Sema4A is implicated in the acceleration of Th17 cell-mediated neuroinflammation in the effector phase.

BACKGROUND: Sema4A is a regulator of helper T cell (Th) activation and differentiation in the priming phase, which plays an important role in the pathogenesis of experimental autoimmune encephalomyelitis (EAE) and multiple sclerosis (MS). However, the role of Sema4A in the effector phase remains elusive. We aimed to investigate the role of Sema4A at the effector phase in adoptively transferred EAE model. Clinical features and cytokine profiles of MS patients with high Sema4A levels were also examined in detail to clarify the correlation between Sema4A levels and disease activity of patients with MS. METHODS: We adoptively transferred encephalitogenic Th1 or Th17 cells to wild type (WT) or Sema4A-deficient (Sema4A KO) mice and assessed severity of symptoms and cellular infiltration within the central nervous system (CNS). In addition, we analyzed clinical and radiological features (n = 201), levels of serum IFN-γ and IL-17A (n = 86), complete remission ratio by IFN-ß (n = 38) in all of relapsing-remitting multiple sclerosis (RRMS) patients enrolled in this study. RESULTS: Sema4A KO recipient mice receiving Th17-skewed WT myelin oligodendrocyte glycoprotein (MOG)-specific encephalitogenic T cells showed a significant reduction in the clinical score compared to the WT recipient mice. However, Sema4A KO recipient mice showed similar disease activity to the WT recipient mice when transferred with Th1-skewed encephalitogenic T cells. Bone marrow chimeric study indicated that Sema4A expressed on hematopoietic cells, but not the CNS resident cells, are responsible for augmenting Th17-mediated neuroinflammation. Additionally, in contrast to comparable IFN-γ levels, IL-17A is significantly higher in RRMS patients with high Sema4A level than those with low Sema4A patients with high Sema4A levels showed earlier disease onset, more severe disease activity and IFN-ß unresponsiveness than those with low Sema4A levels. CONCLUSIONS: Sema4A is involved not only in the Th cell priming but also in the acceleration of Th17 cell-mediated neuroinflammation in the effector phase, which could contribute to the higher disease activity observed in RRMS patients with high serum Sema4A levels.

Single injection of sustained-release prostacyclin analog ONO-1301-MS ameliorates hypoxic toxicity in the murine model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized by several pathologies including oxidative stress, apoptosis, neuroinflammation, and glutamate toxicity. Although multiple reports suggest that ischemia and hypoxia in the spinal cord plays a pivotal role in the pathogenesis of ALS, the precise role of hypoxia in disease progression remains unknown. In this study, we detected higher expression levels of Hypoxia-inducible factor 1-alpha (HIF-1α), a key regulator of cellular responses to hypoxia, in the spinal cord of ALS patients and in the transgenic mice overexpressing the familial ALS-associated G93A SOD1 mutation (mSOD1G93A mice) compared to controls. Single subcutaneous administration of sustained-release prostacyclin analog ONO-1301-MS to mSOD1G93A mice abrogated the expression of HIF-1α in their spinal cords, as well as erythropoietin (EPO) and vascular endothelial growth factor (VEGF), both of which are downstream to HIF-1α. Furthermore, ONO-1301-MS increased the level of mature brain-derived neurotrophic factor (BDNF) and ATP production in the spinal cords of mSOD1G93A mice. At late disease stages, the motor function and the survival of motor neurons of ONO-1301-MS-treated mSOD1G93A mice was significantly improved compared to vehicle-treated mSOD1G93A mice. Our data suggest that vasodilator therapy modulating local blood flow in the spinal cord has beneficial effects against ALS disease progression.

Cerebrospinal fluid mitochondrial DNA in neuromyelitis optica spectrum disorder.

BACKGROUND: Neuromyelitis optica spectrum disorder (NMOSD) is an inflammatory disease of the central nervous system. Although complement-dependent astrocyte damage mediated by anti-aquaporin 4 autoantibody (AQP4-Ab) is well acknowledged to be the core of NMOSD pathogenesis, additional inflammatory cascades may contribute to the establishment of lesion formation. Thus, in this study, we investigated the possible pathogenic role of immune-reactive mitochondrial DNA (mtDNA) in cerebrospinal fluid (CSF) of NMOSD patients. METHODS: Using quantitative polymerase chain reaction, we measured extracellular mtDNA levels in CSF of NMOSD patients positive for AQP4-Ab. Patients with multiple sclerosis or other neurological diseases were examined as controls. Pre- and post-treatment extracellular mtDNA levels were also compared in the NMOSD group. Extracellular mtDNA release from human astrocytes was analyzed in vitro utilizing NMOSD sera, and interleukin (IL)-1ß production was measured in supernatants of mixed glial cells stimulated with DNA fraction of CSF derived from NMOSD patients. Furthermore, specific innate immune pathways mediating the IL-1ß production by mtDNA were investigated in peripheral blood mononuclear cells with selective inhibitors of Toll-like receptor 9 (TLR9) and NOD-like receptor protein 3 (NLRP3) inflammasomes. RESULTS: Extracellular mtDNA level was specifically elevated in acute phase of NMOSD CSF. In vitro studies provided the evidence that mtDNA is released from human astrocytes by NMOSD sera. In addition, DNA fraction isolated from NMOSD CSF promoted secretion of IL-1ß from mixed glial cells. Selective inhibition of TLR9 and NLRP3 inflammasomes revealed that mtDNA-mediated IL-1ß production depends on specific innate immune pathways. CONCLUSION: Extracellular mtDNA is specifically elevated in the CSF of patients with acute phase NMOSD, and mtDNA released by AQP4-Ab-mediated cellular damage elicits the innate immune cascades via TLR9 and NLRP3 inflammasomes pathways. Our study highlights mtDNA-mediated innate immune pathways as a novel therapeutic target for future treatment of NMOSD patients.

Beneficial effects of fingolimod in MS patients with high serum Sema4A levels.

We previously demonstrated that patients with multiple sclerosis (MS) of high serum Sema4A levels are resistant to IFN-ß therapy. To further elucidate the role of serum Sema4A as a biomarker for therapeutic stratification in MS patients, it is important to clarify the efficacy of other disease-modifying drugs (DMD) in those with high serum Sema4A levels. Thus, in this study we investigated whether fingolimod has beneficial effects on MS patients with high Sema4A levels. We retrospectively analyzed annualized relapse rate (ARR) and Expanded Disability Status Scale (EDSS) change in 56 relapsing-remitting multiple sclerosis (RRMS) patients who had been treated with fingolimod, including those who switched from IFN-ß therapy. The levels of Sema4A in the sera were measured by sandwich ELISA. The implications of Sema4A on the efficacy of fingolimod were investigated by administering recombinant Sema4A-Fc and fingolimod to mice with experimental autoimmune encephalomyelitis (EAE). Retrospective analysis of MS cohort (17 high Sema4A and 39 low Sema4A) demonstrated the effectiveness of fingolimod in those with high serum Sema4A levels, showing reduction of ARR (from 1.21 to 0.12) and EDSS progression (from 0.50 to 0.04). Consistent with this observation, improvement in the disease severity of EAE mice receiving recombinant Sema4A-Fc was also observed after fingolimod treatment. These data suggest that fingolimod could serve as a candidate DMD for managing the disease activity of MS patients with high Sema4A levels.

Febuxostat ameliorates secondary progressive experimental autoimmune encephalomyelitis by restoring mitochondrial energy production in a GOT2-dependent manner.

Oxidative stress and mitochondrial dysfunction are important determinants of neurodegeneration in secondary progressive multiple sclerosis (SPMS). We previously showed that febuxostat, a xanthine oxidase inhibitor, ameliorated both relapsing-remitting and secondary progressive experimental autoimmune encephalomyelitis (EAE) by preventing neurodegeneration in mice. In this study, we investigated how febuxostat protects neuron in secondary progressive EAE. A DNA microarray analysis revealed that febuxostat treatment increased the CNS expression of several mitochondria-related genes in EAE mice, most notably including GOT2, which encodes glutamate oxaloacetate transaminase 2 (GOT2). GOT2 is a mitochondrial enzyme that oxidizes glutamate to produce α-ketoglutarate for the Krebs cycle, eventually leading to the production of adenosine triphosphate (ATP). Whereas GOT2 expression was decreased in the spinal cord during the chronic progressive phase of EAE, febuxostat-treated EAE mice showed increased GOT2 expression. Moreover, febuxostat treatment of Neuro2a cells in vitro ameliorated ATP exhaustion induced by rotenone application. The ability of febuxostat to preserve ATP production in the presence of rotenone was significantly reduced by GOT2 siRNA. GOT2-mediated ATP synthesis may be a pivotal mechanism underlying the protective effect of febuxostat against neurodegeneration in EAE. Accordingly, febuxostat may also have clinical utility as a disease-modifying drug in SPMS.

Partial suppression of M1 microglia by Janus kinase 2 inhibitor does not protect against neurodegeneration in animal models of amyotrophic lateral sclerosis.

BACKGROUND: Accumulating evidence has shown that the inflammatory process participates in the pathogenesis of amyotrophic lateral sclerosis (ALS), suggesting a therapeutic potential of anti-inflammatory agents. Janus kinase 2 (JAK2), one of the key molecules in inflammation, transduces signals downstream of various inflammatory cytokines, and some Janus kinase inhibitors have already been clinically applied to the treatment of inflammatory diseases. However, the efficacy of JAK2 inhibitors in treatment of ALS remains to be demonstrated. In this study, we examined the role of JAK2 in ALS by administering a selective JAK2 inhibitor, R723, to an animal model of ALS (mSOD1G93A mice). FINDINGS: Orally administered R723 had sufficient access to spinal cord tissue of mSOD1G93A mice and significantly reduced the number of Ly6c positive blood monocytes, as well as the expression levels of IFN-γ and nitric oxide synthase 2, inducible (iNOS) in the spinal cord tissue. R723 treatment did not alter the expression levels of Il-1ß, Il-6, TNF, and NADPH oxidase 2 (NOX2), and suppressed the expression of Retnla, which is one of the markers of neuroprotective M2 microglia. As a result, R723 did not alter disease progression or survival of mSOD1G93A mice. CONCLUSIONS: JAK2 inhibitor was not effective against ALS symptoms in mSOD1G93A mice, irrespective of suppression in several inflammatory molecules. Simultaneous suppression of anti-inflammatory microglia with a failure to inhibit critical other inflammatory molecules might explain this result.

[Successful treatment of methicillin-resistant staphylococcus aureus meningitis by intrathecal injection of vancomycin].

A 70-year-old Japanese man developed fever, headache, and lumbago, presumably due to an epidural abscess caused by methicillin-resistant Staphylococcus aureus (MRSA) in the L5-S2 region. On the night of admission to our hospital, he showed disorientation to places and abnormal eating behavior, indicating a complication of MRSA meningitis. Cerebrospinal fluid (CSF) examination confirmed this diagnosis. Although he was treated with venous infusion of vancomycin and meropenem, the CSF culture remained positive for MRSA even a week after the treatment, and Gram-positive cocci were also seen in the CSF. An intrathecal injection of vancomycin (10mg/day) was subsequently added, which resulted in absence of the organism on Gram-stained CSF smear and CSF culture a week later. His condition improved without any adverse effects. Vancomycin cannot freely penetrate the blood-brain barrier (BBB); therefore, when administered intravenously, its concentration in the CSF is insufficient. Therefore, intrathecal injection of vancomycin is necessary to achieve the desired bacteriocidal level in the CSF. Thus, intrathecal administration of vancomycin seems a very effective and safe treatment for MRSA meningitis.