Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS) affecting over two million individuals worldwide. It is a major cause of neurodegeneration in young adults, but can also first manifest in late middle age or during childhood. Its aetiology is unknown, however genetic predisposition and environmental factors are known to contribute significantly to disease development. MS exists as a number of sub-types differentiated by age of onset and frequency and severity of episodes of neurological deficits.
It was originally thought that MS was a disease of the CNS white matter (wm), but it is now apparent that the grey matter (gm) is also affected from the earliest disease stage. Using the animal model for MS known as experimental autoimmune encephalomyelitis (EAE), we aim to generate proof-of-concept for pathological and molecular mechanisms underlying the disease and to evaluate candidate MS drugs.
Research areas
Comparative evaluation of EAE variants
Given the complexity of MS and its unclear aetiology, unsurprisingly, no animal model recapitulating the clinical features together with the pathophysiology of MS has been successfully developed to date. Currently, EAE generated by autoimmune-mediated demyelination, is the most widely accepted model. However, EAE itself is a complex disease which is not completely understood. Similarly to MS, it can follow multiple clinical courses and in a number of these variants, clinical profiles apparently mimicking those of MS subtypes, are actually associated with major deviations from MS in terms of hallmarks of pathology. Therefore each variant only reflects certain specific facets of MS.
In view of current developments in large-scale gene expression and protein profiling technology, as well as in drug design, it is important to continue to identify areas of coincidence in disease mechanisms between EAE variants and MS, so that the full capabilities provided by these technologies can be exploited with the aims of further elucidating molecular pathways underlying MS and generating more refined drugs.
Our lab has mapped the spatio-temporal evolution of disease, clinically and pathologically in 4 EAE variants, namely: (a) the NOD/Lt mouse strain induced with peptide 35-55 of myelin oligodendrocyte glycoprotein (MOG35-55), (b) the C57Bl/6 mouse strain induced with MOG35-55, (c) the BALB/c mouse strain induced with peptide 180-199 of proteolipid protein (PLP180-199) and (d) SJL/J mouse strain induced with peptide 139-151 of proteolipid protein (PLP139-151).
Molecular mechanisms underlying wm and gm pathology during neuroinflammation
MS is now regarded as a global CNS disease. Previously thought to be a disease of the wm characterized by inflammatory demyelinating focal white matter lesions, it has become increasingly evident over the last two decades that primary gm damage is a significant component of MS pathophysiology. This type of damage is characterized by severe pathology occurring within the context of less significant inflammation and blood brain barrier (BBB) loss of function. Our investigations of multiple EAE variants have found that gm pathological features are reproduced in the model.
To elucidate the spatio-temporal and pathological relationships between white and grey matter mechanisms, we have developed a range of approaches. These include histology, quantitative confocal microscopy for neuronal, glial and stress markers and laser microdissection followed by expression profiling on disease/pathway-specific arrays. Analyses are performed on wm and adjacent gm regions, for example in the cervical spinal cord and cerebellum. When used over a time course, these approaches enable comparisons of disease evolution at the cellular and molecular levels.
Currently they are being used to evaluate:
The kinetics of BBB breakdown and repair.
The relationship between inflammation, demyelination, axonal/neuronal damage, glial responses and cellular stress in each CNS compartment.
Characterization of the inflammatory environment generated in each compartment, and,
The relationship between stress and neuronal apoptosis.
These combined approaches also allowed the development of a workflow for the evaluation of the efficacy of MS drugs on wm and gm independently.
The role of astrocytes in neuroinflammation
Concurrent with the changing view of MS, evidence of axon-glia and glia-glia networking and communication emerged. These data emphasize the existence and importance of cross-talk and interdependence between the cellular CNS elements, namely astrocytes, microglia, oligodendroglia and nerve cell bodies and axons. A picture of an immensely complex interrelated and constantly communicating multi-dimensional network emerges in which change or damage to one element will inevitably be transmitted throughout the whole system.
Our electron microscopy studies have clearly demonstrated the occurrence of widespread axonal damage and significant astrocyte hypertrophy before large-scale parenchymal invasion of inflammatory cells, particularly T cells and before widespread demyelination. These observations are of particular interest, because the contribution of astrocytes to neuroinflammation is poorly understood and is generally viewed as being secondary to that of microglia.
Further to the above, we have compared astrocytic responses between NOD/Lt and C57Bl/6 variants by quantitative confocal microscopy and quantitative western blotting. These studies demonstrated opposing responses between the two variants, with gradually increasing astrocytic response in NOD/Lt mice with increasing inflammatory infiltration, but a diminishing response in C57Bl/6 mice.
Emotional and cognitive changes during neuroinflammation
It is now increasingly recognized that anxiety and cognition symptoms in MS are primary effects of the disease, rather than psychological consequences of living with a chronic illness. To date, only a limited number of investigations of the molecular basis of these symptoms, based on the EAE model, have been conducted. These studies concluded that hitherto unknown pathological mechanisms underlie these defects.
Our preliminary data, on the other hand, support classical immunopathological processes as the basis of anxiety and cognition problems in EAE. In collaboration with Dr Matthew Hale, Department of Psychology and Counselling, LTU, we have begun investigations of the timing of onset and severity of these deficits in EAE, using well established behavioural approaches including the elevated plus maze and novel object recognition tests. Data will be correlated with cellular and molecular changes in the relevant brain regions, namely the hippocampal formation and striatum. For these investigations we will use the MOG35-55–induced NOD/Lt and C57Bl/6 variants and the PLP180-199-induced BALB/c variant. Additionally the capacity of MS drugs, for example FTY720 (aka Fingolimod or Gilenya), to ameliorate these symptoms will be examined. We will also compare responses between EAE variants on disease development and severity and drug treatment.
