Related Experiment Videos
Comparison of human eosinophil and neutrophil ligands for P-selectin: ligands for P-selectin differ from those for
M Wein1, S A Sterbinsky, C A Bickel
1Department of Medicine, Johns Hopkins University School of Medicine, Johns Hopkins Asthma and Allergy Center, Baltimore, Maryland 21224-6801.
Abstract:
Eosinophils (EOS) and neutrophils (PMN) display different patterns of accumulation during various inflammatory reactions. We hypothesized that EOS and PMN may differ in their ligands for P-selectin, and that these ligands may differ from those previously identified for E-selectin. Recombinant human P-selectin was immobilized on plastic surfaces and adhesion of 51Cr-labeled human EOS or PMN was compared. EOS and PMN adhered in a concentration-dependent fashion, with similar maximal net adhesion. Preincubation with a blocking P-selectin antibody inhibited adhesion of both cell types, whereas a non-blocking antibody did not. To determine if the counterligands were sialylated proteins, cells were treated with various glycosidases and proteases before testing adhesion. Neuraminidase treatment markedly inhibited binding of both cell types, while endo-beta-galactosidase had no significant effect. Pretreatment with several proteases reduced adhesion of both cell types, although they consistently caused a greater inhibition of PMN binding than EOS binding. To determine whether the P-selectin ligands were surface structures whose expression or function may be altered by cell activation, leukocytes were pretreated with various stimuli; only platelet-activating factor (PAF) treatment reduced the capacity of leukocytes to adhere to P-selectin. Thus, the counterligands for P-selectin on EOS and PMN are similar sialylated, protease-sensitive, endo-beta-galactosidase-resistant structures, whose function and/or expression is reduced following treatment with PAF. These characteristics are clearly different than those reported for EOS and PMN ligands for E-selectin, and suggest disparate roles for P-selectin and E-selectin during EOS and PMN recruitment during inflammatory responses in vivo.
Insights
Eosinophils (EOS) and neutrophils (PMN) utilize similar sialylated ligands for P-selectin adhesion, differing from E-selectin ligands. Platelet-activating factor (PAF) reduces this P-selectin binding capacity in both cell types.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Eosinophils (EOS) and neutrophils (PMN) exhibit distinct inflammatory accumulation patterns.
- The specific ligands involved in EOS and PMN adhesion via P-selectin are not fully elucidated.
- Understanding these ligands is crucial for differentiating inflammatory response mechanisms.
Purpose of the Study:
- To investigate and compare the P-selectin counterligands on human eosinophils and neutrophils.
- To determine the biochemical characteristics of these P-selectin ligands.
- To assess the impact of cell activation on P-selectin-mediated adhesion.
Main Methods:
- Immobilization of recombinant human P-selectin on plastic surfaces.
- Measurement of 51Cr-labeled human EOS and PMN adhesion.
- Enzymatic treatments (glycosidases, proteases) and stimulation with platelet-activating factor (PAF) to analyze ligand properties.
Main Results:
- EOS and PMN demonstrated concentration-dependent adhesion to P-selectin, inhibited by a blocking P-selectin antibody.
- Neuraminidase treatment significantly reduced binding, while endo-beta-galactosidase had no effect.
- Protease treatment inhibited adhesion, with greater impact on PMN binding; PAF reduced leukocyte adhesion to P-selectin.
Conclusions:
- P-selectin counterligands on EOS and PMN are similar sialylated, protease-sensitive, and endo-beta-galactosidase-resistant structures.
- These ligands exhibit altered function or expression after PAF treatment.
- The distinct nature of these ligands compared to E-selectin ligands suggests differential roles in inflammatory cell recruitment.