Click here to view next page of this article


Disorders of Red Blood Cell Enzymes, Membranes and Metabolism

During the process of maturation, the erythrocyte loses some of its functions including protein synthesis and oxidative phosphorylation. As a result, the mature red blood cell (RBC) generates energy through anaerobic glycolysis via the Embden-Meyerhof (EM) pathway; through oxidative glycolysis via the hexose monophosphate (HMP) shunt; and through nucleotide salvage pathways.

Enzyme Deficiencies of the Embden-Meyerhof Pathway

Hexokinase (HK) deficiency

HK is the major determinant of glucose consumption and of the formation of all products of glycolysis since it is the first enzyme in the glycolytic pathway. HK is the least active of the glycolytic enzymes and its activity decreases quickly as the RBCs mature. Decreased HK activity may be secondary to a quantitative defect of a normal enzyme or to the inheritance of a functionally abnormal enzyme. Since young RBCs have increased activity, normal HK activity can be seen with increased reticulocytosis.

Glucosephosphate Isomerase (GPI) Deficiency

GPI deficiency is the third most common hemolytic enzymopathy after G6PD and PK deficiencies. This enzyme is encoded by a single gene in all body cells. However, in the majority of cases, hemolytic anemia of variable severity may be the only clinical manifestation since only mature RBCs.

Phosphofructokinase (PFK) Deficiency

PFK is a tetrameric enzyme that is composed of a varying combination of three basic subunit types referred to as M(muscle), L(granulocytes), and F(fibroblasts, platelets). RBC PFK consists of the L4 tetramer. The clinical manifestation of PFK deficiency is dependent on the expression of these subunits in different tissues. PFK deficiency may be associated with myopathy alone, hemolytic anemia alone or the combination.

Triosephosphate Isomerase (TPI) Deficiency

TPI is coded by a single structural gene located on chromosome 12 and is present in all tissues. This enzyme deficiency appears to have an autosomal recessive inheritance and may lead to a progressive multi-system syndrome. Hemolytic anemia and hyperbilirubinemia occur in early life and affected individuals may require frequent blood transfusions. Within the first year of life, neurological manifestations such as spasticity,

Phosphoglycerate Kinase (PGK) Deficiency

This is the only X-linked enzymopathy affecting the EM pathway. PGK deficient males develop normally until early childhood. However, behavioral aberrations, motor regression, and cerebellar tremors become apparent within the first four to five years of life.

Pyruvate Kinase (PK) Deficiency

This is the most common of the enzymatic deficiencies involving the EM pathway. This enzymopathy has a world-wide, multi-racial distribution with over three hundred reported cases. This enzyme catalyzes the conversion of phosphoenol pyruvate (PEP) to pyruvate, and during this process ATP is generated. PK deficiency results in impaired glycolysis and diminished capacity to produce ATP. Clinical expression is highly variable.

Defects of Nucleotide Metabolism

Pyrimidine 5'-Nucleotide (P5N)Deficiency

Ribosomal RNA of reticulocytes is degraded into 5'-nucleotides like cytidine, thymidine and uridine monophosphate by the P5N enzyme. In P5N deficiency, these non-diffusible catabolites form large aggregates which are seen as basophilic stippling on wright-stained blood smear. In lead poisoning, there is inhibition of this enzyme and this results in basophilic stippling also. In the screening test for P5N activity, RBCs are extracted by perchloric acid to remove purine.

Adenylate Kinase (AK) Deficiency

This enzyme catalyzes the formation of ATP and adenosine monophosphate (AMP) from two moles of adenosine diphosphate (ADP). This is a very rare enzymopathy and hemolytic anemia of moderate severity has been described in few subjects. In this enzyme assay, AK reaction is linked to pyruvate kinase and LDH reactions. The decrease in optical density at 340rim due to oxidation of NADH to NAD is measured.

Hyperactivity of Adenosine Deaminase (AD)

This enzyme catalyzes the deamination of adenosine to inosine. AD deficiency has been associated with immune deficiency states and a 40 to 70 fold increased activity of the enzyme has been associated with chronic hemolytic anemia. This enzyme is transmitted by autosomal dominant genes. The enzyme activity is measured by the decrease in optical density due to the conversion of adenosine to inosine at 265 nm.

Enzyme Deficiencies of the Hexose Monophosphate Shunt (HMP)

Glucose-6 Phospate-Dehydrogenase Deficiency (G6PD)

This is the most prevalent inborn error of erythrocytes metabolism affecting more than 150 million people in many geographic areas around the world. The enzymatically active form of G6PD is either a dimer or a tetramer. There is only one structural gene for G6PD in the human genome and it is located in the long arm of the X-chromosome (band Xq28).

Defects in Glutathione Metabolism

Glutathione Reductase (GSSG-R) Deficiency

The activity of this enzyme is dependent on riboflavin in the diet. Patients with riboflavin deficiency, have low enzyme activity that can be restored to normal level with the administration of riboflavin. Genetically determined GSSG-R deficiency has been reported in only few patients. The fluorescent-spot test can be used as a screening method.

Glutathione Peroxidase (GSH-Px) Deficiency

Glutathione peroxidase catalyzes the oxidation of reduced glutathione by peroxides such as hydrogen peroxide and organic hydroperoxides. This enzymopathy results in mild hemolytic anemia. Acquired deficiency of this enzyme has been reported in patients with iron and selenium deficiencies, cirrhosis of the liver, and in Glanzmannís thrombasthenia. In this enzyme assay, GSH-Px reaction is linked to the GSSG-R reaction and a decrease in the absorbance due to the oxidation of NADPH is then measured.

Gamma Glutamyl Cysteine Synthetase (GGCS) Deficiency

This enzyme catalyzes the formation of gamma-glutamyl-cysteine. Deficiency of this enzyme causes a mild congenital hemolytic anemia associated with myopathy, neuropathy, spinocerebellar degeneration, and psychosis developing in early adulthood.

Glutathione Synthetase (GH) Deficiency

Glutathione formation is catalyzed by glutathione synthetase. Severe GSH deficiency has been reported to cause a syndrome characterized by mild hemolytic anemia, metabolic acidosis, cerebral and cerebellar degeneration, mental retardation, and 5-oxoprolinuria. Episodes of hemolysis can be precipitated by consumption of fava beans. In the screening test for this enzyme deficiency, GSH concentration in RBCs is assayed.

Enzyme Deficiencies of Doubtful Clinical Significance

Congenital deficiencies of the RBC enzymes 2-3 diphosphoglycerate mutase, glyceraldehyde-3-phosphate dehydrogenase, enolase, aldolase, lactic dehydrogenase and ATPase have been reported.

Antenatal Diagnosis

Antenatal diagnosis can be achieved by measuring fetal RBC enzyme activity as early as seventeen weeks of pregnancy. Gene probes are available for many RBC enzymopathies. Lestas and colleagues, characterized the normal enzyme activities of more than twelve glycolytic enzymes in fetal RBCs at 17-24 weeks of gestation, and these reference values can be used in the screening for these enzymopathies.