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Abstract
Background Although neuropathological observations suggest that cerebellar cortex is a major site of demyelination in multiple sclerosis (MS), only a few MRI studies on cerebellar cortical pathology in MS are available.
Objective To analyse cerebellar cortical volume (CCV) and leucocortical lesions (CL) in MS, and their impact on clinical disability.
Methods The authors studied 125 patients divided into 38 Clinical Isolated Syndrome (CIS), 35 relapsing remitting multiple sclerosis (RRMS), 27 secondary progressive multiple sclerosis (SPMS) and 25 primary progressive multiple sclerosis, and 32 normal controls (NC). CCV and cerebellar CL number and volume were evaluated by means of Freesurfer software and Double Inversion Recovery, respectively.
Results Compared with NC (mean 113.2±2.6 cm3), the CCV was significantly reduced in CIS (105.4±2.2 cm3, p=0.018), RRMS (104.0±2.0 cm3, p=0.012), SPMS (98.8±2.0 cm3, p<0.001) and PPMS (100.6±2.2 cm3, p<0.001), even after age, gender and mean cortical volume correction. CL were observed in all patient groups and were an independent predictor of CCV and cerebellar dysfunction.
Discussion The authors confirm that the cerebellar cortex is severely and early affected by MS pathology. The monitoring of cerebellar cortical atrophy and CL may help to understand the mechanism underlying disability progression in MS.
- multiple sclerosis
- cerebellum
- grey matter pathology
- MRI
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Introduction
Although multiple sclerosis (MS) is still regarded as a typical white matter (WM) disease,1 several magnetic resonance (MR) imaging studies have demonstrated that grey matter (GM) atrophy is relevant since disease onset, significantly contributes to clinical disability2 3 and preferentially affects some brain regions.4 5
Cerebellar dysfunctions are common in MS, and neuropathological observations indicate that the cerebellar cortex is a major site of demyelination, especially in the primary progressive multiple sclerosis (PPMS) and the secondary progressive multiple sclerosis (SPMS) forms of the disease.6 Moreover, modifications in cerebellar–neocortical connectivity may influence adaptive changes in brain motor control in MS.7 Similar changes in the cerebellum and premotor cortex have been reported in healthy brain during motor learning, thus suggesting that common mechanisms may contribute to normal motor learning and motor recovery in MS.8–10
Only a few MR studies on atrophy of the cerebellar cortex have been performed in MS, and no data are available on the presence of cortical lesions in the cerebellum. The development of fully automated high-resolution software (Freesurfer) for regional cortical volume analysis and the availability of the Double Inversion Recovery (DIR) sequence11 gave us the possibility of studying cerebellar cortical atrophy and CL, and their relationship with cerebral WM and GM pathology, and with disability, in a large group of MS patients.
Methods
Study population
We studied 32 normal controls (NC)(22F/10M) and 125 MS patients (82F/43M) divided into four groups12 13: 38 Clinical Isolated Syndrome (CIS, 25F/13M), 35 Relapsing Remitting Multiple Sclerosis (RRMS, 24F/11M)), 27 SPMS (18F/9M) and 25 PPMS (15F/10M) (table 1). Among the 38 CIS, 20 had evidence of dissemination in space of lesions.12 13 Neurological evaluation was done by means of the Kurtzke's Expanded Disability Status Scale (EDSS).14 Forty-seven patients (34 RRMS/13 SPMS) were treated with immunomodulatory agents (24 IFNβ-1a, 11 IFNβ-1b, 12 GA), 24 (14 SPMS/10 PPMS) with mitoxantrone. The study was approved by the local Ethics Committee.
Demographic, clinical and magnetic resonance characteristics of the study population
Image acquisition
As described in detail elsewhere,15 the following MR sequences were acquired: (1) Fast fluid attenuated inversion recovery (FLAIR), (2) double inversion recovery (DIR) and (3) three anatomical 3D fast field echo (FFE) sequences.
Image analysis
WM lesions
After lesion identification on FLAIR, brain WM lesion volume (T2-WM-LV) was quantified using a semiautomated local thresholding technique based on the Fuzzy C-mean algorithm,15 16 as described elsewhere.17 Brainstem and cerebellar T2-WM-LV were analysed on DIR images with the same procedure.
Cerebellar leucocortical lesions
After CL identification, with particular attention to the possible artefacts, CL number and volume were quantified on DIR, with the same semiautomatic procedure used for brain WM lesions.17 Of course, considering the actual resolution of our images, the possibility that some cortical lesions extend in the juxtacortical white matter cannot be excluded. As a consequence, we prefer to refer to these lesions as leucocortical lesions (CL).
Cerebral and cerebellar cortical volume
Measurements of brain cortical volume (BCV), cerebellar cortical volume (CCV) and cerebellar white matter volume (CWMV) were performed using a completely automatic technique, FreeSurfer (version 4.04)18 19 on a data set of three anatomical 3D-FFE. The maps obtained were not restricted to the voxel resolution of the original data, and thus were capable of detecting submillimetric differences between groups.18 19 After Freesurfer analysis, all images were reviewed to guarantee the accuracy of cerebellar cortical–subcortical segmentation. Procedures for the measurement of cortical thickness have been validated against histological analysis20 and manual measurements.21 Freesurfer morphometric procedures have been demonstrated to show good test–retest reliability across scanner manufacturers and across field strengths.22
Statistical analysis
Between-group differences in demographic, clinical and MR variables were assessed using ANOVA with a Tukey test for multiple comparisons. Between-group differences in MR derived measures were assessed using an ANCOVA model with age and gender as a covariate, and with the Tukey test. The Pearson and Spearman coefficients were used to analyse the correlation among continuous and discrete variables. A stepwise linear regression analysis was performed to assess the relative contribution of age, disease duration, CV, CWMV, cerebellar CL and T2-WM-LV in predicting CCV and to assess the relative contribution of all demographic variables in predicting EDSS and cerebellum Functional System. Backward stepwise analyses were conducted (Wald statistic), with a p value for entry of 0.05 and a p value for removal of 0.1. All statistical analyses were performed using SPSS v.13 and R (SPSS, Chicago, Illinois).
Results
Cerebellar cortical volume
Following age and gender correction, a significant reduction in the CCV compared with NC was observed in CIS (p=0.018), RRMS (p=0.012), SPMS (p<0.001) and PPMS (p<0.001) (see table 1 for more details). CCV was found to be slightly higher in CIS without dissemination in space of the lesions (106.4±2.2 cm3, range 101.0 to 109.6, p=0.018) compared with CIS with dissemination in space of the lesions (104.2±2.2 cm3, range 101.4 to 108.8), but the difference was not significant (p=0.138). Following age, gender and BCV correction, the CCV still resulted significantly lower in CIS (mean 104.2±2.0 cm3, range 100.2 to 108.2, p=0.038), RRMS (mean 104.0±2.0 cm3, range 100.0 to 107.6, p=0.033), SPMS (mean 101.4±2.0 cm3, range 97.4 to 105.4, p=0.006) and PPMS (mean 102.6±2.0 cm3, range 98.4 to 106.8, p=0.019) compared with NC (mean 110.6±2.4 cm3, range 106.4 to 115.2). After correction for disease duration, a high reduction in CCV was observed in PPMS (99.0±2.2 cm3, range 97.4 to 101.2, p=0.01), SPMS (101.2±2.0 cm3, range 99.0 to 102.8, p=0.022) and RRMS (101.8±2.0 cm3, range 98.8 to 103.2, p=0.031), but not in CIS patients (106.2±2.0 cm3, range 101.6 to 109.2, p=0.151), compared with NC (111.4±2.6 cm3, range 107.6 to 116.6). No significant difference was observed between PPMS and SPMS (p=0.071) and between PPMS and RRMS (p=0.078).
Cerebellar cortical lesions
At least one cerebellar CL was found in 13 (34%) CIS, 14 (40%) RRMS, 18 (67%) SPMS and 15 (60%) PPMS (table 1 and figure 1). Leucocortical lesions were equally distributed in the cerebellum without any preferential site. SPMS and PPMS had a significantly higher number and volume of cerebellar CL than RRMS and CIS (p<0.001 for both comparisons). No difference was observed between RRMS and CIS (p=0.121 for number, p=0.098 for volume).
Axial double inversion recovery images of the brain from a patient with relapsing remitting multiple sclerosis (A, C), secondary progressive multiple sclerosis (B) and primary progressive multiple sclerosis (D). White arrows indicate cerebellar cortical lesions that are clearly hyperintense compared with cerebellar grey matter.
Correlation between clinical and MR variables
The CCV strongly correlated with BCV (r=0.540, p<0.001) and with cerebellar CL volume (r=−0.618, p<0.001), while a mild/modest correlation with cerebellar CL number (r=−0.332, p=0.001), brain T2-WM-LV (r=−0.266, p=0.005), cerebellar T2-WM-LV (r=−0.370, p<0.001) and brainstem T2-WM-LV (r=−0.201, p=0.009) was observed. Moreover, CCV strongly correlated with cerebellar FS score (r=−0.644, p<0.001) and moderately with EDSS (r=−0.371, p<0.001), whereas cerebellar CL volume moderately correlated with both cerebellar FS score (r=0.461, p<0.001) and EDSS (r=0.401, p<0.001).
Multivariate analysis
A significant contribution from BCV (β=0.471, p<0.001), age (β=−0.323, p=0.030) and cerebellar CL volume (β=−0.423, p<0.001) as independent predictors of the CCV was found. The CCV (β=−0.601, p<0.001) and the cerebellar CL volume (β=0.512, p<0.001) were found to be the best predictors of cerebellar disability. BCV (β=−0.351, p=0.002), CCV (β=−0.339, p=0.011) and T2-WM-LV (β=0.308, p=0.019) were identified as independent predictors of EDSS.
Discussion
Neuropathological examination of post-mortem brain tissue has revealed that cerebellum is a major predilection site of demyelination in MS, especially in patients with PPMS and SPMS.6 The few available MR studies have demonstrated a significant reduction in cerebellar volume (GM+WM) in MS patients,23 and a marked reduction in CCV in SPMS compared with benign MS24 and CIS.25 In experimental autoimmune encephalomyelitis, the animal model of MS, CCV significantly decreases 48–56 days after disease induction.26 All these observations prompted us to investigate cerebellum cortical pathology in a large number of MS patients with new MR methodologies.
We observed: (1) a widespread pathology (atrophy and CL) in the cerebellar cortex in all MS groups, (2) a low correlation between CCV and WM lesion load and (3) the absence of a significant cerebellar WM atrophy in the early phases of the disease. All together, these findings confirm in vivo the existence of a diffuse and primary GM pathology involving the cerebellar cortex.6
The significant reduction in CCV in all MS groups and its high correlation with the cerebellar FS are particularly interesting, since these confirm the neuropathological findings6 27 and indicate that cortical neurodegeneration is very pronounced in the cerebellum, especially in PPMS patients, which had the lowest CCV values, despite a disease duration similar to that of the other patient groups. Whether CCV contributes to the peculiar clinical course of PPMS deserves further investigation. Indeed, the cerebellum seems to play a major role in brain plasticity, as suggested by the increased connectivity between the cerebellar cortex (the input region of the corticoponto-cerebellar projection system) and the controlateral premotor cortex observed in MS patients.7
Some differences between in vivo and ex vivo6 CL images are probably due to patient selection, in terms of age, disease duration, and disease severity and type. Histological material is often derived from patients with particularly severe and/or long-lasting disease, in which the widespread cortical demyelination may impair the clear identification of CL subtypes. On the other hand, only a minority of CL can be demonstrated by DIR.28
Finally, we would like to stress the results of the multivariate analysis that revealed that cerebellar CL volume is an independent predictor of both CCV and cerebellar disability, thus suggesting that cerebellar CL may play a major role in determining cerebellar cortical atrophy and cerebellar dysfunction. A longitudinal study is currently in progress to clarify the effects of cerebellar CL and atrophy on long-term disability.
References
Footnotes
Competing interests None.
Ethics approval Ethics approval was provided by the University Hosptial of Padua.
Patient consent Obtained.
Provenance and peer review Not commissioned; externally peer reviewed.