Role of Primary and Secondary Capture for Leukocyte Accumulation In Vivo

From the Department of Biomedical Engineering, University of Virginia School of Medicine, Charlottesville.

Correspondence to Klaus Ley, MD, University of Virginia School of Medicine, Department of Biomedical Engineering, Health Sciences Center, Box 377, Charlottesville, VA 22908. E-mail kfl3f{at}virginia.edu

Abstract—Leukocyte accumulation during inflammation depends on the concerted action of selectin and integrin adhesion molecules, which promote capture, rolling, and arrest of these cells on activated endothelium. In addition to interacting with endothelial cells, leukocytes can also adhere to already adherent leukocytes through an L-selectin–dependent mechanism. Initiation of adhesion through this mechanism has been called nucleation and leads to characteristic geometric patterns (ie, clusters and strings) of adherent leukocytes in flow chambers. We have used intravital microscopy of tumor necrosis factor- (TNF-)–treated mouse cremaster muscles to quantitatively investigate the potential role of leukocyte-leukocyte adhesion in initiating and maintaining the leukocyte clusters that are commonly observed in inflamed venules. Our data show that in TNF-–treated venules with diameters between 23 and 108 µm, leukocyte adhesion occurs in clusters that are 19 to 50 µm long and 8 to 44 µm wide. They are almost entirely made up of slow-rolling leukocytes. Of all leukocytes recruited into a cluster (100%), the majority enter the cluster rolling along the endothelium and sharply reduce their velocity in the absence (59%) or presence (15%) of other leukocytes in proximity (one cell diameter). Some of the rolling leukocytes (17%) pass through the cluster without reducing their velocity. Recruitment of leukocytes from the free flow regime into a cluster is a rare event and accounts for only 7 (1.2%) of 476 leukocytes arriving in the cluster. However, of the leukocytes captured from the free flow, 6 initiated contact with a slow-rolling leukocyte rather than making direct contact with the endothelium. Our data show that leukocyte-leukocyte interactions can occur in vivo but are not important for cluster formation. This is confirmed by the observation of normal cluster formation in L-selectin–deficient mice, in which leukocyte-leukocyte interactions under flow are abolished. We conclude that leukocyte-mediated nucleation contributes little to leukocyte recruitment during inflammation in vivo. Cluster formation appears to be dominated by areas of endothelium with a higher expression of E-selectin, because cluster formation is greatly reduced in E-selectin–deficient mice.

Key Words: inflammation • E-selectin • rolling • intravital microscopy • knockout mouse

This article has been cited by other articles:

				E. E. Eriksson No detectable endothelial- or leukocyte-derived L-selectin ligand activity on the endothelium in inflamed cremaster muscle venules J. Leukoc. Biol., July 1, 2008; 84(1): 93 - 103. [Abstract] [Full Text] [PDF]

				B. Petri, M. Phillipson, and P. Kubes The Physiology of Leukocyte Recruitment: An In Vivo Perspective J. Immunol., May 15, 2008; 180(10): 6439 - 6446. [Abstract] [Full Text] [PDF]

				S. Nolan, R. Dixon, K. Norman, P. Hellewell, and V. Ridger Nitric Oxide Regulates Neutrophil Migration through Microparticle Formation Am. J. Pathol., January 1, 2008; 172(1): 265 - 273. [Abstract] [Full Text] [PDF]

				E. Galkina and K. Ley Vascular Adhesion Molecules in Atherosclerosis Arterioscler Thromb Vasc Biol, November 1, 2007; 27(11): 2292 - 2301. [Abstract] [Full Text] [PDF]

				R. Sumagin and I. H. Sarelius A role for ICAM-1 in maintenance of leukocyte-endothelial cell rolling interactions in inflamed arterioles Am J Physiol Heart Circ Physiol, November 1, 2007; 293(5): H2786 - H2798. [Abstract] [Full Text] [PDF]

				C. Abbal, M. Lambelet, D. Bertaggia, C. Gerbex, M. Martinez, A. Arcaro, M. Schapira, and O. Spertini Lipid raft adhesion receptors and Syk regulate selectin-dependent rolling under flow conditions Blood, November 15, 2006; 108(10): 3352 - 3359. [Abstract] [Full Text] [PDF]

				M. B. Kim and I. H. Sarelius Role of shear forces and adhesion molecule distribution on P-selectin-mediated leukocyte rolling in postcapillary venules Am J Physiol Heart Circ Physiol, December 1, 2004; 287(6): H2705 - H2711. [Abstract] [Full Text] [PDF]

				M. M. Burdick and K. Konstantopoulos Platelet-induced enhancement of LS174T colon carcinoma and THP-1 monocytoid cell adhesion to vascular endothelium under flow Am J Physiol Cell Physiol, August 1, 2004; 287(2): C539 - C547. [Abstract] [Full Text] [PDF]

				M. Sperandio, M. L. Smith, S. B. Forlow, T. S. Olson, L. Xia, R. P. McEver, and K. Ley P-selectin Glycoprotein Ligand-1 Mediates L-Selectin-dependent Leukocyte Rolling in Venules J. Exp. Med., May 19, 2003; 197(10): 1355 - 1363. [Abstract] [Full Text] [PDF]

				C. A. St. Hill, S. R. Alexander, and B. Walcheck Indirect capture augments leukocyte accumulation on P-selectin in flowing whole blood J. Leukoc. Biol., April 1, 2003; 73(4): 464 - 471. [Abstract] [Full Text] [PDF]

				M. P. Bernimoulin, X.-L. Zeng, C. Abbal, S. Giraud, M. Martinez, O. Michielin, M. Schapira, and O. Spertini Molecular Basis of Leukocyte Rolling on PSGL-1. PREDOMINANT ROLE OF CORE-2 O-GLYCANS AND OF TYROSINE SULFATE RESIDUE 51 J. Biol. Chem., January 3, 2003; 278(1): 37 - 47. [Abstract] [Full Text] [PDF]

				M. R. King and D. A. Hammer Multiparticle adhesive dynamics: Hydrodynamic recruitment of rolling leukocytes PNAS, December 18, 2001; 98(26): 14919 - 14924. [Abstract] [Full Text] [PDF]

				M. J. Cotter, K. E. Norman, P. G. Hellewell, and V. C. Ridger A Novel Method for Isolation of Neutrophils from Murine Blood Using Negative Immunomagnetic Separation Am. J. Pathol., August 1, 2001; 159(2): 473 - 481. [Abstract] [Full Text]

				E. E. Eriksson, X. Xie, J. Werr, P. Thoren, and L. Lindbom Importance of Primary Capture and L-Selectin-Dependent Secondary Capture in Leukocyte Accumulation in Inflammation and Atherosclerosis in Vivo J. Exp. Med., July 16, 2001; 194(2): 205 - 218. [Abstract] [Full Text] [PDF]

				A. Stucki, A.-S. Rivier, M. Gikic, N. Monai, M. Schapira, and O. Spertini Endothelial cell activation by myeloblasts: molecular mechanisms of leukostasis and leukemic cell dissemination Blood, April 1, 2001; 97(7): 2121 - 2129. [Abstract] [Full Text] [PDF]

				K. E. Norman, A. G. Katopodis, G. Thoma, F. Kolbinger, A. E. Hicks, M. J. Cotter, A. G. Pockley, and P. G. Hellewell P-selectin glycoprotein ligand-1 supports rolling on E- and P-selectin in vivo Blood, November 15, 2000; 96(10): 3585 - 3591. [Abstract] [Full Text] [PDF]

				V. Grabovsky, S. Feigelson, C. Chen, D. A. Bleijs, A. Peled, G. Cinamon, F. Baleux, F. Arenzana-Seisdedos, T. Lapidot, Y. van Kooyk, et al. Subsecond Induction of {alpha}4 Integrin Clustering by Immobilized Chemokines Stimulates Leukocyte Tethering and Rolling on Endothelial Vascular Cell Adhesion Molecule 1 under Flow Conditions J. Exp. Med., August 21, 2000; 192(4): 495 - 506. [Abstract] [Full Text] [PDF]

				D. J. Mitchell, P. Li, P. H. Reinhardt, and P. Kubes Importance of L-selectin-dependent leukocyte-leukocyte interactions in human whole blood Blood, May 1, 2000; 95(9): 2954 - 2959. [Abstract] [Full Text] [PDF]

				S. Neelamegham, A. D. Taylor, H. Shankaran, C. W. Smith, and S. I. Simon Shear and Time-Dependent Changes in Mac-1, LFA-1, and ICAM-3 Binding Regulate Neutrophil Homotypic Adhesion J. Immunol., April 1, 2000; 164(7): 3798 - 3805. [Abstract] [Full Text] [PDF]

				L. Tu, P. G. Murphy, X. Li, and T. F. Tedder L-Selectin Ligands Expressed by Human Leukocytes Are HECA-452 Antibody-Defined Carbohydrate Epitopes Preferentially Displayed by P-Selectin Glycoprotein Ligand-1 J. Immunol., November 1, 1999; 163(9): 5070 - 5078. [Abstract] [Full Text] [PDF]

				S. D. Robinson, P. S. Frenette, H. Rayburn, M. Cummiskey, M. Ullman-Cullere, D. D. Wagner, and R. O. Hynes Multiple, targeted deficiencies in selectins reveal a predominant role for P-selectin in leukocyte recruitment PNAS, September 28, 1999; 96(20): 11452 - 11457. [Abstract] [Full Text] [PDF]

				S. Kanwar, D. A. Steeber, T. F. Tedder, M. J. Hickey, and P. Kubes Overlapping Roles for L-Selectin and P-Selectin in Antigen-Induced Immune Responses in the Microvasculature J. Immunol., March 1, 1999; 162(5): 2709 - 2716. [Abstract] [Full Text] [PDF]

				Y.-C. Lim, K. Snapp, G. S. Kansas, R. Camphausen, H. Ding, and F. W. Luscinskas Important Contributions of P-Selectin Glycoprotein Ligand-1-Mediated Secondary Capture to Human Monocyte Adhesion to P-Selectin, E-Selectin, and TNF-{alpha}-Activated Endothelium Under Flow In Vitro J. Immunol., September 1, 1998; 161(5): 2501 - 2508. [Abstract] [Full Text] [PDF]

				U. Jung, C. L. Ramos, D. C. Bullard, and K. Ley Gene-targeted mice reveal importance of L-selectin-dependent rolling for neutrophil adhesion Am J Physiol Heart Circ Physiol, May 1, 1998; 274(5): H1785 - H1791. [Abstract] [Full Text] [PDF]

				X. Xie, A.-S. Rivier, A. Zakrzewicz, M. Bernimoulin, X.-L. Zeng, H. P. Wessel, M. Schapira, and O. Spertini Inhibition of Selectin-mediated Cell Adhesion and Prevention of Acute Inflammation by Nonanticoagulant Sulfated Saccharides. STUDIES WITH CARBOXYL-REDUCED AND SULFATED HEPARIN AND WITH TRESTATIN A SULFATE J. Biol. Chem., October 27, 2000; 275(44): 34818 - 34825. [Abstract] [Full Text] [PDF]

				T. S. Olson, K. Singbartl, and K. Ley L-selectin is required for fMLP- but not C5a-induced margination of neutrophils in pulmonary circulation Am J Physiol Regulatory Integrative Comp Physiol, April 1, 2002; 282(4): R1245 - R1252. [Abstract] [Full Text] [PDF]