Reports based on magnetic resonance imaging (MRI) of multiple sclerosis (MS) patients and its animal model (experimental autoimmune encephalomyelitis, or EAE) have consistently documented early and extensive grey matter changes. Post-mortem investigations have also demonstrated cortical lesions without inflammation. It is clear that the question of timing and extent of grey matter involvement and in particular, that of direct neuronal injury in MS, must be addressed. This project was specifically aimed as testing the hypothesis that: in early, pre-symptomatic EAE, pathological changes occur in grey matter, expressed in parallel by astrocytes, microglia and neurons.
Our approach was to investigate parameters of neuroinflammation, namely inflammation, demyelination and axonal injury in grey and white matter, by immunostaining of longitudinal sections of the cervical spinal cord in regions C2 to C8.
Inflammation was evaluated by investing the presence of CD3-positive cells, microglial activation and expression of MHC class II. CD3+ cells were found within blood vessels in grey and white matter, from 10 days post disease initiation (dpi). By 16dpi, corresponding with the peak of the first attack, these cells were clearly evident in the grey matter tissue of the brain and spinal cord. However, they were more numerous in white matter. By 10dpi microglia also showed signs of activation. These were more pronounced by 16dpi and more evident in white relative to grey matter. MHC Class II expression was detected and appeared associated with microglia only. Astrocytic reactivity was evident from 7dpi in both grey and white matter. In both regions astrocyte reactivity was very elevated from early stages, especially under the pia mater and around the central canal. This was manifested both by increases in cell body volume and length of processes and continued to occur over the whole time course investigated. Changes to neurons were also detected, with axonal injury evident in the white matter, from 10dpi and continued to increase dramatically. In grey matter, by contrast, levels of immunostaining demonstrated a change in cytoskeletal structure of neurons. Simultaneously in double immunostaining with anti-GFAP, astrocytes immediately adjacent to neuronal cell bodies were found to be highly activated, thus demonstrating a dramatic change in the environment of neurons.
Taken together, these data demonstrate grey matter pathology from pre-clinical changes and suggest
that neuronal function may be compromised (but not necessarily irrevocably destroyed) from the earliest disease stage. They support recent claims that MS should be redefined as a diffuse central nervous system rather that a white matter disease. Further studies are in progress to define grey matter pathology and the extent of neuronal damage. These data will be compared to MRI findings in patients and become significant in terms of predicting disease course.
Updated: 03 January, 2007