Effects of alpha 1-antitrypsin on endotoxin-induced lung inflammation in vivo.
ABSTRACT Previous in vitro experiments demonstrated that acute-phase protein, alpha 1-antitrypsin (AAT), could act either as an enhancer or as a suppressor of lipopolysaccharide (LPS)-induced cell activation depending on treatment time. Here we investigate how AAT regulates inflammatory responses in the short term when administrated post LPS challenge.
Similar experimental setup was used both in vitro and in vivo: human monocytes and neutrophils were stimulated with LPS for 2 h followed by AAT for a total time of 4 h, and C57BL/6 mice were treated intranasally with LPS and 2 h later with AAT and sacrificed after 4 h. Bronchial lavage (BAL) and lung homogenates were analyzed using bio-plex cytokine assay. BAL cell counts were assessed.
Within 4 h, AAT enhanced LPS-induced tumor necrosis factor-alpha (TNFalpha), interleukin (IL)-6, and IL-8 release from monocytes and neutrophils. Mice challenged for 4 h with LPS followed by AAT at 2 h showed no changes in BAL cell counts and higher levels of almost all measured cytokines, specifically RANTES in BAL and IL-12, IL-13, granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), and IL-10 levels in lung homogenates, than in mice treated with LPS only.
Within the short term, AAT enhances the magnitude of LPS-induced specific cytokine/chemokine production, which may play an important role in amplification and resolution of acute-phase inflammatory reactions in vivo.
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ABSTRACT: Background. Severe bacterial infection can cause sepsis, multiple organ dysfunction syndrome (MODS), and death. Human α1-antitrypsin (hAAT) is an antiinflammatory, immune-modulating, and tissue-protective circulating serine-protease inhibitor, with levels that increase during acute-phase responses. It is currently being evaluated as a therapeutic agent for individuals with diabetes and graft-versus-host disease. However, the concern of opportunistic bacterial infections has yet to be addressed. Therefore, we investigated host immune cell responses during acute bacterial infections under conditions of elevated hAAT levels. Methods. Peritonitis and sepsis models were created using wild-type mice and hAAT-transgenic mice. Bacterial loads, MODS, leukopenia, neutrophil infiltration, immune cell activation, circulating cytokine levels, and survival rates were then assessed. Results. hAAT significantly reduced infection-induced leukopenia and liver, pancreas, and lung injury, and it significantly improved 24-hour survival rates. Unexpectedly, bacterial load was reduced. Levels of early proinflammatory mediators and neutrophil influx were increased by hAAT soon after infection but not during sterile peritonitis. Conclusions. hAAT reduces the bacterial burden after infection. Since hAAT does not block bacterial growth in culture, its effects might rely on host immune cell modulation. These outcomes suggest that prolonged hAAT treatment in patients without hAAT deficiency is safe. Additionally, hAAT treatment may be considered a preemptive therapeutic measure for individuals who are at risk for bacterial infections.The Journal of Infectious Diseases 11/2014; 211(9). DOI:10.1093/infdis/jiu620 · 5.78 Impact Factor
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ABSTRACT: One would assume that the anti-inflammatory activity of α1-antitrypsin (AAT) is the result of inhibiting neutrophil enzymes. Yet, AAT exhibits tolerogenic activities that are difficult to explain by serine-protease inhibition, nor by reduced inflammatory parameters. Targets outside the serine-protease family have been identified, supporting the notion that elastase inhibition, the only functional factory release criteria for clinical-grade AAT, is overemphasized. Non-obvious developments in the understanding of AAT biology disqualify it from being a straightforward anti-inflammatory agent: AAT does not block dendritic cell activities, nor does it promote viral and tumor susceptibilities, stunt B-lymphocyte responses or render treated patients susceptible to infections; accordingly, outcomes of elevated AAT do not overlap those attained by immunosuppression. Aside from the acute phase response, AAT rises during the third trimester of pregnancy and also in advanced age. At the molecular level, AAT docks onto cholesterol-rich lipid-rafts and circulating lipid particles, directly binds IL-8, ADAM17 and danger-associated molecular pattern (DAMP) molecules, and its activity is lost to smoke, high glucose levels and bacterial proteases, introducing a novel entity – ‘relative AAT deficiency’. Unlike immunosuppression, AAT appears to help the immune system distinguish between desired responses against authentic threats, and unwanted responses fueled by a positive feedback loop perpetuated by, and at the expense of, inflamed injured innocent bystander cells. With a remarkable clinical safety record, AAT treatment is currently tested in clinical trials for its potential benefit in a variety of categorically distinct pathologies that share at least one common driving force – cell injury.Clinical & Experimental Immunology 10/2014; 179(2). DOI:10.1111/cei.12476 · 3.28 Impact Factor
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ABSTRACT: Our aim was to study the protein expression profiles of placenta obtained from lean and obese pregnant women with normal glucose tolerance at the time of term Caesarean section. We used two-dimensional difference gel electrophoresis (2D-DIGE), utilising narrow-range immobilised pH gradient strips that encompassed the broad pH range of 4-5 and 5-6, followed by MALDI-TOF mass spectrometry of selected protein spots. Western blot and quantitative RT-PCR (qRT-PCR) analyses were performed to validate representative findings from the 2D-DIGE analysis. Eight proteins were altered (six down-regulated and two up-regulated on obese placentas). Annexin A5 (ANXA5), ATP synthase subunit beta, mitochondria (ATPB), brain acid soluble protein 1 (BASP1), ferritin light chain (FTL), heterogeneous nuclear ribonucleoprotein C (HNRPC) and vimentin (VIME) were all lower in obese patients. Alpha-1-antitrypsin (A1AT) and stress-70 protein, mitochondrial (GRP75) were higher in obese patients. Western blot analysis of ANXA5, ATPB, FTL, VIME, A1AT and GRP75 confirmed the findings from the 2D-DIGE analysis. For brain acid soluble protein 1 and HNRPC, qRT-PCR analysis also confirmed the findings from the 2D-DIGE analysis. Immunohistochemical analysis was also used to determine the localisation of the proteins in human placenta. In conclusion, proteomic analysis of placenta reveals differential expression of several proteins in patients with pre-existing obesity. These proteins are implicated in a variety of cellular functions such as regulation of growth, cytoskeletal structure, oxidative stress, inflammation, coagulation and apoptosis. These disturbances may have significant implications for fetal growth and development.Journal of Molecular Endocrinology 02/2012; 48(2):139-49. DOI:10.1530/JME-11-0123 · 3.62 Impact Factor