RESUMO
The adhesion molecules intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) expressed in atherogenic lesions are thought to regulate monocyte diapedesis. To better understand their specific roles we used function-blocking antibodies and examined in a culture model the morphology, motility, and diapedesis of THP-1 cells interacting with human coronary artery endothelial cells. The number of motile THP-1 cells was reduced only when VCAM-1 or both ICAM-1 and VCAM-1 were blocked. Blockade of ICAM-1 and VCAM-1, either separately or together, reduced to the same degree the distance that THP-1 cells traveled. Diapedesis was reduced only during the simultaneous blockade of both adhesion molecules. Blockade of either ICAM-1 or VCAM-1 inhibited pseudopodia formation, but ICAM-1 blockade induced the formation of filopodia. We suggest that the interactions of endothelial ICAM-1 and VCAM-1 with their ligands differentially regulate distinct steps of diapedesis by modulating the ratio of active and inactive forms of small GTPases such as Rho, Rac, and Cdc42.
Assuntos
Quimiotaxia de Leucócito , Endotélio Vascular/fisiologia , Molécula 1 de Adesão Intercelular/fisiologia , Monócitos/imunologia , Receptores de Retorno de Linfócitos/fisiologia , Molécula 1 de Adesão de Célula Vascular/fisiologia , Anticorpos Monoclonais/farmacologia , Arteriosclerose/imunologia , Linhagem Celular , Tamanho Celular , Células Cultivadas , Citoesqueleto/ultraestrutura , Humanos , Integrina alfa4beta1 , Integrinas/fisiologia , Molécula 1 de Adesão Intercelular/imunologia , Antígeno-1 Associado à Função Linfocitária/fisiologia , Microscopia Confocal , Monócitos/citologia , Monócitos/ultraestrutura , Células Tumorais Cultivadas , Molécula 1 de Adesão de Célula Vascular/imunologiaRESUMO
To determine changes in the distribution of cell adhesion molecules during diapedesis of monocytes in situ, we labeled aortic whole mounts from hypercholesterolemic rats with Texas red-phalloidin and antibodies to LFA-1, PECAM-1, or alpha-catenin, and analyzed them by laser scanning confocal microscopy. Monocytes transmigrated through circular openings (transmigration passages) formed by pseudopodia that penetrated between adjacent endothelial cells. Transmigrating monocytes remained spherical above the endothelium, while spreading beneath it. The transmigration passage was lined by F-actin and partially by alpha-catenin, suggesting cadherin-mediated heterotypic interactions. LFA-1 was present in clusters at the monocyte cell surface throughout diapedesis, but was concentrated at the margin of the transmigration passage. PECAM-1 was enriched in the endothelial contact regions where the monocytes transmigrated. PECAM-1 was barely detectable in monocytes before and after diapedesis, but appeared during diapedesis at the cell surface in the parts of the monocyte located above the endothelium. PECAM-1 was enriched near the endothelial cell-cell junctions, but was not detected in parts that spread beneath the endothelium. Our results suggest a major role for LFA-1 during diapedesis and reveal dynamic changes in the distribution of PECAM-1, the actin cytoskeleton, and alpha-catenin during monocyte diapedesis in situ.