Glia Cells In Multiple Sclerosis

The purpose of this study was to examine the roles played by Astrocytes in a case of rapidly Progressive Multiple Sclerosis (MS).

Within early-active and active lesions, HyperTrophic Astrocytes played an important role in lesion pathology through the Phagocytosis of Myelin and Axonal debris and through the internalization of other Glial Cells, including Astrocytes.

In addition to this critical role, HyperTrophic Astrocytes, in areas that lack significant inflammation (within the adjacent Normal-Appearing White Matter).

And, within late ReMyelinating lesions were found to be active in Myelin and Axonal debris Phagocytosis with no evidence of cellular internalization.

HyperTrophic Astrocytes therefore not only play an important role in the pathogenesis of MS lesions but also exert a continued deleterious effect upon tissue in the absence of significant inflammation.

In addition, we found evidence for a significant population of Vimentin-positive, Glial Fibrillary Acidic Protein (GFAP)-negative, bipolar, Astrocyte Precursors within the late ReMyelinating lesions.

Their significance is not known but a possible role may include their participation in the successful ReMyelination of the lesion.

Disease progression in Multiple Sclerosis occurs within the interface of Glial activation and Gliosis.

This study aimed to investigate the relationship between biomarkers of different Glial Cell responses to:

1. Disease dynamics and the clinical subtypes of Multiple Sclerosis
2. Disability
3. Cross-validate these findings in a post-mortem study

To address the first goal, 51 patients with Multiple Sclerosis [20 Relapsing/Remitting (RR), 21 Secondary/Progressive (SP) and 10 Primary/Progressive (PP)] and 51 Neurological control patients were included.

Disability was assessed using the Ambulation Index (AI), the Expanded Disability Status Scale score (EDSS) and the 9-Hole PEG Test (9-HPT). Patients underwent Lumbar Puncture within 7 days of clinical assessment.

Post-mortem Brain tissue (12 Multiple Sclerosis and eight control patients) was classified Histologically and adjacent sites were homogenized for protein analysis.

S100B, Ferritin and Glial-Fibrillary Acidic Protein (GFAP) were quantified in CSF and Brain-tissue homogenate by ELISA (Enzyme-Linked Immunosorbent Assay) techniques developed in-house.

There was a significant trend for increasing S100B levels from PP to SP to RR Multiple Sclerosis (P < 0.05).

S100B was significantly higher in RR Multiple Sclerosis than in control patients (P < 0.01), while Ferritin levels were significantly higher in SP Multiple Sclerosis than in control patients (P < 0.01).

The S100B : Ferritin ratio discriminated patients with RR Multiple Sclerosis from SP, PP or control patients (P < 0.05, P < 0.01 and P < 0.01, respectively).

Multiple Sclerosis patients with poor Ambulation (AI >/=7) or severe disability (EDSS >6.5) had significantly higher CSF GFAP levels than less disabled Multiple Sclerosis or control patients (P < 0.01 and P < 0.001, respectively).

There was a correlation between GFAP levels and Ambulation in SP Multiple Sclerosis (r = 0.57, P < 0.01), and between S100B level and the 9-HPT in PP Multiple Sclerosis patients (r = -0.85, P < 0.01).

The post-mortem study showed significantly higher S100B levels in the acute than in the subacute plaques (P < 0.01), while Ferritin levels were elevated in all Multiple Sclerosis lesion stages.

Both GFAP and S100B levels were significantly higher in the Cortex of Multiple Sclerosis than in control Brain homogenate (P < 0.001 and P < 0.05, respectively).

We found that S100B is a good marker for the Relapsing phase of the disease (confirmed by post-mortem observation) as opposed to Ferritin, which is elevated throughout the entire course.

GFAP correlated with Disability Scales and may therefore be a marker for irreversible damage.

The results of this study have broad implications for finding new and sensitive outcome measures for treatment trials that aim to delay the development of Disability. They may also be considered in future classifications of Multiple Sclerosis patients.

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.

During the past decade, mechanisms involved in the Immune surveillance of the Central Nervous System (CNS) have moved to the forefront of NeuroPathological research.

Mainly because of the recognition that most Neurological Disorders involve activation and, possibly, dysregulation of Microglia, the intrinsic Macrophages of the CNS.

Increasing evidence indicates that, in addition to their well-established Phagocytic function, Microglia may also participate in the regulation of non specific inflammation as well as Adaptive Immune Responses.

This article focuses on the signals regulating Microglia Innate Immune Functions, the role of Microglia in Antigen presentation, and their possible involvement in the development of CNS ImmunoPathology.

Copyright 2001 Wiley-Liss, Inc.

Recent discoveries suggest that the resident cells of the Central Nervous System (CNS) the Nerve Cells and Glia, play a more Immunologically active role than was previously assumed.

NeuroGlial communication is of central interest in virtually all types of pathological conditions that affect the Brain and several features of the activation that results from Nerve Cell damage resemble the type of Innate Immune Reactions that occur in other parts of the body.

In particular, the characteristics of the activation of these CNS cells will affect both the interaction with cells of the Immune System as well as processes related to NeuroDegeneration and Regeneration.

We here review data regarding 3 different aspects of local inflammatory activation in the rat Nervous System:

1. The genetic heterogeneity of Glial activation across inbred strains after Nerve injury
2. Expression of MHC Class I Genes in the CNS
3. NeuroProtective effects of CNS Antigen AutoReactive Immune reactions

Apart from NeuroImmune Diseases such as Experimental AutoImmune Encephalomyelitis & Multiple Sclerosis, these features are also of relevance for a wider range of Neurological Diseases which present pathological signs of inflammation, such as Alzheimer's Dementia, CerebroVascular Diseases and CNS Trauma.

The cellular constituents of the Blood-Brain Barrier (BBB) must make finely tuned, regulatory responses to maintain MicroVascular Homeostasis. The mechanisms by which this task is accomplished are largely unknown.

However, it is thought they involve a series of cross-talk mechanisms among Endothelial Cells (EC), Pericytes (PC), and Astrocytes.

During inflammation, the BBB is exposed to a number of Biological Response modifiers including Cytokines released by infiltrating Leukocytes.

The response to inflammatory Cytokines may alter the normal regulatory function of EC and PC. These changes may account for some of the pathological findings in Central Nervous System (CNS) Inflammatory Disease.

Previous studies have shown that PC and EC may have Immune potential. We have investigated the response of the PC to a variety of ProInflammatory Cytokines.

Primary rat PC constitutively express low levels of InterCellular Adhesion Molecule-1 (ICAM-1) and Major Histocompatibility Complex (MHC) Class I molecule.

Which can be upregulated in response to the Cytokine Interferon-gamma (IFN-γ). IFN-γ also induced the expression of MHC Class II molecule.

After induction of MHC Class II molecule, CNS PC acquired the capacity to present Antigen to primed syngeneic rat T-Lymphocytes. Antigen presentation by PC was comparable to that seen with classic Antigen-Presenting Cells.

A small number of primary PC constitutively express low levels of Vascular Cell Adhesion Molecule-1 (VCAM-1), which was increased on exposure to Tumor Necrosis Factor-alpha (TNF-).

Results suggest that CNS PC respond to Inflammatory Cytokines, are involved in T-Lymphocyte activation, and express Cell Surface Adhesion Molecules (VCAM-1, ICAM-1).

That may provide costimulatory activity. It is likely that CNS PC are important in NeuroImmune Networks at the BBB.

The irreversible loss of function after Axonal injury in the Central Nervous System (CNS) is a result of the lack of NeuroGenesis, poor regeneration, and the spread of damage.

Caused by toxicity emanating from the degenerating Axons to uninjured Neurons in the vicinity.

Now, 100 years after Ramon y Cajal's discovery that CNS Neurons - unlike Neurons of the Peripheral Nervous System - fail to regenerate, it has become evident that:

1. CNS tissue is indeed capable of regenerating, at least in part,
   • provided it acquires appropriate conditions for growth support
2. The spread of damage can be stopped
3. The PostInjury rescue of Neurons thus achieved
4. If ways are found to neutralize the mediators of toxicity either by
   • inhibiting their action
   • or increasing tissue resistance to them

In most physiological systems the processes of tissue maintenance and repair depend on the active assistance of Immune Cells. In the CNS, however, communication with the Immune System is restricted.

The accumulated evidence from our previous studies suggests that the poor PostTraumatic repair and maintenance in the CNS is due at least in part to this restriction.

Key factors in the recovery of injured tissues, but missing or deficient in the CNS, are the processes of recruitment and activation of Immune Cells.

We therefore propose the development of Immune Cell therapies in which the injured CNS is exogenously provided with an adequate number of appropriately activated Immune Cells:

Macrophages for regrowth and AutoImmune T-Cells for maintenance controlled in such a way as to derive maximal benefit with minimal risk of disease.

It is expected that these self-adjusting cells will communicate with the damaged tissue, monitor tissue needs, and control the dynamic course of CNS healing.

NeuroTrophins, secreted in an activity-dependent manner, are thought to be involved in the activity-dependent refinement of Synaptic connections.

Here we demonstrate that in Hippocampal Neurons and the rat pheochromocytoma cell line PC12 application of exogenous NeuroTrophins induces secretion of NeuroTrophins.

An effect that is mediated by the activation of Tyrosine Kinase NeuroTrophin Receptors (Trks).

Like activity-dependent secretion of NeuroTrophins, NeuroTrophin-induced NeuroTrophin secretion requires mobilization of Calcium from IntraCellular stores.

Because NeuroTrophins are likely to be released from both Dendrites and Axons:

NeuroTrophin-induced NeuroTrophin release represents a potential positive feedback mechanism, contributing to the reinforcement and stabilization of Synaptic connections.

We analyzed the inducibility of Major Histocompatibility Complex (MHC) Class II molecules of Astrocytes and Microglia in organotypic Hippocampus slice cultures of Lewis rats.

Treatment with Interferon-gamma (IFN-γ) resulted in the induction of MHC Class II molecules.

On Microglia preferentially in the injured marginal zones of the slice culture, but only sporadically in areas containing intact Neuronal architecture.

In Astrocytes, inducibility of MHC Class II molecules was even more strictly controlled:

IFN-γ treatment induced MHC Class II expression only in the slice culture zones containing degenerated Neurons, and not in the presence of functional Neurons.

After suppression of spontaneous Neuronal activity of the slice culture by the Sodium Channel Blocker TetrodoToxin, MHC Class II molecules on Astrocytes could be induced by IFN-γ in areas with intact Neuronal architecture, and Microglia Cells exhibited a higher level of expression.

These data suggest that loss of Neurons could result in MHC Class II inducibility of Glial Cells, and thus in increased Immune reactivity of Nervous tissue.

There is increasing evidence that NeuroTrophins (NTs) are involved in processes of Neuronal plasticity besides their well-established actions in regulating the survival, differentiation, and maintenance of functions of specific populations of Neurons.

Nerve Growth Factor, Brain-Derived Neurotrophic Factor, NT-4/5, and corresponding AntiBodies dramatically modify the development of the Visual Cortex.

Although the Neuronal elements involved have not yet been identified, complementary studies of other systems have demonstrated that NT synthesis is rapidly regulated by Neuronal activity and that NTs are released in an activity-dependent manner from Neuronal Dendrites.

These data, together with the observation that NTs enhance Transmitter release from Neurons that express the corresponding signal-transducing Trk Receptors, suggest a role for NTs as selective retrograde messengers that regulate Synaptic efficacy.

In this study Microglial Cells isolated from Brain Cell cultures of newborn mice were characterized and investigated for Morphology, their responses to Growth Factors and their functional properties.

The Microglial Cells were Phagocytic, contained nonspecific Esterase activity and expressed Fc (IgG1/2b) and type-3 Complement Receptors.

Scanning electron microscopy revealed that in analogy to Brain tissue two types of Microglial Cells are present in the cultures: the ameboid and the ramified type which both display similar appearance by transmission electron microscopy.

InterLeukin-3 and the Granulocyte-Macrophage Colony-Stimulating Factor were potent Growth Factors for the cultured Microglial Cells.

The cells were negative for Class II Antigens (Ia) of the Major Histocompatibility Antigen Complex.

However, upon treatment with Interferon-gamma (IFN-γ) Microglial Cells became Ia+ and functioned as Antigen-Presenting Cells when tested on Ovalbumin-specific Ia-restricted Helper T-Cells.

Furthermore, Microglial Cells exposed to IFN-γ and EndoToxin developed Tumor Cell CytoToxicity and produced Tumor Necrosis Factor-alpha (TNF-).

Taken together, Microglial Cells share the characteristics of cells of the Macrophage lineage.

Reactive Microglia in the CNS have been implicated in the pathogenesis of White Matter Disorders, such as Periventricular Leukomalacia and Multiple Sclerosis.

However, the mechanism by which activated Microglia kill Oligodendrocytes (OLs) remains elusive.

Here we show that LipoPolySaccharide (LPS)-induced death of developing OLs is caused by Microglia-derived Peroxynitrite, the reaction product of Nitric Oxide (NO) and Superoxide Anion.

Blocking Peroxynitrite formation with Nitric Oxide Synthase Inhibitors, Superoxide Dismutase Mimics, or a decomposition catalyst abrogated the CytoToxicity.

Only Microglia, but not OLs, expressed Inducible NO Synthase (INOS) after LPS challenge; Microglia from INOS knockout mice were not CytoToxic upon activation.

The molecular source for superoxide was identified as the superoxide-generating enzyme NADPH Oxidase. The Oxidase was activated upon LPS exposure, and its inhibition prevented Microglial toxicity toward OLs.

Furthermore, Microglia isolated from mice deficient in the catalytic component of the Oxidase, gp91(phox), failed to induce cell death.

Our results reveal a role for NADPH oxidase in LPS-induced OL death and suggest that Peroxynitrite produced by INOS and NADPH Oxidase in activated Microglia may play an important role in the pathogenesis of White Matter disorders.