Haemoglobin variants

Common haemoglobin variants

• Haemoglobin S: this is the main haemoglobin in people with sickle cell anaemia. About 8% of African Americans carry the Hb S mutation in one of their two beta genes (0.15% of African Americans have sickle cell anaemia). People with haemoglobin variant S have two abnormal beta chains (βS) and two normal alpha chains (α). Haemoglobin S deforms the red blood cells, which take on a sickle shape when exposed to low levels of oxygen (as happens, for example, during exercise). Sickle-shaped red cells can block small blood vessels, causing pain and impairing circulation, reduce the oxygen-carrying capacity of the red cells and shorten the life of the cells. A single abnormal beta copy (βS) does not cause symptoms unless it is combined with another haemoglobin mutation, such as the one that causes HbC (βC).
• Haemoglobin C: about 2-3% of people of West African descent are heterozygous for haemoglobin C (i.e. they have one copy of βC). Haemoglobin C disease (seen in homozygous people, who have two copies of βC) is rare and relatively mild. It usually causes mild haemolytic anaemia and mild to moderate enlargement of the spleen.
• Haemoglobin E: haemoglobin E is one of the most common beta-chain haemoglobin variants in the world. It is prevalent in South-East Asia, especially Cambodia, Laos and Thailand, and in people descended from South-East Asian populations. People who are homozygous for Hb E (i.e. who have two copies of βE) usually have moderate haemolytic anaemia, microcytosis and moderate enlargement of the spleen. A single copy of the haemoglobin E gene does not cause symptoms unless it is combined with another mutation, such as the one that causes beta thalassaemia trait.
• Less common haemoglobin variants
• There are many other variants. Some are silent – they cause no signs or symptoms – while others affect the function and/or stability of the haemoglobin molecule. Examples of these variants include haemoglobin D, haemoglobin G, haemoglobin J, haemoglobin M and haemoglobin Constant Spring, a mutation in the alpha globin gene that results in an alpha (α) chain of abnormal length and an unstable haemoglobin molecule. Further examples of beta-chain variants are:
• Haemoglobin F: Hb F is the main haemoglobin produced by the fetus, and its role is to carry oxygen efficiently in a low-oxygen environment. Production of haemoglobin F stops at birth and falls to adult levels by the age of 1-2 years. Hb F may be raised in several congenital diseases. Levels may be normal to high in beta thalassaemia and are often raised in people with sickle cell anaemia and sickle cell–beta thalassaemia. People with sickle cell disease and raised Hb F levels often have a milder phenotype, because haemoglobin F inhibits the sickling of red cells. Hb F levels are also raised in a rare condition called hereditary persistence of fetal haemoglobin (HPFH). This is a group of inherited disorders in which Hb F levels are raised without the signs or clinical features of thalassaemia. Different mutations cause HPFH in different ethnic groups. Hb F may also be raised in some acquired conditions characterised by impaired red cell production. Leukaemias and other myeloproliferative disorders are also often associated with raised Hb F levels.
• Haemoglobin H: Hb H is an abnormal haemoglobin found in some cases of thalassaemia. It is made up of four beta (β) globin chains and is produced in response to a severe shortage of alpha (α) chains. Although each beta (β) globin chain is normal, the tetramer of four beta chains does not work normally. It has an increased affinity for oxygen, which it holds on to instead of releasing it to the tissues and cells.
• Haemoglobin Barts: Hb Barts develops in the fetus with alpha thalassaemia. It is formed from four gamma (γ) protein chains when alpha chains are lacking, in a similar way to the formation of haemoglobin H. Hb Barts disappears soon after birth as production of gamma chains falls.
• A person may inherit two different abnormal genes, one from each parent. This is known as being a compound heterozygote or double heterozygote. Some clinically significant combinations are listed below.
• Haemoglobin SC disease. Inheriting one beta S gene and one beta C gene causes haemoglobin SC disease. Affected people have mild haemolytic anaemia and moderate enlargement of the spleen. People with haemoglobin SC disease can develop the same vaso-occlusive complications (blockage of blood vessels) seen in sickle cell anaemia, although most cases are less severe.
• Sickle cell–haemoglobin D disease. People with sickle cell–haemoglobin D disease have inherited one copy of the haemoglobin S gene and one of haemoglobin D-Los Angeles (or D-Punjab). These patients may have occasional sickle cell crises and moderate haemolytic anaemia.
• Haemoglobin E–beta thalassaemia. People who are double heterozygous for haemoglobin E and beta thalassaemia have anaemia that can range in severity from mild (or no symptoms) to severe.
• Sickle cell–beta thalassaemia varies in severity depending on the type of beta thalassaemia mutation inherited. Some mutations reduce the production of beta haemoglobin (beta+), while others eliminate it completely (beta0). Sickle cell–beta+ thalassaemia tends to be less severe than sickle cell–beta0 thalassaemia. Patients with sickle cell–beta0 thalassaemia tend to have more irreversibly sickled cells, more frequent vaso-occlusive problems and more severe anaemia than patients with sickle cell–beta+ thalassaemia. It is often difficult to distinguish between sickle cell anaemia and sickle cell–beta0 thalassaemia.

How are they tested?

Laboratory tests
Laboratory testing for haemoglobin variants involves checking whether the red blood cells (RBCs) are “normal”, assessing the haemoglobin contained in the RBCs and analysing the relevant gene mutations or deletions. Each test provides a piece of the puzzle, giving clinicians important information about the variants that may be present. The tests ordered for haemoglobin variants are also used to investigate thalassaemia. Looking for both is important because thalassaemia is sometimes inherited together with a haemoglobin variant.

FBC (full blood count). The FBC is a snapshot of the cells circulating in the blood. Among other things, the FBC will tell the doctor how many red cells are present and how much haemoglobin they contain, and will give an assessment of the size and shape of the red cells present. The mean corpuscular volume (MCV) is a measure of red cell size. A low MCV is often the first sign of thalassaemia. If the MCV is low and iron deficiency has been ruled out, the person may be a carrier of thalassaemia trait or have one of the haemoglobin variants that cause microcytosis (for example Hb E).

Blood smear (also called a peripheral smear, and a manual differential when the white cells are examined). In this test, an experienced laboratory specialist examines under the microscope a thin layer of blood spread on a slide and treated with a special stain. The number and type of white cells, red cells and platelets can be determined and assessed to see whether they are normal and mature. A variety of diseases affect normal blood cell production. Red cells may be:

• Microcytic
• Hypochromic
• Varied in size (anisocytosis) and shape (poikilocytosis)
• Nucleated (not normal in a mature RBC).
• Unevenly filled with haemoglobin (producing “target cells”, which look like a bullseye under the microscope).

The greater the percentage of red cells that look abnormal, the greater the likelihood of an underlying disease and of impaired oxygen-carrying capacity.

Haemoglobin variant tests. These tests identify the type and measure the relative amounts of the haemoglobins present in red cells. Most common variants can be identified using one of these tests or a combination of them. The relative amount of each haemoglobin variant detected can help diagnose combinations of haemoglobin variants and thalassaemia (compound heterozygotes).

DNA analysis. This test is used to look for deletions and mutations in the genes that produce the alpha and beta globins. Family studies may be carried out to assess carrier status and the types of mutations present in other family members. DNA testing is not routine, but it can be used to diagnose haemoglobin variants and thalassaemia and to determine carrier status.

Why are they done?
Testing for haemoglobin variants is carried out to:

• Screen newborns for common haemoglobin variants. In all US states this has become a standard part of the newborn screening programme. Newborns with variants such as Hb S can benefit from early detection and treatment.
• Prenatal screening is also carried out in some areas where mothers are at high risk: those from an ethnic background associated with a high prevalence of haemoglobin variants (such as people of African descent) and those with affected family members. Screening may also be offered together with genetic counselling before pregnancy to determine the potential carrier status of family members.
• Identify variants in family members without symptoms who have an affected child.
• Identify haemoglobin variants in people with unexplained symptoms such as anaemia, microcytosis and/or hypochromia. The test may also be requested as part of an investigation for anaemia.

What should I know?
Blood transfusions can interfere with haemoglobin variant testing. A patient should wait several months after a transfusion before having the test. However, in patients with sickle cell anaemia, the test may be carried out after a transfusion to check whether enough normal haemoglobin has been transfused to reduce the risk of red cell damage due to sickling.

Since prenatal screening programmes began to include haemoglobin variant testing, hundreds of carrier children have been identified. This is due to improved technology, not to an increase in the prevalence of the gene mutations. A child’s health is not affected by having a single mutated copy of the gene, but the availability of this new information has greatly increased the need for information about haemoglobin variants and how they are inherited.