Neuroradiology(1995) 37:568-575 9 Springer-Verlag1995

S. Confort-Gouny J. Vion-Dury B. Chabrol E Nicoli P. J. Cozzone

Received: 7 January 1994 Accepted: 11 July 1994 S.Confort-Gouny- J. Vion-Dury 9 E J. Cozzone (~]) Centre de R6sonanceMagndtique Biologique et M6dicale (CRMBM), URA CNRS 1186, Facult6 de M6dicine de Marseille, 27 Bd J. Moulin, F-13005 Marseille, France B. Chabrol Service de Neurop6diatrie, H6pital d'Enfants, Centre Hospitalo Universitaire de la Timone, 13385 Cedex 5 Marseille, France E Nicoli Service de Neurologie,H6pital Ste Marguerite, 13009 Marseille,France

Localised proton magnetic resonance spectroscopy in X-linked adrenoleukodystrophy

Abstract We have performed localised proton magnetic resonance spectroscopy (MRS) of the brain on four patients with X-linked adrenoleukodystrophy (X-ALD). The spectrum is characterised at the beginning of the disease by a decrease in N-acetylaspartate and phosphocreatine-creatine content. Choline is strongly increased, and lactate can be detected in some cases. A proton signal from the CH 2 groups borne by free intracellular very long chain fatty acids can also be observed. Later in the disease, the levels of all metabolites, in particular NAA, decrease significantly. The progression

Introduction X-linked adrenoleukodystrophy (X-ALD) is a metabolic encephalopathy of childhood characterised by peroxisomal dysfunction leading to abnormal accumulation of very long chain fatty acids (VLCFA) in various organs, including the brain and adrenals [1-4]. The clinical features are progressive intellectual impairment culminating in dementia and blindness, and adrenal insufficiency. CT and MRI [5, 6] show white matter disease spreading forwards from the occipital lobes. MRI usually displays high signal on T2-weighted images, related to demyelination and white matter oedema. Localised proton magnetic resonance spectroscopy (MRS) of the brain provides a new noninvasive technique for exploring cerebral metabolism in vivo, which can be performed following a standard MRI examination [7]. Studies have been reported in children showing metabolic changes on in vivo localised proton MR

of neurometabolism documented by MRS correlates well with MRI and clinical progression on follow-up study. In one case, the metabolic profile recorded by proton MRS was abnormal before any change occurred on MRI. Proton MRS of the brain might be the method of choice for monitoring patients with XALD, to screen presumed cases and to study the effects of treatment.

Key words AdrenoleukodystrophyLocalised proton NMR spectroscopy. Demyelination

spectra in various metabolic encephalopathies [8], including Canavan's disease [9], Leigh's disease [10], infantile neuronal ceroid-lipofuscinosis [11] and ornithine carbamoyl transferase deficiency [12]. Brain metabolism of some peroxisomal disorders has been studied by localised proton MRS [13], and spectra from patients with X-linked ALD have been documented [14]. We present four additional cases of XALD, including serial examinations on two of them, documented by localised proton MRS using short stimulated echo times [15].

Patients and methods Patients We examined four patients using a combined proton MRI/MRS protocol; two were examinedthree times during a 16-monthperiod. At the time of diagnosis,X-ALD was confirmedusingVLCFA assay (Table 1). All patients had adrenal insufficiency.

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Table 1 Clinical data concentration (mg/ml) in Patient; age (years) at 1st MRS

Plasma very long chain fatty acids at diagnosis a

Electrophysiological findings at diagnosis

Neurological findings at 1st MRS

1; 19

C26 C26/C22 C~/C22

1.08 0.072 1.29

Nerve conduction velocity normal; brain stem and visual evoked responses abnormal

Seizures; dementia; aphasia; bradykinesia; brady psychism

2; 10

C26 C26/C22

0.60 0.056

Brain stem and Visual evoked responses abnormal

Cortical blindness; central deafness; cerebellar ataxia; spastic tetraparesis

C24/C22

1.30

C26

0.74

0.058

Nerve conduction slowed; visual evoked responses normal

Cerebellar ataxia

cedc~2 C24/C22

1.34 Visual evoked responses abnormal

Impaired intellect; behavioural disorder; no focal signs

3; 12

4; 7

C26

0.90

C26/022 C24/C22

0.058 1.38

a Normal values: C26 0.26 (+_ 0.08) mg/ml; C26/C220.010 (_+ 0.006); C24/C220.08 (+ 0.0g) Case 1

Case 4

This 19-year-old patient suffered from X-ALD, the manifestations of which began at the age of 7 years, with schooling difficulties, and then acute adrenal insufficiency with coma. A maternal grandfather and a maternal great-uncle were suspected of suffering from adrenomyeloneuropathy and died with psychiatric and neurological disorders. This patient was progressively disabled and, at the time of the M R examination, in addition to encephalopathy and adrenal insufficiency, had hypothyroidism and obesity. CT showed diffuse low density of the cerebral white matter, without contrast enhancement. At the time of the M R examination, he was receiving sodium valproate (1500 rag/day) because of a previous status epilepticus, replacement hormone treatment, and 7-globulins (1 g/kg/15 days). For 8 months he had been on a diet with erucic and oleic acids [16]. He died of a respiratory arrest 3 months after the M R examination.

This 7 year-old boy is the younger brother of patient 3. He presented at the age of 3 years, with language and behaviour disorders and rapid intellectual deterioration. He suffers from a more severe form of the disease than his brother and does not attend school. We performed three M R examinations at the same times as those of his brother. In contrast to his brother, progressive intellectual decline has been observed during the period of M R monitoring. However, no new neurological signs were detected. He has the same diet and treatment as patients 2 and 3.

Case 2

This 10-year-old boy suffered from the age of 8 years from behavioural disorders, decrease in language performance, schooling difficulties, and a squint, related to rapidly progressive X-ALD. His mother is asymptomatic, but one of his three sisters is heterozygotous for X-ALD. He was severely disabled at the time of the M R examination. For more than one year, he had been on a diet with erucic and oleic acids, and treatment with v-globulins (1 g/kg/15 days). It was not possible to perform a second MRS examination. Case 3

X - A L D was detected in this 12 year-old boy at the age of 9 years, after a genetic check of his family following the discovery of X - A L D in an older brother, who is demented. A maternal uncle, previously suspected of having the same disease, died before any investigations could be made. Initial examination revealed only a cerebellar syndrome. This boy has normal intellectual development and can still follow a normal curriculum at school. He has adrenomyeloneuropathy: nerve velocity in the peroneal sciatic nerve is reduced. We performed three successive M R examinations, in June and November 1991 and September 1992. During this 16 month-period, his neurological state was unchanged; his cerebellar syndrome was stable, He is having a diet and treatment similar to that of patient 2.

M R I and localised proton MRS The magnetic resonance examinations was conducted on a 1.5 T images. Standard axial spin-echo (SE) imaging was performed using a double-echo sequence (TE 60 and 120 ms, T R 3600 ms, slice thickness 5 mm (Fig. 1 a). In case 1 only, localisation of the spectroscopic volume of interest (VOI) was performed using the PSIF3D sequence (40 ~ flip angle, TE 7 T R 17 ms, 1 acquisition, slice thickness i mm (Fig. l b ) [17]. Proton MRS was performed at 63 MHz using the stimulated echo acquisition mode (STEAM) sequence combined with a chemical shift selective excitation (CHESS) sequence to suppress the water signal [18]. The stimulated-echo time was 34 ms in case 1 and 20 ms in the others, repetition time 1500 ms, and a spectrum was typically acquired in 256 scans. The VOI was 2 c m x 2 cm x 2 cm = 8 cm 3. Resonances were assigned according to the literature [18-20]. Only one VOI was examined. In all cases, the total duration of the exploration never exceeded 45 min. With patients 3 and 4, particular attention was paid to positioning the spectroscopic VOI at the same site for the three MRS examinations. The power required to deliver a 90 ~ radiofrequency pulse and the load factor of the coil were similar for patients 2, 3, and 4, and for a 6-year-old control subject throughout the follow-up period, and the stability of the system was checked with a phantom before each examination. Processing of spectra Resonances were assigned to INS (inositol and glycine, 3.5 ppm), T A U (taurine and scyllo-inositol; 3.3 ppm), C H O (choline-containing molecules; 3.2 ppm), PCr-Cr (phosphocreatine-creatine;

570

f i g . l a Axial spin-echo image, showing major demyelination in the centrum ovale of patient 1. b Axial PSIF-3D image of the same patient showing the spectroscopic VOI. e Proton spectrum acquired from this area, using a STEAM pulse sequence with a 34 ms stimulated echo. d Control spectrum acquired from the same area, with the same pulse sequence, in a 23 year-old healthy volunteer

CHO

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3 ppm), GLU-GLN (glutamate-glutamine; 2.10-2.45 ppm), NAA (N-acetyl-aspartate and N-acetyl-aspartyl-glutamate; 2.00 ppm). Signals corresponding to lipids and/or proteins were detected at 0.8-1.5 ppm. Signal centered at 1.3 ppm was tentatively assigned to lactate when it was a doublet (J-coupling 7 Hz). No absolute quantitation was performed at this stage of routine application of localised proton MRS. Phase of the spectra was carefully corrected, in the same manner for all spectra, by the same experimenter. The areas of the MR signals were calculated on the spectra using the Siemens baseline correction (spline method using polynomial function between selected points) and area calculation software. In the absence of automatic curve fitting, and because the high-frequency parts of the NAA and INS resonances overlap with resonances from other amino-acids (GLX) we used the integration method described by Alger et al. [21] by beginning the integration at the top of the signal and continuing in the low-frequency direction until the baseline was reached, and multiplying the result by 2. Phase and baseline corrections were standardised (points between polynomial spline functions were always positioned at the same location in the spectra), and comparison of areas calculated using this protocol by various experimenters showed differences of tess than 10 %. In addition, all integrations of resonances on the spectra recorded on the 4 patients were calculated in triplicate, and no significant change (< 10 %) was observed in the 3 runs. The data are presented as ratios of the signal areas of the metabolites of interest.

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Results

M R I of the four patients s h o w e d the typical white matter ( W M ) lesions f o u n d in X - A L D (Table 2). T h e clinical state of the patients c o r r e l a t e d well with the extent of the W M lesions (Fig. 2 a). I n all f o u r patients, X - A L D was associated with m a jor c h a n g e s in brain m e t a b o l i s m . I n patients 1 and 2 , with the m o r e active f o r m s of the disease, a m a r k e d decrease o f N A A resonance, a n d abnormalities in the C H O p e a k were: increase of C H O with r e s p e c t to P C r in case 1, a n d strong r e d u c t i o n of the C H O p e a k in case 2 (Fig. 1 c, 2c). I n addition, the brain spectra disp l a y e d low levels of all metabolites, particularly in patient 2, with the m o s t progressive f o r m of the disease. I n the p a t i e n t with the milder f o r m of the disease (patient 3), the initial s p e c t r u m (Fig. 3 d) was similar to that r e c o r d e d f r o m a h e a l t h y c o n t r o l (Figs. 2d, 4). H o w e v e r , the first s p e c t r u m r e c o r d e d f r o m his b r o t h e r (case 4), w h o has m o r e severe intellectual i m p a i r m e n t and m o r e extensive W M lesions, displayed an increased C H O / N A A ratio (Fig.4), and b r o a d signals w e r e p r e s e n t in the 0.8-1.5 p p m p a r t of the s p e c t r u m (Fig. 5 d). T h e p r o g r e s s i o n of the m e t a b o l i c profiles of the two b r o t h e r s (patients 3, 4) displayed c o m m o n features: N A A / C H O ratio d e c r e a s e d and C H O / P C r - C r a n d INS/

571

Fig. 2,a, b MRI of patient 2, with the most progressive XALD, performed at an advanced stage of the disease. b Also shows the spectroscopic VOI. e,d Localised proton spectra of this patient and a control of about the same age

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P C r - C r ratios increased during the 1 6 - m o n t h p e r i o d of o b s e r v a t i o n (Fig; 4) c o r r e s p o n d i n g to an increase in the C H O r e s o n a n c e and, a d e c r e a s e in N A A and PCr-Cr, the I N S r e s o n a n c e r e m a i n i n g stable t h r o u g h o u t . I n these b o y s , an increase in G L U - G L N r e s o n a n c e c o m p a r e d to the c o n t r o l was s o m e t i m e s o b s e r v e d (Fig. 3 d f, 5 a, c). T h e 0.8-1.5 p p m p a r t of the s p e c t r u m r e c o r d e d f r o m patient 4, was also a b n o r m a l , with two additional resonances, which could be assigned to the C H 2 and

C H 3 r e s o n a n c e s of lipids, and a d o u b l e t s u p e r i m p o s e d at 1.33 p p m which could c o r r e s p o n d to l a c t a t e / J c o m p l i n g constant: = 7 Hz). T h e d e g r a d a t i o n of the p r o t o n spect r u m is c o r r e l a t e d well with the extension of the lesions on M R I and the clinical d e t e r i o r a t i o n of p a t i e n t 4 . T h e r e was a d i s c r e p a n c y b e t w e e n M R S , M R I and clinical features in patient 3 with early m e t a b o l i c changes o n M R S (decrease in the area of N A A a n d increase in that of C H O ) while no n e w clinical or M R I a b n o r m a l i t y

Table 2 White matter lesions shown by T2-weighted MRI (0 no lesion, + small lesions, ++ moderate lesions, +++ extensive lesions) Patient

June 1991 November 1991 September 1992 June 1991 November 1991 September 1992

Corpus call0sum

Parieto-occipital

Anterior

Geniculate bodies

+++

+++

+++

++

Pyramidal, occipitopontiue or lemniscal tracts +/-

+++ + + + ++ +++ +++

+++ +/+/+/+ ++ +++

0 0 0 0 0 0 0

+++ 0 0 0 0 + ++

+++ 0 0 0 +/++ ++

572

Fig. 3 Progression of MRI and proton MRS in patient 3, with a less progressive form of XALD, without intellectual impairment, a,b MRI obtained at the first and third examination, e the VOI position selected for all spectra in the longitudinal study, in the posterior part of the centrum semiovale, d-f Spectra recorded in June and November 1991 and September 1992. Spectral resolution calculated from the water resonance half-linewidth was 5 Hz, 5 Hz and 4 Hz respectively in the VOI. This high resolution accounts for the sharp lines recorded for all 3 spectra. On visual inspection of the NAA and CHO signals variation in line intensity is not readily apparent; significant differences do exist when areas are computed

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was apparent. We have checked that no significant variations in m e t a b o l i c ratios (less than 10 %) were observed on spectra r e c o r d e d in control children over a 24 month-period.

Discussion Few pathological conditions of the brain have b e e n so far studied in details by localised p r o t o n M R S with short stimulated echos; we report four examples of the m e t a -

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bolic profile of X - A L D . The decrease in N A A , PCr-Cr and INS signals in X - A L D indicates b o t h a loss of neurones and complex metabolic lesions in the remaining cells. N A A , often the m o s t intense signal of the p r o t o n spectrum, is said to be a m a r k e r of neuronal activity and viability [7, 20]. The presence of N A A in the active lesion early in the course of X - A L D is in agreem e n t with the pathological findings, characterised in the early stage of the disease, by demyelination without axonal destruction [22, 23]. Destruction of neurons occurs later, accounting for the strong m a r k e d reduction in

573

9 NANCHO [] CHO/PCr [] INS/PCr

and increased CHO. The brain spectra recorded in patients with other peroxisomal disorders such as Zellweger's syndrome or neonatal adrenoleukodystrophy [13], seem to display additional features related to the hepatorenal syndrome in these diseases. Our four patients were at different clinical stages of the disease and we may tentatively suggest a pattern of progression of the metabolic profile of X-ALD, based on our proton MRS observations. During the initial time ] j u n e 91 nov 91 sept 92 I period (patient 3), the disease is characterised by weak, I 1 CONTROL PATIENTN~ 3 progressive, nonspecific demyelination. This induces metabolic changes similar to those observed in multiple sclerosis [22, 25] or HIV-encephalopathy [29-31] with 9 NAA/CHO mainly a decrease in the NAA/CHO ratio. Later, more extensive changes in the spectra occur, with increase in [] CHO/PCr 4" the CHO/PCr-Cr and INS/PCr-Cr ratios, accompanied by the lipid and/or lactate signals. These spectra resem13" " ~ ~ j ~ m NS/PCr ble those recorded in patients with HIV and severe destruction of myelin, as in progressive multifocal 2encephalopathies [30]. The presence of CH 2 or CH 3 resonances could correlate with rapid demyelination and subsequent release of free lipids, as suggested for 9 9 time some MS plaques [25], or the accumulation of VLCFA sept 92 I I _ _ J i june 91 nov 91 in the brain, a process observed in the most aggressive CONTROL PATIENTN~ 4 forms of X-ALD. At the end-stage of the disease, imFig.4 Progression of four metabolic ratios calculated from the MRS in patients 3 (a) and 4 (b). Controlvalues are calculatedfrom pairment of cerebral metabolism and destruction of brain cells in manifest with a drastic reduction of all the spectrum shown in Fig.2 d metabolite signals in the spectra. A significant result of this study is that a diet with erucic and oleic acids, and treatment with 7-globulins NAA in the last spectrum of patient 4 and the spectra of does not prevent the progression of anatomical or metapatients 1 and 2. The increase in CHO signal detected is bolic abnormalities. This is in agreement with the disapthought to indicate demyelination, as in multiple pointing results of the open trial which demonstrated a sclerosis [24, 25]. In X-ALD the increase in the CHO lack of effect of ~ oil" in patients with adrenoresonance is in agreement with the severity and extent leukodystrophy [32]. In patient 3, a bone-marrow transof demyelination observed neuropathologically and by plant is being considered as possible treatment. UnforMRI. In the last spectrum of patient 4, the CHO re- tunately, no histocompatible donor has yet been found. sonance was particularly intense, suggesting very active As suggested by Tsika et al. [17], changes in brain demyelination. The signal centered at 1.35 ppm has metabolism may occur before abnormalities can be debeen tentatively assigned to lactate. During active de- tected on MRI or clinical examination. Although more myelination increased lactate could originate from an examinations are required to support this hypothesis in ischaemic/hypoxic state of the brain due to in- X-ALD it is noteworthy that in HIV-related flammatory processes and local oedema [26, 27], and/or encephalopathies, proton MRS of the brain can reveal from the macrophages infiltrating the X-ALD lesion metabolic defects before the occurrence of any ab[22] which have an anaerobic metabolism generating normality on MRI [33]. We may therefore envision lactate [28]. The PCr-Cr resonance, a metabolic para- monitoring asymptomatic males in families with Xmeter of the brain energy storage system, is reduced at ALD, using repeated noninvasive MRS examinations to the end-stage of the disease, probably in relation to cell detect the onset of metabolic lesions, and perhaps deloss and/or changes in bioenergetic function. No satis- cide on bone marrow transplantation in the very early factory explanation can be proposed at the moment for stage of the disease. Magnetic resonance spectroscopy the variations in inositol, significantly reduced only at a affords a quantitative noninvasive means of monitoring late stage of the disease. the effects of new treatments on the basis of alterations The metabolic changes that we describe here com- of brain metabolism and to demonstrate changes in the plement our preliminary observations and are similar to brain related to the metabolic defect. Staging of brain those reported by Tzika et al. [14], particularly as re- metabolism should facilitate the definition of the critical gards the presence of lactate and mobile lipids, NAA times at which therapeutic decisions must be made.

574

Fig.5 Progression of MRI and proton MRS in patient 4, with a progressive form of X-ALD, and serious intellectual decline over the 16 months of the study, a,b MRI obtained at the first and third examinations, e VOI selected for all spectra. d-f Spectra recorded in June and November 1991 and September 1992. Spectral resolution calculated from the water resonance half-linewidth was 5 Hz, 6 Hz and 4 Hz respectively in the VOI

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Acknowledgements This work is supported by the Administration de l'Assistance Publique ~t Marseille, the Ministry of Health (PHRC), the CNRS (URA 1186), the Association Vaincre les

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Maladies Lysosomales and the Ligue Nationale contre le Cancer. We thank Dr J.M.Franconi, Mr J.Macias and Mr C.Barles (Siemens, France) for their assistance.

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