Unstable Hemoglobinopathies

Medical Experts

Contributor

Agarwal

Professor of Pathology (Clinical), University of Utah

Medical Director, Hematopathology and Special Genetics, ARUP Laboratories

The unstable hemoglobinopathies are rare inherited hemoglobinopathies in which the solubility of hemoglobin (Hb) is altered, which often leads to Hb precipitates (Heinz bodies), reduced red blood cell (RBC) lifespan, and hemolytic anemia. The clinical severity and presentation of the unstable hemoglobinopathies vary widely, and they are frequently underdiagnosed or misdiagnosed. Laboratory testing for unstable hemoglobinopathies consists of an initial evaluation, specialized testing, and genetic testing.

Quick Answers for Clinicians

What is the pathophysiology of unstable hemoglobinopathy?

Most unstable hemoglobinopathies arise from point variants in one of the globin genes and are inherited as autosomal dominant disorders, although de novo variants and other inheritance patterns have been observed.cite#4479 These variants lead to changes in hemoglobin (Hb) structure that cause the Hb to become unstable and precipitate within red blood cells (RBCs). These precipitates, known as Heinz bodies, bind to the membrane of the RBC, causing premature breakdown of the affected cell within the spleen. Hbs Koln and Zurich are the most prevalent unstable Hbs; see the Human Hemoglobin Variants and Thalassemias database for more information on other variants.

How do unstable hemoglobinopathies present?

Unstable hemoglobinopathies may present with congenital Heinz body hemolytic anemia and pigmented urine. Heinz bodies are not specific to unstable hemoglobinopathies and may also be found in enzymopathies and thalassemia. Neonatal hemoglobinopathy syndromes related to pathogenic gamma globin variants (hemoglobin [Hb] Poole and Hb Hasharon) may present with hemolysis, jaundice, and anemia. These syndromes resolve with aging as adult Hb replaces fetal Hb. Other presentations include congenital anemia, mild or minimal anemia with reticulocytosis that is out of proportion to the circulating Hb, and acute hemolysis induced by drugs (eg, sulfonamides) or other oxidants.

What are the hyperunstable hemoglobins, and what laboratory testing challenges do they present?

The hyperunstable hemoglobins (Hbs) are more unstable than typical unstable Hbs, and are thus very rapidly destroyed, which prevents their detection in hemolysate. Hyperunstable hemoglobinopathies present similarly to severe thalassemia but are inherited in an autosomal dominant pattern.

The hyperunstable Hbs are not readily detected by the Hb assays typically used to detect unstable Hbs, such as stability testing, isoelectric focusing (IEF), or high-performance liquid chromatography (HPLC). Because they are difficult to detect, genetic testing is often required for diagnosis.

What is glucose-6-phosphate dehydrogenase deficiency, and how is it diagnosed?

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzymatic disorder of red blood cells (RBCs) and can result in RBC hemolysis. The most frequent condition associated with G6PD deficiency is hemolytic anemia, which can be triggered by bacterial or viral infections, certain antibiotics and malaria medications, or favism (a reaction to the consumption of fava beans or inhalation of fava pollen). As in unstable hemoglobinopathies, Heinz bodies can be observed in G6PD deficiency. G6PD deficiency is diagnosed using enzymatic and molecular genetic tests. For more information, see the Glucose-6-Phosphate Dehydrogenase (G6PD) Deficiency Test Fact Sheet.

Indications for Testing

Testing for unstable hemoglobinopathies is useful in cases of unexplained hemolytic anemia or when there is a known familial pathogenic Hb variant.

Laboratory Testing

Initial Evaluation

The first step in the evaluation of a suspected unstable hemoglobinopathy is a CBC with peripheral smear and reticulocyte count. The initial evaluation usually reveals normocytic anemia (ranging from mild to severe) with nonspecific findings of hemolysis.^, Hemolysis may be chronic or may be induced by oxidative stress, such as exposure to sulfonamide drugs. Brisk reticulocytosis is generally observed. “Bite” or “blister” cells may be observed in the peripheral smear. In severe cases, particularly when hyperunstable Hbs are present, the peripheral smear may reveal anisocytosis, basophilic stippling, Howell-Jolly bodies, and microspherocytes.

If a hemolytic anemia workup is performed, characteristic findings include decreased Hb, elevated unconjugated bilirubin, elevated lactate dehydrogenase, decreased haptoglobin, and disproportionately elevated aspartate aminotransferase (compared with alanine aminotransferase) concentrations.

Specialized Testing

Stability Testing

Stability testing is performed by incubating with isopropanol (isopropanol stability test) or treating with heat at 50° Celsius (heat stability test). A precipitate will form if unstable Hbs are present. A false-positive result may occur if there is much HbF present (as is typical in neonates) or in cases of methemoglobinemia.

Heinz Bodies

Heinz bodies may be detected by supravital staining of erythrocytes in peripheral blood. However, the absence of Heinz bodies does not rule out unstable hemoglobinopathy. Furthermore, Heinz bodies are not specific to the unstable hemoglobinopathies and may be observed in other conditions that lead to Hb precipitates (eg, glucose-6-phosphate dehydrogenase [G6PD] deficiency). Results of this test are unreliable in infants younger than 6 months of age.

Hemoglobin Assays

Hb electrophoresis (eg, isoelectric focusing [IEF]) or high-performance liquid chromatography (HPLC) may reveal increased HbA2 and HbF. However, unstable Hb variants, particularly hyperunstable variants, undergo rapid denaturation and degradation within the erythrocyte, and remaining Hbs may appear relatively normal. Therefore, a normal Hb electrophoresis or HPLC test result does not rule out an unstable hemoglobinopathy.^, Hb electrophoresis should not be repeated in patients with a previous test result who do not require intervention or monitoring.

Genetic Testing

Globin gene sequencing is the only technique that may detect some of the rare unstable Hbs. Therefore, sequencing of the globin genes, including the gamma globin gene in affected neonates, is often needed for a definitive diagnosis.^,