Lung (1990) Suppl:1050-1056
Lung
© Springer-¥erlag New York Inc. 1990
Future Directions for Bronchoalveolar Lavage Stephen I. Rennard Pulmonary and Critical Care Medicine Section, Department of Internal Medicine, University of Nebraska Medical Center, Omaha, Nebraska, USA
Abstract. Bronchoalveolar lavage provides a means to sample the lung. The ability to describe material obtained from the lower respiratory tract has greatly facilitated pulmonary diagnosis, and it is likely that improvements in diagnostic techniques will continue to develop. The ability to accurately measure lower respiratory tract components has permitted staging of lung disease. Such staging will probably provide improved prognostic insight and permit the development of new therapeutic strategies in lung disease. Key words: Bronchoalveolar lavagemStaging of lung disease--Viable cells--Experimental intervention. Introduction Predicting future directions, particularly for a technique used both clinically and for research, is a difficult task. Bronchoalveolar lavage (BAL) has a long history dating back over 100 years [1-3]. It has been in widespread use, however, only since the introduction of the flexible fiberoptic bronchoscope 20 years ago [4]. Through this instrument lavage is easily performed, and it has rapidly made the lower respiratory tract accessible to investigators [5-7]. As a result, BAL has changed the ability of physicians and researchers to approach pulmonary disease. In particular, BAL has made the lung an accessible organ. By virtue of the fact that the procedure can be performed safely and easily and that it is well tolerated, a vast array of material has been made available for diagnostic and research purposes. The procedure of bronchoalveolar lavage is generally performed with the bronchoscope in a wedged position [8-10]. Sterile saline is infused through the Offprint requests to: Dr. S. I. Rennard, Pulmonary and Critical Care Medicine Section, Department of Internal Medicine, University of Nebraska Medical Center, 600 So. 42nd, Omaha, NE 68198-2465, USA.
Future Directions for B A L
1051
biopsy port of the bronchoscope, and this infused saline mixes with the fluid lining the airways and alveolar spaces of the lung. This fluid can then be recovered by aspiration through the biopsy channel of the bronchoscope. The resulting lavage fluid contains some of the material previously present within the lumen of the airways and alveoli and thus represents a direct sampling of the lung. This sampling presents the pulmonologist with several distinctly different opportunities including: 1) description of the material present within the lower respiratory tract and hence diagnosis; 2) quantification of various components present within the lower respiratory tract and hence staging; 3) recovery of viable cells from the lower respiratory tract and hence insight into normal and abnormal cellular physiology; and 4) specific performance of experiments in the lower respiratory tract.
Description and Diagnosis While bronchoalveolar lavage primarily samples the luminal contents, many abnormal processes within the lung are characterized by abnormalities within the air spaces. The material obtained by lavage can, therefore, be of diagnostic use. Pneumocystis carinii pneumonia, for example, is characterized by an exuberant alveolar infiltrate [11]. When stained with an appropriate technique, material covered by lavage can readily yield a diagnosis of pneumocystis carinii. This procedure has almost completely replaced the need for open lung biopsy in the diagnosis of pneumocystis pneumonia. Other pathologic processes can also be present within the airway. For example, cancer infiltrating air spaces or present on the airway surfaces can also be recovered by lavage [9]. Special techniques, for example the use of monoclonal antibodies specific for tumor markers, can greatly aid in the diagnosis of cancer on BAL cytology by increasing the diagnostic yield [12]; it can also result in the diagnosis of infectious processes [9, 13-15], and cytopathologic changes characteristic of viral infections can be readily recognized as well [9, 16-18]. Again, the application of new biological techniques such as the use of cDNA probes for viral specific DNA can greatly increase the sensitivity of diagnostic methods for specific pathogens [19, 20]. It is likely that the appfication of monoclonal antibodies and cDNA probes will increase the power of BAL as a diagnostic tool in the future. This is likely to have the biggest impact in the diagnosis of malignancies and in viral infections.
Measurement and Staging Measurement of lower respiratory tract components permits the "staging" of lower respiratory tract disease. Staging can be very useful in assessing relative severity of a condition and can, therefore, lead to important prognostic and therapeutic decisions. In addition, by being able to evaluate the severity of a disease process, the ability to quantify lower respiratory tract components can permit the assessment of therapeutic interventions.
1052
S . I . Rennard
Sarcoidosis, for example, has been associated with lower respiratory tract lymphocytosis [21, 22]. In particular, T helper cells are found to be increased in both number and percentage [23-25]. This increase can be quantified and therefore permit staging. In general, patients with higher numbers and percentages of lymphocytes are thought to have a more "active" disease which is more likely to progress [26, 27], although a number of controversies persist [28]. Similarly, therapeutic intervention, for example with glucocorticoids, is often associated with a reduction in the magnitude of the lymphocytosis [29]. In a similar fashion, it is possible to assess a great variety of lower respiratory tract components [8-10, 30]. Cigarette smoking is thought to lead to "small airways disease," a condition characterized by accumulation of macrophages, neutrophils and their secretory products in the small airways of the lung [31-33]. The release of secretory products, particularly neutrophil elastase, is thought to lead to destruction of lung tissue and subsequently to the development of pulmonary emphysema [34, 35]. By virtue of its ability to sample the air spaces of the lung, BAL can quantify these potential mediators of disease. It is, for example, possible to measure lower respiratory tract neutrophil elastase as a complex to alpha-1 antitrypsin [36, 37]. A therapeutic intervention, like cigarette smoke reduction, can then be assessed in terms of its ability to reduce the lung's burden of this potentially damaging protease [38]. The use of bronchoalveolar lavage to stage various lung diseases is clearly an area of active investigation. The future will undoubtedly lead to great insights regarding the quantitative nature of lower respiratory tract cells and components with respect to pathophysiologic mechanisms, disease prognosis and therapeutic intervention. While the use of these staging techniques is currently not fully established, it is almost certain that there will be increasingly wide applications in the future.
Function and Pathophysioiogy Bronchoalveolar lavage is capable of recovering viable cells from the lower respiratory tract [8, 9, 39]. These viable cells can be maintained in culture ex vivo, and their functional state can be assessed. A large spectrum of assay systems can be applied to these cells recovered from the lower respiratory tract. These include biological assays, biochemical assays, immunoassays and studies of gene regulation. These studies have permitted the description of release of growth factors, chemotactic factors and cytotoxic factors from lower respiratory tract cells. In addition, it has been possible to characterize and quantify the release of specific mediators in individual disease states. Finally, it has been possible to determine the state of gene activation in cells recovered from the lower respiratory tract. Taken together, these types of studies have greatly helped explain the interactions between the cells in the lower respiratory tract involved in normal health and in disease states. Specific mechanisms of lung injury and repair have been delineated. It is almost certain that these cytokine networks and the various cellular interactions which they mediate will be described in increasing detail in the near future.
Future Directions for BAL
1053
Experimentation and Manipulation Bronchoalveolar lavage not only samples the lung, it creates the possibility of introducing material into the lung. This permits experimental manipulations of the lower respiratory tract. It is possible, for example, to introduce antigens into the sensitive asthmatic directly through the bronchoscope. Using these techniques, Casale et al. and Fick et al. have demonstrated that local application of antigen can lead to the release of histamine from mast cells and to the leakage of albumin from the intravascular spaces into the airway lumen [40, 41]. It is likely that the ability of BAL to introduce materials into the lung will be greatly exploited by investigators in the near future. BAL also opens the horizons for "extracorporeal surgery" of lung cells. Extracorporeal surgery is now routinely performed on kidneys [42]. A kidney can be removed from a patient under general anesthesia, be microdissected removing vascular lesions or small tumor implants, be repaired and then restored to the patient. Similar "surgery" could be performed on lung cells using BAL instead of nephrectomy. For example, BAL cells can be removed from the body, subjected to "repair" mechanisms, and then be reinfused into the lungs. Examples of currently available methods to alter cellular functions in cells obtained from the lung include: 1) specific expansion of subpopulations of lung cells using techniques of cell culture [43]; 2) selection of specific types of lung cells using monoclonal antibodies [44]; 3) activation of lung cells using specific monokines or cytokines in vitro [45, 46]; and 4) introduction of novel genes into the cells obtained from the lung [47, 48]. Examples of the potential for such surgery in lung disease include: 1) the activation of immune and inflammatory effector cells in diseased states where such cells are ineffective, e.g., infections and malignancy; 2) in diseases where lung cells are characterized by inappropriate activation, such as immune and inflammatory lung disease or asthma, it may be possible to introduce specific suppressor cells which could, when reintroduced in vivo, result in a suppression of the abnormal immune and inflammatory responses; 3) it may be possible to introduce a normal gene, and therefore restore to normal function, in an individual with a genetic abnormality, e.g., in alpha-1 protease inhibitor deficiency or in cystic fibrosis; 4) it may be possible to utilize the ability of BAL cells to mediate lung repair processes to control the regulation of such processes. As such, it may be possible to restore function in lungs which have been disrupted by pulmonary fibrosis or by widespread emphysema. Clearly, such applications are beyond current capabilities. Nevertheless, currently available technology suggests that they are within the realm of imagination and represent reasonable long-term future directions.
Summary The ability of lavage to recover viable cells has permitted great insights into the pathophysiologic mechanisms of disease and the normal physiologic mechanisms responsible for lower respiratory tract cellular function. It is likely that
1054
S.I. Rennard
these studies will continue to advance our understanding of the interactions between cells and the mediators which make such interactions possible. Bronchoalveolar lavage also makes the lung accessible to experimental intervention. Such experimental intervention will not only greatly aid in the understanding of physiologic mechanisms of disease states but also raises the possibility that specific therapeutic interventions can be targeted directly to the lung. The rapid advances in biological techniques including cell culture, biochemical analysis, the application of monoclonal antibodies and the application of molecular biological techniques represents a "new biology." The ability of bronchoalveolar lavage to make the lung an accessible organ ensures that this new biology can be readily applied to the lung.
References 1. Bernard J, Gee L, Fick RB (1980) Bronchoalveolar lavage (editorial). Thorax 35:1-8 2. Rogers RM, Braunstein MS, Shuman JR (1972) Role of bronchopulmonary lavage in the treatment of respiratory failure: a review. Chest 62(Suppl):95-106 3. Ramirez-R J, Kieffer RF, Ball WC (1965) Bronchopulmonary lavage in man. Ann Intern Med 63:819-828 4. lkeda S, Yanai N, Ishikawa S (1968) Flexible bronchofiberscope. Keio J Med 17:1-16 5. CantreUET, Warr GA, Busbee DL, et al (1973) Induction of arylhydrocarbon hydroxylase in human pulmonary alveolar macrophages by cigarette smoking. J Clin Invest 52:1881-1884 6. Waldmann RH, Jurgensen PF, Olen GN, et al (1973) Immune response of the human respiratory tract: immunoglobulin levels and influenza virus vaccine antibody response. J Immunol 111:38--41 7. Reynolds HY, Newball HH (1974) Analysis of proteins and respiratory cells obtained from human lungs by bronchial lavage. J Lab Clin Med 84:559-573 8. Reynolds HY (1987) State of the art: bronchoalveolar lavage. Am Rev Respir Dis 135:250263 9. Linder J, Rennard S Bronchoalveolar lavage. Chicago: ASCP Press, pp 1-196 10. Costabel U, Danel C, Haslam P, Higgenbottam T, Klech H, Pohl W, Rennard S, Rossi G, Rust M, Semenzato Get al (1989) Technical recommendations and guidelines for bronchoalveolar lavage (BAL). Report of the S.E.P. Task Group on BAL. Eur Respir J 2:561-585 11. Hartman B, Koss M, Hui A, et al (1985) Pneumocystis carinii pneumonia in the acquired immunodefic.iency syndrome (AIDS): diagnosis with bronchial brushings, biopsy, and bronchoalveolar lavage. Chest 87:603-607 12. Radio SJ, Rennard SI, Kessinger A, Vaughan WP, Linder J (1989) Breast carcinoma in bronchoalveolar lavage. Arch Pathol Lab Med 113:333-336 13. Stover DE, Zaman MB, Hajdu SI, et al (1984) Bronchoalveolar lavage in the diagnosis of diffuse pulmonary infiltrates in the immunosuppressed host. Ann Intern Med 101:1-7 14. Young JA, Hopkin JM, Cuthbertson WP (1984) Pulmonary infiltrates in immunocompromised patients: diagnosis by cytological examination of bronchoalveolar lavage fluid. J Clin Pathol 37:390-397 15. Springmeyer SC, Hackman RC, Holle R, et al (1986) Use of bronchoalveolar lavage to diagnose acute diffuse pneumonia in the immunocompromised host. J Infect Dis 154:604-610 16. Legge RIt, Thompson AB, Linder J, Woods GL, Robbins RA, Moulton AL, Rennard SI (1988) Acyclovir responsive herpetic tracheobronchitis. Amer J Med 85:561-563 17. Pierce CH, Knox AW (1960) Ciliocytophthoria in sputum from patients with adenovirus infections. Proc Soc Exp Biol Med 104:492--495 18. Warner NE, McGrew EA, Nanos S (1964) Cytologic study of the sputum in cytomegalic inclusion disease. Acta Cytol 8:311-315
Future Directions for BAL
1055
19. Myerson D, Hackman RC, Myers JD (1984) Diagnosis of cytomegaloviral pneumonia by in situ hybridization. J Infect Dis 150:272-277 20. Masih A, Rennard SI, Binkley LI, et al (1987) Detection of cytomegalovirus in bronchoalveolar lavage specimens by cytology, tissue culture, fluorescent monoclonal antibodies and in situ hybridization. Acta Cytol 31:648 21. Crystal RG, Roberts WC, Hunninghake GW, et al (1981) Pulmonary sarcoidosis: a disease characterized and perpetuated by activated lung T lymphocytes. Ann Intern Med 94:73 22. Venet A (1984) Immunology of sarcoidosis. Ann Med Intern 135:113 23. Hunninghake GW, Crystal RG (1981) Pulmonary sarcoidosis: a disorder mediated by excess helper T-lymphocyte activity at sites of disease activity. N Engl J Med 305:429-434 24. Costabel U, Zaiss A, Wagner DJ, et al (1988) Value of bronchoalveolar lavage lymphocyte subpopulations for the diagnosis of sarcoidosis. In: Sarcoidosis and other granulomatus disorders. Amsterdam: Excepta Medica, Grassi C, Rizzato G, Possi E, eds. p 429 25. Ainslie G, du Bois RM, Poulter LW (1989) Relation between immunocytological features of bronchoalveolar lavage fluid and clinical indicies in sarcoidosis. Thorax 44:501 26. Keogh BA, Hunninghake GW, Line BR, et al (1983) The alveolitis of pulmonary sarcoidosis-evaluation of natural history and alveolitis dependent changes in lung function. Am Rev Respir Dis 128:256 27. Costabel U, Bross KJ, Gujman J, et al (1986) Predictive value of bronchoalveolar lavage T cell subsets for the course of pulmonary sarcoidosis. Am NY Acad Sci 465:418 28. Ward K, O'Connor C, Odium C, Fitzgerald MS (1989) Prognostic value of bronchoalveolar lavage in sarcoidosis: the critical influence of disease presentation. Thorax 44:6 29. Pinkston P, Saltini C, MuUer-Quernheim J, Crystal RG (1987) Corticosteroid therapy suppresses spontaneous interleukin 2 release and spontaneous proliferation of lung T lymphocytes of patients with active pulmonary sarcoidosis. J Immunol 139:755-760 30. Reynolds HY, Chretian J (1984) Respiratory tract fluids: analysis of content and contemporary use of understanding lung diseases. DM 30:1-103 31. Macklem PT (1972) Obstruction in small airways--a challenge to medicine. Am J Med 52:721724 32. Macklem PT, Thurlbeck WM, Fraser RG (1971) Chronic obstructive disease of small airways. Ann Intern Med 74:167-177 33. Niewoehner DE, Kleinerman J, Rice DB (1974) Pathologic changes in the peripheral airways of young cigarette smokers. N Engl J Med 291:755-758 34. Lieberman J (1973) Involvement of leukocytic proteases in emphysema and antitrypsin deficiency. Arch Environ Health 27:196-200 35. Janoff A (1985) Elastases and emphysema. Current assessment of the protease-antiprotease hypothesis. Am Rev Respir Dis 132:417-433 36. Brower MS, Harpel PC (1983) Alpha-1 antitrypsin-human leukocyte elastase complexes in blood: quantification by an enzyme-linked differential antibody immunoassay and comparison with alpha 2-plasmin inhibitor-plasmin complexes. Blood 842-849 37. Jochum M, PeUetier A, Boudier C, Pauli G, Bieth JH (1985) The concentration of leukocyte elastase-a 1-proteinase inhibition complex in bronchoalveolar lavage fluids of healthy human subjects. Am Rev Respir Dis 132:913-914 38. Fujita J, Daughton DM, Stahl MG, Rennard SI (1987) Neutrophil elastase burden is decreased after smoking reduction. Am Rev Respir Dis 135:A147 39. Daniele RP (1988) Immunology and immunologic diseases of the lung. Boston: Blackwell Scientific Publications 40. Casale TB, Wood D, Richerson HB, Zehr B, Zavala D, Hunninghake GW (1987) Direct evidence of a role for mast cells in the pathogenesis of antigen-induced bronchoconstriction. J Clin Invest 80:1507-1511 41. Fick RB Jr, Metzger WJ, Richerson HB, Zavala DC, Moseley PL, Schoderbek WE, Hunninghake GW (1987) Increased bronchovascular permeability after allergen exposure in sensitive asthmatics. J Appt Physiol 63:1147-1155 42. Radomski JS, ,larrell BE, Carabasi RA, Yang SL (1987) Renal autotransplantation and extracorporeal reconstruction for complicated benign and malignant diseases of the urinary tract. J Cardiovasc Surg 28:413-419
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43. Yano T, Yasumoto K, Nomoto K (1989) Generation and expansion of lymphokine-activated killer cells from lymph node lymphocytes in human lung cancer. Eur J Canc Clin Oncol 25:201-208 44. Saltini C, Spurzem JR, Lee JJ, Pinkston P, Crystal RG (1986) Spontaneous release of interleukin 2 by lung T lymphocytes in active pulmonary sarcoidosis is primarily from the Leu3+DR+ T cell subset. J Clin Invest 77:1962-1970 45. Iida J, Nishi. N, Saiki I, Mizukoshi N, lshihara C, Tokura S, Azuma 1 (1989) Macrophage activation and host augmentation against Sendai herpes simplex virus (HSV) infections with synthetic polypeptides in mice. Int J lmmunopharmacol 22:249-258 46. Jensen WA, Rose RM, Burke RH Jr, Anton K, RemoId HG (1988) Cytokine activation of antibacterial activity in human pulmonary macrophages: comparison of recombinant interferon-gamma and granulocyte-macrophage colony-stimulating factor. Cell Immunol 177:369377 47. Davis LG, Dibner MD, Battey JF (1986) Basic methods in molecular biology. New York: Elsevier Science Publishing Co. Inc. 48. Watson JD, Tooge J, Kurtz DT (1983) Recombinant DNA, a short course. New York: Scientific American Books
Lung
© Springer-¥erlag New York Inc. 1990
Future Directions for Bronchoalveolar Lavage Stephen I. Rennard Pulmonary and Critical Care Medicine Section, Department of Internal Medicine, University of Nebraska Medical Center, Omaha, Nebraska, USA
Abstract. Bronchoalveolar lavage provides a means to sample the lung. The ability to describe material obtained from the lower respiratory tract has greatly facilitated pulmonary diagnosis, and it is likely that improvements in diagnostic techniques will continue to develop. The ability to accurately measure lower respiratory tract components has permitted staging of lung disease. Such staging will probably provide improved prognostic insight and permit the development of new therapeutic strategies in lung disease. Key words: Bronchoalveolar lavagemStaging of lung disease--Viable cells--Experimental intervention. Introduction Predicting future directions, particularly for a technique used both clinically and for research, is a difficult task. Bronchoalveolar lavage (BAL) has a long history dating back over 100 years [1-3]. It has been in widespread use, however, only since the introduction of the flexible fiberoptic bronchoscope 20 years ago [4]. Through this instrument lavage is easily performed, and it has rapidly made the lower respiratory tract accessible to investigators [5-7]. As a result, BAL has changed the ability of physicians and researchers to approach pulmonary disease. In particular, BAL has made the lung an accessible organ. By virtue of the fact that the procedure can be performed safely and easily and that it is well tolerated, a vast array of material has been made available for diagnostic and research purposes. The procedure of bronchoalveolar lavage is generally performed with the bronchoscope in a wedged position [8-10]. Sterile saline is infused through the Offprint requests to: Dr. S. I. Rennard, Pulmonary and Critical Care Medicine Section, Department of Internal Medicine, University of Nebraska Medical Center, 600 So. 42nd, Omaha, NE 68198-2465, USA.
Future Directions for B A L
1051
biopsy port of the bronchoscope, and this infused saline mixes with the fluid lining the airways and alveolar spaces of the lung. This fluid can then be recovered by aspiration through the biopsy channel of the bronchoscope. The resulting lavage fluid contains some of the material previously present within the lumen of the airways and alveoli and thus represents a direct sampling of the lung. This sampling presents the pulmonologist with several distinctly different opportunities including: 1) description of the material present within the lower respiratory tract and hence diagnosis; 2) quantification of various components present within the lower respiratory tract and hence staging; 3) recovery of viable cells from the lower respiratory tract and hence insight into normal and abnormal cellular physiology; and 4) specific performance of experiments in the lower respiratory tract.
Description and Diagnosis While bronchoalveolar lavage primarily samples the luminal contents, many abnormal processes within the lung are characterized by abnormalities within the air spaces. The material obtained by lavage can, therefore, be of diagnostic use. Pneumocystis carinii pneumonia, for example, is characterized by an exuberant alveolar infiltrate [11]. When stained with an appropriate technique, material covered by lavage can readily yield a diagnosis of pneumocystis carinii. This procedure has almost completely replaced the need for open lung biopsy in the diagnosis of pneumocystis pneumonia. Other pathologic processes can also be present within the airway. For example, cancer infiltrating air spaces or present on the airway surfaces can also be recovered by lavage [9]. Special techniques, for example the use of monoclonal antibodies specific for tumor markers, can greatly aid in the diagnosis of cancer on BAL cytology by increasing the diagnostic yield [12]; it can also result in the diagnosis of infectious processes [9, 13-15], and cytopathologic changes characteristic of viral infections can be readily recognized as well [9, 16-18]. Again, the application of new biological techniques such as the use of cDNA probes for viral specific DNA can greatly increase the sensitivity of diagnostic methods for specific pathogens [19, 20]. It is likely that the appfication of monoclonal antibodies and cDNA probes will increase the power of BAL as a diagnostic tool in the future. This is likely to have the biggest impact in the diagnosis of malignancies and in viral infections.
Measurement and Staging Measurement of lower respiratory tract components permits the "staging" of lower respiratory tract disease. Staging can be very useful in assessing relative severity of a condition and can, therefore, lead to important prognostic and therapeutic decisions. In addition, by being able to evaluate the severity of a disease process, the ability to quantify lower respiratory tract components can permit the assessment of therapeutic interventions.
1052
S . I . Rennard
Sarcoidosis, for example, has been associated with lower respiratory tract lymphocytosis [21, 22]. In particular, T helper cells are found to be increased in both number and percentage [23-25]. This increase can be quantified and therefore permit staging. In general, patients with higher numbers and percentages of lymphocytes are thought to have a more "active" disease which is more likely to progress [26, 27], although a number of controversies persist [28]. Similarly, therapeutic intervention, for example with glucocorticoids, is often associated with a reduction in the magnitude of the lymphocytosis [29]. In a similar fashion, it is possible to assess a great variety of lower respiratory tract components [8-10, 30]. Cigarette smoking is thought to lead to "small airways disease," a condition characterized by accumulation of macrophages, neutrophils and their secretory products in the small airways of the lung [31-33]. The release of secretory products, particularly neutrophil elastase, is thought to lead to destruction of lung tissue and subsequently to the development of pulmonary emphysema [34, 35]. By virtue of its ability to sample the air spaces of the lung, BAL can quantify these potential mediators of disease. It is, for example, possible to measure lower respiratory tract neutrophil elastase as a complex to alpha-1 antitrypsin [36, 37]. A therapeutic intervention, like cigarette smoke reduction, can then be assessed in terms of its ability to reduce the lung's burden of this potentially damaging protease [38]. The use of bronchoalveolar lavage to stage various lung diseases is clearly an area of active investigation. The future will undoubtedly lead to great insights regarding the quantitative nature of lower respiratory tract cells and components with respect to pathophysiologic mechanisms, disease prognosis and therapeutic intervention. While the use of these staging techniques is currently not fully established, it is almost certain that there will be increasingly wide applications in the future.
Function and Pathophysioiogy Bronchoalveolar lavage is capable of recovering viable cells from the lower respiratory tract [8, 9, 39]. These viable cells can be maintained in culture ex vivo, and their functional state can be assessed. A large spectrum of assay systems can be applied to these cells recovered from the lower respiratory tract. These include biological assays, biochemical assays, immunoassays and studies of gene regulation. These studies have permitted the description of release of growth factors, chemotactic factors and cytotoxic factors from lower respiratory tract cells. In addition, it has been possible to characterize and quantify the release of specific mediators in individual disease states. Finally, it has been possible to determine the state of gene activation in cells recovered from the lower respiratory tract. Taken together, these types of studies have greatly helped explain the interactions between the cells in the lower respiratory tract involved in normal health and in disease states. Specific mechanisms of lung injury and repair have been delineated. It is almost certain that these cytokine networks and the various cellular interactions which they mediate will be described in increasing detail in the near future.
Future Directions for BAL
1053
Experimentation and Manipulation Bronchoalveolar lavage not only samples the lung, it creates the possibility of introducing material into the lung. This permits experimental manipulations of the lower respiratory tract. It is possible, for example, to introduce antigens into the sensitive asthmatic directly through the bronchoscope. Using these techniques, Casale et al. and Fick et al. have demonstrated that local application of antigen can lead to the release of histamine from mast cells and to the leakage of albumin from the intravascular spaces into the airway lumen [40, 41]. It is likely that the ability of BAL to introduce materials into the lung will be greatly exploited by investigators in the near future. BAL also opens the horizons for "extracorporeal surgery" of lung cells. Extracorporeal surgery is now routinely performed on kidneys [42]. A kidney can be removed from a patient under general anesthesia, be microdissected removing vascular lesions or small tumor implants, be repaired and then restored to the patient. Similar "surgery" could be performed on lung cells using BAL instead of nephrectomy. For example, BAL cells can be removed from the body, subjected to "repair" mechanisms, and then be reinfused into the lungs. Examples of currently available methods to alter cellular functions in cells obtained from the lung include: 1) specific expansion of subpopulations of lung cells using techniques of cell culture [43]; 2) selection of specific types of lung cells using monoclonal antibodies [44]; 3) activation of lung cells using specific monokines or cytokines in vitro [45, 46]; and 4) introduction of novel genes into the cells obtained from the lung [47, 48]. Examples of the potential for such surgery in lung disease include: 1) the activation of immune and inflammatory effector cells in diseased states where such cells are ineffective, e.g., infections and malignancy; 2) in diseases where lung cells are characterized by inappropriate activation, such as immune and inflammatory lung disease or asthma, it may be possible to introduce specific suppressor cells which could, when reintroduced in vivo, result in a suppression of the abnormal immune and inflammatory responses; 3) it may be possible to introduce a normal gene, and therefore restore to normal function, in an individual with a genetic abnormality, e.g., in alpha-1 protease inhibitor deficiency or in cystic fibrosis; 4) it may be possible to utilize the ability of BAL cells to mediate lung repair processes to control the regulation of such processes. As such, it may be possible to restore function in lungs which have been disrupted by pulmonary fibrosis or by widespread emphysema. Clearly, such applications are beyond current capabilities. Nevertheless, currently available technology suggests that they are within the realm of imagination and represent reasonable long-term future directions.
Summary The ability of lavage to recover viable cells has permitted great insights into the pathophysiologic mechanisms of disease and the normal physiologic mechanisms responsible for lower respiratory tract cellular function. It is likely that
1054
S.I. Rennard
these studies will continue to advance our understanding of the interactions between cells and the mediators which make such interactions possible. Bronchoalveolar lavage also makes the lung accessible to experimental intervention. Such experimental intervention will not only greatly aid in the understanding of physiologic mechanisms of disease states but also raises the possibility that specific therapeutic interventions can be targeted directly to the lung. The rapid advances in biological techniques including cell culture, biochemical analysis, the application of monoclonal antibodies and the application of molecular biological techniques represents a "new biology." The ability of bronchoalveolar lavage to make the lung an accessible organ ensures that this new biology can be readily applied to the lung.
References 1. Bernard J, Gee L, Fick RB (1980) Bronchoalveolar lavage (editorial). Thorax 35:1-8 2. Rogers RM, Braunstein MS, Shuman JR (1972) Role of bronchopulmonary lavage in the treatment of respiratory failure: a review. Chest 62(Suppl):95-106 3. Ramirez-R J, Kieffer RF, Ball WC (1965) Bronchopulmonary lavage in man. Ann Intern Med 63:819-828 4. lkeda S, Yanai N, Ishikawa S (1968) Flexible bronchofiberscope. Keio J Med 17:1-16 5. CantreUET, Warr GA, Busbee DL, et al (1973) Induction of arylhydrocarbon hydroxylase in human pulmonary alveolar macrophages by cigarette smoking. J Clin Invest 52:1881-1884 6. Waldmann RH, Jurgensen PF, Olen GN, et al (1973) Immune response of the human respiratory tract: immunoglobulin levels and influenza virus vaccine antibody response. J Immunol 111:38--41 7. Reynolds HY, Newball HH (1974) Analysis of proteins and respiratory cells obtained from human lungs by bronchial lavage. J Lab Clin Med 84:559-573 8. Reynolds HY (1987) State of the art: bronchoalveolar lavage. Am Rev Respir Dis 135:250263 9. Linder J, Rennard S Bronchoalveolar lavage. Chicago: ASCP Press, pp 1-196 10. Costabel U, Danel C, Haslam P, Higgenbottam T, Klech H, Pohl W, Rennard S, Rossi G, Rust M, Semenzato Get al (1989) Technical recommendations and guidelines for bronchoalveolar lavage (BAL). Report of the S.E.P. Task Group on BAL. Eur Respir J 2:561-585 11. Hartman B, Koss M, Hui A, et al (1985) Pneumocystis carinii pneumonia in the acquired immunodefic.iency syndrome (AIDS): diagnosis with bronchial brushings, biopsy, and bronchoalveolar lavage. Chest 87:603-607 12. Radio SJ, Rennard SI, Kessinger A, Vaughan WP, Linder J (1989) Breast carcinoma in bronchoalveolar lavage. Arch Pathol Lab Med 113:333-336 13. Stover DE, Zaman MB, Hajdu SI, et al (1984) Bronchoalveolar lavage in the diagnosis of diffuse pulmonary infiltrates in the immunosuppressed host. Ann Intern Med 101:1-7 14. Young JA, Hopkin JM, Cuthbertson WP (1984) Pulmonary infiltrates in immunocompromised patients: diagnosis by cytological examination of bronchoalveolar lavage fluid. J Clin Pathol 37:390-397 15. Springmeyer SC, Hackman RC, Holle R, et al (1986) Use of bronchoalveolar lavage to diagnose acute diffuse pneumonia in the immunocompromised host. J Infect Dis 154:604-610 16. Legge RIt, Thompson AB, Linder J, Woods GL, Robbins RA, Moulton AL, Rennard SI (1988) Acyclovir responsive herpetic tracheobronchitis. Amer J Med 85:561-563 17. Pierce CH, Knox AW (1960) Ciliocytophthoria in sputum from patients with adenovirus infections. Proc Soc Exp Biol Med 104:492--495 18. Warner NE, McGrew EA, Nanos S (1964) Cytologic study of the sputum in cytomegalic inclusion disease. Acta Cytol 8:311-315
Future Directions for BAL
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