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The spherocytic Haemolytic Anaemias

2001; Wiley; Volume: 112; Issue: 4 Linguagem: Inglês

10.1046/j.1365-2141.2001.02440.x

1365-2141

Charles H. Packman,

Neonatal Health and Biochemistry

The diagnosis of haemolytic anaemia is made when circulating red blood cell (RBC) survival is shortened. Today, the laboratory findings in haemolytic anaemia and the distinction between inherited and acquired forms are studied by preclinical medical students the world over. Although haemolytic anaemia is much less common than anaemia as a result of marrow failure or bleeding, general physicians are well familiar with the characteristic findings in the condition and, by the time the patient is seen by a haematologist, the diagnosis is usually already made. Among the spherocytic haemolytic anaemias, hereditary spherocytosis is the most common inherited haemolytic anaemia and autoimmune haemolytic disease is the most common acquired form. The two diseases share many features that were known to clinicians in the early part of the twentieth century: jaundice and splenomegaly on physical examination, spherocytes and reticulocytosis on the blood film, increased osmotic fragility of RBCs, and clinical improvement following splenectomy. This constellation of clinical features was generally designated haemolytic or acholuric jaundice (icterus), acholuric referring to the characteristic absence of bile in the urine. A congenital or familial form of haemolytic anaemia was often diagnosed in children with a family history of jaundice, splenomegaly or haemolytic anaemia. However, spontaneous cases in children without a family history and the initial appearance of the disease in adults were common enough to cause confusion, as acquired cases were often diagnosed on the basis of acuity and absent family history in adult patients. No one seriously questioned the existence of familial or congenital forms of haemolytic anaemia, but many questioned the existence of acquired forms other than those obviously as a result of infection or toxic exposure. It was not until the rediscovery of the principle of anti-globulin testing and its application to diagnosis of patients with spherocytic haemolytic anaemia that the existence of acquired spherocytic haemolytic anaemia was accepted and the means to easily distinguish the inherited and acquired forms was at hand. It is perhaps a unique aspect of these disorders that the underlying principles of our best current diagnostic tests, i.e. spherocytes on the blood film, the osmotic fragility test and the anti-globulin test, have been known for nearly a century or more and applied, with a few refinements, for over half a century. This review will trace the evolution of understanding of the spherocytic haemolytic anaemias from the earliest case descriptions to the middle of the twentieth century when it all came together. Perhaps the first case of haemolytic anaemia in the medical literature was described by the physician Claudius Galen (Dreyfus, 1942). The patient, a slave of the emperor Marcus Aurelius, hunted snakes. He was bitten by a viper and, in spite of treatment with the 'usual drugs', his skin 'turned the colour of a ripe leek.' Galen prescribed theriaca, which cured the patient. Galen explained, 'The spleen produces colours of this type, darker even than those produced by the liver. They are difficult to explain, but easy to recognize if you have seen them often.' The first recognizable reports of haemolytic anaemia were undoubtedly cases of paroxysmal cold haemoglobinuria because of the dramatic presentation. Johannis Actuarius, a court physician at Constantinople during the late thirteenth century described azure, livid and black urine following chilling of certain individuals (Major, 1945). The syndrome was well known by physicians of the mid-nineteenth century. A relationship to jaundice was recognized and the absence of RBCs from the dark urine was noted. These patients were thought to suffer from a disorder of the kidneys. Congenital or familial cases Towards the end of the nineteenth century, physicians began to note certain chronically icteric patients who had no bile in the urine, no evidence of liver disease and often, but not always, splenomegaly and a family history of jaundice. Vanlair and Masius (1871) described the case of a young woman who developed icterus, recurrent attacks of left upper quadrant abdominal pain and splenomegaly shortly after giving birth. The stools were not light coloured, but rather deeply pigmented. The patient's mother and sister were also icteric, and the sister's spleen was enlarged. The most remarkable aspect of this paper lies in their description of the blood findings. Although they made no mention of anaemia and had no concept of haemolysis as a pathological process, they unmistakably described spherocytes with remarkable clarity. The authors noted that some of the erythrocytes, which they called microcytes, were smaller than normal cells, 3–4 μm in diameter, spherical in shape (elle est parfaitement spherique) and the contours were completely smooth. Their drawing of the patient's blood cells and a companion drawing of normal cells is reproduced in Fig 1. With remarkable discernment, Vanlair and Masius stated, 'The jaundice of our patient appears to be a peculiar type of icterus. The fact that the patient's mother and sister had a slight jaundice and that the sister had an enlarged spleen may indicate that this condition is one disease entity.' Reproduction of a camera lucida drawing from the classic paper of Vanlair and Masius (1871), de la Microcythemie. Left, sang microcytique from their propositus with microcythemia. Right, sang normal. Murchison (1877), in his textbook, Clinical Lectures on Diseases of the Liver, describes a man who was 'born yellow' and who suffered from gout. His urine contained bile pigments and the liver was slightly enlarged. The patient's mother and brother also suffered from gout and jaundice. Unfortunately, there is no mention of their spleens or characteristics of their blood. Murchison was struck by the persistent and hereditary nature of the jaundice in the absence of obstruction of the bile duct. Hunter (1888), who made important contributions to the recognition of pernicious anaemia, may have been the first to classify anaemias by mechanism, i.e. haemorrhage, decreased production and increased destruction. He classified pernicious anaemia and the experimental anaemia produced in animals by destructive agents such as pyrogallic acid as haemolytic. He distinguished haemorrhagic and haemogenic anaemias from haemolytic anaemias by the presence of 'the small yellow microcytes, perfectly spherical in form and deep in colour which point to an anaemia of haemolytic origin.' Wilson's cases (Wilson, 1890; Wilson & Stanley, 1893) almost certainly represent hereditary haemolytic anaemia. Six consanguinous relatives in four generations of a family exhibited pallor, enlarged spleens and icterus. Several of them described attacks of chills, fever and increasing jaundice, reminiscent of the crises of what would later be called haemolytic jaundice. Sir William Gull had seen the propositus, 'Mrs. A.P.' and two of her children in 1857; he thought they had 'ague cake from catching malaria at Worthing.' Thirty years later, Wilson was less certain, stating (Wilson, 1890), '…the more I see and think of these cases, the less do they remind me of malaria.' Wilson measured RBC counts ranging from 3–4 million and haemoglobin levels ranging from 50–60% in three of the patients. The family tree of Mrs. A. P. (Wilson, 1890) is depicted in Fig 2. The family tree of 'Mrs. A. P.'Wilson (1890) studied six individuals from four generations of this consanguineous family. The sole surviving member of the family, 'Mr R. T.', was reported to have an abnormal osmotic fragility test when studied almost 40 years later (Campbell, 1926). Cases marked with an asterisk show enlarged spleen and sallow complexion. 'Mrs. T', the eldest daughter of Wilson's propositus, was sallow from the age of 5 or 6 years and exhibited splenomegaly from age 7 years. She died from complications of pregnancy and childbirth at age 34 years. Permission for a limited post-mortem examination was granted. The spleen weighed almost 2 lbs and measured 6·5 by 5 inches. Microscopically there was thickening of the capsule and trabeculae and engorgement with RBCs. The gall bladder was forgotten in the rush to complete the examination, a charming, but unfortunate, omission, given her history of occasional attacks of jaundice and pain that had been attributed to biliary concretions. Wilson attributed her death to 'rapidly progressing anaemia, dependent upon an active haemolysis of splenic origin.' By this time, Wilson was firmly convinced that his patients were not suffering from malaria. He thought that all the patients in this family were suffering from the same disease in spite of the differences in clinical history and pointed out that near relatives of these patients could suffer unknown from the condition. Thirty-five years later, Dr Wilson introduced the only survivor of his six patients to Dr J. M. H. Campbell (1926). 'Mr R. T.' was the son of 'Mrs. T.' and the grandson of Wilson's propositus, 'Mrs. A. P.' Mr R. T. was 44 years old at the time and an active tennis player. He was deeply jaundiced and the spleen projected more than 4 inches below the costal margin. The haemoglobin measured 80%, the plasma indirect Van den Bergh test was positive and the urine contained urobilin, but no bile pigment. Most importantly, the RBC exhibited increased fragility in hypotonic saline, confirming the diagnosis of familial acholuric jaundice. Minkowski (1900, 1905) reported important observations of two families with familial jaundice, splenomegaly and urobilinuria. The affected members of these families were in otherwise good health and one even became a physician. One of the patients died of pneumonia and, at post-mortem examination, Minkowski demonstrated the integrity of the liver, pigment gall stones and splenomegaly (1 kg), as well as siderosis of the kidneys. He stated 'We are concerned with a peculiar inborn affection which is characterized by permanent jaundice, the continued presence of urobilin in the urine, hyperplasia of the spleen and increased iron in the kidneys. It is evidently hereditary in origin and does not appear to affect length of life…this affection constitutes a peculiar anomaly of the metabolism of the blood pigment, perhaps following a primary alteration of the spleen.' The blood of Minkowski's patient was described as normal; he did not offer an opinion regarding the increased iron in the kidneys. Minkowski was probably unaware of Wilson's patients, but his clinical descriptions and the post-mortem findings agree closely with the observations of Wilson. Around the same time, Bettman in Heidelberg and Krannhals of Riga (Dreyfus, 1942) published other accounts of anaemia in patients with familial jaundice. Krannhals measured RBC counts ranging from 2·8 to 3·5 million and noted polychromatophilia for the first time. He also commented on the fact that the stools were coloured in spite of the jaundice. He postulated that the RBCs may have been haemolysed by the bile pigments in the serum. Barlow & Shaw (1902) similarly described two patients with familial jaundice and anaemia; they also receive credit for the first description of chronic leg ulcers in this disease. Osmotic fragility and reticulocytosis During the first decade of the new century, two important discoveries rounded out the clinical picture of haemolytic icterus: increased osmotic fragility of RBCs and reticulocytosis. Credit for priority in their description is generally given to the French investigator, Chauffard (Fig 3) and co-workers. Chauffard (1907) noted that the RBCs of several patients, but not those of normal subjects, were haemolysed by hypotonic saline. Almost 40 years earlier, Malassez (Dreyfus, 1942), who developed fluids for counting blood cells, had observed that RBCs from various patients lysed at different rates in 0·06% saline. In Chauffard's osmotic fragility test, RBCs were placed in a series of tubes containing successively decreasing concentrations of saline, not greatly different from current practice. The osmotic fragility was expressed as the concentration of saline at which haemolysis began and at which haemolysis was complete. Chauffard recognized that the liver was not at fault and that the disorder was as a result of haemolysis. He wrote, 'Perhaps after this clinical and haematologic inquiry, the cause of the haemolytic theory could be considered as won.' This observation finally enabled physicians to distinguish hepatic and haemolytic jaundice, as Ribbierre (Dreyfus, 1942) had recently (in 1903) demonstrated that red cells from patients with hepatic jaundice are resistant to osmotic stress. Anatole Chauffard. From the United States National Library of Medicine, History of Medicine Division. Of course, Chauffard and co-workers had discovered the in vitro pathophysiological expression of the spherical microcytes first described by Vanlair and Masius almost 40 years earlier. They were probably unaware of the work of these early investigators and they certainly made no association between microcytic spherical cells and increased osmotic fragility. That correlation was noted much later by Haden (1934). Naegli is often credited with first use of the term 'spherocyte.' However, according to Crosby (1952), two British army officers, Christophers and Bentley, were the first. They were assigned to India to study blackwater fever and made very careful descriptions of spherocytes in a monograph published in 1909. Naegli also proposed that the spherocyte was pathognomonic of congenital haemolytic icterus and was the result of abnormal erythropoiesis in the marrow. Dameshek & Schwartz (1940) wryly note, 'Like many other statements made by Naegli, this was soon accepted as gospel…' and the effect was to constrict thinking about haemolytic icterus for the next 15 or 20 years. In fact, many authorities began to doubt the existence of an acquired type of haemolytic icterus, regarding the disease as a variation on the congenital form. About a year after his description of increased osmotic fragility in congenital haemolytic icterus, Chauffard & Fiessinger (1907) and Chauffard (1908) stained RBCs from patients with haemolytic icterus with Pappenheim's solution and noted large numbers of cells containing a peculiar basophilic granulation or reticulum, which they called 'granular degeneration.' Ehrlich (Dreyfus, 1942) had first described this special staining method in 1881 and noted increased numbers of reticular cells in anaemic patients. Vaughan (1903) noted these granular cells constituted about 1% of the RBCs in normal subjects. Chauffard had hoped to explain the anatomical lesion that underlay the increased fragility of the RBCs. What he actually discovered, or rediscovered, was the reticulocytosis that is now a hallmark of haemolytic anaemia. Chauffard's drawing of a blood smear stained with Pappenheim stain from a patient with familial haemolytic icterus (Chauffard, 1908) is shown in Fig 4. Drawing of a blood smear (Pappenheim stain) as seen by Chauffard (1908). The granular appearing cells are reticulocytes from a patient with familial haemolytic icterus. Acquired cases Georges Hayem (1898)) (Fig 5) is generally given credit for the first description of acquired haemolytic icterus. In his paper entitled 'Sur une variete particulaire d'ictere chronique', he describes the several known varieties of jaundice and then continues, 'Among these cases, I will alert you to a chronic jaundice very particular to certain dyspeptics that manifest themselves by a yellow chamois colouration of the skin and most notably of the palms of the hands. Above all, this jaundice has the unique characteristic of the absence of biliary pigment in the urine while blood serum clearly shows Gmelin's reaction. I have observed several examples.' Hayem documented anaemia and splenomegaly in his patients, but considered the disease an affliction of the liver and placed great emphasis on the digestive disorders for which his patients had initially consulted him. He proposed the name 'chronic infectious icterus of the paroxysmal type, with splenomegaly.' His was the first clear description of the acholuric nature of the jaundice. Georges Hayem. From the United States National Library of Medicine, History of Medicine Division. At about the same time as Chauffard reported his cases of congenital haemolytic icterus, his colleagues Widal et al (1907a, b, c, 1909) (Fig 6) published a series of papers on a haemolytic icterus that was apparently neither congenital nor familial, that could appear gradually or suddenly during the course of various diseases or could be unassociated with any underlying disease. These cases were considered similar to those described by Hayem 10 years earlier. In contrast to Chauffard's patients, whom he described as more icteric than ill (icterique plutot que vraiment malade), Widal's patients were severely anaemic and ill. They were all adults who lacked a family history or past history of icterus. The patients exhibited reticulocytosis, but the alterations in the fragility test were less marked than in the congenital form. Autoagglutination was observed in some of these patients and Chauffard & Vincent (1909) later described isohaemolysins in the sera of two of Widal's patients. For a time thereafter, two types of haemolytic icterus were distinguished, the congenital or familial type of Chauffard and Minkowski (Dr Wilson's precedence notwithstanding), and the acquired type of Hayem and Widal. Fernand Widal. From the United States National Library of Medicine, History of Medicine Division. Splenectomy The postulated role of the spleen in the pathogenesis of the anaemia led to attempts to cure the disease by splenectomy. Reviews of haemolytic icterus by Tileston (1922), Campbell (1926) and Dameshek & Schwartz (1940) credit Micheli in 1911 with the first published report of a successful splenectomy, in a patient with the inherited form of the disease. However, priority probably goes to Sir Spencer Wells who operated successfully on a patient in 1887, subsequently reported by Dawson in 1914. The patient undoubtedly had familial haemolytic icterus, as two family members were affected, and the patient whose spleen was removed had an abnormal osmotic fragility test when studied by Dawson over 20 years after the operation. Elliot & Kanavel (1915) reported 47 splenectomized cases from the literature and one of their own. Of these, 16 were thought to be acquired cases, 23 were congenital and nine cases were unclassified. Only two patients died, one shortly after surgery, the other 6 weeks after surgery, of sepsis. The 46 surviving patients were relieved of their jaundice and presumed to be cured. Correction of the osmotic fragility test after surgery was the exception rather than the rule. Interestingly, their table of patients showed that only in acquired cases did the osmotic fragility correct after splenectomy, but the authors apparently failed to notice this correlation. Giffen (1917) reported nine curative splenectomies out of 12 attempted. Two patients died of operative complications and one patient with acquired disease initially responded to splenectomy, but relapsed after 2 years. In the last patient, the osmotic fragility test returned to normal during remission, in sharp contrast to the seven cured patients studied after splenectomy, whose osmotic fragility remained abnormal. Mayo (1917) further demonstrated the safety of the procedure, reporting one fatality in 19 operations. Thus, Tileston (1922) was able to recommended splenectomy for patients with congenital or familial haemolytic jaundice, if the symptoms warranted, and for primary cases of the acquired type. World War I brought a halt to investigation and case reports of haemolytic icterus. After a hiatus of over a decade, Lederer (1925, 1930) rediscovered the acquired syndrome and published accounts of six cases manifesting acute onset, rapid course, fever, acholuric jaundice, splenomegaly, severe anaemia and leucocytosis. Several of these cases exhibited a remarkable response to blood transfusion. The blood film exhibited reticulocytosis, anisocytosis, poikilocytosis and microcytes. Platelet counts were generally normal. Fragility studies and serology were not done. Brill (1926) described another similar case of his own in which the patient was near death until he received blood transfusions, after which he promptly recovered. He cites a clinically similar case reported by Macintosh and Cleland from Australia in which the patient died without transfusion. Brill thought his case and the Australian case resembled the recently reported cases of Moschkowitz, which the latter called 'acute pleichromic anaemia with hyaline thrombosis of terminal arterioles and capillaries', and which we now call thrombotic thrombocytopenic purpura (TTP). Indeed, Moschkowitz thought his cases resembled those of Lederer, although Lederer (1930) disagreed. Lederer was probably right; the constellation of acute haemolysis, fever and remarkable response to blood transfusion could cause one to consider TTP, but the normal platelet counts in his patients militate against the diagnosis. Strangely, neither Lederer nor Brill were aware of the contributions of the French and German investigators in the early part of the century. Lederer's 'discovery' was hailed as a new disease entity, in spite of the extensive review of haemolytic icterus by Tileston (1922), acknowledging the work of the European investigators just 3 years earlier. The cases of Lederer and Brill and other anecdotal reports were accepted as cases of acute haemolytic icterus and, for a time, the eponyms 'Lederer' or 'Lederer–Brill' were applied to these cases. Dyke and Young (1938) published a series of cases of 'macrocytic haemolytic anaemia' characterized by splenomegaly and haemolytic anaemia with increased indirect bilirubin, increased RBC fragility, reticulocytosis and macrocytosis. Microcytes or spherocytes were not prominent. The patients exhibited either an acute fulminant course, or a protracted relapsing and remitting course. Splenectomy was of doubtful benefit for the four patients in whom it was performed. They distinguished their cases from the congenital cases of Chauffard, in that the latter exhibited prominent microcytosis and a predictable response to splenectomy. Lederer's acute cases were thought to be a subtype of their own. Others applied the eponym 'Dyke–Young' to the chronic macrocytic cases to distinguish them from the acute transient cases of the 'Lederer–Brill' type. We now understand that they were probably variations of one and the same disease. Dameshek and Schwartz (1940) published a remarkable review of acquired haemolytic icterus. Ninety-six pages in length with 380 references from the world literature, it is the most extensive review of the subject extant, with the exception of whole books devoted to the topic. They identified 81 articles that described cases fitting their concept of acute (acquired) haemolytic icterus. Based on their own clinical observations of haemolysins in some patients, cases reported in the literature, including those of Chauffard and co-workers, and their own experiments involving injection of varying amounts of haemolytic serum into guinea pigs, they proposed that all cases of haemolytic icterus were as a result of haemolysins. The differences in clinical manifestations, ranging from mild congenital cases to fulminant acute haemoglobinurias, were accounted for by the dose of haemolysin. Figure 7 from their review illustrates their hypothesis. The relationship between concentration of haemolysin and the various clinical syndromes of haemolysis, as conceived by Dameshek and Schwartz (1940). These investigators hypothesized that differences between apparent familial cases and acquired cases of haemolytic anaemia lay primarily in the blood concentration of haemolysin. [Used with permission. Dameshek, W. and Schwartz, S. O., Acute Hemolytic Anemia (Acquired Hemolytic Icterus, Acute Type), Medicine, volume 19, pp 231–327, copyright Lippincott, Williams and Wilkins, 1940.] Thus, by the end of the 1930s, a clinical syndrome called haemolytic icterus or acholuric jaundice was well recognized. A number of eponyms had been applied to one or another variation of the clinical syndrome, but none gained widespread use. The congenital and frequently familial form was generally accepted. A presumably acquired form was well described, but not so well accepted, despite recent evidence to the contrary. Both types of cases held several clinical and haematological features in common. Both were characterized by clinical icterus and hyperbilirubinaemia without bilirubinuria. The spleen was usually palpable. Anaemia varied from mild to severe. Reticulocytosis and spherocytosis were present and the red cells exhibited increased fragility in hypotonic saline. Distinguishing features of the congenital and acquired forms lay in the presence or absence of family history, age of onset (younger age more probably congenital) and tempo of the disease, more acute types thought to be acquired. Serological studies for haemolysins and agglutinins were almost never reported. Splenectomy was consistently beneficial in the congenital and familial cases, but of inconsistent benefit in cases thought to be acquired. However, exceptions to all these distinctions were well described. What was sorely needed to distinguish congenital and acquired haemolytic icterus, if the latter indeed existed, was an understanding of aetiology and pathogenesis; it was soon to come. It was the War and the anticipated need for blood transfusions for large numbers of civilian and military casualties that brought together in Great Britain the talented and imaginative investigators who finally defined congenital and acquired haemolytic icterus. The distinctions between these two disorders are based on studies of RBC survival and on the anti-globulin test. Studies of RBC survival The first means of studying RBC survival was developed by Winifred Ashby (1919), a predoctoral fellow at the Mayo clinic. At that time, it was known that the RBCs of group I (now group AB) were agglutinated by serum from all other groups and Group I serum had no agglutinating properties for the cells of other groups. RBCs from groups II and III (groups A and B) were mutually agglutinated by each other's sera. Group IV (group O) serum agglutinated the RBCs of the other groups, but Group IV RBCs were not agglutinable by those sera. As it was possible to transfuse patients with RBCs not of their own group, Ashby reasoned it would be possible to measure the lifespan of transfused RBCs by periodically taking specimens of a patient's blood after transfusion, quantitatively agglutinating the patient's cells from the specimen and then counting the remaining cells which were the transfused cells. She applied her method in several subjects and found that the lifespan of transfused RBCs in the circulation extended for 30 d or more. Dacie & Mollison (1943) first applied Ashby's technique in patients with haemolytic anaemia over 20 years after her publication. They were able to show that normal RBCs transfused into patients with familial haemolytic anaemia survived normally, for approximately 100–120 d. The survival curves from their paper are shown in Fig 8. In sharp contrast, Loutit & Mollison (1946) noted that normal RBCs transfused into patients with acquired haemolytic anaemia exhibited markedly reduced survival. The RBC survival curve in one such patient is shown in Fig 9. As shown, following splenectomy, RBC survival improved. Dacie and Mollison, using the Ashby technique, were the first to demonstrate that normal RBCs survive normally in patients with familial haemolytic anaemia. The figure shows survival of blood from normal donors after transfusion to six patients with familial haemolytic anaemia. Case 3 was an Rh-negative patient who was later found to have developed an alloantibody to Rh (Loutit & Mollison, 1946), accounting for the shortened survival of transfused Rh-positive RBC. Although not shown in the figure, survival in cases 2 and 5 was followed to completion and found to exceed 100 days in each case. The dotted lines indicate the limits of survival in a group of normal recipients (Mollison, unpublished observations). [Figure and legend used with permission. Dacie, J. V. & Mollison, P. L., Survival of normal erythrocytes after transfusion to patients with familial haemolytic anaemia (acholuric jaundice). The Lancet, volume i, May 1, 1943, pp 550–552, copyright by The Lancet Ltd, 1943]. Survival of transfused normal RBCs in case 5 of acquired acholuric jaundice. Dotted line, survival graph before remission. Continuous line, survival graph after remission, following splenectomy. Figure and legend reproduced with permission. Loutit, J. F. & Mollison, P.L., Haemolytic icterus (acholuric jaundice), congenital and acquired. Journal of Pathology and Bacteriology, volume 58, pp 711–728, copyright by John Wiley & Sons 1946. Loutit & Mollison (1946) also transfused RBCs from patients with congenital and acquired haemolytic icterus into normal recipients and followed their survival. The RBCs from patients with congenital acholuric jaundice, including those from a patient who had undergone splenectomy, exhibited short survival. Cells from four patients with acquired disease exhibited normal survival. The RBC survival curves from these patients are shown in Fig 10. It is interesting to note the initial rapid destruction of RBCs followed by a normalization of the slope of the survival curve in one of the patients with acquired disease. As we now know, in acquired haemolytic anaemia, the red cells are coated with antibody. The initial rapid des