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Roles of the Neutrophil and Other Mediators in Adult Respiratory Distress Syndrome

      The adult respiratory distress syndrome (ARDS) and transfusion-related acute lung injury (TRALI) are characterized by diffuse, acute lung injury. Most likely, TRALI is a type of ARDS although it is associated with a much lower morbidity and mortality than found with classic ARDS. For years, the pathogenesis of ARDS has been explained by the complement hypothesis in which pulmonary neutrophilic sequestration and degranulation follow complement-mediated neutrophil chemotaxis. A definitive role for the neutrophil in diffuse, acute lung injury, however, has not been established. Although numerous chemoattractants for neutrophils are generated in the lungs and, through degranulation and formation of toxic oxygen free radicals, the neutrophil is fully capable of causing tissue injury, substantial evidence refutes the requirement for neutrophils in diffuse, acute lung injury. Other potential factors in the pathogenesis of ARDS include primary endothelial cell injury, alveolar macrophage activity, and hemostatic disorders.
      A definitive role for the neutrophil in diffuse, acute lung injury is controversial and has not yet been established. Results of many in vitro studies, animal experiments, and in vivo human studies suggest that neutrophils cause acute lung injury in certain settings.
      • Thommasen HV
      The role of the polymorphonuclear leukocyte in the pathogenesis of the adult respiratory distress syndrome.
      • Tate RM
      • Repine JE
      Neutrophils and the adult respiratory distress syndrome.
      Neutropenia, from various causes, apparently has prevented or at least attenuated the development of adult respiratory distress syndrome (ARDS) in both experimental animal systems and in vivo human studies. In contrast, mounting evidence suggests that the neutrophil is neither necessary nor sufficient in the pathogenesis of diffuse, acute lung injury.

      CLINICAL CHARACTERISTICS OF ARDS AND TRANSFUSION-RELATED ACUTE LUNG INJURY

      Diffuse, acute lung injury characterizes both ARDS and transfusion-related acute lung injury (TRALI). Most likely, TRALI is a form of ARDS, albeit with an unusually good prognosis. The mortality associated with TRALI is less than 10%. This outcome is in contrast to the overall 50 to 60% mortality associated with ARDS from well-substantiated causes. The explanation for this difference is unclear. Morbidity is also lower in TRALI than in ARDS. For example, approximately 80% of patients with TRALI were noted to have rapid resolution of pulmonary infiltrates and return of arterial blood gas values to normal within 96 hours after the initial respiratory insult. In 17% of patients, however, pulmonary infiltrates were noted to persist for at least 7 days after the transfusion reaction.
      • Popovsky MA
      • Abel MD
      • Moore SB
      Transfusion-related acute lung injury associated with passive transfer of antileukocyte antibodies.
      • Popovsky MA
      • Moore SB
      Diagnostic and pathogenetic considerations in transfusion-related acute lung injury.
      More than one etiologic factor may play a role in the development of ARDS in a given patient. For example, the need for transfusion of blood products is likely in a patient who has sustained major trauma, and both the trauma and the transfusion may be instrumental in the development of ARDS. Thus, TRALI can be diagnosed when transfusion of blood products is the sole etiologic factor and other known causes of ARDS have been excluded.
      “ARDS” is a descriptive term applied to diffuse, acute lung lesions that result in noncardiogenic pulmonary edema. It was first clearly described in 1967 by Ashbaugh and associates,
      • Ashbaugh DG
      • Bigelow DB
      • Petty TL
      • Levine BE
      Acute respiratory distress in adults.
      who recognized similarities between acute respiratory distress in adults and hyaline membrane disease in infants. The numerous precipitating events include pulmonary infections, overdose of narcotics, severe trauma, pancreatitis, and gram-negative septicemia.
      • Balk R
      • Bone RC
      The adult respiratory distress syndrome.
      ARDS is a common disorder; it is estimated to affect approximately 150,000 patients each year in the United States.
      TRALI is characterized by acute respiratory distress that typically occurs within 4 hours after a transfusion. It is an unusual and often unsuspected cause of diffuse, acute lung injury. In the past, many cases of acute pulmonary edema as a consequence of transfusion have most likely been misdiagnosed as circulatory overload. The clinical similarity between TRALI and ARDS is suggested by the observation that multiple transfusions have been the only substantiated etiologic factor in three separate series of patients with ARDS.
      • Maunder RJ
      Clinical prediction of the adult respiratory distress syndrome.
      The pathogenesis of TRALI is presumed to be due primarily to passive transfusion and reaction of donor granulocyte or lymphocyte antibodies with recipient granulocytes. “Toxic” products released from the granulocytes lead to acute, edematous lung injury (Fig. 1). In 36 TRALI reactions, Popovsky and Moore
      • Popovsky MA
      • Moore SB
      Diagnostic and pathogenetic considerations in transfusion-related acute lung injury.
      demonstrated that 89% were correlated with the passive transfusion of donor granulocyte antibodies and 72% were associated with donor lymphocyte antibodies. HLA-specific antibodies were identified in 65% of cases. In a minority of cases, however, the pathogenesis may have been due to reaction of recipient granulocyte or lymphocyte antibodies with donor granulocytes. The type of blood product transfused, the volume of plasma, and the titer of antibody may not be directly correlated with the occurrence or the severity of TRALI.
      Figure thumbnail gr1
      Fig. 1Presumed pathogenesis of transfusion-related acute lung injury.
      Implicated donors of these granulocyte antibodies are those who, because of previous exposure to alloantigens, have had an opportunity to develop antibodies against those antigens that also may be carried on granulocytes. These donors typically include multiparous women or persons with a history of previous transfusions of blood.
      Clinical manifestations of both ARDS and TRALI include acute onset of respiratory failure, tachypnea, cyanosis, and chest rales on auscultation. Additional findings include diffuse, bilateral interstitial (early) and alveolar (later) infiltrates on a chest roentgenogram, decreased arterial oxygen pressure (PaO2 less than 50 mm Hg with a fractional concentration of oxygen in inspired gas [FiO2] of more than 0.6), increased alveolar-arterial oxygen pressure gradient, normal pulmonary capillary wedge pressure, overall decreased lung compliance (less than 50 ml/cm H2O), and increased shunt fraction and dead-space ventilation. The shunt fraction and dead-space ventilation may exceed 50% in severe ARDS.
      • Balk R
      • Bone RC
      The adult respiratory distress syndrome.
      In both ARDS and TRALI, mechanical ventilation, positive end-expiratory pressure, and supplemental oxygen are often used with the goal of achieving a PaO2 of 60 mm Hg and an oxygen saturation of at least 90%.

      ROLE OF THE NEUTROPHIL IN DIFFUSE ACUTE LUNG INJURY

      The Complement Hypothesis.

      For several years, the pathogenesis of ARDS has been explained by the “complement hypothesis.” This hypothesis suggests that an inciting event—;for example, septicemia—;is followed by complement activation to generate C5a, a complement breakdown product and potent chemoattractant for neutrophils. C5a presumably attracts large numbers of neutrophils into a pulmonary vascular bed that already functions normally as a major reservoir for marginated neutrophils. Acute, edematous lung injury results from release of “toxic” products from the neutrophils (Fig. 2). Proof both to support and to refute this simple hypothesis exists.
      Figure thumbnail gr2
      Fig. 2Release of toxic products from neutrophils.
      In 1968, Kaplow and Goffinet
      • Kaplow LS
      • Goffinet JA
      Profound neutropenia during the early phase of hemodialysis.
      published their observations on four patients undergoing 33 sessions of hemodialysis. All showed appreciable temporary neutropenia within 15 minutes after the start of dialysis. Blood total leukocyte counts became normal within 1 hour but an increase in band neutrophils was detected up to 5 hours after dialysis. Craddock and colleagues
      • Craddock PR
      • Fehr J
      • Dalmasso AP
      • Brigham KL
      • Jacob HS
      Hemodialysis leukopenia: pulmonary vascular leukostasis resulting from complement activation by dialyzer cellophane membranes.
      suggested that it was the cellophane membrane of the dialysis tubing that activated complement. (The polysaccharides zymosan, inulin, and endotoxin are known to activate complement; cellophane also is a polysaccharide and would be expected to activate complement in a similar fashion.) In the 34 patients in the study by Craddock and co-workers, severe, transient neutropenia and monocytopenia developed within the first 20 minutes of dialysis. Pulmonary vascular leukostasis followed. Total hemolytic complement and serum C3 levels decreased; however, serum C1 levels decreased only minimally, a result that suggests activation of complement predominantly through the alternate pathway (which bypasses C1 activation).
      Hammerschmidt and associates
      • Hammerschmidt DE
      • Weaver LJ
      • Hudson LD
      • Craddock PR
      • Jacob HS
      Association of complement activation and elevated plasma-C5a with adult respiratory distress syndrome: pathophysiological relevance and possible prognostic value.
      suggested that complement activation predicted the development of ARDS secondary to sepsis, pancreatitis, or severe trauma. Of 33 patients in whom ARDS developed, 31 had increased serum C5a values compared with only 5 of 28 patients who did not have ARDS (P<0.00001). Even after excluding patients with sepsis (which commonly causes an increase in serum complement level), a highly statistically significant difference was still noted. The authors concluded that serum C5a levels were highly predictive of ARDS.
      Evidence to refute this conclusion was published by Duchateau and colleagues
      • Duchateau J
      • Haas M
      • Schreyen H
      • Radoux L
      • Sprangers I
      • Noel FX
      • Braun M
      • Lamy M
      Complement activation in patients at risk of developing the adult respiratory distress syndrome.
      and Weinberg and co-workers
      • Weinberg PF
      • Matthay MA
      • Webster RO
      • Roskos KV
      • Goldstein IM
      • Murray JF
      Biologically active products of complement and acute lung injury in patients with sepsis syndrome.
      in 1984. The former found that abnormal consumption of C3 as well as increased C5a-like activity occurred in 84% and 86%, respectively, of patients with ARDS. Even though the increased C5a-like activity was closely associated with clinical conditions that predispose to ARDS, the authors concluded that changes in complement components were no more predictive for the development of ARDS than were the initial clinical manifestations.
      The studies by Weinberg and associates
      • Weinberg PF
      • Matthay MA
      • Webster RO
      • Roskos KV
      • Goldstein IM
      • Murray JF
      Biologically active products of complement and acute lung injury in patients with sepsis syndrome.
      in 40 patients revealed that serum des-Arg-C3a and des-Arg-C5a levels were increased in 35 and 38 patients, respectively, with suspected sepsis. des-Arg-C3a and des-Arg-C5a are biochemically active, yet stable, cleavage products of C3a and C5a, respectively.
      • Cooper NR
      The complement system.
      Mean levels of these complement components did not differ between patients without or with mild to moderate lung injury and patients with severe lung injury (ARDS). Neither did the mean values differ in comparisons of patients whose initial lung injury worsened and those whose injury resolved. Thus, measurements of these complement breakdown products were not thought to be predictive of either the course or the severity of the lung injury.
      Because of the characteristics of the acute-phase response, serum complement levels in patients in whom ARDS develops are unlikely to differ significantly from levels in critically ill patients who are predisposed to the development of ARDS but do not have this disorder. The acute-phase response is a generalized reaction to either localized or systemic diseases. It is primarily mediated by interleukin 1, a polypeptide secreted from various types of cells including blood cells and tissue macrophages. Two of the many clinical and cellular manifestations of interleukin 1 are an increase in the number of both mature and immature circulating neutrophils and increased secretion of hepatic acute-phase proteins. The latter include complement components that are likely to be secreted in substantial quantities in patients who are predisposed to or have ARDS.
      • Maury CPJ
      Interleukin 1 and the pathogenesis of inflammatory diseases.
      • Dinarello CA
      • Mier JW
      Lymphokines.
      In addition to complement activation, pulmonary sequestration of neutrophils with subsequent release of their “toxic” products is central to the complement hypothesis. Evidence for this premise ranges from in vitro biochemical and physiologic studies of neutrophil function to histologic examination of lung biopsy specimens from patients with ARDS in the setting of severe neutropenia. Fundamental questions to be considered are the following: Is pulmonary sequestration of neutrophils both necessary and sufficient for the development of ARDS? If so, by what mechanisms are neutrophils recruited to the critical site of involvement? Once at that site, what does the neutrophil use to mediate cellular and tissue injury? The first question is best considered in the context of answers to the last two questions.

      Neutrophil Chemotaxis.

      Chemotactic agents recruit neutrophils into the pulmonary vasculature (Fig. 2). These agents create a concentration gradient along which the neutrophil moves toward its destination. Neutrophils alter their direction in response to a change in chemical gradient of as little as 0.1% in concentration.
      • Boxer GJ
      • Curnutte JT
      • Boxer LA
      Polymorphonuclear leukocyte function.
      Indirect experimental evidence from in vitro and animal models suggests a host of diverse chemotactic agents for neutrophils that may function in humans: complement cascade components, biomolecules intimately associated with coagulation, arachidonic acid-derived metabolites, lymphokines, and numerous other proteins and metabolites. C5a probably is the best known chemotactic agent for the neutrophil. This cleavage product of C5 induces neutrophil aggregation and degranulation and stimulates oxidative metabolism.
      • Smith EL
      • Hill RL
      • Lehman IR
      • Lefkowitz RJ
      • Handler P
      • White A
      Factor Bb, another complement cascade component, is part of a proteolytic complex that cleaves C3 and C5 and thus generates additional chemotactic agents.
      Fibrinogen, fibrinopeptide B, plasminogen activator, kallikrein, platelet-derived growth factor, platelet factor IV, and platelet-activating factor are not only chemotactic for neutrophils but also prominent contributors in hemostasis. Disturbances in hemostasis may be critical in the pathogenesis of ARDS. Fibrinopeptide B is generated by the thrombin-catalyzed conversion of fibrinogen to fibrin in clot formation. Plasminogen activator is secreted by both macrophages and endothelial cells and cleaves plasminogen proteolytically; the resultant product, plasmin, lyses fibrin and activates C1, C3, and Hageman factor (factor XII of the intrinsic pathway of coagulation).
      • Smith EL
      • Hill RL
      • Lehman IR
      • Lefkowitz RJ
      • Handler P
      • White A
      Activated Hageman factor, in turn, catalyzes the conversion of prekallikrein to the chemoattractant kallikrein. The cycle is completed when kallikrein autocatalytically activates additional Hageman factor. Kallikrein also enzymatically cleaves kininogen to generate the potent vasodilator bradykinin. The greatly increased vascular permeability that occurs as a result of this vasodilatation is probably instrumental in generation of the pulmonary edema characteristic of ARDS and TRALI.
      Platelet-derived growth factor is chemotactic for both macrophages and neutrophils at concentrations of approximately 700 pmol/liter and 32 pmol/liter, respectively.
      • Deuel TF
      • Senior RM
      • Huang JS
      • Griffin GL
      Chemotaxis of monocytes and neutrophils to platelet-derived growth factor.
      It also is a potent mitogen toward mesenchymal cells and may be instrumental in the fibroblastic proliferation characteristic of the organizing phase of ARDS. Platelet factor IV promotes coagulation by inhibiting the interaction between heparin-antithrombin III complexes and serine protease coagulation factors.
      • Smith EL
      • Hill RL
      • Lehman IR
      • Lefkowitz RJ
      • Handler P
      • White A
      Platelet-activating factor not only is chemotactic for and an activator of neutrophils but also activates platelets, induces smooth muscle contraction, and has vasoactive effects approximately 1,000 times more intense than those of histamine. Infusion of this factor into rabbits has been shown to induce pronounced neutropenia, pulmonary platelet sequestration, and thrombocytopenia.
      • Pinckard RN
      Platelet-activating factor.
      Other chemotactic agents for neutrophils are the Escherichia coli-derived N-formylmethionylleucylphenylalanine tripeptide,
      • Malech HL
      • Gallin JI
      Neutrophils in human diseases.
      • Nauseef WM
      • De Alarcon P
      • Bale JF
      • Clark RA
      Aberrant activation and regulation of the oxidative burst in neutrophils with Mo1 glycoprotein deficiency.
      lymphocyte- or macrophage-derived lymphokines such as interleukin 1,
      • Dinarello CA
      • Mier JW
      Lymphokines.
      immunoglobulin fragments, collagen fragments, prostaglandins, and leukotrienes (in particular, leukotriene B4).
      • Thommasen HV
      The role of the polymorphonuclear leukocyte in the pathogenesis of the adult respiratory distress syndrome.
      • Maury CPJ
      Interleukin 1 and the pathogenesis of inflammatory diseases.
      • Smith EL
      • Hill RL
      • Lehman IR
      • Lefkowitz RJ
      • Handler P
      • White A
      • Curnutte JT
      • Boxer LA
      Clinically significant phagocytic cell defects.
      Production of leukotriene B4 is stimulated by platelet-activating factor. This leukotriene is a potent arachidonic acid-derived chemoattractant (active at a concentration of 0.1 μM), is a potentiator of oxygen free-radical generation in stimulated neutrophils, and augments neutrophil endothelial adherence.
      • Gay JC
      • Beckman JK
      • Brash AR
      • Oates JA
      • Lukens JN
      Enhancement of chemotactic factor-stimulated neutrophil oxidative metabolism by leukotriene B4.
      • Hoover RL
      • Karnovsky MJ
      • Austen KF
      • Corey EJ
      • Lewis RA
      Leukotriene B4 action on endothelium mediates augmented neutrophil/endothelial adhesion.
      The last phenomenon is believed to be especially critical to neutrophil-mediated cellular injury.
      • Nauseef WM
      • De Alarcon P
      • Bale JF
      • Clark RA
      Aberrant activation and regulation of the oxidative burst in neutrophils with Mo1 glycoprotein deficiency.
      Interleukin 1 is secreted by neutrophils and macrophages and is chemotactic for neutrophils, lymphocytes, and monocytes.
      • Maury CPJ
      Interleukin 1 and the pathogenesis of inflammatory diseases.
      • Dinarello CA
      • Mier JW
      Lymphokines.
      It is the primary mediator of the acute-phase response. Among its diverse biologic properties, it induces neutrophilia, stimulates synthesis of collagenases, and activates macrophages. Interleukin 1 also stimulates release of lactoferrin (a potential attenuator of neutrophil-mediated acute lung injury) from neutrophilic granules and induces a procoagulant state through its activation of endothelial cells.
      The foregoing discussion illustrates the plethora of potential chemoattractants for neutrophils. Various biomolecules generated under numerous conditions are capable of recruiting neutrophils into the lung. The importance of this recruitment, however, is only relative. The lung functions normally as a major steady-state reservoir of neutrophils.

      Degranulation and Toxic Oxygen Free-Radical Generation.

      Once at its destination, the neutrophil is well equipped to cause cellular and tissue injury and thus can radically alter pulmonary homeostasis. The neutrophil mediates inflammation and phagocytosis primarily by two mechanisms—;degranulation and oxygen free-radical generation (Fig. 2). Degranulation involves both the spillover of lysosome-stored granular contents into the extracellular environment (leading to inflammation) and fusion of the lysosome with a phagosome to form a phagolysosome in the process of intracellular phagocytosis.
      Human neutrophilic lysosomal granules contain a wide variety of biomolecules that mediate various effects. The neutrophilic proteases elastase, collagenase, and cathepsins degrade lung tissue and assist in amplification of the inflammatory process. Destruction of collagen, elastin, and basement membrane in ARDS is well substantiated.
      • Demling RH
      The role of mediators in human ARDS.
      Elastase is a proteolytic enzyme with a wide substrate specificity besides elastin. For example, it generates bradykinin from kininogen and cleaves C3 and C5.
      • Turino GM
      • Rodriquez JR
      • Greenbaum LM
      • Mandl I
      Mechanisms of pulmonary injury.
      It also degrades its own inhibitor (α1-proteinase inhibitor) as well as fibronectin, a potent opsonin and basement membrane stabilizer. Collagenase cleaves the carboxy-terminal quarter of the collagen molecule and thereby increases its susceptibility to further degradation by other proteases.
      • Smith EL
      • Hill RL
      • Lehman IR
      • Lefkowitz RJ
      • Handler P
      • White A
      Other lysosomal contents include cationic proteins, lysozyme, lactoferrin, and myeloperoxidase. Cationic proteins, which include both the bactericidal/membrane-active protein and the defensins, noncatalytically destroy bacteria by binding to their negatively charged cell surfaces.
      • Elsbach P
      • Weiss J
      • Franson RC
      • Beckerdite-Quagliata S
      • Schneider A
      • Harris L
      Separation and purification of a potent bactericidal/permeability-increasing protein and a closely associated phospholipase A2 from rabbit polymorphonuclear leukocytes.
      • Ganz T
      • Selsted ME
      • Szklarek D
      • Harwig SSL
      • Daher K
      • Bainton DF
      • Lehrer RI
      Defensins: natural peptide antibiotics of human neutrophils.
      Lysozyme is a β-N-acetylglucosaminidase that hydrolyzes native intracellular and phagocytized mucopolysaccharides. The cationic proteins and lysozyme play no known role in acute lung injury. Iron-saturated lactoferrin probably functions as a substrate-catalyst in the generation of the potent reactant hydroxyl free radical.
      • Badwey JA
      • Karnovsky ML
      Active oxygen species and the functions of phagocytic leukocytes.
      In contrast, iron chelators such as iron-free lactoferrin or deferoxamine have been shown to attenuate complement-dependent and neutrophil-mediated acute lung injury significantly in various animal models.
      • Till GO
      • Ward PA
      Oxygen radicals in complement and neutrophil-mediated acute lung injury.
      Myeloperoxidase, an abundant component of neutrophilic azurophilic granules, constitutes 5% of the dry weight of the neutrophil.
      • Smith EL
      • Hill RL
      • Lehman IR
      • Lefkowitz RJ
      • Handler P
      • White A
      On release into the phagosome, this heme-containing enzyme catalyzes the peroxidation of hydrogen peroxide and chloride ion with generation of water and the hypochlorite anion (OCl):
      H2O2 + Cl → H2O + OCl


      The hypochlorite ion inactivates the major antiprotease α1-proteinase inhibitor, acts as substrate for generation of hydroxyl free radicals, and is a potent inhibitor of bacterial respiration. It is able to oxidize adenine nucleotides (adenosine monophosphate, diphosphate, and triphosphate), cytochromes, and iron-sulfur proteins rapidly and irreversibly.
      • Albrich JM
      • McCarthy CA
      • Hurst JK
      Biological reactivity of hypochlorous acid: implications for microbicidal mechanisms of leukocyte myeloperoxidase.
      The myeloperoxidase-hydrogen peroxide-halide system is not considered entirely necessary for either neutrophil-mediated acute lung injury or microbicidal activity. For example, neutrophils from patients with congenital myeloperoxidase deficiency still display increased oxygen consumption, superoxide anion and hydrogen peroxide production, and substantial microbicidal activity. Thus, the morbidity in this enzyme deficiency manifests itself primarily as candidal infections in patients with diabetes mellitus rather than as diverse infections.
      • Malech HL
      • Gallin JI
      Neutrophils in human diseases.
      • Badwey JA
      • Karnovsky ML
      Active oxygen species and the functions of phagocytic leukocytes.
      The second mechanism by which neutrophils mediate inflammation is through oxygen free-radical generation (Fig. 2). Although direct evidence of a role for oxygen free radicals in acute lung injury in humans has not been found, these radicals have been strongly implicated as causal factors in acute lung injury in animal models. Various scavengers of these free radicals, such as mannitol, dimethylthiourea, and dimethyl sulfoxide, have been shown to be capable of inhibiting neutrophil-mediated edema in isolated animal lungs.
      • Tate RM
      • Repine JE
      Neutrophils and the adult respiratory distress syndrome.
      • Ward PA
      • Till GO
      • Harherill JR
      • Annesley TM
      • Kunkel RG
      Systemic complement activation, lung injury, and products of lipid peroxidation.
      • Brigham KL
      Role of free radicals in lung injury.
      Oxygen free radicals have been implicated as primary mediators in the pathogenesis of various biologic processes other than ARDS, including reperfusion injury after myocardial ischemia and normobaric oxygen toxicity. Southorn and Powis
      • Southorn PA
      • Powis G
      Free radicals in medicine. I. Chemical nature and biologic reactions. II. Involvement in human disease.
      recently published excellent articles on the chemistry and biology of free radicals as well as their involvement in human disease.
      Oxygen free-radical generation is considered both necessary and sufficient for adequate in vivo neutrophil-mediated microbicidal activity. For example, neutrophils from patients with chronic granulomatous disease fail to produce oxygen free radicals. These patients usually succumb to recurrent infections, most often caused by catalase-positive bacteria or fungi.
      • Malech HL
      • Gallin JI
      Neutrophils in human diseases.
      • Badwey JA
      • Karnovsky ML
      Active oxygen species and the functions of phagocytic leukocytes.
      The microbial catalase efficiently catabolizes any microbicidal hydrogen peroxide that may be generated in the phagocyte. Without hydrogen peroxide-mediated generation of either hypochlorite ion or toxic oxygen free radicals, cellular microbicidal activity is severely limited.
      Oxygen free radicals are potent destroyers of biologic molecules and tissues and are easily generated within neutrophils by efficient enzyme systems. When large numbers of activated neutrophils collect at an inflammatory site, such as occurs in ARDS, extracellular generation and spillover from intracellular generation ensue, results that overwhelm available antioxidant mechanisms and produce diffuse tissue damage.
      • Malech HL
      • Gallin JI
      Neutrophils in human diseases.
      • Gallin JI
      Neutrophil specific granules: a fuse that ignites the inflammatory response.

      IS MEDIATION OF NEUTROPHILS NECESSARY AND SUFFICIENT IN DIFFUSE ACUTE LUNG INJURY?

      Supporting Evidence.

      It has long been known that the lung functions as a major reservoir of marginated neutrophils.
      • Heinemann HO
      • Fishman AP
      Nonrespiratory functions of mammalian lung.
      As Andrewes
      • Andrewes FW
      The behaviour of the leucocytes in infection and immunity.
      stated in 1910, “… if the bone marrow is the birthplace of these cells [polymorphonuclear leukocytes] and the spleen their ultimate tomb, while the blood is their means of transit, the lung may serve as a week-end at the seaside, where they may recuperate their energies.” A steady state exists such that factors that increase or decrease pulmonary blood flow increase or decrease, respectively, the flux from the marginating to the circulating intravascular pool of granulocytes.
      • Thommasen HV
      The role of the polymorphonuclear leukocyte in the pathogenesis of the adult respiratory distress syndrome.
      Thommasen and associates,
      • Thommasen HV
      • Russell JA
      • Boyko WJ
      • Hogg JC
      Transient leucopenia associated with adult respiratory distress syndrome.
      in a study of 40 patients at high risk for the development of ARDS, suggested that frequent leukocyte counts in these patients might predict the development of ARDS. Of the 10 patients in whom ARDS developed, 8 were proved to have leukopenia by serial leukocyte counts. In contrast, only 4 of the remaining 30 patients in whom ARDS did not develop had leukopenia.
      In contrast, Fein and colleagues
      • Fein AM
      • Lippmann M
      • Holtzman H
      • Eliraz A
      • Goldberg SK
      The risk factors, incidence, and prognosis of ARDS following septicemia.
      compared the leukocyte counts in 116 patients with sepsis; ARDS developed in 21 of them. No significant difference in leukocyte counts was noted between those with and those without the development of ARDS. (The study by Thommasen and co-workers
      • Thommasen HV
      • Russell JA
      • Boyko WJ
      • Hogg JC
      Transient leucopenia associated with adult respiratory distress syndrome.
      also showed no correlation between the decrease in leukocyte counts and the development of ARDS in a subgroup of patients with sepsis.)
      Craddock and associates
      • Craddock PR
      • Fehr J
      • Brigham KL
      • Kronenberg RS
      • Jacob HS
      Complement and leukocyte-mediated pulmonary dysfunction in hemodialysis.
      demonstrated that lung dysfunction did not occur in a patient with agranulocytosis after hemodialysis. This outcome was in distinct contrast to the lung dysfunction that occurred in other nonneutropenic patients undergoing the same procedure.
      Clinical studies have been performed in an attempt to prove that pulmonary sequestration of neutrophils does occur in ARDS. For example, when lung scans with indium-111-labeled granulocytes were performed on a healthy volunteer and a patient with ARDS, more radioactivity was detected in the lung fields of the patient with ARDS than in the healthy volunteer, a finding that indicates pulmonary sequestration of neutrophils in the patient with ARDS.
      • Brigham KL
      Mechanisms of lung injury.
      Other evidence implicates neutrophil mediation of ARDS. Neutropenia has been shown to attenuate the diffuse, acute lung injury in dogs infused with glass bead microemboli,
      • Johnson A
      • Malik AB
      Effect of granulocytopenia on extravascular lung water content after microembolization.
      in hyperoxic-induced lung injury in guinea pigs,
      • Hosea S
      • Brown E
      • Hammer C
      • Frank M
      Role of complement activation in a model of adult respiratory distress syndrome.
      and in sheep with bacteremia.
      • Heflin Jr, AC
      • Brigham KL
      Prevention by granulocyte depletion of increased vascular permeability of sheep lung following endotoxemia.
      Also, accumulation of neutrophils in lung biopsy specimens or postmortem specimens from patients with ARDS is common but not universal.
      • Bachofen M
      • Weibel ER
      Structural alterations of lung parenchyma in the adult respiratory distress syndrome.
      • Zimmerman GA
      • Renzetti AD
      • Hill HR
      Functional and metabolic activity of granulocytes from patients with adult respiratory distress syndrome.
      Neutrophils, their secretory products, and chemotactic factors for neutrophils are present in increased numbers and concentration in the airways of patients with ARDS.
      • Rinaldo JE
      Mediation of ARDS by leukocytes: clinical evidence and implications for therapy.
      Weiland and colleagues
      • Weiland JE
      • Davis WB
      • Holter JF
      • Mohammed JR
      • Dorinsky PM
      • Gadek JE
      Lung neutrophils in the adult respiratory distress syndrome: clinical and pathophysiologic significance.
      analyzed pulmonary lavage fluid from 11 patients with ARDS and found that two-thirds of the cells recovered from the lavage fluid were neutrophils. Similarly, Fowler and co-workers
      • Fowler AA
      • Walchak S
      • Giclas PC
      • Henson PH
      • Hyers TM
      Characterization of antiproteinase activity in the adult respiratory distress syndrome.
      noted a mean of 92 to 95% neutrophils and 4 to 6% alveolar macrophages in ARDS lavage fluid.
      Lee and associates
      • Lee CT
      • Fein AM
      • Lippmann M
      • Holtzman H
      • Kimbel P
      • Weinbaum G
      Elastolytic activity in pulmonary lavage fluid from patients with adult respiratory-distress syndrome.
      detected high levels of neutrophil-derived elastase in pulmonary la vage fluid from 12 of 23 patients with ARDS. Concomitantly, α1-proteinase inhibitor activity was decreased in eight of nine patients with high elastolytic activity and ARDS, presumably through elastase-mediated inactivation of the antiprotease. (Oxygen free radicals could also mediate the inactivation.)
      Cochrane and colleagues
      • Cochrane CG
      • Spragg R
      • Revak SD
      Pathogenesis of the adult respiratory distress syndrome: evidence of oxidant activity in bronchoalveolar lavage fluid.
      noted that 15 of 17 bronchoalveolar lavage specimens from patients with ARDS showed a partial to total decrease in α1-proteinase inhibitor activity. They hypothesized that inactivation occurred by oxidation or proteolytic cleavage of the antiprotease or by complex formation with neutrophil-derived elastase.
      In contrast to these studies, however, Fowler and co-workers
      • Fowler AA
      • Walchak S
      • Giclas PC
      • Henson PH
      • Hyers TM
      Characterization of antiproteinase activity in the adult respiratory distress syndrome.
      noted that bronchoalveolar lavage fluid analyzed within 12 hours after the onset of ARDS contained large, not small, amounts of antiproteases. They hypothesized that only later in the course of ARDS might extensive neutrophilic autolysis occur and overwhelm the antiproteases with neutrophil-derived proteases.
      The relative numbers of neutrophils and macrophages in lavage fluid from healthy volunteers and from patients with ARDS argue for a specific neutrophil mediation in some cases of ARDS. More than 90% of the cells in lavage fluid from healthy volunteers are alveolar macrophages, with neutrophils a minor cellular constituent.
      • Fowler AA
      • Walchak S
      • Giclas PC
      • Henson PH
      • Hyers TM
      Characterization of antiproteinase activity in the adult respiratory distress syndrome.
      • Lee TH
      Interactions between alveolar macrophages, monocytes, and granulocytes: implications for airway inflammation.
      The reverse is often the case in lavage fluid from patients with ARDS.
      Although these bronchoalveolar lavage studies implicate the neutrophil as a primary mediator of pulmonary injury in ARDS, certain drawbacks are present. For instance, how accurately do cells in airspaces reflect inflammatory processes on the endothelial side of the alveolocapillary membrane? Is the recovery of neutrophils from airspaces specific for or predictive of ARDS? Hyers and associates
      • Hyers TM
      • Fowler AA
      • Stephenson AH
      • Dettenmeier PA
      • Fisher B
      • Webster RO
      The appearance of neutrophils and metabolites of arachidonic acid in bronchial fluid of patients at risk for ARDS (abstract).
      reported that patients predisposed to ARDS and those with ARDS showed a neutrophil influx into airspaces regardless of whether ARDS developed or not. Do secondary, superimposed pulmonary infections (which commonly occur in ARDS) cause neutrophils to enter airways, or does the presence of the neutrophil truly reflect a primary, neutrophil-mediated lung injury?
      • Bell RC
      • Coalson JJ
      • Smith JD
      • Johanson Jr, WG
      Multiple organ system failure and infection in adult respiratory distress syndrome.
      Control experiments with rigorous exclusion of infection in patients studied by bronchoalveolar lavage have not been published. Another consideration questions the effect of life-support measures such as mechanical ventilation and supplementary inspired oxygen on the results of bronchoalveolar lavage studies.
      The foregoing discussion suggests that the neutrophil may be only a marker of diffuse acute lung injury rather than a prominent mediator of such pulmonary injury.

      Refuting Evidence.

      Although substantial evidence is available from in vitro experiments, animal models, and in vivo human studies to support the idea that the neutrophil is a necessary and sufficient cellular mediator in the genesis of diffuse acute lung injury, other studies in the past several years have produced substantial evidence that discounts this basic tenet. Several in vivo human studies have shown that neutropenia does not prevent the development of ARDS in patients (Table 1).
      Table 1Occurrence of Adult Respiratory Distress Syndrome in Patients With Severe Neutropenia
      AML = acute myeloid leukemia; ARDS = adult respiratory distress syndrome; CML = chronic myeloid leukemia.
      ReferencePatients with ARDS (no.)Neutrophils (no./μl)Underlying disease
      Ognibene et al
      • Ognibene FP
      • Martin SE
      • Parker MM
      • Schlesinger T
      • Roach P
      • Burch C
      • Shelhamer JH
      • Parrillo JE
      Adult respiratory distress syndrome in patients with severe neutropenia.
      50–500Sepsis
      Braude et al
      • Braude S
      • Apperley J
      • Krausz T
      • Goldman JM
      • Royston D
      Adult respiratory distress syndrome after allogeneic bone-marrow transplantation: evidence for a neutrophil-independent mechanism.
      5100–1,900CML
      Maunder et al
      • Maunder RJ
      • Hackman RC
      • Riff E
      • Albert RK
      • Springmeyer SC
      Occurrence of the adult respiratory distress syndrome in neutropenic patients.
      47–75Hodgkin's disease, CML, aplastic anemia, AML
      Laufe et al
      • Laufe MD
      • Simon RH
      • Flint A
      • Keller JB
      Adult respiratory distress syndrome in neutropenic patients.
      30–910Bacteremia
      Rinaldo & Borovetz
      • Rinaldo JE
      • Borovetz H
      Deterioration of oxygenation and abnormal lung microvascular permeability during resolution of leukopenia in patients with diffuse lung injury.
      61,000Leukemia
      * AML = acute myeloid leukemia; ARDS = adult respiratory distress syndrome; CML = chronic myeloid leukemia.
      Ognibene and colleagues
      • Ognibene FP
      • Martin SE
      • Parker MM
      • Schlesinger T
      • Roach P
      • Burch C
      • Shelhamer JH
      • Parrillo JE
      Adult respiratory distress syndrome in patients with severe neutropenia.
      examined 11 patients who had severe neutropenia and ARDS precipitated by sepsis. Five of these 11 patients showed diffuse alveolar damage, a histopathologic characteristic of ARDS. A complete absence of neutrophils in both the vascular bed and the pulmonary interstitium was noted. After ruling out local pneumonitis, antibiotic therapy, and chemotherapeutic agents as primary causal factors for the histologic changes noted, the authors concluded “… that ARDS can occur in the setting of severe neutropenia, without pulmonary neutrophil infiltration.”
      Braude and co-workers
      • Braude S
      • Apperley J
      • Krausz T
      • Goldman JM
      • Royston D
      Adult respiratory distress syndrome after allogeneic bone-marrow transplantation: evidence for a neutrophil-independent mechanism.
      described five patients with chronic myeloid leukemia in whom ARDS developed subsequent to bone marrow transplantation. Histologic examination of lung tissue at autopsy on all patients showed no detectable intrapulmonary sequestration of neutrophils but diffuse alveolar damage, formation of hyaline membranes, intra-alveolar hemorrhage, and only mild interstitial inflammation.
      Maunder and associates
      • Maunder RJ
      • Hackman RC
      • Riff E
      • Albert RK
      • Springmeyer SC
      Occurrence of the adult respiratory distress syndrome in neutropenic patients.
      presented results on four patients with various hematologic diseases in whom ARDS developed while they had severe neutropenia. Histologic examination of lung tissue from these patients revealed 5 to 32 chloroacetate esterase-positive cells per 100 high-power fields (both granulocytes and mast cells are chloroacetate esterase-positive).
      • Yam LT
      • Li CY
      • Crosby WH
      Cytochemical identification of monocytes and granulocytes.
      Diffuse alveolar damage, epithelial degeneration, hyaline membranes, alveolar hemorrhage, and interstitial edema were noted in the lung biopsy specimens. The respiratory failure was not thought to be due to infection, radiation pneumonitis, or anti-neoplastic agent therapy.
      Laufe and colleagues
      • Laufe MD
      • Simon RH
      • Flint A
      • Keller JB
      Adult respiratory distress syndrome in neutropenic patients.
      studied three patients who had ARDS, bacteremia, and neutropenia. In addition to hyaline membranes, interstitial edema, and regenerative pneumatocytes, histologic sections of lung and bone marrow from two of the patients demonstrated an absence of neutrophils.
      Rinaldo and Borovetz
      • Rinaldo JE
      • Borovetz H
      Deterioration of oxygenation and abnormal lung microvascular permeability during resolution of leukopenia in patients with diffuse lung injury.
      studied six patients with leukemia who initially had acute respiratory failure and concurrent leukopenia attributable to chemotherapy-induced marrow aplasia but who eventually had resolution of their leukopenia. All patients showed diffuse alveolar infiltrates without cardiomegaly or evidence of congestive heart failure. Four of the six patients had clinical evidence of pulmonary dysfunction within 96 hours preceding the resolution of the leukopenia. The authors concluded that diffuse lung injury can occur in patients with leukopenia.

      OTHER PROPOSED FACTORS IN THE PATHOGENESIS OF ARDS

      Other factors that may be primary in the genesis of diffuse, acute lung injury include endothelial or alveolar epithelial cell damage, alveolar macrophages, disturbances in hemostasis, and loss of fibronectin.
      Evidence refuting the premise that the mere presence of neutrophils is sufficient to cause acute lung injury is available from experiments on endothelium and alveolar epithelium. A fundamental premise in these experiments is that intense surface contact, accomplished by spreading of neutrophils on susceptible endothelial or epithelial cells, is a prerequisite for activation of neutrophils or intense, neutrophil-mediated acute lung injury.
      Neutrophils layered on optimally adherent, closed monolayers of cultured endothelial cells remained loosely attached to the endothelium. They also exhibited minimal release of granules and hexose monophosphate shunt activity (necessary for oxygen free-radical generation). Only when the neutrophils were exposed to a plastic surface that stimulated them to adhere tightly and to spread on that surface did intense hexose monophosphate shunt activity and release of granules occur.
      • Fehr J
      • Moser R
      • Leppert D
      • Groscurth P
      Antiadhesive properties of biological surfaces are protective against stimulated granulocytes.
      A primary role for prerequisite spreading of neutrophils on cells in the pathogenesis of acute lung injury is also suggested by the discovery of a family of cell-surface glycoproteins designated as the CDw18 membrane antigen complex.
      • Nauseef WM
      • De Alarcon P
      • Bale JF
      • Clark RA
      Aberrant activation and regulation of the oxidative burst in neutrophils with Mo1 glycoprotein deficiency.
      One component of this complex, the iC3b receptor (CR3), functions as a receptor for the central complement component in the complement cascade, C3b. Functionally, this receptor enhances adherence and phagocytosis of opsonized microorganisms. It also facilitates adherence and spreading of the neutrophil to surfaces (Fig. 2). Decreased spontaneous movement, chemotaxis, and aggregation, decreased adherence and inefficient spreading on exposed surfaces such as endothelium, and inefficient phagocytosis of opsonized microorganisms characterize neutrophils from patients with iC3b receptor deficiency.
      • Malech HL
      • Gallin JI
      Neutrophils in human diseases.
      • Nauseef WM
      • De Alarcon P
      • Bale JF
      • Clark RA
      Aberrant activation and regulation of the oxidative burst in neutrophils with Mo1 glycoprotein deficiency.
      Clinically, these patients are prone to various severe, recurrent bacterial infections and have a depressed inflammatory response.
      Ismail and associates
      • Ismail G
      • Morganroth ML
      • Todd III, RF
      • Boxer LA
      Prevention of pulmonary injury in isolated perfused rat lungs by activated human neutrophils preincubated with anti-Mo1 monoclonal antibody.
      showed that incubation of neutrophils with anti-Mo1, a monoclonal antibody specifically directed against the iC3b receptor, significantly attenuated sequestration of neutrophils and lung injury in rats stimulated by phorbol myristate acetate. (This agent has been shown to cause oxygen free-radical-mediated acute lung injury.) Despite this, neutrophil degranulation and superoxide production occurred and thus were considered not to be factors in attenuation of the lung injury. Neither was the attenuation thought to be due to a nonspecific effect of the monoclonal antibody.
      Similarly, Simon and colleagues
      • Simon RH
      • DeHart PD
      • Todd III, RF
      Neutrophil-induced injury of rat pulmonary alveolar epithelial cells.
      noted that anti-Mo1-treated, activated neutrophils prevented spreading of neutrophils on alveolar epithelial cells and inhibited epithelial cell injury. Extensive contact between the neutrophils and epithelial cells seemed to be a requirement for injury. Furthermore, the alveolar epithelial cells were killed in an oxygen metabolite-independent manner by the stimulated neutrophils.
      Brigham and Meyrick
      • Brigham KL
      • Meyrick B
      Granulocyte-dependent injury of pulmonary endothelium: a case of miscommunication?.
      hypothesized that interactions of neutrophils with pulmonary vascular endothelium disrupt endothelial integrity only if endothelial cell structure or function was previously altered. Infusion of zymosan-activated plasma into sheep induced an early sequestration of neutrophils in lung capillaries and caused only mild endothelial injury. (Zymosan is a polysaccharide activator of complement.) Endothelial cell recovery occurred within 4 hours despite the fact that the neutrophils remained sequestered in the lung circulation. In contrast, infusion of endotoxin resulted in pulmonary sequestration of neutrophils with pronounced, persistent injury of endothelial cells. In other words, pulmonary endothelial cell injury was not an obligatory consequence of pulmonary sequestration of neutrophils. Indeed, even though stimulated granulocytes were intimately associated with endothelial cells, no structural evidence of endothelial cell injury or of increased permeability was noted. This concept is supported by studies published by Bachofen and Weibel.
      • Bachofen M
      • Weibel ER
      Alterations of the gas exchange apparatus in adult respiratory insufficiency associated with septicemia.
      Electron microscopic morphologic analysis of lungs from patients with ARDS showed leukocytes to be sequestered in the interstitium. The endothelial layer, however, was intact, and no gross damage of the pulmonary capillaries was detected.
      Perhaps endothelial damage by endotoxin, direct oxygen toxicity, or hypoxemia is necessary before neutrophils are capable of continuing their deleterious effects on the alveolocapillary membrane. For example, endotoxin in nanogram amounts directly injures bovine pulmonary artery endothelium, even in the absence of neutrophils or any other cell type.
      • Meyrick BO
      • Ryan US
      • Brigham KL
      Direct effects of E. coli endotoxin on structure and permeability of pulmonary endothelial monolayers and the endothelial layer of intimal explants.
      Acute lung injury in endotoxic shock may result from the combined effects of activated neutrophils and tumor necrosis factor (cachectin).
      • Tracey KJ
      • Beutler B
      • Lowry SF
      • Merryweather J
      • Wolpe S
      • Milsark IW
      • Hariri RJ
      • Fahey III, TJ
      • Zentella A
      • Albert JD
      • Shires GT
      • Cerami A
      Shock and tissue injury induced by recombinant human cachectin.
      One of the earliest and most characteristic lesions of acute oxygen toxicity is endothelial cell injury.
      • Kistler GS
      • Caldwell PRB
      • Weibel ER
      Development of fine structural damage to alveolar and capillary lining cells in oxygen-poisoned rat lungs.
      • Crapo JD
      • Barry BE
      • Foscue HA
      • Shelburne J
      Structural and biochemical changes in rat lungs occurring during exposures to lethal and adaptive doses of oxygen.
      Bowman and associates
      • Bowman CM
      • Butler EN
      • Repine JE
      Hyperoxia damages cultured endothelial cells causing increased neutrophil adherence.
      demonstrated that hyperoxia (95% O2) was directly toxic to cultured, bovine pulmonary artery endothelial cells. In addition, significantly more neutrophils adhered to these endothelial cells than to cells exposed to normoxic conditions. Bronchoalveolar lavage studies in humans have shown that short exposure times to hyperoxic conditions (several hours) are sufficient to induce “leaks” in the alveolocapillary membranes as well as to stimulate release of fibroblast proliferation factors from alveolar macrophages.
      • Davis WB
      • Rennard SI
      • Bitterman PB
      • Crystal RG
      Pulmonary oxygen toxicity: early reversible changes in human alveolar structures induced by hyperoxia.
      Shasby and co-workers
      • Shasby DM
      • Fox RB
      • Harada RN
      • Repine JE
      Reduction of the edema of acute hyperoxic lung injury by granulocyte depletion.
      demonstrated that the severity of lung injury in rabbits exposed to hyperoxic conditions for 72 hours was significantly correlated with the number of neutrophils in pulmonary lavage fluid and histologic lung sections. Inducing a sustained granulocytopenia, however, diminished the lung injury in the rabbits.
      Webster, Henson, and their associates
      • Webster RO
      • Larsen GL
      • Mitchell BC
      • Goins AJ
      • Henson PM
      Absence of inflammatory lung injury in rabbits challenged intravascularly with complement-derived chemotactic factors.
      • Henson PM
      • Larsen GL
      • Webster RO
      • Mitchell BC
      • Goins AJ
      • Henson JE
      Pulmonary microvascular alterations and injury induced by complement fragments: synergistic effect of complement activation, neutrophil sequestration, and prostaglandins.
      concluded that factors other than chemotactic agents were necessary to cause complement-mediated lung injury in rabbits. Even with surgical manipulation, anesthesia, and intubation, lung injury did not ensue. In rabbits that were intubated, infused with a complement activator, and subjected to transient hypoxia, however, severe acute lung injury occurred.
      The alveolar macrophage is capable of playing a primary role in pulmonary injury. It constitutes more than 90% of the total cells in normal airways, as evaluated by bronchoalveolar lavage. This cell is fully capable of phagocytosis, the respiratory burst in oxygen free-radical generation, degranulation, and synthesis and secretion of a wide variety of biomolecules including complement components, platelet-activating factors, interleukin 1, and arachidonic acid-derived metabolites.
      • Tate RM
      • Repine JE
      Neutrophils and the adult respiratory distress syndrome.
      • Duchateau J
      • Haas M
      • Schreyen H
      • Radoux L
      • Sprangers I
      • Noel FX
      • Braun M
      • Lamy M
      Complement activation in patients at risk of developing the adult respiratory distress syndrome.
      • Nathan CF
      • Murray HW
      • Cohn ZA
      The macrophage as an effector cell.
      Also, this cell is known to augment the oxidative activity of the neutrophil by secreting heat-stable neutrophil-activating factors.
      • Lee TH
      Interactions between alveolar macrophages, monocytes, and granulocytes: implications for airway inflammation.
      Although investigators are uncertain about whether intravascular coagulation and platelet aggregation are primary causes or simply the effects of diffuse, acute lung injury, some evidence supports the concept that disturbances in hemostasis may be the primary factor in the pathogenesis of diffuse, acute lung injury.
      • Turino GM
      • Rodriquez JR
      • Greenbaum LM
      • Mandl I
      Mechanisms of pulmonary injury.
      • Malik AB
      • Johnson A
      • Tahamont MV
      Mechanisms of lung vascular injury after intravascular coagulation.
      Thrombocytopenia occurs with a high incidence in patients with acute respiratory failure.
      • Schneider RC
      • Zapol WM
      • Carvalho AC
      Platelet consumption and sequestration in severe acute respiratory failure.
      Increased platelet consumption and pulmonary platelet sequestration have also been noted.
      • Hechtman HB
      • Lonergan EA
      • Shepro D
      Platelet and leukocyte lung interactions in patients with respiratory failure.
      Other studies, however, have noted the same incidence of thrombocytopenia in critically ill patients without ARDS as in patients with ARDS.
      • Thommasen HV
      • Russell JA
      • Boyko WJ
      • Hogg JC
      Transient leucopenia associated with adult respiratory distress syndrome.
      • Haynes JB
      • Hyers TM
      • Giclas PC
      • Franks JJ
      • Petty TL
      Elevated fibrin(ogen) degradation products in the adult respiratory distress syndrome.
      The fibrinogen degradation product D concentration is substantially increased in patients with ARDS in comparison with patients who have similar predisposing conditions but do not have pulmonary insufficiency. In addition, increased levels of this product are found in almost all severely traumatized or burned patients before the onset of pulmonary dysfunction. Infusion of this product into animals produces terminal severe respiratory distress.
      • Manwaring D
      • Thorning D
      • Curreri PW
      Mechanisms of acute pulmonary dysfunction induced by fibrinogen degradation product D.
      In contrast, neither administration of heparin nor depletion of platelets or coagulation factors altered steady-state pulmonary hemodynamics in sheep models, a result that suggests that intravascular coagulation does not play a major role in lung injury.
      • Binder AS
      • Nakahara K
      • Ohkuda K
      • Kageler W
      • Staub NC
      Effect of heparin or fibrinogen depletion on lung fluid balance in sheep after emboli.
      • Binder AS
      • Kageler W
      • Perel A
      • Flick MR
      • Staub NC
      Effect of platelet depletion on lung vascular permeability after microemboli in sheep.
      Disseminated intravascular coagulation has developed in some patients with ARDS, accompanied by profound thrombocytopenia and fibrin-platelet microthrombi evident histologically.
      • Bone RC
      • Francis PB
      • Pierce AK
      Intravascular coagulation associated with the adult respiratory distress syndrome.
      • El-Kassimi FA
      • Al-Mashhadani S
      • Abdullah AK
      • Akhtar J
      Adult respiratory distress syndrome and disseminated intravascular coagulation complicating heat stroke.
      (Even in some patients with ARDS who did not have disseminated intravascular coagulation, however, platelet counts decreased by at least 50% during the illness.) In contrast, Bachofen and Weibel
      • Bachofen M
      • Weibel ER
      Alterations of the gas exchange apparatus in adult respiratory insufficiency associated with septicemia.
      demonstrated by electron microscopic morphologic analysis that widespread disseminated intravascular coagulation was not present in lungs from patients who had died of ARDS.
      Blockade of the reticuloendothelial system may enhance formation of pulmonary edema as a result of decreased clearance of products from disseminated intravascular coagulation. This outcome, in turn, may be due to decreased serum fibronectin levels. Fibronectin is a potent, opsonic α2-glycoprotein whose serum level closely correlates with reticuloendothelial cell phagocytosis. Both endothelial cells and fibroblasts synthesize and secrete cell-surface fibronectin, which mediates various processes including cell spreading on surfaces and both intercellular and basement membrane adhesion. Proteolysis of cell-surface fibronectin by granular contents of activated neutrophils may initiate detachment of endothelial cells from one another, followed by an increase in microvascular permeability and interstitial edema.
      • Saba TM
      • Jaffe E
      Plasma fibronectin (opsonic glycoprotein): its synthesis by vascular endothelial cells and role in cardiopulmonary integrity after trauma as related to reticuloendothelial function.
      Lastly, the role of the neutrophil in diffuse, acute lung injury has important therapeutic implications. If neutrophils indeed mediate ARDS, administration of anti-inflammatory drugs is appropriate. If, however, the chief cause of death in patients with ARDS is superinfection or sepsis-related multiple organ failure rather than respiratory failure, anti-inflammatory agents may be detrimental because of suppression of the bactericidal functions of the neutrophil.

      CONCLUSION

      Several questions remain unsettled about the role of the neutrophil in diffuse, acute lung injury. Is the neutrophil a sufficient mediator of this injury? The answer is a qualified “yes.” The neutrophil possesses an adequate armamentarium for tissue destruction, and many chemoattractants exist to recruit neutrophils for this purpose. Although substantial, direct evidence does not yet exist, these mechanisms are probably operative in humans. They have been well proved in both in vitro and animal models. In addition, defects in one or several of these mechanisms in humans leads to disease.
      Is endothelial or epithelial damage a prerequisite for neutrophil-mediated acute lung injury? The “sufficiency” of neutrophil mediation probably is dependent on other cellular damage. The mere presence of activated neutrophils in the pulmonary vascular bed and interstitium does not signify tissue damage.
      Is the neutrophil a necessary mediator of acute lung injury? Evidence indicates that the answer is “perhaps not.” ARDS has been well substantiated in patients with severe neutropenia, and severe respiratory distress has been confirmed in neutropenic animal models. In addition, the alveolar macrophage is at least equal to the neutrophil in its potential to destroy tissue. The macrophage may be the primary mediator of lung injury in the setting of neutropenia. In addition, the role of the neutrophil and of the macrophage in acute lung injury may not be mutually exclusive. Perhaps the neutrophil initiates lung injury and the macrophage propagates it.
      Various clinically related questions about ARDS and TRALI also remain unsettled. For example, is TRALI a cause of ARDS? What differences exist in the course of TRALI to explain its relatively low morbidity and mortality in comparison with, for example, sepsis-related ARDS? Because of the variety of potential mediators of acute lung injury, is early, specific detection of acute lung injury feasible?
      Answers to these questions depend on continued in vitro, animal, and human experimentation and assessment. Having an animal model that mimics TRALI probably would make it possible to answer many questions about both its pathogenesis and treatment. Such a model would necessitate presensitization of a donor animal to specific alloantigens on transfused neutrophils or lymphocytes. Blood containing appropriate antibodies from this donor animal could then be transfused into a recipient animal.
      Clinical and anatomic data on patients with ARDS abound in comparison with the relative lack of data on TRALI. Continued clinical monitoring of patients with TRALI may identify as yet unknown major differences between TRALI and ARDS from other causes. In addition, data derived from gross, microscopic, and electron microscopic analyses of lungs from patients with TRALI may assist in this endeavor.
      Detection in plasma, lung secretions, or lung tissue of antibodies specific for several of the proposed mediators in acute lung injury would provide both qualitative and quantitative proof of their involvement. This experimental approach probably would be applicable to in vivo and postmortem studies. Ideally, identification of early markers of diffuse, acute lung injury would allow early attempts at prevention. Direct detection of free radicals is likely to be extremely difficult because their life spans are measured in fractions of a second. At best, one might attempt indirect quantitation of lipid peroxides or other products of free radical-mediated catabolism of lipids, nucleic acids, or proteins as proof of their role in acute lung injury.

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