Methods: Eosinophil surface expression was quantified by flow cytometry in whole blood and on purified eosinophils stimulated with IL-5, GM-CSF (10ng/ml) or SEB (5ug/ml) or exposed to hypoxia (3% O2). Eosinophil apoptosis and migration were measured by AnnexinV-FITC/7-AAD staining and using transwell system, respectively.
Results: Expression of Siglec-7 on eosinophils in blood was low compared to CD300a and Siglec-8 (GM DMFI 70 vs 577 and 448, respectively), was similar between eosinophilic (n=13) and normal (n=16) donors (GM DMFI 52 vs. 88), and did not correlate with peripheral eosinophil count. Eosinophil expression of Siglec-7 was positively correlated with CD300a expression (r=0.42, p=0.025). Siglec-7 expression on purified eosinophils was unaffected by stimulation with IL-5, GM-CSF, SEB or hypoxia (which is known to upregulate CD300a expression). In contrast to anti-Siglec-8 antibodies, which induced eosinophil apoptosis after overnight priming with IL-5 or GM-CSF, anti-Siglec-7 antibodies had no effect on eosinophil viability under the same conditions (n=5). Finally, whereas eosinophil migration towards eotaxin decreased by 37-64% in the presence of anti-Siglec-8 antibody in all 4 donors tested, anti-siglec-7 antibody had no appreciable effect.
Conclusions: Siglec7 is weakly expressed on eosinophils from normal and eosinophilic donors. In contrast to Siglec-8 and CD300a, which have been shown to affect eosinophil function in vitro, the role of Siglec-7 remains to be elucidated.