Extracellular Gelsolin Binds Lipoteichoic Acid and Modulates Cellular Response to Proinflammatory Bacterial Wall Components This information is current as Robert Bucki, Fitzroy J. Byfield, Alina Kulakowska, of September 23, 2021. Margaret E. McCormick, Wieslaw Drozdowski, Zbigniew Namiot, Thomas Hartung and Paul A. Janmey J Immunol 2008; 181:4936-4944; ; doi: 10.4049/jimmunol.181.7.4936 http://www.jimmunol.org/content/181/7/4936 Downloaded from References This article cites 59 articles, 18 of which you can access for free at: http://www.jimmunol.org/content/181/7/4936.full#ref-list-1 http://www.jimmunol.org/ Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication by guest on September 23, 2021 *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2008 by The American Association of Immunologists All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Extracellular Gelsolin Binds Lipoteichoic Acid and Modulates Cellular Response to Proinflammatory Bacterial Wall Components1 Robert Bucki,2* Fitzroy J. Byfield,* Alina Kulakowska,† Margaret E. McCormick,* Wieslaw Drozdowski,† Zbigniew Namiot,‡ Thomas Hartung,§ and Paul A. Janmey* The various functions of gelsolin in extracellular compartments are not yet clearly defined but include actin scavenging and antiin- flammatory effects. Gelsolin was recently reported to bind endotoxin (LPS) from various Gram-negative bacteria with high affinity. In this study we investigate whether gelsolin also interacts with bacterial wall molecules of Gram-positive bacteria such as lipoteichoic acid (LTA) and whether gelsolin’s interaction with bacterial lipids from Gram-negative or Gram-positive bacteria affects their cellular inflammatory responses. A peptide based on the PPI binding site of gelsolin (160–169) binds purified LTA at the same molecular Downloaded from ratio that it binds phosphatidylinositol 4,5-bisphosphate. The OD of recombinant human plasma gelsolin was found to decrease following the addition of purified LTA, and the binding of gelsolin to LTA inhibits F-actin depolymerization by gelsolin. Simul- taneously, the ability of LTA to activate translocation of NF-␬B, E-selectin expression, and adhesion of neutrophils to LTA-treated human aortic endothelial cells was compromised by gelsolin. Gelsolin was able to partially inhibit LPS- or LTA-induced release of IL-8 from human neutrophils but was unable to prevent Gram-positive Bacillus subtilis or Gram-negative Pseudomonas aeruginosa growth and had no effect on the antibacterial activity of the cathelicidin-derived antibacterial peptide LL37. These data http://www.jimmunol.org/ suggest that extracellular gelsolin is involved in the host immune recognition of LTA or LPS following release of these molecules from the bacterial outer membrane during cell division or attack by drugs and immune components. The Journal of Immunology, 2008, 181: 4936–4944. elsolin is an ϳ84-kDa actin-binding protein first identi- proposed is adult respiratory distress syndrome (ARDS)3 (8). Ac- fied in the cytoplasm. Intracellular gelsolin is involved tin release in excess of the actin scavenger molecules’ capacities G in the remodeling of actin filaments associated with affects the lung because of its size and rich blood flow through cell shape changes and movement (1, 2). Cells from gelsolin- narrow vessels, leading to the characteristic inflammatory null mice exhibit a variety of motility and cytoskeletal defects. changes of ARDS. In established ARDS cases, plasma gelsolin by guest on September 23, 2021 Gelsolin-null fibroblasts have pronounced actin stress fibers, a levels were found to be on average 30% of normal values (4). phenotype consistent with an inability to sever and remodel Recently, repletion with exogenous plasma gelsolin was found actin filaments (3). The functions of extracellular gelsolin are to be beneficial in mice subjected to hyperoxia (9), a condition less well defined. Initially its involvement in scavenging F-actin that often results in ARDS development. Furthermore, the treat- polymers released into the circulation during cell death was the ment of mice with exogenous plasma gelsolin significantly only role considered (4, 5). This preventative nature of gelsolin blunted neutrophil recruitment to the lungs (9), and gelsolin was is an idea supported by observations that a reduction in plasma able to attenuate vascular permeability associated with burn in- gelsolin as well as the detection of circulating gelsolin-actin jury in rats (5). Reduction in plasma gelsolin levels was also complexes were reported in a variety of human and animal in- observed in patients with sepsis, myocardial infarction, hepati- jury states (4, 6, 7). tis, myonecrosis (10), and trauma (11, 12). However, an insuf- One of the most important clinical examples of secondary injury ficiency in the blood F-actin buffering system has not been ob- in which the pathological implication of circulating F-actin was served, and the involvement of plasma gelsolin in inflammatory mediator transport has been proposed (13). Plasma gelsolin binds LPS from various bacteria, and some LPS-induced cellular functions are neutralized by gelsolin and by *University of Pennsylvania, Institute for Medicine and Engineering, Philadelphia, PA 19104; †Department of Neurology and ‡Department of Physiology, Medical Uni- a peptide based on gelsolin residues 160–169 (14). The gelsolin P2 versity of Bialystok, Bialystok, Poland; §European Union Joint Research Centre, In- peptide (residues 150–169) binds LPS with greater affinity than it stitute for the Protection and Security of the Citizen, the Traceability, Risk and Vul- binds lysophosphatidic acid (LPA) (15), a potent extracellular ag- nerability Assessment Unit (TRiVA), Ispra, Italy; and Department of Biochemical Pharmacology, University of Konstanz, Konstanz, Germany onist that influences endothelial cell migration and proliferation Received for publication October 2, 2007. Accepted for publication July 24, 2008. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance 3 Abbreviations used in this paper: ARDS, adult respiratory distress syndrome; CSF, with 18 U.S.C. Section 1734 solely to indicate this fact. cerebrospinal fluid; DLS, dynamic light scattering; HAEC, human aorta endothelial 1 cells; LBP, LPS-binding protein; LPA, lysophosphatidic acid; LTA, lipoteichoic acid; This work was supported by the National Institutes of Health (Grant AR38910), the Malp-2, macrophage-activating lipopeptide-2; MIC, minimal inhibitory concentra- Cystic Fibrosis Foundation, and the Medical University of Bialystok Grant 3-44903L. tion; PAF, platelet-activating factor; PBP10, rhodamine B-QRLFQVKGRR; PIP2, 2 Address correspondence and reprint requests to Dr. Robert Bucki, University of phosphatidylinositol 4,5-bisphosphate; RhB, rhodamine B; rhGSN, recombinant hu- Pennsylvania, Institute for Medicine and Engineering, 1010 Vagelos Research Lab- man plasma gelsolin. oratories, 3340 Smith Walk, Philadelphia, PA 19104. E-mail address: buckirob@ mail.med.upenn.edu Copyright © 2008 by The American Association of Immunologists, Inc. 0022-1767/08/$2.00 www.jimmunol.org The Journal of Immunology 4937 (16). Plasma gelsolin was also found to interfere with platelet- (L4015), and Bacillus subtilis (L3265) were purchased from Sigma- activating factor (PAF)-induced cellular activation in vitro, sug- Aldrich. According to the manufacturer’s quality control, the preparation Ͻ gesting a protective mechanism for gelsolin in vivo (17). These of LTA contained 1 ng of LPS/1 mg of LTA, and therefore LPS would contribute Ͻ10 pg/ml culture medium in the highest LTA concentration findings suggest a new role for plasma gelsolin in inflammatory used. Purified LTA from S. aureus was prepared as described previously response and, in this context, actin may directly affect gelsolin (23, 31). To calculate molar concentrations of LPS and LTA, we have used binding to LPS, LPA, or PAF. the lowest range of their reported molecular mass in buffer without divalent LPS and lipoteichoic acid (LTA) represent the major virulence cations (stock solution was made in H2O). Recombinant human plasma gelsolin (rhGSN) was obtained from Biogen Idec. Solution of human al- factors of Gram-negative and Gram-positive bacteria, respectively. bumin was from Baxter Healthcare. Human aortic endothelial cells LTA concentrations can reach higher levels at infectious sites (HAECs) were obtained from BioWhittaker. ELISA kit for IL-8 determi- compared with LPS. Reported local tissue concentrations of LTA nation was from BioLegend. Heat-inactivated P. aeruginosa (PAO1) and can be as high as 26 ␮g/ml (18), which may be associated with the B. subtilis (American Type Culture Collection (ATCC) 6051) were ob- 8 7 ␮ tained by autoclaving their suspension (10 CFU/ml) in PBS for1hat fact