RED CEll MEMBRANE DEFECTS
The normal red cell membrane consists of a lipid bilayer crossed by integral proteins with an underlying lattice of proteins (or cytoskeleton), including spectrin, actin, ankyrinand protein 4.1, attached to the integral proteins.
Hereditary spherocytosis (H 5)
HS is the commonest inherited haemolytic anaemia in northern Europeans, affecting 1 in 5000. It is inherited in an autosomal dominant manner but in 25% of patients neither parent is affected and it is presumed that HS hasoccurred by spontaneous mutation. HS is due to a defect in the red cell membrane, resulting in the cells losing part of the cell membrane as they pass through the spleen, possibly because the lipid bilayer is inadequately supported by the cytoskeleton. The surface-to-volume ratio decreases, and the cells become spherocytic. Spherocytes are more rigid and less deformable than normal red cells. They are unable to pass through the splenic microcirculation and they die. Several defects in the cell membrane have been identified in HS. The best characterized is a deficiency in the structural protein spectrin, but quantitative defects in other membrane proteins have been identified such as a deficiency of ankyrin. There may also be functional abnormalities of membrane proteins such as defective binding of spectrin to protein 4.1. The abnormal red cell membrane in HS is associated functionally with an increased permeability to sodium, and this requires an increased rate of active transport of sodium out of the cells which is dependent on ATP produced by glycolysis.
The condition may present with jaundice at birth. However, the onset of jaundice can sometimes be delayed for many years and some patients may go through life with no symptoms and are only detected during family studies. The patient may eventually develop anaemia, splenomegaly and ulcers on the leg. As in many haemolytic anaemias, the course of the disease may be interrupted by aplastic, haemolytic and megaloblastic crises. Aplastic anaemia usually occurs after infections, particularly with parvovirus whereas megaloblastic anaemia is the result of folate depletion due to the hyperactivity of the bone marrow.
Chronic haemolysis leads to the formation of pigment gallstones.
ANAEMIA is usually mild, but occasionally can be severe. BLOOD FILM shows spherocytes and reticulocytes. HAEMOLYSIS is evident, e.g. the serum bilirubin and urinary urobilinogen will be raised.
OSMOTIC FRAGILITY: when red cells are placed in solutions of increasing hypotonicity, they take in water, swell, and eventually lyse. Spherocytes tolerate hypotonic solutions less well than normal biconcave red cells. Osmotic fragility tests are infrequently carried out in routine practice, but may be useful to confirm a suspicion of spherocytosis on a blood film.
DIRECT ANTIGLOBULIN (COOMBS’) TEST is negative in spherocytosis, virtually ruling out autoimmune haemolytic anaemia where spherocytes are also commonly present.
The spleen, which is the site of cell destruction, should be removed in all but the mildest cases. The decision about splenectomy in symptomless patients is difficult, but a raised bilirubin and especially the presence of gallstones should encourage splenectomy. It is best to postpone splenectomy until after childhood, as sudden overwhelming fatal infections, usually due to encapsulated organisms such as pneumococci, may occur. Following splenectomy, the spherocytosis is reduced and Hb usually returns to normal as the red cells are no longer destroyed. Folate deficiency often occurs in chronic haemolysis with rapid cell turnover. Folate levels should be monitored, or folic acid can be given prophylactically.
This disorder of the red cell membrane is inherited in an autosomal dominant manner. The red cells are elliptical. It is a similar condition to HS but milder clinically. Only a minority of patients have anaemia and only occasional patients require splenectomy.
Stomatocytes are red cells in which the pale central area appears slit-like. Their presence in large numbers may occur in a hereditary haemolytic anaemia associated with a membrane defect but excess alcohol intake is a common cause.
Normal adult Hb (Hb A) has two polypeptide globin chains, the a and f3 chains, which have 141 and 146 amino acids, respectively. These are folded so that haem molecules can be held within the fold and are yet able to combine reversibly with oxygen. In early embryonic life, haemoglobins Gower 1, Gower 2 and Portland predominate. Later, fetal haemoglobin (Hb F), which has two a and two ‘I chains, is produced. There is increasing synthesis of f3 chains from 13 weeks of gestation and at term there is 80% Hb F and 20% Hb A. The switch from Hb F to Hb A occurs 3-6 months after birth when the genes for ‘I chain production are further suppressed and there is rapid increase in the synthesis of f3 chains. The exact mechanism responsible for the switch remains unknown. There is little Hb F produced (normally less than 1%) from 6 months after birth. The 0 chain is synthesized just before birth and Hb A2 (0’282) remains at a level of about 2% throughout adult life.
Globin chains are synthesized in the same way as any protein (see Chapter 2). Four globin chain genes are required to control a-chain production. Two are present on each haploid genome (genes derived from one parent). These are situated close together on chromosome b16. The genes controlling the production of E, ‘I, 0 and, f3 chains are close together on chromosome 11. The globin genes are arranged on chromosomes 16 and 11 in the order in which they are expressed.
Abnormal haemoglobins Abnormalities occur in:
• Globin chain production, e.g. thalassaemia
• Structure of the globin chain, e.g. sickle cell disease
• Combined defects of globin chain production and structure, e.g. sickle cell f3-thalassaemia Genetic defects in haemoglobin are the commonest of all genetic disorders.
The thalassaemias (Greek thalassa = sea) are anaemias originally found in people living on the shores of the Mediterranean but are now known to affect people throughout the world.
Normally there is balanced (l : 1) production of a and f3 chains. The defective synthesis of globin genes in thalassaemia leads to ‘imbalanced’ globin chain production,
leading to precipitation of globin chains within the red cell precursors and resulting in ineffective erythropoiesis. Precipitation of globin chains in mature red cells leads to haemolysis.
In homozygous f3-thalassaemia either no normal f3 chains are produced (f30) or f3-chain production is very reduced (fJ”). There is an excess of a chains which precipitate in erythroblasts and red cells causing ineffective erythropoiesis and haemolysis. The excess a chains combine with whatever f3, 0 and ‘I chains are produced, resulting in increased quantities of Hb A2 and Hb F and, at best, small amounts of Hb A. In heterozygous f3-thalassaemia there is usually symptomless microcytosis with or without mild anaemia. Table 6.11 shows the findings in the homozygote and heterozygote for the common types of f3-thalassaemia.
The molecular errors accounting for over 100 genetic defects leading to f3-thalassaemia genes have been characterized. Unlike o-thalassaemia, the defects are mainly point mutations rather than gene deletions. The mutations result in defects in transcription, RNA splicing and modification, translation via frame shifts and nonsense codons producing highly unstable J3-globin which cannot be utilized.