Diagnosis and epidemiology of red blood cell enzyme disorders

The red blood cell possess an active metabolic machinery that provides the cell with energy to pump ions against electrochemical gradients, to maintain its shape, to keep hemoglobin iron in the reduced (ferrous) form, and to maintain enzyme and hemoglobin sulfhydryl groups. The main source of metabolic energy comes from glucose. Glucose is metabolized through the glycolytic pathway and through the hexose monophosphate shunt. Glycolysis catabolizes glucose to pyruvate and lactate, which represent the end products of glucose metabolism in the erythrocyte. Adenosine diphosphate (ADP) is phosphorylated to adenosine triphosphate (ATP), and nicotinamide adenine dinucleotide (NAD)+ is reduced to NADH in glycolysis. 2,3Bisphosphoglycerate, an important regulator of the oxygen affinity of hemoglobin, is generated during glycolysis by the Rapoport-Luebering shunt. The hexose monophosphate shunt oxidizes glucose-6-phosphate, reducing NADP+ to reduced nicotinamide adenine dinucleotide phosphate (NADPH). The red cell lacks the capacity for de novo purine synthesis but has a salvage pathway that permits synthesis of purine nucleotides from purine bases. The red cell contains high concentrations of glutathione, which is maintained almost entirely in the reduced state by NADPH through the catalytic activity of glutathione reductase. Glutathione is synthesized from glycine, cysteine, and glutamic acid in a two-step process that requires ATP as a source of energy. Catalase and glutathione peroxidase serve to protect the red cell from oxidative damage. Erythrocyte enzyme deficiencies may lead to hemolytic anemia(1); expression of the defect in other cell lines may lead to pathologic changes such as myopathy and neuromuscular abnormalities. Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common erythrocyte enzyme defect(2). In some populations, more than 20 percent of people may be affected by this enzyme deficiency. In common polymorphic forms, such as G6PD Aor G6PD Mediterranean, hemolysis occurs only during the stress imposed by infection or administration of oxidative drugs, and in some individuals upon ingestion of fava beans. Patients with uncommon, functionally very severe, genetic variants of G6PD experience chronic hemolysis, a disorder designated hereditary nonspherocytic hemolytic anemia (HNSHA).(3) Hereditary nonspherocytic hemolytic anemia also occurs as a consequence of other enzyme deficiencies, the most common of which is pyruvate kinase (PK) deficiency.(4-6) Deficiencies of glucosephosphate isomerase(6), triosephosphate isomerase(7), and pyrimidine 52 -nucleotidase deficiency(8) are included among the relatively rare causes of HNSHA. In the case of some deficiencies, notably those of glutathione synthetase(9), triosephosphate isomerase(7), and phosphoglycerate kinase(10), the defect is expressed throughout the body, and neurologic and other defects may be a prominent part of the clinical syndrome. Diagnosis is best achieved by determining red cell enzyme activity with a quantitative assay or a screening test(11). However, since the maturation of reticulocytes into erythrocytes is associated with a rapid decrease in the activity of several enzymes reticulocytosis, which is normally present, represents an important pitfall in the diagnosis of red cell enzyme deficiencies.(12) Except for the basophilic stippling of erythrocytes which is characteristic for pyrimidine 5 nucleotidase deficiency, red cell morphology is of little or no help in differentiating red cell enzyme deficiencies from another. Many different mutations have been defined in most of the enzyme deficiencies, in particular in PK deficiency (www.pklrmutationdatabase.com). Accurate diagnosis is necessary for genetic counseling and is helpful in recommendations for treatment, since patients with some enzyme deficiencies tend to respond more favorably to splenectomy than do others. Most red cell enzyme defects are transmitted as autosomal recessive disorders, while G6PD and phosphoglycerate kinase deficiencies are X linked. There are no exact and verified figures regarding the occurrence of red blood cell enzyme disorders, other than the number of cases reported in literature (Table 1). Basically, this is due to the lack of a certified registry. Also, like in disorders of the red cell membrane, some enzyme disorders will be difficult to identify because they are either very rare or clinically mild. This latter fact may, for instance, explain the discrepancy between the estimated number of cases affected by PK deficiency (i.e. 1:20,000 in the general white population) and the true number of identified cases. The European NEtwork for Rare and Congenital Anaemias (ENERCA) enabled a comparison of these numbers.(13)It was concluded that both in The Netherlands and Italy, 2 countries with a large and well-characterized database of patients with PK deficiency the true frequency was, in fact, about 10 times lower than predicted. As stated, this may either be due to a high number of patients showing a mild to very mild clinical picture or to a lack of awareness, or both. A similar situation might be applicable to haemolytic anaemia due to pyrimidine-5’-nucleotidase deficiency. A recently conducted survey by ENERCA also brought to light that another important issue contributes to the limited knowledge on epidemiology of red cell enzyme disorders. This concerns the fact there are Correspondence: Richard Van Wijk E-mail: r.vanwijk@umcutrecht.nl

The red blood cell possess an active metabolic machinery that provides the cell with energy to pump ions against electrochemical gradients, to maintain its shape, to keep hemoglobin iron in the reduced (ferrous) form, and to maintain enzyme and hemoglobin sulfhydryl groups.The main source of metabolic energy comes from glucose.Glucose is metabolized through the glycolytic pathway and through the hexose monophosphate shunt.Glycolysis catabolizes glucose to pyruvate and lactate, which represent the end products of glucose metabolism in the erythrocyte.Adenosine diphosphate (ADP) is phosphorylated to adenosine triphosphate (ATP), and nicotinamide adenine dinucleotide (NAD)+ is reduced to NADH in glycolysis.2,3-Bisphosphoglycerate, an important regulator of the oxygen affinity of hemoglobin, is generated during glycolysis by the Rapoport-Luebering shunt.The hexose monophosphate shunt oxidizes glucose-6-phosphate, reducing NADP+ to reduced nicotinamide adenine dinucleotide phosphate (NADPH).The red cell lacks the capacity for de novo purine synthesis but has a salvage pathway that permits synthesis of purine nucleotides from purine bases.
The red cell contains high concentrations of glutathione, which is maintained almost entirely in the reduced state by NADPH through the catalytic activity of glutathione reductase.Glutathione is synthesized from glycine, cysteine, and glutamic acid in a two-step process that requires ATP as a source of energy.Catalase and glutathione peroxidase serve to protect the red cell from oxidative damage.
Erythrocyte enzyme deficiencies may lead to hemolytic anemia(1); expression of the defect in other cell lines may lead to pathologic changes such as myopathy and neuromuscular abnormalities.Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common erythrocyte enzyme defect (2).In some populations, more than 20 percent of people may be affected by this enzyme deficiency.In common polymorphic forms, such as G6PD A-or G6PD Mediterranean, hemolysis occurs only during the stress imposed by infection or administration of oxidative drugs, and in some individuals upon inges-tion of fava beans.Patients with uncommon, functionally very severe, genetic variants of G6PD experience chronic hemolysis, a disorder designated hereditary nonspherocytic hemolytic anemia (HNSHA).( 3) Hereditary nonspherocytic hemolytic anemia also occurs as a consequence of other enzyme deficiencies, the most common of which is pyruvate kinase (PK) deficiency.(4-6)Deficiencies of glucosephosphate isomerase (6), triosephosphate isomerase (7), and pyrimidine 52 -nucleotidase deficiency(8) are included among the relatively rare causes of HNSHA.In the case of some deficiencies, notably those of glutathione synthetase(9), triosephosphate isomerase (7), and phosphoglycerate kinase (10), the defect is expressed throughout the body, and neurologic and other defects may be a prominent part of the clinical syndrome.
Diagnosis is best achieved by determining red cell enzyme activity with a quantitative assay or a screening test (11).However, since the maturation of reticulocytes into erythrocytes is associated with a rapid decrease in the activity of several enzymes reticulocytosis, which is normally present, represents an important pitfall in the diagnosis of red cell enzyme deficiencies.(12) Except for the basophilic stippling of erythrocytes which is characteristic for pyrimidine 5 nucleotidase deficiency, red cell morphology is of little or no help in differentiating red cell enzyme deficiencies from another.
Many different mutations have been defined in most of the enzyme deficiencies, in particular in PK deficiency (www.pklrmutationdatabase.com).Accurate diagnosis is necessary for genetic counseling and is helpful in recommendations for treatment, since patients with some enzyme deficiencies tend to respond more favorably to splenectomy than do others.Most red cell enzyme defects are transmitted as autosomal recessive disorders, while G6PD and phosphoglycerate kinase deficiencies are X linked.
There are no exact and verified figures regarding the occurrence of red blood cell enzyme disorders, other than the number of cases reported in literature (Table 1).Basically, this is due to the lack of a certified registry.Also, like in disorders of the red cell membrane, some enzyme disorders will be difficult to identify because they are either very rare or clinically mild.This latter fact may, for instance, explain the discrepancy between the estimated number of cases affected by PK deficiency (i.e.1:20,000 in the general white population) and the true number of identified cases.The European NEtwork for Rare and Congenital Anaemias (ENERCA) enabled a comparison of these numbers.(13)It was concluded that both in The Netherlands and Italy, 2 countries with a large and well-characterized database of patients with PK deficiency the true frequency was, in fact, about 10 times lower than predicted.As stated, this may either be due to a high number of patients showing a mild to very mild clinical picture or to a lack of awareness, or both.A similar situation might be applicable to haemolytic anaemia due to pyrimidine-5'-nucleotidase deficiency.
A recently conducted survey by ENERCA also brought to light that another important issue contributes to the limited knowledge on epidemiology of red cell enzyme disorders.This concerns the fact there are currently only a very limited number of laboratories in the EU capable of performing the necessary tests, either on the biochemical level or on the genetic level, required to diagnose red cell enzyme disorders.Whereas a considerable number of laboratories are performing diagnostic tests for detection of the two most frequently occurring red cell enzyme disorders, i.e. deficiencies of G6PD and PK, only very few laboratories offer the complete panel of tests for detection of theother 12 rare enzyme disorders of the red blood cell (Table 1).This fact probably strongly contributes to the relatively high amount of patients with hereditary haemolytic anaemia that remain undiagnosed.In addition, it affirms the belief among ENERCA members that the true worldwide population frequency of the rare and very rare enzyme disorders of the red blood cell may, in fact, be significantly higher than that reported in literature.This may for instance be true for a deficiency of glutathione synthetase.Further investigations into the frequency of red blood cell enzyme deficiencies are therefore warranted and should be encouraged.