Correspondence : P.-F. Pradat, D é partement des Maladies du Syst è me Nerveux, Groupe hospitalier Piti é -Salp ê tri è re, 47 � 83, Boulevard de l'H ô pital, 75651 Paris, France. E-mail: pierre-francois.pradat@psl.aphp.fr

(Received 3 January 2012 ; accepted 5 June 2012 )

Amyotrophic Lateral Sclerosis and Frontotemporal Degeneration, 2013; 14: 30–38

ISSN 2167-8421 print/ISSN 2167-9223 online © 2013 Informa HealthcareDOI: 10.3109/17482968.2012.701308

symptoms and lower amplitude of compound action potential amplitudes of the sural nerve (2,6). Assess-ing in vivo the damage of the spinal sensory pathway would bring additional insights for understanding the physiopathology of the disease and its various phenotypes.

Magnetic resonance diffusion tensor imaging (DTI) characterizes water diffusion in the white matter and can probe CST degeneration in the brain (5,7 � 12) and spinal cord (13,14). Notably, Nair et al. found larger DTI abnormalities in more caudal parts of the spinal cord, suggesting that degeneration

Introduction

Amyotrophic lateral sclerosis (ALS) is a fatal neuro-degenerative disease characterized by combined effects on upper motor neurons and lower motor neurons. In conjunction with the progressive damage of the corticospinal tract (CST), autopsy cases and animal studies showed involvement of sensory path-ways (1,2), which was further confi rmed by morpho-metric measures in the somatosensory cortex (3) as well as functional imaging (4) and structural imaging in the white matter (5). Moreover, electrophysiolog-ical measurements in ALS patients showed sensory

ORIGINAL ARTICLE

Involvement of spinal sensory pathway in ALS and specifi city of cord atrophy to lower motor neuron degeneration

JULIEN COHEN-ADAD 1,2,10 , MOHAMED-MOUNIR EL MENDILI 1 , R É GINE MORIZOT-KOUTLIDIS 3 , ST É PHANE LEH É RICY 4,5, 6 , VINCENT MEININGER 5,7 , SOPHIE BLANCHO 8 , SERGE ROSSIGNOL 9 , HABIB BENALI 1 & PIERRE-FRAN Ç OIS PRADAT 1,7

1 UMR-678, INSERM-UPMC, Groupe Hospitalier Piti é -Salp ê tri è re, Paris, France, 2 A.A. Martinos Centre for Biomedical Imaging, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts, USA, 3 D é partement d ’ Explorations Fonctionnelles Neurologiques, AP-HP, Groupe Hospitalier Piti é -Salp ê tri è re, Paris, France, 4 Centre de Neuroimagerie de Recherche – CENIR, Paris, 5 Université Pierre et Marie Curie-Paris 6, Paris, France , 6 ICM – Institut du Cerveau et de la Moelle Epiniere, Paris, 7 D é partement des Maladies du Syst è me Nerveux, AP-HP, Groupe Hospitalier Piti é -Salp ê tri è re, Paris, France, 8 Institut pour la Recherche sur la Moelle Epini è re et l'Enc é phale, France, 9 GRSNC, Faculty of Medicine, Universit é de Montr é al, Montreal, and 10 Department of Electrical Engineering, Ecole Polytechnique de Montreal, QC, Canada

Abstract Our objective was to demonstrate that ALS patients have sensory pathway involvement and that local cord atrophy refl ects segmental lower motor neuron involvement. Twenty-nine ALS patients with spinal onset and twenty-one healthy controls were recruited. Diffusion tensor imaging (DTI), magnetization transfer and atrophy index were measured in the spinal cord, com-plemented with transcranial magnetic stimulations. Metrics were quantifi ed within the lateral corticospinal and the dorsal seg-ments of the cervical cord. Signifi cant differences were detected between patients and controls for DTI and magnetization transfer metrics in the lateral and dorsal segments of the spinal cord. Fractional anisotropy correlated with ALSFRS-R ( p � 0.04) and motor threshold ( p � 0.02). Stepwise linear regression detected local spinal cord atrophy associated with weakness in the corresponding muscle territory, i.e. C4 level for deltoid and C7 level for hand muscles. In conclusion, impairment of spinal sensory pathways was detected at an early stage of the disease. Our data also demonstrate an association between muscle defi cits and local spinal cord atrophy, suggesting that atrophy is a sensitive biomarker for lower motor neurons degeneration.

Key words: ALS , spinal cord , diffusion tensor imaging , magnetization transfer , atrophy

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Spinal sensory impairment and atrophy in ALS 31

of the CST follows a retrograde pattern ( ‘ dying-back ’ ). Magnetization transfer imaging is sensitive to water bound to macromolecules and correlates with myelin content (15,16), as shown in ALS patients (17,18). While the specifi city of DTI and magnetiza-tion transfer to white matter pathology is still under debate (19,20), their combination can potentially improve spinal cord assessment (21,22).

In addition to CST impairment, the degeneration of lower motor neurons occurs at various spinal levels, and gives rise to signs and symptoms such as weak-ness, atrophy, cramps and fasciculations. Detecting local spinal atrophy in metameres corresponding to specifi c altered muscle might have a signifi cant impact for the future non-invasive detection and objective quantifi cation of lower motor neuron degeneration.

This study combines DTI, magnetization transfer, atrophy measurements in the spinal cord of 29 ALS patients, along with clinical evaluation and electro-physiological measurements. We hypothesized that: 1) patients have sensory pathway involvement that could be detected with MRI; and that 2) local cord atrophy refl ects segmental lower motor neuron involvement.

Material and methods

Subjects

Patients with ALS ( n � 29, seven females, mean age 53 � 10 years, median disease duration 1.4 years) and age-matched control subjects ( n � 21, 11 females, mean age 52 � 13 years) were recruited from the Paris ALS Centre and were diagnosed with probable, laboratory-probable or defi nite ALS according to El Escorial criteria (23). None of the patients had sen-sory signs or symptoms. Patients had sporadic ALS except for one female and one male with SOD1-linked familial ALS. All patients had spinal onset (upper limb, 16; lower limb, 11; upper limb � lower limb, 2). Exclusion criteria were signifi cant acute and chronic medical condition, signifi cant psychia-tric or neurological history (other than ALS for patients) – and standard contraindications to MRI. The local ethics committee of our institution approved all experimental procedures of the study, and written informed consent was obtained from each participant.

Patients were clinically assessed on the day of MRI and scored on muscle testing using the Medical Research Council (MRC) score (see Table I). MRC scores were 8/3 for the deltoid and 6/3 for the pol-licis brevis (median/interquartile). The revised ALS Functional Rating Scale (ALSFRS-R) (24) was also evaluated on the same day. The ALSFRS-R was 38/11.5 (median/interquartile).

TMS measurements

TMS was performed on 25 subjects within two weeks of the MRI. One patient died before TMS examina-tion and three patients could not come back for the

TMS examination. TMS was performed using a MAGSTIM 200 device, delivering monophasic stim-ulation through a round coil (9 cm diameter). Sub-jects had no contraindications to TMS. Responses of hand muscle (adductor digiti minimi) were recorded with surface electrodes using a KPnet system (Natus/Dantec, Denmark). The stimulation was fi rst applied at the cervical level (C7-D1) to excite the root near its exit from the spinal cord (25). The peripheral con-duction time, and the motor evoked potential (MEP) amplitude were measured to assess lower motor neu-rons impairment. Higher Facilitation Motor Thresh-old (FMT) has been associated with the degeneration of CST (26). We defi ned the FMT as the minimal stimulus intensity that evoked a response of at least twice the mean activity amplitude. Intensity was dec-remented � then incremented � in a 10%-steps para-digm, starting at 100%. The minimal FMT amplitude was selected out of four trials. The FMT was 60/43.12% (median/interquartile).

The latency of the response obtained at FMT enabled measurement of the total conduction time. The central conduction time was calculated by sub-tracting the total conduction time and the peripheral conduction time. Peripheral conduction time was 13.33/1.59 ms and the central conduction time was 11.91/5.71 ms (median/interquartile).

MRI acquisition . Acquisitions were conducted us-ing a 3T MRI system (TIM Trio, Siemens Health-care, Erlangen, Germany) and a neck/spine coil. For an exhaustive description of the MRI acquisition parameters, the reader is referred to (22).

Anatomical. Cord atrophy was assessed using a sagittal T2-weighted 3D turbo spin echo (TSE) with slab selective excitation: 52 slices, fi eld of view (FOV) � 280 mm, TR/TE � 1500/120 ms, voxel size � 0.9 � 0.9 � 0.9 mm 3 , R � 3 acceleration factor, acquisition time ∼ 6 min.

Table I. Medical Research Council (MRC) scale for muscle strength.

Grade 5 Muscle contracts normally against full resistance.

Grade 4 Muscle strength is reduced but muscle contraction can still move joint against resistance.

Grade 3 Muscle strength is further reduced such that the joint can be moved only against gravity with the examiner's resistance completely removed. As an example, the elbow can be moved from full extension to full fl exion starting with the arm hanging down at the side.

Grade 2 Muscle can move only if the resistance of gravity is removed. As an example, the elbow can be fully fl exed only if the arm is maintained in a horizontal plane.

Grade 1 Only a trace or fl icker of movement is seen or felt in the muscle or fasciculations are observed in the muscle.

Grade 0 No movement is observed

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32 J. Cohen-Adad et al.

DTI. Cardiac-gated DTI data were acquired using a single-shot EPI sequence with monopolar scheme and eight axial slices covering C2 to T2 vertebral levels. Parameters were: FOV � 128 mm, TR/TE � ∼ 700/96 ms, voxel size � 1 � 1 � 5 mm 3 , R � 2, b-value � 1000 s/mm 2 , 64 directions, four repetitions.

Magnetization transfer. 3D gradient echo images with slab-selective excitation were acquired with and without magnetization transfer saturation pulse (Gaussian envelop, duration � 9984 μ s, frequency offset � 1200 Hz). Parameters were: axial orienta-tion, 52 slices (covering the same C2-T2 region as for the DTI scans), FOV � 230 mm, TR/TE � 28/3.2 ms, voxel size � 0.9 � 0.9 � 2 mm 3 , acquisition time ∼ 5 min for each volume.

Data processing

Atrophy measurement. Cord area was measured on the anatomical T2-TSE images in the middle of ver-tebral levels C4, C5, C6 and C7 using the semi-automatic method described in (27). The plane perpendicular to the spinal cord was resampled to maximize the accuracy of area measurements (28).

DTI. Motion correction was applied slice-by-slice using FSL FLIRT (29) with three degrees of free-dom (Tx, Ty, Rz). Diffusion tensor and its related metrics were estimated voxel-wise: fractional aniso-tropy (FA), axial ( λ // ) and radial ( λ ̂ ) diffusivities and mean diffusivity.

Magnetization transfer. Gradient echo data with and without magnetization transfer pulse were coregis-tered using the non-linear algorithm available in FSL (FNIRT). Magnetization transfer ratio was comput-ed voxel-wise following the equation [(S 0 � S MT )/S 0 ] � 100, where S 0 and S MT represent the signal without and with the magnetization transfer pulse, respectively.

ROI-based analysis. Regions of interest (ROI) were manually defi ned on each slice and using geometry-based information (22). To avoid any user bias, ROIs were defi ned on the T2-weighted b � 0 EPI (for DTI analysis) and on the 3D gradient-echo T1-weighted image (for magnetization transfer analysis). The lateral portion of the cord (including CST) and the dorsal col-umns were circumscribed from C2 to T2 (Figure 1).

Statistical analysis

Statistical analysis was conducted with Matlab (The Mathworks, MA, USA).

Differences between controls and patients. Gaussian distribution of cord area measurements, DTI and magnetization transfer metrics were assessed with-in patients and controls using a χ 2 goodness of fi t ( p < 0.05). A two-way ANOVA was performed to assess cord atrophy in patients versus controls and to evaluate the effect of vertebral level (C4 to C7). Post hoc analyses used one-tailed t -test, given that ALS patients would exhibit decrease of spinal cord area. Difference in DTI and magnetization transfer metrics between patients and controls was assessed using a linear regression. Metrics tested were: FA, λ // , λ

^ , mean diffusivity and magnetization transfer

ratio in the lateral and dorsal ROIs. Given that cord atrophy could lead to greater partial volume effect in patients, cord area was added as a regressor.

To investigate whether posterior pathways were also hampered at an early stage of the disease, we selected a subset of 12 patients with early onset (mean age 52 � 9 years, median/maximum disease duration 9/12.6 months) and 12 age-matched con-trols (mean age 53 � 11 years) and re-ran linear regressions in these two sub-populations.

Correlations with clinical disability scores and TMS. Multiple regression analyses (forward-stepwise algo-rithm) were performed to fi nd the best predictor of

Figure 1. Regions of interest for quantifying metrics in the spinal cord. ROIs were anatomically defi ned on the mean diffusion-weighted data (for DTI metrics) and on the T1-weighted image (for magnetization transfer). ROIs were selected in the lateral segment (red) to include most of the lateral CST and in the dorsal segment (green) to include most of the dorsal columns. D: Dorsal; V: Ventral; R: Right; L: Left. The number of selected voxels in the lateral segments from T2 to C2 were: 19.6 � / � 3.7, 21.9 � / � 4.0, 29.7 � / � 5.5, 38.0 � / � 6.9, 42.6 � / � 7.1, 40.2 � / � 5.8, 35.8 � / � 6.1, 30.8 � / � 4.6 (mean � / � SD across subjects). The number of selected voxels in the dorsal segments were: 9.8 � / � 2.2, 9.1 � / � 2.5, 10.1 � / � 2.5, 12.5 � / � 2.1, 13.7 � / � 2.6, 14.7 � / � 2.5, 14.4 � / � 2.9, 13.4 � / � 2.7.

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Spinal sensory impairment and atrophy in ALS 33

clinical disability and CST impairment. Dependent variables were: ALSFRS-R, FMT, central conduc-tion time and peripheral conduction time. Predictors were: age, FA, λ // , λ ̂ , mean diffusivity, magnetiza-tion transfer ratio and cord area. The probability for a predictor to enter the stepwise model was based on a Fisher ' s test, with a p -value set to 0.05.

Relationship between atrophy level and muscle defi cits. The specifi city of atrophy at a given vertebral level in relation to the muscle MRC score and TMS was tested using a stepwise linear regression model. The hypothesis was that defi cit of the deltoid muscle (C5 spinal level, equivalent C4 vertebral level) is associ-ated with atrophy at C4 vertebral level and that defi -cit of the abductor pollicis brevis or adductor digiti minimi (C8 spinal level, equivalent C7 vertebral level) is associated with atrophy at C7 vertebral level. Dependent variables were muscle scores (deltoid and abductor pollicis brevis) and the MEP ampli-tude (adductor digiti minimi) and predictors were cord area at levels C4, C5, C6 and C7.

Dying-back versus dying-forward hypothesis. FA and mag-netization transfer ratio were calculated from C2 to T2, in the lateral ROI (14). A relationship between MRI metrics and the vertebral level was tested using Spear-man ' s correlation. For magnetization transfer ratio, since we disposed of 52 slices – as opposed to eight slic-es for DTI � we computed the nominal vertebral level by interpolating the C3-T1 region into 52 samples.

Results

Comparison patients/controls

Table II lists mean cord area for controls and patients from C4 to C7. Two-way ANOVA demonstrated a signifi cant difference between patients and age-matched controls (reduction of cord area in patients, F � 27.82, p < 10 -6 ) and between vertebral levels ( F � 40.67, p < 10 -6 ). The interaction between popu-lation and vertebral level was not signifi cant ( F � 0.23, p � 0.8). Two-sample one-tailed t -tests of level-wise analysis revealed signifi cant difference of cord area at C4, C5 and C6 levels (Bonferroni-corrected).

Figure 2A shows plots of MRI metrics in the lateral segments. Linear regression between the two populations demonstrated decrease in FA (beta � 0.53, p � 2 � 10 -6 ), increase in λ ̂ (beta � 7.7 � 10 -4 ,

p � 0.005) and decrease in magnetization transfer ratio (beta � 30.4, p � 0.008) in patients. No signifi cant dif-ference was detected for λ // (beta � 1.9 � 10 -3 , p � 0.1) and mean diffusivity (beta � 1.1 � 10 -3 , p � 0.13).

Figure 2B shows MRI metrics measured in the dorsal segment (ascending fi bres). Linear regression demonstrated signifi cant differences in the dorsal col-umn with a decrease in FA (beta � 0.56, p � 0.002) and increase in λ ̂ (beta � 7.8 � 10 -4 , p � 0.02) in patients. The two patients with modifi ed SOD1 mutation-linked familial ALS are circled in red in Fig-ure 2. These two patients exhibited particularly low FA compared to the other ALS patients. When remov-ing these two patients from the regression analysis, there was still a signifi cant difference between the two populations for FA (beta � 0.56, p � 0.005) and λ ̂ (beta � 8.1 � 10 -4 , p � 0.03). The 12 patients with early onset showed signifi cant difference in the dorsal column, but only for FA (beta � 0.65, p � 0.01).

Correlations with clinical disability and TMS

Results of stepwise linear regression revealed that FA measured in the lateral segments was the best predic-tor of the ALSFRS-R (beta � 45.54, StdErr � 20.59, p � 0.036) and FMT (beta � � 194.09, StdErr � 70.49, p � 0.012). No predictor was found for the peripheral conduction time and central conduction time. Figure 3 shows plots and linear line slope of FA versus ALS-FRS-R (R � 0.38, p � 0.04) and FMT (R � � 0.47, p � 0.02).

Cord atrophy and muscle defi cits

Table III shows the results of the stepwise regression analysis that tested the specifi city of local spinal cord atrophy to muscle score (deltoid and hand muscle). Spinal cord area at C4 vertebral level was asso ciated with the deltoid MRC score (beta � 0.15, StdErr � 0.06, p � 0.017). Spinal cord area at C7 vertebral level (equivalent C8 spinal level) was asso-ciated with abductor pollicis brevis MRC score (beta � 0.19, StdErr � 0.06, p � 0.005) and adductor digiti minimi MEP amplitude (beta � 0.08, StdErr � 0.04, p � 0.048).

Dying-back versus dying-forward hypotheses

Figure 4 shows the FA and magnetization transfer ratio (controls/patients) plotted against the nominal vertebral level. Spearman ' s correlation demonstrated signifi cant relationship for FA ( r � – 0.83, p � 0.015) and magnetization transfer ( r � – 0.51, p � 0.008). The negative correlation indicates a decrease of these metrics towards the more caudal segments.

Discussion

Multi-parametric MRI of the spinal cord in ALS patients demonstrated: 1) DTI and magnetization

Table II. Spinal cord atrophy. Mean � / � standard deviation of cord area (in mm 2 ) at four different vertebral levels, measured from the anatomical T2-weighted image. p -values resulting from one-tailed two-sample t -tests (Bonferroni corrected). Signifi cant values ( p < 0.05) are marked with ( * ).

C4 C5 C6 C7

Controls 81.8 � 8.0 81.7 � 8.5 78.5 � 8.6 65.8 � 7.7ALS 76.0 � 8.0 75.4 � 8.2 71.3 � 7.0 61.3 � 6.2 p -value 0.0321( * ) 0.0279( * ) 0.0048( * ) 0.0530

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Figure 2. Regression analysis between ALS patients and controls . A. Individual MRI metrics averaged in the lateral (CST) segments of the spinal cord in controls and patients. MTR stands for Magnetization Transfer Ratio. B. Individual plots of MRI metrics averaged in the dorsal columns. The mean is represented as a thick black line and the standard deviation as a light grey rectangle. Group differences were assessed using linear regression (normal distribution). Levels of signifi cance are indicated as: * p < 0.05, * * p < 0.005, * * * p < 0.0005. Patients with SOD1 gene are circled in red.

Figure 3. Correlations. A. Plots of ALSFRS-R versus FA. B . Plots of FMT versus FA. Correlation coeffi cients and p -values are derived from Pearson ' s correlation.

transfer abnormalities in the lateral (CST) and dor-sal (sensory) segments of the cervical spinal cord; 2) spinal cord atrophy associated with the correspond-ing muscle defi cit; and 3) correlations between FA and electrophysiological measurements (TMS) and functional impairment (ALSFRS-R).

Spinal cord biomarkers in ALS

In ALS, most neuroimaging studies were conducted in the brain (30) and provided non-invasive markers characterizing the involvement of the motor struc-tures classically involved (the CST) as well as

non-motor structures (sensory systems). In this study, spinal cord imaging provided additional infor-mation on the involvement of the lower motor neurons and central sensory pathways. Using multi-parametric MRI, we quantifi ed the degree of local spinal cord atrophy, which probably refl ects the degeneration of lower motor neurons (but not only, as discussed later), and the degree of white matter lesions using DTI and magnetization transfer, which refl ect spinal pathways abnormalities. Our results confi rm previous DTI studies (13,14) and add fur-ther evidence from magnetization transfer measure-ments. FA was decreased and radial diffusivity was

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Spinal sensory impairment and atrophy in ALS 35

Table III. Relationship between muscle defi cits and local cord atrophy. p -values (truncated at 2 digits) resulting from multiple regression analyses (stepwise model). Signifi cant values are marked with ( * ). The best predictor of deltoid defi cit is spinal cord atrophy at C4 vertebral level. The best predictor of abductor pollicis brevis defi cit and adductor digiti minimi MEP amplitude (from TMS measurement) is spinal cord atrophy at C7 vertebral level.

Dependent variable

Predictors: Atrophy index

C4 C5 C6 C7

Deltoid (MRC) 0.01( * ) 0.25 0.41 0.82Abductor pollicis brevis

(MRC)0.86 0.14 0.40 0.00( * )

Adductor digiti minimi (TMS)

0.77 0.38 0.66 0.04( * )

Figure 4. ‘ Dying-back ’ hypothesis. A. FA ratio (control/patient) plotted versus the nominal vertebral level. The ratio increases toward the caudal direction, suggesting larger degeneration toward caudal segments. B. Magnetization transfer ratio (MTR, control/patient) versus the nominal vertebral level. Spearman ' s correlation assessed relationship between the two variables. The reason there are more samples in panel B compared to panel A is that MTR measurements are derived from the T1-weighted scans (with and without MT pulse). As opposed to the DTI scans that only have one slice per vertebra, the T1 scans have about 2 � 3 times more slices per vertebra.

increased, suggesting secondary demyelination (31). Relative preservation of axonal architecture might then explain the lack of MD and axial diffusivity changes.

Multiple regression analysis showed signifi cant correlations between FA measured in the lateral seg-ments and both ALSFRS-R and FMT. Correlations were positive for ALSFRS-R and negative for FMT, i.e. lower FA was associated with lower ALSFRS-R (i.e. larger functional defi cits) and higher electrical threshold to evoke muscular response with TMS. These results are in line with previous studies report-ing that FA in the CST correlates with disease severity (9,12,30). However, correlations with mag-netization transfer and with DTI metrics other than FA were non-signifi cant, which may be related to the somewhat lower sensitivity of these metrics compared to FA. This lack of correlation may have also been due to the use of the total ALSFRS-R score, which includes bulbar scores and may therefore be less spe-cifi c to spinal involvement. However, we performed the same analyses using the ALSFRS-R score with-out including the bulbar score and have not found any signifi cant MRI predictor. We explain these neg-ative results by the somewhat low specifi city of the

ALSFRS score and the relatively small population used here.

Despite the relatively high in-plane spatial resolu-tion used here (1 � 1 mm 2 ), partial volume effect was inevitably present in the lateral and dorsal ROI. The lateral ROI, which aimed at including most of the descending CST, probably included ascending spi-nothalamic and spinocerebellar tracts. It is possible that the degeneration of the spinocerebellar tract (see next section) partially contributed to MRI differences within the lateral ROI.

Abnormalities in the spinal sensory pathway

Previous studies have shown that neurodegeneration in ALS is not restricted to the primary motor system but also extends to sensory cortical areas (2 � 5) and to the spinocerebellar system (1,32). At the spinal level, convincing evidence for impairment of central sensory pathways came from autopsy studies (1,2). In patients with ALS, electrophysiological and path-ological fi ndings also indicate a pattern of sensory peripheral axonal loss that predominantly affects the large-calibre myelinated fi bres (2,6). Our study shows impairment in the dorsal columns, in patients within a year of onset of symptoms. However, none of our patients showed any clinical evidence of pro-prioceptive, or any other sensory, defi cit. The dis-crepancy between the clinical evaluation and the radiological fi ndings might arise from the fact that MRI is sensitive to abnormalities of a sub-type of fi bres related to the amyotrophy, i.e. type Ia fi bres originating from muscle spindles. Hence, it is pos-sible that only these fi bres degenerated and produced MRI abnormalities, whereas most large fi bres remained intact, explaining the absence of sensory symptoms in this cohort. An impairment of the sen-sory pathways, with a degeneration of the dorsal funiculus, has been detected in an animal model of SOD1-linked ALS (33), and can occur early in the course of the disease (34). Two recent studies in a

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36 J. Cohen-Adad et al.

mouse model of another motor neuron degenerative disease, spinal muscular atrophy, demonstrated an early impairment of proprioceptive inputs originat-ing from muscle spindles to motor neurons (35,36). Abnormal somatosensory evoked potentials have also been found in humans (37 � 41). All these fi nd-ings support the involvement of sensory neurons and their axonal projections in the central and peripheral nervous pathology of ALS, extending the established concept of a multi-system degenerative process.

In the present study, the two patients having a mutation of the SOD1 gene had a lower FA and magnetization transfer ratio in the dorsal columns compared to the other patients. Those two patients had a classical ALS phenotype and had a combina-tion of upper and lower motor neuron signs, without sensory signs or symptoms. Another DTI study showed different CST involvement in a group of disability-matched SOD1 patients (42). These obser-vations support that atypical sensory signs and symp-toms are more frequent in SOD1-linked ALS (43,44). Further studies must be conducted to confi rm/deny this possibility.

Dying-back versus dying-forward hypotheses

Previous studies investigated whether axonal degen-eration in ALS progresses anterogradely ( ‘ dying-forward ’ ) (45 � 48) or from the extremity to the cell bodies ( ‘ dying-back ’ hypothesis) (14,49 � 51). Here we investigated the pattern of CST degeneration along the cervical spinal cord and confi rmed the observation from Nair et al. (14), i.e. FA appeared to be lower at more caudal levels of the spinal cord, with additional evidence from magnetization transfer mea-surements. Our results are also consistent with the study of Iwata et al. (52), in which the greatest reduc-tion in FA occurred in the distal portions of the intracranial corticospinal tract of ALS patients.

Greater degeneration in more caudal segments of the cord could be explained by 1) neurotoxic events close to the lower motor neuron that invade termi-nals of the CST and induce a dying back reaction; or 2) a process affecting the cell body in the cortex that might lead to a degeneration of more distal terminals fi rst (53).

Cord atrophy and muscle defi cit

Atrophy at C5 spinal level was associated with del-toid defi cits and atrophy at C8 spinal level was asso-ciated with defi cits in hand muscles. Although atrophy may partly be explained by degeneration of white matter pathways (via degeneration of upper motor neurons), the specifi c association between C8 atrophy and MEP amplitude of the adductor digiti minimi provides a strong evidence for grey matter atrophy, given that MEP amplitude obtained after stimulation at the level of C8 nerve root refl ects impairment of lower motor neurons, not CST. Our

fi ndings suggest that regional spinal cord atrophy as measured using MRI is potentially a sensitive bio-marker of anterior horn cell degeneration in ALS. Specifi city for grey matter degeneration may come with higher spatial resolution.

Acknowledgements

We thank Kevin Nigaud, Alexandre Vignaud and Eric Bardinet for helping with the acquisition and Henrik Lundell for providing the code to measure the cord area. We are grateful for the contribution of the patients and their relatives. We thank the clini-cians of the Paris ALS Centre, Ga ë lle Bruneteau, Lucette Lacomblez, Marie-Violaine Lebouteux, Timothee Lenglet and Fran ç ois Salachas. We thank the reviewers for their very insightful comments. This study was supported by the Association Fran -ç aise contre les Myopathies (AFM), the Institut pour la Recherche sur la Moelle é pini è re et l'Enc é phale (IRME), the National MS Society and the Sensori-Motor Rehabilitation Research Team (SMRRT) of the Canadian Institute of Health Research.

Declaration of interest: The authors report no confl icts of interest. The authors alone are respon-sible for the content and writing of the paper.

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