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KMID : 1161520010050030253
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Animal Cells and Systems
2001 Volume.5 No. 3 p.253 ~ p.262
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Analysis of the molecular event of ICAM-1 interaction with LFA-1 during leukocyte adhesion using a reconstituted mammalian cell expression model
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Han Weon-Cheol
Kim Kwon-Seop
Park Jae-Seung
Hwang Sung-Yeoun
Moon Hyung-Bae
Chung Hun-Taeg
Jun Chang-Duk
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Abstract
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Ligand?receptor clustering event is the most important step in leukocyte adhesion and spreading on endothelial cells. Intercellular adhesion mole?cule?1 (ICAM?1) has been shown to enhance leukocyte adhesion, but the molecular event during the process of adhesion is unclear. To visualize the dynamics of ICAM?1 movement during adhesion, we have engineered stable Chinese hamster ovary cell lines expressing ICAM?1 fused to a green fluorescent protein (IC1GFP/CHO) and examined them under the fluorescence microscopy. The transfection of IC1GFP alone in these cells was sufficient to support the adhesion of K562 cells that express aLβ2 (LFA?1) integrin stimulated by CBR LFA?1/2 mAb. This phenomenon was mediated by ICAM?1?LFA?1 interactions, as an mAb that specifically inhibits ICAM?1?LFA?1 interaction (RR1/1) completely abolished the adhesion of LFA?1+ cells to IC1_GFP/CHO cells. We found that the characteristic of adhesion was followed almost immediately (∼10min) by the rapid accumulation of ICAM?1 on CHO cells at a tight interface between the two cells. Interestingly, IC1_GFP/CHO cells projected plasma membrane and encircled approximately half surface of LFA?1+ cells, as defined by confocal microscopy. This unusual phenomenon was also confirmed on HUVEC after adhesion of LFA?1+ cells. Neither cytochalas in D nor 2,3?butanedione 2?monoxime an inhibitor of myosin light chain kinase blocked LFA?1?mediated ICAM?1 clustering, indicating that actin cytoskeleton and myosin?dependent contractility are not necessary for ICAM?1 clustering. Taken together, we suggest that leukocyte adhesion to endothelial cells induces specialized form of ICAM?1 clustering that is distinct from immunological synapse mediated by T cell interaction with antigen presenting cells.
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KEYWORD
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Clustering, ICAM-1, Integrin, LFA-1, MLCK
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