MS - Abstracts 9c-2g1

The elimination of inflammatory cells within the Central Nervous System (CNS) by Apoptosis plays an important role in protecting the CNS from Immune-mediated damage.

T-Cells, B-Cells, Macrophages, and Microglia all undergo Apoptosis in the CNS. The Apoptotic elimination of CNS-reactive T-Cells is particularly important, as these cells can recruit and activate other inflammatory cells.

T-Cell Apoptosis contributes to the resolution of CNS inflammation and clinical recovery from attacks of Experimental AutoImmune EncephaloMyelitis (EAE), an animal model of the DeMyelinating disease Multiple Sclerosis (MS).

T-Cell Apoptosis in the CNS in EAE occurs in both an Antigen-specific and an Antigen-nonspecific manner.

In Antigen-specific T-Cell Apoptosis, it is proposed that T-Cells that recognize their Antigen in the CNS, such as CNS-reactive T-Cells, are deleted by the process of activation-induced Apoptosis after activation of the T-Cell Receptor.

This may result from the Ligation of T-Cell death receptors (such as CD95 (Fas) or Tumor Necrosis Factor (TNF) Receptor 1) by CD95 Ligand (CD95L) or TNF expressed by the same T-Cell or possibly by Microglia, Astrocytes or Neurons.

Inadequate costimulation of the T-Cell by Antigen-Presenting Glial Cells may render T-Cells susceptible to activation-induced Apoptosis.

T-Cells expressing CD95 may also die in an Antigen-nonspecific manner after interacting with Glial Cells expressing CD95L.

Other mechanisms for Antigen-nonspecific T-Cell Apoptosis include the endogenous release of GlucoCorticoSteroids, deprivation of InterLeukin-2, and the release of Nitric Oxide by Macrophages or Glia.

Apoptosis of AutoReactive T-Cells in the CNS is likely to be important in preventing the development of AutoImmune CNS diseases such as MS.

Copyright 2001 Wiley-Liss, Inc.

Disruption of the Tight Junctions (TJs) of the Blood-Brain Barrier (BBB) is a hallmark of many CNS pathologies, including Stroke, HIV Encephalitis, Alzheimer's Disease, Multiple Sclerosis and Bacterial Meningitis.

Furthermore, systemic-derived inflammation has recently been shown to cause BBB tight junctional disruption and increased ParaCellular permeability.

The BBB is capable of rapid modulation in response to physiological stimuli at the CytoSkeletal level, which enables it to protect the Brain Parenchyma and maintain a homeostatic environment.

By allowing the 'loosening' of TJs and an increase in ParaCellular permeability, the BBB is able to 'bend without breaking'; thereby, maintaining structural integrity.

Objectives
We aimed to investigate associations between NeuroPsychological indices and normalized volumes of SupraTentorial structures, and the area of the Corpus Callosum.

Materials And Methods
We studied 40 patients with Clinically Definite MS, using 3D-acquired MRI (MPRAGE, Magnetization Prepared Rapid Acquisition Gradient Echo) and stereology.

Subjects underwent a NeuroPsychological battery interrogating multiple Cognitive domains, from which a global Cognitive Index Score (CIS) was derived.

Results
White Matter volumes were significantly correlated with CIS (rho=-0.59, P<0.0001) and with many of the individual Cognitive tests.

CIS was also significantly correlated with the Corpus Callosal area (rho=-0.49, P<0.002). Gray Matter volumes did not significantly correlate with any Cognitive test.

Conclusions
These volume/function relationships presumably reflect the effects of SubCortical Axonal and Myelin loss on the Neural Networks that subserve Cognition.

If serial MRI volume estimations can index accumulating Cognitive deficits, this simple technique may be useful in therapeutic trials.

Sixty eight patients with verified Multiple Sclerosis (MS) (mean EDSS score 3.1 +/- 1.0) and 50 healthy donors have been investigated.

Thirty five patients had Relapsing/Remitting, 25 - Secondary/Progressive, 8 - Primary/Progressive course.

The remission was in 38, decompensation - in 20, relapse - in 10 patients.

Lymphocyte subpopulations were investigated using MonoClonal AntiBodies (Moscow) to the following Antigens: CD3⁺ (Naive T-Lymphocytes), CD4⁺ (T-Helpers), CD8⁺ (CytoToxic Lymphocytes), CD20⁺ (T-Supressors), CD25⁺ (IL-2 Receptor), CD16⁺ (Natural Killers), CD95⁺ (activated T-Cells ready to Apoptosis).

Cytokines and Tumor Necrosis Factor-alpha (TNF-) levels were measured using ELISA test. HLA Antigens were investigated by standard LymphoCytoToxic test.

In MS we found a fall of CD3⁺, CD4⁺, CD8⁺, CD20⁺ and CD16⁺, but an increase of CD4⁺/CD8⁺, CD95⁺, CD25⁺. The CD95⁺ level correlated with CD4⁺, CD4⁺/CD8⁺ and CD16⁺.

In MS spontaneous IL-2, IL-6, IL-8 and TNF- production was raised, and stimulated IL-6 and IL-8 secretion was reduced. IL-4, IL-6, IL-8, TNF- and IL-1 beta Serum production in vivo was elevated.

We found an increase of CD3⁺, CD4⁺, CD16⁺, CD25⁺, but a decrease of IL-1 (p < 0.01) spontaneous production and IL-6, IL-8, TNF- stimulated secretion in DR2⁺ MS patients, comparing to DR2^- patients and controls.

In DR2^- patients as compared to DR2⁺ patients and controls, all Lymphocyte subpopulations levels, especially CD8⁺ (p < 0.001) one, were decreased, but spontaneous IL-8 (p < 0.01) production was increased.

The data obtained indicate Lymphocyte Apoptosis activation, targeting promoted Lymphocyte destruction, and suggest T-Helper Type-1 reaction prevalence in MS.

July 1, 1997 was stipulated as the day for estimating the prevalence of Multiple Sclerosis within the city of Sao Paulo. The patients were identified via various sources, including associated universities and magnetic resonance services of the city of Sao Paulo.

The area covered by the study has a population of 9,380,000, mainly white and of European origin, with a large number of immigrants from Spain and Portugal.

The patients were classified in accordance with the criteria of Poser et al. (1983), and only those with defined Multiple Sclerosis were registered.

The study gave a prevalence of 15.0/10 (5%) inhabitants, or three times the value obtained in a similar study in 1990.

This increase reveals the larger number of cases encompassed by the study, and is attributed to the use of more detailed recording methods, improvements in diagnosis, and better conditions for treatment.

Objective
To study VesicoSphincteric Dysfunction in 108 patients with Multiple Sclerosis.

Methods
We reviewed the clinical records of 108 patients with Multiple Sclerosis and analyzed those with voiding symptoms ascribable to Multiple Sclerosis.

These patients underwent complete Urodynamic assessment and complementary tests according to their symptoms. The Blaivas classification was used for the clinical classification of Multiple Sclerosis.

Results
64 of the 108 patients presented voiding symptoms ascribable to Multiple Sclerosis (59.2%).

The clinical features presented as episodes in 75% and were progressive in 25% of the cases. In 6% of the patients, the voiding symptoms were the first symptoms of Multiple Sclerosis.

Urodynamic assessment showed Detrusor HyperReflexia in 73% of the patients, HypoReflexia in 14%, and 13% showed normal Urodynamics. All complications were infective; no patient showed Upper Urinary Tract complications.

Conclusions
VesicoSphincteric Dysfunction in Multiple Sclerosis is frequent. Most of the patients present Bladder HyperReflexia.

The Urological complications are usually infective. Involvement of the Upper Urinary Tract is rare.

Objective
The EtioPathoGenesis in AudioVestibular symptoms can be elusive, despite extensive differential diagnosis.

This article addresses the value of Magnetic Resonance Imaging (MRI) in analysis of the complete AudioVestibular pathway.

Study Design, Setting & Patients
Retrospective evaluation. Tertiary referral center. Consecutive sample of 354 patients (mean age 49 years, range 8 to 86 years) with AudioVestibular Disorders.

Intervention
Contrast-enhanced MRI of the head with thin-slice investigation of the Inner Ear, Internal Auditory Meatus, and CerebelloPontine Angle.

Main Outcome Measure
All MRIs were evaluated by experienced independent investigators.

Statistical analysis was performed using the Statistical Package of Social Sciences data analysis 9.0.

Results
MRI abnormalities were seen in 122 of 354 patients (34.5%). The MRIs revealed the following:

1. 4 pathologic conditions (1.1%) of
   • the Cochlea/Labyrinth
2. 23 abnormalities (6.5%)
   • at the Internal Auditory Meatus/CerebelloPontine Angle
3. 12 pathologic lesions (3.4%) that involved
   • the Central AudioVestibular Tract at the BrainStem
4. 78 MicroAngiopathic changes of the Brain (22%)
5. 3 focal HyperIntensities of the Brain that turned out to be
   • the first evidence of Multiple Sclerosis in 2 patients
6. Sarcoidosis in 1 patient
7. 1 Temporal Metastasis

Other pathologic conditions, such as Parotid Gland or Petrous Bone Apex Tumors, were unrelated to the AudioVestibular symptoms.

Conclusions
This study indicates that contrast-enhanced MRI can be used to assess a significant number of different pathologic conditions in patients with AudioVestibular Disorders.

Although Interferon-beta (IFN-ß) has shown a significant clinical benefit in Multiple Sclerosis (MS), its mechanism of action remains unclear.

We found that IFN-ß treatment of patients with MS resulted in a significant increase in Apoptotic Cell Death (measured by Annexin V staining and nuclear fragmentation) of Monocyte-derived Macrophages.

As compared with cells derived from patients before treatment. Stimulation of the cells with IFN-ß in vitro resulted in an even further increase of Annexin V binding, as well as increased Fas (CD95⁺, APO-1) expression.

However, no increased Fas expression, Apoptotic Monocytes, or MonoCytopenia were observed upon in vivo treatment.

This indicates that IFN-ß does not deliver a death signal to Monocytes but rather primes for subsequent Macrophage Apoptosis upon activation or differentiation.

Despite the strength of the association of Multiple Sclerosis (MS) and Human Leukocyte Antigen (HLA)-DR2, other Genetic elements could have a role in the pathophysiology of MS.

We investigated possible associations with PolyMorphic susceptibility Genes located within the HLA complex, i.e., Heat-Shock Protein (HSP) 70-1, HSP70-2, and HSP70-hom in Japanese patients with MS.

Furthermore, we analyzed the influence of HSP70 gene PolyMorphisms on the severity of the disease, clinical course, Magnetic Resonance Imaging findings, and OligoClonal Bands in the CerebroSpinal Fluid, and HLA in MS patients.

The results of the present study indicated that there were no significant differences in the distribution of all HSP70 Genotypes and Allele frequencies between Japanese MS patients and controls.

In MS patients, there were no associations between HSP70 Gene PolyMorphisms and the clinical data.

Moreover, there were no significant differences in HSP70 Genotype or Allele frequencies between MS patients positive for HLA-DRB1*1501 Alleles and matched controls.

Our data indicate that HSP70 Gene PolyMorphisms are not relevant in the susceptibility to or the severity of Japanese MS patients.

The AntiApoptosis protein FLIP (Fas-associated death domain-like InterLeukin-1ß-converting enzyme inhibitory protein) has been recently identified as a potent regulator of T-Lymphocyte susceptibility to Apoptosis.

In a prospective study, we evaluated the expression of FLIP and other Apoptosis regulatory proteins in ex vivo activated T-Lymphocytes from MS patients, before and serially after treatment with Interferon-ß.

We also investigated the long-term effects of Interferon-ß on T-Cell Apoptosis in a cross-sectional study of MS patients receiving chronic drug therapy.

Treatment with Interferon-ß reduced the expression of FLIP isoforms in activated T-Lymphocytes. This reduced expression correlated with augmented T-Cell susceptibility to Apoptosis and with clinical response to treatment.

In contrast, Interferon-ß therapy did not alter cellular expression of the AntiApoptotic protein Bcl-2. This downregulatory effect of Interferon-ß on cellular FLIP expression was maintained following long-term therapy.

Our findings suggest that Interferon-ß therapy exerts a regulatory effect on peripheral T-Lymphocytes through a ProApoptosis mechanism that involves the downregulation of cellular FLIP expression.

Little is known about the involvement of Cytokines in the pathogenesis of Primary/Progressive (PP) Multiple Sclerosis (MS).

We evaluated in this cross-sectional study whether IL-18, IL-12p35, IL-12p40, TNF- IFN-, IL-10, IL-4, TGF-ß, IL-12Rbeta1, and IL-12Rß2 mRNA expression in unstimulated white blood cells showed significant differences.

Between, Relapsing/Remitting (RR), Secondary/Progressive (SP) and PP MS patients, and healthy controls.

All clinical subtypes showed unique mRNA expression patterns as compared to the controls.

Both RR and SP patients displayed increased levels of IL-12p40, IL-18, and TGF-ß mRNA compared to controls, whereas PP patients showed only increased IL-18 mRNA levels.

Both in PP and SP patients, IFN- and IL-10 mRNA were decreased compared to RR patients and controls.

PP patients were unique in that they showed decreased IL-12Rß1 mRNA.

In conclusion, our data show that the assessment of Cytokine (Receptor) mRNA profiles is useful to discriminate between the different clinical subtypes.

And, suggest that different Cytokines are involved in the pathogenesis of PP MS as compared to RR and SP MS.

Objectives
We investigated whether therapy of Multiple Sclerosis (MS) with Glatiramer Acetate (GA) involves the modulation of programmed cell death (Apoptosis) in disease-relevant T-Helper Lymphocytes.

Material And Methods
Blood was drawn from 15 Relapsing/Remitting MS patients both before (baseline) as well as 6, 12, and 18 weeks after GA therapy and from 15 healthy controls.

Detection of Apoptosis was performed in response to in vitro stimulation with GA, Myelin Basic Protein or medium alone.

Results
T-Helper Lymphocytes from untreated MS patients displayed an overall increased Apoptosis susceptibility in vitro, compared to controls.

During subsequent GA therapy, Apoptosis vulnerability of these T-Cells in MS patients significantly declined under the initial baseline before treatment, and was finally equal in treated patients and controls.

GA itself had no direct Apoptosis-modulatory properties in vitro.

Conclusion
Our findings indicate that therapy of Multiple Sclerosis with Glatiramer Acetate presumably involves the compensation of altered Apoptosis in T-Helper Lymphocytes.