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In-Vitro Suppression of IL-6 and IL-8 Release from Human Pulmonary Epithelial Cells by Non-Anticoagulant Fraction of Enoxaparin
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In-Vitro Suppression of IL-6 and IL-8 Release from Human Pulmonary Epithelial Cells by Non-Anticoagulant Fraction of Enoxaparin
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In-Vitro Suppression of IL-6 and IL-8 Release from Human Pulmonary Epithelial Cells by Non-Anticoagulant Fraction of Enoxaparin
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Journal Article
In-Vitro Suppression of IL-6 and IL-8 Release from Human Pulmonary Epithelial Cells by Non-Anticoagulant Fraction of Enoxaparin
2015
Overview
Enoxaparin, a mixture of anticoagulant and non-anticoagulant fractions, is widely used as an anticoagulant agent. However, it is also reported to possess anti-inflammatory properties. Our study indicated that enoxaparin inhibits the release of IL-6 and IL-8 from A549 pulmonary epithelial cells. Their release causes extensive lung tissue damage. The use of enoxaparin as an anti-inflammatory agent is hampered due to the risk of bleeding associated with its anticoagulant fractions. Therefore, we aimed to identify the fraction responsible for the observed anti-inflammatory effect of enoxaparin and to determine the relationship between its structure and biological activities.
A549 pulmonary epithelial cells were pre-treated in the presence of enoxaparin and its fractions. The levels of IL-6 and IL-8 released from the trypsin-stimulated cells were measured by ELISA. The anticoagulant activity of the fraction responsible for the effect of enoxaparin was determined using an anti-factor-Xa assay. The fraction was structurally characterised using nuclear magnetic resonance. The fraction was 2-O, 6-O or N-desulfated to determine the position of sulfate groups required for the inhibition of interleukins. High-performance size-exclusion chromatography was performed to rule out that the observed effect was due to the interaction between the fraction and trypsin or interleukins.
Enoxaparin (60 μg/mL) inhibited the release of IL-6 and IL-8 by >30%. The fraction responsible for this effect of enoxaparin was found to be a disaccharide composed of α-L-iduronic-acid and α-D-glucosamine-6-sulfate. It (15 μg/mL) inhibited the release of interleukins by >70%. The 6-O sulphate groups were responsible for its anti-inflammatory effect. The fraction did not bind to trypsin or interleukins, suggesting the effect was not due to an artefact of the experimental model.
The identified disaccharide has no anticoagulant activity and therefore eliminates the risk of bleeding associated with enoxaparin. Future in-vivo studies should be designed to validate findings of the current study.
Publisher
Public Library of Science,Public Library of Science (PLoS)
Subject
Alveolar Epithelial Cells - drug effects
/ Alveolar Epithelial Cells - metabolism
/ Asthma
/ Bleeding
/ Cell Proliferation - drug effects
/ Cell Survival - drug effects
/ Dose-Response Relationship, Drug
/ Enzyme-linked immunosorbent assay
/ Epithelial Cells - drug effects
/ Epithelial Cells - metabolism
/ Humans
/ Interleukin-6 - biosynthesis
/ Interleukin-8 - biosynthesis
/ Lungs
/ Medicine
/ NMR
/ Pharmacy
/ Proteins
/ Rodents
/ Sulfates
/ Trypsin
