Beta 2-integrin and L-selectin are obligatory receptors in neutrophil aggregation [see comments]

Abstract

We have recently found that antibodies to L-selectin, the homing receptor on neutrophils, are as effective as those to beta 2-integrin at blocking formyl peptide-stimulated aggregation. Therefore, we investigated the requirements for expression of L-selectin and beta 2- integrin on adjacent cells during aggregation. Fluorescence flow cytometry allowed characterization of aggregates on the basis of size and color, as well as antibody binding to these two adhesive molecules. Formyl peptide-stimulated aggregate formation was measured for individual populations fluorescently labeled red (LDS-751) or green (CD44-FITC), and interpopulation red-green cell conjugates. Blocking either the beta 2-integrin or L-selectin adhesive epitope with monoclonal antibody on individual cell populations resulted in an approximately 50% reduction in two-color aggregation as compared with that in unblocked samples. Shedding the L-selectin on a cell population by preincubation with complexes of lipopolysaccharide and its plasma membrane binding protein also decreased aggregation to a control population by approximately 50%. We examined the aggregation of neutrophils from patients genetically deficient in beta 2-integrin and clinically leukocyte adhesion deficient (LAD). LAD adhesion to normal neutrophils was dependent on the expression of L-selectin on LAD cells and beta 2-integrin on normal cells. Thus, the minimum requirement for adhesion between two mixed populations of neutrophils was that one population expressed the beta 2-integrin and the other expressed the L- selectin adhesive epitope.

Mimicking the inflammatory cell adhesion cascade by nucleic acid aptamer programmed cell‐cell interactions

“…We have demonstrated for the first time that MSCs engineered with nucleic acid aptamers on their surface can specifically engage their respective protein receptors on a substrate or on adjacent cells. We chose P‐ and L‐selectin as target receptors because they are expressed on inflamed endothelial cells and leukocytes, respectively, and serve major roles in mediating leukocyte‐endothelium, leukocyte‐leukocyte, and leukocyte‐platelet interactions in the inflammatory cell adhesion cascade (4, 5, 9–11). We have demonstrated that MSCs engineered with aptamers, which target adhesion receptors on leukocytes, can recapitulate 3 key types of cell‐cell interactions under flow conditions that represent critical components of the inflammatory cell adhesion cascade (Scheme 1).…”

“…In addition, neutrophils that adhere on activated endothelium further capture free‐flowing neutrophils via interactions between L‐selectin and its ligands ( i.e. , PSGL‐1), which are both expressed on neutrophils (4, 5, 42). We first validated these native properties of neutrophils using the parallel flow chamber assay and observed robust neutrophil rolling, adhesion, and secondary tethering events on P‐selectin‐coated surfaces (data not shown), confirming that P‐selectin ligands and L‐selectin expressed on neutrophils are viable and functional and confirming the reliability of using such an assay to study MSC‐neutrophil interactions, as described below.…”

“…Interestingly, unlike interactions between L‐aptamer‐MSCs on L‐selectin‐coated surfaces that were sustained well above 1.5 dyn/cm 2 , L‐aptamer‐MSCs and neutrophil interactions were only effective under flow conditions at shear stresses of 0.5 dyn/cm 2 or lower. It is unclear whether this is due to cell‐cell interactions being ineffective at higher shear stresses and/or the shedding of L‐selectins from neutrophil surfaces at higher shear stresses (4, 5, 42, 43).…”

“…This event includes 3 types of cell‐cell interactions: between circulating inflammatory cells and endothelial cells (ECs; refs. 1, 3), between circulating inflammatory cells and inflammatory cells arrested on ECs (4, 5), and between 2 or more circulating inflammatory cells in blood circulation (6, 7), which form cellular complexes and subsequently adhere on ECs. These intercellular interactions, which collectively deliver and accumulate immune cells to sites of inflammation, are mediated by cell surface adhesion molecules and, in particular, selectins and their ligands (8).…”

“…L‐selectin and its ligands ( e.g. , PSGL‐1) are exclusively responsible for leukocyte‐leukocyte interactions (4, 5).…”