Alexander Zarbock

Leukocyte ligands for endothelial selectins: specialized glycoconjugates that mediate rolling and signaling under flow

Reversible interactions of glycoconjugates on leukocytes with P- and E-selectin on endothelial cells mediate tethering and rolling of leukocytes in inflamed vascular beds, the first step in their recruitment to sites of injury. Although selectin ligands on hematopoietic precursors have been identified, here we review evidence that PSGL-1, CD44, and ESL-1 on mature leukocytes are physiologic glycoprotein ligands for endothelial selectins. Each ligand has specialized adhesive functions during tethering and rolling. Furthermore, PSGL-1 and CD44 induce signals that activate the β2 integrin LFA-1 and promote slow rolling, whereas ESL-1 induces signals that activate the β2 integrin Mac-1 in adherent neutrophils. We also review evidence for glycolipids, CD43, L-selectin, and other glycoconjugates as potential physiologic ligands for endothelial selectins on neutrophils or lymphocytes. Although the physiologic characterization of these ligands has been obtained in mice, we also note reported similarities and differences with human selectin ligands.

GDF-15 is an inhibitor of leukocyte integrin activation required for survival after myocardial infarction in mice

Inflammatory cell recruitment after myocardial infarction needs to be tightly controlled to permit infarct healing while avoiding fatal complications such as cardiac rupture. Growth differentiation factor-15 (GDF-15), a transforming growth factor-β (TGF-β)–related cytokine, is induced in the infarcted heart of mice and humans. We show that coronary artery ligation in Gdf15-deficient mice led to enhanced recruitment of polymorphonuclear leukocytes (PMNs) into the infarcted myocardium and an increased incidence of cardiac rupture. Conversely, infusion of recombinant GDF-15 repressed PMN recruitment after myocardial infarction. In vitro, GDF-15 inhibited PMN adhesion, arrest under flow and transendothelial migration. Mechanistically, GDF-15 counteracted chemokine-triggered conformational activation and clustering of β2 integrins on PMNs by activating the small GTPase Cdc42 and inhibiting activation of the small GTPase Rap1. Intravital microscopy in vivo in Gdf15-deficient mice showed that Gdf-15 is required to prevent excessive chemokine-activated leukocyte arrest on the endothelium. Genetic ablation of β2 integrins in myeloid cells rescued the mortality of Gdf15-deficient mice after myocardial infarction. To our knowledge, GDF-15 is the first cytokine identified as an inhibitor of PMN recruitment by direct interference with chemokine signaling and integrin activation. Loss of this anti-inflammatory mechanism leads to fatal cardiac rupture after myocardial infarction.

Rolling on E- or P-selectin induces the extended but not high-affinity conformation of LFA-1 in neutrophils

Human blood neutrophils rolling on E- or P-selectin reduced their rolling velocity when intercellular adhesion molecule (ICAM)-1 was available. Similar to mouse neutrophils, this was dependent on P-selectin glycoprotein ligand 1 (PSGL1), alpha(L)beta(2) integrin, the Src family tyrosine kinase FGR and spleen tyrosine kinase SYK. Blocking phospholipase C or p38 MAP kinase attenuated, but did not abolish the velocity reduction. To test expression of integrin activation epitopes, we adapted an immobilized reporter assay and developed a new homogeneous microfluidics-based reporter antibody binding assay. Rolling on E- or P-selectin induced the extension reporter epitopes KIM127 and NKI-L16, but not the high affinity reporter epitope monoclonal antibody (mAb) 24. This enabled rolling neutrophils to bind to immobilized extension reporter, but not activation reporter antibodies and allowed binding of soluble KIM127 during rolling. We conclude that human neutrophil rolling on E- or P-selectin induces the extended alpha(L)beta(2) integrin conformation through signaling triggered by PSGL-1 engagement.

Crucial role of SLP-76 and ADAP for neutrophil recruitment in mouse kidney ischemia-reperfusion injury

Leukocyte recruitment to the kidney during acute injury is mediated by E-selectin–mediated rolling and requires SLP-76 and the adaptor protein ADAP.

Leukocyte integrin activation and deactivation: novel mechanisms of balancing inflammation

Leukocyte recruitment into tissue forms the basis of immune surveillance and direct immune defense. It proceeds in a cascade-like fashion. The first contact of leukocytes with the endothelium is mediated by selectins and their counter receptors, followed by rolling and integrin-mediated arrest. While rolling, neutrophils collect different inflammatory signals which can activate several signaling pathways leading to leukocyte adhesion to the endothelium and transmigration through the blood vessel wall into the inflamed tissue. Whereas inflammatory reactions are beneficial and necessary for host defense, they need to be balanced and controlled to prevent harmful consequences and tissue destruction. In this article, we discuss the different signaling pathways that ensure rapid and efficient integrin activation on leukocytes. In addition, we report on a recently identified novel endogenous mechanism that counteracts and balances integrin activation, thereby limiting leukocyte recruitment and the extent of inflammation. Further investigation of this new mechanism may allow providing new approaches for the development of the next generation of anti-inflammatory drugs.

T lymphocyte rolling and recruitment into peripheral lymph nodes is regulated by a saturable density of L-selectin (CD62L)

L‐selectin mediates tethering and rolling of lymphocytes in high endothelial venules (HEV) of lymph nodes (LN) and of leukocytes at inflammatory sites. We used transgenic mice expressing varying levels of wild‐type or a non‐cleavable mutant form of L‐selectin on T cells to determine the relationship between L‐selectin density, tethering and rolling, and migration into LN. T cells expressing supraphysiological levels of either wild‐type or non‐cleavable L‐selectin showed rolling parameters similar to C57BL/6 T cells in hydrodynamic flow assays and during rolling in Peyers patch HEV. In contrast, PMA‐ or antigen‐activated T cells and L‐selectin+/– T cells expressing subphysiological levels of L‐selectin showed reduced numbers of rolling cells with increased rolling velocity. Short‐term homing studies showed that elevated expression of L‐selectin above physiological levels had no effect on T cell migration to LN; however, low L‐selectin expression resulted in reduced T cell homing to LN. Thus, T lymphocyte migration into LN is regulated by the density of cell surface L‐selectin. In addition, there is a saturable density of L‐selectin required for optimal homing to PLN in C57BL/6 mice, the L‐selectin level on circulating naive T cells promotes optimal homing, and increased expression above saturating levels promotes no further increase in T cell recruitment.