Role of Siglec-7 in Apoptosis in Human Platelets
Figure 1
Expression of Siglec-7 in human platelets.
(A), (B) Siglec-7 expression in platelets was analyzed by flow cytometry (n = 20). Platelets (stimulated with TRAP or vehicle control) were labeled with anti-CD41a and anti-Siglec-7 mAbs. Membrane expression of Siglec-7+ was increased after TRAP-induced platelet activation (*significant difference, of %CD41a+Siglec-7+ between TRAP-induced platelets activation vs resting platelets, t-test, p<0.05). (C) Membrane and intracellular localization of Siglec-7 in platelets. Platelets before and after permeabilization (stimulated with TRAP or vehicle control) were labeled with anti-CD41a, anti-CD62P (positive control for permeabilized platelets) and anti-Siglec-7 mAbs. Gating on the CD41a+ population, intra-platelet expression of Siglec-7 is significantly higher than its membrane expression (in both stimulated and unstimulated) (*, # significant difference (t-test, p<0.05) between %CD41a+CD62P+ or %CD41a+Siglec-7+, respectively, vs non-permeabilized resting platelets. (D) Distribution of Siglec-7 in platelets analyzed by confocal microscopy. Immunofluorescence labeling with anti–CD41a, and anti-Siglec-7 mAbs, and overlay (top to bottom). Siglec-7 staining in permeabilized (right panel) and non-permeabilized platelets (left panel) shows intracellular expression is more important than membrane expression. Scale bars = 6 µm. (E) Labeling with anti-tubulin and anti Siglec-7 antibodies and overlay (top to bottom). Siglec-7 is expressed on platelet membranes and tubulin is stained in permeabilized platelets. Tubulin was labeled to demarcate platelet borders and Siglec-7 is mostly observed in intracellular compartments. Scale bars = 10 µm.