ABSTRACTS: Oral Presentation Abstracts
Introduction: The understanding of the mechanisms by which eosinophils migrate into the intestinal epithelium can provide alternative therapeutic targets for conditions characterized by eosinphilic cryptitis and crypt abscesses. However, the details of how these eosinophils adhere and migrate into the intestinal epithelium are not known. The lack of data is due, in part, to the relative scarcity of human eosinophils for large-scale studies. HL-60 differentiated eosinophils present an alternative option by which studies can be done to elucidate eosinophil function and chemotaxis. Eosinophil migration is dependent on adhesion molecule such as selectins. The tetrasaccharide sialyl Lewis(x) (sLe(x)) binds to all 3 selectins, so compounds that mimic sLe(x), such as TBC1269, are potential antagonists. Human eosinophils express L-selectin and P-selectin counterligand PSGL-1. We hypothesize that eosinophils migrate from the basolateral to the apical surface of intestinal epithelium through the orchestrated effects of selectins. This study was performed to examine the inhibitory effect of sLex mimetic (which antagonizes all selectins) on the intestinal epithelial migration of HL-60 differentiated eosinophils. Consequently, the role of individual selectins during the process of migration would be evaluated.
Methods: TBC1269 was added to fluorescently labeled HL-60 clone 15 eosinophils in incremental amounts. Subsequently, blocking antibodies towards L-selectin and PSGL-1 were used in a similar manner. HL-60 eosinophils were allowed to migrate into T-84 monolayers. The number of migrated HL-60 cells was calculated by comparing fluorescence to known cell densities.
Results: The number of HL-60 eosinophils undergoing migration was significantly lower in the presence of TBC1269. This effect was concentration-dependant and near complete inhibition of migration was seen at a TBC1269 concentration of 10mg/ml (2.95X105±0.15X105 migration in the absence of TBC1269 and 0.71X105±0.04X105 migration in the presence of TBC1269, p<0.0001, n=8, control [no fMLP gradient]=0.68X105±0.04X105). In addition, HL-60 eosinophil migration was significantly lower in the presence of the blocking antibodies to PSGL-1 and L-selectin (39.2% and 51.6% inhibition, respectively). Simultaneous blocking of PSGL-1 and L-selectin resulted in inhibition of 76.0% of the migration.
Conclusion: The results of this study suggest a major role for selectins in the intestinal epithelial migration of differentiated HL-60 eosinophils. Tetrasaccharide sialyl Lewis(x), L-selectin, and the P-selectin counterligand PSGL-1 could be potential therapeutic targets.