To target specific sites of inflammation within the body, leukocytes must first bind to the endothelial lining of the vasculature through selectins and/oralpha4 integrins, leading to an alpha4beta1 or beta2 integrin dependent arrest and emigration into the extravascular space. Control of leukocyte accumulation appears to be exerted through several mechanisms including selectin molecular biophysics and integrin affinity modulation by inflammatory signals that lead to arrest of the rolling leukocyte on the blood vessel wall. We hypothesize that the intrinsic selectin and alpha4 integrin bond lifetimes exert a controlliIig effect on leukocyte sensing of endothelial cell presented signaling chemokines that ultimately leads to arrest. We will address this hypothesis by quantification of the dependence of the molecular bond mechanics of an L-selectin and P-selectin shared ligand, P-selectin Glycoprotein Ligand-1 (PSGL-1) on the binding pocket structure and the cytoplasmic domain structure. Once rolling, we hypothesize that selectin and alpha4beta1 bond lifetimes control chemokine signaling and that cooperative effects of selectin and alpha4 integrins are important in mononuclear cell arrest through endothelial ligands such as VCAM-1 or ICAM-1. G-protein coupled receptors bind chemokines in parallel+ with adhesion receptor signaling, leading to beta1 and beta2 integrin avidity increases for endothelial ligands. The process of leukocyte arrest involves in part integrin-cytoskeletal interactions and possibly increases in integrin affinity for endothelial ligands. We will test small molecule antagonists of beta1/beta2 high affinity binding to VCAM-1 and ICAM-1 for their influence on the arrest of rolling leukocytes under flow conditions. The combination of small molecule antagonists of beta1 and beta2 integrin function, PSGL-1 mutants, flow chamber assays to create physiologically relevant forces on the flowing leukocyte, and high speed videomicroscopy (2 ms time resolution) to capture the dynamics of chemokine- and selectin-induced leukocyte arrest on purified leukocyte adhesion receptor ligands is unique and we believe will contribute to the understanding of the complex processes that govern leukocyte accumulation in tissues. Such insight may lead to identification of more effective molecular targets for anti-inflammatory therapeutics.
| Ham, Anthony S; Klibanov, Alexander L; Lawrence, Michael B (2009) Action at a distance: lengthening adhesion bonds with poly(ethylene glycol) spacers enhances mechanically stressed affinity for improved vascular targeting of microparticles. Langmuir 25:10038-44 |
| Ham, Anthony Sang Won; Goetz, Douglas J; Klibanov, Alexander L et al. (2007) Microparticle adhesive dynamics and rolling mediated by selectin-specific antibodies under flow. Biotechnol Bioeng 96:596-607 |
| Mougin, K; Ham, A S; Lawrence, M B et al. (2005) Construction of a tethered poly(ethylene glycol) surface gradient for studies of cell adhesion kinetics. Langmuir 21:4809-12 |
