MS Abstracts 01a-2g3

Objective
Our aim was to determine if the resonance intensity of Choline-containing compounds (Cho) measured, using Proton Magnetic Resonance Spectroscopy (Spectroscopy), was increased in pre-lesional Normal Appearing White Matter (NAWM).

In patients with Multiple Sclerosis (MS) relative to NAWM that remained stable in subsequent scans.

Background
The Cho peak in MR Spectra is associated with membrane PhosphoLipids and increases in acute MS plaques, possibly even before the appearance of MRI-visible MS lesions.

Methods
Three combined proton MRI and Spectroscopy imaging examinations of the Corpus Callosum and adjacent PeriVentricular White Matter were performed on 12 MS patients at intervals of 6 months.

Proton Density (PD) images were visually matched across 3 time points and the Lesion Volume in each Voxel of the volume of interest was determined.

The Voxels were subdivided into four groups based on the presence or absence of lesion at baseline and change or no change in Lesion Volume on the subsequent scan.

Results
We found a significantly higher baseline Cho/Creatine (Cr) ratio in NAWM Voxels that displayed MRI visible lesions 6 months later than NAWM Voxels that remained unchanged (1.57 +/- 0.30 and 1.37 +/- 0.33, respectively, p < 0.001).

The 12-month interval data revealed similar pre-lesional elevated Cho/Cr, (1.51 +/- 0.29 versus 1.39 +/- 0.32, p = 0.009).

Voxels that contained lesion at baseline and increased in lesion volume at 6 months, also showed a significantly higher Cho/Cr ratio than those whose lesion volume did not change (1.60 +/- 0.32 and 1.49 +/- 0.36, respectively, p = 0.043).

Conclusions
The results of this study are consistent with focal pre-lesional Myelin membrane pathology in the NAWM at least 12 months before lesions become visible on conventional MRI.

This could reflect altered Myelin chemistry or the presence of inflammation as seen in Experimental Allergic Encephalomyelitis.

Adult Oligodendrocyte Precursor Cells (OPCs) make up around 5-8% of the Glial cell population in the CNS.

Their function in the undamaged CNS is largely unknown, but their Processes are in contact with Nodes of Ranvier and Synapses, suggesting a regulatory role at these structures.

The cells divide slowly, and constitute approximately 70% of cells labelled following a pulse injection of Bromodeoxyuridine. In the injured CNS the cells form a reactive Glial population that undergoes Hypertrophy and Mitosis, probably driven by a variety of Growth Factors and Cytokines.

In response to DeMyelination they divide and are thought to differentiate to provide new Oligodendrocytes to replace those that have been lost.

However, ReMyelination fails during the later stages of Multiple Sclerosis, and it is not clear whether this is as a result of a depletion of adult OPCs, inhibition within the Glial Scar, or damage to the Axons that prevents Myelination.

Adult OPCs are also activated and proliferate following other forms of CNS damage, such as mechanical injury, ExcitoToxicity and Viral infection.

The cells produce several of the Chondroitin Sulphate Proteoglycans that might inhibit Axon regeneration.

Myelin Basic Protein (MBP)-reactive T-Cells are potentially involved in the pathogenesis of Multiple Sclerosis (MS), and can be depleted by subcutaneous inoculations with irradiated autologous MBP-reactive T-Cells (T-Cell Vaccination).

This preliminary open label study was undertaken to evaluate whether depletion of MBP-reactive T-Cells would be clinically beneficial to patients with MS.

Fifty-four patients with Relapsing/Remitting (RR) MS (n=28) or Secondary/Progressive (SP) MS (n=26) were immunized with irradiated autologous MBP-reactive T-Cells and monitored for changes in rate of relapse, Expanded Disability Scale score (EDSS) and MRI lesion activity over a period of 24 months.

Depletion of MBP-reactive T-Cells correlated with a reduction (40%) in rate of relapse in RR-MS patients as compared with the pre-treatment rate in the same cohort.

However, the reduction in EDSS was minimal in RR-MS patients while the EDSS was slightly increased in SP-MSS patients over a period of 24 months.

Serial semi-quantitative MRI examinations suggest stabilization in lesion activity as compared with baseline MRI.

The findings suggest some potential clinical benefit of T-Cell Vaccination in MS and encourage further investigations to evaluate the treatment efficacy of T-Cell Vaccination in controlled trials.

Previous work has demonstrated potentially adaptive Cortical plasticity that increases with Brain Injury in patients with Multiple Sclerosis.

However, animal studies showing use-dependent changes in Motor Cortex organization suggest that functional changes also may occur in response to disability.

We therefore wished to test whether Brain Injury and disability lead to distinguishable patterns of activation with hand movement in patients with Multiple Sclerosis.

By employing a passive as well as an active movement task, we also wished to test whether these changes were independent of voluntary recruitment and thus more likely to reflect true functional reorganization.

Fourteen patients [Extended Disability Status Score (EDSS) 0-7.5] with Relapsing/Remitting Multiple Sclerosis were selected on the basis of pathology load and hand functional impairment for three study groups:

1. Low Diffuse Central Brain Injury (DCBI) as assessed from relative N-AcetylAspartate concentration (a marker of Axonal integrity) and normal hand function (n = 6)
2. Greater DCBI and normal hand function (n = 4)
3. Greater DCBI and impaired hand function (n = 4)

Functional MRI (fMRI) was used to map Brain activation with a four-finger and both one-finger passive and active flexion-extension movement tasks for the three groups.

Considering all the patients, we found increased activity in IpsiLateral PreMotor and IpsiLateral Motor Cortex (IMC) and in the IpsiLateral Inferior Parietal Lobe with increasing global disability (as assessed from the EDSS score).

These changes appear to define true functional reorganization, as fMRI activations in IMC (r = 0.87, P < 0.001) and in the ContraLateral Motor Cortex (r = 0.67, P < 0.007) were highly correlated between active and passive single finger movements.

We attempted to disambiguate any distinct effects of disability and Brain injury by direct contrasts between patients differing predominantly in one or the other.

To make these contrasts as powerful as possible, we used impairment of finger tapping as a measure of disability specific to the hand tested.

A direct contrast of patients matched for DCBI, but differing in hand disability (group 3 - group 2) showed greater BiLateral Primary and Secondary Somatosensory Cortex activation with greater disability alone.

A contrast matched for hand disability, but differing in DCBI (group 2 - group 1) showed a different pattern of changes with relative IpsiLateral PreMotor Cortex and BiLateral Supplementary Motor Area activity.

We conclude that the pattern of Brain activity with finger movements changes both with increasing DCBI and with hand disability in patients with Multiple Sclerosis, and that these changes are distinct.

Those related directly to disability may reflect responses to altered patterns of use. As injury- and disability-related activation changes are found even with passive finger movements, they may reflect true Brain reorganization.

We demonstrate here the feasibility of Antigen-specifically redirecting T-Cells against AutoReactive T-Lymphocytes and thereby treating a model AutoImmune Disease.

We created and transgenically expressed on T-Cells a heterodimeric chimeric receptor that genetically links an AutoAntigenic Peptide, its restricting MHC, and the signal transduction domain of the T-Cell Receptor (TCR) zeta-chain.

Engagement of the chimeric receptor by the TCR of AutoReactive T-Cells activated the receptor-modified T-Cells in vitro and in vivo, inducing proliferation and CytoLysis.

Adoptively transferred receptor-modified T-Cells prevented and treated a model AutoImmune Disease, Experimental Allergic Encephalomyelitis (EAE), even after Epitope spreading had diversified the AutoAntigenic response.

Treatment reduced disease severity and increased survival of affected animals, and was durable for >75 days.

The receptor-modified cells acted both by strongly attenuating T-Cell response to AutoAntigen as well as by shifting the residual response from an ImmunoPathologic Th1 to a protective Th2 format.

Multiple Sclerosis seems to be an AutoImmune Disease of unknown Etiology affecting the White Matter of the CNS. It is generally accepted that the AutoImmune Response is directed against specific components of Myelin.

We show here that Proteasome, a ubiquitous Protease complex composed of 14 different subunits, is a target for AutoAntibodies (IgG and IgM classes) present in the Serum (66%, 73 out of 110) and in the CSF (61%, 16 out of 26) of patients with Multiple Sclerosis.

Using recombinant Proteasomal subunits we demonstrate the presence of specific AutoAntibodies against subunits C2, C8, C9 and C5 in Multiple Sclerosis patients.

Recombinant C2 constructs allow us to localize an ImmunoDominant AutoEpitope recognized by the Sera of Multiple Sclerosis patients within the C-terminal of C2 Proteasomal subunit (251-Depaekadepmeh-263).

In addition, two constructs of the recombinant Proteasomal subunits C2 and C8 were also used to study the proliferation of peripheral blood MonoNuclear Cells from Multiple Sclerosis patients; 12 out of 30 (40%) Multiple Sclerosis patients show positive proliferation with one or both of these recombinant subunits.

The high prevalence of AntiProteasome AutoAntibodies in Multiple Sclerosis Sera compared with Sera from patients with other chronic inflammatory conditions:

1. Systemic Lupus Erythematosus (35%, 35 out of 100)
2. primary Sjogren's Syndrome (16%, 5 out of 31)
3. Vasculitis (0 out of 20)
4. Sarcoidosis (7%, 1 out of 13)
5. Behcet's Disease (19%, 4 out of 21)

Suggest that Humoral AutoReactivity to Proteasome could be a useful test in Multiple Sclerosis patients that may be of help in the diagnosis and/or progression of this chronic inflammatory disease.

Finally, these results suggest that some global abnormality in B and/or T-Cell function is also involved in the chronic inflammatory response observed in Multiple Sclerosis patients, as it is frequently observed in other human organ-specific AutoImmune Diseases.

A strong association exists between Multiple Sclerosis (MS) and the DRB1*1501 haplotype, in most populations. Linkage of Multiple Sclerosis (MS) with the MHC or HLA region on Chromosome 6p21 has previously been observed in DRB1*1501 positive MS families.

A group of 13 Israeli multiplex MS families with a very low frequency of DRB1*1501 haplotype were examined in this study. Association and a linkage test were performed in order to identify a non-DRB1*1501 effect of HLA on susceptibility for MS.

MS multiplex families and healthy controls were molecularly typed for six highly polymorphic markers located within the MHC region: DRB1, DQA1 and DQB1, BAT-2, MIB and D6S248.

Data analyzes included:
1. An association study comparing the patient group with both healthy relative, and healthy control groups
2. Transmission test for linkage disequilibrium (TDT) of the MS-associated alleles in the multiplex families

3. Multipoint non-parametric linkage (NPL) and parametric LOD score analyzes using the GENEHUNTER program

The DRB1*1303 allele was significantly more frequent among the MS patients. There was a trend towards transmission disequilibrium of DRB1*1303, but was not statistically significant.

Allele sharing and LOD score analyzes revealed no evidence for linkage. The high frequency of DRB1*1303 observed in our family patients provides evidence to support the association with this allele that previously described in sporadic non-Ashkenazi MS patients.

Thus, DRB1*1303 may serve as genetic risk factor for MS. Our study exemplifies the genetic heterogeneity in MS as there is a genetic effect of HLA on MS susceptibility in our low frequency DRB1*1501 patients.

Conventional MRI (cMRI) is limited in its ability to provide specific information about pathology in MS.

Measures commonly derived from cMRI include T₂ lesions, T₁-enhanced lesions, Atrophy, and possibly T₁-HypoIntense lesions, which have been extensively investigated in many clinical trials.

Better MRI measures are needed to advance our understanding of MS and design ideal clinical trials. This article reviews the strengths and weaknesses of the major MRI-based methods used to monitor MS evolution and submits that

1) metrics derived from Magnetization Transfer MRI, Diffusion-weighted MRI, and Proton MRS should be implemented to achieve reliable specific in vivo quantification of MS pathology;

2) targeted multiparametric MRI protocols rather than generic application of cMRI should be used in all possible clinical circumstances and trials; and

3) reproducible quantitative MR measures should ideally be used for the assessment of patients but are essential for clinical trials.

The clinical effects of Glatiramer Acetate (GA), an approved therapy for Multiple Sclerosis, are thought to be largely mediated by a T-Helper 1 (Th1) to T-Helper 2 (Th2) shift of GA-reactive T-Lymphocytes.

Current theories propose that activated GA-reactive Th2 cells penetrate the CNS, release AntiInflammatory Cytokines such as InterLeukin (IL-4), IL-5, and IL-10.

And thus inhibit neighboring inflammatory cells by a mechanism termed 'Bystander Suppression'.

We demonstrate that both GA-specific Th2 and Th1 Cells produce the NeuroTrophin Brain-Derived NeuroTrophic Factor (BDNF).

As the signal-transducing receptor for BDNF, the full-length 145 Tyrosine Kinase Receptor (trk) B, is expressed in Multiple Sclerosis lesions.

It is likely that the BDNF secreted by GA-reactive Th2 and Th1 has NeuroTrophic effects in the Multiple Sclerosis target tissue.

This may be an additional mechanism of action of GA, and may be relevant for therapies with altered Peptide Ligands in general.

To demonstrate that GA-reactive T-Cells produce BDNF, we selected four GA-specific, long-term T-Cell lines (TCLs).

Which were characterized according to their Cytokine profile by IntraCellular double-fluorescence flow cytometry.

Three TCLs (isolated from a normal subject) had the phenotypes Th1, Th1/Th0, and Th0; the fourth, derived from a GA-treated patient, had the phenotype Th2.

To demonstrate BDNF production, we used a combination of RT-PCR (Reverse Transcription-Polymerase Chain Reaction) and two specially designed techniques for BDNF protein detection:

One was based on ELISA (Enzyme-Linked Immunosorbent Assay) of supernatants from co-cultures of GA-specific TCLs plus GA-pulsed Antigen-Presenting Cells.

And, the other on the direct IntraCellular staining of BDNF in individual T-Cells and flow cytometric analysis.

The different assays and different TCLs yielded similar, consistent results.

All four GA-specific T-Cell lines, representing the major different TH phenotypes, could be stimulated to produce BDNF.

Although NeuroPhysiologic doctrine has traditionally referred to "the" voltage-gated Sodium Channel, it is now clear that there are at least nine Genes that encode molecularly and physiologically distinct Sodium Channels.

Mutations of Sodium Channel genes provide a basis for Genetic ChanneloPathies.

Dysregulated expression of Sodium Channels due to alterations in activity of nonmutated channel genes, on the other hand, can produce acquired ChanneloPathies.

Two examples of Acquired ChanneloPathies are discussed in this article.

Recent research has established that Peripheral Nerve Injury can provoke an Acquired ChanneloPathy in Spinal Sensory Neurons:

1. Axonal transection triggers the turning-off of some previously active Sodium Channel Genes
2. The turning-on of at least one previously silent Sodium Channel Gene, a set of molecular changes that can result in HyperExcitability of these cells

Emerging evidence also suggests that an Acquired ChanneloPathy.

Characterized by abnormal expression of Sensory Neuron specific Sodium Channels that can alter Impulse trafficking within Purkinje Cells, may contribute to the PathoPhysiology of MS.

Subtype-specific drugs that selectively modulate various types of channels probably will soon be developed.

The Acquired ChanneloPathies associated with Nerve injury and MS may thus represent prototype disorders that present therapeutic opportunities.

Background
The authors have recently described that Intrathecal IgM Synthesis (ITMS) correlates with a higher disability in patients with Clinically Definite MS (CDMS).

Objective
To follow-up a group of patients with MS in the initial stages of the disease to evaluate if the presence of ITMS correlates with a worse evolution.

Methods
OligoClonal IgM Bands were performed in 22 patients with MS with a mean of 1.14 months of evolution.

Patients were followed for a period ranging from 6 to 36 months (mean, 21.4 months). During follow-up, time to conversion to CDMS, number of relapses, and changes in Expanded Disability Status Scale (EDSS) score were evaluated.

Results
Patients were divided into two groups according to the presence (Group 1, 10 patients) or absence (Group 2, 12 patients) of ITMS.

No clinical differences were observed between the groups at inclusion in the study.

During the follow-up, the probability of conversion to CDMS was greater in Group 1 (90% of the patients had converted to CDMS after 8 months of follow-up) than in Group 2 (51% of patients had converted to CDMS after 36 months of follow-up) (p = 0.0001).

Patients from Group 1 had more relapses (mean, 2.0) than those from Group 2 (mean, 0.58) (p = 0.02). At the end of the study, patients from Group 1 had higher EDSS scores (mean, 1.70) than those from Group 2 (mean, 0.79) (p = 0.02).

Conclusion
The presence of OligoClonal IgM Bands in CSF can be a prognostic marker in the early phases of MS.

Multiple Sclerosis is still regarded primarily as a disease of the White Matter. However, recent evidence suggests that there may be significant involvement of Gray Matter.

Here, we have used Magnetic Resonance Imaging and Magnetic Resonance Spectroscopy in vivo and HistoPathology postmortem to estimate Thalamic Neuronal loss in patients with Multiple Sclerosis.

Our results show that Neuronal loss in Multiple Sclerosis can be substantial (30-35% reduction).

We conclude that a NeuroDegenerative pathology may make a major contribution to the genesis of symptoms in Multiple Sclerosis.