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Abstract: Objective: To review the topic of alpha-1 antitrypsin (AAT) deficiency. Method: Narrative literature review. Results: Much work has been carried out on this condition with many questions being answered but still further questions remain. Discussion and conclusions: AAT deficiency is an autosomal co-dominantly inherited disease which affects the lungs and liver predominantly. The clinical manifestations, prevalence, genetics, molecular pathophysiology, screening and treatment recommendations are summarised in this review. [Copyright &y& Elsevier]

Summary: Objective: To review the topic of alpha-1 antitrypsin (AAT) deficiency. Method: Narrative literature review. Results: Much work has been carried out on this condition with many questions being answered but still further questions remain. Discussion and conclusions: AAT deficiency is an autosomal co-dominantly inherited disease which affects the lungs and liver predominantly. The clinical manifestations, prevalence, genetics, molecular pathophysiology, screening and treatment recommendations are summarised in this review. [Copyright &y& Elsevier]

Z α[sub1]-antitrypsin (ZAAT) deficiency is a disease associated with emphysematous lung disease and also with liver disease. The liver disease of AAT deficiency is associated with endoplasmic reticulum (ER) stress. SEPS1 is a selenoprotein that, through a chaperone activity, decreases ER stress. To determine the effect of SEPS1 on ER stress in ZAAT deficiency, we measured activity of the grp78 promoter and levels of active ATF6 as markers of the unfolded protein response in HepG2 cells transfected with the mutant form of AAT, a ZAAT transgene. We evaluated levels of NFκB activity as a marker of the ER overload response. To determine the effect of selenium supplementation on the function of SEPS1, we investigated glutathione peroxidase activity, grp78 promoter activity, and NF.B activity in the presence or absence of selenium. SEPS1 reduced levels of active ATF6. Overexpression of SEPS1 also inhibited grp78 promoter and NFκB activity, and this effect was enhanced in the presence of selenium supplementation. This finding demonstrates a role for SEPS1 in ZAAT deficiency and suggests a possible therapeutic potential for selenium supplementation. [ABSTRACT FROM AUTHOR]

Neutrophil Elastase (NE) is a pro-inflammatory protease present at higher than normal levels in the lung during inflammatory disease. NE regulates IL-8 production from airway epithelial cells and can activate both EGFR and TLR4. TACE/ADAM17 has been reported to trans-activate EGFR in response to NE. Here, using 16HBE14o-humanbronchial epithelial cells we demonstrate anew mechanism by which NE regulates both of these events. A high molecular weight soluble metalloprotease activity detectable only in supernatants from NE-treated cells by gelatin and casein zymography was confirmed to be meprin alpha by Western immunoblotting. In vitro studies demonstrated the ability of NE to activate meprin alpha, which in turn could release soluble TGFα and induce IL-8 production from 16HBE14o- cells. These effects were abrogated by actinonin, a specific meprin inhibitor. NE-induced IL-8 expression was also inhibited by meprin alpha siRNA. Immunoprecipitation studies detected EGFR/TLR4 complexes in NE-stimulated cells overexpressing these receptors. Confocal studies confirmed colocalization of EGFR and TLR4 in 16HBE14o-cells stimulated with meprin alpha. NFκB was also activated via MyD88 in these cells by meprin alpha. In bronchoalveolar lavage fluid from NE knock-out mice infected intra-tracheally with Pseudomonas aeruginosa meprin alpha was significantly decreased compared with control mice, and was significantly increased and correlated with NE activity, in bronchoalveolar lavage fluid from individuals with cystic fibrosis but not healthy controls. The data describe a previously unidentified lung metalloprotease meprin alpha, and its role in NE-induced EGFR and TLR4 activation and IL-8 production. [ABSTRACT FROM AUTHOR]

We have demonstrated that bronchoalveolar lavage fluid from chronic obstructive pulmonary disease patients contains higher levels of interferon-y compared with controls. Interferon-γ is a potent inducer of various cathepsins and matrix metalloproteases. Therefore, we postulated that interferon-y could induce protease expression by macrophages in acute and chronic lung disease. Chronic obstructive pulmonary disease patients had greater levels of cathepsin S and matrix metalloprotease-12 in their bronchoalveolar lavage fluid. Macrophages incubated with chronic obstructive pulmonary disease bronchoalveolar lavage fluid exhibited increased expression of cathepsin S and matrix metalloprotease-12, which was inhibited by the addition of interferon-γ-neutralizing immunoglobulin. Human secretory leukocyte protease inhibitor is an 11.7-kDa cationic non-glycosylated antiprotease synthesized and secreted by cells at the site of inflammation. We have demonstrated that secretory leukocyte protease inhibitor can inhibit interferon-γ-induced cathepsin S production by macrophages. Pretreatment of macrophages with secretory leukocyte protease inhibitor inhibited interferon-γ-induced inhibitor κB β degradation and activation of nuclear factor κB. Secretory leukocyte protease inhibitor may prove to be therapeutically important as a potential inhibitor of protease expression in chronic obstructive pulmonary disease. [ABSTRACT FROM AUTHOR]

Background Colonization by Aspergillus fumigatus in patients with cystic fibrosis (CF) can cause A fumigatus sensitization and/or allergic bronchopulmonary aspergillosis (ABPA), which affects pulmonary function and clinical outcomes. Recent studies show that specific allergens upregulate the surface-expressed basophil marker CD203c in sensitized subjects, a response that can be readily measured by using flow cytometry. Objective We sought to identify A fumigatus sensitization in patients with CF by using the basophil activation test (BAT). Methods Patients with CF attending Beaumont Hospital were screened for study inclusion. BAT was used to identify A fumigatus sensitization. Serologic (total and A fumigatus –specific IgE), pulmonary function, and body mass index measurements were performed. Results The BAT discriminates A fumigatus –sensitized from nonsensitized patients with CF. Persistent isolation of A fumigatus in sputum is a significant risk factor for A fumigatus sensitization. Levels of the A fumigatus –stimulated basophil activation marker CD203c inversely correlated with pulmonary function and body mass index in A fumigatus –sensitized but not nonsensitized patients with CF. Total and A fumigatus –specific IgE, but not IgG, levels are increased in A fumigatus –sensitized patients with CF and ABPA when compared with those in A fumigatus –sensitized and nonsensitized patients with CF without ABPA. Itraconazole treatment did not affect A fumigatus sensitization. Conclusion Combining the BAT with routine serologic testing allows classification of patients with CF into 3 groups: nonsensitized, A fumigatus –sensitized, and ABPA. Accurate and prompt identification of A fumigatus –associated clinical status might allow early and targeted therapeutic intervention, potentially improving clinical outcomes. [ABSTRACT FROM AUTHOR]

A high intrapulmonary protease burden is characteristic of cystic fibrosis (CF), and the resulting dysregulation of the protease/anti-protease balance has serious implications for inflammation in the CF lung. Because of this inflammation, micro-bleeds can occur releasing hemoglobin into the lung. The aim of this study was to investigate the effect of the protease-rich environment of the CF lung on human hemoglobin and to assess the proinflammatory effect of heme on CF bronchial epithelium. Here, we show that the Pseudomonas proteases (Pseudomonas elastase and alkaline protease) and the neutrophil proteases (neutrophil elastase (NE) and proteinase-3) are capable of almost complete degradation of hemoglobin in vitro but that NE is the predominant protease that cleaves hemoglobin in vivo in CF bronchoalveolar lavage fluid. One of the effects of this is the release of heme, and in this study we show that heme stimulates IL-8 and IL-10 protein production from ΔF508 CFBE41o- bronchial epithelial cells. In addition, heme-induced IL-8 expression utilizes a novel pathway involving meprin, EGF receptor, and MyD88. Meprin levels are elevated in CF cell lines and bronchial brushings, thus adding to the proinflammatory milieu. Interestingly, α1-antitrypsin, in addition to its ability to neutralize NE and protease-3, can also bind heme and neutralize heme-induced IL-8 from CFBE41o- cells. This study illustrates the proinflammatory effects of micro-bleeds in the CF lung, the process by which this occurs, and a potential therapeutic intervention. [ABSTRACT FROM AUTHOR]

The serine anti-protease elafin is expressed by monocytes, alveolar macrophages, neutrophils, and at mucosal surfaces and possesses antimicrobial activity. It is also known to reduce lipopolysaccharide-induced neutrophil influx into murine alveoli as well as to abrogate lipopolysaccharide-induced production of matrix metalloprotease 9, macrophage inhibitory protein 2, and tumor necrosis factor-α by as-yet unidentified mechanisms. In this report we have shown that elafin inhibits the lipopolysaccharide-induced production of monocyte chemoattractant protein-1 in monocytes by inhibiting AP-i and NP-κB activation. Elafin prevented lipopolysaccharide-induced phosphorylation of AP-1, c-Jun, and JNK but had no effect on phosphorylation of p38. The lipopolysaccharide-induced degradation of IL-1R-associated kinase 1, IκBα and IκBβ was inhibited by elafin but phosphorylation of IκBα was unaffected. Polyubiquitinated protein including polyubiquitinated IκBα was shown to accumulate in the presence of elafin. These results suggest that inhibition by elafin of lipopolysaccharide-induced AP-1 and NF-κB activation occurs via an effect on the ubiquitin-proteasome pathway. [ABSTRACT FROM AUTHOR]

A highly branched (1 → 3,1 → 6)-β-d-glucan was isolated from the microalga Isochrysis galbana Parke (Isochrysidales, Haptophyta). The polysaccharide structure was analyzed by methylation and Smith degradation, as well as by ESI and MALDI TOF mass spectrometry and NMR spectroscopy. The glucan was shown to contain a (1 → 6)-linked backbone, where every residue is substituted at position 3 by Glc, which in turn may be substituted at C-6 by a single Glc or by rather short (up to tetrasaccharide) oligosaccharide chains. All the 3-linked Glc residues are present in these side chains. In the biological activity experiments it was demonstrated that the polysaccharide directly inhibits the proliferation of U937 human leukemic monocyte lymphoma cells and therefore has potential anti-tumor activity. [ABSTRACT FROM AUTHOR]

Abstract: Long non-coding RNAs (lncRNAs) have emerged recently as key regulatory molecules with diverse roles at almost every level of the regulation of gene expression. The roles of these RNAs in the pathogenesis of cystic fibrosis (CF); a lethal multisystem, autosomal recessive disorder have yet to be explored. Our aim was to examine the expression profile of lncRNA, in the airway epithelium of people with CF. We examined the expression of 30,586 lncRNAs by microarray (Human LncRNA Array v3.0, Arraystar, Inc.), in vivo in bronchial cells isolated from endobronchial brushings obtained from CF and non-CF individuals. In total, we identified 1,063 lncRNAs with differential expression between CF and non-CF individuals (fold change ≥3, p ≤0.001). The microarray also contained probes for ∼26,109 protein coding transcripts, of which 720 were differentially expressed between CF and non-CF brush samples (fold change ≥3, p ≤0.001). Confirmation of a selection of differentially expressed coding mRNA and lncRNA transcripts such as XIST and TLR8 was achieved using qRT-PCR. Gene ontology bioinformatics analysis highlighted that many processes over-represented in the CF bronchial epithelium are related to inflammation. These data show a significantly altered lncRNA and mRNA expression profile in CF bronchial cells in vivo. Dysregulation of some of these lncRNAs may play important roles in the chronic infection and inflammation that exists in the lungs of people with CF. This article is part of a directed issue entitled: Cystic fibrosis: from o-mics to cell biology, physiology, and therapeutic advances. [Copyright &y& Elsevier]

Elafin is a neutrophil serine protease inhibitor expressed in lung and displaying anti-inflammatory and anti-bacterial properties. Previous studies demonstrated that some innate host defense molecules of the cystic fibrosis (CF) and chronic obstructive pulmonary disease airways are impaired due to increased proteolytic degradation observed during lung inflammation. In light of these findings, we thus focused on the status of elafin in CF lung. We showed in the present study that elafin is cleaved in sputum from individuals with CF. Pseudomonas aeruginosa-positive CF sputum, which was found to contain lower elafin levels and higher neutrophil elastase (NE) activity compared with P. aeruginosa-negative samples, was particularly effective in cleaving recombinant elafin. NE plays a pivotal role in the process as only NE inhibitors are able to inhibit elafin degradation. Further in vitro studies demonstrated that incubation of recombinant elafin with excess of NE leads to the rapid cleavage of the inhibitor. Two cleavage sites were identified at the N-terminal extremity of elafin (Val-5—Lys-6 and Val-9—Ser-10). Interestingly, purified fragments of the inhibitor (Lys-6—Gln-57 and Ser-10—Gln-57) were shown to still be active for inhibiting NE. However, NE in excess was shown to strongly diminish the ability of elafin to bind lipopolysaccharide (LPS) and its capacity to be immobilized by transglutamination. In conclusion, this study provides evidence that elafin is cleaved by its cognate enzyme NE present at excessive concentration in CF sputum and that P. aeruginosa infection promotes this effect. Such cleavage may have repercussions on the innate immune function of elafin. [ABSTRACT FROM AUTHOR]