Contributions
Abstract: P780
Type: Poster Sessions
Abstract Category: Pathology and pathogenesis of MS - Neurodegeneration
Introduction: An essential characteristic of MS is multifocal demyelination with relative axonal sparing. Clostridium perfringens epsilon toxin (ETX) is a candidate environmental agent for new lesion formation in MS. ETX is known to alter the blood-brain barrier, but its effects in the central nervous system (CNS) are controversial and still being elucidated.
Objectives: To determine the specificity of ETX for oligodendrocytes/myelin in organotypic cultures and in vivo. To define the molecular mechanism by which ETX causes selective demyelination with sparing of other neural elements.
Aims: To determine if ETX causes demyelination with preservation of axons and other neural elements. To determine if ETX causes demyelination through a glutamate-dependent mechanism as suggested in the literature. To determine if ETX-mediated demyelination requires formation of a heptameric pore. To determine if neutralizing antibodies to ETX prevent demyelination.
Methods: Mouse cerebellum organotypic cultures were treated with ETX in the presence or absence of relevant antagonists or agonists. To improve reliability of this method, we utilized transgenic mice expressing GFP off the PLP promoter such that myelination and demyelination in slices could be followed in real time. Stereotactic injections of ETX or vehicle into rat dorsal columns were performed for in vivo validation.
Results: ETX induced robust demyelination in cerebellar explants. The effect of ETX was completely dependent on formation of a heptameric pore as MβCD, a cholesterol depleting agent, protected against toxin-mediated demyelination. Metabotropic and ionotropic glutamate antagonists had no effect on toxin-mediated demyelination. Antibodies against ETX that block binding to its receptor or prevent heptamerization also prevented demyelination.
Conclusions: ETX causes selective CNS demyelination through generation of a heptameric pore complex on oligodendrocytes/myelin. Our data does not support glutamate toxicity as a mechanism of ETX-mediated demyelination as previously reported. Demyelination with axonal preservation was observed in both organotypic cultures and in vivo models. ETX oligomerization is required for demyelination, as disruption of lipid raft structures and antibodies that inhibit toxin oligomerization prevented demyelination.Taken together, these results support the role of ETX in formation of MS lesions.
Disclosure: Research was generously supported by the National Multiple Sclerosis Society and the National Institutes of Health. Funding was provided by U.S. patent No. 9,758,573, ´Methods to Protect Against and Treat Multiple Sclerosis.´
Abstract: P780
Type: Poster Sessions
Abstract Category: Pathology and pathogenesis of MS - Neurodegeneration
Introduction: An essential characteristic of MS is multifocal demyelination with relative axonal sparing. Clostridium perfringens epsilon toxin (ETX) is a candidate environmental agent for new lesion formation in MS. ETX is known to alter the blood-brain barrier, but its effects in the central nervous system (CNS) are controversial and still being elucidated.
Objectives: To determine the specificity of ETX for oligodendrocytes/myelin in organotypic cultures and in vivo. To define the molecular mechanism by which ETX causes selective demyelination with sparing of other neural elements.
Aims: To determine if ETX causes demyelination with preservation of axons and other neural elements. To determine if ETX causes demyelination through a glutamate-dependent mechanism as suggested in the literature. To determine if ETX-mediated demyelination requires formation of a heptameric pore. To determine if neutralizing antibodies to ETX prevent demyelination.
Methods: Mouse cerebellum organotypic cultures were treated with ETX in the presence or absence of relevant antagonists or agonists. To improve reliability of this method, we utilized transgenic mice expressing GFP off the PLP promoter such that myelination and demyelination in slices could be followed in real time. Stereotactic injections of ETX or vehicle into rat dorsal columns were performed for in vivo validation.
Results: ETX induced robust demyelination in cerebellar explants. The effect of ETX was completely dependent on formation of a heptameric pore as MβCD, a cholesterol depleting agent, protected against toxin-mediated demyelination. Metabotropic and ionotropic glutamate antagonists had no effect on toxin-mediated demyelination. Antibodies against ETX that block binding to its receptor or prevent heptamerization also prevented demyelination.
Conclusions: ETX causes selective CNS demyelination through generation of a heptameric pore complex on oligodendrocytes/myelin. Our data does not support glutamate toxicity as a mechanism of ETX-mediated demyelination as previously reported. Demyelination with axonal preservation was observed in both organotypic cultures and in vivo models. ETX oligomerization is required for demyelination, as disruption of lipid raft structures and antibodies that inhibit toxin oligomerization prevented demyelination.Taken together, these results support the role of ETX in formation of MS lesions.
Disclosure: Research was generously supported by the National Multiple Sclerosis Society and the National Institutes of Health. Funding was provided by U.S. patent No. 9,758,573, ´Methods to Protect Against and Treat Multiple Sclerosis.´