Welcome to the IBRO 2023 Interactive Programme

Abstract Body

The bioactive lipid sphingosine-1-phosphate (S1P) is crucial for brain development. Its absence is embryonically lethal but a defective degradation of S1P is also harmful causing brain malformations and early postnatal death. Likewise, findings regarding the role of S1P in the initiation and progression of diverse neurodegenerative diseases are rather contrasting. Therefore, we generated a mouse model in which S1P-lyase (SGPL1), the enzyme which irreversibly cleaves S1P yielding ethanolamine phosphate (EAP) and hexadecenal, was inactivated specifically in neural cells (SGPL1^fl/fl/Nes). In humans, the gene SGPL1 maps to a region prone to mutations in several human cancers and also in the SGPL1 Insufficiency Syndrome (SPLIS) characterized by several symptoms including peripheral and central neurological defects. Indeed, accumulation of S1P in the brain of SGPL1^fl/fl/Nes mice affected synaptic architecture and plasticity whereas the drop of EAP and consequently of phosphatidylethanolamine blocked neuronal autophagy at its early stages. Hence aggregate-prone proteins such as the amyloid precursor protein (APP) and alpha synuclein were entrapped in neurons along with profound deficits in cognitive skills observed in vivo. In response to neuronal damages, microglial cells were activated propagating neuroinflammation and neuropathology. We also observed an age-dependent activation of astrocytes. The underlying molecular mechanism is rather complex and involves on the one hand a calcium dependent increase of histone acetylation. On the other hand, S1P stimulates glycolysis in SGPL1-deficient cells. In astrocytes which usually are glycolytic S1P stimulated the TCA cycle via S1PR₂ and S1PR₄ while lactate dehydrogenase was considerably decreased. As a consequence, more ATP was generated in line with a significant rise of extracellular ADP that turned out to induce via the purinoreceptor P2Y1 the activation of the NLRP3 inflammasome. Our findings may contribute to a better understanding of S1P function in brain pathologies and to the development of drugs targeting S1P metabolism.

Abstract Body

Therapeutics that promote oligodendrocyte survival and remyelination are needed to restore neurological function in demyelinating diseases such as multiple sclerosis. Sphingosine 1-phosphate (S1P) receptor agonists Fingolimod, Siponimod, Ozanimod, Ponesimod, and others are clinical immunosuppressants used to treat multiple sclerosis. Clinical evidence suggests that these drugs also have direct neuroprotective actions in the CNS, however experimental evidence to support this hypothesis is lacking. S1P is an essential lipid metabolite that signals through a family of five G-protein coupled receptors. Our recent research demonstrates that endogenous S1P, synthesised by the enzyme sphingosine kinase 2 (SphK2), is an important mediator of oligodendrocyte survival and essential for remyelination after treatment with the oligodendrocyte toxin cuprizone. Mice lacking functional SphK2 also show loss of myelin integrity with ageing, further indicative of a requirement for endogenous S1P synthesis in myelin maintenance. Interestingly, SphK2 is required for myelin synthesis by mature oligodendrocytes, but not for oligodendrocyte progenitor cell proliferation and differentiation. Treatment of mice with the S1P receptor 1 and 5 agonist Siponimod protects against cuprizone-mediated oligodendrocyte loss and demyelination, and reverses the effect of SphK2 deficiency on oligodendrocyte loss. However, Siponimod does not reverse the inhibitory effect of SphK2 deficiency on remyelination. Our results therefore indicate that both endogenous and pharmacological activation of S1P receptors 1 and/or 5 protects against oligodendrocyte loss and demyelination, whereas the requirement for endogenous S1P synthesis in remyelination is independent of these receptors. This work establishing the requirement for endogenous S1P in protection and maintenance of oligodendrocyte functions is fundamental to understanding the potential neuroprotective properties of clinical S1P receptor agonists, and should guide clinical decision making.

Abstract Body

Astrocytes are specialized glial cells that possess an intricate morphology with highly ramified cellular processes interacting closely with individual synapses and other brain cells. Astrocytes perform a range of essential developmental and homeostatic functions in neural circuits, and it is thought that these functions depend on their remarkable morphological complexity and close association with neural elements. However, we know surprisingly little about the molecular and genetic mechanisms that underlie astrocyte morphogenesis or how astrocytes modulate brain circuits in vivo. By leveraging a combination of molecular-genetic tools and live-imaging techniques in Drosophila and zebrafish, we sought to identify evolutionarily conserved signaling pathways that regulate astrocyte development and function in vivo. In Drosophila, we found that RNAi-mediated depletion of Trapped in endoderm 1 (Tre1) GPCR in astrocytes resulted in severely reduced astrocyte morphological complexity. We showed that endogenous Tre1 is highly enriched in astrocytes and validated the astrocyte morphology phenotypes in Tre1 genetic mutants. We demonstrated that Tre1 acts cell autonomously to direct the elaboration of fine astrocyte processes into the synaptic neuropil, by genetically interacting with small GTPase Rac1 to regulate the actin cytoskeleton. Furthermore, we found this is a conserved signaling modality across species, as loss of the functional vertebrate GPCR analog sphingosine-1-phosphate receptor 1 (s1pr1) in zebrafish led to disrupted astrocyte morphology similar to that observed in Drosophila mutants. Using a combination of live-imaging and pharmacology approaches in zebrafish, we found that S1pr1 is required throughout astrocyte development to control astrocyte process outgrowth and dynamics. Together, our data reveal a crucial and conserved Tre1/S1pr1-signaling in governing astrocyte morphogenesis in vivo.

Abstract Body

CNS mechanisms of S1P receptor signaling in multiple sclerosis

Yasuyuki Kihara, Valerie Tan, Anis Shahnaee, Deepa Jonnalagadda, Carter Palmer, Christine Liu, and Jerold Chun

Sphingosine 1-phosphate (S1P) is a lysophospholipid whose biological actions are primarily mediated by a subfamily of G protein-coupled receptors (GPCRs) referred to as S1P_1-5. S1P receptor small-molecule modulators now account for four FDA-approved medicines (fingolimod, siponimod, ponesimod, and ozanimod) for the treatment of multiple sclerosis (MS), where they are best characterized as immunomodulators of lymphocyte trafficking. However, all receptor subtypes show brain expression during development and/or during adult life, and an expanding range of experimental data support their involvement in normal and disease processes that likely underlie their efficacy in MS. Earlier work identified astrocyte activities mediated by S1P₁ that contributed to the efficacy of fingolimod (FTY720) independent of lymphocyte trafficking effects. Astrocytes have many functions within the central nervous system (CNS) that can be altered by receptor-mediated S1P signaling. Single nucleus RNAseq (sn-RNAseq) identified several pathways that could be accessed by S1P receptor modulators, including those that involve vitamin B₁₂. Vitamin B₁₂ deficiency has been associated with MS and MS-like symptoms historically, and current work identified molecular elements that connect S1P signaling with that of vitamin B₁₂. In addition, S1P signaling has been linked to myelination, and recent studies identified myelination effects that can be influenced by S1P receptor modulators, including those that can protect or possibly promote myelination/re-myelination, at least in animal models. These results implicate a range of CNS mechanisms that underlie effects of S1P receptor modulators, including therapeutic effects on progressive MS that involve neurodegeneration, which may be generalizable to other CNS indications.