N-terminal-pro-b-type natriuretic peptide levels and cardiac hemosiderosis in adolescent β-thalassemia major patients.

BACKGROUND: Iron-induced cardiomyopathy remains the leading cause of mortality in patients with β-thalassemia major. Iron overload cardiomyopathy, which may be reversible through iron chelation, is characterized by early diastolic dysfunction. Amino-terminal pro-brain natriuretic peptide (NT-proBNP) is a sensitive biomarker of diastolic dysfunction. AIM: The aim of the study is to evaluate the diagnostic value of NT-proBNP as a surrogate marker of iron overload examined with magnetic resonance imaging T2-star (MRI T2*).
METHODS: Sixty-eight β-thalassemia major patients (10-18 years) with no signs of heart failure underwent NT-proBNP measurement before routine transfusion. All participants prospectively underwent cardiac MRI T2* examination within 3 months (median 19 days). Patients were divided as cardiac hemosiderosis (cardiac MRI T2* <20 ms) and nonhemosiderosis (cardiac MRI T2* >20 ms).
RESULTS: Of 68 patients, the male-to-female ratio was 1:1.1 and the median age was 14.1 years (range: 10-17.8 years). NT-proBNP levels were not different between hemosiderosis and nonhemosiderosis patients (P = 0.233). Further receiver operating characteristic analysis resulted in no significant correlation of NT-proBNP and MRI T2* (area under the curve 0.393, P = 0.233).
CONCLUSION: Measurement of NT-proBNP levels cannot be used for early detection of cardiac iron overload in adolescent with β-thalassemia major.

INTRODUCTION

Thalassemia is the most common single-gene disorder in the world with 4.5% population of the world having carrier gene and 300.000–500.000 homozygotes born each year.[1] Chronic hemolysis in β-thalassemia major necessitates regular blood transfusion to maintain adequate tissue oxygenation and leads to increase the risk of iron overload.[2] Lifelong regular transfusion in combination with increase intestinal iron absorption will cause hemochromatosis in various organs.[23] Hemochromatosis alone or in combination with immunogenetic factors is the leading mechanism of heart failure development in β-thalassemia.[456] Although recent advances in iron-chelating agents, iron-related cardiomyopathy remains the leading cause of mortality and morbidity in thalassemia major patients, and once heart failure occurs, the prognosis is poor.[789] Therefore, accurate measurement of cardiac iron is important clinically as detecting early cardiac dysfunction, and adequate chelating agent will reverse the disease process.[101112]

Iron overload cardiomyopathy is characterized by early diastolic dysfunction which precedes systolic dysfunction.[13] Serum ferritin level is widely used in the evaluation of thalassemia patient's iron state yet it is not a reliable indicator of cardiac iron.[14] Cardiac magnetic resonance imaging T2-star (MRI T2*) is an easy and highly reproducible measurement technique which correlates better with cardiac iron concentration.[1516] However, it is relatively expensive and it is not widely available in developing countries. Doppler echocardiography and tissue Doppler imaging can detect early cardiac dysfunction, but their clinical use is limited due to high operator dependence and poor correlation with cardiac iron.[17]

Diastolic dysfunction in β-thalassemia major patients is mainly caused by cardiac iron overload.[13] Brain natriuretic peptide (BNP) and amino-terminal pro-BNP (NT-proBNP) are released after increased cardiac stress and volume overload.[18] The increase of NT-proBNP is detected early in the course of the disease and seems to be a reliable indicator for the early detection of cardiac hemosiderosis in adult β-thalassemia patients.[192021] However, the clinical use of this assay in children, particularly in adolescents, is limited.

The aim of this study was to explore the possible use of serum NT-proBNP levels in adolescents with β-thalassemia major to detect cardiac hemosiderosis as assessed by MRI T2*.

METHODS

Study population

Sixty-eight adolescent patients from Cipto Mangunkusumo Hospital and Tangerang District hospital, aged 10–18 years (36 males and 32 females) with β-thalassemia major, were enrolled in this study during June to November 2017. Inclusion criteria were asymptomatic β-thalassemia major patients with pretransfusion hemoglobin level above 7 g/dL. Patients with clinical sign and symptoms of heart failure and impaired renal and liver function were excluded from the study. All patients were receiving regular blood transfusion every 2–3 weeks and chelation therapy, which had been started before the age of 5 years. At the time of sample collection, 42 patients were receiving deferiprone (DFP) chelation therapy, 10 deferasirox (DFX), 6 combination therapy of DFP and DFX, 9 combination therapy of DFP and deferoxamine (DFO), and 1 combination therapy of DFO and DFX. The local ethics committee approved the study protocol, and written informed consent was obtained from all patients and parents.

Amino-terminal pro-brain natriuretic peptide measurements

All blood samples for NT-proBNP measurement were collected just before their scheduled packed red blood cells transfusion along with routine blood examination and serum ferritin. Samples were collected in tube and were centrifuged at 3000 rpm for 10 min within 2 h. Sera were extracted and stored in −80°C until the analysis day. NT-proBNP was measured by electrochemiluminescence assay technique using the Roche e411 cobas immunoassay analyzer (Roche Diagnostics).

Ferritin measurement

Ferritin levels were determined by electrochemiluminescence technique using the Roche e411 cobas immunoassay analyzer (Roche Diagnostics). The mean serum ferritin value was derived from the mean obtained at 3-month interval over the previous year.

Cardiac iron concentration

T2* MRI was performed at the Department of Radiology, Cipto Mangunkusumo Hospital, using 1.5 Tesla MRI scanner (Siemens Avanto Germany). Myocardial T2* was analyzed using dedicated software (Thalassemia-Tools; Cardiovascular Imaging Solutions, London, United Kingdom) with regions of interest in ventricular septum. Each image was acquired during 11–13 s breath-hold, using a gradient echo sequence. The repetition time was 200 ms, the flip angle used was 20°, echo times was 1.3–23 ms, the base resolution matrix was 128 pixels, the field of view was 39.7 cm and 19.7 cm, and the sampling bandwidth was 125 kHz. Results of cardiac T2* were categorized as severe (T2* <10 ms), mild to moderate (10 ms < T2*<20 ms), and acceptable (T2* >20 ms). The imaging procedure was performed within 3 months after laboratory examination. Clinical chemists and radiologists were blinded to any information regarding patient's medical records.

Statistical analyses

Values are expressed as mean ± standard deviation as indicated. All tests were carried out using SPSS Version 23 software (IBM Corp., NY, USA). The calculation was performed using Pearson's and Spearman's correlation coefficient, independent t-test, Mann–Whitney test, and receiver operating characteristic curve analysis. Statistical significance was set at P < 0.005.

RESULTS

We studied 68 patients (mean age: 13.9 ± 2.3, 52.9% [n = 36] males, 47.1% [n = 32] females) with transfusion-dependent β-thalassemia major. Patients demographic and baseline characteristics are presented in Table 1.

Table 1

Baseline characteristics

Fifty-five (80.8%) of the 68 patients had no evidence of cardiac hemosiderosis, corresponding to T2* >20 ms, whereas 13 (19.1%) patients had cardiac hemosiderosis, corresponding to T2* <20 ms. Of 13 patients with cardiac hemosiderosis, 2 of 13 had severe cardiac hemosiderosis, corresponding to T2*<10 ms. Mean ferritin serum levels were increased in patients with cardiac hemosiderosis (T2* <20 ms) compared to nonsiderosis patients (T2*>20 ms) but were statistically insignificant (P = 0.376) [Table 2]. There were three patients in cardiac hemosiderosis group who had mean ferritin serum <2500 ng/dL.

Table 2

Laboratory examination difference across cardiac iron loading

There was no difference of NT-proBNP levels between patients with cardiac hemosiderosis and no cardiac hemosiderosis [Table 2]. There were four patients in cardiac hemosiderosis patients whose NT-proBNP levels was higher than the upper limit of normal (160 pg/mL).[22] There was weak yet insignificant correlation between NT-proBNP levels and age (r = 0.07, P = 0.569).

Further ROC analyses resulted in no significant result for NT-proBNP to detect cardiac hemosiderosis (area under the curve [AUC]: 0.39, P = 0.23, confidence interval [CI] 95%: 0.190–0.596) and severe cardiac hemosiderosis (AUC: 0.54, P = 0.85, CI: 95% 0–1) [Figures 1 and 2].

Figure 1

Receiver operating characteristic curve for predicting cardiac hemosiderosis (T2* <20 ms)

Figure 2

Receiver operating characteristic curve for predicting severe cardiac hemosiderosis (T2* <10 ms)

DISCUSSION

This is the first study assessing NT-proBNP levels exclusively in adolescent β-thalassemia patients. The most important finding of the present study is the inability of NT-proBNP to detect cardiac hemosiderosis in adolescent β-thalassemia patients. Furthermore, we demonstrate that NT-proBNP levels do not differ significantly between patients with cardiac hemosiderosis and normal cardiac iron loading.

β-thalassemia major cardiomyopathy is characterized by early diastolic dysfunction which precedes systolic dysfunction.[1323] Recent advances in chelation therapy has improved cardiac morbidity so the onset of heart failure occurs at around the third decade.[24] Thalassemia cardiomyopathy is silent in young patients while the early diagnosis is crucial since early intervention could reverse the disease process.[10111225] Cardiac iron overload mainly occurs at the epicardium, and hence, the conventional endocardial biopsy does not reflect the mean cardiac iron.[16] Measuring cardiac iron loading has become possible using MRI T2* and is well correlated with cardiac iron loading in vivo.[1626]

The prevalence of cardiac hemosiderosis in this study (19.1%) was lower than that in other published studies. A study in Hong Kong[27] found 50% abnormal cardiac iron loading while a study in Iran[28] showed a value of 58%. Cardiac iron loading can be detected as early as 6 years of age in a study in China[29] while previous study[30] in our center reported that the youngest abnormal cardiac T2* result was 8 years of age. Although cardiac MRI T2* is the best method to detect cardiac iron overload, there is limited availability in the developing countries. Hence, ferritin serum remains the primary screening tool in our country. In our study, there were three patients with cardiac hemosiderosis who had mean ferritin serum <2500 ng/mL and most patients with high ferritin value did not have cardiac hemosiderosis proven by MRI T2*. Therefore, ferritin levels do not necessarily correlate with cardiac iron overload, and low ferritin value is not free from cardiac complication.[15] The nature of ferritin as an acute-phase reactant and higher prevalence of hepatitis coexistence may limit its use as cardiac marker of hemosiderosis.[3132]

Natriuretic peptides are secreted in response to increased cardiac volume and pressure overload. The release results in improved myocardial relaxation in response to myocardial stretch through vasoconstriction as well as sodium and water retention.[33] Previous studies dealing with NT-proBNP in β-thalassemia patients have shown that NT-proBNP is a sensitive biomarker to detect systolic and latent diastolic dysfunction, while NT-proBNP is slightly more sensitive than BNP due to its longer half-life.[19203034]

A previous study from Tanner et al. observed that BNP was weakly related to myocardial iron and only abnormal in high values of cardiac MRI T2*.[10] Association of natriuretic peptide and cardiac iron was documented for the first time in the study of 187 asymptomatic β-thalassemia major patients by Delaporta et al.[21] They reported that NT-proBNP levels were significantly higher in patients with cardiac hemosiderosis (T2* <20 ms) and the correlation of NT-proBNP and cardiac MRI T2* was only significant in cardiac hemosiderosis patients.[21] Our present study observed that there was no significant difference between NT-proBNP levels and cardiac hemosiderosis, and NT-proBNP levels cannot be used to discriminate cardiac hemosiderosis status. Another study from Mehrzad et al. also found that NT-proBNP levels did not correlate with cardiac hemosiderosis while significant increase was only found in severe cardiac hemosiderosis patients (T2*<10 ms).[35] Most of the reported studies only include adult population as their participants and very few include children and adolescents.[102135]

A study from Kremastinos et al.[19] suggested that NT-proBNP levels are related to age and this increase only became significant in the third decade of life. Kremastinos' study included a very wide range of participant's age (5 until 46 years). Another study by Balkan et al.[20] reported that NT-proBNP secretion begins early in the phase of the disease before overt diastolic dysfunction. They also did not find any correlation between the increase of NT-proBNP and diastolic parameter indices mainly due to narrow age range in the third decade of life. The reason for insignificant result in the present study may be due to earlier age's participants when the increase of NT-proBNP levels has not occurred. We presume that the increase of NT-proBNP in thalassemia patient may not be evident in adolescent population due to better chelation strategies.[82436] The evidence of diastolic dysfunction as an early sign of cardiomyopathy could not be appreciated since we did not assess diastolic parameters using more sensitive tools such as tissue Doppler imaging.[1937] It is important to note also that some variables may affect plasma NT-proBNP levels, including the assay used, age, body mass index (lower levels with higher body mass index), and genetic factors.[38] Another potential bias was the duration of NT-proBNP measurements and imaging evaluation. Sample collection of NT-proBNP could not be drawn at the same time of imaging evaluation (mean duration: 19 days). This reason as well as earlier age of participants may lead to lack of significant discrimination of NT-proBNP and cardiac hemosiderosis.[1921]

Therefore, it is still important to perform MRI T2* to detect cardiac iron loading. The sensitivity and specificity of MRI T2* are appreciable and it is the standard of care most thalassemia centres according to thalassemia international federation's guideline.[39] MRI T2* could identify individuals at risk of iron overload cardiomyopathy before it becomes evident.[1516] Detection of early cardiac dysfunction is of utmost importance since appropriate chelation therapy could allow complete recovery of heart function.[10111239]

CONCLUSION

NT-proBNP levels do not differ between adolescent β-thalassemia patients with cardiac hemosiderosis and no cardiac hemosiderosis. Consequently, this biomarker is unlikely to be valuable as an early detection tool of cardiac hemosiderosis in adolescent β-thalassemia patients.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.