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Infection and Immunity, February 2009, p. 725-732, Vol. 77, No. 2
0019-9567/09/$08.00+0 doi:10.1128/IAI.01305-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Inhibition of Complement and CD14 Attenuates the Escherichia coli-Induced Inflammatory Response in Porcine Whole Blood
Ebbe Billmann Thorgersen,1*
Anne Pharo,1
Karin Haverson,2
Anne K. Axelsen,3
Peter Gaustad,3
Girish J. Kotwal,4,
Georgia Sfyroera,5 and
Tom Eirik Mollnes1
Institute of Immunology, Rikshospitalet University Hospital and University of Oslo, Oslo, Norway,1 Department of Clinical Veterinary Science, University of Bristol, Bristol BS40 5DU, United Kingdom,2 Department of Medical Microbiology, Rikshospitalet University Hospital and University of Oslo, Oslo, Norway,3 Division of Medical Virology, IIDMM, University of Cape Town, HSC, Cape Town 7925, South Africa,4 Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 191045
Received 24 October 2008/ Accepted 18 November 2008
The innate immune response is a double-edged sword in systemic inflammation and sepsis. Uncontrolled or inappropriate activation can damage and be lethal to the host. Several studies have investigated inhibition of downstream mediators, including tumor necrosis factor alpha (TNF-
) and interleukin-1β (IL-1β). Emerging evidence indicates that upstream inhibition is a better therapeutic approach for attenuating damaging immune activation. Therefore, we investigated inhibition of two central innate immune pathways, those of complement and CD14/Toll-like receptor 4 (TLR4)/myeloid differentiation protein 2 (MD-2), in a porcine in vitro model of Escherichia coli-induced inflammation. Porcine whole blood anticoagulated with lepuridin, which did not interfere with the complement system, was incubated with E. coli lipopolysaccharide (LPS) or whole bacteria. Inhibitors of complement and CD14 and thus the LPS CD14/TLR4/MD-2 receptor complex were tested to investigate the effect on the inflammatory response. A broad range of inflammatory readouts were used to monitor the effect. Anti-CD14 was found to saturate the CD14 molecule on granulocytes and completely inhibited LPS-induced proinflammatory cytokines in a dose-dependent manner. Anti-CD14 significantly reduced the levels of the E. coli-induced proinflammatory cytokines TNF- and IL-1β, but not IL-8, in a dose-dependent manner. No effect on bacterial clearance was seen. Vaccinia complement control protein and smallpox inhibitor of complement enzymes, two Orthopoxvirus-encoded complement inhibitors, completely inhibited complement activation. Furthermore, these agents almost completely inhibited the expression of wCD11R3, which is associated with CD18 as a β2 integrin, on porcine granulocytes and decreased IL-8 levels significantly in a dose-dependent manner. As expected, complement inhibition reduced bacterial clearance. We conclude that inhibition of complement and CD14 attenuates E. coli-induced inflammation and might be used as a therapeutic regimen in gram-negative sepsis along with appropriate treatment with antibiotics.
* Corresponding author. Mailing address: Institute of Immunology, Rikshospitalet University Hospital, N-0027 Oslo, Norway. Phone: 47 23073014. Fax: 47 23073510. E-mail: ebbtho{at}rr-research.no
Published ahead of print on 1 December 2008.
Present address: Kotwal Bioconsulting, LLC, and InFlaMed, Inc., Louisville, KY 40241.
Infection and Immunity, February 2009, p. 725-732, Vol. 77, No. 2
0019-9567/09/$08.00+0 doi:10.1128/IAI.01305-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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