In vitro effect of ketones and hyperglycemia on feline hemoglobin oxidation and D- and L-lactate production

Abstract

Abstract Objectives To investigate in vitro effects of ketosis and hyperglycemia on feline erythrocyte Heinz body formation, reduced glutathione (GSH) concentration, and D-and L-lactate production, and to identify potential metabolic mechanisms of oxidative stress in diabetic cats. Design Washed feline erythrocytes suspended in buffers containing normal or high glucose concentration were incubated with various concentrations of ketone bodies and tested at defined time intervals for Heinz bodies, GSH concentration, and D- and L-lactate production. Animals Three healthy female domestic cats. Procedure Erythrocytes were washed, suspended in buffers containing 5 mM glucose (simulates euglycemia) or 25 mM glucose (simulates hyperglycemia), and incubated with acetone (5 and 10 mM), acetoacetate (5 and 10 mM), or β-hydroxybutyrate (5 and 25 mM) for 24 hours at 37 C. Aliquots were stained with new methylene blue for Heinz bodies, and assayed spectrophotometrically for GSH and D- and L-lactate concentrations. Experiments were done in triplicate. Data were analyzed, using ANOVA with repeated measures. Results Neither high glucose concentration nor ketosis had direct effects on Heinz body formation or GSH values. Glutathione decreased to 89% of initial values over the 24-hour period in all samples. High glucose concentration also had no effect on erythrocyte D-lactate production; however, the rate of D-lactate production was slightly increased in samples containing 25 mM β-hydroxybutyrate. Linear L-lactate production confirmed metabolic viability of the erythrocyte suspensions. Samples in high glucose concentration produced L-lactate at a slightly higher (1.2×) rate than did samples in normal glucose concentration. Conclusions High glucose concentration and ketosis do not account directly for oxidative damage and glyoxalase induction in feline erythrocytes in vitro, although high concentrations of β-hydroxybutyrate may stimulate D-lactate formation. (Am J Vet Res 1996;57:463–467)