Microvascular responses to experimental sickle cell anemia

Evidence suggests that endothelial cells assume an inflammatory phenotype in sickle cell disease (SCD) that is characterized by increased blood cell-endothelial cell adhesion and oxidative stress. The nature of the linkage between these vascular responses in SCD remains unclear. The objectives of the studies in this dissertation were: (1) to compare the expression of E- and P-selectin, quantified in different regional vascular beds, between sickle cell transgenic (betaS) mice and their wild-type counterparts and to define the cells that exhibit the altered selectin expression, (2) to assess the contribution of platelet- and endothelial cell-associated P-selectin to the exaggerated leukocyte-endothelial cell and platelet-endothelial cell adhesion that is observed in the cerebral microvasculature of betaS mice under basal conditions and following hypoxia-reoxygenation, and (3) to identify enzymatic and non-enzymatic sources of the oxidative and nitrosative stress that contribute to the blood cell-endothelial cell adhesion elicited in betaS mice.;Dual-radiolabeled antibody experiments revealed that the expression of E- and P-selectin are differentially increased in some vascular beds of unstimulated beta S mice. Intravital fluorescence microscopy revealed significantly elevated leukocyte-endothelial cell and platelet-endothelial cell adhesion in the cerebral microvasculature of betaS mice that were intensified by hypoxia-reoxygenation but attenuated by genetic deficiency of vascular P-selectin. Genetic over-expression of endothelial cell superoxide dismutase and genetic deficiency of endothelial NADPH oxidase each resulted in a reduction of hypoxia-reoxygenation induced blood cell adhesion in cerebral venules of betaS mice. Iron chelation similarly attenuated the betaS-associated adhesion responses, while xanthine oxidase inhibition did not. Nitric oxide synthase (NOS) inhibition and genetic deficiency of endothelial cell eNOS each normalized the adhesion responses, while a nitric oxide donor and genetic over-expression of vascular eNOS each exacerbated blood cell adhesion. A precursor for the eNOS cofactor tetrahydrobiopterin attenuated betaS-associated adhesion responses.;These findings suggest that SCD is associated with dysfunctional microvascular endothelium. Increased endothelial cell P-selectin expression and an imbalance between endothelial cell NOS-derived NO and superoxide, that is initiated by oxidative species derived from NADPH oxidase and catalytically active iron, appear to induce the pro-inflammatory and pro-thrombogenic phenotype that is assumed by the cerebral microvasculature in SCD.