Antigens and mechanisms of immune-mediated encephalitis
BACKGROUND: The autoimmune encephalitis represents a new category of immune- mediated diseases of the brain that are mediated by antibodies against neurotransmitter receptors, ion channels or neuronal cell-surface proteins. Among the many encephalitis considered idiopathic, there are subtypes that a...
| Autor: | |
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| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2018 |
| País: | España |
| Institución: | CBUC, CESCA |
| Repositorio: | TDR. Tesis Doctorales en Red |
| OAI Identifier: | oai:www.tdx.cat:10803/664343 |
| Acceso en línea: | http://hdl.handle.net/10803/664343 |
| Access Level: | acceso abierto |
| Palabra clave: | Neuroimmunologia Neuroinmunología Neuroimmunology Receptors neurals Neurorreceptores Neural receptor Sinapsi Sinapsis Synapses Excitació (Fisiologia) Excitación (Fisiología) Excitation (Physiology) Encefalitis Encephalitis Ciències de la Salut 616.8 |
| Sumario: | BACKGROUND: The autoimmune encephalitis represents a new category of immune- mediated diseases of the brain that are mediated by antibodies against neurotransmitter receptors, ion channels or neuronal cell-surface proteins. Among the many encephalitis considered idiopathic, there are subtypes that are probably immune-mediated and are pending to be discovered. In the last 11 years, 16 subtypes of idiopathic encephalitis have been found to be immune mediated. The recognition of these disorders is important because despite being potentially lethal, they are curable if promptly recognized and treated. On a more basic level, the study of these diseases has uncovered novel antibody-mediated mechanisms of synaptic dysfunction that lead to changes in memory and behavior, epilepsy, abnormal movements, or decreased level of consciousness. OBJECTIVES: 1) Identify patients with encephalitis of unclear etiology but whose clinical features and initial investigations strongly suggest an antibody-mediated cause; 2) characterize the new autoantibodies associated with these disorders along with the neuronal target antigens, and develop unambiguous diagnostic tests, and 3) determine in animal models how autoantibodies alter the level and function of neuronal synaptic antigens (NMDAR and LGI1) potentially resulting in an impairment of memory and behavior. METHODS: Rat brain sections and primary cultures of dissociated rat hippocampal neurons served to identify patients whose serum and CSF samples contained novel neuronal antibodies. Techniques of tissue immunohistochemistry, neuronal immunocytochemistry and cell-based assays were used to identify the antibodies. Cell- surface neuronal protein immunoprecipitation with patients’ antibodies was used to isolate the target antigen, which was subsequently characterized by mass spectrometry. Cerebroventricular transfer of patients’ antibodies to mice through subcutaneously implanted mini-osmotic pumps was used to determine the pathogenic effect of patients’ antibodies on the corresponding synaptic targets and how these changes altered memory and behavior. An extensive combination of techniques was used for these studies, including quantitative confocal microscopy analysis of the levels of synaptic targets, immunoprecipitation of brain-bound antibodies, immunoblot of precipitated proteins, electrophysiology on acute sections of mice hippocampus, and a comprehensive panel of standard behavioral tests. All studies were conducted with sets of mice at different time points during the antibody infusion (disease phase) and after the infusion was stopped (recovery phase). RESULTS: (1) Two novel autoimmune encephalitis were identified: anti-GABAaR encephalitis and anti-neurexin-3α encephalitis. Anti-GABAaR encephalitis can affect children and adults, associates with prominent seizures and highly suggestive MRI abnormalities, and is treatable with immunotherapy. In some patients the immune response is triggered by the presence of a tumor. Anti-neurexin-3α encephalitis manifests with a less distinctive syndrome, often associates with seizures and is also treatable with immunotherapy. No tumor association has been identified. (2) Immunoprecipitation of the target antigen of anti-GABAaR encephalitis demonstrated that the epitopes are mainly located in the alpha1 and beta3 subunits, and less frequently in the γ2 subunit. Whereas the antibodies against alpha1 and beta3 subunits are disease relevant, the presence of additional antibodies to γ2 does not modify the disease phenotype. Patients’ GABAaR antibodies cause a decrease in the total and synaptic levels of GABAaR clusters, supporting their pathogenicity. (3) Immunoprecipitation of the target antigen of patients with anti-neurexin-3α encephalitis demonstrated that the epitopes are specifically located in neurexin-3α, but not in its postsynaptic ligand LRRTM2. Patients’ antibodies cause a reduction of the total number of synapses as well as the levels of the presynaptic protein Bassoon and the post-synaptic protein Homer1, supporting the pathogenicity of the antibodies. (4) Recombinant expression of the indicated subunits of the GABAaR and neurexin-3α in HEK293 cells can be used as a test to diagnose patients with these autoimmune encephalitis. (5) The infusion of patients’ NMDAR antibodies into the cerebroventricular system of mice, cause memory deficits, anhedonia, and depressive-like behavior. The infused antibodies specifically bind to brain NMDAR resulting in a highly specific reduction of the density of these receptors at synaptic and extrasynaptic levels. The behavioral and molecular effects caused by patients’ antibodies are reversible upon stopping the infusion of antibodies. (6) The administration of ephrin-B2 antagonizes the pathogenic effects of patients’ NMDAR antibodies in all the investigated paradigms, including memory, depressive-like behavior, density of cell-surface and synaptic NMDAR and EphB2, and long-term synaptic plasticity. These findings reveal a strategy beyond immunotherapy to antagonize patients’ antibody effects. (7) The antibodies of patients with anti-LGI1 encephalitis abrogate the binding of the neuronal secreted LGI1 with the presynaptic ADAM23 and with the postsynaptic ADAM22. (8) The infusion of patients’ LGI1 antibodies into the cerebroventricular system of mice cause protracted memory deficits, together with a decrease of presynaptic Kv1.1 potassium channels and post- synaptic AMPAR. These structural effects associate with impairment of synaptic plasticity and increase of neuronal excitability, which are in line with the models of genetic depletion of LGI1. CONCLUSIONS: (1) Anti-GABAaR and anti-neurexin-3α encephalitis are two new forms of antibody-mediated encephalitis for which there is evidence of antibody-mediated pathogenicity. These findings support the concept that among the many types of encephalitis of unclear etiology, there are some that are immune mediated but are still pending to characterize. (2) Direct neuronal antigen precipitation using patients’ antibodies is an excellent strategy to isolate and characterize disease-relevant antigens, and subsequently develop diagnostic screening tests. (3) My studies have contributed to develop two animal models of antibody-mediated symptoms (NMDAR, LGI1) and uncover the underlying antibody-mediated changes in synaptic function and plasticity. Both models fulfill the Witebsky’s criteria for antibody-mediated disease, and provide the basis for modeling other antibody-mediated neurological disorders. |
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