R Kapoor

Demyelination: the role of reactive oxygen and nitrogen species.

This review summarises the role that reactive oxygen and nitrogen species play in demyelination, such as that occurring in the inflammatory demyelinating disorders multiple sclerosis and Guillain‐Barré syndrome. The concentrations of reactive oxygen and nitrogen species (e.g. superoxide, nitric oxide and peroxynitrite) can increase dramatically under conditions such as inflammation, and this can overwhelm the inherent antioxidant defences within lesions. Such oxidative and/or nitrative stress can damage the lipids, proteins and nucleic acids of cells and mitochondria, potentially causing cell death. Oligodendrocytes are more sensitive to oxidative and nitrative stress in vitro than are astrocytes and microglia, seemingly due to a diminished capacity for antioxidant defence, and the presence of raised risk factors, including a high iron content. Oxidative and nitrative stress might therefore result in vivo in selective oligodendrocyte death, and thereby demyelination. The reactive species may also damage the myelin sheath, promoting its attack by macrophages. Damage can occur directly by lipid peroxidation, and indirectly by the activation of proteases and phospholipase A2. Evidence for the existence of oxidative and nitrative stress within inflammatory demyelinating lesions includes the presence of both lipid and protein peroxides, and nitrotyrosine (a marker for peroxynitrite formation). The neurological deficit resulting from experimental autoimmune demyelinating disease has generally been reduced

Electrically active axons degenerate when exposed to nitric oxide

Axonal degeneration is a major cause of permanent deficit in inflammatory neurological diseases such as multiple sclerosis. Axons undergo degeneration specifically at the site of the inflammatory lesions, suggesting that locally produced inflammatory factors mediate the phenomenon. One such factor is nitric oxide (NO), which we have previously reported can cause reversible conduction block in axons. Here we confirm these observations and extend them to show that axons exhibit the early stages of wallerian degeneration if they are conducting impulses at physiological frequencies while they are exposed to the low micromolar concentrations of NO that are likely to occur at sites of inflammation. Rat dorsal roots were concurrently exposed in vivo to both NO and sustained impulse activity at 1, 50, or 100 Hz. Although our in vivo observations necessarily focused on the more acute responses, morphological examination of exposed roots at the end of the recording period revealed nodal and paranodal changes consistent with acute wallerian degeneration in roots stimulated at 50 or 100 Hz. This interpretation was confirmed in a few experiments that were prolonged to permit more obvious indicators of degeneration to develop. In these experiments the formation of myelin ovoids and frank axonolysis occurred in more than 95% of fibers. Roots stimulated at only 1 Hz appeared normal. We propose that the combination of normal impulse traffic and NO at sites of inflammation may cause axonal degeneration and that electrical activity may therefore be an important factor in causing permanent disability in patients with neuroinflammatory disorders. Ann Neurol 2001;49:470–476

Blockers of sodium and calcium entry protect axons from nitric oxide-mediated degeneration.

Axonal degeneration can be an important cause of permanent disability in neurological disorders in which inflammation is prominent, including multiple sclerosis and Guillain–Barré syndrome. The mechanisms responsible for the degeneration remain unclear, but it is likely that axons succumb to factors produced at the site of inflammation, such as nitric oxide (NO). We previously have shown that axons exposed to NO in vivo can undergo degeneration, especially if the axons are electrically active during NO exposure. The axons may degenerate because NO can inhibit mitochondrial respiration, leading to intraaxonal accumulation of Na+ and Ca2+ ions. Here, we show that axons can be protected from NO‐mediated damage using low concentrations of Na+ channel blockers, or an inhibitor of Na+/Ca2+ exchange. Our findings suggest a new strategy for axonal protection in an inflammatory environment, which may be effective in preventing the accumulation of permanent disability in patients with neuroinflammatory disorders. Ann Neurol 2003

Axonal protection using flecainide in experimental autoimmune encephalomyelitis

Axonal degeneration is a major cause of permanent neurological deficit in multiple sclerosis (MS), but no current therapies for the disease are known to be effective at axonal protection. Here, we examine the ability of a sodium channel–blocking agent, flecainide, to reduce axonal degeneration in an experimental model of MS, chronic relapsing experimental autoimmune encephalomyelitis (CR‐EAE). Rats with CR‐EAE were treated with flecainide or vehicle from either 3 days before or 7 days after inoculation (dpi) until termination of the experiment at 28 to 30 dpi. Morphometric examination of neurofilament‐labeled axons in the spinal cord of CR‐EAE animals showed that both flecainide treatment regimens resulted in significantly higher numbers of axons surviving the disease (83 and 98% of normal) compared with controls (62% of normal). These findings indicate that flecainide and similar agents may provide a novel therapy aimed at axonal protection in MS and other neuroinflammatory disorders.

Lamotrigine for neuroprotection in secondary progressive multiple sclerosis: a randomised, double-blind, placebo-controlled, parallel-group trial

BACKGROUND Partial blockade of voltage-gated sodium channels is neuroprotective in experimental models of inflammatory demyelinating disease. In this phase 2 trial, we aimed to assess whether the sodium-channel blocker lamotrigine is also neuroprotective in patients with secondary progressive multiple sclerosis. METHODS Patients with secondary progressive multiple sclerosis who attended the National Hospital for Neurology and Neurosurgery or the Royal Free Hospital, London, UK, were eligible for inclusion in this double-blind, parallel-group trial. Patients were randomly assigned via a website by minimisation to receive lamotrigine (target dose 400 mg/day) or placebo for 2 years. Treating physicians, evaluating physicians, and patients were masked to treatment allocation. The primary outcome was the rate of change of partial (central) cerebral volume over 24 months. All patients who were randomly assigned were included in the primary analysis. This trial is registered with ClinicalTrials.gov, NCT00257855. FINDINGS 120 patients were randomly assigned to treatment (87 women and 33 men): 61 to lamotrigine and 59 to placebo. 108 patients were analysed for the primary endpoint: 52 in the lamotrigine group and 56 in the placebo group. The mean change in partial (central) cerebral volume per year was -3.18 mL (SD -1.25) in the lamotrigine group and -2.48 mL (-0.97) in the placebo group (difference -0.71 mL, 95% CI -2.56 to 1.15; p=0.40). However, in an exploratory modelling analysis, lamotrigine treatment seemed to be associated with greater partial (central) cerebral volume loss than was placebo in the first year (p=0.04), and volume increased partially after treatment stopped (p=0.04). Lamotrigine treatment reduced the deterioration of the timed 25-foot walk (p=0.02) but did not affect other secondary clinical outcome measures. Rash and dose-related deterioration of gait and balance were experienced more by patients in the lamotrigine group than the placebo group. INTERPRETATION The effect of lamotrigine on cerebral volume of patients with secondary progressive multiple sclerosis did not differ from that of placebo over 24 months, but lamotrigine seemed to cause early volume loss that reversed partially on discontinuation of treatment. Future trials of neuroprotection in multiple sclerosis should include investigation of complex early volume changes in different compartments of the CNS, effects unrelated to neurodegeneration, and targeting of earlier and more inflammatory disease. FUNDING Multiple Sclerosis Society of Great Britain and Northern Ireland.

Progressive grey matter atrophy in clinically early relapsing-remitting multiple sclerosis.

Brain atrophy appears to occur in patients with multiple sclerosis (MS) in excess of that associated with normal ageing, and may be observed early in the clinical course of the disease. The dynamics and tissue specificity of this process remain unclear. This preliminary study explored the evolution of brain grey matter (GM) and white matter (WM) volume loss (as fractions of total intracranial volumes) in 13 subjects with relapsing-remitting MS (mean disease duration 1.9 years at first scan), compared with nine normal control (NC) subjects. Subjects were scanned every six months for 18 months. In MS compared with NC subjects, significant differences in WM fractional volumes were observed at baseline (mean-5.8%, P/0.008) but no apparent progressive WM tissue loss was detected. In contrast, while no significant differences in GM fractional volumes were observed at baseline, there was significantly greater time-related volume loss in MS compared with NC subjects over the follow-up period (circa-0.0086 per year in MS subjects,-0.0021 per year in the NC subjects, difference 0.010). These results suggest that while both GM and WM atrophy are seen early in the clinical course of MS, they may not occur concurrently and may evolve at different rates.

Effects of intravenous methylprednisolone on outcome in MRI-based prognostic subgroups in acute optic neuritis

Treatment of acute optic neuritis with steroids has been shown to hasten visual recovery without affecting the final degree of recovery. However, MRI-clinical studies indicate that patients with long optic nerve lesions, particularly those that involve the nerve within the optic canal, may have a worse prognosis for recovery of vision. Partly because such lesions could lead to swelling and subsequent ischemic optic nerve damage, steroids could have a selective beneficial effect on this subgroup of patients. The present randomized trial was designed to test this possibility. Sixty-six patients with acute optic neuritis received IV saline or IV methylprednisolone. The clinical, psychophysical, electrophysiologic, and MRI outcomes were assessed after 6 months. Patients with short lesions presented earlier than those with long lesions (involving three or more 5-mm-thick slices of any part of the optic nerve, as well as its intracanalicular portion), and lesion length was significantly less in patients presenting within a week of onset of symptoms. Lesions also tended to lengthen during follow-up in individual patients. Treatment did not limit lesion length in either the long or short lesion subgroup and had no significant effect on final visual outcome. We conclude that steroids do not improve visual outcome or lesion length in patients with acute optic neuritis.

Increasing normal–appearing grey and white matter magnetisation transfer ratio abnormality in early relapsing–remitting multiple sclerosis

Abnormalities within normal–appearing grey and white matter (NAGM and NAWM) occur early in the clinical course of multiple sclerosis (MS) and can be detected in–vivo using the magnetisation transfer ratio (MTR). To better characterize the rates of change in both tissues and to ascertain when such changes begin, we serially studied a cohort of minimally disabled, early relapsing–remitting MS patients, using NAGM and NAWM MTR histograms. Twenty–three patients with clinically definite early relapsing–remitting MS (mean disease duration at baseline 1.9 years), and 19 healthy controls were studied. A magnetisation transfer imaging sequence was acquired yearly for two years. Twenty–one patients and 10 controls completed followup. NAWM and NAGM MTR histograms were derived and mean MTR calculated. A hierarchical regression analysis, adjusting for brain parenchymal fraction,was used to assess MTR change over time. MS NAWM and NAGM MTR were significantly reduced in comparison with controls at baseline and, in patients, both measures decreased further during follow–up: (–0.10pu/year, p = 0.001 and –0.18pu/year, p < 0.001 respectively). The rate of MTR decrease was significantly greater in NAGM than NAWM (p = 0.004). Under the assumption that such changes are linear, backward extrapolation of the observed rates of change suggested that NAWM abnormality began before symptom onset. We conclude that increasing MTR abnormalities in NAWM and NAGM are observed early in the course of relapsing–remitting MS. It is now important to investigate whether these measures are predictive of future disability, and consequently, whether MTR could be used as a surrogate marker in therapeutic trials.

Evidence for grey matter MTR abnormality in minimally disabled patients with early relapsing-remitting multiple sclerosis

Objectives: To establish whether magnetisation transfer ratio (MTR) histograms are sensitive to change in normal appearing grey matter (NAGM) in early relapsing-remitting multiple sclerosis (RRMS) in the absence of significant disability; and to assess whether grey or white matter MTR measures are associated with clinical measures of impairment in early RRMS Methods: 38 patients were studied (mean disease duration 1.9 years (range 0.5 to 3.7); median expanded disability status scale (EDSS) 1.5 (0 to 3)), along with 35 healthy controls. MTR was determined from proton density weighted images with and without MT presaturation. SPM99 was used to generate normal appearing white matter (NAWM) and NAGM segments of the MTR map, and partial voxels were minimised with a 10 pu threshold and voxel erosions. Mean MTR was calculated from the tissue segments. Atrophy measures were determined using a 3D fast spoiled gradient recall sequence from 37 patients and 17 controls. Results: Mean NAGM and NAWM MTR were both reduced in early RRMS (NAGM MTR: 31.9 pu in patients v 32.2 pu in controls; p<0.001; NAWM MTR: 37.9 v 38.3 pu, p = 0.001). Brain parenchymal fraction (BPF) correlated with NAGM MTR, but when BPF was included as a covariate NAGM MTR was still lower in the patients (p = 0.009). EDSS correlated with NAGM MTR (r = 0.446 p = 0.005). Conclusions: In early RRMS, grey matter MTR abnormality is apparent. The correlation with mild clinical impairment (in this essentially non-disabled cohort) suggests that NAGM MTR could be a clinically relevant surrogate marker in therapeutic trials.

Phenytoin for neuroprotection in patients with acute optic neuritis: a randomised, placebo-controlled, phase 2 trial

BACKGROUND Acute demyelinating optic neuritis, a common feature of multiple sclerosis, can damage vision through neurodegeneration in the optic nerve and in its fibres in the retina. Inhibition of voltage-gated sodium channels is neuroprotective in preclinical models. In this study we aimed to establish whether sodium-channel inhibition with phenytoin is neuroprotective in patient with acute optic neuritis. METHODS We did a randomised, placebo-controlled, double-blind phase 2 trial at two UK academic hospitals in London and Sheffield. Patients with acute optic neuritis aged 18-60 years, presenting within 2 weeks of onset, with visual acuity of 6/9 or worse, were randomly assigned (1:1) by minimisation via a web-based service to oral phenytoin (maintenance dose 4 mg/kg per day if randomised before or on July 16, 2013, and 6 mg/kg per day if randomised on or after July 17, 2013) or placebo for 3 months, stratified by time from onset, centre, previous multiple sclerosis diagnosis, use of disease-modifying treatment, and use of corticosteroids for acute optic neuritis. Participants and treating and assessing physicians were masked to group assignment. The primary outcome was retinal nerve fibre layer (RNFL) thickness in the affected eye at 6 months, adjusted for fellow-eye RNFL thickness at baseline, analysed in a modified intention-to-treat population of all randomised participants who were followed up at 6 months. Safety was analysed in the entire population, including those who were lost to follow-up. The trial is registered with ClinicalTrials.gov, number NCT 01451593. FINDINGS We recruited 86 participants between Feb 3, 2012, and May 22, 2014 (42 assigned to phenytoin and 44 to placebo). 29 were assigned to phenytoin 4 mg/kg and 13 to phenytoin 6 mg/kg. Five participants were lost to follow-up, so the primary analysis included 81 participants (39 assigned to phenytoin and 42 to placebo). Mean 6-month RNFL thickness in the affected eye at 6 months was 81.46 μm (SD 16.27) in the phenytoin group (a mean decrease of 16.69 μm [SD 13.73] from baseline) versus 74.29 μm (15.14) in the placebo group (a mean decrease of 23.79 μm [13.97] since baseline; adjusted 6-month difference of 7.15 μm [95% CI 1.08-13.22]; p=0.021), corresponding to a 30% reduction in the extent of RNFL loss with phenytoin compared with placebo. Treatment was well tolerated, with five (12%) of 42 patients having a serious adverse event in the phenytoin group (only one, severe rash, was attributable to phenytoin) compared with two (5%) of 44 in the placebo group. INTERPRETATION These findings support the concept of neuroprotection with phenytoin in patients with acute optic neuritis at concentrations at which it blocks voltage-gated sodium channels selectively. Further investigation in larger clinical trials in optic neuritis and in relapsing multiple sclerosis is warranted. FUNDING US National Multiple Sclerosis Society, Multiple Sclerosis Society of Great Britain and Northern Ireland, Novartis, UK National Institute for Health Research (NIHR), and NIHR UCLH/UCL Biomedical Research Centre.