Syndecan-4 mediates antithrombin-induced chemotaxis of human peripheral blood lymphocytes and monocytes.
Antithrombin inhibits chemokine-induced migration of neutrophils by activating heparan sulfate proteoglycan-dependent signaling. Whether antithrombin affects migration of other types of leukocytes is not known. We investigated the effects of antithrombin on spontaneous and chemokine-triggered migration of lymphocytes and monocytes from human peripheral blood in modified Boyden chamber micropore filter assays. Lymphocyte and monocyte populations from human peripheral blood were purified using magnetic antibody cell sorting. The signaling mechanisms required for antithrombin-dependent migration were studied using signaling enzyme blockers. Expression of heparan sulfate proteoglycan core protein was studied by RT-PCR and flow cytometry. The antithrombins used were Kybernin P from human plasma and a monoclonal-antibody-purified preparation from this plasma. Pretreatment of lymphocytes and monocytes with antithrombin inhibited chemotaxis toward optimal concentrations of interleukin-8 or Rantes (regulated upon activation normal T-cell expressed and activated) at concentrations of antithrombin as low as 10 nU/ml. In the absence of the chemokines, direct exposure of cells to gradients of antithrombin stimulated migration. Effects of antithrombin were abolished by pretreating cells with heparinase-1, chondroitinase, sodium chlorate and anti-syndecan-4 antibodies. Expression of syndecan-4 mRNA and protein in monocytes and lymphocytes was demonstrated in RT-PCR and anti-syndecan-4 immunoreactivity assays, respectively. In the presence of pentasaccharide, antithrombin lost its effect on cells. Data indicate that antithrombin directly inhibits chemokine-stimulated migration of monocytes and lymphocytes via the effects of its heparin-binding site on cell surface syndecan-4 by activation of protein kinase C and Rho signaling.
Kaneider
Nicole C
NC
Division of General Internal Medicine, Department of Internal Medicine, University of Innsbruck, Anichstrasse 35, A-6020 Innsbruck, Austria.
Reinisch
Christina M
CM
Dunzendorfer
Stefan
S
Römisch
Jürgen
J
Wiedermann
Christian J
CJ
Wiederman
Christian J
CJ
D000911:Antibodies, Monoclonal / Q000276:immunology; D000991:Antithrombins / Q000494:pharmacology*; D002478:Cells, Cultured; D018946:Chemokine CCL5 / Q000378:metabolism; D002634:Chemotaxis, Leukocyte; D002704:Chlorates / Q000378:metabolism; D002810:Chondroitinases and Chondroitin Lyases / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology*; D019812:Heparan Sulfate Proteoglycans / Q000378:metabolism; D019763:Heparin Lyase / Q000378:metabolism; D006801:Humans; D016209:Interleukin-8 / Q000378:metabolism; D008214:Lymphocytes / Q000187:drug effects / Q000276:immunology*; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D009000:Monocytes / Q000187:drug effects / Q000276:immunology*; D011509:Proteoglycans / Q000276:immunology / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D053671:Syndecan-4; D014471:Umbilical Veins / Q000166:cytology
D000911:Antibodies, Monoclonal; D000991:Antithrombins; D018946:Chemokine CCL5; D002704:Chlorates; D019812:Heparan Sulfate Proteoglycans; D016209:Interleukin-8; D008562:Membrane Glycoproteins; D011509:Proteoglycans; D012333:RNA, Messenger; C508304:SDC4 protein, human; D053671:Syndecan-4; D002810:Chondroitinases and Chondroitin Lyases; D019763:Heparin Lyase; C032706:sodium chlorate