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dc.contributor.authorNilsson, Per
dc.contributor.authorJohnson, Christina
dc.contributor.authorQuach, Huy Quang
dc.contributor.authorMacpherson, Alex
dc.contributor.authorDurrant, Oliver
dc.contributor.authorPischke, Soeren
dc.contributor.authorFure, Hilde
dc.contributor.authorLandsem, Anne
dc.contributor.authorBergseth, Grete
dc.contributor.authorSchjalm, Camilla
dc.contributor.authorHaugaard-Kedström, Linda M.
dc.contributor.authorHuber-Lang, Markus
dc.contributor.authorvan den Elsen, Jean
dc.contributor.authorBrekke, Ole-Lars
dc.contributor.authorMollnes, Tom Eirik
dc.date.accessioned2021-10-15T06:34:13Z
dc.date.available2021-10-15T06:34:13Z
dc.date.created2021-09-20T12:17:31Z
dc.date.issued2021
dc.identifier.citationJournal of Immunology. 2021, 207 (6), 1641-1651.en_US
dc.identifier.issn0022-1767
dc.identifier.urihttps://hdl.handle.net/11250/2823150
dc.description.abstractThrombin activation of C5 connects thrombosis to inflammation. Complement research in whole blood ex vivo necessitates anticoagulation, which potentially interferes with the inflammatory modulation by thrombin. We challenged the concept of thrombin as an activator of native C5 by analyzing complement activation and C5 cleavage in human whole blood anticoagulated with Gly-Pro-Arg-Pro (GPRP), a peptide targeting fibrin polymerization downstream of thrombin, allowing complete endogenous thrombin generation. GPRP dose-dependently inhibited coagulation but allowed for platelet activation in accordance with thrombin generation. Spontaneous and bacterial-induced complement activation by Escherichia coli and Staphylococcus aureus, analyzed at the level of C3 and C5, were similar in blood anticoagulated with GPRP and the thrombin inhibitor lepirudin. In the GPRP model, endogenous thrombin, even at supra-physiologic concentrations, did not cleave native C5, despite efficiently cleaving commercially sourced purified C5 protein, both in buffer and when added to C5-deficient serum. In normal serum, only exogenously added, commercially sourced C5 was cleaved, whereas the native plasma C5 remained intact. Crucially, affinity-purified C5, eluted under mild conditions using an MgCl2 solution, was not cleaved by thrombin. Acidification of plasma to pH ≤ 6.8 by hydrochloric or lactic acid induced a C5 antigenic change, nonreversible by pH neutralization, that permitted cleavage by thrombin. Circular dichroism on purified C5 confirmed the structural change during acidification. Thus, we propose that pH-induced conformational change allows thrombin-mediated cleavage of C5 and that, contrary to previous reports, thrombin does not cleave plasma C5 in its native form, suggesting that thrombin cleavage of C5 may be restricted to certain pathophysiological conditions.en_US
dc.language.isoengen_US
dc.publisherAmerican Association of Immunologistsen_US
dc.titleA conformational change of complement C5 is required for thrombin-mediated cleavage, revealed by a novel ex vivo human whole blood model preserving full thrombin activityen_US
dc.typePeer revieweden_US
dc.typeJournal articleen_US
dc.description.versionpublishedVersionen_US
dc.rights.holderCopyright © 2021 by The American Association of Immunologists, Inc. This article is distributed under The American Association of Immunologists, Inc., Reuse Terms and Conditions for Author Choice articles.en_US
dc.source.pagenumber1641-1651en_US
dc.source.volume207en_US
dc.source.journalJournal of Immunologyen_US
dc.source.issue6en_US
dc.identifier.doi10.4049/jimmunol.2001471
dc.identifier.cristin1935958
cristin.ispublishedtrue
cristin.fulltextoriginal
cristin.qualitycode2


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