Effect of an anti-sulfatide single-chain antibody probe on platelet function.
Sulfatide (galactocylceramide-3'-sulfate), a cell surface glycosphingolipid interacts with several cell adhesion molecules including fibrinogen, von Willebrand factor (VWF), P-selectin, thrombospondin (TSP) and laminin, which are involved in haemostasis. We have used a sulfatide-specific single-chain fragment variable (scFv) antibody probe PA38 and sulfatide-deficient mice to investigate the role of membrane sulfatide in platelet function. PA38 bound to platelets and binding increased following platelet activation. Sulfatide was localized as a large cluster towards the center of the platelet surface when examined in a confocal microscope. PA38 (20 microg/ml) inhibited the adhesion of activated platelets to fibrinogen, VWF, P-selectin, TSP1 and laminin by 30%, 30%, 75%, 20% and 35%, respectively, compared to a control scFv (p < 0.05). Furthermore, PA38 inhibited collagen, ADP, thrombin and ristocetin-induced platelet aggregation in PRP by 25%, 30%, 18% and 20%, respectively, compared to the control scFv (p < 0.05). In a PFA-100 platelet function assay, PA38 prolonged the occlusion time by 25% (p < 0.05). Under flow PA38 decreased the thrombus formation on collagen by 31%, (p < 0.01). Sulfatide-deficient mice displayed an extended lag-phase in collagen-induced platelet aggregation compared to wild type (p < 0.05), though in-vivo haemostasis did not differ significantly. Thus, this study provides new evidence for a role for membrane sulfatide in platelet function.
Guchhait
Prasenjit
P
Thrombosis Research Section, Dept. of Medicine, One Baylor Plaza, BCM286, N1319, Houston, Texas 77030, USA. guchhait@bcm.tmc.edu
Shrimpton
Corie N
CN
Honke
Koichi
K
Rumbaut
Rolando E
RE
Lopez
Jose A
JA
Thiagarajan
Perumal
P
D000818:Animals; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects* / Q000378:metabolism; D002553:Cerebroside-Sulfatase / Q000235:genetics / Q000378:metabolism; D004195:Disease Models, Animal; D018056:Hemorheology; D006487:Hemostasis / Q000187:drug effects; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000494:pharmacology* / Q000627:therapeutic use; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D010973:Platelet Adhesiveness / Q000187:drug effects*; D010974:Platelet Aggregation / Q000187:drug effects*; D010979:Platelet Function Tests; D013433:Sulfoglycosphingolipids / Q000276:immunology / Q000378:metabolism*; D013927:Thrombosis / Q000097:blood / Q000517:prevention & control
D007135:Immunoglobulin Variable Region; D013433:Sulfoglycosphingolipids; D002553:Cerebroside-Sulfatase