrial accidents rather than the more modest exposures associated with cigarette smoking. Unfortunately, there have been no studies of airways reactivity and permeability before and after such events that would provide the data needed to test such a hypothesis. In summary, there is ample evidence that the airways of patients suffering from asthma contain an inflammatory response that is consistent with that seen in tissue covered by a mucosal surface. This response can be demonstrated by both autopsy and biopsy studies and is present even in mild asthmatic patients. Although this inflammatory process is associated with abnormalities in the epithelium and hypertrophy of the airway muscle, neither of these provides a clear explanation for the observed changes in airways function. These latter changes are most readily explained by normal smooth muscle contraction and shortening acting in series with a thickened submucosa and luminal exudate to narrow the airway caliber.
REFERENCES 1. Armour CL, Lazar NM, Schellenberg RR, et al: A comparison of in vivo and in vitro human airway reactivity to histamine. Am Rev Respir Dis 129:907-910, 1984 2. Beasley R, Roche WR, Roberts JA, et al: Cellular events in the bronchi in mild asthma and after bronchial provocation. Am Rev Respir Dis 139:806-817, 1989 3. Benson MK: Bronchial hyperreactivity. Br J Dis Chest 69:227-239, 1975 4. Boucher HC, Johnson J, Inoue S, et al: The effect of cigarette smoke on the permeability of guinea pig airways. Lab Invest 43:94-100, 1980 5. Bullen SS: Correlation of clinical and autopsy findings in 176 cases of asthma. J Allerg Clin Immunol 23:193-203, 1952 6. Burns AH, Hosford SP, Dunn LA, et al: Hespiratory epithelial permeability after cigarette smoke exposure in guinea pigs. J Appl Physiol 66:2109-2116, 1989 7. Cerrina J, Ladurie ML, Labat C, et al: Comparison of human bronchial muscle responses to histamine in vivo with histamine and isoproteronol in vitro. Am Hev Hespir Dis 134:51-61, 1986 8. Cudz E, Levison H, Cooper DM: Ultrastructure of airways in children with asthma. Histopathology 2:407-421, 1978
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9. DUBBili ~.,.IS, Massarella GR, Anderson JA: A comparison of the quantitative anatomy of the bronchi in normal subjects, in status asthmaticus, in chronic bronchitis and in emphysema. Thorax 24: 176-179, 1969 10. Dunnill MS: The pathology of asthma with special reference to changes in the bronchial mucosa. J Clin Path 13:27-33, 1960 11. Elwood HK, Kennedy SM, Belzberg A, et al: Respiratory mucosal permeability in asthma. Am Rev Respir Dis 128:523-527, 1983 12. Florey HvV: Secretion of mucus and inflammation of mucus membranes. In General Pathology. London, Lloyd Luke (Medical Books) Ltd, 1962, pp 167-197 13. Florey HW: Inflammation. In General Pathology. London, Lloyd Luke (Medical Books) Ltd, 1962, pp 1-167 14. Freedman BJ: The functional geometry of the bronchi. Bull Physiopathol Resp 8:545551, 1972 15. Heard BE, Hossain S: Hyperplasia of bronchial muscle in asthma. J Pathol 110:319-331, 1971 16. Hogg JC: Bronchial mucosal permeability and its relationship to airways hyperreactivity. J Allergy Clin Immunol 67:421-425, 1981 17. Houston JC, De Vavasquez S, Trounce JR: A clinical and pathological study offatal cases of status asthmaticus. Thorax 8:207-213, 1953 18. Huber HL, Koessler KK: The pathology of bronchial asthma. Arch Int Med 30:689-760, 1922 19. Hulbert WC, McLean T, Hogg JC: The effect of acute airway inflammation on bronchial reactivity in guinea pigs. Am Rev Respir Dis 132:7-11, 1985 20. Hulbert WC, Walker DC, Jackson A, et al: Airway permeability to horseradish peroxidase in guinea pigs: The repair phase after injury by cigarette smoke. Am Rev Respir Dis 123:320-326, 1981 21. James AL, Hogg JC, Dunn LA, et al: The use of internal perimeter to compare airway size and calculate smooth muscle shortening. Am Rev Respir Dis 138: 136-139, 1988 22. James AL, Pare PD, Hogg JC: Mechanisms of airway narrowing in asthma. Am Hev Respir Dis 139:242-246, 1989 23. Jones JG, Lawler P, Crawley JCW, et al: Increased alveolar epithelial permeability in cigarette smokers. Lancet 1:66-68, 1980 24. Kennedy SM, Elwood RK, Wiggs BJ, et al: Increased airway mucosal permeability of smokers: Relationship to airways reactivity. Am Rev Respir Dis 129:143-148, 1984 25. Messer l, Peters GA, Bennet W A: Cause of death and pathological findings in 304 cases of bronchial asthma. Dis Chest 38:616-624, 1960 26. Moreno RH, Hogg lC, Pare PD: Mechanisms of airway narrowing. Am Rev Respir Dis 133: 1171-1180, 1986 27. Nadel lA: Structure and function relationship in the airways. Medicina Thoracalis 22:231, 1965 28. Naylor B: The shedding of the mucosa of the bronchial tree in asthma. Thorax 17:69-72, 1962 29. Neiwoehner DE, Kleinerman l, Rice DB: Pathologic changes in the peripheral airways of young cigarette smokers. N Engl 1 Med 291:755-758, 1974 30. Reid L, lones R: Bronchial mucosal cells. Fed Proc 38:191, 1979 31. Richards W, Patrick JR: Death fi'om asthma in children. Am J Dis Child 110:4-21, 1965 32. Salvato G: Some histological changes in chronic bronchitis and asthma. Thorax 23:168172, 1968 33. Simani IAS, Inoue S, Hogg lC: Penetration of the respiratory epithelium of guinea pigs following exposure to cigarette smoke. Lab Invest 31:75-87, 1974 :34. Takizawa T, Thurlbeck WM: Muscle and mucous gland size in the major bronchi of patients with chronic bronchitis, asthma and asthmatic bronchitis. Am Rev Hespir Dis 104:331-336, 1971 35. Unger L: Pathology of bronchial asthma. South Med 1 38:513-522, 1945 36. Vincenc KS, Black lL, Yan K, et al: A comparison of in vivo and in vitro responses to histamine in human airway. Am Rev Respir Dis 128:875-879, 1983
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37. Widdicombe JG: Reflex control of tracheobronchial smooth muscle in experimental and human asthma. In Austen KF, Lichtenstein (eds): AsthmaII: Pathology, immunopharmacology and treatment. Second International Symposium. New York, Academic Press, 1977, pp 225-231
Address reprint requests to James G. Hogg, MD, PhD, FRCP(C) UBC Pulmonary Research Laboratory st. Paul's Hospital 1081 Burrard Street Vancouver, British Columbia V6Z 1Y6 Canada