Cleavage of C3 by Neutral Proteases from Granulocytes in Pleural Empyema

Abstract

The possibility of direct inactivation of C3 by granular enzymes from polymorphonuclear leukocytes (PMNLs) in pleural empyema was examined. As a group, pleural empyema from 10 patients with purulent effusions and a positive bacteriologic culture cleaved significantly more 125I-labeled C3 bound to Sepharose (18.4070 ± 7.3070) than did 19 sterile pleural effusions (2.4070 ± 0.9070; P« 0.001) and sonicates from bacterial strains commonly found in empyema (1.4070 ± 0.2070). Granular enzymes from 7 x 10 6 PMNLs cleaved 78.5070 of 125I-labeled C3 bound to Sepharose. When proteolysis of 125I-labeled C3 after incubation with pleural empyema or PMNL granular enzymes was examined with polyacrylamide gel electrophoresis, breakdown products were similar. Granulocyte elastase-like activity was detected in four samples of pleural empyema. Granulocyte elastase inhibitors, as well as 10070 human serum, effectively suppressed cleavage of C3 and elastase-like activity. In pleural empyemas, granular enzymes from PMNLs, especially elastase, apparently contribute to low complement-mediated opsonic activity by direct inactivation of C3.Pleural empyema is a closed-space infection defined by the simultaneous presence of large numbers of live microorganisms and polymorphonuclear leukocytes (PMNLs). The disease is further characterized by its chronic evolution and its poor response to medical and sometimes even surgical therapy.Work from our laboratory [1] has shown that the persistence of live microorganisms despite the presence of large numbers of PMNLs in pleural empyemas can be partially explained by a marked decrease in complement-mediated opsonic activity in culture-positive pleural effusions. This deficiency in opsonic activity was further shown to be accompanied by the local generation of increased amounts of C3d, the inactive breakdown product of C3.Serum complement is of major importance in the humoral immune response to pyogenic infections, and C3 is currently viewed as the target molecule of both the classical and the alternative pathways of complement activation [1][2][3]. Its acReceived for publication March 10, 1981, and in revised form June 22, 1981. This work was supported in part by grant no. 3.836-0.79 from the Swiss National Research Foundation.We thank Drs. M. Baggiolini and B. Dewald for advice, suggestions, and revision of the manuscript and F. Michaud for secretarial assistance.Please address requests for reprints to Dr. F. A. Waldvogel, Department of Medicine, University Hospital, 121I Geneva 4, Switzerland.499 tivation leads to the production of the biologically active fragment C3b, and the coating of microorganisms with C3b promotes their recognition by PMNLs. Activation proceeds through the cleavage of native C3 by either the classical (C42) or the alternative (C3b, Bb) pathway convertase of C3 or both [2,3]. Recently it has been shown in vitro [4][5][6][7][8] that C3, as well as C5, can be split by complement-unrelated enzymes, such as purified neutral proteases from PMNLs. These reports have shown that the...

Pathogenesis of Foreign Body Infection: Description and Characteristics of an Animal Model

“…Either decreased opsonization of S. aureus might be a consequence of the imbalance between the rate of coating of bacteria with functionally active C3 and the logarithmic growth of the organisms, or cleavage of the opsonically active factor by fJ-I-H-globulin, C3b inactivator, and proteases [47][48][49][50][51] could have occurred during the development of a purulent infection. Loss of opsonic activity, as well as proteolytic cleavage of C3, has been recently demonstrated in several exudates rich in PMNLs [13,15,24]. Finally, our experiments showed that PMNLs attracted into the infected tissue cages displayed normal phagocytic bactericidal function, a finding which is apparently in contradiction with our observations in sterile tissue cages.…”

“…Although polymorphonuclear leukocytes (PMNLs) accumulating around a foreign body appear microscopically normal, their functional status as well as the opsonizing capacity of the surrounding fluid have never been explored because of the lack of an appropriate animal model. In the present communication we describe the development of an animal model that reproduces the clinical characteristics of a foreign body infection and thus allows the in vitro evaluation of the various phagocytic steps in its neighborhood [13][14][15][16].…”

Granulocyte Neutral Proteases and Pseudomonas Elastase as Possible Causes of Airway Damage in Patients with Cystic Fibrosis

“…Bronchial lavage fluid was obtained from six patients by bronchoscopy on the same day that the sputum samples were obtained. Pathogens isolated from sputum included: Haemophilus influenzae [18], Streptococcus pneumoniae [8], Escherichia coli [1], Proteus vulgaris [1], S aureus [5], and Aspergillus [1]. Three patients had mixed infections.…”

“…Granular enzymes were prepared from PMNs collected from fresh heparinized blood of normal adult volunteers, according to the method of Baggiolini [21] and modified as previously described [8]. They had an elastolytic activity of 17.4 UE and granulocyte elastase-like activity of 0.59 UTGP (see below for definition of units).…”

“…These enzymes are indeed able to destroy important structural proteins of the lung and its airways such as elastin, collagen, and proteoglycans [4,5] in vitro; in addition, they can inactivate important opsonins such as complement component C3 [6][7][8] and IgG and IgM immunoglobulins [9][10][11]. It is also well established that they are released extracellularly during phagocytosis [12].…”