Platelet–Leukocyte–Endothelial Cell Interactions after Middle Cerebral Artery Occlusion and Reperfusion

Abstract

Summary:The adhesion of both leukocytes and platelets to microvascular endothelial cells has been implicated in the pathogenesis of ischemia/reperfusion (I/R) injury in several vascular beds. The objectives of this study were to (1) assess the platelet-leukocyte-endothelial cell interactions induced in the cerebral microvasculature by middle cerebral artery occlusion (MCAO)/reperfusion, and (2) define the molecular determinants of the prothrombogenic and inflammatory responses in this model of focal I/R. MCAO was induced for 1 hour in wild-type (WT) mice, WT mice treated with a monoclonal antibody (mAb) to either P-selectin or GPIIb/IIIa, and in P-selectin) chimeras. Isolated platelets labeled with carboxyfluorescein diacetate succinimidyl ester (CFDASE) were administered intravenously and observed with intravital fluorescence microscopy. Leukocytes were observed after intravenous injection of rhodamine 6G. One hour of MCAO followed by 1 hour of reperfusion resulted in the rolling and adhesion of leukocytes in venules, and after 4 hours of reperfusion, the adhesion of both leukocytes and platelets was detected. Although both the P-selectin and GPIIb/IIIa mAbs significantly reduced the adhesion of leukocytes and platelets at 4 hours of reperfusion, the antiadhesive effects of the P-selectin mAb were much greater. The leukocyte and platelet adhesion responses were significantly attenuated in both P-sel −/− →WT and WT→P-sel −/− bone marrow chimeras, compared with WT→WT chimeras. Neutropenia, induced by antineutrophil serum treatment, also reduced the recruitment of leukocytes and platelets after cerebral I/R. These findings implicate a major role for both platelet-associated and endothelial cellassociated P-selectin, as well as neutrophils in the inflammatory and prothrombogenic responses in the microcirculation after focal cerebral I/R.

Platelet Recruitment to the Inflamed Glomerulus Occurs via an αIIbβ3/GPVI-Dependent Pathway

“…25,29 Conversely, in other models, platelet accumulation within the inflamed microvasculature is facilitated via interaction with adherent leukocytes. 23,24 In the model of glomerulonephritis used in the present study, our findings indicate that both mechanisms are active, and that recruitment of one cell type facilitates the other. Previously we have found that platelet depletion inhibits leukocyte recruitment and glomerular injury.…”

“…Indeed, leukocytes have been shown to promote platelet adhesion in other inflammatory models. 23,24 In mice treated with anti-GBM antibody, which typically have elevated neutrophil counts, treatment with neutrophil-depleting antibody RB6-8C5 Figure 3. Anti-GBM antibody alters the morphology of the glomerular endothelium within ten minutes.…”

“…20 Platelets have also been shown to interact with the inflamed endothelium via both endothelial and platelet P-selectin. [23][24][25][26] However, in the glomerulus, capillary endothelial cells do not express P-selectin. 2,27,28 Moreover, we observed that P-selectin blockade did not abolish platelet recruitment in the inflamed glomerulus, indicating that P-selectin was not involved in this process.…”

Measuring and interpreting platelet-leukocyte aggregates

“…In a murine model of cerebral artery occlusion, IVM studies showed platelet-leukocyte interactions within post-ischemic venules (88). These events might be facilitated via GPIIb/IIIa or P- and L-selectin, as inhibition of these molecules in an experimental model of ischemic stroke and reperfusion reduces the increase in PLAs (89).…”

Deficiency of PAR4 Attenuates Cerebral Ischemia/Reperfusion Injury in Mice

“…P-selectin secreted from platelet-a granules and Weibel-Palade bodies of endothelial cells is critical for leukocyte rolling. P-selectin from platelets is important for the formation of plateletleukocyte aggregation and recruitment of leukocytes to the endothelial surface (Ishikawa et al, 2004). Thrombin activates the PAR4 receptor and causes PAR4 deletion attenuates cerebral ischemic injury Y Mao et al P-selectin secretion in platelets.…”