Some recent studies suggest that in progressive multiple sclerosis, neurodegeneration may occur independently from inflammation. The aim of our study was to analyse the interdependence of inflammation, neurodegeneration and disease progression in various multiple sclerosis stages in relation to lesional activity and clinical course, with a particular focus on progressive multiple sclerosis. The study is based on detailed quantification of different inflammatory cells in relation to axonal injury in 67 multiple sclerosis autopsies from different disease stages and 28 controls without neurological disease or brain lesions. We found that pronounced inflammation in the brain is not only present in acute and relapsing multiple sclerosis but also in the secondary and primary progressive disease. T- and B-cell infiltrates correlated with the activity of demyelinating lesions, while plasma cell infiltrates were most pronounced in patients with secondary progressive multiple sclerosis (SPMS) and primary progressive multiple sclerosis (PPMS) and even persisted, when T- and B-cell infiltrates declined to levels seen in age matched controls. A highly significant association between inflammation and axonal injury was seen in the global multiple sclerosis population as well as in progressive multiple sclerosis alone. In older patients (median 76 years) with long-disease duration (median 372 months), inflammatory infiltrates declined to levels similar to those found in age-matched controls and the extent of axonal injury, too, was comparable with that in age-matched controls. Ongoing neurodegeneration in these patients, which exceeded the extent found in normal controls, could be attributed to confounding pathologies such as Alzheimer's or vascular disease. Our study suggests a close association between inflammation and neurodegeneration in all lesions and disease stages of multiple sclerosis. It further indicates that the disease processes of multiple sclerosis may die out in aged patients with long-standing disease.

Multiple sclerosis is a chronic inflammatory disease of the central nervous system, associated with demyelination and neurodegeneration. The mechanisms of tissue injury are currently poorly understood, but recent data suggest that mitochondrial injury may play an important role in this process. Since mitochondrial injury can be triggered by reactive oxygen and nitric oxide species, we analysed by immunocytochemistry the presence and cellular location of oxidized lipids and oxidized DNA in lesions and in normal-appearing white matter of 30 patients with multiple sclerosis and 24 control patients without neurological disease or brain lesions. As reported before in biochemical studies, oxidized lipids and DNA were highly enriched in active multiple sclerosis plaques, predominantly in areas that are defined as initial or ‘prephagocytic’ lesions. Oxidized DNA was mainly seen in oligodendrocyte nuclei, which in part showed signs of apoptosis. In addition, a small number of reactive astrocytes revealed nuclear expression of 8-hydroxy-d-guanosine. Similarly, lipid peroxidation-derived structures (malondialdehyde and oxidized phospholipid epitopes) were seen in the cytoplasm of oligodendrocytes and some astrocytes. In addition, oxidized phospholipids were massively accumulated in a fraction of axonal spheroids with disturbed fast axonal transport as well as in neurons within grey matter lesions. Neurons stained for oxidized phospholipids frequently revealed signs of degeneration with fragmentation of their dendritic processes. The extent of lipid and DNA oxidation correlated significantly with inflammation, determined by the number of CD3 positive T cells and human leucocyte antigen-D expressing macrophages and microglia in the lesions. Our data suggest profound oxidative injury of oligodendrocytes and neurons to be associated with active demyelination and axonal or neuronal injury in multiple sclerosis.

Multiple sclerosis is a chronic inflammatory disease of the central nervous system, associated with demyelination and neurodegeneration. The mechanisms of tissue injury are poorly understood, but recent data suggest that mitochondrial injury may play an important role in this process. Mitochondrial injury can be triggered by reactive oxygen and nitric oxide species, and we recently provided evidence for oxidative damage of oligodendrocytes and dystrophic axons in early stages of active multiple sclerosis lesions. In this study, we identified potential sources of reactive oxygen and nitrogen species through gene expression in carefully staged and dissected lesion areas and by immunohistochemical analysis of protein expression. Genome-wide microarrays confirmed mitochondrial injury in active multiple sclerosis lesions, which may serve as an important source of reactive oxygen species. In addition, we found differences in the gene expression levels of various nicotinamide adenine dinucleotide phosphate oxidase subunits between initial multiple sclerosis lesions and control white matter. These results were confirmed at the protein level by means of immunohistochemistry, showing upregulation of the subunits gp91phox, p22phox, p47phox, nicotinamide adenine dinucleotide phosphate oxidase 1 and nicotinamide adenine dinucleotide phosphate oxidase organizer 1 in activated microglia in classical active as well as slowly expanding lesions. The subunits gp91phox and p22phox were constitutively expressed in microglia and were upregulated in the initial lesion. In contrast, p47phox, nicotinamide adenine dinucleotide phosphate oxidase 1 and nicotinamide adenine dinucleotide phosphate oxidase organizer 1 expression were more restricted to the zone of initial damage or to lesions from patients with acute or early relapsing/remitting multiple sclerosis. Double labelling showed co-expression of the nicotinamide adenine dinucleotide phosphate oxidase subunits in activated microglia and infiltrated macrophages, suggesting the assembly of functional complexes. Our data suggest that the inflammation-associated oxidative burst in activated microglia and macrophages plays an important role in demyelination and free radical-mediated tissue injury in the pathogenesis of multiple sclerosis.

In multiple sclerosis (MS), diffuse degenerative processes in the deep grey matter have been associated with clinical disabilities. We performed a systematic study in MS deep grey matter with a focus on the incidence and topographical distribution of lesions in relation to white matter and cortex in a total sample of 75 MS autopsy patients and 12 controls. In addition, detailed analyses of inflammation, acute axonal injury, iron deposition and oxidative stress were performed. MS deep grey matter was affected by two different processes: the formation of focal demyelinating lesions and diffuse neurodegeneration. Deep grey matter demyelination was most prominent in the caudate nucleus and hypothalamus and could already be seen in early MS stages. Lesions developed on the background of inflammation. Deep grey matter inflammation was intermediate between low inflammatory cortical lesions and active white matter lesions. Demyelination and neurodegeneration were associated with oxidative injury. Iron was stored primarily within oligodendrocytes and myelin fibres and released upon demyelination. In addition to focal demyelinated plaques, the MS deep grey matter also showed diffuse and global neurodegeneration. This was reflected by a global reduction of neuronal density, the presence of acutely injured axons, and the accumulation of oxidised phospholipids and DNA in neurons, oligodendrocytes and axons. Neurodegeneration was associated with T cell infiltration, expression of inducible nitric oxide synthase in microglia and profound accumulation of iron. Thus, both focal lesions as well as diffuse neurodegeneration in the deep grey matter appeared to contribute to the neurological disabilities of MS patients.

The causes of incomplete remyelination in progressive multiple sclerosis are unknown, as are the pathological correlates of the different clinical characteristics of patients with primary and secondary progressive disease. We analysed brains and spinal cords from 51 patients with progressive multiple sclerosis by planimetry. Thirteen patients with primary progressive disease were compared with 34 with secondary progressive disease. In patients with secondary progressive multiple sclerosis, we found larger brain plaques, more demyelination in total and higher brain loads of active demyelination compared with patients with primary progressive disease. In addition, the brain density of plaques with high-grade inflammation and active demyelination was highest in secondary progressive multiple sclerosis and remained ∼18% higher than in primary progressive multiple sclerosis after adjustments for other plaque types and plaque number (P < 0.05). Conversely, the proportion of remyelinated shadow plaques (P < 0.05) and the overall remyelination capacity (P < 0.01) per brain were higher in primary, compared with secondary, progressive multiple sclerosis. By contrast, there were no group differences in the brain load or frequency of low-grade inflammatory plaques with slowly expanding demyelination. Spinal cord lesion loads and remyelination capacity were also comparable in the two patient groups. Remyelinated areas were more vulnerable than the normal-appearing white matter to new demyelination, including active demyelination in secondary progressive multiple sclerosis. ‘Recurrent’ slowly expanding demyelination, affecting remyelinated areas, and the load of slowly expanding demyelination correlated with incomplete remyelination in both groups. In turn, incomplete remyelination in the spinal cord correlated with higher disease-related disability (determined retrospectively; r = −0.53; P < 0.05 for remyelination capacity versus disease severity). By contrast, such a correlation was not observed in the brain. We propose that regulatory and reparative properties could protect the white matter of the brain in patients with primary progressive multiple sclerosis. These patients may, thereby, be spared symptoms until the spinal cord is affected. By contrast, recurrent active demyelination of repaired myelin could explain why similar symptoms often develop in consecutive relapses in relapsing-remitting/secondary progressive multiple sclerosis. Our data also indicate that slowly expanding demyelination may irreparably destroy normal and repaired myelin, supporting the concept of slowly expanding demyelination as an important pathological correlate of clinical progression.