The role of oxidation in hyperglycemia-mediated erythrocyte phospholipid asymmetry

Diabetes Mellitus is a chronic wide spread disease affecting 23.6 million people in the United States alone (2007 estimate). If left untreated, individuals with diabetes suffer from numerous vascular complications (such as heart disease and stroke), which can ultimately result in death. The primary symptom of diabetes is hyperglycemia, which has been shown to be the underlying cause of these complications. In the erythrocyte membrane, hyperglycemia induces a loss of phospholipid asymmetry, which promotes cell adhesion and triggers blood clotting factors, contributing to vascular obstruction. Hyperglycemia also accelerates the rate of transbilayer phospholipid movement (flip-flop) across the erythrocyte membrane. The mechanism(s) by which hyperglycemia causes the loss of phospholipid asymmetry and increased rate of lipid flip-flop are unknown, but may be due to oxidative stress. The goal of this research was to determine the mechanism by which hyperglycemia induces the loss of asymmetry, increases rates of phospholipid flip-flop and to determine if the effects of hyperglycemia on membrane structure are permanent or transient. Treatment of isolated non-diabetic human erythrocytes with hyperglycemic concentrations of glucose resulted in a loss of asymmetry, detected by an array of methods. An increase in flip-flop was also demonstrated and could be suppressed by the use of the antioxidants ascorbic acid and alpha-tocopherol. Using various methods to measure phospholipid asymmetry and lipid flip-flop a change in erythrocytes incubated in euglycemic levels of glucose was observed, suggesting that the overnight incubation might be the cause of these effects. The change in phospholipid asymmetry of eugylcemic treated cells could be eliminated upon incubation in a dialysis cassette or in cell-free plasma.;Erythrocytes isolated from diabetic rats (induced by streptozotocin injection) did not exhibit permanent loss of asymmetry. An increase in flip-flop was also observed in diabetic rats, which could be prevented by antioxidant administration. Taken together, these data suggest that the effects of hyperglycemia on membrane structure are transient, provide support that hyperglycemia affects erythrocyte membranes through oxidative stress and suggests a possible therapeutic approach for the treatment of diabetic patients.