L. Szur

The Pattern of Erythrocyte Destruction in Haemolytic Anaemia, as Studied with Radioactive Chromium

THE introduction of isotope techques and the availability of a suitable isotope label have greatly facilitated the study of the life-span and the sites of destruction of erythrocytes in haemolytic anaemia. Radioactive chromium (51Cr) has several advantages which make it especially suitable for this purpose : the patient’s erythrocytes can be labelled and re-introduced into his own circulation, and so studied in their natural environment; chromium liberated by erythrocyte destruction is not uthzed to relabel other erythrocytes, and the specific activity of the radloactive chromium is sufficiently high for the labelling to be attained with a small and apparently non-toxic amount of chromium. Furthermore, in addition to determining the intensity of haemolysis by measuring the rate of elimination of erythrocytes from the circulation, it is possible to learn somethmg of the nature of the haemolytic process by studying the pattern of elimination. The effect of treatment on the survival of a patient’s erythrocytes can also be studied and the sites of their destruction determined by counting in vim, with special reference to the role of the spleen and liver. Jandl, Greenberg, Yonemoto and Castle (1956), Hughes Jones and Szur (1957) and others have shown that the in vivo counting techque can be of value, too, in predicting the response to splenectomy. In the present paper we describe the results of survival and in vivo counting studies, using 51Cr,.[carried out on patients suffering from hereditary spherocytosis, hereditary elliptocytosis, hereditary non-spherocytic haemolytic anaemia, sickle-cell disease, paroxysmal nocturnal haemoglobinuria and auto-immune acquired haemolytic anaemia.

The Influence of Anaemia, Polycythaemia and Splenomegaly on the Relationship between Venous Haematocrit and Red-Cell Volume

A FAIRLY close relationship has been observed between venous haematocrit (packed cell volume, PCV) and the red-cell volume in normal and in anaemic subjects. However, this is not invariable, and the relationship may be upset by a number of factors. These include a disproportionate increase in plasma volume, as has been demonstrated in cirrhosis (Bateman, Slhorr and Elgvin, 1949), nephritis (Eisenberg, 1959) and pregnancy (Verel, Bury and Hope, 1956). Under these circumstances the PCV cannot be regarded as a reliable index of red-cell mass. Anaemia, polycythaemia and splenomegaly are also thought to influence the relationship of PCV and red-cell volume. The present study was undertaken in order to evaluate the effect of these factors on this relationship.

Red-cell production and destruction in myelosclerosis.

MYELOSCLEROSIS is a condition in which there is replacement of the bone marrow by fibrous tissue (myelofibrosis) or by cancellous bone (myelosclerosis). Almost invariably this is associated with splenomegaly, and hepatomegaly is also frequently found. Extramedullary haemopoiesis is a characteristic feature of the condition and is the main cause of enlargement of the liver and spleen. The peripheral blood shows a leucoerythroblastic picture, sometimes with considerable numbers of nucleated red cells. Reticulocytosis is common and there is marked variation in the shape of the red cells, teardrop poikilocytes being especially characteristic (Cook, Franklin, Hamilton and Fowler, 1953). Hueck (1879) is generally held to have been the first to report a case of myelosclerosis in the medical literature. Lack of agreement in the past on the definition, aetiology and pathogenesis of the condition makes a review of the disease difficult. Whilst nomenclature and criteria for diagnosis have been simplified in recent years, there is still considerable controversy over the aetiology and nature of the syndrome. These aspects have recently been reviewed by Leonard, Israels and Wilkinson (1957), who describe the clinical features and findings in 28 cases, and by Linman and Bethel1 (1957), who report on 56 cases of their own. The aetiology of myelosclerosis remains unknown. In most instances there is no apparent cause; it is said, however, to follow a variety of diseases and poisons which have been tabulated in detail by Erf and Herbut (1944). Wyatt and Sommers (1950) found similarities between myelosclerosis and cirrhosis of the liver and concluded that the condition might be due to toxic agents or deficiencies, causing injury and necrosis of the marrow, followed by reactive hyperplasia of surviving cells, fibrous overgrowth and ultimate progression to organ failure and death. In recent years idiopathic myelosclerosis has been considered to be closely linked to conditions like polycythaemia Vera and myeloid leukaemia (Vaughan and Harrison, 1939; Merskey, 1949; Dameshek, 1951), and thus to take its place amongst the myeloproliferative disorders. The cause and extent of the splenic, and the less frequent hepatic enlargement, and of the extramedullary haemopoiesis which develops in these and occasionally in other sites (such as the kidney, lymph nodes, etc.) are of great importance, both for the understanding of the pathogenesis of myelosclerosis and for its treatment. Broadly speaking there are two schools of thought on this subject. The older view was that the extramedullary blood formation is compensatory in nature, due to the fibrosis and the resultant inadequacy of the marrow (Donhauser, 1908); among the more recent supporters of this view are Leonard, IsraZls and Wilkinson (1957). The other view is that it is primary rather than compensatory in nature, of autochthonous origin, and, like the bone marrow hyperplasia, part of the generalized proliferative process (Rosenthal, 1950; Hutt, Pinniger and Wetherley-Mein, 1953 ; Wasser-

Elution Correction in 51Cr Red Cell Survival Studies

Summary. Red cell survival has been studied in 13 normal adults using simultaneous labelling with DF32P and 51Cr; elution correction factors have been determined for the51 Cr labelling in which ACD NIH formula A has been used as an anticoagulant with ICSH recommended method. The mean rate of 51Cr elution was 1.0% per day and there was no demonstrable early loss of either51 Cr or DF32P. The mean value for mean cell lifespan was found to be 111.2 days with an SD of 20.3 days.

Studies of splenic function in the myeloproliferative disorders and generalized malignant lymphomas.

The relative importance of splenic red‐cell pooling, sequestration and cell destruction in the causation of anaemia has been studied in 29 patients—16 with generalized lymphoproliferative disease, 12 with myeloproliferative disease and one with idiopathic autoimmune haemolytic anaemia.

SURFACE COUNTING IN THE ASSESSMENT OF SITES OF RED CELL DESTRUCTION

It is now well over a decade since in-viuo surface counting techniques, using red cells labelled with isotopes, were developed for the determination of sites of red-cell destruction, and it should be possible to make at least an interim assessment of their value and limitations. Although a number of isotopes have been investigated, Chromium-51, which emits a y ray of 0.32 meV and whose HVL in tissue is about 9 cm has, in spite of a number of drawbacks, been used almost exclusively. The determination of sites of red-cell destruction is based on the finding that the destruction ~f~~Crlabe l led red cells within an organ is associated with a measurable and progressive accumulation of the isotope in the organ. In this context ‘destruction’ needs to be clearly distinguished from ‘sequestration’ with which it is frequently confused. Sequestration refers to the retention of intact red cells within the organ for variable periods of time; whilst destruction of these cells often follows, either in that organ or elsewhere, this is not inevitable. Investigations based on these techniques have proved of considerable value in haematology. They have enlarged our theoretical understanding of the mechanism of haemolysis in various haemolytic disorders, and of the destruction of normal red cells. They have also been of value in the study of the removal of incompatible cells from the circulation and of the sequestration or lysis of cells altered by immune mechanisms, by exposure to various metals, or by other forms of damage such as heat and sulphydryl inhibitors (e.g. Hughes Jones et al, 1957; Jandl etal, 1957; Lewis etal, 1960a; Crome& Mollison, 1964). They have thus led to the development of more sophisticated techniques employed, for example, in splenic scanning. From a practical clinical point of view, information about the sites of red-cell destruction has proved of considerable value in the management of haemolytic anaemia and especially in decisions as to whether splenectomy should be undertaken. It cannot be stressed too strongly that surface-counting data must be considered in conjunction with all the clinical, haematological and biochemical findings in an individual patient. Nevertheless, the latter may be of little help in predicting the outcome of splenectomy. This was stressed by Chertkow & Dacie (1956) and Allgood & Chaplin (1967) in autoimmune haemolytic anaemia and again by Goldberg et a1 (1966), who reviewed the results of splenectomy in 182 cases collected from the literature.

Quantitative Measurement of Splenic and Hepatic Red‐Cell Destruction

A method has been developed by means of which independent measurement can be made of the amount of red‐cell destruction occurring in the spleen and the liver. The technique involves a standard red‐cell survival study and surface‐counting measurements together with quantitative scanning of the spleen and liver with 113mIn colloid in order to calibrate the surface counter. The rate of destruction in each organ is obtained by fitting the measured uptake curve for the organ to a theoretical uptake curve by computer. In addition, if whole body counting is also performed, the amount of red‐cell destruction occurring in the rest of the reticulo‐endothelial system may be deduced. Results are given for measurements on a series of 11 patients.

52Fe Studies of the Effects of Treatment on Erythropoiesis in Megaloblastic Anaemia

Summary. 52Fe has been used to study iron turnover and the extent of haemopoietic tissue in five patients (age range 13–74 yr) with severe megaloblastic anaemia, before and after treatment. All the untreated patients showed extension of haemopoietic marrow, and this was most marked in the youngest subjects. One patient (aged 60) showed a considerable degree of extramedullary haemopoiesis in the spleen. The extent of haemopoietic tissue decreased after therapy in all cases and appeared to reach normal by about 4–5 mth.

Studies of Splenic Function by Means of Radioisotope-Labelled Red Cells

Among John Dacie’s many contributions to haematology, one of the most important has undoubtedly been his work on the haemolytic anaemias. This encompassed almost all aspects of the subject including classification, aetiology, pathogenesis, prognosis and treatment. It was inevitable, therefore, that early in his studies he should have considered the role of the spleen in the mechanism of haemolysis and the value of splenectomy as a form of treatment. Thus, in 1943 he published a paper on ‘Familial haemolytic anaemia (acholuric jaundice), with particular reference to changes in fragility produced by splenectomy’, which contained many important insights about the role of the spleen in hereditary spherocytosis. Other noteworthy papers dealt with the results of splenectomy in autoimmune haemolytic anaemia (Chertkow & Dacie, 1956; Dacie, 1963) and his four-volume book on the haemolytic anaemias contains many sections on the spleen. When radioactive isotopes became available for the study of red-cell life span and sites of red-cell destruction, Dacie encouraged developments in t h i s field, participated in, and suggested a number of experiments (e.g. Lewis et al, 1960a, b). Many of the studies reported in this paper owe a great deal to discussion which the authors have had with him and the encouragement which he has given them. The spleen has always been something of an organ of mystery in the history of medicine. Since Pliny’s doubtful claim that physicians in antiquity removed the spleen of athletes in order to improve their ‘wind’ (see Crosby, 1963), many fanciful and sometimes amusing ideas have been propounded about the role and function of this organ. Although considerable gaps remain in our understanding of splenic function and of the mechanisms by which many of the observed phenomena take place, a great deal has been learned in recent years. Undoubtedly isotope labelling techniques have played a major part in the formulation ofpresentday concepts, For a variety of reasons these concepts are based primarily on investigations involving the red cells, although some very interesting and important investigations have been carried out using labelled platelets or white cells. The in vivo isotope studies with labelled red cells can usefully be divided into four groups: I. Studies of red-cell production and, to a lesser extent, of destruction and sequestration, with the use of the radioisotopes of iron. It. Visualization of the spleen by the use of labelled red cells damaged in various ways, so as to ensure their selective uptake by the spleen. This is frequently combined with measurement of the rate of clearance of these damaged cells from the circulation.