Autoimmune mechanisms involved in narcolepsy with cataplexy – ImmunitySleep

Sleep is a fundamental neurological behavior for normal brain function. Sleep is a highly complex behavior regulated by many genes, environment, and gene-environment interactions. Sleep disorders are frequent with public health, socioeconomic and psychological consequences. Narcolepsy with cataplexy (NC) is a disabling orphan disorder and the most severe cause of hypersomnia in human, caused by the specific loss of hypocretin (also called orexin) neurons within the hypothalamus. Several evidences argue for an autoimmune pathogenesis with recent studies underlying an association between H1N1 influenza virus vaccination and NC. We aim in this proposal to test some hypotheses regarding the immune mechanisms that could induce NC, using a variety of complementary approaches in a solid consortium involving clinicians, neurobiologists and immunologists with effective between-partner collaborations. This project is based on a translational approach using both human and animal models to study the autoimmune hypothesis of NC. Task 1 will be dedicated to the coordination of the project. Task 2 refers to the in-depth study of cases of narcolepsy induced or not by H1N1 vaccination. We aim at continuing to diagnose, phenotype, and collect samples from recent onset H1N1-exposed NC patients, non-exposed NC patients, and age- and gender-matched healthy controls. We will analyze biological samples from 2 French case-control biobanks, Narcobank and Narcoflu, to determine the neuroinflammation characteristics (cytokines, chemokines, vitamin D), the presence of specific antibodies and the genetic predisposition factors in patients with NC depending on the exposure to pandemic H1N1 flu-vaccine, in comparison to non-exposed patients and healthy controls. Task 3 will test whether H1N1 elicits an adaptive T cell response that inadvertently would cross-react with proteins expressed by hypocretin neurons. We collected peripheral blood mononuclear cells from NC and controls to analyze proliferation and differentiation of T cells upon stimulation with hypocretin or hemagglutinin (HA) peptides from the pandemic influenza virus, and to test the potential involvement of molecular mimicry in NC following H1N1 vaccination. Task 4 aims at investigating the mechanisms involved in antibody-mediated neuroinflammation that could lead to NC. We will study an original rat model of chronic infusion of IgG purified from H1N1-induced or non-exposed recent-onset NC patients. Pathogenicity will be evaluated by sleep analyses, loss of hypocretin neurons and identification of mechanisms of neuroinflammation using immunofluorescence and proteomic approaches. Task 5 tests whether activation of cellular immunity is capable of inducing NC in a unique mouse model of autoimmune disease where hypocretin neurons selectively express the HA gene from an H1N1 viral strain. We recently induced a selective loss of 75% of hypocretin neurons following injection of ex vivo activated HA-specific CD8 T cells. Early immune response will be dissected by histopathological studies and functional analysis of the CNS-infiltrating immune cells. Detection of sleep abnormalities and cataplexy will be done. Upon vaccination or infection of the mice, we will further analyze CD4 and CD8 T cell proliferation and differentiation by flow cytometry and ex vivo functional assays, and characterize the CNS-infiltrating immune cells using immunofluorescence. Altogether these preclinical and clinical research studies have the objectives to decipher the autoimmune mechanisms able to kill hypocretin neurons and investigate how environmental factors such as vaccination trigger an autoimmune response against CNS neurons. The strong position of the 3 academic partners in the domain of sleep, sleep disorders and immunology, and more largely in public health domain, is a major advantage for the success of the project. Our results may lead to new ways to treat such pathologies and to new patentable molecules.