Immunoneutralization of glycoprotein Ib alpha attenuates endotoxin-inducedinteractions of platelets and leukocytes with rat venular endothelium in vivo

This study aimed to examine molecular mechanisms for endotoxin-induced adhe sive changes in platelets in vivo. Platelets labeled with carboxyfluorescei n diacetate succinimidyl ester were visualized in rat mesenteric venules th rough intravital microscopy assisted by a high-speed fluorescence video ima ger at 1000 frames per second or by a normal-speed intensifier under monito ring of erythrocyte velocity. Leukocyte rolling was examined by normal-spee d transmission video images. The velocity of platelets traveling along the centerline of venules followed that of erythrocytes, whereas that measured at the periendothelial space was significantly smaller than the erythrocyte velocity; a majority of these cells exhibited transient but notable rollin g with endothelium. Administration of endotoxin increased the density elf p eriendothelial platelets and reduced the rolling velocities of platelets an d leukocytes in venules, All events were attenuated by anti-rat P-selectin monoclonal antibody s789G or by anti-human glycoprotein (GP) Ib alpha monoc lonal antibody GUR83/35, which blocks ristocetin-induced aggregation of rat platelets. Isolated rat platelets injected into endotoxin-pretreated rats were able to roll on the venules, This event was attenuated by pretreatment of platelets in vitro with GUR83/35 but not with s789G, suggesting involve ment of endothelial P-selectin and platelet GP Ib alpha in the endotoxin-in duced responses. Furthermore, isolated human platelets showed similar rolli ng interactions with endotoxin-preexposed rat venules, and pretreatment of the platelets with GUR83/35, but not with s789G, significantly reduced such interactions. Our results provide the first evidence for involvement of GP Ib alpha in endotoxin-induced microvascular rolling of platelets and leuko cytes, and this system serves as a potentially useful tool to examine GP Ib alpha-associated function of human platelets in vivo.