Blair Wood

Cyclooxygenase-2 expression in oligodendrocytes increases sensitivity to excitotoxic death

BackgroundWe previously found that cyclooxygenase 2 (COX-2) was expressed in dying oligodendrocytes at the onset of demyelination in the Theilers murine encephalomyelitis virus-induced demyelinating disease (TMEV-IDD) model of multiple sclerosis (MS) (Carlson et al. J.Neuroimmunology 2006, 149:40). This suggests that COX-2 may contribute to death of oligodendrocytes.ObjectiveThe goal of this study was to examine whether COX-2 contributes to excitotoxic death of oligodendrocytes and potentially contributes to demyelination.MethodsThe potential link between COX-2 and oligodendrocyte death was approached using histopathology of MS lesions to examine whether COX-2 was expressed in dying oligodendrocytes. COX-2 inhibitors were examined for their ability to limit demyelination in the TMEV-IDD model of MS and to limit excitotoxic death of oligodendrocytes in vitro. Genetic manipulation of COX-2 expression was used to determine whether COX-2 contributes to excitotoxic death of oligodendrocytes. A transgenic mouse line was generated that overexpressed COX-2 in oligodendrocytes. Oligodendrocyte cultures derived from these transgenic mice were used to examine whether increased expression of COX-2 enhanced the vulnerability of oligodendrocytes to excitotoxic death. Oligodendrocytes derived from COX-2 knockout mice were evaluated to determine if decreased COX-2 expression promotes a greater resistance to excitotoxic death.ResultsCOX-2 was expressed in dying oligodendrocytes in MS lesions. COX-2 inhibitors limited demyelination in the TMEV-IDD model of MS and protected oligodendrocytes against excitotoxic death in vitro. COX-2 expression was increased in wild-type oligodendrocytes following treatment with Kainic acid (KA). Overexpression of COX-2 in oligodendrocytes increased the sensitivity of oligodendrocytes to KA-induced excitotoxic death eight-fold compared to wild-type. Conversely, oligodendrocytes prepared from COX-2 knockout mice showed a significant decrease in sensitivity to KA induced death.ConclusionsCOX-2 expression was associated with dying oligodendrocytes in MS lesions and appeared to increase excitotoxic death of oligodendrocytes in culture. An understanding of how COX-2 expression influences oligodendrocyte death leading to demyelination may have important ramifications for future treatments for MS.

Anti-Yo Antibody Uptake and Interaction with Its Intracellular Target Antigen Causes Purkinje Cell Death in Rat Cerebellar Slice Cultures: A Possible Mechanism for Paraneoplastic Cerebellar Degeneration in Humans with Gynecological or Breast Cancers

Anti-Yo antibodies are immunoglobulin G (IgG) autoantibodies reactive with a 62 kDa Purkinje cell cytoplasmic protein. These antibodies are closely associated with paraneoplastic cerebellar degeneration in the setting of gynecological and breast malignancies. We have previously demonstrated that incubation of rat cerebellar slice cultures with patient sera and cerebrospinal fluid containing anti-Yo antibodies resulted in Purkinje cell death. The present study addressed three fundamental questions regarding the role of anti-Yo antibodies in disease pathogenesis: 1) Whether the Purkinje cell cytotoxicity required binding of anti-Yo antibody to its intraneuronal 62 kDa target antigen; 2) whether Purkinje cell death might be initiated by antibody-dependent cellular cytotoxicity rather than intracellular antibody binding; and 3) whether Purkinje cell death might simply be a more general result of intracellular antibody accumulation, rather than of specific antibody-antigen interaction. In our study, incubation of rat cerebellar slice cultures with anti-Yo IgG resulted in intracellular antibody binding, and cell death. Infiltration of the Purkinje cell layer by cells of macrophage/microglia lineage was not observed until extensive cell death was already present. Adsorption of anti-Yo IgG with its 62 kDa target antigen abolished both antibody accumulation and cytotoxicity. Antibodies to other intracellular Purkinje cell proteins were also taken up by Purkinje cells and accumulated intracellularly; these included calbindin, calmodulin, PCP-2, and patient anti-Purkinje cell antibodies not reactive with the 62 kDa Yo antigen. However, intracellular accumulation of these antibodies did not affect Purkinje cell viability. The present study is the first to demonstrate that anti-Yo antibodies cause Purkinje cell death by binding to the intracellular 62 kDa Yo antigen. Anti-Yo antibody cytotoxicity did not involve other antibodies or factors present in patient serum and was not initiated by brain mononuclear cells. Purkinje cell death was not simply due to intraneuronal antibody accumulation.

Neuronal uptake of anti-Hu antibody, but not anti-Ri antibody, leads to cell death in brain slice cultures

BackgroundAnti-Hu and anti-Ri antibodies are paraneoplastic immunoglobulin (Ig)G autoantibodies which recognize cytoplasmic and nuclear antigens present in all neurons. Although both antibodies produce similar immunohistological labeling, they recognize different neuronal proteins. Both antibodies are associated with syndromes of central nervous system dysfunction. However, the neurological deficits associated with anti-Hu antibody are associated with neuronal death and are usually irreversible, whereas neurological deficits in patients with anti-Ri antibody may diminish following tumor removal or immunosuppression.MethodsTo study the effect of anti-Hu and anti-Ri antibodies on neurons, we incubated rat hippocampal and cerebellar slice cultures with anti-Hu or anti-Ri sera from multiple patients. Cultures were evaluated in real time for neuronal antibody uptake and during prolonged incubation for neuronal death. To test the specificity of anti-Hu antibody cytotoxic effect, anti-Hu serum IgG was incubated with rat brain slice cultures prior to and after adsorption with its target Hu antigen, HuD.ResultsWe demonstrated that: 1) both anti-Hu and anti-Ri antibodies were rapidly taken up by neurons throughout both cerebellum and hippocampus; 2) antibody uptake occurred in living neurons and was not an artifact of antibody diffusion into dead cells; 3) intracellular binding of anti-Hu antibody produced neuronal cell death, whereas uptake of anti-Ri antibody did not affect cell viability during the period of study; and 4) adsorption of anti-Hu antisera against HuD greatly reduced intraneuronal IgG accumulation and abolished cytotoxicity, confirming specificity of antibody-mediated neuronal death.ConclusionsBoth anti-Hu and anti-Ri antibodies were readily taken up by viable neurons in slice cultures, but the two antibodies differed markedly in terms of their effects on neuronal viability. The ability of anti-Hu antibodies to cause neuronal death could account for the irreversible nature of paraneoplastic neurological deficits in patients with this antibody response. Our results raise questions as to whether anti-Ri antibody might initially induce reversible neuronal dysfunction, rather than causing cell death. The ability of IgG antibodies to access and react with intracellular neuronal proteins could have implications for other autoimmune diseases involving the central nervous system.

The role of the prostaglandin E2 receptors in vulnerability of oligodendrocyte precursor cells to death

BackgroundActivity of cyclooxygenase 2 (COX-2) in mouse oligodendrocyte precursor cells (OPCs) modulates vulnerability to excitotoxic challenge. The mechanism by which COX-2 renders OPCs more sensitive to excitotoxicity is not known. In the present study, we examined the hypothesis that OPC excitotoxic death is augmented by COX-2-generated prostaglandin E2 (PGE2) acting on specific prostanoid receptors which could contribute to OPC death.MethodsDispersed OPC cultures prepared from mice brains were examined for expression of PGE2 receptors and the ability to generate PGE2 following activation of glutamate receptors with kainic acid (KA). OPC death in cultures was induced by either KA, 3′-O-(Benzoyl) benzoyl ATP (BzATP) (which stimulates the purinergic receptor P2X7), or TNFα, and the effects of EP3 receptor agonists and antagonists on OPC viability were examined.ResultsStimulation of OPC cultures with KA resulted in nearly a twofold increase in PGE2. OPCs expressed all four PGE receptors (EP1–EP4) as indicated by immunofluorescence and Western blot analyses; however, EP3 was the most abundantly expressed. The EP3 receptor was identified as a candidate contributing to OPC excitotoxic death based on pharmacological evidence. Treatment of OPCs with an EP1/EP3 agonist 17 phenyl-trinor PGE2 reversed protection from a COX-2 inhibitor while inhibition of EP3 receptor protected OPCs from excitotoxicity. Inhibition with an EP1 antagonist had no effect on OPC excitotoxic death. Moreover, inhibition of EP3 was protective against toxic stimulation with KA, BzATP, or TNFα.ConclusionTherefore, inhibitors of the EP3 receptor appear to enhance survival of OPCs following toxic challenge and may help facilitate remyelination.

Voltage-gated calcium channel autoimmune cerebellar degeneration: Case and study of cytotoxicity.

Objectives: To describe response to treatment in a patient with autoantibodies against voltage-gated calcium channels (VGCCs) who presented with autoimmune cerebellar degeneration and subsequently developed Lambert-Eaton myasthenic syndrome (LEMS), and to study the effect of the patients autoantibodies on Purkinje cells in rat cerebellar slice cultures. Methods: Case report and study of rat cerebellar slice cultures incubated with patient VGCC autoantibodies. Results: A 53-year-old man developed progressive incoordination with ataxic speech. Laboratory evaluation revealed VGCC autoantibodies without other antineuronal autoantibodies. Whole-body PET scans 6 and 12 months after presentation detected no malignancy. The patient improved significantly with IV immunoglobulin G (IgG), prednisone, and mycophenolate mofetil, but worsened after IV IgG was halted secondary to aseptic meningitis. He subsequently developed weakness with electrodiagnostic evidence of LEMS. The patients IgG bound to Purkinje cells in rat cerebellar slice cultures, followed by neuronal death. Reactivity of the patients autoantibodies with VGCCs was confirmed by blocking studies with defined VGCC antibodies. Conclusions: Autoimmune cerebellar degeneration associated with VGCC autoantibodies may precede onset of LEMS and may improve with immunosuppressive treatment. Binding of anti-VGCC antibodies to Purkinje cells in cerebellar slice cultures may be followed by cell death. Patients with anti-VGCC autoantibodies may be at risk of irreversible neurologic injury over time, and treatment should be initiated early.

Comparative neuronal uptake and cytotoxicity of anti-Hu and anti-Ri antibodies in rat cerebellar and hippocampal slice cultures

WCN 2013 No: 1897 Topic: 36 — Other Topic Comparative neuronal uptake and cytotoxicity of anti-Hu and anti-Ri antibodies in rat cerebellar and hippocampal slice cultures J.E. Greenlee, S.A. Clawson, B. Wood, K.E. Hill, N.G. Carlson. Neurology Service, Veterans Affairs Medical Center, Salt Lake City, UT, USA; Neurology, University of Utah Health Sciences Center, Salt Lake City, UT, USA; Research Service, Veterans Affairs Medical Center, Salt Lake City, UT, USA; Neurobiology and Anatomy, University of Utah School of Medicine, Salt Lake City, UT, USA; Center on Aging, University of Utah School of Medicine, Salt Lake City, UT, USA Background: Anti-Hu and anti-Ri are paraneoplastic autoantibodies recognizing intracellular antigens present in essentially all neurons. At autopsy, brains of patients with anti-Hu antibody show neuronal destruction. In contrast, anti-Ri antibody is less clearly associated with neuronal death, and patients with anti-Ri antibodies may respond to treatment. We have demonstrated that anti-Yo antibodies, associated with paraneoplastic cerebellar degeneration, accumulate intracellularly in cerebellar Purkinje cells in slice cultures of rat cerebellum and that antibody accumulation is followed by cell death. The present study was conducted to determine whether anti-Hu and anti-Ri antibodies are also taken up by neurons and whether uptake of either antibody is cytotoxic. Objective: To evaluate neuronal uptake and cytotoxicity of anti-Hu and anti-Ri antibodies in slice cultures of rat cerebellum and hippocampus. Materials and methods: Rat cerebellar and hippocampal slice cultures were incubated with anti-Hu or anti-Ri antibodies and evaluated over time for antibody uptake and for cell death. Specificity of anti-Hu cytotoxicity was confirmed by adsorbing anti-Hu IgGwith recombinant HuD protein. Results: Anti-Hu and anti-Ri antibodies accumulated in cerebellar and hippocampal neurons. Anti-Hu antibodies produced cell death which was significantly reduced by adsorption of anti-Hu IgG with recombinant HuD protein. In contrast, neurons accumulating anti-Ri antibodies showed no evidence of cell death as compared to controls. Conclusions: Anti-Hu and anti-Ri antibodies entered and accumulated in cerebellar and hippocampal neurons. Anti-Hu antibody associated neuronal death involved reactivity with HuD protein. Anti-Ri antibody did not affect neuronal viability and may cause neuronal dysfunction rather than cell death. doi:10.1016/j.jns.2013.07.2239 Abstract — WCN 2013 No: 1902 Topic: 36 — Other Topic An evaluation of the risks involved in ischemic encephalopathy among Mongolian children T. Sosorburam, B. Batbayar. Anesthesiology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China; Children Hospital, Ulan-Bator, Mongolia Background: Ischemic encephalopathy is one of the emerging issues among third world countries. Hypoxia seems to be the main cause of this problem, but many risk factors are associated in causation of hypoxic ischemic encephalopathy (HIE). The aim of our study was to evaluate these risk factors among Mongolian children. Methods: This study was carried out at 3 childrens hospitals, 85 neonates diagnosed with HIE over a year. Efforts were made to evaluate the questionnaire including details on parity, pre and postnatal histories, and specially the events of birth from labour till delivery of placenta. Results: There was a remarkable decreased antenatal hospital visits and almost 52% reported that they visited just once or twice during the whole pregnancy period. In 21%, there was history of increased mother age above 40 years while 62% reported that some incidents of hypoxia occurred at the time of birth. 20% used drugs or medication during the course of pregnancy without the consultation of the physician.31 % had a history of prolonged 2nd stage of labour. 49% were delivered by unskilled birth attendants. Conclusion: The data clearly showed that there are risks that can be avoided easily and a leading cause of mental retardation can be prevented. Lack of health awareness and decreased consultation from physicians on time seems to be the bulk of the problem in addition to lack of trained birth staff and health facilities. Efforts should be made to educate the mothers of child bearing ages, and counsel them to make the antenatal hospital visits more frequent as possible. doi:10.1016/j.jns.2013.07.2240 Abstract — WCN 2013 No: 1907 Topic: 36 — Other Topic Different emotions and strong stimuli: How do we make choices? WCN 2013 No: 1907 Topic: 36 — Other Topic Different emotions and strong stimuli: How do we make choices? C.V.D.S. Vilharba, A.J. Godoy. Marketing, University Sao Judas, Sao Paulo, Brazil; Neurology, University City of Sao Paulo, Sao Paulo, Brazil Our understanding of the way the brain makes a decision is still incomplete. We decided to analyze the relationships between color and different emotions and sound and different emotions and their influences to make a choice. Forty-eight undergraduate students of marketing courses were interviewed. We presented 7 objects with different colors while they were listening to a “sad” song and again while listening to a “happy” song. In each case they had to choose one of them. Then we presented six identical objects associated with different sounds while listening to those songs. Forty-six percent of the students selected the red color during the sad moment (SM) and 40% during the happy moment (HM). None of them chose white during SM (however 10% selected white during HM). During HM the “forgotten” color was orange (2% of the students). Twenty-nine percent of the volunteers chose the object associated with the sound of a bomb during SM (the same sound was selected only by 12% during HM). Only 2% selected the sound of the wind (during SM or HM). Our results suggest that strong stimuli (visual or auditory) activate many neural circuits, making someone choose a red object or one associated with the sound of a bomb. Nevertheless a so called Abstracts / Journal of the Neurological Sciences e629 (2013) e629–e678 e645