Clinical Imaging 37 (2013) 631–636

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Clinical Imaging

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Original articles

Clinical MRI for iron detection in Parkinson's disease☆

Maija Rossi a,⁎,1, Hanna Ruottinen b,1, Seppo Soimakallio a,c, Irina Elovaara b,c, Prasun Dastidar a,c

a Medical Imaging Centre, Department of Radiology, Tampere University Hospital, Tampere, Finlandb Department of Neurology and Rehabilitation, Tampere University Hospital, Tampere, Finlandc University of Tampere, Tampere Medical School, Tampere, Finland

☆ This study was supported by the Alfred KordelinResearch Funding of the Tampere University Hospita⁎ Corresponding author. Medical Imaging Centre, Dep

University Hospital, Teiskontie 35, 33520 Tampere, post3-311-65204; fax: +358-3-311-65586.

E-mail address: maija.rossi@pshp.fi (M. Rossi).1 The authors contributed equally in the study.

0899-7071/$ – see front matter © 2013 Elsevier Inc. Alhttp://dx.doi.org/10.1016/j.clinimag.2013.02.001

a b s t r a c t

a r t i c l e i n f o

Article history:Received 31 October 2012Received in revised form 3 January 2013Accepted 7 February 2013

Keywords:IronMagnetic resonance imagingParkinson's disease

We studied nonheme iron in Parkinson's disease (PD) using clinically available MRI in 36 patients and 21healthy volunteers. The subjects underwent thorough clinical investigation, including 3-T MRI. QuantitativeR2* was able to reflect symptoms of PD. In addition, the clinically used susceptibility-weighted imagingdifferentiated between controls and patients, whereas T2-weighted imaging did not. Disease-related changeswere present not only in substantia nigra but also in globus pallidus. Such changes are associated withneurodegeneration, reflecting the severity of motor impairment.

© 2013 Elsevier Inc. All rights reserved.

1. Introduction

Parkinson's disease (PD) is an incurable neurodegenerativedisease. Common symptoms of the disease include slowness inmovement (bradykinesia), muscular stiffness (rigidity), restingtremor, and postural instability. The symptoms originate from thedegeneration of neuromelanin-containing dopaminergic neurons inthe pars compacta of the substantia nigra (SN) that participate inregulation of voluntary movements. Here, information is transferredbetween neurons via dopamine, and cell-death-induced loss ofdopamine results in the known symptoms of PD [1]. Magneticresonance imaging (MRI) of PD patients shows few changes whencompared to healthy individuals, but there may be slight volumetricchanges, especially in the SN [2–4].

Normal aging is associatedwith the accumulation of nonheme ironthat is contained in ferritin; especially in the globus pallidus (GP), rednuclei, SN, and putamen [5–8], and it is vital in the synthesis ofdopamine [9]. Normally, iron is stored in compounds such as ferritin.However, if iron is present in an unbound form in tissues, it becomestoxic andmay lead to cell death [8–11]. In patients with PD, significantiron accumulation in the SN has been observed [2,12–23]. In addition,

Foundation and Competitivel.artment of Radiology, Tamperebox 2000, Finland. Tel.: +358-

l rights reserved.

the caudate nuclei of PD patients may have increased iron levelscompared to those of normal age-matched controls [21]. Controver-sies arise as to whether iron levels in the putamen increase[8,19,20,23,24], decrease [2,12–14], or remain unchanged [15–17].Similar controversies exist for GP iron increase [7,21], decrease[17,18], and neutrality [15]. The eventual decrease in putaminal ironmay correlate with disease duration [2,18].

The field of study has been extensively studied in previousliterature using sophisticated imaging methods. Therefore, we feelthat the next step would be to show their applicability in clinicalenvironment. The purpose of this study is to determine the R2* andsusceptibility-weighted imaging (SWI)-given contrast (i.e., putativeiron content) in the basal ganglia of PD patients using MRI methodsreadily available for clinical use. Correlations with clinical character-istics and symptoms alongwith a comparisonwith healthy volunteerswill be presented.

2. Materials and methods

2.1. Clinical characteristics of the patients

The Ethics Committee of Tampere University Hospital approvedthe study, and all of the patients gave their informed consent. Thirty-seven patients with PD (age range=42–86, mean=69, males:females=19:18) were recruited for the study. The patients werereferred from local health centers to the university hospital for clinicaldiagnosis. To be included in the study, the patients were required tohave two or more of the following symptoms: resting tremor,bradykinesia or hypokinesia, rigidity, or postural instability. Patientswere excluded from the study if they had Alzheimer's disease or any

632 M. Rossi et al. / Clinical Imaging 37 (2013) 631–636

other type of dementia that was diagnosed within 1 year precedingthe study. The other exclusion criteria included contra indications forMRI, alcohol or drug addiction, pregnancy, or severe general illnessessuch as cardiac, lung, or gastrointestinal disease; liver or kidneymalfunction; active malignant neoplasm; and neurological or psychi-atric disease. In addition, 1 patient was excluded due to claustropho-bia that led to discontinuation of the MRI examination. At the time ofentering the study, levodopa medication had been started with4 patients. These patients were at initial stages of the disease, andthere was no on–off state variation.

All of the patients underwent a thorough clinical examination andneuropsychological tests. The examination included the UnifiedParkinson's Disease Rating Scale (UPDRS) motor examination [25].The motor examination of the UPDRS scoring includes evaluations ofspeech, facial expression, resting tremor (face; upper and lowerlimbs), action or postural tremor, rigidity (neck; upper and lowerlimbs), finger taps, hand movements, rapid alternating movements ofthe hands, leg agility, rising from a chair, posture, gait, posturalstability, and body bradykinesia and hypokinesia. Each of these wasmeasured on a scale from 0 to 4, and the leg and hand evaluationswere measured bilaterally. The total range of the UPDRS motor scorewas thus from 0 to 108. Alzheimer's disease was excluded using theMini-Mental State Examination.

Twenty-one healthy volunteers (age range=58–80, mean=66,males:females=4:17) were recruited for the study. The volunteerswere recruited among hospital employees and the patients' spouses,and they had similar education level. The inclusion criterion was aclinical examination with no abnormal findings. The exclusioncriterion was a history of cerebrovascular attack. The volunteersunderwent MRI examinations similar to the PD patients.

2.2. MRI protocol

Imaging was performed using a 3-T MRI device (Siemens TrioTim,Erlangen, Germany) with a standard body coil for transmitting and a12-channel head coil for receiving. We acquired the MapIt, SWI, andT2-weighted sequences. The MapIt sequence provided by themanufacturer (Siemens, Erlangen, Germany) was used to map T2*[26,27]. The imaging parameters were as follows: voxel size=0.29 *0.29 * 4 mm3 (interpolated from 0.57 * 0.57 * 4 mm3), field of view=220 * 220 mm2, slice gap=0.8 mm, TR=422 ms, TE1/TE2/TE3/TE4/

Fig. 1. Examples of ROI settings in a 71-year-old female with a 4-month symptom duration in(4) SN compacta; and red nucleus (5) in a susceptibility-weighted image. The ROI placeme

TE5=4.18/11.32/18.46/25.60/32.74 ms, flip angle=60°, and numberof slices=11. Interpolation was performed for better visualization;this did not affect the quantitative measurements through the partialvolume effect because the outer voxels of the structures wereexcluded from the regions of interest (ROIs). Three-dimensionalSWI [28,29] was acquired with voxel size=0.90 * 0.90 * 1.5 mm3, fieldof view=172.5 * 230 mm2, TR=27 ms, TE=20 ms, and flip angle=15°. Typically, 96 slices were acquired, and we used the SWI imageswithout a minimum intensity projection. Instead of the phase images,we used the clinical SWI data (magnitude images multiplied fourtimes with the phase mask). In five patients, the MapIt sequence wasnot acquired because it was not available at the time the first patientswere scanned. We also used clinical three-dimensional T2-weightedimaging (T2WI), using the sampling perfection with applicationoptimized contrasts using different flip-angle evolution (SPACE)acquisition method [30]. The imaging parameters were voxel size=0.60 * 0.60 * 3 mm3, field of view=172.5 * 230 mm2, TR=3200 ms,TE=354 ms, and flip angle=120°.

2.3. Image analysis

Image analysis was performed using the free software programImageJ 1.42q (National Institutes of Health, USA). All of the imageswere analyzed by the same person (MR) under the supervision of anexperienced neuroradiologist (PD); the investigators were blinded topatient status. The ROIs were placed bilaterally in a single slice, wherethey were best observed. The ROIs included the lateral and medial SNpars reticulata (SNr) and compacta (SNc), red nuclei, nucleusdentatus, caudate nucleus, anterior and posterior putamen, anteriorand posterior GP, thalamus, and basilar pons; and for contrastmeasurements, the genu of the corpus callosum (CC). Differentiationbetween the lateral and medial SN, as well as between the anteriorand posterior sections of both the putamen and GP, was similar to thatused by Martin et al. [16]. The ROIs were drawn freehand to includethe whole structure within a single slice but to exclude structureborders to avoid partial volume effects. Examples of ROI settings attwo levels are shown in Fig. 1 and Fig. 2. Image coregistration was notused, but the ROIs were carefully set for visual agreement between thesequences to assure identical ROIs.

Brain atrophy was analyzed using FMRIB Software Library (FSL)v5.0 (FMRIB Analysis Group, Oxford, UK) on the T2WI as their contrast

a (A) T2* map and (B) SWI image. Medial (1) and lateral (2) SNr; medial (3) and lateralnts mimicked those of Martin et al. [16].

Fig. 2. Examples of ROI settings in a 63-year-old male with a 24-month symptomduration in a T2* map. Genu of the CC (1); thalamus (2); caudate nucleus (3); anterior(4) and posterior (5) putamen; anterior (6) and posterior (7) GP in a T2* map. Theputamen and GP were measured one slice below, but they are presented here asexamples. The ROI placements mimicked those of Martin et al. [16].

633M. Rossi et al. / Clinical Imaging 37 (2013) 631–636

gave better segmentation results than our T1WI. First, the brainextraction tool [31] was used with the option of robust brain centerestimation and fractional intensity threshold of 0.2. Next, the resultswere inspected and hand edited where necessary. Segmentation wasthen computed with the FMRIB's Automated Segmentation Tool [32]with five tissue-type classes. These five classes were interpreted asnonbrain tissue, white matter (two classes), grey matter, andcerebrospinal fluid (CSF). For whole brain–CSF separation using ourMRI data, the results were more reliable when initially five classeswere used rather than three or four. Afterwards, the grey matter andthe two white matter classes were added together, and CSF wasseparated from the whole-brain volume. Finally, eventual age-relatedlesions were hand edited not to be included in CSF, and the resultingvolumes were then recorded.

2.4. Statistical analysis

The MRI results were analyzed with respect to the clinicalcharacteristics and symptoms of the patients. We used R2*=1/T2* toconform to the previous literature. Because both SWI and T2WI werenonquantitative, the signal intensity of each structure was analyzed asthe contrast against the genu of the CC, c=(Sa−SgCC)/(Sa+SgCC),where Sa and SgCC are the signal intensities of the concerned structureand genu of the CC, respectively. The clinical parameters used in thecorrelation analysis included age, symptomatic disease duration, and

Table 1MRI differences between patients and healthy volunteers measured using Student's t test f

P value

Medial SN compacta 0.001a (R2*); 0.013a (SWI); 0.874 (T2WI) HePat

Lateral SN compacta b0.001a (R2*); 0.122 (SWI); 0.982 (T2WI) HePat

Anterior GP 0.059 (R2*); 0.025a (SWI); 0.223 (T2WI) HePat

Posterior GP 0.708 (R2*); 0.660 (SWI); 0.029a (T2WI) HePat

In R2*, nhealthy=21 and nPD=34; in SWI, nhealthy=21 and nPD=36.T2WI=T2-weighted imaging.

a Statistically significant, P values corrected for multiple comparisons.

the UPDRS motor score. In addition, although they were alreadyincluded in the UPDRS motor score, resting tremor, rigidity, andhypokinesia were individually tested against the MRI results. Groupcomparisons were tested by gender (male/female), postural instabil-ity (stable/unstable), whether the patient had already receivedtherapy at the time of imaging (yes/no), and the side of symptomaticdisease onset (right/left). The clinical characteristics were testedagainst the MRI parameter averaged over the left and right lobes.However, for comparisons between the MRI and symptoms, theclinical scores for the side with maximum symptom severity weretested against the contralateral brain lobe.

The normality of the data distribution was tested using a one-sample Kolmogorov–Smirnov test. Group comparisons were per-formed using Student's t test for independent samples, and correla-tions were tested using Pearson's correlation test. All statistical testingwas conducted with IBM SPSS Statistics 20.0.0 (SPSS, Chicago, IL,USA). The significance level was initially set at Pb .05. We thenperformed the Bonferroni post hoc test to correct multiple compar-isons and present the corrected P values only.

3. Results

Thirty patients were diagnosed with clinically definite PD, and 7were diagnosed with highly probable PD. One patient was excludeddue to a considerably longer duration of symptoms (78 months)compared to the others (median=12 months, range=3 to 41months) to reduce statistical outliers. The 36 PD patients included18 males and 18 females, aged from 42 to 86 years (median=71years), and having their UPDRS motor scores between 9 and 49(median=21). These patients included two of the first five patientsnot imaged with the MapIt sequence. Therefore, the patient popula-tions for R2* and SWI differ slightly. We found no significantdifferences in the statistical results between the SWI and clinicaldata when testing with all 36 patients or with the 34 patients imagedwith the MapIt sequence. Examples of the image quality in both theT2* map and SWI are shown in Figs. 1 and 2. Therapy had been startedin 15 patients at the time of MRI. Among these patients the duration ofthe treatment was very short, that is, only few months. No differencein the MRI findings was found between groups.

3.1. Comparison to healthy volunteers

Differences between the patients and healthy volunteers areshown in Table 1 for both R2* and SWI. Relaxation and susceptibilityeffects were enhanced in the SNc and anterior GP of the patients. InT2WI the results diverged, with only GP posterior differing betweenPD patients (contrast=98%±8%) and healthy volunteers (contrast=93%±5%) (P=.022). Brain atrophy was similar between patients andhealthy volunteers (P=.842).

or independent samples

R2* (1/s) SWI contrast (%) T2WI contrast (%)

althy 43±7 9.9±6.5 18.5±5.5ient 51±10 5.7±5.9 18.7±5.5althy 42±6 9.1±5.2 17.4±5.6ient 50±10 7.2±3.5 17.4±5.8althy 62±17 −18±13 −18±13ient 72±18 −26±12 −26±12althy 50±8 −9±5 −7±5ient 52±13 −10±7 −3±8

Fig. 3. R2* correlation with the UPDRS motor score in the anterior GP.

634 M. Rossi et al. / Clinical Imaging 37 (2013) 631–636

3.2. R2* and symptoms

We found modest correlations between the UPDRS motor scoreand the R2*, averaged over the two hemispheres of a given structure,in the basilar pons (but compromised by image heterogeneities) andanterior GP (Fig. 3). However, in the anterior GP, the correlation waslargely dependent on two outliers whose UPDRS motor score wasapproximately 50, as presented in the figure. Tendency to correlatewas found between age and R2* in posterior putamen and anterior GP.A correlation between R2* and the duration of symptoms wasobserved in the medial SNc. The results are presented in Table 2.

In the subanalysis of individual symptoms, the extent of thetremor correlated with the R2* of the medial (r=0.426, P=.027) andlateral (r=0.423, P=.029) SNr and tended to correlate with the R2* ofthe anterior GP, caudate nucleus, and the anterior putamen at the sideof maximum symptoms. No correlations were found between R2* andrigidity or hypokinesia. Patients with postural instability (n=16) hadan increased R2* (82±17 1/s) in the anterior GP compared to patientsfree of this deficit (61±10 1/s) (P=.004). The mere existence of atremor, rigidity, and hypokinesia were not tested because nearly all ofthe patients had these symptoms, and statistical group comparisonswere not considered appropriate.

3.3. SWI and symptoms

For the SWI signal strength, in contrast to the genu of the CC, therewas no significant correlation with the UPDRS motor score, disease

Table 2Pearson correlation and (P value) between MRI interhemispheric averages and patient cha

UPDRS III Duration of PD

R2⁎ SWI T2WI R2⁎

Basilar pons −0.45⁎(0.019) −0.12(1.000) 0.18(0.703) −0.21(0.515)Red nucleus −0.03(1.000) −0.04(1.000) 0.01(1.000) 0.13(1.000)Medial SNr 0.17(0.794) −0.27(0.269) −0.03(1.000) 0.34(0.103)Lateral SNr 0.04(1.000) 0.06(1.000) −0.16(0.779) 0.33(0.121)Medial SNc 0.27(0.280) 0.20(0.550) 0.09(1.000) 0.43⁎(0.023)Lateral SNc 0.36(0.091) −0.33(0.117) 0.12(1.000) 0.31(0.176)Nucleus dentatus −0.28(0.691) 0.01(1.000) 0.17(0.748) 0.12(1.000)Caudate nucleus 0.10(1.000) −0.13(1.000) 0.04(1.000) 0.09(1.000)Anterior putamen 0.14(0.954) −0.16(0.807) −0.08(1.000) 0.11(1.000)Posterior putamen 0.16(0.820) −0.04(1.000) −0.01(1.000) 0.03(1.000)Anterior GP 0.40⁎(0.045) −0.33(0.126) 0.17(0.740) −0.02(1.000)Posterior GP −0.01(1.000) 0.14(0.918) 0.13(1.000) −0.19(0.660)Thalamus 0.01(1.000) −0.17(0.770) 0.24(0.381) −0.08(1.000)

T2WI=T2-weighted imaging.⁎ Statistically significant, P values corrected for multiple comparisons.

duration, or age in any of the ROIs (Table 2). No correlation betweenthe signal and tremor, rigidity, or hypokinesia was found.

We found that patients with postural instability had a lower signalin contrast to the genu of the CC in the anterior GP (−32±14%) thanthe patients without this deficit (−19±6%) (P=.005).

3.4. T2-weighted imaging and symptoms

In contrast to the genu of the CC, nucleus dentatus (r=0.490, P=.004), caudate nucleus (r=0.396, P=.022), and posterior GP (r=0.366, P=.036) were correlated with hypokinesia. No correlationswith the extent of tremor or rigidity were found. Age was correlatedwith the contrast in caudate nucleus and putamen posterior (Table 2).

3.5. Quantitative T2*

We present the median T2* averaged over the two hemispheres ofall of the patients in Fig. 4. The shortest T2* was measured in the SNr,and the longest was measured in the caudate nucleus. The differencebetween the anterior and posterior parts of the putamen and of the GP(Pb .001) and the difference between the medial and lateral SNr weresignificant (P=.005). These results indicate that there were signalgradients within the structures. However, the difference between themedial and lateral SN was not significant in the pars compacta (P=.653). The nucleus dentatus was within the measured slices in only 22out of the 32 patients. The range of interindividual variation in theparameters is shown in Fig. 4. The standard deviation was relativelyhigh (6.8 ms) in the basilar pons that suffered from heterogeneities.The SNc was small and may have suffered from partial volume effectsdespite careful ROI selection.

4. Discussion

Neurodegenerative PD is presumed to progress in six pathophys-iological stages [33]. At Stage 3, the disease progresses to the SNc, andthe first symptoms emerge [33]. The disease is associated withincreased iron deposits in the SNc [2,7,12–21,23,34]. Althoughmeasurements of the field-dependent relaxation increase haveshown a good correlation with brain iron content [35,36], the methodsuffers from the major disadvantage of requiring imaging in two ormore fields. Parametric mapping is used to noninvasively indicateputative brain iron alterations associated with the disease.

Clinical environments are often equipped with 1.5-T devices, butthe number of 3-T devices is constantly increasing. For example, ouruniversity hospital has five MRI scanners, two of which operating at3 T. The different voxel sizes, including variation in the slice thickness,

racteristics

Age

SWI T2WI R2⁎ SWI T2WI

−0.07(1.000) 0.16(0.716) −0.07(1.000) 0.09(1.000) −0.07(1.000)−0.06(1.000) −0.02(1.000) 0.12(1.000) −0.15(0.884) −0.21(0.479)−0.09(1.000) 0.03(1.000) −0.03(1.000) −0.12(1.000) −0.16(0.779)

0.00(1.000) 0.04(1.000) −0.30(0.185) −0.13(1.000) −0.08(1.000)0.25(0.321) 0.24(0.378) 0.08(1.000) 0.02(1.000) −0.11(1.000)

−0.04(1.000) 0.17(0.711) 0.04(1.000) −0.15(0.842) −0.28(0.226)−0.01(1.000) 0.01(1.000) −0.35(0.457) 0.03(1.000) −0.16(0.819)−0.03(1.000) 0.02(1.000) 0.34(0.110) −0.16(0.814) −0.43⁎(0.021)−0.07(1.000) −0.02(1.000) 0.22(0.487) −0.13(1.000) −0.35(0082)−0.12(1.000) 0.08(1.000) 0.38(0.056) −0.14(0.942) −0.40⁎(0.034)

0.03(1.000) 0.10(1.000) 0.37(0.073) −0.26(0.293) −0.15(0.845)−0.01(1.000) −0.05(1.000) 0.07(1.000) −0.04(1.000) −0.02(1.000)

0.24(0.361) 0.27(0.249) −0.08(1.000) −0.10(1.000) −0.04(1.000)

Fig. 4. T2* averaged over the two hemispheres in 34 patients and 21 age-matchedhealthy volunteers. A significant difference was observed in the SNc. In the anterior GP,the difference was significant in SWI (not shown) but not in T2*. The medians (lines),interquartile range (boxes), 95% confidence intervals (T-bars), and outliers (spheres)are presented. In nucleus dentatus npatients = 22.

635M. Rossi et al. / Clinical Imaging 37 (2013) 631–636

may slightly affect the results through different amount of partialvolume effects, especially in small structures. However, as ourintention was to study the use of real clinical sequences, thesedimensions were used as they are in the daily routine in this hospital.To minimize effects on the results, structure borders were avoided inthe analysis. Small structures are vulnerable to partial volume effectsand require very careful ROI placement. It is through consistent resultsin publications by different research groups that we can considerthese results reliable.

In clinical environments, the putative iron concentration can beindicated using R2*, SWI, and T2WI [37]. In this study, these sequenceswere associated with symptoms, age, and disease duration in 36patients with PD. However, we did find better correlations with thequantitative R2* thanwith the clinical, nonquantitative SWI and T2WI.Better correlations have been achieved with SWI using phase mapsinstead of the clinical images [16,36,38–40]. However, phase maps aregenerally not used by clinicians in hospital environments and werenot the interest of this study. Our aim is to confirm our data in anongoing 2-year follow-up study using the same MRI sequences.

In 98 human autopsies, which excluded patients with cerebrovas-cular and neuropsychiatric disorders, the nonheme iron contentwas highest in the GP and red nucleus, followed by the SN [5]. Theiron content increases with age in the first two decades of life. Inputamen, the increase slightly continues at older age [5,36], seen inour study as a weak correlation in the posterior part in R2* and T2WI.In our PD patient cohort, the index of the putative iron content,measured with R2* and SWI, was elevated in the SNc and anterior GPcompared to controls.

There are contradictions between studies as to whether MRIdoes [16,19,41,42] or does not [20] correlate with symptoms in theSNc. Unlike other researchers, Antonini [20] placed the ROI in thehypointense area that is usually referred to as the pars reticulata inother studies, including ours. Another study also placed the ROI in

the hypointense area and found a correlation to motor symptoms[43]. We found no correlation between UPDRS III and the lateralSNc, but there was a modest correlation in the anterior GP.However, this correlation was dependent on two patients withhigh UPDRS motor scores and should therefore be consideredpreliminary. These structures' MRI results, along with the rednucleus, were also correlated with the extent of tremor. Previousliterature also shows correlations in the GP [16] but not in theputamen [16,19], which is consistent with our study. In the basilarpons, the results are inconclusive due to field heterogeneities.Postural instability affected the MRI results in the anterior GP. Theiron levels of the GP and the putamen have already been shown tobe associated with other symptoms, namely, tremor [44]. There-fore, we may assume that the iron content of GP may be ofimportance in the pathogenesis of PD.

An increasing gradient in the iron content from the anterior to theposterior putamen [19,20] was confirmed in our study. We alsoshowed an inverse gradient in the GP. The discrepancies in putaminaliron behavior in PD patients cannot, however, be explained byinterstudy variation in the ROI placement combined with thegradient. One study found a difference between PD patients andhealthy controls in the total putamen but not in the posteriorputamen [20]. Studies that divided the putamen into anterior andposterior areas still disagree on this issue [16,19]. It has beensuggested that the controversial iron content decrease depends ondisease duration [2]. Similarly, due to the progressivity of PD, R2* andSWI may be expected to reflect increasing neurodegeneration withtime in the SN, although earlier investigations failed to find such acorrelation [15,20]. Although we could not show iron level changes inthe putamen in relation to either disease duration or healthyvolunteers, the putative iron content of the medial SNc, indicated byR2*, was correlated with the symptomatic duration of the disease. Thedifferences compared to the earlier study on the relations between SNiron and disease duration [35] are probably due to differences in bothimaging methods and ROI definitions. Indeed, what was defined as SNin the earlier study [15,20] was defined as the SNr in our study. Here,we also found only a very slight tendency toward correlation.

The T2* fitting procedure used in the MapIt sequence assumes amonoexponential decay. This assumption, however, may over-estimate R2* through signal losses because intravoxel gradientslead to nonexponential decay, such as sinc-formed decay, and thisoverestimation is directly related to the estimation of iron content[45,46]. Fitting to positive magnitude images may also lead to biasthrough a nonzero mean of noise. However, the SNr observed in theimages was high. In addition, susceptibility gradients (thoseunrelated to iron) interfered with the R2* relaxation determination,which further increased R2*. Further, the expected increase in thewater content of the diseased tissue due to neuronal loss may affectthe relation between R2* and iron content [8]. Increasing watercontent decreases cellular complexity, static field fluctuations, andR2 and R2*. These decreases conversely affect the cellular ironincreases that are caused by PD and detected byMRI andmay, hence,decrease the expected linear correlation between iron content andT2*. It has been suggested that measuring the field-dependentrelaxation [43,47] or measuring R2'=R2*−R2 compensates for thiseffect [42,48]. However, one study between simple motor reactiontimes and R2' or R2* indicated that R2* may not be a worse predictorof iron content than R2' [42].

In conclusion, both R2* and SWI detected differences betweenpatients and controls, but T2WI did not. The quantitative, clinicallyseldom used R2* was more powerful than the two clinical sequencesin reflecting clinical characteristics in PD patients who normally donot show considerable changes on MRI. These changes are probablycaused by increased iron deposits and were present not only in the SNbut also in the GP. Such early changes may help when planning andmonitoring neuroprotective therapy for PD.

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