Hemoglobinopathies and Thalassemias

Ed Uthman, MD

Diplomate, American Board of Pathology

This is a document in a five-part series
on blood cells and anemia:
1. Blood cells and the CBC
2. Anemia: Pathophysiologic Consequences,
Classification, and Clinical Investigation
3. Nutritional Anemias and
Anemia of Chronic Disease
4. Hemolytic Anemias
5. Hemoglobinopathies and Thalassemias

I. Introduction

These conditions comprise a very large number of genetic biochemical/ physiological entities, most of which are academic curiosities whose major effect on medicine is to add to the surfeit of useless scientific information. However, several of these conditions (e.g., sickle cell anemia, hemoglobin SC disease, and some thalassemias) are common major life-threatening diseases, and some others (e.g., most thalassemias, hemoglobin E disease, and hemoglobin O disease) are conditions that produce clinically noticeable -- if not serious -- effects and can cause the unaware physician a lot of frustration and the hapless patient a lot of expense and inconvenience. We will study a few hemoglobinopathies and thalassemias of special importance. It should be kept in mind, though, that there are literally hundreds of diseases in these categories.

II. Definitions

Hemoglobinopathy: A genetic defect that results in abnormal structure of one of the globin chains of the hemoglobin molecule. Although the suffix "-pathy" would conjure an image of "disease," most of the hemoglobinopathies are not clinically apparent. Others produce asymptomatic abnormal hematologic laboratory findings. A very few produce serious disease. The genetic defect may be due to substitution of one amino acid for another (as with the very common Hb S and Hb C and the great majority of the other abnormal hemoglobins), deletion of a portion of the amino acid sequence (Hb Gun Hill), abnormal hybridization between two chains (Hb Lepore), or abnormal elongation of the globin chain (Hb Constant Spring). The abnormal chain that results may be the alpha chain (Hb GPhiladelphia), beta chain (Hb S, Hb C), gamma chain (Hb FTexas), or delta chain (Hb A2Flatbush). These abnormal hemoglobins can have a variety of physiologically significant effects, discussed below in greater depth, but the most severe hemoglobinopathies (Hb S and Hb C diseases) are characterized by hemolysis.

Thalassemia: A genetic defect that results in production of an abnormally low quantity of a given hemoglobin chain or chains. The defect may affect the alpha, beta, gamma, or delta chain, or may affect some combination of the beta, gamma, and delta chain in the same patient (but never the alpha and beta chain together). The result is an imbalance in production of globin chains and the production of an inadequate number of red cells. The cells which are produced are hypochromic/microcytic and contain a surfeit of the unaffected chains which cannot stoichiometrically "mate" with the inadequate supply of thalassemic chains. These "bachelor" chains can produce adverse effects on the red cell and lead to destruction of the red cell in the marrow (ineffective erythropoiesis) and in the circulation (hemolysis). Note that these two definitions are not mutually exclusive -- some hemoglobinopathies may also be thalassemias, in that a structurally abnormal hemoglobin (hemoglobinopathy) may also be underproduced (thalassemia). Some, but not all, hemoglobinopathies and thalassemias are hemolytic anemias. These nosologic concepts are summarized by the Venn diagram below.


III. Pathophysiology of hemoglobinopathies

Messing around with the amino acid sequence of a globin chain has something of a red kryptonite effect. While some positions on the protein chain can tolerate a lot of substitutions without compromising the physiologic integrity of hemoglobin, other positions are very sensitive to amino acid substitutions. For instance, substitution of valine or lysine for glutamate at position 6 of the beta chain produces hemoglobins S and C, respectively, which form intraerythrocytic tactoids (see below) and crystals (again respectively) that cause premature destruction of the rbc (hemolysis). On the other hand, substitution of glutamate, asparagine, and threonine for lysine at position 59 of the beta chain produces, respectively, hemoglobins IHigh Wycombe, JLome, and JKaoshiung, all of which are physiologically indistinguishable from normal Hb A. Without venturing too deeply into tedious stereochemistry, we can say that abnormal globin structure can functionally manifest itself in one or more of the following ways:

IV. Specific hemoglobinopathies

A. Hemoglobin S and sickle cell disease

1. Epidemiology and genetics

The Hb S gene is found primarily in populations of native tropical African origin (which include most African-Americans). The incidence of the gene in some African populations is as high as 40%; in African-Americans the incidence is 8%. The gene is also found with less frequency in non-Indo-European aboriginal peoples of India and in the Middle East. Rare cases have been reported in Caucasians of Mediterranean descent. The gene established itself in the tropical African population presumably because its expression in heterozygotes (sickle cell trait) affords some protection against the clinical consequences of Plasmodium falciparum infestation. Unfortunately, homozygous expression produces sickle cell disease, which is a chronic hemolytic anemia and vaso-occlusive condition that usually takes the life of the patient.

Hemoglobin S has the peculiar characteristic of expressing its biochemical instability by precipitating out of solution and forming up into long microtubular arrays called tactoids. The erythrocytes which contain the Hb S stretch around the tactoids to form the characteristic long, pointed, slightly curved cells called (with somewhat liberal imagination) "sickle cells." Only the deoxygenated form of Hb S (deoxy-Hb S) makes tactoids. The greater the proportion of Hb S in the cell, the greater is the propensity to sickle. Therefore, persons with 100% Hb S (being homozygotes) sickle under everyday conditions, while typical heterozygotes (who usually have about 30-40% Hb S) do not sickle except possibly under extraordinary physiologic conditions. Since Hb S is a beta chain mutation, the disease does not manifest itself until six months of age; prior to that time the Hb S is sufficiently "watered down" by Hb F (alpha2gamma2), which of course has no beta chain.

In post-infancy individuals homozygous for the Hb S gene, 97+% of the hemoglobin is Hb S, the remainder being the normal minor hemoglobin, Hb A2 (alpha2delta2). Several coexisting genetic "abnormalities" (actually godsends) prevalent in African populaitons may ameliorate the course of the disease:

  1. alpha-thalassemia carriers (which comprise 20% of African-Americans!) have a lower MCHC than normal individuals. It has been suggested that a low MCHC is beneficial in decreasing the vaso-occlusive properties of sickled cells. These sickle cell patients live longer and have a milder disease than do non-thalassemic patients. Thalassemia is discussed in greater detail below.

  2. Hereditary persistence of fetal hemoglobin (HPFH) has established itself in the black population and allows Hb F to so dilute the Hb S that sickling does not occur or is less prominent. In these people the Hb F gene does not "turn off" in infancy but persists indefinitely.

  3. G-6-PD deficiency has been suggested as an ameliorative condition for sickle cell disease. This is controversial; the pathophysiologic basis of any such effect must be pretty obscure.

2. Clinical findings

Sickle cell anemia is a particularly bad disease in that not only is it a hemolytic anemia, but also a vaso-occlusive condition. The clinical findings can then be divided into one of these two groups:

B. Hemoglobin C

The gene for Hb C is also prevalent in the African-American population but with less frequency (2-3%) than that of the sickle cell gene. Hb C does not form tactoids, but intracellular blunt ended crystalloids. The result is decreased rbc survival time; however, hemolysis is not as severe as in sickle cell disease, and the vaso-occlusive phenomena, so devastating in sickle cell disease, are not generally noted. Like sickle cell trait, the Hb C trait is asymptomatic. Homozygotes (and some heterozygotes) for Hb C often have many target cells (codocytes) in the peripheral smear, but the crystals, although pathognomonic, are only occasionally seen. The prognosis of homozygous Hb C disease is excellent.

An individual may inherit a Hb S gene from one parent and a Hb C gene from the other. The result of this double whammy is Hb SC disease. The clinical severity of this condition is intermediate between that of sickle cell disease and Hb C disease, except that visual damage due to retinal vascular lesions is characteristically worse in SC disease than in sickle cell anemia. The intracellular bodies that occur upon hemoglobin destabilization in SC disease are curious hybrids of the blunt-ended crystalloids of Hb C and the sharp-pointed tactoids of Hb S, in that they often have one pointed end and one blunt end, thus vaguely resembling arrowheads.

C. Hemoglobin E

This is a very common beta chain mutation among Southeast Asians. The Thai and Khmer groups have the highest frequency, followed by Burmese and Malays, then Vietnamese and Bengalis. The gene does not occur in ethnic Han Chinese or Japanese. The heterozygous state is asymptomatic but causes microcytosis without anemia, thus resembling some cases of beta thalassemia minor (see below). The homozygous state has more severe microcytosis and hypochromia, but little, if any, anemia (this is also reminiscent of thalassemia minor). Hemoglobin E should always be considered working up an unexplained microcytosis in a member of one of the affected ethnic groups.

V. Thalassemia

A. Genetics

Understanding the thalassemias can be facilitated by reviewing the genesis of the normal post-embryonal hemoglobins:

Globin chain genes

Chromosome 16 contains the genes for the all-important alpha chain. The genes for all of the other important globin chains are on chromosome 11, where they are closely linked. The linkage means (if you will briefly abuse yourself by recalling basic genetics) that the genes tend to be inherited as a group, as opposed to non-linked (or distantly linked) genes which assort independently due to crossing over during gametogenesis. Because of the linkage, a mutation that affects the rate of production of the beta chain not uncommonly affects rate of production of the adjacent delta chain. An individual carrying such a mutation would then have a gene for "deltabeta thalassemia." He or she could pass on the deltabeta thalassemia gene to offspring but would essentially never, say, pass a delta thalassemia gene to one child and a beta thalassemia gene to another. Conversely, since the genome for the alpha chain is on a completely different chromosome than the genes for all the other chains, one would expect no mutation in a chromosome 11 chain gene (delta,beta,gamma) to affect alpha chain production. Moreover, if some poor shlimazel happened to inherit an alpha thalassemia gene from one parent and a beta thalassemia gene from another, he would not tend to pass both abnormal genes on as a unit to his or her offspring. One kid (out of a representative Mendelian sibship of four) would get the abnormal alpha gene, one would get the abnormal beta , one would get neither, and one would get both.

B. Biochemistry and pathophysiology

But enough of Mendel! We're in med school to learn about hemoglobin, right? Whatever the genetics, the clinical problem in the thalassemias is the inability to maintain a balance between the synthesis rate of one type of globin chain vis-à-vis that of its mate. Even though thalassemias have been described for all four of the above chains, we will consider only those that involve the beta chain (the beta thalassemias and deltabeta thalassemias) and the alpha chain (alpha thalassemias). It will be useful to review what kind of hemoglobins you can build by mixing and matching globin chains:

Globin chain
A alpha2beta2 The only physiologically important adult hemoglobin in normal individuals. Includes the post-translational glycosylated hemoglobins A1a, A1b, and A1c, the last being important in monitoring diabetics.
F alpha2gamma2 The major physiologic hemoglobin in post embryonal fetuses. Adapted best for lowered intrauterine O2 tension because of its left-shifted Hb-O2 dissociation curve (allowing O2 to be more readily picked up from maternal circulation). Production normally turns off in early infancy. Proportion of circulating Hb F fades to insignificance at about 6 months of age.
A2 alpha2delta2 Medical philosopher's proof of the existence of God (and God's love of physicians). Apparently put here solely as a marker for doctors trying to figure out whether a patient has iron deficiency anemia or beta thalassemia. Normally less than 3% of circulating hemoglobin (thus physiologically insignificant), Hb A2is slightly elevated in most beta thalassemias, but normal or decreased in iron deficiency, thus making it a nifty marker for evaluating microcytic, hypochromic anemias.
Gower 1
Gower 2
Very early normal embryonal hemoglobins that disappear after 8 weeks of gestation. The only one of clinical importance is Hb Portland, which may be seen at birth in cases of the severest form of alpha thalassemia.
H beta4 Abnormal hemoglobin produced in cases of alpha thalassemia, when excess beta chains decide to get it on with each other, there not being enough alpha chains to go around. Intrinsically unstable, Hb H produces Heinz bodies in the erythrocytes and subsequent hemolysis.
Bart's gamma4 Delinquent youth gang analogue of Hb H. This abnormal hemoglobin is found in infants with alpha thalassemia. Detecting presence of Hb Bart's in cord blood may be the only practical way to screen for the very large number of individuals who are silent carriers of one type of alpha thalassemia (see below).

C. Beta thalassemia

Although this is the classic form of thalassemia it is not the most common. The first description was written by Dr. Thomas Cooley in 1925. The term "Cooley's anemia" has been used synonymously with clinically severe forms of beta thalassemia, although the remainder of Cooley's career was so undistinguished as to cause some to suggest that his name is not worthy of eponymous immortality. Cooley's anemia was a fatal microcytic anemia of children of Mediterranean descent. The name "thalassemia" was coined to reflect the original geographic home of the target population ( "thalassa" is the classical Greek name for the Mediterranean Sea). Over the years, it became clear that many other groups (Africans, African-Americans, Arabs, Indians, and Southeast Asians) are affected. In fact, thalassemias in general tend to affect races of people that hail from a tropical belt that girdles the Mediterranean and extends all the way through the Indian subcontinent to Southeast Asia.

There are a multiplicity of different beta thalassemia genes that give rise to a clinically heterogeneous spectrum ranging from asymptomatic expression to classical, deadly Cooley's anemia. It is convenient to group the various beta thalassemias into two groups, based on the amount of beta globin chain production:

Although these genes are remarkably varied in their effect on beta chain synthesis rate, one can make up some useful rules of thumb:

  1. Individuals heterozygous for any of the beta thalassemia genes are either silent carriers or have minimal clinical effects, usually manifested as a borderline anemia (Hct ~ 35 cL/L) with disproportionate microcytosis (MCV ~ 60 fL) and a reciprocally high rbc count (~ 6 x 106/µL). The Hb A2 is increased. This clinical presentation is called thalassemia minor. It makes for interesting wine-tasting party conversation if you have this condition, and all that your friends can muster is chronic fatigue syndrome. Your kids should have no problems if you just marry a Teuton, Slav, Balt, or Lapp.

  2. Individuals homozygous for all of the beta thalassemia genes [except the beta+ (Negro) gene] have severe anemia and some or all of the pathophysiological consequences given in the diagram below. This is classic Cooley's anemia and is termed thalassemia major. This is bad news.

  3. Individuals homozygous for the beta+ (Negro) gene and several other miscellaneous types of mildly behaving genes have a relatively mild clinical anemia called thalassemia intermedia. These patients may require transfusion, but only later in life than is the case in the very sick children with thalassemia major.

The pathophysiology of beta thalassemia major is best understood by going and getting yourself a beer (or politically correct beverage), watching a little TV, doing one or two other chores to postpone the inevitable, and then sitting down to study the next diagram.

Diagram: pathogenesis of thalassemia major

While studying the illustration, consider the following observations concerning beta thalassemia:

  1. Since there is a decrease in the synthesis of beta chains, there is a net decreased synthesis of Hb A. With less Hb A available to fill the red cells, the result is microcytic anemia. Whereas in iron deficiency microcytosis occurs because there is not enough heme, in thalassemia the same thing occurs because there is not enough globin.

  2. Since the body cannot make enough beta chains, it makes a feeble attempt to compensate by trying to churn out delta chains. The result is increased Hb A2, which can be measured easily and inexpensively by column chromatography. This is a pretty specific test for the diagnosis of beta thalassemia. Pitfall: both beta and delta chains are decreased in deltabeta thalassemia, which is not rare and presents like beta thalassemia, except that the Hb A2 is not elevated. You would expect this since Hb A2 contains delta chains).

  3. In some cases of beta thalassemia, there is attempt at compensation by maintaining some production of Hb F. This has some pathophysiologic consequences (as shown above) and also provides a laboratory marker to assist in diagnosis. Retention of Hb F production is not as common as increased rbc Hb A2 content.

  4. In severe forms of thalassemia, the anemia due to failure to make adequate amounts of Hb A is compounded by the hemolysis, ineffective erythropoiesis, and extramedullary hematopoiesis brought on by precipitation of alpha4 tetramers (which are unstable). In classic Cooley's anemia, the ineffective erythropoiesis dominates the clinical picture by producing tremendous expansion of the marrow space, manifested by the so-called "tower skull" with an x-ray showing innumerable vertical bony striae between the inner and outer tables of the calvarium. This radiographic feature is fancifully called the "hair-on-end appearance" by radiologists, and the "guy-who-accidentally-sat-on-a-Van-de-Graaff-generator appearance" by those wacky electrical engineers. Extramedullary hematopoiesis and hemolysis causes splenomegaly, which produces hypersplenism, and more hemolysis.

  5. The high turnover state caused by the tremendous erythroproliferative activity causes wastage of folate and may produce a complicating megaloblastic anemia. Another effect of the high turnover state is hyperuricemia (due to catabolism of the purine content of cellular DNA).

  6. Classically in thalassemia major, the treatment is the cause of death. The children are maintained by transfusions until about age ten years, at which time they start to show symptoms of excess iron loading. This happens because the transfusion bypasses the body's normal gastrointestinal mechanism of iron intake and excretion. The iron is poured into the bloodstream directly; the body cannot excrete it fast enough. First iron (as hemosiderin) fills the cytoplasm of the RES phagocytes and then starts to be deposited in the parenchymal cells of just about every organ of the body. The pancreas, liver, myocardium, adrenals, and gonads are among the organs most sensitive to iron toxicity. The clincial result is diabetes mellitus, hepatic cirrhosis, congestive heart failure, adrenal insufficiency, and failure to undergo puberty. Death used to occur in the second or third decade of life, the most common immediate cause being complications of heart failure. Nowadays, thal major patients live longer because of advances in chelation therapy. To achieve such longevity, they must submit to daily subcutaneous injections of the parenteral chelating agent, deferoxamine. These injections are given by pump, usually overnight, and last 9 to 12 hours each. An oral chelating drug in presently under development in Europe.

D. Alpha thalassemia

The alpha thalassemias include the most common of all hemoglobinopathies and thalassemias. One form of alpha thalassemia is very common in African-Americans. Fortunately this form is so mild that its very detection is almost impossible in adult heterozygotes, and even homozygotes are asymptomatic with mild laboratory abnormalities. Yet another form of alpha thalassemia, fortunately uncommon, produces the most severe disease of all the hemoglobinopathies and thalassemias and usually takes the life of its victim even before birth. Surely alpha thalassemia is a disease of extremes!

It is helpful to consider two concepts concerning alpha thalassemia:

Diagram: pathogenesis of thalassemia major

In the above diagram, the normal diplotype is compared with the heterozygous state for alpha thalassemia-2. The latter occurs in 20% of all African-Americans. It produces no symptoms and no abnormalities on routine laboratory tests. The only practical way to detect it is to screen all black infants at birth for Hb Bart's (gamma4), and even this technique will miss some individuals. It is thought that the reason for the high prevalence of this gene in the black population is that it may be an "anti-sickle cell" gene. It turns out that Hb S homozygotes have a milder course of sickle cell anemia if they also are silent carriers for alpha thalassemia.

Diagram: pathogenesis of thalassemia major

The the two diplotypes given above, two of the four alpha genes are trashed by "bad guy" mutations. Such individuals are usually not anemic but may have microcytosis. They become victims of the medical system when they are subjected to expensive and time consuming testing in a quixotic search for some serious hematologic condition that does not exist. About the only way to make a diagnosis of one of these conditions is to eliminate other causes of microcytosis and/or check rbc indices on all available blood (no pun intended) relatives. Amidst all this shenanigans lurks a somber note -- if an alpha thalassemia-1 heterozygote finds his or her true love in another alpha thalassemia-1 heterozygote, one-fourth of the issue of such a union will suffer the always lethal homozygous state.

Diagram: pathogenesis of thalassemia major

The double heterozygote on the left has "hemoglobin H disease," so named because of the presence of a significant proportion of the hemoglobin composed of four beta chains. Affected infants will, of course, show some Hb Bart's as well. These people have a hemolytic anemia which varies from very mild to that which clinically resembles beta thalassemia major.

The alpha thalassemia-1 homozygote, on the right, is allowed no production of alpha chains. The only hemoglobins present are Hb Bart's, Hb H, and Hb Portland. Most of the affected die in utero or within hours after birth. Autopsy shows massive extramedullary hematopoiesis in virtually every parenchymatous organ of the body. The severe anemia causes congestive heart failure and subsequent massive total body edema, termed "hydrops fetalis." Parenthetically, it should be noted that hydrops (from which springs the term "dropsy") is not limited to thalassemia but is seen in any conditon that causes severe heart failure in utero, such as in the anemia due to alloimmune hemolytic disease of the newborn.

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