Scand J Haematol(l982) 28,169-174

Iron Overload in a Non-Transfused Patient with Thalassaemia Intermedia

ANTONIO CELADA

Department of Medicine, University Hospital of Geneva, Geneva, Switzerland

A patient with thalassaemia intermedia and haemosiderosis is reported. This patient did not receive transfusions or iron therapy. The iron absorption and the plasma iron turnover (PIT) were increased. Transfusions were carried out in order to decrease the amount of abnormal erythropoiesis. After that, the erythropoietin and PIT were decreased to normal levels and the iron absorption also returned to normal. The data presented suggest that increased erythropoiesis was responsible for the abnormaly high iron absorption and subsequently for the haemosiderosis of the patient presented.

Key words: erythropoiesis - iron absorption - iron overload - thalassaemia

Accepted for publication September 28, 1981

Correspondence to: A. Celada, Division of Hematology, RM-10, Department of Medicine, School of Medicine, University of Washington, Seattle, Washington 98195, USA

In patients with thalassaemia intermedia the marked genetic heterogeneity causes a variety of genetic combinations which pro- duce the different clinical pictures. The Hb concentration can vary considerably in asymptomatic carriers from normal to low values. However, the levels of Hb are con- siderably higher than those found in homo- zygous @-thalassaemia and these patients rarely need regular blood transfusions (Weatherall & Clegg 1972).

Recently, an increase in iron deposits in patients with thalassaemia intermedia has been reported, as well as an increase in iron absorption (Pippard et a1 1979). In some reports, patients with thalassaemia inter- media and haemosiderosis have been de-

scribed (Bannerman et al 1967, Bhamara- pravati et a1 1967). The reason for iron overload in these patients has been stated to be increased iron absorption.

In this study is presented a patient with thalassaemia intermedia and iron overload, who had not had any transfusion, iron treatment or alcohol intake. The iron ab- sorption was studied in relation to erythro- poiesis before and after transfusion the- rapy *

METHODS I

Iron absorption The whole-body counter previously described (Celada et a1 1978a) was used to measure iron absorption. The

003~553X/82/010169-06 $02.50/0 0 1982 Munksgaard, Copenhagen

170 ANTONIO

methodology has been described elswhere (Celada et a1 1978b). In brief, the inorganic iron test dose con- sisted of 10 pmol 59Fe as a ferrous citrate with an approximate activity of 1 pCi + 100 p o l / l (+) ascor- bate in 0.001 mol/l HCI. A test dose was given in the morning after a 12-h fast and after measurement of

CELADA

background radioactivity. 4 h after ingestion of radio- active iron the 100% value of total body radioactivity was measured. The third determination of total body radioactivity was carried out 14 d later. The following formula was used to calculate iron absorption:

cpm day 14 - cpm B day 1 cpm day 1 - cpm B day 11

Absorption (%) = x 100 x F.

F = correction factor decay for 59Fe by a comparison with a standard, B = background.

Ferro kinetics 1 ml of transferrin-bound 55Fe. PIT was calculated Iron absorption and plasma iron turnover (PTI) were according to Finch et a1 (1970) using the plasma iron, measured on the same day, similarly as previously radio-iron disappearance rate, and the haematocrit described for animals (Celada et a1 1979). After an according to the following formula: overnight fast, PTI was determined by i.v. injection of

Plasma iron (pg/dl plasma) (100 - PCV x 0.9) PIT (mg/dl whole blood/d) = X

T% (min) 100

The erythron iron turnover (EIT) was calculated as described by Finch et a1 (1970) from the following formula:

EIT (mg/dl whole blood/d) = PIT - plasma iron x plasmatocrit x 0.003 5 .

The 55 Fe content was determined by liquid scintillation counting, using the method of Eakins & Brown (1966).

Haematological and serological determinations Erythrocyte count, haematocrit (PCV), Hb and globu- lar indices were done with a laser ray counter (Hemac 630 L, Ortho Diagnostics Instruments, New Jersey). The spectrophotometric method using bathopenantro- line as reagent, suggested by the International Commit- tee for Standardization in Haematology (1971) was used for measuring serum iron. For the total iron- binding capacity the method of Ramsey (1957) was used. Urine iron content was measured colorimetrically using bathophenantroline sulfonate. Reticulocytes were counted in 2000 red cells stained with methylene blue and counterstained with Wright’s stain. Serum folate, vitamin B-12 levels and ferritin were measured using a commercial radioassay (Simultrac Radioassay Kit, Schwarz/Mann, New York and Ramco Laboratories, Houston, Texas, respectively). Erythropoietin was measured by a haemagglutination inhibition assay (J. L. C . Clinical Research Corporation, Knoxville, Tenessee, USA).

CASE REPORT

The patient (EM) was a male of Italian origin, born in 1936. In the past he was splenectomized at the age of 7 because of ‘anaemia’. In 1972, the patient was hospita- lized in the University Hospital of Geneva due to vesicular lithiasis but surgery was not performed. The patient did not drink alcohol regularly and his alimen- tary habits appeared normal. During this hospitaliza- tion the diagnosis of thalassaemia intermedia was made. At this time, this Hb was 77 g/l, MCV was 68 fl. In the blood smear, the erythrocytes were microcytic, hypochromic and, some with a basophilic stippling. Also, Howell-Jolly bodies and target cells were ob- served. The number of erythroblasts per 100 leuco- cytes was 115. Serum iron was 26.85 pmol/l and trans- ferrin saturation with iron was 86%. Liver function tests were within normal limits. Erythrocyte Hb showed Hb-F 18% (normal values 0 .25475%) and Hb-A2 4.6% (1-3%). Bone marrow aspiration showed a hyperplasia of erythroblasts and increased iron in RES. At this time his liver extended 5 cm. Haemato- logical studies of the parents showed a microcytosis in

THALASSAEMIA INTERMEDIA AND HAEMOSIDEROSIS 171

Figure 1. X-ray photo of humerus showing diminution of bone density and medullary cavity expansion.

both but no anaemia; S-iron and transferrin saturation were within normal limits. Hb of the patient’s father showed Hb-F 0.52% and Hb-A2 4.2% and his mother Hb-F 0.94% and Hb-A2 4.3%. In both, the diagnosis of thalassaemia minor was made. Hb of the patient fluctuated between 46 and 81 g/l, without any trans- fusions. The patient was periodically treated with folic acid and vitamin B-12.

In Jan 1980 the patient was hospitalized for respira- tory infection with a congestive heart failure which responded to red cell transfusions. At that time, the patient showed an excessive skin pigmentation of gray colour, his liver extended 11 cm and he showed a bilateral oedema in the lower limbs. Hb was 32 g/l ; there were 563 erythroblasts per 100 leucocytes. Ra- diography showed a severe cardiomegaly. Skeletal X- rays showed typical expansion of the medullary cavity throughout the skeleton as well as a diminution of bone density (Figure 1). The computed tomography showed an increased liver density, from 70-95 U.H. in the 20 measures which had been performed (Figure 2). Glucose, electrolytes, total protein, BUN, creatinine, calcium and phosphorus were within normal limits. Total bilirubin was 26 mg/l, y-glutamintransferase 70 U/1, alkaline phosphatase 57 U/1, aspartate amino- transferase 34 U/1 and alanine amino-transferase 20 U/1. Glucose tolerance curve was normal. Serum iron was 22.3 pmol/l and total iron-binding capacity of 24.6 pmol/l, S-folate 12.8 pg/l, S-vitamin B-12 783 ng/l and S-feritin 5 823 kg/1. A decreased fragility of erythrocytes was observed. The prothrombin time was 25%, fibrinogen 1.2 g/l, the partial thrombo-

Figure 2. Computed tomog showing increased size and of the liver.

:rap hY density

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172 ANTONIO CELADA

plastin time 37 s (normal 25-30) and platelet count 161 x lO9/l. After injection 500 rng desferrioxamine the urinary iron excretion in 24 h was 22 rng.

RESULTS

Two months after the last hospitalization, ferrokinetics and iron absorption were mea- sured when the levels of Hb were lowered (Table 1). At that time, iron absorption was dramatically increased over normal values. Also, the PIT and EIT were markedly eleva- ted suggesting an increase of erythropoiesis. 2 weeks after the beginning of this study, when the measurement of iron ab- sorption was completed, the patient had been transfused to the normal levels of Hb. 5 d later, a second measurement of ferroki- netics and iron absorption was performed (Tablel). Iron absorption as well as PIT and EIT were decreased to the normal limits. These changes accompanied a de- crease of erythropoietin. The numbers of reticulocytes, erythroblasts, bilirubin and the percentage of Hb-F were decreased but

not to normal limits, probably in relation to the dilution by the red blood cells introdu- ced by the transfusions. Finally, plasma iron, transferrin saturtion and ferritin were unchanged as well as the levels of S-folate and S-vitamin B-12.

DISCUSSION

The diagnosis of thalassaemia intermedia was made on the basis of clinical and haema- tological data, Hb electrophoresis and the family study. Both parents had thalas- saemia minor without clinical expression. The levels of ferritin and transferrin satura- tion showed no iron loading.

In the last hospitalization, the patient showed a typical picture of haemosiderosis with an excessive skin pigmentation, hepa- tomegaly and cardiomegaly. The haemosi- derosis was confirmed by S-ferritin, trans- ferrin saturation, S-iron and the sideruria after desferrioxamine injection. The in- crease in density in liver tissue by computed tomography suggested an increase in iron

TABLE 1 Effect of red cell transfusions on erythropoiesis and iron absorption

Before After Normal transfusions transfusions limits

Haemoglobin (g/l) Reticulocytes (%) Erythroblasts (/lo0 leucocyte Haemoglobin F (%) Haemoglobin A2 (%) Bilirubin (mg/l) Erythropoietin (MIV/ml) Plasma iron (pmol/l) Transferrin saturation (%) Ferritin (pg/l) PIT (mg/d) whole blood/d) EIT (mg/dl whole blood/d) Iron absorption (%) S-folate (pg/l) S-vitamin B-12 (ng/l)

62 15.7

: S ) 115 18.0 4.6 21.5 450.0 23.52

96 5 320 3.25 2.82 95.0 14.0 535

141 5 36 3.2 3.6 12.6 75.0 22.48

98 4 840 0.93 0.83 20.0 18.0 460

155k17 0.920.3

0- 1 0.25-0.75

1.0-3.0 5.3f2.6

55.76f33.91 18.64f5.85

32k6.6 126f89

0.77 f 0.50 0.56k0.36 24.30k9.46 11.34f5.21 470f155

THALASSAEMIA INTERMEDIA AND HAEMOSIDEROSIS 173

deposits (Mills et a1 1977, Houang et a1 1979). The abnormal level of S-enzymes and the altered coagulation tests showed an impairment of the liver that was probably secondary to iron toxicity. It is impossible to know if the cardiomegaly was secondary to anaemia or to the iron deposit in the myocardium. In patients with thalassaemia major, it has been suggested that the erythropoietic activity can be responsible for the decreased bone mass (Pootrakul et a1 1981). On the other hand, in patients with iron overload, osteoporosis has been de- scribed related to a depletion in tissue ascorbic acid and operating through its oxidative destruction by iron deposits (Charlton et a1 1973). Cardiomegaly and bone lesions may be due to the effect of the thalassaemia and iron overload together.

The origin of haemosiderosis in the pa- tient presented is probably related to an increased iron absorption. The patient had not received transfusions or iron therapy. In basal conditions, when the Hb was 62 g/l, the iron absorption was greatly in- creased; similarly, erythropoiesis was in- creased. In thalassaemia intermedia there was an increased erythropoiesis inversely correlated with the degree of anaemia (Pearson et a1 1960, Malamos et a1 1961, Gallo et a1 1975, Rotoli 1976, Cazzola et a1 1979). After transfusions, in order to re- duce erythropoietic stimulation of the erythroid marrow, the PIT of patients with thalassaemia major decreased to nearly normal limits (Cavill et a1 1978, Pootrakul et a1 1981). Also, the iron absorption in these patients decreased after transfusion therapy (Erlandson et a1 1962, Heinrich et a1 1973, Necheles et a1 1974, Wyss et a1 1977). In our patient, both erythropoiesis and iron ab- sorption decreased to normal levels after red blood cell transfusions when the Hb was 14

g/l. Correspondingly erythropoietin was de- creased. This experiment suggests that in thalassaemia intermedia patients, iron ab- sorption increases secondarily to the active erythropoiesis which is responsible for the iron overload.

ACKNOWLEDGEMENT

The constructive criticism of Professor Finch was highly appreciated.

Clement A.

REFERENCES

Bannerman R, Keusch G, Kreimer-Bimbaum M, Vance V K & Vaughan S (1967) Thalassemia inter- media with iron overload, cardiac failure, diabetes mellitus, hypopituitarism and porphyrinuria. Am J Med 42,476-86.

Bhamarapravati N, Na-Nakom S, Wasi P & Tuchinda S (1967) Pathology of abnormal hemoglobin diseases seen in Thailand. I. Pathology of beta-thalassemia hemoglobin E disease. Am J Clin Path01 47,745-58.

Cavill I, Richetts C, Jacobs A & Letsky E (1978) Erythropoiesis and the effect of transfusion in homo- zygous B-thalassemia. N Engl J Med 298,776-78.

Cazzola M, Alessandrino P, Barosi G, Morandi S & Stafanelli M (1979) Quantitative evaluation of the mechanisms of the anemia in heterozygous B-thalas- semia. Scand J Haematol23,107-14.

Celada A, Rudolf H & Donath A (1978b) Effect of a single ingestion of alcohol on iron absorption. Am J Hematol5,225-37.

Celada A, Rudolf H & Donath A (1979) Effect of experimental chronic alcohol ingestion and folic acid deficiency on iron absorption. Blood 54,96&15.

Celada A, Rudolf H, Herreros V & Donath A (1978a) Inorganic iron absorption in subjects with iron defi- ciency anemia, achylia gastrica and alcoholic cirrhosis using a whole-body counter. Acta Haematol (Basel) 60, 182-92.

Charlton R W, Bothwell T H & Seftel H C (1973) Dietary iron overload. Clin Haematol2,383-403.

Eakins J D & Brown D A (1966) An improved method for the simultaneous determinations of iron-55 and iron-59 in blood by liquid scintillation countings. Int J Appl Radiat Isot 17, 391-97.

174 ANTONIO CELADA

Erlandson M E, Walden B, Stern G, Hilgartner M W, Wehman J & Smith C H (1962) Studies on congeni- tal hemolytic syndromes. IV. Gastrointestinal ab- sorption of iron. Blood 19, 359-78.

Finch C A, Deubelbeiss K, Cook J D , Eschback J W, Harker L A, Funk D D, Marsaglia G , Hillman R S, Slichter S, Adamson J W, Ganzoni A & Giblett E R (1970) Ferrokinetics in man. Medicine 49, 17-53.

Gallo E, Pich P, Ricco G , Saglio G, Camaschella C & Mazz U (1975) The relationship between anemia, fecal stercobilinogen, erythrocyte survival and glo- bin synthesis in heterozygotes for f3-thalassemia. Blood 46,69%98.

Heinrich H C, Gabbe E E, Oppitz K H, Whang D H, Bender-Gotze C,Schafer K H, Schroter W & Pfau A A (1973)Absorption of inorganic and food iron in children with homozygous and heterozygous B-thalassemia. Z Kinderheilkd 115, 1-22.

Houang M T W, Arozena X, Skalicka A, Huehns E R & Shaw D G (1979) Correlations between computed tomographic values and liver iron content in thalas- semia major iron overload. Lancet i, 1322-23.

International Committee for Standardization in Haematology. (1971). Proposed recommendations for measurement of serum iron in human blood. Br J Haernatol20, 451-53.

Malamos B, Belcher E H, Gyftaki E & Binopoulos D (1961) Simultaneous radioactive tracer studies of erythropoiesis and red cell destruction in thalas- semia. Br J Haematol7,411-29.

Mills S R, Doppman J L & Nienhuis A W (1977) Computed tomography in the diagnosis of disor- ders of excessive iron storage of the liver. J Comput Assit Tomogr 1, 101-04.

Necheles T F, Chung G, Sabbali R & Shitten D (1974) Intensive transfusion therapy in thalassemia major: an eight year follow-up. Ann N Y Acad Sci 232, 179-85.

Pearson H A, McFarland W & King E R (1960) Erythrokinetics studies in thalassemia trait. J Lab Clin Med 56,866-73.

Pippard M J, Callender S T, Warner G T & Weather all D J (1979) Iron absorption and loading in B-thalassaemia intermedia. Lancet ii, 819-21.

Pootrakul P, Hungsprenges S, Fucharoen S, Baylink D, Thompson E, English E, Lee M, Burnell J & Finch C A (1981) Relation between erythropoiesis and bone matabolism in thalassemia. N Engl J Med 304, 1470-73.

Ramsey V N M (1957) The determination of the total iron binding capacity of serum. Clin Chim Acta 2

Rotoli B (1976) Thalassemia in Italy: treatment of Cooley disease and iron kinetics in heterozygotes. Birth Defects 12,5341.

Weatherall D J & Clegg J B (1972). The Thalus- saemia Syndromes, 2nd edn. Blackwell Scientific Publications, Oxford.

Wyss M, Celada A, Martin D & Jeannet M (1977) Multiple transfusions of HLA compatible blood in thalassemia major. Helv Paediat Acta 32, 451-59.

221-25.

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