Objective: Cortical gray matter (GM) pathology, involving demyelination and neurodegeneration, associated with meningeal inflammation, could be important in determining disability progression in multiple sclerosis (MS). However, we need to know more about how cortical demyelination, neurodegeneration, and meningeal inflammation contribute to pathology at early stages of MS to better predict long-term outcome. Methods: Tissue blocks from short disease duration MS (n = 12, median disease duration = 2 years), progressive MS (n = 21, disease duration = 25 years), non-diseased controls (n = 11), and other neurological inflammatory disease controls (n = 6) were quantitatively analyzed by immunohistochemistry, immunofluorescence, and in situ hybridization. Results: Cortical GM demyelination was extensive in some cases of acute MS (range = 1-48% of total cortical GM), and subpial lesions were the most common type (62%). The numbers of activated (CD68 + ) microglia/macrophages were increased in cases with subpial lesions, and the density of neurons was significantly reduced in acute MS normal appearing and lesion GM, compared to controls (p < 0.005). Significant meningeal inflammation and lymphoid-like structures were seen in 4 of 12 acute MS cases. The extent of meningeal inflammation correlated with microglial/macrophage activation (p < 0.05), but not the area of cortical demyelination, reflecting the finding that lymphoid-like structures were seen adjacent to GM lesions as well as areas of partially demyelinated/remyelinated, cortical GM. Interpretation: Our findings demonstrate that cortical demyelination, neuronal loss, and meningeal inflammation are notable pathological hallmarks of acute MS and support the need to identify early biomarkers of this pathology to better predict outcome. ANN NEUROL 2018;84:829-842 M ultiple sclerosis (MS) is a highly variable and lifechanging condition, characterized by inflammation, demyelination, and neurodegeneration. MS typically presents as a relapsing-remitting disease, where bouts of acute inflammation and new, active, demyelinating lesions are associated with neurological impairment. Lesions of the gray matter (GM) occur at all stages of the disease, and the extent of cortical GM pathology predicts conversion to clinically definite MS and associates with cognitive and motor disability, epilepsy, and an earlier transition to the progressive phase. 1 Recent findings suggest that subpial GM lesions of the neocortex, the principal lesion type of the MS cortex, are related, at least in part, to inflammatory activity in the overlying leptomeninges. 2 We and others have shown how the degree of meningeal inflammation correlates with cortical microglial activation, neuritic and neuronal degeneration, and demyelination in primary progressive and View this article online at wileyonlinelibrary.com.

The complement pathway has potential contributions to both white (WM) and grey matter (GM) pathology in Multiple Sclerosis (MS). A quantitative assessment of complement involvement is lacking. Here we describe the use of Tissue MicroArray (TMA) methodology in conjunction with immunohistochemistry to investigate the localization of complement pathway proteins in progressive MS cortical GM and subcortical WM. Antibodies targeting complement proteins C1q, C3b, regulatory proteins C1 inhibitor (C1INH, complement receptor 1 (CR1), clusterin, factor H (FH) and the C5a anaphylatoxin receptor (C5aR) were utilised alongside standard markers of tissue pathology. All stained slides were digitised for quantitative analysis. We found that numbers of cells immunolabelled for HLA-DR, GFAP, C5aR, C1q and C3b were increased in WM lesions (WML) and GM lesions (GML) compared to normal appearing WM (NAWM) and GM (NAGM), respectively. The complement regulators C1INH, CR1, FH and clusterin were more abundant in WM lesions, while the number of C1q+ neurons were increased and the number of C1INH+, clusterin+, FH+ and CR1+ neurons decreased in GM lesions. The number of complement component positive cells (C1q, C3b) correlated with complement regulator expression in WM, but there was no statistical association between complement activation and regulator expression in the GM. We conclude that TMA methodology and quantitative analysis provides evidence of complement dysregulation in MS GML, including an association of the numerical density of C1q+ cells with tissue lesions. Our work confirms that complement activation and dysregulation occur in all cases of progressive MS and suggest that complement may provide potential biomarkers of the disease.

Aim: Multiple sclerosis (MS) is an inflammatory demyelinating and neurodegenerative disease. Much of the complex symptomatology relates to pathology outside the classic white matter plaque, whereby lesions of the cortical grey matter, which are difficult to resolve by conventional clinical imaging, are in part predictive of outcome. We investigated the extent of grey matter pathology in whole coronal macrosections to reassess the contribution of cortical pathology to total demyelinating lesion area in progressive MS. Methods: Twenty-two cases of progressive MS were prepared as whole bi-hemispheric macrosections for histology, immunostaining and quantitative analysis of lesion number and relative area, leptomeningeal inflammation and microglial/macrophage activation. Results: Cortical grey matter demyelination was seen in all cases, which was more extensive than in white and deep grey matter (hippocampus, thalamus and basal ganglia) and accounted for 0.8%-60.2% of the entire measurable cortical ribbon. The pattern of cortical grey matter demyelination was predominantly subpial (mean 90.9%, range 60%-100%, of total cortical grey matter lesion area) and cases with the largest areas of subpial cortical lesions had more and larger deep grey matter lesions, greater numbers of activated microglia/macrophages, both in lesions as well as in normal cortical grey matter, together with elevated leptomeningeal inflammation and lymphoid-like structures. White matter lesion area was unchanged when compared with the progressive MS cases with little subpial cortical demyelination. Conclusion: Analysis of whole coronal macrosections reveals cortical demyelination is more extensive than reported by conventional histological methods. Cases of progressive MS with substantial subpial cortical demyelination that is independent of underlying white matter lesion area support the implications that these lesions may in-part arise through different pathogenetic mechanisms. Biomarkers and/or imaging correlates of this subpial pathology are required if we are to fully comprehend the clinical disease process.

CSF levels of sCD59 are not a biomarker of demyelinating diseases. High levels of sCD59 in CSF relative to plasma suggest an intrathecal source; CD59 expression in brain parenchyma was low, but expression was strong on choroid plexus (CP) epithelium, immediately adjacent the CSF, suggesting that this is the likely source.

BackgroundThe extent of cortical pathology is an important determinant of multiple sclerosis (MS) severity. Cortical demyelination and neurodegeneration are related to inflammation of the overlying leptomeninges, a more inflammatory CSF milieu and with parenchymal microglia and astroglia activation. These are all components of the compartmentalised inflammatory response. Compartmentalised inflammation is a feature of progressive MS, which is not targeted by disease modifying therapies. Complement is differentially expressed in the MS CSF and complement, and complement receptors, are associated with demyelination and neurodegeneration.MethodsTo better understand if complement activation in the leptomeninges is associated with underlying cortical demyelination, inflammation, and microglial activation, we performed a neuropathological study of progressive MS (n = 22, 14 females), neuroinflammatory (n = 8), and non-neurological disease controls (n = 10). We then quantified the relative extent of demyelination, connective tissue inflammation, complement, and complement receptor positive microglia/macrophages.ResultsComplement was elevated at the leptomeninges, subpial, and within and around vessels of the cortical grey matter. The extent of complement C1q immunoreactivity correlated with connective tissue infiltrates, whilst activation products C4d, Bb, and C3b associated with grey matter demyelination, and C3a receptor 1+ and C5a receptor 1+ microglia/macrophages closely apposed C3b labelled cells. The density of C3a receptor 1+ and C5a receptor 1+ cells was increased at the expanding edge of subpial and leukocortical lesions. C5a receptor 1+ cells expressed TNFα, iNOS and contained puncta immunoreactive for proteolipid protein, neurofilament and synaptophysin, suggesting their involvement in grey matter lesion expansion.InterpretationThe presence of products of complement activation at the brain surfaces, their association with the extent of underlying pathology and increased complement anaphylatoxin receptor positive microglia/macrophages at expanding cortical grey matter lesions, could represent a target to modify compartmentalised inflammation and cortical demyelination.