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Cyanate, which is in equilibrium with urea, combines with the alpha-amino group of the aminoterminal valine of hemoglobin in an irreversible, specific carbamylation reaction. Partial carbamylation (0.72 residues/hemoglobin tetramer) as determined by cyanate-(14)C incorporation or hydantoin analysis diminishes the in vitro sickling phenomenon. Since cyanate may react not only with hemoglobin but also with functional groups of other red blood cell proteins, the in vitro effect of cyanate was studied on sickle cells. Cells were incubated with 10 mM KCl (control) or 10 mM KNCO (carbamylated) for 1 hr, washed, and resuspended in autologous plasma. Glycolysis, ATP and 2,3-diphosphoglyceric acid (DPG) stability, autohemolysis, and osmotic fragility were not affected by carbamylation. Potassium loss in carbamylated cells (2.8 mmol/liter) was less than in control cells (9.0 mmol/liter). Pyruvate kinase activity of carbamylated cells was decreased ( approximately 25%) but the activities of other glycolytic enzymes were similar to those of control cells. Oxygen affinity of carbamylated sickle, normal, and DPG-depleted normal cells increased, and was a sensitive index of the degree and duration of reaction with cyanate. The reactivity of carbamylated cells to DPG was similar to control cells. DPG-depleted carbamylated cells regenerated DPG and increased the P(50) when incubated with pyruvate, inosine, and phosphate. The Bohr effect of normal and of sickle cells was not affected (Deltalog P(50)/Delta pH=-0.48 and -0.53, respectively) after carbamylation. The reserve buffering capacity of plasma offset the slightly diminished ( approximately 15%) CO(2) capacity of carbamylated cells so that whole blood CO(2) capacity, pH, and P(CO2) were normal. These studies provide further support for the potential clinical use of cyanate in treating and preventing the anemia and painful crises of sickle cell disease.

The Effects of Cyanate In Vitro on Red Blood Cell Metabolism and Function in Sickle Cell Anemia