A finite life span is a distinct characteristic of red cells. Hence, a logical, time-honored classification of anemias is in three groups: (1) decreased production of red cells, (2) increased destruction of red cells, and (3) acute blood loss. Decreased production is covered in Chaps. 126, 128, and 130; increased destruction and acute blood loss are covered in this chapter.
All patients who are anemic as a result of either increased destruction of red cells or acute blood loss have one important element in common: the anemia results from overconsumption of red cells from the peripheral blood, whereas the supply of cells from the bone marrow is normal (indeed, it is usually increased). On the other hand, these two groups differ in that physical loss of red cells from the bloodstream or from the body itself, as in acute hemorrhage, is fundamentally different from destruction of red cells within the body, as in hemolytic anemias. Therefore, the clinical aspects and pathophysiology of anemia in these two groups of patients are quite different, and they will be considered separately.
With respect to primary etiology, anemias due to increased destruction of red cells, which we know as hemolytic anemias (HAs), may be inherited or acquired; from a clinical point of view, they may be more acute or more chronic, and they may vary from mild to very severe; the site of hemolysis may be predominantly intravascular or extravascular. With respect to mechanisms, HAs may be due to intracorpuscular causes or to extracorpuscular causes (Table 129-1). But before reviewing the individual types of HA, it is appropriate to consider what they have in common.
TABLE 129-1Classification of Hemolytic Anemiasa ||Download (.pdf) TABLE 129-1Classification of Hemolytic Anemiasa
| ||Intracorpuscular Defects ||Extracorpuscular Factors |
|Hereditary || |
|Familial (atypical) hemolytic-uremic syndrome |
|Acquired ||Paroxysmal nocturnal hemoglobinuria (PNH) || |
Mechanical destruction (microangiopathic)
GENERAL CLINICAL AND LABORATORY FEATURES
The clinical presentation of a patient with anemia is greatly influenced in the first place by whether the onset is abrupt or gradual, and HAs are no exception. A patient with autoimmune HA or with favism may be a medical emergency, whereas a patient with mild hereditary spherocytosis or with cold agglutinin disease may be diagnosed after years. This is due in large measure to the remarkable ability of the body to adapt to anemia when it is slowly progressing (Chap. 77).
What differentiates HAs from other anemias is that the patient has signs and symptoms arising directly from hemolysis (Table 129-2). At the clinical level, the main sign is jaundice; in addition, the patient may report discoloration of the urine. In many cases of HA, the spleen is enlarged, because it is a preferential site of hemolysis; and in some cases, the liver may be enlarged as well. In all severe congenital forms of HA, there may also be skeletal changes due to overactivity of the bone marrow (although they are never as severe as they are in thalassemia).
TABLE 129-2Features Common to Most Patients with a Hemolytic Disorder ||Download (.pdf) TABLE 129-2Features Common to Most Patients with a Hemolytic Disorder
|General examination ||Jaundice, pallor |
|Other physical findings ||Spleen may be enlarged; bossing of skull in severe congenital cases |
|Hemoglobin level ||From normal to severely reduced |
|MCV, MCH ||Usually increased |
|Reticulocytes ||Increased |
|Bilirubin ||Increased (mostly unconjugated) |
|LDH ||Increased (up to 10 time normal with intravascular hemolysis) |
|Haptoglobin ||Reduced to absent (if hemolysis in part intravascular) |
The laboratory features of HA are related to hemolysis per se and the erythropoietic response of the bone marrow. Hemolysis regularly produces an increase in unconjugated bilirubin and aspartate aminotransferase (AST) in the serum; urobilinogen will be increased in both urine and stool. If hemolysis is mainly intravascular, the telltale sign is hemoglobinuria (often associated with hemosiderinuria); in the serum, there is hemoglobin, lactate dehydrogenase (LDH) is increased, and haptoglobin is reduced. In contrast, the bilirubin level may be normal or only mildly elevated. The main sign of the erythropoietic response by the bone marrow is an increase in reticulocytes (a test all too often neglected in the initial workup of a patient with anemia). Usually the increase will be reflected in both the percentage of reticulocytes (the more commonly quoted figure) and the absolute reticulocyte count (the more definitive parameter). The increased number of reticulocytes is associated with an increased mean corpuscular volume (MCV) in the blood count. On the blood smear, this is reflected in the presence of macrocytes; there is also polychromasia, and sometimes one sees nucleated red cells. In most cases, a bone marrow aspirate is not necessary in the diagnostic workup; if it is done, it will show erythroid hyperplasia. In practice, once an HA is suspected, specific tests will usually be required for a definitive diagnosis of a specific type of HA.
The mature red cell is the product of a developmental pathway that brings the phenomenon of differentiation to an extreme. An orderly sequence of events produces synchronous changes, whereby the gradual accumulation of a huge amount of hemoglobin in the cytoplasm (to a final level of 340 g/L, i.e., about 5 mM) goes hand in hand with the gradual loss of cellular organelles and of biosynthetic abilities. In the end, the erythroid cell undergoes a process that has features of apoptosis, including nuclear pyknosis and actual loss of the nucleus. However, the final result is more altruistic than suicidal; the cytoplasmic body, instead of disintegrating, is now able to provide oxygen to all cells in the human organism for some remaining 120 days of the red cell life span.
As a result of this unique process of differentiation and maturation, intermediary metabolism is drastically curtailed in mature red cells (Fig. 129-1); for instance, cytochrome-mediated oxidative phosphorylation has been lost with the loss of mitochondria (through a process of physiologic autophagy); therefore, there is no backup to anaerobic glycolysis, which in the red cell is the only provider of adenosine triphosphate (ATP). Also the capacity of making protein has been lost with the loss of ribosomes. This places the cell’s limited metabolic apparatus at risk, because if any protein component deteriorates, it cannot be replaced, as it would be in most other cells; and in fact the activity of most enzymes gradually decreases as red cells age. At the same time, during their long time in circulation, various red cell components inevitably accumulate damage; in senescent red cells, the membrane protein band 3 molecules (see below and Fig. 129-1), having bound hemichromes on their intracellular domains, tend to cluster. Now they bind anti–band 3 IgG antibodies (present in most people) and C3 complement fragments; thus they become opsonized and are eventually removed by phagocytosis in the reticuloendothelial system.
Red blood cell (RBC) metabolism. The Embden-Meyerhof pathway (glycolysis) generates ATP for energy and membrane maintenance. The generation of NADPH maintains hemoglobin in a reduced state. The hexose monophosphate shunt generates NADPH that is used to reduce glutathione, which protects the red cell against oxidant stress. Regulation of 2,3-bisphosphoglycerate levels is a critical determinant of oxygen affinity of hemoglobin. Enzyme deficiency states in order of prevalence: glucose 6-phosphate dehydrogenase (G6PD) > pyruvate kinase > glucose-6-phosphate isomerase > rare deficiencies of other enzymes in the pathway. The more common enzyme deficiencies are encircled.
Another consequence of the relative simplicity of red cells is that they have a very limited range of ways to manifest distress under hardship; in essence, any sort of metabolic failure will eventually lead either to structural damage to the membrane or to failure of the cation pump. In either case, the life span of the red cell is reduced, which is the definition of a hemolytic disorder. If the rate of red cell destruction exceeds the capacity of the bone marrow to produce more red cells, the hemolytic disorder will manifest as HA.
Thus, the essential pathophysiologic process common to all HAs is an increased red cell turnover; and in many HAs, this is due at least in part to an acceleration of the senescence process described above. The gold standard for proving that the life span of red cells is reduced (compared to the normal value of about 120 days) is a red cell survival study, which can be carried out by labeling the red cells with 51Cr and measuring residual radioactivity over several days or weeks: however, this classic test is now available in very few centers, and it is rarely necessary. If the hemolytic event is transient, it does not usually cause any long-term consequences, except for an increased requirement for erythropoietic factors, particularly folic acid. However, if hemolysis is recurrent or persistent, the increased bilirubin production favors the formation of gallstones. If a considerable proportion of hemolysis takes place in the spleen, as is often the case, splenomegaly may become increasingly a feature, and hypersplenism may develop, with consequent neutropenia and/or thrombocytopenia.
The increased red cell turnover also has metabolic consequences. In normal subjects, the iron from effete red cells is very efficiently recycled by the body; however, with chronic intravascular hemolysis, the persistent hemoglobinuria will cause considerable iron loss, needing replacement. With chronic extravascular hemolysis, the opposite problem, iron overload, is more common, especially if the patient needs frequent blood transfusions. Chronic iron overload will cause secondary hemochromatosis; this will cause damage particularly to the liver, eventually leading to cirrhosis, and to the heart muscle, eventually causing heart failure.
Compensated Hemolysis Versus Hemolytic Anemia
Red cell destruction is a potent stimulus for erythropoiesis, which is mediated by erythropoietin (EPO) produced by the kidney. This mechanism is so effective that in many cases the increased output of red cells from the bone marrow can fully balance an increased destruction of red cells. In such cases, we say that hemolysis is compensated. The pathophysiology of compensated hemolysis is similar to what we have just described, except there is no anemia. This notion is important from the diagnostic point of view, because a patient with a hemolytic condition, even an inherited one, may present without anemia; and it is also important from the point of view of management, because compensated hemolysis may become “decompensated,” i.e., anemia may suddenly appear, in certain circumstances, for instance in pregnancy, folate deficiency, or renal failure interfering with adequate EPO production. Another general feature of chronic HAs is seen when any intercurrent condition, such as an acute infection, depresses erythropoiesis. When this happens, in view of the increased rate of red cell turnover, the effect will be predictably much more marked than in a person who does not have hemolysis. The most dramatic example is infection by parvovirus B19, which may cause a rather precipitous fall in hemoglobin—an occurrence sometimes referred to as aplastic crisis.
INHERITED HEMOLYTIC ANEMIAS
There are three essential components in the red cell: (1) hemoglobin, (2) the membrane-cytoskeleton complex, and (3) the metabolic machinery necessary to keep hemoglobin and the membrane-cytoskeleton complex in working order. Diseases caused by abnormalities of hemoglobin, or hemoglobinopathies, are covered in Chap. 127. Here we will deal with diseases of the other two components.
Hemolytic Anemias due to Abnormalities of the Membrane-Cytoskeleton Complex
The detailed architecture of the red cell membrane is complex, but its basic design is relatively simple (Fig. 129-2). The lipid bilayer incorporates phospholipids and cholesterol, and it is spanned by a number of proteins that have their hydrophobic transmembrane domain(s) embedded in the membrane; most of these proteins also extend to both the outside (extracellular domains) and the inside of the cell (cytoplasmic domains). Other proteins are tethered to the membrane through a glycosylphosphatidylinositol (GPI) anchor; these have only an extracellular domain, and they include ion channels, receptors for complement components, and receptors for other ligands. The most abundant red cell membrane proteins are glycophorins and the so-called band 3, an anion transporter. The extracellular domains of many of these proteins are heavily glycosylated, and they carry antigenic determinants that correspond to blood groups. Underneath the membrane, and tangential to it, is a network of other proteins that make up the cytoskeleton. The main cytoskeletal protein is spectrin, the basic unit of which is a dimer of α-spectrin and β-spectrin. The membrane is physically linked to the cytoskeleton by a third set of proteins (including ankyrin and the so-called band 4.1 and band 4.2), which thus make these two structures intimately connected to each other.
The red cell membrane. In this figure, one sees, within the lipid bilayer, several membrane proteins, of which band 3 (anion exchanger 1 [AE1]) is the most abundant; the α-β spectrin dimers that associate to form most of the cytoskeleton; and several proteins (e.g., ankyrin) that connect the membrane to the cytoskeleton. In addition, as examples of glycosylphosphatidylinositol (GPI)-linked proteins, one sees acetylcholinesterase (AChE) and the two complement-regulatory proteins CD59 and CD55. The (nonrealistic) shapes of the protein moieties of the GPI-linked proteins are meant to indicate that they can be very different from each other and that, unlike with the other membrane proteins shown, the entire polypeptide chain is extracellular. Branched lines symbolize carbohydrate moiety of proteins. The molecules are obviously not drawn to the same scale. Additional explanations can be found in the text. (From N Young et al: Clinical Hematology. Copyright Elsevier, 2006; with permission.)
The membrane-cytoskeleton complex is so integrated that, not surprisingly, an abnormality of almost any of its components will be disturbing or disruptive, causing structural failure, which results ultimately in hemolysis. These abnormalities are almost invariably inherited mutations; thus, diseases of the membrane-cytoskeleton complex belong to the category of inherited HAs. Before the red cells lyse, they often exhibit more or less specific morphologic changes that alter the normal biconcave disk shape. Thus, the majority of the diseases in this group have been known for over a century as hereditary spherocytosis and hereditary elliptocytosis. Over the past 20 years, their molecular basis has been elucidated; it has emerged that both conditions can arise from mutations in several genes with considerable overlap (Fig. 129-3).
Hereditary spherocytosis (HS), hereditary elliptocytosis (HE), and hereditary stomatocytosis (HSt) are three morphologically distinct forms of congenital hemolytic anemia. It has emerged that each one can arise from mutation of one of several genes and that different mutations of the same gene can give one or another form. (See also Table 129-3.)
HEREDITARY SPHEROCYTOSIS (HS)
This is a relatively common type of genetically determined HA, with an estimated frequency of at least 1 in 5000. Its identification is credited to Minkowksy and Chauffard, who, at the end of the nineteenth century, reported families who had the presence of numerous spherocytes in the peripheral blood (Fig 129-4A). In vitro studies revealed that the red cells were abnormally susceptible to lysis in hypotonic media; indeed, the presence of osmotic fragility became the main diagnostic test for HS. Today we know that HS, thus defined, is genetically heterogeneous; i.e., it can arise from a variety of mutations in one of several genes (Table 129-3). It has been also recognized that the inheritance of HS is not always autosomal dominant (with the patient being heterozygous); indeed, some of the most severe forms are instead autosomal recessive (with the patient being homozygous).
Peripheral blood smear from patients with membrane-cytoskeleton abnormalities. A. Hereditary spherocytosis. B. Hereditary elliptocytosis, heterozygote. C. Elliptocytosis, with both alleles of the α-spectrin gene mutated.
TABLE 129-3Inherited Diseases of the Red Cell Membrane-Cytoskeleton Complex ||Download (.pdf) TABLE 129-3Inherited Diseases of the Red Cell Membrane-Cytoskeleton Complex
|Gene ||Chromosomal Location ||Protein Produced ||Disease(s) with Certain Mutations (Inheritance) ||Comments |
|SPTA1 ||1q22-q23 ||α-Spectrin ||HS (recessive) ||Rare |
| || || ||HE (dominant) ||Mutations of this gene account for about 65% of HE. More severe forms may be due to coexistence of an otherwise silent mutant allele. |
|SPTB ||14q23-q24.1 ||β-Spectrin ||HS (dominant) ||Rare |
| || || ||HE (dominant) ||Mutations of this gene account for about 30% of HE, including some severe forms. |
|ANK1 ||8p11.2 ||Ankyrin ||HS (dominant) ||May account for majority of HS. |
|SLC4A1 ||17q21 ||Band 3; also known as AE (anion exchanger) or AE1 ||HS (dominant) ||Mutations of this gene may account for about 25% of HS. |
| || || ||Southeast Asia ovalocytosis (dominant) ||Polymorphic mutation (deletion of 9 amino acids); clinically asymptomatic; protective against Plasmodium falciparum. |
| || || ||Stomatocytosis ||Certain specific missense mutations shift protein function from anion exchanger to cation conductance. |
|EPB41 ||1p33-p34.2 ||Band 4.1 ||HE (dominant) ||Mutations of this gene account for about 5% of HE, mostly with prominent morphology but no hemolysis in heterozygotes; severe hemolysis in homozygotes. |
|EPB42 ||15q15-q21 ||Band 4.2 ||HS (recessive) ||Mutations of this gene account for about 3% of HS. |
|RHAG ||6p21.1-p11 ||Rhesus antigen ||Chronic nonspherocytic hemolytic anemia (recessive) || |
Very rare; associated with total loss of all Rh antigens.
A specific mutation causes overhydrated stomatocytosis.
|PIEZO1 ||16q23-q24 ||PIEZO1 ||Dehydrated hereditary stomatocytosis (dominant) ||Also known as xerocytosis with pseudohyperkalemia. Patients may present with perinatal edema. PIEZO1 is a mechanosensitive cation channel. |
Clinical presentation and diagnosis
The spectrum of clinical severity of HS is broad. Severe cases may present in infancy with severe anemia, whereas mild cases may present in young adults or even later in life. The main clinical findings are jaundice, an enlarged spleen, and often gallstones; indeed, it may be the finding of gallstones in a young person that triggers diagnostic investigations.
The variability in clinical manifestations that is observed among patients with HS is largely due to the different underlying molecular lesions (Table 129-3). Not only are mutations of several genes involved, but also individual mutations of the same gene can also give very different clinical manifestations. In milder cases, hemolysis is often compensated (see above), and this may cause variation in time even in the same patient, due to the fact that intercurrent conditions (e.g., pregnancy, infection) may cause decompensation. The anemia is usually normocytic, with the characteristic morphology that gives the disease its name. An increased mean corpuscular hemoglobin concentration (MCHC) on an ordinary blood count report should raise the suspicion of HS, because HS is almost the only condition in which this abnormality occurs. It has been apparent for a long time that the spleen plays a special role in HS through a dual mechanism. On one hand, like in many other HAs, the spleen itself is a major site of destruction; on the other hand, transit through the splenic circulation makes the defective red cells more spherocytic and, therefore, accelerates their demise, even though that may take place elsewhere.
When there is a family history, it is usually easy to make a diagnosis based on features of HA and typical red cell morphology. However, there may be no family history for at least two reasons. First, the patient may have a de novo mutation, i.e., a mutation that has taken place in a germ cell of one of his parents or early after zygote formation. Second, the patient may have a recessive form of HS (Table 129-3). In such cases, more extensive laboratory investigations are required, including osmotic fragility, the acid glycerol lysis test, the eosin-5′-maleimide (EMA)–binding test, and SDS-gel electrophoresis of membrane proteins; these tests are usually carried out in laboratories with special expertise in this area. Sometimes a definitive diagnosis can be obtained only by molecular studies demonstrating a mutation in one of the genes underlying HS (Table 129-3).
TREATMENT Hereditary Spherocytosis
We do not have a causal treatment for HS; i.e., no way has yet been found to correct the basic defect in the membrane-cytoskeleton structure. Given the special role of the spleen in HS (see above), it has long been thought that an almost obligatory therapeutic measure was splenectomy. Because this operation may have more than trivial consequences, today we have more articulate recommendations, based on disease severity (having found out, whenever possible, about the outcome of splenectomy in the patient’s relatives with HS), as follows. In mild cases, avoid splenectomy. Delay splenectomy until puberty in moderate cases or until 4–6 years of age in severe cases. Antipneumococcal vaccination before splenectomy is imperative, whereas penicillin prophylaxis after splenectomy is controversial. Along with splenectomy, cholecystectomy should not be regarded as automatic; it should be carried out, usually by the laparoscopic approach, when clinically indicated.
HEREDITARY ELLIPTOCYTOSIS (HE)
HE is at least as heterogeneous as HS, both from the genetic point of view (Table 129-3, Fig. 129-3) and from the clinical point of view. Again, it is the shape of the red cells (Fig. 129-4B) that gives the name to the condition, but there is no direct correlation between the elliptocytic morphology and clinical severity. In fact, some mild or even asymptomatic cases may have nearly 100% elliptocytes, whereas in severe cases, all kinds of bizarre poikilocytes can predominate. Clinical features and recommended management are similar to those outlined above for HS. Although the spleen may not have the specific role it has in HS, in severe cases, splenectomy may be beneficial. The prevalence of HE causing clinical disease is similar to that of HS. However, an in-frame deletion of nine amino acids in the SLC4A1 gene encoding band 3, causing the so-called Southeast Asia ovalocytosis, has a frequency of up to 7% in certain populations, presumably as a result of malaria selection; it is asymptomatic in heterozygotes and probably lethal in homozygotes.
Disorders of Cation Transport
These rare conditions with autosomal dominant inheritance are characterized by increased intracellular sodium in red cells, with concomitant loss of potassium; indeed, they are sometimes discovered through the incidental finding, in a blood test, of a high serum K+ (pseudohyperkalemia). In patients from some families, the cation transport disturbance is associated with gain of water; as a result, the red cells are overhydrated (low MCHC), and on a blood smear, the normally round-shaped central pallor is replaced by a linear-shaped central pallor, which has earned this disorder the name stomatocytosis (Fig. 129-3). In patients from other families, instead, the red cells are dehydrated (high MCHC), and their consequent rigidity has earned this disorder the name xerocytosis. One would surmise that in these disorders the primary defect may be in a cation transporter; indeed, xerocytosis results from mutations in PIEZO1. In other patients with stomatocytosis, mutations are found in other genes also related to solute transport (Table 129-3), including SLC4A1 (encoding band 3), the Rhesus gene RHAG, and the glucose transporter gene SLC2A1 responsible for a special form called cryohydrocytosis. Hemolysis can vary from relatively mild to quite severe. From the practical point of view, it is important to know that in stomatocytosis, splenectomy is strongly contraindicated because it has been followed in a significant proportion of cases by severe thromboembolic complications.
When there is an important defect in the membrane or in the cytoskeleton, hemolysis is a direct consequence of the fact that the very structure of the red cell is abnormal. Instead, when one of the enzymes is defective, the consequences will depend on the precise role of that enzyme in the metabolic machinery of the red cell, which, in first approximation, has two important functions: (1) to provide energy in the form of ATP and (2) to prevent oxidative damage to hemoglobin and to other proteins by providing sufficient reductive potential; the key molecule for this is NADPH.
ABNORMALITIES OF THE GLYCOLYTIC PATHWAY
Because red cells, in the course of their differentiation, have sacrificed not only their nucleus and their ribosomes, but also their mitochondria, they rely exclusively on the anaerobic portion of the glycolytic pathway for producing energy in the form of ATP. Most of the ATP is required by the red cell for cation transport against a concentration gradient across the membrane. If this fails, due to a defect of any of the enzymes of the glycolytic pathway (Table 129-4), the result will be hemolytic disease.
TABLE 129-4Red Cell Enzyme Abnormalities Causing Hemolysis ||Download (.pdf) TABLE 129-4Red Cell Enzyme Abnormalities Causing Hemolysis
| ||Enzyme (Acronym) ||Chromosomal Location ||Prevalence of Enzyme Deficiency (Rank) ||Clinical Manifestations Extra-Red Cell ||Comments |
|Glycolytic Pathway || || || || || |
| ||Hexokinase (HK) ||10q22 ||Very rare || ||Other isoenzymes known |
| ||Glucose 6-phosphate isomerase (G6PI) ||19q31.1 ||Rare (4)a ||NM, CNS || |
| ||Phosphofructokinase (PFK) ||12q13 ||Very rare ||Myopathy || |
| ||Aldolase ||16q22-24 ||Very rare || || |
| ||Triose phosphate isomerase (TPI) ||12p13 ||Very rare ||CNS (severe), NM || |
| ||Glyceraldehyde 3-phosphate dehydrogenase (GAPD) ||12p13.31-p13.1 ||Very rare ||Myopathy || |
| ||Diphosphoglycerate mutase (DPGM) ||7q31-q34 ||Very rare || ||Erythrocytosis rather than hemolysis |
| ||Phosphoglycerate kinase (PGK) ||Xq13 ||Very rare ||CNS, NM ||May benefit from splenectomy |
| ||Pyruvate kinase (PK) ||1q21 ||Rare (2)a || ||May benefit from splenectomy |
|Redox || || || || || |
| ||Glucose 6-phosphate dehydrogenase (G6PD) ||Xq28 ||Common (1)a ||Very rarely granulocytes ||In almost all cases, only AHA from exogenous trigger |
| ||Glutathione synthase ||20q11.2 ||Very rare ||CNS || |
| ||γ-Glutamylcysteine synthase ||6p12 ||Very rare ||CNS || |
| ||Cytochrome b5 reductase ||22q13.31-qter ||Rare ||CNS ||Methemoglobinemia rather than hemolysis |
|Nucleotide Metabolism || || || || || |
| ||Adenylate kinase (AK) ||9q34.1 ||Very rare ||CNS || |
| ||Pyrimidine 5′-nucleotidase (P5N) ||3q11-q12 ||Rare (3)a || ||May benefit from splenectomy |
Pyruvate kinase deficiency
Abnormalities of the glycolytic pathway are all inherited and all rare. Among them, deficiency of pyruvate kinase (PK) is the least rare, with an estimated prevalence in most populations of the order of 1:10,000. However, very recently, a polymorphic PK mutation (E277K) was found in some African populations, with heterozygote frequencies of 1–7%, suggesting that this may be another malaria-related polymorphism. The clinical picture of homozygous (or compound biallelic) PK deficiency is that of an HA that often presents in the newborn with neonatal jaundice; the jaundice persists, and it is usually associated with a very high reticulocytosis. The anemia is of variable severity; sometimes it is so severe as to require regular blood transfusion treatment, whereas sometimes it is mild, bordering on a nearly compensated hemolytic disorder. As a result, the diagnosis may be delayed, and in some cases, it is made, for instance, in a young woman during her first pregnancy, when the anemia may get worse. The delay in diagnosis may be also helped by the fact that the anemia is remarkably well tolerated, because the metabolic block at the last step in glycolysis causes an increase in bisphosphoglycerate (or DPG; Fig. 129-1), a major effector of the hemoglobin-oxygen dissociation curve; thus, the oxygen delivery to the tissues is enhanced, a remarkable compensatory feat.
TREATMENT Pyruvate Kinase Deficiency
The management of PK deficiency is mainly supportive. In view of the marked increase in red cell turnover, oral folic acid supplements should be given constantly. Blood transfusion should be used as necessary, and iron chelation may have to be added if the blood transfusion requirement is high enough to cause iron overload. In these patients, who have more severe disease, splenectomy may be beneficial. There is a single case report of curative treatment of PK deficiency by bone marrow transplantation from an HLA-identical PK-normal sibling. This seems a viable option for severe cases when a sibling donor is available. Rescue of inherited PK deficiency through lentiviral-mediated human PK gene transfer has been successful in mice. Prenatal diagnosis has been carried out in a mother who had already had an affected child.
Other glycolytic enzyme abnormalities
All of these defects are rare to very rare (Table 129-4), and all cause hemolytic anemia with varying degrees of severity. It is not unusual for the presentation to be in the guise of severe neonatal jaundice, which may require exchange transfusion; if the anemia is less severe, it may present later in life, or it may even remain asymptomatic and be detected incidentally when a blood count is done for unrelated reasons. The spleen is often enlarged. When other systemic manifestations occur, they can involve the central nervous system (sometimes entailing severe mental retardation, particularly in the case of triose phosphate isomerase deficiency), the neuromuscular system, or both. This is not altogether surprising, if we consider that these are housekeeping genes. The diagnosis of hemolytic anemia is usually not difficult, thanks to the triad of normomacrocytic anemia, reticulocytosis, and hyperbilirubinemia. Enzymopathies should be considered in the differential diagnosis of any chronic Coombs-negative hemolytic anemia. Unlike with membrane disorders where the red cells show characteristic morphologic abnormalities, in most cases of glycolytic enzymopathies, these are conspicuous by their absence. A definitive diagnosis can be made only by demonstrating the deficiency of an individual enzyme by quantitative assays; these are carried out in only a few specialized laboratories. If a particular molecular abnormality is already known in the family, then one could test directly for that defect at the DNA level, thus bypassing the need for enzyme assays. Of course the time may be getting nearer when a patient will present with her or his exome already sequenced, and we will need to concentrate on which genes to look up within the file. The principles for the management of these conditions are similar as for PK deficiency. In one case of phosphoglycerate kinase deficiency, allogeneic bone marrow transplantation (BMT) effectively controlled the hematologic manifestations but did not reverse neurologic damage.
ABNORMALITIES OF REDOX METABOLISM
Glucose 6-phosphate dehydrogenase (G6PD) deficiency
G6PD is a housekeeping enzyme critical in the redox metabolism of all aerobic cells (Fig. 129-1). In red cells, its role is even more critical, because it is the only source of NADPH, which directly and via glutathione (GSH) defends these cells against oxidative stress (Fig. 129-5). G6PD deficiency is a prime example of an HA due to interaction between an intracorpuscular cause and an extracorpuscular cause, because in the majority of cases hemolysis is triggered by an exogenous agent. Although a decrease in G6PD activity is present in most tissues of G6PD-deficient subjects, in other cells, the decrease is much less marked than in red cells, and it does not seem to impact on clinical expression.
Diagram of redox metabolism in the red cell. 6PG, 6-phosphogluconate; G6P, glucose 6-phosphate; G6PD, glucose 6-phosphate dehydrogenase; GSH, reduced glutathione; GSSG, oxidized glutathione; Hb, hemoglobin; MetHb, methemoglobin; NADP, nicotinamide adenine dinucleotide phosphate; NADPH, reduced nicotinamide adenine dinucleotide phosphate.
The G6PD gene is X-linked, and this has important implications. First, because males have only one G6PD gene (i.e., they are hemizygous for this gene), they must be either normal or G6PD deficient. By contrast, females, who have two G6PD genes, can be either normal or deficient (homozygous) or intermediate (heterozygous). As a result of the phenomenon of X chromosome inactivation, heterozygous females are genetic mosaics, with a highly variable ratio of G6PD-normal to G6PD-deficient cells and an equally variable degree of clinical expression; some heterozygotes can be just as affected as hemizygous males. The enzymatically active form of G6PD is either a dimer or a tetramer of a single protein subunit of 514 amino acids. G6PD-deficient subjects have been found invariably to have mutations in the coding region of the G6PD gene (Fig. 129-5). Almost all of the approximately 180 different mutations known are single missense point mutations, entailing single amino acid replacements in the G6PD protein. In most cases, these mutations cause G6PD deficiency by decreasing the in vivo stability of the protein; thus, the physiologic decrease in G6PD activity that takes place with red cell aging is greatly accelerated. In some cases, an amino acid replacement can also affect the catalytic function of the enzyme.
Among these mutations, those underlying chronic nonspherocytic hemolytic anemia (CNSHA; see below) are a discrete subset. This much more severe clinical phenotype can be ascribed in some cases to adverse qualitative changes (for instance, a decreased affinity for the substrate, glucose 6-phosphate) or simply to the fact that the enzyme deficit is more extreme, because of a more severe instability of the enzyme. For instance, a cluster of mutations map at or near the dimer interface, and clearly they compromise severely the formation of the dimer.
G6PD deficiency is widely distributed in tropical and subtropical parts of the world (Africa, Southern Europe, the Middle East, Southeast Asia, and Oceania) (Fig. 129-6) and wherever people from those areas have migrated. A conservative estimate is that at least 400 million people have a G6PD deficiency gene. In several of these areas, the frequency of a G6PD deficiency gene may be as high as 20% or more. It would be quite extraordinary for a trait that causes significant pathology to spread widely and reach high frequencies in many populations without conferring some biologic advantage. Indeed, G6PD is one of the best-characterized examples of genetic polymorphisms in the human species. Clinical field studies and in vitro experiments strongly support the view that G6PD deficiency has been selected by Plasmodium falciparum malaria, by virtue of the fact that it confers a relative resistance against this highly lethal infection. Different G6PD variants underlie G6PD deficiency in different parts of the world. Some of the more widespread variants are G6PD Mediterranean on the shores of that sea, in the Middle East, and in India; G6PD A– in Africa and in Southern Europe; G6PD Vianchan and G6PD Mahidol in Southeast Asia; G6PD Canton in China; and G6PD Union worldwide. The heterogeneity of polymorphic G6PD variants is proof of their independent origin, and it supports the notion that they have been selected by a common environmental agent, in keeping with the concept of convergent evolution (Fig. 129-6).
Epidemiology of glucose 6-phosphate dehydrogenase (G6PD) deficiency throughout the world. The different shadings indicate increasingly high levels of prevalence, up to about 20%; the different colored symbols indicate individual genetic variants of G6PD, each one having a different mutation. (From L Luzzatto et al, in C Scriver et al [eds]: The Metabolic & Molecular Bases of Inherited Disease, 8th ed. New York, McGraw-Hill, 2001.)
The vast majority of people with G6PD deficiency remain clinically asymptomatic throughout their lifetime; however, all of them have an increased risk of developing neonatal jaundice (NNJ) and a risk of developing acute HA (AHA) when challenged by a number of oxidative agents. NNJ related to G6PD deficiency is very rarely present at birth; the peak incidence of clinical onset is between day 2 and day 3, and in most cases, the anemia is not severe. However, NNJ can be very severe in some G6PD-deficient babies, especially in association with prematurity, infection, and/or environmental factors (such as naphthalene-camphor balls, which are used in babies’ bedding and clothing), and the risk of severe NNJ is also increased by the coexistence of a monoallelic or biallelic mutation in the uridyl transferase gene (UGT1A1; the same mutations are associated with Gilbert’s syndrome). If inadequately managed, NNJ associated with G6PD deficiency can produce kernicterus and permanent neurologic damage.
AHA can develop as a result of three types of triggers: (1) fava beans, (2) infections, and (3) drugs (Table 129-5). Typically, a hemolytic attack starts with malaise, weakness, and abdominal or lumbar pain. After an interval of several hours to 2–3 days, the patient develops jaundice and often dark urine. The onset can be extremely abrupt, especially with favism in children. The anemia is moderate to extremely severe, usually normocytic and normochromic, and due partly to intravascular hemolysis; hence, it is associated with hemoglobinemia, hemoglobinuria, high LDH, and low or absent plasma haptoglobin. The blood film shows anisocytosis, polychromasia, and spherocytes typical of hemolytic anemias. The most typical feature of G6PD deficiency is the presence of bizarre poikilocytes, with red cells that appear to have unevenly distributed hemoglobin (“hemighosts”) and red cells that appear to have had parts of them bitten away (“bite cells” or “blister cells”) (Fig. 129-7). A classical test, now rarely carried out, is supravital staining with methyl violet, which, if done promptly, reveals the presence of Heinz bodies (consisting of precipitates of denatured hemoglobin and hemichromes), which are regarded as a signature of oxidative damage to red cells (they are also seen with unstable hemoglobins). LDH is high, and so is the unconjugated bilirubin, indicating that there is also extravascular hemolysis. The most serious threat from AHA in adults is the development of acute renal failure (this is exceedingly rare in children). Once the threat of acute anemia is over and in the absence of comorbidity, full recovery from AHA associated with G6PD deficiency is the rule.
TABLE 129-5Drugs That Carry Risk of Clinical Hemolysis in Persons with Glucose 6-Phosphate Dehydrogenase Deficiency ||Download (.pdf) TABLE 129-5Drugs That Carry Risk of Clinical Hemolysis in Persons with Glucose 6-Phosphate Dehydrogenase Deficiency
Peripheral blood smear from a glucose 6-phosphate dehydrogenase (G6PD)-deficient boy experiencing hemolysis. Note the red cells that are misshapen and called “bite” cells. (From MA Lichtman et al: Lichtman's Atlas of Hematology: http://www.accessmedicine.com. Copyright © The McGraw-Hill Companies, Inc. All rights reserved.)
Although it was primaquine (PQ) that led to the discovery of G6PD deficiency, this drug has not been very prominent subsequently, because it is not necessary for the treatment of life-threatening P. falciparum malaria. Today there is a revival of interest in PQ because it is the only effective agent for eliminating the gametocytes of P. falciparum (thus preventing further transmission) and eliminating the hypnozoites of Plasmodium vivax (thus preventing endogenous relapse). In countries aiming to eliminate malaria, there may be a call for mass administration of PQ; this ought to be associated with G6PD testing. At the other end of the historic spectrum, the latest addition to the list of potentially hemolytic drugs (Table 129-5) is rasburicase; again G6PD testing ought to be made mandatory before giving this drug because fatal cases have been reported in newborns with kidney injury and in adults with tumor lysis syndrome.
A very small minority of subjects with G6PD deficiency have chronic nonspherocytic hemolytic anemia (CNSHA) of variable severity. The patient is nearly always a male, usually with a history of NNJ, who may present with anemia, unexplained jaundice, or gallstones later in life. The spleen may be enlarged. The severity of anemia ranges in different patients from borderline to transfusion dependent. The anemia is usually normomacrocytic, with reticulocytosis. Bilirubin and LDH are increased. Although hemolysis is, by definition, chronic in these patients, they are also vulnerable to acute oxidative damage, and therefore the same agents that can cause AHA in people with the ordinary type of G6PD deficiency will cause severe exacerbations in people with CNSHA associated with G6PD deficiency. In some cases of CNSHA, the deficiency of G6PD is so severe in granulocytes that it becomes rate-limiting for their oxidative burst, with consequent increased susceptibility to some bacterial infections.
The suspicion of G6PD deficiency can be confirmed by semiquantitative methods often referred to as screening tests, which are suitable for population studies and can correctly classify male subjects, in the steady state, as G6PD normal or G6PD deficient. However, in clinical practice, a diagnostic test is usually needed when the patient has had a hemolytic attack; this implies that the oldest, most G6PD-deficient red cells have been selectively destroyed, and young red cells, having higher G6PD activity, are being released into the circulation. Under these conditions, only a quantitative test can give a definitive result. In males, this test will identify normal hemizygotes and G6PD-deficient hemizygotes; among females, some heterozygotes will be missed, but those who are at most risk of hemolysis will be identified. Of course, G6PD deficiency also can be diagnosed by DNA testing.
TREATMENT G6PD Deficiency
The AHA of G6PD deficiency is largely preventable by avoiding exposure to triggering factors of previously screened subjects. Of course, the practicability and cost-effectiveness of screening depend on the prevalence of G6PD deficiency in each individual community. Favism is entirely preventable in G6PD-deficient subjects by not eating fava beans. Drug-induced hemolysis can be prevented by testing for G6PD deficiency before prescribing; in most cases, one can use alternative drugs. When AHA develops and once its cause is recognized, in most cases, no specific treatment is needed. However, if the anemia is severe, it may be a medical emergency, especially in children, requiring immediate action, including blood transfusion. This has been the case with an antimalarial drug combination containing dapsone (called Lapdap, introduced in 2003) that has caused severe acute hemolytic episodes in children with malaria in several African countries; after a few years, the drug was taken off the market. If there is acute renal failure, hemodialysis may be necessary, but if there is no previous kidney disease, recovery is the rule. The management of NNJ associated with G6PD deficiency is no different from that of NNJ due to other causes.
In cases with cnsha, if the anemia is not severe, regular folic acid supplements and regular hematologic surveillance will suffice. It will be important to avoid exposure to potentially hemolytic drugs, and blood transfusion may be indicated when exacerbations occur, mostly in concomitance with intercurrent infection. In rare patients, regular blood transfusions may be required, in which case appropriate iron chelation should be instituted. Unlike in HS, there is no evidence of selective red cell destruction in the spleen; however, in practice, splenectomy has proven beneficial in severe cases.
Other abnormalities of the redox system
As mentioned above, GSH is a key player in the defense against oxidative stress. Inherited defects of GSH metabolism are exceedingly rare, but each one can give rise to chronic HA (Table 129-4). A rare, peculiar, usually self-limited severe HA of the first month of life, called infantile poikilocytosis, may be associated with deficiency of glutathione peroxidase (GSHPX) due not to an inherited abnormality, but to transient nutritional deficiency of selenium, an element essential for the activity of GSHPX.
PYRIMIDINE 5′-NUCLEOTIDASE (P5N) DEFICIENCY
P5N is a key enzyme in the catabolism of nucleotides arising from the degradation of nucleic acids that takes place in the final stages of erythroid cell maturation. How exactly its deficiency causes HA is not well understood, but a highly distinctive feature of this condition is a morphologic abnormality of the red cells known as basophilic stippling. The condition is rare, but it probably ranks third in frequency among red cell enzyme defects (after G6PD deficiency and PK deficiency). The anemia is lifelong, of variable severity, and may benefit from splenectomy.
Familial (Atypical) Hemolytic-Uremic Syndrome
The term familial (atypical) hemolytic-uremic syndrome is used to designate a group of rare disorders, mostly affecting children, characterized by microangiopathic HA with presence of fragmented erythrocytes in the peripheral blood smear, thrombocytopenia (usually mild), and acute renal failure. (The word atypical is part of the phrase for historical reasons; it was hemolytic-uremic syndrome [HUS] caused by infection with Escherichia coli producing the Shiga toxin that was regarded as typical). The genetic basis of atypical HUS (aHUS) has been elucidated. Studies of >100 families have revealed that those family members who developed HUS had mutations in any one of several genes encoding complement regulatory proteins: complement factor H (CFH), CD46 or membrane cofactor protein (MCP), complement factor I (CFI), complement component C3, complement factor B (CFB), and thrombomodulin. Thus, whereas all other inherited HAs are due to intrinsic red cell abnormalities, this group is unique in that hemolysis results from an inherited defect external to red cells (Table 129-1). Because the regulation of the complement cascade has considerable redundancy, in the steady state, any of the above abnormalities can be tolerated. However, when an intercurrent infection or some other trigger activates complement through the alternative pathway, the deficiency of one of the complement regulators becomes critical. Endothelial cells get damaged, especially in the kidney; at the same time, and partly as a result of this, there will be brisk hemolysis (thus, the more common Shiga toxin–related HUS [Chap. 149] can be regarded as a phenocopy of aHUS). aHUS is a severe disease, with up to 15% mortality in the acute phase and up to 50% of cases progressing to end-stage renal disease. Not infrequently, aHUS undergoes spontaneous remission; but because its basis is an inherited abnormality, it is not surprising that, given renewed exposure to a trigger, the syndrome will tend to recur; when it does, the prognosis is always serious. The standard treatment has been plasma exchange, which will supply the deficient complement regulator. The anti-C5 complement inhibitor eculizumab (see below) was found to greatly ameliorate the microangiopathic picture, with improvement in platelet counts and in renal function, thus abrogating the need for plasma exchange. It remains to be seen for how long eculizumab treatment will have to be continued in individual patients and whether it will influence the controversial issue of kidney (and liver) transplantation.
ACQUIRED HEMOLYTIC ANEMIA
Mechanical Destruction of Red Cells
Although red cells are characterized by the remarkable deformability that enables them to squeeze through capillaries narrower than themselves for thousands of times in their lifetime, there are at least two situations in which they succumb to shear, if not to wear and tear; the result is intravascular hemolysis, resulting in hemoglobinuria (Table 129-6). One situation is acute and self-inflicted, march hemoglobinuria. Why sometimes a marathon runner may develop this complication, whereas on another occasion, this does not happen, we do not know (perhaps her or his footwear needs attention). A similar syndrome may develop after prolonged barefoot ritual dancing or intense playing of bongo drums. The other situation is chronic and iatrogenic (it has been called microangiopathic hemolytic anemia). It takes place in patients with prosthetic heart valves, especially when paraprosthetic regurgitation is present. If the hemolysis consequent on mechanical trauma to the red cells is mild, and if the supply of iron is adequate, the loss may be largely compensated; if more than mild anemia develops, reintervention to correct regurgitation may be required.
TABLE 129-6Diseases and Clinical Situations with Predominantly Intravascular Hemolysis ||Download (.pdf) TABLE 129-6Diseases and Clinical Situations with Predominantly Intravascular Hemolysis
| ||Onset/Time Course ||Main Mechanism ||Appropriate Diagnostic Procedure ||Comments |
|Mismatched blood transfusion ||Abrupt ||Nearly always ABO incompatibility ||Repeat cross-match || |
|Paroxysmal nocturnal hemoglobinuria (PNH) ||Chronic with acute exacerbations ||Complement (C)-mediated destruction of CD59(−) red cells ||Flow cytometry to display a CD59(−) red cell population ||Exacerbations due to C activation through any pathway |
|Paroxysmal cold hemoglobinuria (PCH) ||Acute ||Immune lysis of normal red cells ||Test for Donath-Landsteiner antibody ||Often triggered by viral infection |
|Septicemia ||Very acute ||Exotoxins produced by Clostridium perfringens ||Blood cultures ||Other organisms may be responsible |
|Microangiopathic ||Acute or chronic ||Red cell fragmentation ||Red cell morphology on blood smear ||Different causes ranging from endothelial damage to hemangioma to leaky prosthetic heart valve |
|March hemoglobinuria ||Abrupt ||Mechanical destruction ||Targeted history taking || |
|Favism ||Acute ||Destruction of older fraction of G6PD-deficient red cells ||G6PD assay ||Triggered by ingestion of large dish of fava beans, but trigger can be infection or drug instead |
By far the most frequent infectious cause of HA, in endemic areas, is malaria (Chap. 248). In other parts of the world, the most frequent direct cause is probably Shiga toxin–producing E. coli O157:H7, now recognized as the main etiologic agent of HUS, which is more common in children than in adults (Chap. 149). Life-threatening intravascular hemolysis, due to a toxin with lecithinase activity, occurs with Clostridium perfringens sepsis, particularly following open wounds, septic abortion, or as a disastrous accident due to a contaminated blood unit. Rarely, and if at all in children, HA is seen with sepsis or endocarditis from a variety of organisms. In addition, bacterial and viral infections can cause HA by indirect mechanisms (see above section on G6PD deficiency and Table 129-6).
These can arise through at least two distinct mechanisms. (1) There is a true autoantibody directed against a red cell antigen, i.e., a molecule present on the surface of red cells. (2) When an antibody directed against a certain molecule (e.g., a drug) reacts with that molecule, red cells may get caught in the reaction, whereby they are damaged or destroyed. Because the antibodies involved differ in optimum reactivity temperatures, they are classified in the time-honored categories of “cold” and “warm” (Table 129-7). Autoantibody-mediated HAs may be seen in isolation (when they are called idiopathic) or as part of a systemic autoimmune disorder such as systemic lupus erythematosus. Here we discuss the most distinctive clinical pictures.
TABLE 129-7Classification of Acquired Immune Hemolytic Anemias ||Download (.pdf) TABLE 129-7Classification of Acquired Immune Hemolytic Anemias
| ||Type of Antibody |
|Clinical Setting ||Cold, Mostly IgM, Optimal Temperature 4–30°C ||Warm, Mostly IgG, Optimal Temperature 37°C; or Mixed |
|Primary ||CAD ||AIHA (idiopathic) |
|Secondary to viral infection || |
|Secondary to other infection ||Mycoplasma infection: paroxysmal cold hemoglobinuria || |
|Secondary to/associated with other disease || |
Chronic inflammatory disorders (e.g., IBD)
After allogeneic HSCT
|Secondary to drugs: drug-induced immune hemolytic anemia ||Small minority (e.g., with lenalidomide) ||Majority: currently most common culprit drugs are cefotetan, ceftriaxone, piperacillin |
| ||Drug-dependent: antibody destroys red cells only when drug present (e.g., rarely penicillin) |
| ||Drug-independent: antibody can destroy red cells even when drug no longer present (e.g., methyldopa) |
AUTOIMMUNE HEMOLYTIC ANEMIA (AIHA)
Once a red cell is coated by an autoantibody (see  above), it will be destroyed by one or more mechanisms. In most cases, the Fc portion of the antibody will be recognized by the Fc receptor of macrophages, and this will trigger erythrophagocytosis. Thus, destruction of red cells will take place wherever macrophages are abundant, i.e., in the spleen, liver, or bone marrow; this is called extravascular hemolysis (Fig. 129-8). Because of the special anatomy of the spleen, this organ is particularly efficient in trapping antibody-coated red cells, and often this is the predominant site of red cell destruction. In some cases, the nature of the antibody is such (usually an IgM antibody) that the antigen-antibody complex on the surface of red cells is able to activate complement (C); as a result, a large amount of membrane attack complex will form, and the red cells may be destroyed directly; this is known as intravascular hemolysis.
Mechanism of antibody-mediated immune destruction of red blood cells (RBCs). ADCC, antibody-dependent cell-mediated cytotoxicity. (From N Young et al: Clinical Hematology. Philadelphia, Elsevier, 2006; with permission.)
AIHA is a serious condition; without appropriate treatment, it may have a mortality of approximately 10%. The onset is often abrupt and can be dramatic. The hemoglobin level can drop, within days, to as low as 4 g/dL; the massive red cell removal will produce jaundice; and sometimes the spleen is enlarged. When this triad is present, the suspicion of AIHA must be high. When hemolysis is (in part) intravascular, the telltale sign will be hemoglobinuria, which the patient may report or about which we must enquire or test for. The diagnostic test for AIHA is the direct antiglobulin test developed in 1945 by R. R. A. Coombs and known since by this name. The beauty of this test is that it detects directly the pathogenetic mediator of the disease, i.e., the presence of antibody on the red cells themselves. When the test is positive, it clinches the diagnosis; when it is negative, the diagnosis is unlikely. However, the sensitivity of the Coombs test varies depending on the technique that is used, and in doubtful cases, a repeat in a specialized lab is advisable; the term Coombs-negative AIHA is a last resort. In some cases, the autoantibody has a defined identity; it may be specific for an antigen belonging to the Rhesus system (it is often anti-e). In many cases, it is regarded as “nonspecific” because it reacts with virtually all types of red cells.
When AIHA develops in a person who is already known to have, for instance, systemic lupus or chronic lymphocytic leukemia (Table 129-7), we call it a complication; conversely, when AIHA presents on its own, it may be a pointer to an underlying condition that we ought to seek out. In both cases, what brings about AIHA remains, as in other autoimmune disorders, obscure. In some cases, AIHA can be associated, on first presentation or subsequently, with autoimmune thrombocytopenia (Evans’ syndrome).
TREATMENT Autoimmune Hemolytic Anemia
Severe acute AIHA can be a medical emergency. The immediate treatment almost invariably includes transfusion of red cells. This may pose a special problem because, if the antibody involved is nonspecific, all of the blood units cross-matched will be incompatible. In these cases, it is often correct, paradoxically, to transfuse incompatible blood, with the rationale being that the transfused red cells will be destroyed no less but no more than the patient’s own red cells, but in the meantime, the patient stays alive. A situation like this requires close liaison and understanding between the clinical unit treating the patient and the blood transfusion/serology lab. Whenever the anemia is not immediately life-threatening, blood transfusion should be withheld (because compatibility problems may increase with each unit of blood transfused), and medical treatment started immediately with prednisone (1 mg/kg per day), which will produce a remission promptly in at least one-half of patients. Rituximab (anti-CD20) was regarded as second-line treatment, but it is increasingly likely that a relatively low dose (100 mg/wk × 4) of rituximab together with prednisone will become a first-line standard. It is especially encouraging that this approach seems to reduce the rate of relapse, a common occurrence in AIHA. For patients who do relapse or are refractory to medical treatment, one may have to consider splenectomy, which, although it does not cure the disease, can produce significant benefit by removing a major site of hemolysis, thus improving the anemia and/or reducing the need for other therapies (e.g., the dose of prednisone). Since the introduction of rituximab, azathioprine, cyclophosphamide, cyclosporine, and intravenous immunoglobulin have become second- or third-line agents. In very rare severe refractory cases, either autologous or allogeneic hematopoietic stem cell transplantation may have to be considered.
PAROXYSMAL COLD HEMOGLOBINURIA (PCH)
PCH is a rather rare form of AIHA occurring mostly in children, usually triggered by a viral infection, usually self-limited, and characterized by the involvement of the so-called Donath-Landsteiner antibody. In vitro, this antibody has unique serologic features; it has anti-P specificity and binds to red cells only at a low temperature (optimally at 4°C), but when the temperature is shifted to 37°C, lysis of red cells takes place in the presence of complement. Consequently, in vivo there is intravascular hemolysis, resulting in hemoglobinuria. Clinically the differential diagnosis must include other causes of hemoglobinuria (Table 129-6), but the presence of the Donath-Landsteiner antibody will prove PCH. Active supportive treatment, including blood transfusion, is needed to control the anemia; subsequently, recovery is the rule.
COLD AGGLUTININ DISEASE (CAD)
This designation is used for a form of chronic AIHA that usually affects the elderly and has special clinical and pathologic features. First, the term cold refers to the fact that the autoantibody involved reacts with red cells poorly or not at all at 37°C, whereas it reacts strongly at lower temperatures. As a result, hemolysis is more prominent the more the body is exposed to the cold. The antibody is usually IgM; usually it has an anti-I specificity (the I antigen is present on the red cells of almost everybody), and it may have a very high titer (1:100,000 or more has been observed). Second, the antibody is produced by an expanded clone of B lymphocytes, and sometimes its concentration in the plasma is high enough to show up as a spike in plasma protein electrophoresis, i.e., as a monoclonal gammopathy. Third, because the antibody is IgM, CAD is related to Waldenström’s macroglobulinemia (WM) (Chap. 136), although in most cases, the other clinical features of this disease are not present. Thus, CAD must be regarded as a form of WM (i.e., as a low-grade mature B cell lymphoma) that manifests at an earlier stage precisely because the unique biologic properties of the IgM that it produces give the clinical picture of chronic HA.
In mild forms of CAD, avoidance of exposure to cold may be all that is needed to enable the patient to have a reasonably comfortable quality of life; but in more severe forms, the management of CAD is not easy. Blood transfusion is not very effective because donor red cells are I positive and will be rapidly removed. Immunosuppressive/cytotoxic treatment with azathioprine or cyclophosphamide can reduce the antibody titer, but clinical efficacy is limited, and in view of the chronic nature of the disease, the side effects may prove unacceptable. Unlike in AIHA, prednisone and splenectomy are ineffective. Plasma exchange will remove antibody and is, therefore, in theory, a rational approach, but it is laborious and must be carried out at frequent intervals if it is to be beneficial. The management of CAD has changed significantly with the advent of rituximab; although its impact on CAD is not as great as on AIHA, up to 60% of patients respond, and remissions may be more durable with a rituximab-fludarabine combination. Given the long clinical course of CAD, it remains to be seen with what schedule or periodicity these agents will need to be administered.
A number of chemicals with oxidative potential, whether medicinal or not, can cause hemolysis even in people who are not G6PD deficient (see above). Examples are hyperbaric oxygen (or 100% oxygen), nitrates, chlorates, methylene blue, dapsone, cisplatin, and numerous aromatic (cyclic) compounds. Other chemicals may be hemolytic through nonoxidative, largely unknown mechanisms; examples include arsine, stibine, copper, and lead. The HA caused by lead poisoning is characterized by basophilic stippling; it is in fact a phenocopy of that seen in P5N deficiency (see above), suggesting it is mediated at least in part by lead inhibiting this enzyme.
In these cases, hemolysis appears to be mediated by a direct chemical action on red cells. But drugs can cause hemolysis through at least two other mechanisms. (1) A drug can behave as a hapten and induce antibody production; in rare subjects, this happens, for instance, with penicillin. Upon a subsequent exposure, red cells are caught, as innocent bystanders, in the reaction between penicillin and antipenicillin antibodies. Hemolysis will subside as soon as penicillin administration is stopped. (2) A drug can trigger, perhaps through mimicry, the production of an antibody against a red cell antigen. The best known example is methyldopa, an antihypertensive agent no longer in use, which in a small fraction of patients stimulated the production of the Rhesus antibody anti-e. In patients who have this antigen, the anti-e is a true autoantibody, which then causes an autoimmune HA (see below). Usually this will gradually subside once methyldopa is discontinued.
Severe intravascular hemolysis can be caused by the venom of certain snakes (cobras and vipers), and HA can also follow spider bites.
Paroxysmal Nocturnal Hemoglobinuria (PNH)
PNH is an acquired chronic HA characterized by persistent intravascular hemolysis subject to recurrent exacerbations. In addition to hemolysis, there is often pancytopenia and a distinct tendency to venous thrombosis. This triad makes PNH a truly unique clinical condition; however, when not all of these three features are manifest on presentation, the diagnosis is often delayed, although it can always be made by appropriate laboratory investigations (see below).
PNH has about the same frequency in men and women and is encountered in all populations throughout the world, but it is a rare disease; its prevalence is estimated to be approximately 5 per million (it may be somewhat less rare in Southeast Asia and in the Far East). There is no evidence of inherited susceptibility. PNH has never been reported as a congenital disease, but it can present in small children or as late as in the seventies, although most patients are young adults.
The patient may seek medical attention because, one morning, she or he passed blood instead of urine (Fig. 129-9). This distressing or frightening event may be regarded as the classical presentation; however, more frequently, this symptom is not noticed or is suppressed. Indeed, the patient often presents simply as a problem in the differential diagnosis of anemia, whether symptomatic or discovered incidentally. Sometimes, the anemia is associated from the outset with neutropenia, thrombocytopenia, or both, thus signaling an element of bone marrow failure (see below). Some patients may present with recurrent attacks of severe abdominal pain defying a specific diagnosis and eventually found to be related to thrombosis. When thrombosis affects the hepatic veins, it may produce acute hepatomegaly and ascites, i.e., a full-fledged Budd-Chiari syndrome, which, in the absence of liver disease, ought to raise the suspicion of PNH.
Consecutive urine samples from a patient with paroxysmal nocturnal hemoglobinuria (PNH). The variation in the severity of hemoglobinuria within hours is probably unique to this condition.
The natural history of PNH can extend over decades. Without treatment, the median survival is estimated to be about 8–10 years; in the past, the most common cause of death has been venous thrombosis, followed by infection secondary to severe neutropenia and hemorrhage secondary to severe thrombocytopenia. Rarely (estimated 1–2% of all cases), PNH may terminate in acute myeloid leukemia. On the other hand, full spontaneous recovery from PNH has been documented, albeit rarely.
LABORATORY INVESTIGATIONS AND DIAGNOSIS
The most consistent blood finding is anemia, which may range from mild to moderate to very severe. The anemia is usually normomacrocytic, with unremarkable red cell morphology. If the MCV is high, it is usually largely accounted for by reticulocytosis, which may be quite marked (up to 20%, or up to 400,000/μL). The anemia may become microcytic if the patient is allowed to become iron deficient as a result of chronic urinary blood loss through hemoglobinuria. Unconjugated bilirubin is mildly or moderately elevated; LDH is typically markedly elevated (values in the thousands are common); and haptoglobin is usually undetectable. All of these findings make the diagnosis of hemolytic anemia compelling. Hemoglobinuria may be overt in a random urine sample; if it is not, it may be helpful to obtain serial urine samples, because hemoglobinuria can vary dramatically from day to day and even from hour to hour. The bone marrow is usually cellular, with marked to massive erythroid hyperplasia, often with mild to moderate dyserythropoietic features (not to be confused with myelodysplastic syndrome). At some stage of the disease, the marrow may become hypocellular or even frankly aplastic (see below).
The definitive diagnosis of PNH must be based on the demonstration that a substantial proportion of the patient’s red cells have an increased susceptibility to complement (C), due to the deficiency on their surface of proteins (particularly CD59 and CD55) that normally protect the red cells from activated C. The sucrose hemolysis test is unreliable; in contrast, the acidified serum (Ham) test is highly reliable but is carried out only in a few labs. The gold standard today is flow cytometry, which can be carried out on granulocytes as well as on red cells. A bimodal distribution of cells, with a discrete population that is CD59 and CD55 negative, is diagnostic of PNH. In PNH patients, this population is at least 5% of the total red cells and at least 20% of the total granulocytes.
Hemolysis in PNH is mainly intravascular and is due to an intrinsic abnormality of the red cell, which makes it exquisitely sensitive to activated C, whether it is activated through the alternative pathway or through an antigen-antibody reaction. The former mechanism is mainly responsible for chronic hemolysis in PNH; the latter explains why the hemolysis can be dramatically exacerbated in the course of a viral or bacterial infection. Hypersusceptibility to C is due to deficiency of several protective membrane proteins (Fig. 129-10), of which CD59 is the most important, because it hinders the insertion into the membrane of C9 polymers. The molecular basis for the deficiency of these proteins has been pinpointed not to a defect in any of the respective genes, but rather to the shortage of a unique glycolipid molecule, GPI (Fig. 129-2), which, through a peptide bond, anchors these proteins to the surface membrane of cells. The shortage of GPI is due in turn to a mutation in an X-linked gene, called PIG-A, required for an early step in GPI biosynthesis. In virtually each patient, the PIG-A mutation is different. This is not surprising, because these mutations are not inherited; rather, each one takes place de novo in a hemopoietic stem cell (i.e., they are somatic mutations). As a result, the patient’s marrow is a mosaic of mutant and nonmutant cells, and the peripheral blood always contains both PNH cells and normal (non-PNH) cells. Thrombosis is one of the most immediately life-threatening complications of PNH and yet one of the least understood in its pathogenesis. It could be that deficiency of CD59 on the PNH platelet causes inappropriate platelet activation; however, other mechanisms are possible.
The complement cascade and the fate of red cells. A. Normal red cells are protected from complement activation and subsequent hemolysis by CD55 and CD59. These two proteins, being GPI-linked, are missing from the surface of PNH red cells as a result of a somatic mutation of the X-linked PIG-A gene that encodes a protein required for an early step of the GPI molecule biosynthesis. B. In the steady state, PNH erythrocytes suffer from spontaneous (tick-over) complement activation, with consequent intravascular hemolysis through formation of the membrane attack complex (MAC); when extra complement is activated through the classical pathway, an exacerbation of hemolysis will result. C. On eculizumab, PNH erythrocytes are protected from hemolysis from the inhibition of C5 cleavage; however, upstream complement activation may lead to C3 opsonization and possible extravascular hemolysis. GPI, glycosylphosphatidylinositol; PNH, paroxysmal nocturnal hemoglobinuria. (From L Luzzatto et al: Haematologica 95:523, 2010.)
BONE MARROW FAILURE (BMF) AND RELATIONSHIP BETWEEN PNH AND APLASTIC ANEMIA (AA)
It is not unusual that patients with firmly established PNH have a previous history of well-documented AA; indeed, BMF preceding overt PNH is probably the rule rather than the exception. On the other hand, sometimes a patient with PNH becomes less hemolytic and more pancytopenic and ultimately has the clinical picture of AA. Because AA is probably an organ-specific autoimmune disease, in which T cells cause damage to hematopoietic stem cells, the same may be true of PNH, with the specific proviso that the damage spares PNH stem cells. PIG-A mutations can be demonstrated in normal people, and there is evidence from mouse models that PNH stem cells do not expand when the rest of the bone marrow is normal. Thus, we can visualize PNH as always having two components: failure of normal hematopoiesis and massive expansion of a PNH clone. Findings supporting this notion include skewing of the T cell repertoire and the demonstration of GPI-reactive T cells in patients with PNH.
TREATMENT Paroxysmal Nocturnal Hemoglobinuria
Unlike other acquired hemolytic anemias, PNH may be a lifelong condition, and most patients receive supportive treatment only, including transfusion of filtered red cells1 whenever necessary, which, for some patients, means quite frequently. Folic acid supplements (at least 3 mg/d) are mandatory; the serum iron should be checked periodically, and iron supplements should be administered as appropriate. Long-term glucocorticoids are not indicated because there is no evidence that they have any effect on chronic hemolysis; in fact, they are contraindicated because their side effects are considerable and potentially dangerous. A major advance in the management of PNH has been the development of a humanized monoclonal antibody, eculizumab, which binds to the complement component C5 near the site that, when cleaved, will trigger the distal part of the complement cascade leading to the formation of membrane attack complex (MAC). In an international, placebo-controlled, randomized trial of 87 patients (so far the only controlled therapeutic trial in PNH) who had been selected on grounds of having severe hemolysis making them transfusion-dependent, eculizumab proved effective and was licensed in 2007. Eculizumab, by abrogating complement-dependent intravascular hemolysis, significantly improves the quality of life of PNH patients. One would expect that the need for blood transfusion would also be abrogated; indeed, this is the case in about one-half of patients, in many of whom there is also a rise in hemoglobin levels. In the remaining patients, however, the anemia remains sufficiently severe to require blood transfusion. One reason for this is that, once the distal complement pathway is blocked, red cells no longer destroyed by the MAC become opsonized by complement (C3) fragments and undergo extravascular hemolysis (Fig. 129-10). The extent to which this happens depends in part on a genetic polymorphism of the complement receptor CR1. Based on its half-life, eculizumab must be administered intravenously every 14 days. The only form of treatment that currently can provide a definitive cure for PNH is allogeneic BMT. When an HLA-identical sibling is available, BMT should be offered to any young patient with severe PNH; the availability of eculizumab has decreased significantly the proportion of patients receiving BMT.
For patients with the PNH-AA syndrome, immunosuppressive treatment with antithymocyte globulin and cyclosporine A may be indicated, especially in order to relieve severe thrombocytopenia and/or neutropenia in patients in whom these were the main problem(s); of course, this treatment will have little or no effect on hemolysis. Any patient who has had venous thrombosis or who has a genetically determined thrombophilic state in addition to PNH should be on regular anticoagulant prophylaxis. With thrombotic complications that do not resolve otherwise, thrombolytic treatment with tissue plasminogen activator may be indicated.
ANEMIA DUE TO ACUTE BLOOD LOSS
Blood loss causes anemia by two main mechanisms: (1) by the direct loss of red cells; and (2) if the loss of blood is protracted, it will gradually deplete iron stores, eventually resulting in iron deficiency. The latter type of anemia is covered in Chap. 126; here we are concerned with the former type, i.e., posthemorrhagic anemia, which follows acute blood loss. This can be external (e.g., after trauma or obstetric hemorrhage) or internal (e.g., from bleeding in the gastrointestinal tract, rupture of the spleen, rupture of an ectopic pregnancy, subarachnoid hemorrhage). In any of these cases, after the sudden loss of a large amount of blood, there are three clinical/pathophysiologic stages. (1) At first, the dominant feature is hypovolemia, which poses a threat particularly to organs that normally have a high blood supply, like the brain and the kidneys; therefore, loss of consciousness and acute renal failure are major threats. It is important to note that at this stage an ordinary blood count will not show anemia, because the hemoglobin concentration is not affected. (2) Next, as an emergency response, baroreceptors and stretch receptors will cause release of vasopressin and other peptides, and the body will shift fluid from the extravascular to the intravascular compartment, producing hemodilution; thus, the hypovolemia gradually converts to anemia. The degree of anemia will reflect the amount of blood lost. If after 3 days the hemoglobin is, for example, 7 g/dL, it means that about half of the entire blood has been lost. (3) Provided bleeding does not continue, the bone marrow response will gradually ameliorate the anemia.
The diagnosis of acute posthemorrhagic anemia (APHA) is usually straightforward, although sometimes internal bleeding episodes (e.g., after a traumatic injury), even when large, may not be immediately obvious. Whenever an abrupt fall in hemoglobin has taken place, whatever history is given by the patient, APHA should be suspected. Supplementary history may have to be obtained by asking the appropriate questions, and appropriate investigations (e.g., a sonogram or an endoscopy) may have to be carried out.
TREATMENT Anemia Due to Acute Blood Loss
With respect to treatment, a two-pronged approach is imperative. (1) In many cases, the blood lost needs to be replaced promptly. Unlike with many chronic anemias, when finding and correcting the cause of the anemia is the first priority and blood transfusion may not be even necessary because the body is adapted to the anemia, with acute blood loss the reverse is true; because the body is not adapted to the anemia, blood transfusion takes priority. (1) While the emergency is being confronted, it is imperative to stop the hemorrhage and to eliminate its source.
A special type of APHA is blood loss during and immediately after surgery, which can be substantial (e.g., up to 2 L in the case of a radical prostatectomy). Of course with elective surgical procedures, the patient’s own stored blood may be available (through preoperative autologous blood donation), and in any case, blood loss ought to have been carefully monitored/measured. The fact that this blood loss is iatrogenic dictates that ever more effort should be invested in optimizing its management.
A Holy Grail of emergency medicine for a long time has been the idea of a blood substitute that would be universally available, suitable for all recipients, easy to store and to transport, safe, and as effective as blood itself. Two main paths have been pursued: (1) fluorocarbon synthetic chemicals that bind oxygen reversibly, and (2) artificially modified hemoglobins, known as hemoglobin-based oxygen carriers (HBOCs). Although there are numerous anecdotal reports of the use of both approaches in humans, and although HBOCs have reached the stage of phase 2–3 clinical trials, no “blood substitute” has yet become standard treatment.