Pyruvate Kinase Deficiency Workup

Updated: Apr 28, 2014
  • Author: Hassan M Yaish, MD; Chief Editor: George T Griffing, MD  more...
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Workup

Approach Considerations

Prenatal enzymatic testing is not optimal, because a large amount of fetal blood is required, and the test has a high rate of false-negative results.

The minimal tests required to guide the investigation of pyruvate kinase deficiency include the following:

  • Complete blood count (CBC)
  • Differential blood count
  • Reticulocyte count
  • Serum bilirubin level study
  • Peripheral blood film examination

Normochromic, normocytic, or macrocytic anemia, together with reticulocytosis in the absence of blood loss, is suggestive of hemolysis. A negative Coombs test result helps to exclude immune hemolysis.

An elevated direct bilirubin level in the presence of indirect hyperbilirubinemia is not unusual in individuals with pyruvate kinase deficiency and does not necessarily indicate cholestasis, primary liver disease, or biliary obstruction. [22, 23]

Other findings include the following:

  • Normoblastic erythroid hyperplasia of the bone marrow
  • Extramedullary hematopoiesis
  • Splenic and hepatic hemosiderosis and splenic congestion
  • Reticuloendothelial hyperplasia
  • Erythrophagocytosis
  • Increased iron stores

Enzyme activity rate

The enzyme activity rate in most patients is 5-25% of normal, but false-negative results can occur, especially when the deficiency results from a compound heterozygous mutation; kinetic variables are not measured accurately under such circumstances. Measurement of the intermediates (2,3-diphosphoglycerol and glucose-6-phosphate) proximal to the enzyme defect helps to confirm the diagnosis.

Imaging studies

In severe anemia, radiographs may demonstrate findings of marrow expansion. Ultrasonography is occasionally required to document gallbladder stones, which are known to complicate all forms of hemolytic anemias.

Follow-up

Periodic follow-up care is required in patients with pyruvate kinase deficiency to monitor their hemoglobin level and reticulocyte count and to look for possible gallstones.

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Cell Indices

The hematocrit value in pyruvate kinase deficiency ranges from 17-37%. Lower values occur in early childhood and during the neonatal period, with a 3- to 9-point rise after early childhood. Erythrocytes are usually normochromic and normocytic. High reticulocyte counts may raise the mean cell volume, giving the impression that the anemia is macrocytic. Monitor the hematocrit value carefully during times of physiologic stress.

The reticulocyte count in pyruvate kinase deficiency may be increased by 5-15%. Paradoxically, a reticulocyte count of as high as 70% may occur after splenectomy. Leukocyte and platelet counts range from normal to slightly increased, unless the patient develops hypersplenism, which may result in lower cell lines.

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Cell Morphology

Morphologic abnormalities are not a prominent finding in pyruvate kinase deficiency, but hallmarks of accelerated erythropoiesis, such as polychromatophilia, anisocytosis, poikilocytosis, and nucleated red blood cells, may be present.

Following splenectomy, the following may be observed:

  • Siderocytes
  • Target cells
  • Pappenheimer bodies
  • Howell-Jolly bodies
  • Crenated red blood cells
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Hemoglobin Indices

Concurrent with the hematocrit value, the hemoglobin concentration in pyruvate kinase deficiency varies from 6-12 g/dL, with a lower concentration early in life. Hemoglobin electrophoresis reveals normal hemoglobin, as well as normal levels of F and A2 hemoglobins.

Hemoglobin metabolic indices

Indirect hyperbilirubinemia reflects the severity of the hemolytic process. levels of 6 mg/dL are not uncommon, and levels greater than 20 mg/dL have been reported. Haptoglobin is reduced in proportion to disease severity.

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Hemolytic Anemia Studies

Depending on the severity of the anemia, erythrocytes have a moderately to severely reduced lifespan in pyruvate kinase deficiency. Radiochromium labeling reveals an immediate period of destruction, followed by a shortened lifespan for the remainder of labeled cells. The results of this test can help to determine whether a patient is a candidate for splenectomy, because a high rate of immediate destruction suggests significant splenic activity.

Additional findings in hemolytic anemia include the following:

  • Osmotic fragility of erythrocytes is normal
  • Coombs test result is negative
  • Ham test result is negative
  • Donath-Landsteiner antibody is absent
  • Cold agglutinins are absent
  • Incubated Heinz body formation is usually abnormal
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Enzyme and PCR Assays

A precise diagnosis of pyruvate kinase deficiency depends on the detection of the deficient enzyme. [25] Enzyme assays, as well as deoxyribonucleic acid (DNA) analysis with a polymerase chain reaction (PCR) assay or single-strand conformation polymorphism, can be used to confirm the diagnosis and identify the carrier state, if necessary.

Enzyme assays, however, are not always accurate; the typical selective removal of very deficient red cells from the circulation can leave only normal cells. The assay may also be affected by the fact that pyruvate kinase activity is usually normal in white cells, platelets, and other tissues in patients with pyruvate kinase deficiency hemolytic anemia.

Deoxyribonucleic acid (DNA) analysis can be limited as well, owing to the large number of gene mutations that cause pyruvate kinase deficiency.

Targeted next-generation sequencing (NGS) has been very valuable in confirming the diagnosis of pyruvate kinase deficiency, as well as identifying new novel undescribed mutations in the gene, which result in hemolytic anemia.

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