Although sickle cell disease is a disease related to abnormal hemoglobin iside the red cell, a variety of morphologcal and functional abnormalities have been identified in the membranes of sickle cells called irreversibly sickled cells (ISC). These ISC are highly dehydrated and have reduced deformability, altered membrane phospholipid organization, hypercoagulability, and increased tendency to adhere to vascular endothelium. These abnormalities of ISC as well as a relationship between ISC counts and reduced red cell survival, conjunctival blood vessel abnormalities, and occurrence of leg ulcers suggest a potential role for ISC in the pathogenesis of sickle cell disease. The aim of this project is to discover whether there is any correlation between the degree of membrane lipid peroxidaition, altered lipid asymmetry and hypercoagulability in varying density fractions of sickle cells separated on Stractan density gradients and whether these membrane abnormalities are present only in the ISC due to sickling or are abnormalities that accrue gradually to membrane damage. In vitro cellular dehydration induced by Nystatin treatment to discoid sickle cells will be used to discriminate predominant role of membrane damage versus cellular dehydration per se in the hypercoagulability, altered membrane lipid asymmetry and peroxidative membrane lipid damage of sickle cells. This project also will explore whether exogenous peroxidant threat in vitro to discoid sickle cells can generate ISC comparable to in vivo-formed ISC fractionated from sickle blood. Standard techniques for analyses of membrane lipid peroxidation, membrane phospholipid organization, coagulability, and proteins will be utilized and have been standardized in this laboratory. Data from at least 20 sickle cell patients will be analyzed statistically.