Pediatrics

Hemolytic Anemia Due to Enzyme Deficiencies

OVERVIEW: What every practitioner needs to know

Are you sure your patient has hemolytic anemia due to enzyme deficiencies? What are the typical findings for this disease?

Hemolytic anemias may be due to factors external to the red blood cell (e.g., circulating autoantibodies to RBCs in Immune mediated hemolysis) or factors intrinsic to the red blood cell (e.g., RBC membrane defects or enzyme defects).

Hemolytic anemia may be due to an enzyme deficiency within the red blood cells. Deficiencies of most of the enzymes of the oxidative hexose monophosphate shunt and the anaerobic Embden-Meyerhof pathways have been described but Glucose 6 Phosphate Dehydrogenase (G-6-PD) deficiency and Pyruvate Kinase (PK) deficiency are the most common.

G-6-PD is an X-linked inherited disorder, and is the most common, affecting persons of Mediterranean, African, Asian, or Middle Eastern decent. Several hundred variants of the G-6-PD enzyme have been described resulting in a wide variation in the severity of the hemolytic anemia.

The clinical presentations of G-6-PD deficiency can vary from acute hemolytic anemia, to chronic hemolytic anemia, neonatal hyperbilirubinemia, or the absence of any clinical symptoms. Individuals who inherit the most common forms of G-6-PD deficiency (G6PD A- or G6PD Mediterranean) generally have no clinical signs or symptoms when they are in a normal and healthy steady state. Symptoms associated with G-6-PD deficiency only become manifest under conditions of oxidative stress. Acute hemolysis can be precipitated in patients with G-6-PD deficiency during the stress induced by a variety of acute infections, upon exposure to certain medications, or after ingestion of fava beans. Under these conditions, a nonspherocytic hemolytic anemia typically develops 1 to 3 days later.

Patients will develop an acute drop in hemoglobin, sudden hemoglobinuria characterized by dark brown to black "Coca-Cola" colored urine, and jaundice. Acute hemolysis may cause back or abdominal pain. The jaundice is secondary to a rise in unconjugated bilirubin.

Common infectious agents precipitating acute hemolysis in patients with G-6-PD deficiency include Salmonella, E-coli, beta-hemolytic Streptococci, viral hepatitis, influenza A, and rickettsia. Certain drugs and exposure to fava beans may also precipitate acute hemolysis.

Hereditary non-spherocytic hemolytic anemia may also occur as a consequence of other enzyme deficiencies. The most common and best defined deficiency of the Embden-Myerhof pathway is pyruvate kinase (PK) deficiency. Clinically affected individuals are homozygote or double heterozygotes for abnormal PK genes. Like G-6-PD deficiency, the clinical spectrum of individuals with the homozygous PK deficiency ranges from severe neonatal anemia and hyperbilirubinemia requiring exchange transfusion to a fully compensated, mild chronic hemolytic anemia. However, some cases of severe PK deficiency require lifelong chronic transfusion with steady state hemoglobin levels as low as 5gm/dl. These patients commonly have chronic jaundice, splenomegaly, and gallstones.

What other disease/condition shares some of these symptoms?

Drug induced hemolytic anemias associated with an unstable hemoglobin may present with clinical laboratory features similar to G-6-PD deficiency, including the hallmark “hemighost” cell on the peripheral blood smear. Hemoglobin electrophoresis and an isopropol stability test may be performed to confirm an unstable hemoglobin. Additionally, the fluorescent spot test is only positive in G-6-PD deficiency (see below).

Other enzyme deficiencies affecting the pentose phosphate shunt, such as deficiencies of GSH synthetase, may mimic G-6-PD deficiency.

Rare causes of hereditary nonsperocytic hemolytic anemia include deficiencies of glucosephosphate isomerase, triosphosphate isomerase, and pyrimidine 5' nucleotidase .

What caused this disease to develop at this time?

The majority of patients with variants of G-6-PD only manifest symptoms under conditions of oxidative stress, most commonly caused by infection due to Salmonella, E-coli, beta-hemolytic Streptococci, influenza A, viral hepatitis, and rickettsial pathogens. Other important metabolic stressors that can induce hemolysys in these patients include administration of an oxidative drug or ingestion of fava beans.

Hemolysis begins 1 to 3 days following the exposure to the oxidative stress, and resolves within 4 to 7 days. Older erythrocytes, having the greater enzyme deficiency, will undergo hemolysis first. Once the population of older erythrocytes has hemolyzed, younger red cells and reticulocytes - which have higher levels of enzyme activity - are able to sustain the oxidative damage without hemolysis. Therefore, despite the ongoing infection or exposure to the oxidative drug, hemolysis will end.

Pathophysiology

The main source of metabolic energy within the red cell comes from glucose which is metabolized through the glycolytic pathway and through the hexose monophospate shunt. G-6-PD is the central enzyme in the hexose monophosphate shunt oxidizing glucose-6 phosphate and converting NADP+ to its reduced form of NADPH, which in turn is required for methemoglobin reductase activity and the maintenance of the normal level of reduced glutathione (GSH). NADPH and GSH protect hemoglobin against oxidative stress and injury.

During an oxidative stress in patients with G-6-PD deficiency, oxidant metabolites denature hemoglobin resulting in RBC injury and the formation of Heinz bodies - precipitated particles of denatured hemoglobin and stromal protein. Red cells containing Heinz bodies are rapidly removed from the circulation when they circulate through the spleen.

What laboratory studies should you request to help confirm the diagnosis? How should you interpret the results?

In the normal steady state, light microscopy of G-6-PD deficient red cells is normal. Under oxidative stress with hemolysis, varying degrees of anemia and reticulocytosis are the main laboratory features.

Heinz bodies are present in erythrocytes on an unstained peripheral smear. The morphologic hallmark of severe oxidative injury to red cells in G-6-PD deficiency is the presence of "hemighost" erythrocytes. Methemalbuminemia, hemoglobinemia, hemoglobinuria, unconjugated bilirubinemia, and decreased haptoglobin and GSH levels may be noted.

The range of hemoglobin in PK deficient individuals is generally 6 to 10gm/dl with a reticulocyte count of 5% to 10%. The peripheral blood smear has nonspecific abnormalities including a macrocytosis.

Diagnosing an RBC enzyme deficiency usually begins with screening tests for G-6-PD and PK activity, followed by an isopropanol stability test to detect an unstable hemoglobin. The simplest screening test is a fluorescent spot test which is based on the fluorescence of NADPH, generated by an adequate amount of G-6-PD enzyme. The screening test may be performed on a whole blood sample or dried blood on filter paper, which makes the test amenable for screening newborns.

If these relatively simple tests are not diagnostic, it is prudent to submit a blood sample to a reference laboratory capable of assaying for all erythrocyte enzyme activities. Elevated red cell 2,3 DPG levels and elevated concentration of 3-phosphoglyceric acid are helpful in the diagnosis of PD deficiency. Reference laboratories are capable of assaying for all erythrocyte enzyme activities. Diagnosis is confirmed by demonstration of decreased enzyme activity through quantitative spectrophotometric analysis.

If you are able to confirm that the patient has hemolytic anemia due to enzyme deficiencies, what treatment should be initiated?

Most individuals with G-6-PD deficiency usually do not require specific therapy. The mainstay in the management of patients with G-6-PD deficiency is the avoidance of oxidative stressors. Individuals should be counseled to avoid drugs that are reported to induce hemolytic episodes. Those drugs most commonly associated with hemolytic episodes in G-6-PD include the following:

Sulfonamides

Trimethoprim-sulfamethoxazole

Nitrofurantoin

Naladixic acid

Chloramphenicol

Antimalarials (primaquine, pamaquine, cholorquine, quinacrine)

Phenacetin

Vitamin K analogs

Methylene bBlue

Probenecid

Acetylsalicylic acid

Phenazopyridine

Occasionally, the rate of hemolysis is severe enough to warrant a blood transfusion. During episodes of hemolysis with hematuria, good renal blood flow and brisk urine output should be maintained to avoid tubular necrosis or renal failure.

Infants with neonatal jaundice may require phototherapy or even exchange transfusion.

Splenectomy is generally not recommended in G-6-PD deficiency as it is often ineffective. However, splenectomy is recommended for patients with PK deficiency who are severely affected and require transfusion either regularly or intermittently. Hematology consult should be obtained.

What are the adverse effects associated with each treatment option?

Following splenectomy in patients with PK deficiency the reticulocyte count may rise to 30% to 90%, the most profound reticulocytosis of any of the enzymopathies. The peripheral blood smear post-splenectomy often shows crenated or spiculated spherocytes and marked polychromasia.

Splenectomy carries with it an increased risk of life threatening sepsis from encapsulated organisms, especially Steptococcus pneumonia. This risk has been reduced by deferring splenectomy until 5-9 years of age (if possible) together with vaccination against S. pneumoniae with both Prevnar and Pneumovax, Haemophilus influenza B, and N. Meningititis, and antibiotic prophylaxis with penicillin post splenectomy. When possible, those vaccines should be administered several weeks before splenectomy to ensure optimal antibody response and protection.

The optimal duration of prophylactic antibiotic therapy post splenectomy is unknown. Recomendations range from at least 5 years post splenectomy to lifetime.

What causes this disease and how frequent is it?

G-6-PD deficiency, with varying degrees of severity, occurs with increased frequency throughout the Mediterranean, Middle East, Asia, and Africa. The mutations encoding G-6-PD are located on the distal long arm of the X chromosome. The normal enzyme is referred to as G-6-PD B; the most common variant forms include G-G-PD A- (very common in West Africa and African-American males) and G-6-PD Mediterranean (see in people of Italian, Greek, Spanish, Arabic, or Jewish descent).

Black males in the United States are commonly affected with a prevalence of approximately 10% (G-6-PD A- variant). The distribution is similar to that of thalassemia and was thought to confer a selective advantage against endemic malaria infection. More than 400 mutations have been identified.

Other clinical manifestations that might help with diagnosis and management

Although G-6-PD newborn screening is not part of the standard recommended panel, several states and military hospitals have included it in their newborn panels. Parents of newborns may, therefore, inquire about the significance of these results.

What complications might you expect from the disease or treatment of the disease?

Patients with G-6-PD who contract viral hepatitis may develop very severe hemolysis and the complication of renal failure.

Acute tubular necrosis may complicate severe hemolytic episodes, underscoring the importance of a forced alkaline diuresis.

Chronic hemolysis is less commonly seen, and is usually caused by a sporadic gene mutation. The severity of the chronic hemolysis can vary from a mild to a transfusion-dependent anemia. Exposure to an oxidative stress can cause superimposed acute hemolysis.

Neonatal hyperbilirubinemia with kernicterus is rarely seen, with neonatal jaundice typically occurring on days 4-7. The mechanism is not completely understood. Although hemolysis may play a role, the hyperbilirubinemia is likely exacerbated by impairment of bilirubin conjugation and clearance by the liver. Infants with Gilbert's disease and G-6-PD deficiency are particularly susceptible to hyperbilirubinemia.

Favism with acute hemolysis after ingestion of fava beans is usually seen in severe G-6-PD (Asian and Mediterranean populations), and only rarely seen in patients with the G-6-PD A- variant. The mechanism is not well understood.

What is the evidence?

Frank, Jennifer E. "Diagnosis and Management of G6PD Deficiency". American Family Physician. vol. Volume 72. October 1, 2005.

Chan, TK.

Ongoing controversies regarding etiology, diagnosis, treatment

The antioxidant properties of Vitamin E have not consistently been shown to provide benefit in the reduction of hemolysis.

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