Secretory phospholipases A2 stimulate mucus secretion, induce airway inflammation, and produce secretory hyperresponsiveness to neutrophil elastase in ferret trachea

2007 ◽  
Vol 292 (1) ◽  
pp. L62-L67 ◽  
Author(s):  
Kosuke Okamoto ◽  
Jung-Soo Kim ◽  
Bruce K. Rubin
Keyword(s):  
Airway Inflammation ◽  
Neutrophil Elastase ◽  
Lavage Fluid ◽  
Water Soluble ◽  
Mucus Secretion ◽  
Human Neutrophil Elastase ◽  
Lipoxygenase Inhibitor ◽  
Phospholipases A2

Secretory phospholipases A2 (sPLA2) are increased in the bronchoalveolar lavage fluid of patients with asthma and acute respiratory distress syndrome. Intratracheal sPLA2 instillation induces acute lung injury in the rat and guinea pig. We hypothesized that sPLA2 would stimulate mucus secretion in vitro and that intratracheal sPLA2 exposure would induce mucus hypersecretion and airway inflammation in the ferret trachea in vivo. In vitro, porcine pancreatic sPLA2 at a concentration of 0.5 or 5 U/ml significantly increased mucous glycoconjugate (MG) secretion from the excised ferret trachea. P-bromophenacylbromide (a sPLA2 inhibitor), quercetin (a lipoxygenase inhibitor), or MK-886 (a 5-lipoxygenase inhibitor), each at 10−4 M, significantly reduced sPLA2-induced MG secretion. sPLA2-stimulated MG secretion was decreased in Ca2+-free medium. In vivo, ferrets were intubated for 30 min once per day for 3 days using an ETT coated with 20 units of porcine pancreatic sPLA2 mixed in water-soluble jelly. Constitutive MG secretion increased 1 day after sPLA2 exposure and returned to control 5 days later. Human neutrophil elastase (HNE) at 10−8 M increased MG secretion in the sPLA2-exposed trachea compared with that in the control trachea, but methacholine at 10−7 M did not. sPLA2-induced secretory hyperresponsiveness continued for at least 5 days after sPLA2 exposure ended. sPLA2 increased tracheal inflammation, MG secretion, and secretory hyperresponsiveness to HNE probably through enzymatic action rather than by activation of its receptor.

Peptidyl .alpha.-ketoheterocyclic inhibitors of human neutrophil elastase. 3. In vitro and in vivo potency of a series of peptidyl .alpha.-ketobenzoxazoles

1995 ◽  
Vol 38 (20) ◽  
pp. 3972-3982 ◽  
Author(s):  
Philip D. Edwards ◽  
Mark A. Zottola ◽  
Matthew Davis ◽  
Joseph Williams ◽  
Paul A. Tuthill
Keyword(s):  
Neutrophil Elastase ◽  
Human Neutrophil ◽  
Human Neutrophil Elastase

scholarly journals Novel and Modified Neutrophil Elastase Inhibitor Loaded in Topical Formulations for Psoriasis Management

Pharmaceutics ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 358 ◽  
Author(s):  
Andreia Nunes ◽  
Joana Marto ◽  
Lídia Maria Gonçalves ◽  
Sandra Simões ◽  
Rita Félix ◽  
...  
Keyword(s):  
Serine Protease ◽  
Neutrophil Elastase ◽  
In Vitro Cytotoxicity ◽  
Human Neutrophil Elastase ◽  
Therapeutic Effects ◽  
Neutrophil Elastase Inhibitor ◽  
Analytical Methodology ◽  
Elastase Inhibitor

Human neutrophil elastase (HNE) is a serine protease that degrades matrix proteins. An excess of HNE may trigger several pathological conditions, such as psoriasis. In this work, we aimed to synthesize, characterize and formulate new HNE inhibitors with a 4-oxo-β-lactam scaffold with less toxicity, as well as therapeutic index in a psoriasis context. HNE inhibitors with 4-oxo-β-lactam scaffolds were synthesized and characterized by NMR, FTIR, melting point, mass spectrometry and elemental analysis. In vitro cytotoxicity and serine protease assays were performed. The compound with the highest cell viability (AAN-16) was selected to be incorporated in an emulsion (AAN-16 E) and in a microemulsion (AAN-16 ME). Formulations were characterized in terms of organoleptic properties, pH, rheology, droplet size distribution, in vitro drug release and in vivo psoriatic activity. All compounds were successfully synthesized according to analytical methodology, with good yields. Both formulations presented suitable physicochemical properties. AAN-16 E presented the most promising therapeutic effects in a murine model of psoriasis. Overall, new HNE inhibitors were synthesized with high and selective activity and incorporated into topical emulsions with potential to treat psoriasis.

scholarly journals Effects of Amoxicillin, Gentamicin, and Moxifloxacin on the Hemolytic Activity of Staphylococcus aureus In Vitro and In Vivo

2001 ◽  
Vol 45 (1) ◽  
pp. 196-202 ◽  
Author(s):  
Dieter Worlitzsch ◽  
Hayal Kaygin ◽  
Andrea Steinhuber ◽  
Axel Dalhoff ◽  
Konrad Botzenhart ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Neutrophil Elastase ◽  
Human Neutrophil Elastase ◽  
Mssa Strain ◽  
Subinhibitory Concentrations ◽  
High Level ◽  
Strain Dependent ◽  
Negative Mutant

ABSTRACT In Staphylococcus aureus infection hemolysis caused by the extracellular protein α-toxin encoded by hla is thought to contribute significantly to its multifactorial virulence. In vitro, subinhibitory concentrations of β-lactam antibiotics and fluoroquinolones increase the levels of hla and α-toxin expression, whereas aminoglycosides decrease the levels ofhla and α-toxin expression. In the present study we investigated the effects of subinhibitory concentrations of amoxicillin, gentamicin, and moxifloxacin on hla and α-toxin expression and total hemolysis of S. aureusstrain 8325-4, a high-level α-toxin producer, and its α-toxin-negative mutant, DU 1090, in vitro and in a rat model of chronic S. aureus infection. The levels of expression ofhla and α-toxin and total hemolysis did not differ significantly when amoxicillin, gentamicin, or moxifloxacin was added to cultures of S. aureus strain 8325-4. In vivo, strain 8325-4 induced a significantly increased level of hemolysis in infected pouches compared to that in uninfected control pouches, but the hemolysis was reduced to control levels by treatment with doses of amoxicillin, gentamicin, or moxifloxacin that reduced bacterial numbers by 2 orders of magnitude. Additionally, the effects of subinhibitory concentrations of the three antibiotics on total hemolysis of four methicillin-resistant S. aureus and three methicillin-sensitive S. aureus (MSSA) clinical isolates were assessed in vitro. A significant increase in total hemolysis was observed for only one MSSA strain when it was treated with amoxicillin but not when it was treated with moxifloxacin or gentamicin. When purified α-toxin was incubated with purified human neutrophil elastase, α-toxin was cleaved nearly completely. The results suggest that the penicillin-induced increases in S. aureusα-toxin expression are strain dependent, that reduction of bacterial numbers in vivo counteracts this phenomenon effectively, and finally, that in localized S. aureus infections α-toxin activity is controlled by neutrophil elastase.

Oxidation-resistant variants of recombinant anti-leucoprotease are better inhibitors of human-neutrophil-elastase-induced emphysema in hamsters than natural recombinant antileucoprotease

1991 ◽  
Vol 81 (6) ◽  
pp. 777-784 ◽  
Author(s):  
A. Rudolphus ◽  
R. Heinzel-Wieland ◽  
V. A. M. M. Vincent ◽  
D. Saunders ◽  
G. J. Steffens ◽  
...  
Keyword(s):  
Neutrophil Elastase ◽  
Human Neutrophil ◽  
Site Directed Mutagenesis ◽  
Human Neutrophil Elastase ◽  
Polymorphonuclear Leucocytes ◽  
Inhibitory Effects ◽  
Oxidative Inactivation ◽  
Methionine Residues

1. Antileucoprotease, being sensitive to oxidative inactivation, can be produced by recombinant techniques. Via site-directed mutagenesis, two mutants of recombinant antileucoprotease were produced in which one or more of the oxidation-sensitive methionine residues were replaced by leucine: in rALP242, methionine-73 was replaced by leucine, and in rALP231, leucine was substituted for four methionine residues. In vitro, native antileucoprotease and the recombinant antileucoprotease preparations have similar inhibitory characteristics towards human neutrophil elastase. We hypothesized that replacement of methionine residues in the antileucoprotease molecule would result in a reduced oxidation sensitivity of the mutants. 2. After incubation of recombinant antileucoprotease and its mutants with increasing dosages of cis-platinum(II)diammine dichloride, we observed that native antileucoprotease and recombinant antileucoprotease were inactivated by this reagent to the same extent. Compared with this, rALP242 was less inactivated, whereas the inhibitory capacity of rALP231 was not influenced by cis-platinum(II)diammine dichloride at all. 3. After incubation of recombinant antileucoprotease, rALP242 and rALP231 with triggered polymorphonuclear leucocytes, which are thought to produce an excess of oxidants, we measured residual inhibitory activities towards human neutrophil elastase of 10%, 55% and 87%, respectively. 4. In vivo, the inhibitory effects of intratracheally administered rALP242 and rALP231 towards human-neutrophil-elastase-induced emphysema were significantly greater than that of recombinant antileucoprotease. There were no significant differences between the mutants. With respect to secretory cell metaplasia and haemorrhage, rALP231 tended to be a better inhibitor than recombinant antileucoprotease and rALP242. 5. We conclude that the recombinant antileucoprotease mutants are less sensitive to oxidation and consequently inhibit human-neutrophil-elastase-induced emphysema to a greater extent than recombinant antileucoprotease.

Scutellarin Attenuates Human-Neutrophil-Elastase-Induced Mucus Production by Inhibiting the PKC-ERK Signaling Pathway in Vitro and in Vivo

2011 ◽  
Vol 39 (06) ◽  
pp. 1193-1206 ◽  
Author(s):  
De-Peng Jiang ◽  
Qi Li ◽  
Jie Yang ◽  
Juliy M. Perelman ◽  
Victor P. Kolosov ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Neutrophil Elastase ◽  
Human Neutrophil ◽  
Erk Signaling ◽  
Control Group ◽  
Human Neutrophil Elastase ◽  
Dependent Manner ◽  
Mucus Production

The aim of this study was to investigate the influence of scutellarin on mucus production induced by human neutrophil elastase (HNE) and the possible in vitro and in vivo mechanisms. To this purpose, cells were incubated with saline, scutellarin or gefitinib for 60 min and exposed to 0.1 μM HNE for 24 h. After being pretreated respectively with saline, scutellarin or gefitinib, rats were challenged intratracheally with HNE by means of nebulization for 30 days. The expression of mucin (MUC) 5AC, protein kinase C (PKC), and extracellular signal-regulated kinase 1/2 (ERK1/2) was assessed by ELISA, RT-PCR or Western blotting. The results showed that scutellarin inhibited MUC5AC mRNA and protein expressions induced by HNE in a concentration-dependent manner in vitro. In the in vivo model, scutellarin significantly attenuated MUC5AC mRNA expression and goblet cell hyperplasia in rats treated with HNE for 30 days, as well as decreased the phosporylation of PKC and ERK1/2 compared to the HNE control group. Therefore, our study showed that scutellarin could prevent mucus hypersecretion by inhibiting the PKC-ERK signaling pathway. Inhalation scutellarin may be valuable in the treatment of chronic inflammatory lung disease.

Induction of ferroptosis-like cell death of eosinophils exerts synergistic effects with glucocorticoids in allergic airway inflammation

Thorax ◽  
2020 ◽  
Vol 75 (11) ◽  
pp. 918-927 ◽  
Author(s):  
Yanping Wu ◽  
Haixia Chen ◽  
Nanxia Xuan ◽  
Lingren Zhou ◽  
Yinfang Wu ◽  
...  
Keyword(s):  
Cell Death ◽  
Synergistic Effect ◽  
Airway Inflammation ◽  
Synergistic Effects ◽  
Morphological Characteristics ◽  
Allergic Airway Inflammation ◽  
Lavage Fluid ◽  
Allergic Disorders

IntroductionEosinophils are critical in allergic disorders, and promoting eosinophil death effectively attenuates allergic airway inflammation. Ferroptosis is a recently described novel form of cell death; however, little is known about ferroptosis in eosinophils and related diseases. This study aimed to investigate the effects of ferroptosis-inducing agents (FINs) on eosinophil death and allergic airway inflammation, and to explore their potential synergistic effect with glucocorticoids (GCs).MethodsEosinophils isolated from the peripheral blood of humans or mice were incubated with FINs, and eosinophil ferroptosis was assessed. The in vivo effects of FINs alone or in combination with dexamethasone (DXMS) were examined in a mouse model of allergic airway inflammation. Bronchoalveolar lavage fluid and lung tissue were collected to examine airway inflammation.ResultsTreatment with FINs time and dose dependency induced cell death in human and mouse eosinophils. Interestingly, FINs induced non-canonical ferroptosis in eosinophils, which generated morphological characteristics unique to ferroptosis and was iron dependent but was independent of lipid peroxidation. The antioxidants glutathione and N-acetylcysteine significantly attenuated FIN-induced cell death. Treatment with FINs triggered eosinophil death in vivo and eventually relieved eosinophilic airway inflammation in mice. Furthermore, FINs exerted a synergistic effect with DXMS to induce eosinophil death in vitro and to alleviate allergic airway inflammation in vivo.ConclusionsFINs induced ferroptosis-like cell death of eosinophils, suggesting their use as a promising therapeutic strategy for eosinophilic airway inflammation, especially due to the advantage of their synergy with GCs in the treatment of allergic disorders.

IDENTIFICATION OF A NEUTROPHIL ELASTASE CLEAVAGE SITE ON THE Act -CHAIN OF PRIMATE FIBRINOGEN

1987 ◽  
Author(s):  
J Weitz ◽  
S Landman ◽  
S Birken
Keyword(s):  
Neutrophil Elastase ◽  
Cleavage Site ◽  
High Performance ◽  
Human Neutrophil Elastase ◽  
Amino Acid Sequence Analysis ◽  
Specific Radioimmunoassay ◽  
Human Thrombin ◽  
The Mean

Human neutrophil elastase (HNE) cleaves the Aα21-22 bond of fibrinogen thus releasing the fibrinopeptide A (FPA)-containing fragment Aαl-21. Plasma Aal-21 levels reflect in vivo HNE activity and peptide levels are increased in cigarette smokers and patients with chronic lung disease. To further explore the HNE-fibrinogen interaction, we set out to develop an animal model. The digestion of purified baboon and marmoset fibrinogen by human thrombin, HNE and extracts of baboon and marmoset neutrophils was monitored with a specific radioimmunoassay for human FPA. Thrcmbin produced quantitative release (2 mol/mol fibrinogen) of FPA. In contrast, HNE and the neutrophil extracts did not release FPA, but rather, produced quantitative release of a larger, FPA-containing fragment. Immunochemically, this fragment was clearly distinguishable from FPA in that in vitro thrombin treatment increased its immunoreactivity 1,000-fold (thrombin increasable FPA or TIFPA). TIFPA release by the neutrophil extracts was blocked by α1-proteinase inhibitor, a specific HNE inhibitor (MeO-Suc-Ala2-Pro-ValCH2Cl) and an anti-HNE IgG, indicating that elastase was the responsible proteinase and that there was homology between the human and primate enzymes. The products of HNE and neutrophil extract proteolysis of the primate fibrinogens were then separated by high performance liquid chromatography and the TIFPA-containing fractions were subjected to amino acid sequence analysis. The FPA-containing fragments each consisted of 21 amino acids, had minor substitutions when compared with human A α] -21 [Baboon: Aα(3) Ser - Thr; Marmoset Aα(l) Ala - Thr, Aα(3) Ser - Thr, Aα(ll) Glu - Ala], and exhibited complete crossreactivity with the human peptide. Using the TIFPA assay, there was good recovery of primate or human Aαl-21 added to primate blood and the mean peptide level in 8 healthy marmosets was similar to that in man (0.5 nM and 0.4 nM, respectively). In conclusion, (1) the Aα;21 -22 bond of baboon and marmoset fibrinogen is a cleavage site for human and primate elastase, (2) baboon and marmoset Aal-21 can be measured with the assay for the human peptide, and (3) the primate serves as a useful model for the study of elastase-fibrinogen interactions.

scholarly journals Effects of human neutrophil elastase (HNE) on neutrophil function in vitro and in inflamed microvessels

Blood ◽  
1993 ◽  
Vol 82 (7) ◽  
pp. 2188-2195 ◽  
Author(s):  
RC Woodman ◽  
PH Reinhardt ◽  
S Kanwar ◽  
FL Johnston ◽  
P Kubes
Keyword(s):  
Neutrophil Elastase ◽  
Human Neutrophil ◽  
Neutrophil Migration ◽  
Specific Inhibitor ◽  
Neutrophil Infiltration ◽  
Cathepsin G ◽  
Neutrophil Adhesion ◽  
Human Neutrophil Elastase

Abstract The primary objective of this study was to test the hypothesis that human neutrophil elastase (HNE) affects neutrophil infiltration (adhesion and emigration) into inflamed vessels. To determine whether HNE contributes to neutrophil adhesion in vivo, intravital microscopy was used to study neutrophil-endothelial cell interactions in single inflamed postcapillary venules. Superfusion of platelet-activating factor (PAF) (100 nmol/L) onto the mesentery caused an increase in neutrophil-neutrophil interactions, neutrophil adhesion to postcapillary venules, and cellular emigration out of the vasculature. Both L658 758 (an elastase-specific inhibitor), and Eglin C (an elastase and cathepsin G inhibitor) significantly attenuated all of these parameters in vivo. To further characterize the mechanism(s) involved, various in vitro parameters were assessed. HNE, but not trypsin, caused a dose-dependent (0.01 to 1.0 microgram/mL) increase in the expression of the beta subunit (CD18) of the CD11/CD18 adhesive glycoprotein complex on neutrophils. An HNE-dependent increase in CD11b expression was also observed; however, HNE did not affect the expression of other neutrophil adhesion molecules (L-selectin), superoxide production, or degranulation. PAF-enhanced CD18 expression on neutrophils and neutrophil migration were both abolished by L658 758 but PAF-induced neutrophil adhesion to endothelial monolayers was not affected by the antiproteinase. The in vitro data suggest that the antiproteinases do not directly prevent neutrophil adhesion in vivo but may be important in other CD18-dependent events such as neutrophil- neutrophil interaction or neutrophil infiltration (chemotaxis). These results translate into an important, rate-limiting role for elastase in the process of leukocyte infiltration and accumulation in inflamed microvessels.

Further Sesquiterpene Lactones from Viguiera robusta and the Potential Anti-Inflammatory Activity of a Heliangolide: Inhibition of Human Neutrophil Elastase Release

2008 ◽  
Vol 63 (7-8) ◽  
pp. 533-538 ◽  
Author(s):  
Nilton S. Arakawa ◽  
Karin Schorr ◽  
Sérgio R. Ambrósio ◽  
Irmgard Merfort ◽  
Fernando B. Da Costa
Keyword(s):  
Neutrophil Elastase ◽  
Human Neutrophil ◽  
Sesquiterpene Lactones ◽  
Human Neutrophil Elastase ◽  
Reference Drug ◽  
Aggregation Factor ◽  
Anti Inflammatory ◽  
Isolated Compound

In addition to known heliangolides, a new eudesmanolide was isolated from the leaf rinse extract of Viguiera robusta (Asteraceae). Structural elucidation was based on spectral analysis. It is the first report on eudesmanolides in members of the subgenus Calanticaria of Viguiera. In this work, the main isolated compound, the furanoheliangolide budlein A, besides its previously reported in vitro and in vivo anti-inflammatory activities, inhibited human neutrophil elastase release. The inhibition was at the concentration of (16.83± 1.96) μm for formylated bacterial tripeptide (fMLP) stimulation and (11.84±1.62) μm for platelet aggregation factor (PAF) stimulation, being slightly less active than the reference drug parthenolide. The results are important to demonstrate the potential anti-inflammatory activities of sesquiterpene lactones and corroborate the previous studies using other targets.

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