Cytochrome b5 Reductase T116S Mutation and Hemolysis in Sickle Cell Disease.

Deficiency of NADH-cytochrome b5 reductase (cytb5r, EC 1.6.2.2) is responsible for congenital methemoglobinemia. This enzyme exists in soluble and membrane-bound forms. The soluble erythrocytic cytb5r isoenzyme is involved in cytochrome b5 reduction and in erythrocyte methemoglobin reduction; the membrane-bound microsomal enzyme participates in a fatty acid desaturation complex and in drug metabolism. The cytb5r isoforms are the product of a single gene locus, DIA1 (or CYB5R3), on chromosome 22. More then 40 mutations which cause methemoglobinemia have been reported to date; the majority are missense mutations and are associated with mild type I methemoglobinemia. The CYBR5 T116S mutation is the most common genetic polymorphism among African Americans known (gene frequency as high as 20%) and it has not yet been detected in other ethnic and racial groups. This polymorphism is not associated with methemoglobinemia and its functional significance is not yet known. We studied the relationship of CYBR5 T116S with the degree of hemolysis and the tricuspid regurgitation velocity (which correlates with systolic pulmonary artery pressure) in patients with sickle cell disease.

Two hundred sixty one children and adolescents with hemoglobin SS were recruited at three tertiary medical centers and studied at steady state. Patients with other sickle genotypes were excluded from this analysis of CYBR5 T116S. Principal component analysis was used to develop a hemolytic component from reticulocyte count and concentrations of lactate dehydrogenase, aspartate aminotransferase and bilirubin. PCR was used to determine the presence of the CYBR5 T116S mutation. Multivariate models were employed to determine the independent effects of this genotype on degree of hemolysis and tricuspid regurgitation velocity.

Ninety-eight of the patients (38%) were CYBR5 T116S heterozygotes and 26 (10%) were homozygotes, consistent with Hardy-Weinberg equilibrium. Both heterozygosity (beta = -0.4) and homozygosity (beta = -0.5) were associated with reduction in the hemolytic component (N = 261; P for trend = 0.002) (Figure 1). This relationship persisted after adjusting for α-thalassemia, hemoglobin F percent and hydroxyurea treatment in a subset of 113 patients with all of this information available (P for trend = 0.037) and it also persisted in a subset of 87 patients with no α-globin gene deletion who were not being treated with hydroxyurea (P for trend = 0.029). In none of these analyses did G6PD^-202/-376 have an effect on hemolysis. Both heterozygosity (beta = -0.04) and homozygosity (beta = -0.14) for the CYBR5 T116S mutation were also associated with lower tricuspid regurgitation velocity (P for trend = 0.024).

CYBR5 T116S is a common polymorphism among patients with sickle cell disease that appears to be associated with less hemolysis and lower tricuspid regurgitation velocity. We speculate that this polymorphism may be related to a previously reported subpopulation of African Americans with increased cytochrome b5 reductase activity, and that increased anti-oxidant activity may explain the polymorphism's hemolysis-reducing effect. Functional studies to investigate this possibility are planned.

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No relevant conflicts of interest to declare.