Within the cytoplasm of cells, metabolically active iron is present physiologically in low-molecular-weight forms destined for incorporation into functional compounds or, if present in amounts exceeding cellular requirements, for storage (36
). Excess iron is normally first stored within the protein shell of ferritin, which is soluble and diffusely distributed within cells (12
). As the amount of cellular iron to be stored increases further, the iron is gathered within insoluble aggregates and clumps of varying sizes identified as hemosiderin (13
). As the total amount of tissue iron increases, the proportion stored as hemosiderin rises, from trace amounts in normal individuals to 90% or more in patients with severe iron overload (37
). Increases in the amount of cytosolic iron that exceed the cellular capacity for safe storage are believed to result in oxidative damage, ultimately leading to cell death, organ injury and failure (20
). Recent studies have provided compelling evidence that iron entry and exit from ferritin are the result of an equilibrium based on the concentration of cytosolic iron (18
), suggesting that ferritin iron may serve as indicator of cellular toxicity. Moreover, all three iron-chelating agents in clinical use, deferasirox, deferiprone and deferoxamine, decrease the intracellular concentration of ferritin iron, although by different mechanisms. Therefore, non-invasive measurement of cellular ferritin iron may provide early warning of iron-induced toxicity, and permit rapid monitoring of the effectiveness of iron-chelating regimes.
In the presence of both ferritin and hemosiderin iron, the analytic form of the signal decay, as shown in Eq. 
, can be regarded as a product of the monoexponential factor and a non-monoexponential factor that describes the more complex effects of diffusion in the spatially inhomogeneous field generated by aggregated hemosiderin iron (21
). Most importantly, the effects of ferritin and hemosiderin iron on the signal decay are largely reflected in RR2 and A, respectively. The findings in the current study demonstrate the ability of RR2 to detect myocardial iron changes associated with a 1-week suspension of iron-chelating therapy in transfusion-dependent thalassemia patients. The current results also show that RR2 after resumption of chelation (Day 14) normalized to the baseline level (Day 0). Moreover, RR2 is shown to be more sensitive in detecting changes in myocardial iron than the commonly used relaxation rates including R2* and R2. These initial results suggest the feasibility of better characterization of myocardial storage iron, particularly ferritin iron, using the new transverse relaxation index RR2. This is likely a result of its superior sensitivity to soluble ferritin iron, which is in close equilibrium with the cytosolic iron pool that is expected to increase during the 1-week suspension of chelation and then decrease after resumption of iron-chelating therapy (18
). Furthermore, along with the hemosiderin iron index A, RR2 approach may allow more accurate measurements of total tissue storage iron burden (21
Clinically, determination of T2* (= 1/R2*) in the interventricular septum has been used directly as an index of total myocardial storage iron (8
). However, Anderson et al showed that myocardial T2*, which is predominately sensitive to hemosiderin iron, changed only slowly after months of intensive iron-chelating therapy, despite significantly improved cardiac function within weeks (11
). In line with this finding, no significant difference in R2* was found after suspending and resuming chelation therapy in the current study. Myocardial hemosiderin iron, as estimated by R2*, would be expected to change little during the 1-week periods in our study. It should be noted that methods based on a single metric, such as R2* or R2, have an associated uncertainty reflecting variations in the relative amounts of ferritin and hemosiderin iron (40
). In particular, these methods cannot accurately quantify ferritin iron levels in patients with high iron levels, in which ferritin iron can only be a small fraction of the total iron storage. It is noteworthy that because myocardial R2* increases prominently at 3T as compared with 1.5T, it is technically challenging to use R2* as an iron overload index at 3T where the shortest TE and interecho time can be limited by the peak RF power and gradient strength. More importantly, R2* measurement at high magnetic field (> 1.5T) is more vulnerable to increased B0 inhomogeneity likely resulted from the increased susceptibility and shimming related effects.
With recent advances in the development of multi-echo FSE sequences that permit accurate acquisitions of multi-echo spin-echo signals in the heart during a single breathhold (27
), the RR2 measurement by acquiring spin-echo signal decays with three or more different interecho times has become feasible and practical with a clinically acceptable examination time. Given the specific interecho time dependence predicted by Eq. 
, use of two or more signal decays with different interecho times for fitting could improve the accuracy of RR2 measurement (22
). Acquiring signal decays with three different interecho times as used in this study would be a reasonable compromise between accuracy and scan time for clinical applications. In addition, signal-to-noise ratio increase at 3T can facilitate the acceleration of breathhold acquisition sequence by use of large SENSE factor and half-Fourier sampling as demonstrated in the current study. Such reduction of breathhold time can be a major advantage in patient study, likely leading to more patient comfort and reliable measurements at 3T. On the other hand, it should be noted that for images with long echo times, signal-to-noise ratio could be compromised by increased R2 at 3T compared to 1.5T (34
). Because R2* at high field increases substantially and its quantitation is vulnerable to increased B0 inhomogeneity, the rapid myocardial R2 and RR2 measurement protocols demonstrated in this study may constitute a robust alternative to the traditional R2* measurement for assessment of cardiac iron overload at 3T. Note that patient recruitment in this study was based on prior measurements of T2* (= 21.9 ± 3.1 ms) at 1.5T with selection criterion (i.e., T2* = 20 +/− 5 ms) to exclude patients with severe iron overload. This was to ensure sufficient signal-to-noise ratio in the signal decays at 3T, consequently improving the data and curve-fitting consistency and quality for RR2, R2 and R2* measurements. However, this selection criterion could also be a limitation of the current study because RR2 method could not be verified in patients outside this range of T2* (20 +/− 5 ms at 1.5T). In addition, such selection criterion also limited the number of patients studied (N
= 8) at our institution. Future study without T2* range restriction in a larger number of patients is warranted to further evaluate and validate the RR2 technique.
In conclusion, the experimental findings in this study demonstrate that RR2 measurement can detect myocardial iron changes associated with brief (1 week) changes in iron-chelating therapy in patients with transfusion-dependent thalassemia. While further studies in larger numbers of patients with wider ranges of iron loading over longer periods of time are needed, these initial results suggest that estimation of myocardial ferritin iron level by measuring RR2 may serve as a promising new means to rapidly evaluate the effectiveness of iron-chelating regimes in thalassemia patients. RR2 measurement may also allow monitoring the risk of iron-induced toxicity in patients with iron overload and be helpful in the evaluation of new candidate iron chelators.