Microglia and infiltrating leukocytes are considered major producers of tumor necrosis factor (TNF), which is a crucial player in cerebral ischemia and brain inflammation. We have identified a neuroprotective role for microglial-derived TNF in cerebral ischemia in mice. We show that cortical infarction and behavioral deficit are significantly exacerbated in TNF-knock-out (KO) mice compared with wild-type mice. By using in situ hybridization, immunohistochemistry, and green fluorescent protein bone marrow (BM)-chimeric mice, TNF was shown to be produced by microglia and infiltrating leukocytes. Additional analysis demonstrating that BM-chimeric TNF-KO mice grafted with wild-type BM cells developed larger infarcts than BM-chimeric wild-type mice grafted with TNF-KO BM cells provided evidence that the neuroprotective effect of TNF was attributable to microglial-not leukocyte-derived TNF. In addition, observation of increased infarction in TNF-p55 receptor (TNF-p55R)-KO mice compared with TNF-p75R and wild-type mice suggested that microglialderived TNF exerts neuroprotective effects through TNF-p55R. We finally report that TNF deficiency is associated with reduced microglial population size and Toll-like receptor 2 expression in unmanipulated brain, which might also influence the neuronal response to injury. Our results identify microglia and microglial-derived TNF as playing a key role in determining the survival of endangered neurons in cerebral ischemia.

Reactive gliosis is a prominent feature of neurodegenerative and neuroinflammatory disease in the CNS, yet the stimuli that drive this response are not known. There is growing appreciation that signaling through Toll-like receptors (TLRs), which is key to generating innate responses to infection, may have pathogen-independent roles. We show that TLR2 was selectively upregulated by microglia in the denervated zones of the hippocampus in response to stereotactic transection of axons in the entorhinal cortex. In mice lacking TLR2, there were transient, selective reductions in lesion-induced expression of cytokines and chemokines. Recruitment of T cells, but not macrophages, was delayed in TLR2-deficient mice, as well as in mice lacking TNFR1 (tumor necrosis factor receptor 1). TLR2 deficiency also affected microglial proliferative expansion, whereas all of these events were unaffected in TLR4-mutant mice. Consistent with the fact that responses in knock-out mice had all returned to wild-type levels by 8 d, there was no evidence for effects on neuronal plasticity at 20 d. These results identify a role for TLR2 signaling in the early glial response to brain injury, acting as an innate bridge to neuroinflammation.

ObjectiveAntibodies to cell surface central nervous system proteins help to diagnose conditions which often respond to immunotherapies. The assessment of antibody assays needs to reflect their clinical utility. We report the results of a multicentre study of aquaporin (AQP) 4 antibody (AQP4-Ab) assays in neuromyelitis optica spectrum disorders (NMOSD).MethodsCoded samples from patients with neuromyelitis optica (NMO) or NMOSD (101) and controls (92) were tested at 15 European diagnostic centres using 21 assays including live (n=3) or fixed cell-based assays (n=10), flow cytometry (n=4), immunohistochemistry (n=3) and ELISA (n=1).ResultsResults of tests on 92 controls identified 12assays as highly specific (0–1 false-positive results). 32 samples from 50 (64%) NMO sera and 34 from 51 (67%) NMOSD sera were positive on at least two of the 12 highly specific assays, leaving 35 patients with seronegative NMO/spectrum disorder (SD). On the basis of a combination of clinical phenotype and the highly specific assays, 66 AQP4-Ab seropositive samples were used to establish the sensitivities (51.5–100%) of all 21 assays. The specificities (85.8–100%) were based on 92 control samples and 35 seronegative NMO/SD patient samples.ConclusionsThe cell-based assays were most sensitive and specific overall, but immunohistochemistry or flow cytometry could be equally accurate in specialist centres. Since patients with AQP4-Ab negative NMO/SD require different management, the use of both appropriate control samples and defined seronegative NMOSD samples is essential to evaluate these assays in a clinically meaningful way. The process described here can be applied to the evaluation of other antibody assays in the newly evolving field of autoimmune neurology.

ObjectivesIn a prospective phase IV trial of the first-line oral treatment dimethyl fumarate (DMF), we examined dynamics of neurofilament light (NFL) chain in serum, plasma and cerebrospinal fluid (CSF) samples collected over 12 months from relapsing-remitting multiple sclerosis (RRMS) patients. NFL changes were related to disease activity.MethodsWe examined NFL levels by single-molecule array in 88 CSF, 348 plasma and 131 sera from treatment-naïve RRMS patients (n=52), healthy controls (n=23) and a placebo group matched by age, sex and NFL (n=52). Plasma/sera were collected at baseline, and 1, 3, 6 and 12 months after DMF. CSF samples were collected at baseline and 12 months after DMF.ResultsNFL concentration in CSF, plasma and serum correlated highly (p<0.0001 for all), but plasma levels were only 76.9% of paired serum concentration. After 12 months of DMF treatment, NFL concentration decreased by 73%, 69% and 55% in the CSF, serum and plasma (p<0.0001, respectively). Significant reduction in blood was observed after 6 and 12 months treatment compared with baseline (p<0.01 and p<0.0001, respectively) and to placebo (p<0.0001). Patients with NFL above the 807.5 pg/mL cut-off in CSF had 5.0-times relative risk of disease activity (p<0.001).ConclusionsThis study provides Class II evidence that first-line DMF reduces NFL in both blood and CSF after 6 months and normalises CSF levels in 73% of patients. High NFL concentration in CSF after a year reflected disease activity. NFL levels were higher in serum than in plasma, which should be considered when NFL is used as a biomarker.