Background
The transfusion dependent anaemias cause a substantial burden of morbidity and mortality, of which thalassaemia is the commonest single gene disorder worldwide [
1]. The transfusions that are necessary for survival cause tissue iron loading, and this ultimately results in heart failure as the major cause of death [
2,
3]. Histopathological studies of patients with cardiac siderosis have indicated that replacement myocardial fibrosis is prominent and causative of the heart failure [
4,
5], but these data were based on patients who died 50 years ago or more and prior to the modern era of iron chelation treatment. These historical data have now become controversial because it is now recognised that the risk of developing heart failure in beta-thalassaemia major is closely associated with myocardial siderosis [
6], heart failure may be fully reversed with aggressive iron-chelation treatment [
7], and improvement in ventricular function occurs in concert with myocardial iron reduction implying a causal relationship [
8‐
10]. In other non-transfusional cardiomyopathies such as hypertrophic cardiomyopathy [
11], dilated cardiomyopathy [
12], and arrhythmogenic right/left ventricular cardiomyopathy [
13,
14], macroscopic replacement cardiac fibrosis is a prominent feature and is associated with deteriorating cardiac function and an adverse outcome [
15‐
18]. However, improvement in left ventricular function with treatment is limited or not possible in these cardiomyopathies, which may relate to the underlying myopathy, but also because the replacement fibrosis appears to be permanent. Therefore, we hypothesized that the modern clinical finding of reversible ventricular dysfunction in myocardial siderosis is not driven by replacement fibrosis. To investigate this, we analysed hearts from transfusion dependent patients to examine for cardiac fibrosis by histology and relate the findings to iron overload cardiomyopathy.
Discussion
This study failed to show replacement cardiac fibrosis as a cause of death or transplantation related to heart failure in patients with transfusion dependent anaemia, including most importantly beta-thalassaemia major patients. These findings are in marked contrast to the major historical pathological series in transfusion dependent patients. In 1964 [
4], post-mortem data published by Engle on 41 chronic anemia patients (39 with thalassemia major, 2 with aplastic anemia), showed 26 (63%) had heart failure with a further 6 (15%) having cardiac enlargement and an abnormal ECG. These data clearly indicated that the heart was the target lethal organ in beta-thalassaemia major. Engle noted that focal myocyte degeneration and fibrosis was extensive, and was an “adequate explanation for the progressive cardiac enlargement and heart failure”. In 1971 [
5], Buja reported post-mortem findings on 19 patients with cardiac iron deposits (8 with aplastic anemia, 4 with chronic myelocytic leukemia, 3 with idiopathic hemochromatosis, 2 with chronic lymphocytic leukemia, 1 with acute lymphocytic leukemia, 1 with sickle cell anemia). Heart failure occurred in 14 (67%) patients. The left ventricle was affected by extensive myocardial replacement fibrosis in 6 (29%) patients and by focal interstitial fibrosis in 4 (21%) patients. The papillary muscles were affected by focal interstitial fibrosis in a further 6 (29%) patients. In only 3 patients (14%), was the heart not affected by fibrosis. Buja listed a further 21 reports dating from 1933 to 1967 as showing myocardial fibrosis in patients with cardiac iron loading [
5].
The cause of cardiac fibrosis in iron overload conditions is not fully explained. The most important effect appears to be that myocytes can suppress proliferation of cardiac fibroblasts by cumulative effects on late G1 events leading to DNA synthesis, and these effects are diminished with myocyte iron accumulation, which encourages cardiac fibrosis [
34,
35]. However, a less supporting study of mRNA in ex-vivo cardiac myocytes showed iron level dependent reductions in expression of transforming growth factor-β1 (TGF-B1), biglycan, and collagen type I, which was accompanied by a reduction in TGF-B1 bioactivity, which does not obviously support iron-driven cardiac fibrogenesis [
36]. Animal models support a link between cardiac iron and cardiac fibrosis. Cardiac fibrosis was prominent in double knock-out mice for beta 2 microglobulin (B2m, deficiency of which causes increased gut uptake of iron through impairment of the HFE-B2m complex) and recombinase activator gene 1 (Rag1, deficiency of which causes absence of B and T lymphocytes) which was not seen in B2m and Rag1 single-knockout mice or control mice of the same age, implying that lymphocytes play a role in cardiac fibrosis which is additive to cardiac iron loading alone [
37]. Other iron loading animal models also show cardiac fibrosis, although this was not prominent [
38,
39].
The explanation for the apparent change in prevalence of replacement cardiac fibrosis over 40 years can only be subject to speculation. One obvious possibility is the introduction of the iron chelator deferoxamine, which came into widespread clinical use in the 1970’s. Although not explicitly stated, the patients in Engle and Buja’s papers would not have received such treatment based on the period of the patient post-mortems (1950–1963, and 1953–1969 respectively). This suggests that deferoxamine might impair the development cardiac fibrosis that occurs with myocardial siderosis. This is plausible as deferoxamine is known to stabilise liver fibrosis in association with reduced liver iron loading [
40]. There is also experimental evidence which supports this position. Control of cardiac iron may in itself prevent cardiac fibrosis by suppressing fibroblast proliferation [
34,
35] but other direct supportive evidence for a protective effect of deferoxamine comes from studies of angiotensin II in normal and iron loaded rats in which the development of cardiac fibrosis could be prevented by deferoxamine [
41]. Since angiotensin II causes cardiac fibrosis and is increased in cardiac failure, this effect could be clinically significant. However, this explanation is not completely sufficient because patients 3 and 4 in our series, did not receive chelation but did not have significant myocardial fibrosis.
Although the use of deferoxamine seems the most likely factor distinguishing the historical from our modern cardiac findings, other possibilities exist. Another change in treatment of thalassemia major patients is the use of increased numbers of blood transfusions per year since the 1970’s, which suppresses ineffective erythropoiesis and bony abnormality. A typical modern regime for an adult includes the transfusion of up to 50 units of blood per year (0.4 mg/kg/day transfusional iron burden) [
42]. Engle documented a transfusion rate on average of approximately 13.4 units/year in 26 patients who developed heart failure [
4], consistent with a >3 fold increase in transfusion rate for modern patients. Cardiac fibrosis could therefore have been stimulated by anemia related myocardial hypoxia in historical patient cohorts, particularly in the setting of left ventricular hypertrophy and dilatation [
43]. Increased transfusions with improved tissue oxygenation might have played a role therefore in reducing myocardial fibrosis. The change in cardiac fibrosis might also be related to an apparent reduction in recurrent pericarditis seen in the last 40 years. Engle reported 19 of 41 (46%) patients had 33 recognised episodes of pericarditis [
4]. Pericarditis was not a focus of Buja’s paper [
5]. Pericarditis is recognized in thalassemia major in the modern era, but at a far lower incidence of <5% [
44]. It is likely that pericarditis is now less frequent because of the widespread use of iron chelation therapy, but the genesis of the pericarditis in iron overload is not well understood and other factors might be important. Myocarditis is also recognised as a cause of myocardial fibrosis [
45], and has been documented from a modern series as occurring in 4.5% of beta-thalassemia major patients [
46]. Myocarditis was not documented in the historical series and direct comparisons with the modern findings are therefore not possible. However, the fact that the detailed historical papers did not report myocardial inflammation, would not suggest that it was prominent or common, and therefore a reduction in prevalence seems unlikely. It should be noted that in the modern environment, approximately 2% of cases of heart failure in thalassemia major have low levels of cardiac iron [
47], and these are thought to be caused by myocarditis [
48], in which cardiac fibrosis may play a significant role [
45], which is in addition to the myocardial infection and inflammation. Another possibility is that a historical factor in blood transfusion practice was associated with cardiac fibrosis in the past that has now decreased. The reduced transmission of cardiotrophic viruses might be a possibility, of which one candidate virus is hepatitis C, which has been implicated in myocarditis and cardiac fibrosis [
49‐
51]. Reports of infrequent cardiac fibrosis in beta-thalassemia major using the non-invasive in-vivo technique of late gadolinium enhancement (LGE) CMR showed infrequent minor fibrosis (24%, 2% and 15.8% of patients) in patients without heart failure [
52‐
54], but the extent of fibrosis was limited (3.9%, 0.4%, 1.3%) [
52‐
54]. In comparison to other disease settings it is not clear whether this minor fibrosis would be sufficient to cause significant LV dysfunction, where on average each 1% of infarcted myocardium assessed by LGE leads to only a modest 0.67% reduction in ejection fraction [
55]. However, the difference in prevalence of minor cardiac fibrosis between centers might be explained by the different prevalence of hepatitis C infection [
52,
53]. Despite this possibility however, extensive cardiac fibrosis causing heart failure is not seen in hepatitis C infection, and it is unlikely that changes in transfusional infections can explain the change over time in replacement cardiac fibrosis.
Further possible factors that might influence the difference in cardiac fibrosis over time are: 1) the occurrence of diabetes in thalassaemia major, which has association with the presence of cardiac fibrosis and development of heart failure [
56]. However, seven of our 10 patients had confirmed diabetes without significant cardiac fibrosis. 2) the use of inhibitors of the renin-angiotensin-aldosterone system such as angiotensin converting enzyme inhibitors, angiotensin receptor blockers (ARB) and mineralocorticoid receptor antagonists (MRA) which are known potent inhibitors of myocardial fibrosis [
57‐
60]. Only 2 of our patients were recorded as being on such treatment, although due to the retrospective nature of data collection in this series, the accuracy of drug treatments may not be ideal. 3) Finally, in older transfusion dependent patients, the pattern of heart failure and the relationship to cardiac fibrosis may differ in comparison with younger patients, with development of heart failure with preserved ejection fraction [
61]. This cardiac pathology is not completely understood but restrictive physiology is involved.
Limitations
The absence of in-vivo CMR does not allow the comparison of late gadolinium enhancement with the histological findings. The technique of T1 mapping was likewise not possible in-vivo in this population [
62]. This is inevitable given the international source of the post-mortem hearts. This was a small study in comparison with historical reports, which reflects the general trend towards reduction of patients undergoing post-mortem.
Acknowledgements
This project was supported by the NIHR Cardiovascular Biomedical Research Unit of Royal Brompton and Harefield NHS Foundation Trust and Imperial College, London. Funding was also received from National Institutes of Health Grant Award 5 R01 DK066084-02, and the British Heart Foundation. JMW acknowledges support from Department of Health’s NIHR Biomedical Research Centres funding scheme at UCLH.