Anemias due to isoagglutinins (isoantibodies)

These anemias are hemolytic reactions caused by antibodies within the various blood group systems. The classification, symptomatology, and diagnostic procedures necessary for detection of such reactions and identification of the etiology are discussed in Chapter 9 and Chapter 11.

Anemias due to autoagglutinins (autoantibodies)

Autoagglutinins are antibodies produced by an individual against certain of his or her own body cells. This discussion concerns autoantibodies produced against his or her own RBCs. The anemia associated with this condition has been called autoimmune hemolytic anemia or acquired hemolytic anemia.
Autoantibodies of the autoimmune hemolytic anemias form two general categories: those that react best in vitro above room temperature (37°C, warm autoantibodies) and those that react best in vitro at cold temperatures (cold autoantibodies or cold agglutinins). For each type there are two general etiologies, idiopathic and secondary to some known disease.

Warm autoantibodies are IgG antibodies usually directed against Rh antigens on the RBC membrane. They comprise about 50%-70% of Coombs’-positive autoantibodies. The presence of the autoantibody and the RBC antigen against which it reacts can often (not always) be proven by detaching (eluting) the antibody from affected RBC. Clinical disease from warm autoantibodies is more frequent than clinical abnormality from cold autoantibodies, and the idiopathic variety is twice as frequent as that secondary to known disease. Clinically, anemia due to warm-reacting autoantibodies appears at any age and may be either chronic or acute. When chronic, it is often low grade. When acute, it is often severe and fatal. The laboratory signs are those of any hemolytic anemia and depend on the degree of anemia. Thus, there are varying degrees of reticulocyte elevation. The direct Coombs’ test result is usually, although not always, positive. Most patients have spherocytes in peripheral blood, especially if the anemia is acute; and splenomegaly is frequent.

Cold agglutinins are IgM antibodies usually directed against the I antigen on RBC membranes. These comprise about 15%-30% of Coombs’-positive autoantibodies. Complement can often be detected on affected RBC but no antibody usually can be eluted. Clinical disease from cold-reacting agglutinins is seen much less frequently than hemolytic disease from warm-reacting autoantibodies. Cold agglutinin disease is seen predominantly in adults, particularly in the elderly. The most common cause of symptomatic hemolytic anemia induced by cold agglutinins is mycoplasma infection. After mycoplasma-induced disease, the idiopathic and the secondary forms occur in nearly equal incidences. Clinically, the disease is often worse in cold weather. Raynaud’s phenomenon is common. Splenomegaly is not common. Laboratory abnormalities are not as marked as in the warm autoantibody type, except for a usually positive direct Coombs’ test result, and the anemia tends to be less severe. The reticulocyte count is usually increased but often only slightly. Spherocytes are more often absent than present. WBCs and platelets are usually normal unless altered by underlying disease. However, exceptions to these statements may occur, with severe hemolytic anemia present in all its manifestations. As noted in the discussion of mycoplasma pneumonia (Chapter 14), cold agglutinins may occur in many normal persons but only in titers up to 1:32. In symptomatic anemia due to cold agglutinins the cold agglutinin titer is almost always more than 1:1,000.

Paroxysmal cold hemoglobinuria (PCH)

Paroxysmal cold hemoglobinuria is a rare syndrome in which an antibody (Donath-Landsteiner antibody) of the IgG class binds to and sensitizes RBCs at cold temperatures and then produces complement-activated RBC lysis at warmertemperatures. PCH comprises about 2%-5% of Coombs’-positive autoantibodies, much more common in children than in adults. Paroxysmal cold hemoglobinuria was originally associated with syphilis, but more cases occur idiopathically or following viral infection than from syphilis. Hemoglobinuria is produced after patient exposure to cold temperatures and may be accompanied by back or leg pain, chills, and cramps, similar to symptoms of hemolytic transfusion reaction. The IgG-specific Coombs’ reagent produces positive direct Coombs’ test results at cold temperatures and Coombs’ reagents containing non-gamma non-IgG-specific antibody (sometimes called broad-spectrum Coombs’ reagent) produce positive direct Coombs’ test results at the usual Coombs’ test temperature of 37°C. The major diagnostic procedure for paroxysmal cold hemoglobinuria is the Donath-Landsteiner test, in which the development of hemolysis in patient and normal blood is compared at cold temperature.

Secondary acquired autoimmune hemolytic anemia

The causes of acquired hemolytic anemia of the secondary type, either warm or cold variety, can be divided into three main groups. The first group in order of frequency is leukemia and lymphoma; most often chronic lymphocytic leukemia, to a lesser extent lymphocytic lymphoma, and occasionally Hodgkin’s disease. The second group in order of frequency is collagen disease, notably lupus erythematosus. The third group is a miscellaneous collection of systemic diseases in which overtly hemolytic anemia rarely develops but may do so from time to time. These diseases include viral infections, severe liver disease, ovarian tumors, and carcinomatosis. It should be emphasized that in all three disease groups, anemia is a common or even frequent finding, but the anemia is usually not hemolytic, at least not of the overt or symptomatic type.

Drug-induced hemolytic anemia

Drug-induced hemolytic anemia is sometimes included with the autoimmune hemolytic anemias. However, in most cases antibodies are formed primarily against the drug, and action against the RBC is secondary to presence of the drug on the RBC surface. These cause about 10%-20% of Coombs’-positive autoantibodies. There are four basic mechanisms proposed, as follows:

Combination of the drug with antidrug antibody to form an immune complex that is adsorbed onto RBCs, often activating complement. Quinidine is the best-known drug of this type. The antiquinidine antibody is of the IgM class.
Binding of the drug to the RBC membrane and acting as a hapten. Penicillin (in very large doses, і10 million units/day for 7 days or more) is the major drug of this type, although abnormality develops in fewer than 3% of these cases.
Nonspecific coating of RBC by drug with absorption of various proteins. The antibiotic cephalothin has been shown to act by this mechanism. A positive direct Coombs’ test result is produced by antibodies against proteins absorbed onto the cell or onto cephalothin. There is no hemolysis, however. Cephalothin may occasionally act as a hapten and in these cases may be associated with hemolytic anemia.
Unknown mechanism. a-Methyldopa is the predominant drug of this type and may be the most common agent associated with drug-induced hemolytic anemia. The antimethyldopa antibody is of the IgG class and usually has Rh group specificity. Besides coating of RBCs, a-methyldopa–treated patients may have circulating autoantibodies demonstrated by an indirect Coombs’ test, which is unusual for other drugs. Patients taking a-methyldopa may also develop a syndrome resembling systemic lupus erythematosus, with antinuclear antibodies and lupus erythematosus cells (Chapter 23). Up to 25% of patients (literature range 10%-36%) develop a positive direct Coombs’ test, and about 1% (literature range 0%-5%) develop hemolytic anemia. The direct Coombs’ test result remains positive 1-24 months after the end of therapy.

Laboratory investigation of possible drug-induced hemolytic anemia is usually difficult for the ordinary laboratory. The procedure usually involves washing off (eluting) the antibody from the RBC, if possible, and trying to determine whether the antibody has specificity against drug-coated RBCs rather than normal RBCs.

Traumatic (microangiopathic) hemolytic anemia

This category includes several diseases that produce hemolytic anemia with many schistocytes, the schistocytes being formed through some kind of trauma. Representative conditions are disseminated intravascular coagulation and thrombotic thrombocytopenic purpura (in which RBCs strike fibrin clots in small vessels), the hemolytic-uremic syndrome (thrombi in renal glomerular capillaries and small vessels), the cardiac prosthesis syndrome (in which RBCs are damaged while passing through the artificial heart valve), and hemolytic anemia associated with vascular grafts and some long-term indwelling catheters. The same type of hemolytic anemia may be found in a few patients with malignancy (most commonly gastric carcinoma), in Zieve’s syndrome associated with cirrhosis, in the first few hours after extensive severe burns, and in Clostridium welchii septicemia. As noted in Chapter 2, schistocytes can be found in smaller numbers in other conditions. Microangiopathic hemolytic anemia is discussed in greater length in Chapter 8.

Paroxysmal nocturnal hemoglobinuria (PNH)

Patients with paroxysmal nocturnal hemoglobinuria (PNH) develop an acquired blood cell membrane defect in which RBCs, WBCs, and platelets demonstrate abnormal sensitivity to the effect of activated serum complement. This is manifest by hemolytic anemia, granulocytopenia, and thrombocytopenia. Not all patient RBCs have the same degree of abnormality, and resistance to lysis varies from relatively normal to markedly abnormal. It is often associated with aplastic anemia and is said to develop in 5%-10% of these patients without regard to the cause of the marrow depression (with the exception that PNH is not associated with radiation marrow damage). It may appear either at the beginning of aplasia, during the aplastic period, or during recovery. About 50% of cases develop without prior evidence of aplastic marrow. It may also develop in some patients with erythroleukemia, myelofibrosis, or refractory anemia.

RBCs that are abnormally sensitive to complement have markedly decreased acetylcholinesterase levels, but this is not thought to be the cause of the defect in PNH.

Paroxysmal nocturnal hemoglobinuria most often affects young or middle-aged adults, with the usual age range being 10-60 years. The disease presents as hypoplastic anemia in about 25% of cases, as an episode of abdominal pain in about 10%, and with hemoglobinuria in about 50%. Clinically, there is a chronic hemolytic anemia, with crisis episodes of hemoglobinuria occurring most often at night. However, hemoglobinuria is present at disease onset only in about 50% of cases. Another 20% develop it within 1 year, and eventually it occurs in more than 90% of patients. Anemia is usually of moderate degree except during crisis, when it may be severe. A crisis is reflected by all the usual laboratory parameters of severe hemolysis, including elevated plasma hemoglobin levels. No spherocytosis or demonstrable antibodies are present. The disease gets its name because hemoglobinuric episodes turn urine collected during or just after sleep to red or brown due to large amounts of hemoglobin. Urine formed during the day is clear. Stimuli known to precipitate attacks in some patients include infections, surgery, and blood transfusion.

Laboratory findings. In addition to anemia, leukopenia (granulocytopenia) is present in about 50% of patients, and some degree of thrombocytopenia is present in about 70%. This is in contrast to most other hemolytic anemias, in which hemolysis usually provokes leukocytosis. The MCV is elevated in about 83%, normal in about 13%, and decreased in about 5%. The reticulocyte count is elevated in about 90%. Loss of iron in the urine (in the form of hemoglobin and hemosiderin) leads to chronic iron deficiency in some patients. For some reason the kidney in PNH is not damaged by the hemoglobin or by renal tubular cell deposition of hemosiderin.

Venous thrombosis is frequent in PNH, and patients have a considerably increased tendency toward infection (predominantly lung and urinary tract). There may be episodes of abdominal pain related to venous thrombosis.

Tests for paroxysmal nocturnal hemoglobinuria. A good screening test is a urine hemosiderin examination. However, a positive urine hemosiderin value may be obtained in many patients with chronic hemolytic anemia of various types and also may be produced by frequent blood transfusions, especially if these are given over periods of weeks or months. A much more specific test is the acid hemolysis (Ham) test. The RBCs of PNH are more susceptible to hemolysis in acid pH. Therefore, serum is acidified to a certain point that does not affect normal RBCs but will hemolyze the RBCs of PNH. Another widely used procedure is the sugar-water (sucrose hemolysis) test, which is easier to perform than the Ham test and may be more sensitive. It is based on evidence that RBCs in PNH are more susceptible to hemolysis in low ionic strength media than normal RBCs. Many laboratories screen with the sugar-water test and confirm a positive result with the Ham test. The sugar-water test is apt to produce more weak positive reactions in patients who do not have verifiable PND than does the Ham test. In my experience (also reported by others) there occasionally is discrepancy between results of the sugar-water test and the Ham test in the same patient, resulting in diagnostic problems.

Hemolytic anemia due to toxins

Chemical. Lead poisoning is the most frequent cause in this group. Ingestion of paint containing lead used to be frequent in children and still happens occasionally. Auto battery lead, gasoline fumes, and homemade whiskey distilled in lead-containing apparatus are the most common causes in adults. It takes several weeks of chronic exposure to develop symptoms unless a large dose is ingested. The anemia produced is most often mild to moderate, and the usual reason for seeking medical treatment is development of other systemic symptoms, such as convulsions from lead encephalopathy, abdominal pain, or paresthesias of hands and feet. The anemia is more often hypochromic but may be normochromic; it is usually normocytic. Basophilic stippling of RBCs is often very pronounced and is a classic diagnostic clue to this condition. Basophilic stippling may occur in any severe anemia, especially the hemolytic anemias, but when present to an unusual degree should suggest lead poisoning unless the cause is already obvious. The stippled cells are reticulocytes, which, for some unknown reason, appear in this form in these patients. However, in some patients, basophilic stippling is minimal or absent. Tests useful in lead poisoning for screening purposes or for diagnosis are discussed in Chapter 35.

Other chemicals were mentioned in the discussion of G-6-PD deficiency anemia. Benzene toxicity was discussed in the section on hypoplastic bone marrow anemias. Other chemicals that often produce a hemolytic anemia if taken in sufficient dose include naphthalene, toluene, phenacetin, and distilled water given intravenously. Severe extensive burns often produce acute hemolysis to varying degrees.

Bacterial. Clostridium welchii septicemia often produces a severe hemolytic anemia with spherocytes. Hemolytic anemia is rarely seen with tuberculosis. The anemia of infection is usually not overtly hemolytic, although there may be a minor hemolytic component (not demonstrable by the usual laboratory tests).

Hemolytic anemia due to parasites

Among hemolytic anemias due to parasites, malaria is by far the most frequent. It must be considered in persons who have visited endemic areas and who have suggestive symptoms or no other cause for their anemia. The diagnosis is made from peripheral blood, best obtained morning and afternoon for 3 days. Organisms within parasitized RBCs may be few and often are missed unless the laboratory is notified that malaria is suspected. A thick-drop special preparation is the method of choice for diagnosis. With heavy infection, the parasites may be identified on an ordinary (thin) peripheral blood smear. A hemolytic anemia is produced with the usual reticulocytosis and other laboratory abnormalities of hemolysis. Most patients have splenomegaly. Bartonella infection occurs in South America, most often in Peru. This is actually a bacterium rather than a parasite, but in many textbooks it is discussed in the parasite category. The organisms infect RBCs and cause hemolytic anemia clinically similar to malaria. Babesiosis is an uncommon protozoan infection of RBC similar in some respects to malaria. This condition is discussed in Chapter 18.


Hypersplenism is a poorly understood entity whose main feature is an enlarged spleen associated with a deficiency in one or more blood cell elements. The most common abnormality is thrombocytopenia, but there may be a pancytopenia or any combination of anemia, leukopenia, and thrombocytopenia. Hypersplenism may be primary or, more commonly, secondary to any disease that causes splenic enlargement. However, splenic enlargement in many cases does not produce hypersplenism effects. Portal hypertension with secondary splenic congestion is the most common etiology; the usual cause is cirrhosis. If anemia is produced in hypersplenism, it is normocytic and normochromic without reticulocytosis. Bone marrow examination in hypersplenism shows either mild hyperplasia of the deficient peripheral blood element precursors or normal marrow.

Several mechanisms have been proposed to explain the various effects of hypersplenism. To date, the weight of evidence favors sequestration in the spleen. In some cases, the spleen may destroy blood cells already damaged by immunologic or congenital agents. In some cases, the action of the spleen cannot be completely explained.