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Background: The role of the innate immune system in tubal implantation remains undefined. This study compared expression of two key mediators of innate immunity, secretory leukocyte protease inhibitor (SLPI) and elafin, in the uterine decidua of women with intrauterine and tubal pregnancies.Methods: Uterine decidua was collected from women (18-45 years) undergoing surgical termination of pregnancy (n = 7), surgical management of spontaneous abortion (n = 6) and tubal pregnancy (n = 10). Using quantitative RT-PCR and immunohistochemistry, mRNA and protein expression patterns of SLPI and elafin were compared.Results: Relative SLPI mRNA expression was significantly higher in decidua of women with tubal pregnancy (12.37 +/- 2.66) compared with spontaneous abortion (5.09 +/- 2.22, P < 0.0185). There was no difference demonstrated in elafin mRNA expression. SLPI and elafin protein expression were demonstrated in the decidual leukocyte populations and epithelium. There was no obvious qualitative difference in levels of SLPI and elafin protein expression or their distribution in the uterine decidua of women with termination of pregnancy, spontaneous abortion or tubal pregnancy.Conclusions: Herein we demonstrate novel differences in gene expression of uterine decidua of tubal pregnancy compared with spontaneous abortion thereby contributing further to current knowledge of mechanisms involved in extrauterine implantation. The altered expression of SLPI may be a consequence of, or predispose to, tubal pregnancy. [ABSTRACT FROM AUTHOR]

Recent evidence shows that human neutrophil elastase inhibitors can be synthesized locally at mucosal sites. In addition to efficiently targeting bacterial and host enzymes, they can be released in the interstitium and in the lumen of mucosa, where they have been shown to have antimicrobial activities, and to activate innate immune responses. This review will address more particularly the pleiotropic functions of low-molecular-mass neutrophil elastase inhibitors [SLPI (secretory leucocyte proteinase inhibitor) and elafin] and, more specifically, their role in the development of the adaptive immune response.

Staphylococcus aureus is a pathogen often found in pneumonia and sepsis. In the context of the resistance of this organism to conventional antibiotics, an understanding of the regulation of natural endogenous antimicrobial molecules is of paramount importance. Previous studies have shown that both human and mouse airways express a variety of these molecules, including defensins, cathelicidins, and the four-disulfide core protein secretory leukocyte protease inhibitor. We demonstrate here by culturing mouse tracheal epithelial cells at an air-liquid interface that, despite the production of Defb1, Defb14, and Defr1 in this system, these cells are unable to clear S. aureus when exposed to this respiratory pathogen. Using an adenovirus (Ad)-mediated gene transfer strategy, we show that overexpression of elafin, an anti-elastase/antimicrobial molecule (also a member of the four-disulfide core protein family), dramatically improves the clearance of S. aureus. In addition, we also demonstrate that this overexpression is efficient in vivo and that intratracheal instillation of Ad-elafin significantly reduced the lung bacterial load and demonstrates concomitant anti-inflammatory activity by reducing neutrophil numbers and markers of lung inflammation, such as bronchoalveolar lavage levels of tumor necrosis factor and myeloperoxidase. These findings show that an increased antimicrobial activity phenotype is provided by the elafin molecule and have implications for its use in S. aureus-associated local and systemic infections.

Antimicrobial molecules are ancient and essential small cationic molecules of the host defence system which are found in a wide variety of species. They display antimicrobial activity against a wide range of bacteria, fungi and viruses, an activity that has been mostly attributed to the disruption of microbial membranes. In this article, we will review the "classical" functions of 3 classes of antimicrobial molecules, namely defensins, cathelicidins, and the four-disulfide core proteins secretory leukocyte proteinase inhibitor (SLPI) and elafin. In addition to the study of their expression in a variety of cell types and the regulation of their production, we will also describe novel properties of these molecules that have been highlighted by recent studies. These include their ability to chemoattract a variety of inflammatory, immune and other cell types (neutrophils, macrophages, monocytes, lymphocytes, mast cells, epithelial cells) in vitro and in vivo. In addition, we will discuss the potential use of these newly discovered properties for therapeutic or vaccination purposes, using protein- or gene-transfer based methodologies. Finally, we will examine in an extensive fashion the strategies used by microorganisms to circumvent and subvert host defence mechanisms, such as the modifications of cell membranes and walls, the secretion of inactivating proteins and proteases and the down-regulation of expression of antimicrobial molecules. Increased understanding of the mechanisms used by both the host and the microbes to 'win the battle' may ultimately lead to new therapeutic strategies aimed to treat infectious diseases.

Annexins are widely distributed and have been described in lung as well as in other cells and tissues. Annexin I (ANX AI) is a member of the calcium-dependent phospholipid binding protein family. Besides its anti-inflammatory function, ANX AI has been involved in several mechanisms such as the Erk repression pathway or apoptosis. To investigate the role of ANX AI on apoptosis in broncho-alveolar cells, we have constructed a plasmid containing the ANX AI full length cDNA. Transfected BZR cells displayed a higher level of both forms of ANX AI (37 and 33 kDa) as well as a decrease in cell viability (two-fold versus cells transfected with an empty vector). In order to analyse the endogenous ANX AI processing during stimulus-induced apoptosis, BZR cells were treated with a commonly used inducer, i.e. C2 ceramides. In these conditions, microscopic analysis revealed chromatin condensation in dying cells and the Bcl-2, Bcl-x(L)/Bax mRNA balance was altered. Caspase-3 is one of the key executioners of apoptosis, being responsible for the cleavage of many proteins such as the nuclear enzyme poly(ADP-ribose) polymerase (PARP). We demonstrate that caspase-3 was activated after 4 h treatment in the presence of ceramide leading to the cleavage of PARP. Dose-response experiments revealed that cell morphology and viability modifications following ceramide treatment were accompanied by an increase in endogenous ANX AI processing. Interestingly, in both ceramide and transfection experiments, the ANX AI cleaved form was enhanced whereas pre-treatment with the caspase inhibitor Z-VAD-fmk abolished ANX AI cleavage. In conclusion, this study demonstrates a complex regulatory role of caspase-dependent apoptosis where ANX AI is processed at the N-terminal region which could give susceptibility to apoptosis upon ceramide treatment.

The control of lung inflammation is of paramount importance in a variety of acute pathologies, such as pneumonia, the acute respiratory distress syndrome, and sepsis. It is becoming increasingly apparent that local innate immune responses in the lung are negatively influenced by systemic inflammation. This is thought to be due to a local deficit in cytokine responses by alveolar macrophages and neutrophils following systemic bacterial infection and the development of a septic response. Recently, using an adenovirus-based strategy which overexpresses the human elastase inhibitor elafin locally in the lung, we showed that elafin is able to prime lung innate immune responses. In this study, we generated a novel transgenic mouse strain expressing human elafin and studied its response to bacterial lipopolysaccharide (LPS) when the LPS was administered locally in the lungs and systemically. When LPS was delivered to the lungs, we found that mice expressing elafin had lower serum-to-bronchoalveolar lavage ratios of proinflammatory cytokines, including tumor necrosis factor alpha (TNF-alpha), macrophage inflammatory protein 2, and monocyte chemoattractant protein 1, than wild-type mice. There was a concomitant increase in inflammatory cell influx, showing that there was potential priming of innate responses in the lungs. When LPS was given systemically, the mice expressing elafin had reduced levels of serum TNF-alpha compared to the levels in wild-type mice. These results indicate that elafin may have a dual function, promoting up-regulation of local lung innate immunity while simultaneously down-regulating potentially unwanted systemic inflammatory responses in the circulation.

Chronic obstructive pulmonary disease (COPD) is a common cause of morbidity and mortality. The term is heterogenous and encompasses a number of distinct but often overlapping phenotypes including chronic bronchitis, small airways obstruction, emphysema and in some individuals, a systemic component. Although there have been significant advances in understanding the pathophysiology of COPD, understanding of the role of the inflammation in the pathogenesis of the condition remains in its infancy. Indeed, cytokines that are known to orchestrate the inflammatory response in asthma and other inflammatory diseases are only beginning to be reported in COPD. In this review, we highlight the potential role of cytokines in the development of mucus hypersecretion observed in chronic bronchitis and the morphological changes observed in the small airways and airspaces contributing to the development of airflow limitation and respiratory failure respectively. We report evidence that exacerbations are linked to increased expression of pro-inflammatory cytokines and that the wasting and skeletal muscle dysfunction observed in some patients is most probably related to the presence of a systemic inflammatory response. In addition transgenic and gene therapy technology has been used to explore the temporal and co-ordinated role of cytokines in the development of COPD animal models. Enhanced understanding of the events involved in the pathogenesis of COPD will lead to the development of therapy with potential to modify the observed progressive decline in lung function and impact on the development of the illness.

Low-molecular-mass neutrophil elastase inhibitors have been shown to be important in the control of lung inflammation. In addition to inhibiting the enzyme neutrophil elastase, these low-molecular-mass compounds (10 kDa) have been shown to have other activities. For example, secretory leucocyte proteinase inhibitor (SLPI) and elastase-specific inhibitor/SKALP (skin-derived antileucoproteinase)/elafin have also been shown to have "defensin"-like antimicrobial activities. Indeed, these inhibitors have antimicrobial properties in vitro against bacteria, fungi and, potentially, HIV. In addition, we have shown, using an adenovirus-mediated gene transfer overexpression strategy, that elafin is also active against Pseudomonas aeruginosa infection in mice in vivo. The mechanism of action is currently under investigation. In addition to these direct or indirect effects on microbes, it has been shown that lipopolysaccharide is able to up-regulate SPLI production in macrophages in vitro, and that the addition of recombinant SLPI to human monocytes or the transfection of macrophages with SPLI can down-regulate pro-inflammatory mediators such as tumour necrosis factor, presumably to limit self-damaging excessive inflammation. Using viral gene transfer vectors, we are currently investigating the potential of these inhibitors in various models of inflammation in vivo.

During acute pulmonary infection, tissue injury may be secondary to the effects of bacterial products or to the effects of the host inflammatory response. An attractive strategy for tissue protection in this setting would combine antimicrobial activity with inhibition of human neutrophil elastase (HNE), a key effector of neutrophil-mediated tissue injury. We postulated that genetic augmentation of elafin (an endogenous inhibitor of HNE with intrinsic antimicrobial activity) could protect the lung against acute inflammatory injury without detriment to host defense. A replication-deficient adenovirus encoding elafin cDNA significantly protected A549 cells against the injurious effects of both HNE and whole activated human neutrophils in vitro. Intratracheal replication-deficient adenovirus encoding elafin cDNA significantly protected murine lungs against injury mediated by Pseudomonas aeruginosa in vivo. Genetic augmentation of elafin therefore has the capacity to protect the lung against the injurious effects of both bacterial pathogens resistant to conventional antibiotics and activated neutrophils.

Lipopolysaccharide (LPS) is a mediator of inflammatory lung injury. Selective augmentation of host defense molecules such as elafin (an elastase inhibitor with antimicrobial activity) at the onset of pulmonary inflammation is an attractive potential therapeutic strategy. The aim of this study was to determine whether elafin expression could be induced by LPS administered after transfection with adenovirus (Ad) encoding human elafin downstream of the murine cytomegalovirus (CMV) promoter (known to be potentially responsive to LPS). In addition, we aimed to determine the effect of local elafin augmentation on neutrophil migration to the lung. LPS significantly up-regulated elafin expression from pulmonary epithelial cells transfected with Ad-elafin in vitro. In murine airways expression of human elafin was achieved using doses low enough (3 x 10(7) plaque forming units) to circumvent overt vector-induced inflammation. LPS significantly up-regulated human elafin secretion in murine airways treated with Ad-elafin [117 ng/ml in bronchoalveolar lavage fluid (BALF) after LPS administration, 5.9 ng/ml after PBS, p < 0.01)]. Over-expression of elafin significantly augmented LPS-mediated neutrophil migration into the airways in vivo (1.30 x 10(6) neutrophils in BALF after Ad-elafin/LPS treatment, 0.54 x 10(6) after Ad-lacZ/LPS (p < 0.05), 0.63 x 10(6) after PBS/LPS (p < 0.05)) and significantly enhanced human neutrophil migration in vitro. These data suggest novel functions for elafin in neutrophil migration, and that judicious selection of promoters may allow single, low-dose adenoviral administration to effect inflammation-specific expression of potentially therapeutic transgenes.

The rapidly expanding field of antimicrobial peptides is one that is attracting increasing interest from research groups around the world. The importance of antimicrobial agents in providing alternatives to conventional antibiotics has been highlighted in recent years by the emergence of a number of multidrug-resistant pathogens, such as methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE). Indeed, bacteria refractory to treatment by all known antibiotics are now a reality and the need for developing novel antimicrobial agents is urgent. This meeting brought together researchers working in a number of varied, but ultimately related areas. The functional diversity and putative mechanisms of action of antimicrobial peptides were discussed in depth, along with recent developments in the design of synthetic peptides with enhanced antimicrobial properties. Several ongoing studies were described, ranging from research into cystic fibrosis to work in the food industry. It was emphasized that cationic antimicrobial peptides have a range of properties to offer the world of scientific research and may play an important role in the ongoing battle against pathogenic microorganisms. Oral presentation sessions of the conference were co-chaired by Dr Deirdre A Devine (University of Leeds, UK) and Dr David G Smith (University of Edinburgh, UK).

The proteinase-antiproteinase hypothesis still receives support from clinical and experimental observations in a range of inflammatory lung diseases. The function of these molecules appears to be broader than originally believed and further research is likely to lead to an improved understanding of their role in the regulation of both the beneficial and detrimental effects in inflammatory response and the maintenance of the homeostasis in the normal lung. Thus the potential for the development as therapeutic tools is likely to become more attractive as improved drug development and delivery mechanisms appear.

Secretory leukocyte proteinase inhibitor and elafin are two low-molecular-mass elastase inhibitors that are mainly synthesized locally at mucosal sites. It is thought that their physicochemical properties allow them to efficiently inhibit target enzymes, such as neutrophil elastase, released into the interstitium. Historically, in the lung, these inhibitors were first purified from secretions of patients with chronic obstructive pulmonary disease and cystic fibrosis. This suggested that they might be important in controlling excessive neutrophil elastase release in these pathologies. They are upregulated by 'alarm signals' such as bacterial lipopolysaccharides, and cytokines such as interleukin-1 and tumor necrosis factor and have been shown to be active against Gram-positive and Gram-negative bacteria, so that they have joined the growing list of antimicrobial 'defensin-like' peptides produced by the lung. Their site of synthesis and presumed functions make them very attractive candidates as potential therapeutic agents under conditions in which the excessive release of elastase by neutrophils might be detrimental. Because of its natural tropism for the lung, the use of adenovirus-mediated gene transfer is extremely promising in such applications.

Inappropriate release of proteases from inflammatory and stromal cells can lead to destruction of the lung parenchyma. Antiproteinases such as alpha-1-proteinase inhibitor (alpha1-Pi), secretory leukocyte proteinase inhibitor (SLPI) and elastase-specific inhibitor (elafin) control excess production of human neutrophil elastase. In the present study, the concentrations of alpha1-Pi, SLPI and elafin found in bronchoalveolar lavage (BAL) fluid from control subjects, patients at risk of developing acute respiratory distress syndrome (ARDS) and patients with established ARDS were determined. Levels of all three inhibitors were raised in patients compared with normal subjects. SLPI was increased in the group of patients who were at risk of ARDS and went on to develop the condition, compared with the "at-risk" group who did not progress to ARDS (p=0.0083). Alpha1-Pi and elafin levels were similar in these two populations. In patients with established ARDS, both alpha1-Pi and SLPI levels were significantly increased, compared to patients at risk of ARDS who did (p=0.0089) or did not (p=0.0003) progress to ARDS. The finding of increased antiproteinases shortly before the development of acute respiratory distress syndrome provide further evidence for enhanced inflammation prior to clinical disease.

The lung is a readily accessible target organ for gene therapy. To date, therapeutic gene delivery has largely focused on introducing functional, corrective genes in lung diseases arising from single gene defects such as cystic fibrosis. More recently interest has centred on gene therapy as a potential therapeutic tool in modulating complex pathological processes such as pulmonary inflammation. Genetic modification of critical components of the inflammatory process may be beneficial-for example, overexpressing anti-elastase genes may circumvent elastase mediated lung damage in emphysema. With the development of improved viral and liposome vectors and the evolution of effective adjuvant immunosuppression to obviate host immune responses--for example, using selective cytokines and blockers of T cell surface activation--the potential exists to target therapeutic doses of transgene to deficient or dysregulated cells. Furthermore, increased understanding of tissue-specific promoter regions and of mechanisms controlling regulation of gene expression offer the potential for close control of therapeutic gene expression within the lung. Continuing refinements in these technologies will provide new therapeutic strategies in inflammatory lung disease.

Alpha-1 proteinase inhibitor (A1-Pi) is the main serine proteinase inhibitor found in human plasma and is a potent elastase inhibitor in various tissues, including lung. A1-Pi is expressed and induced in liver during inflammatory responses but can also be produced by epithelial cells. Since hepatocyte A1-Pi production is stimulated by interleukin-6 (IL-6) and other gp130-cytokines, such as leukemia inhibitory factor (LIF) and oncostatin M (OM), we investigated the role of these cytokines in regulating A1-Pi in lung epithelial cells. We show that OM, a monocyte and T cell product, can specifically and potently induce A1-Pi production in lung-derived A549 alveolar (epithelial) cells, as well as in liver-derived HepG2 cells. Both A1-Pi protein (as detected by ELISA and Western blots) and mRNA levels were enhanced 20-fold to 30-fold in A549 cells. OM was also able to stimulate the expression of tissue inhibitor of metalloproteinase-1 in these cells. Interestingly, other members of the IL-6 family (IL-6 and LIF) had little or no effect on A549 cells, and proinflammatory cytokines, such as IL-1 beta and tumor necrosis factor-alpha (TNF-alpha) also had no stimulatory effect on A1-Pi synthesis in A549 cells. Costimulation with IL-1 beta resulted in a decrease in A1-Pi production from OM-stimulated A549 cells. However, IL-6 production was synergistically enhanced. OM was also able to stimulate A1-Pi production from a bronchial epithelial primary cell line, whereas an intestinal epithelial cell line HT29 responded to IL-6 but not OM. These results suggest that lung levels A1-Pi could be derived not only from liver and inflammatory cells but also from epithelial cells, which can be upregulated on stimulation by OM. This may have implications for regulation of local activity of human neutrophil elastase (HNE) in such diseases as emphysema and cystic fibrosis.

Secretory leukocyte proteinase inhibitor (SLPI) is the main neutrophil elastase (HLE) inhibitor found in the upper airways during pulmonary inflammation. It has been shown to be synthesized and secreted in vitro by epithelial cells and has been localized in tracheal glands and bronchiolar epithelial cells by immunocytochemistry. In this study, using immunodetection and immunopurification techniques with specific anti-SLPI immunoglobulin G (IgG), we show that SLPI is present as a native 14-kDa molecule in neutrophil cytosol. In addition, we demonstrate that SLPI is the major inhibitor of HLE present in neutrophil cytosol because pre-incubation with specific anti-SLPI IgG was able to inhibit completely the anti-HLE activity of the cytosol. SLPI can be secreted (probably in an inactive form) by neutrophils and its secretion is enhanced when the cells are stimulated with phorbol myristate acetate (PMA). Elafin, an elastase-specific inhibitor, is also present in minute amounts in neutrophil cytosol and its secretion can be up-regulated. The presence of SLPI in the cytosol of neutrophils may serve as a protective screen against proteinases spilling from azurophilic granules. An alternative or supplementary role may be the maintenance of a differentiated phenotype.

Using adenoviral-mediated gene transfer techniques, the murine granulocyte-macrophage colony-stimulating factor (GM-CSF) transgene is efficiently targeted to and highly expressed by the respiratory epithelium of rat lung. This lung tissue-directed expression of GM-CSF induces accumulation of both eosinophils and macrophages at early stages and an irreversible fibrotic reaction at later stages. These tissue responses to GM-CSF appear to be distinct from those induced by other proinflammatory cytokines, interleukin (IL)-5, IL-6, macrophage inflammatory protein-2 (MIP-2), or RANTES overexpressed in the lung. These findings clearly demonstrate that GM-CSF is more than a hematopoietic cytokine in the lung and may play a pivotal role in the multiple pathological processes underlying numerous respiratory illnesses, including asthma. In this overview, the differences in tissue responses induced by GM-CSF and other individual cytokines are highlighted. In addition, the mechanisms by which GM-CSF and other individual cytokines are highlighted. In addition, the mechanisms by which GM-CSF contributes to the development of eosinophilia, macrophage granuloma, and fibrosis are discussed in conjunction with the recent findings from us and others.

Human bronchial mucous secretions have been shown to contain inhibitors of serine proteinases secreted by neutrophils. The role of these inhibitors is probably to control the enzymes secreted in the airways and in the lung interstitium. Three of these inhibitors have been identified and characterized: alpha 1-proteinase inhibitor, mucus proteinase inhibitor, and elafin. The elafin molecule, a 6.0 kD inhibitor of serine proteinases shows homology with mucus proteinase inhibitor. We recently isolated both molecules in bronchial secretions. In this report, we present evidence for the existence of a precursor of the elafin molecule. We have cloned and sequenced the gene for this precursor and show that it is composed of three exons. The coding information for a 117 amino acid precursor protein of elafin (inclusive of the signal peptide) is contained in the first two exons. This was confirmed at the mRNA and protein levels. By Northern Blot analysis we detected a 800 bp long product, and by immunoaffinity we detected in sputum and in cultured epithelial cell supernatant (NCI-H322 cell line) a 12 kD protein species cross-reacting with anti-elafin IgG. The finding of possible cross-linking function for the precursor in addition to its antiproteinase activity indicates a possible role for this molecule as a cross-linker agent in the extracellular matrix.

The regulation of proteinases secreted by neutrophils is very important for the prevention of tissue injury. We recently described the isolation of elafin from bronchial secretions, a new elastase-specific inhibitor that is also found in the skin of patients with psoriasis. In this study, we investigated the secretion of elafin and mucus proteinase inhibitor (MPI), another inhibitor showing sequence similarity with elafin, in two lung carcinoma cell lines, NCI-H322 and A549, which have features of Clara cells and type II alveolar cells, respectively. The results presented show that the two inhibitors are produced when the cells are cultured either in serum-free or in serum-containing media. MPI was detected immunologically as a unique molecule of M(r) 14 kD, in accordance with previous studies. Conversely, one or two elafin-immunoreactive species were detected depending on the cell line: a 12- to 14-kD species was observed in the A549 cell line, regardless of the culture conditions, whereas in the NCI-H322 cell line we detected a 6-kD species in serum-containing (10% fetal calf serum) conditions and a 12- to 14-kD species in serum-free conditions. The 12- to 14-kD molecule probably represents an active precursor of elafin. Whether the cleavage of the 12- to 14-kD precursor giving rise to the elafin molecule is of any physiologic significance is not known. In showing for the first time that MPI and elafin (and its precursor) are secreted by the A549 cell line, this report implicates the type II alveolar cell in the defense of the peripheral lung against the neutrophil elastase secreted during inflammation.

The calcium-binding proteins calgranulins A and B co-purified with an elastase-specific inhibitor after the affinity and cation-exchange chromatography of the perchloric acid-soluble fraction of pooled sputum collected from patients with chronic obstructive pulmonary disease (Sallenave, J.-M. and Ryle, A.P. (1991) Biol. Chem. Hoppe-Seyler 372, 13-21). The calgranulins were separated from the inhibitor by reverse-phase FPLC. Protein blot analysis of the calgranulin fraction in the absence of reducing agent revealed a band of 25 kDa corresponding to the disulphide-bonded heterodimerization of the two monomer components. Similar results were obtained from the immunoprecipitation and protein blot analysis of plasma, granulocytes and cultured epithelial cells. This implies that the calgranulins exist in the heterodimeric form in secretions in vivo. Their association with pancreatic elastase during the affinity chromatography stage of purification implicates them in the tissue destruction elicited by the inflammatory response in chronic obstructive pulmonary diseases.

Evidence is presented that the elastase-specific inhibitor of Mr 2500 (Sallenave, J.-M. & Ryle, A.P. (1991) Biol. Chem. Hoppe-Seyler 372, 13-21) is a biologically active fragment of a larger molecule described in the skin of patients with psoriasis (Wiedow, O., Shroder, J.-M., Gregory, H., Young, J.A. & Christophers, E. (1990) J. Biol. Chem. 265, 14791-14795) which the authors called elafin. We also describe the purification of the complete elafin molecule from bronchial secretions from a patient suffering from bronchial carcinoma, thus showing that the elafin, like the mucus proteinase inhibitor (MPI), is not of single origin but is probably a marker of inflammation (chronic obstructive pulmonary diseases, psoriasis...) present in different tissues.

Elastase-specific inhibitor (ESI) was purified from sputum of patients with chronic bronchitis and compared with mucus proteinase inhibitor (MPI, BrI) isolated, without the use of affinity chromatography on an enzyme, from non-purulent sputum of a patient with bronchial carcinoma. The N-terminal sequence of 27 residues of the latter was determined and showed serine as the only N-terminus. The partial N-terminal amino-acid sequence of ESI shows some homology with MPI, especially around the reactive site of MPI for human neutrophil elastase. This region could therefore be the reactive site of ESI. The thermodynamic and kinetic constants of the reactions of ESI with human neutrophil elastase and with porcine pancreatic elastase show that ESI is a fast-acting inhibitor.

A high-Mr (Mr 750,000) alpha 1-macroglobulin, obtained from Salamandra salamandra, is described. Salamander alpha 1-macroglobulin is composed of two monomers of equal Mr, which are composed of two polypeptide chains, each of Mr 180,000, linked by disulfide bonds. The molecular parameters of this protein, its binding to trypsin and inactivation by methylamine suggest that salamander alpha 1-macroglobulin is closely related to human alpha 2-macroglobulin and to other related proteins described in the animal kingdom.