Characterizing a novel mouse model linking sphingolipid metabolism to multi-organ fibrosis - PROJECT SUMMARY Nearly half of all deaths in the industrialized world are caused by fibrosis of critical organs and tissues. Despite this fact, few available treatments prevent the progression of fibrosis and its lethal consequences. Critical gaps in our understanding of fibrosis remain, particularly what factors cause this process to switch from a self-limited homeostatic response to injury to become a progressive “chronic wound-healing” pathology, and conversely what endogenous factors help to keep fibrosis in check. Novel fibrosis-prone mouse models afford the potential to elucidate these factors and thereby reveal new anti-fibrotic therapeutic targets for a wide range of diseases. S1P is a bioactive lipid formed in the final step of sphingolipid metabolism that has been implicated in the pathophysiology of fibrosis. S1P lyase (SPL), encoded by SGPL1, is a vitamin B6-dependent enzyme that catalyzes the irreversible degradation of S1P and guards the only exit point of sphingolipid metabolism. Recently, we discovered SPL insufficiency syndrome (SPLIS), a rare genetic disorder that most commonly manifests as glomerulosclerosis. To interrogate the broader role of SPL in fibrosis, we used gene editing to generate a novel SPLIS mouse model harboring the most prevalent SPLIS-associated SGPL1 mutation, SPL R222Q, which is vitamin B6-responsive. When maintained on a diet low in vitamin B6, SPLR222Q mice exhibit SPL insufficiency, accumulate S1P and other sphingolipids in tissues and blood and, over time, develop glomerulosclerosis, whereas these phenotypes are prevented by providing a diet high in vitamin B6. In addition to kidney fibrosis, we showed that SPLR222Q mice spontaneously develop lung fibrosis and are more sensitive to bleomycin-induced pulmonary fibrosis than are wild type (WT) mice. Conversely, upregulation of SPL using an adeno-associated virus mediated SGPL1 gene therapy (AAV-SPL) prevented glomerulosclerosis in Sgpl1 knockout mice and attenuated bleomycin-induced pulmonary fibrosis in WT mice. Altogether, our findings strongly suggest that SPLIS is a harbinger of a broader anti-fibrotic role played by SPL in human physiology. We hypothesize that SPL is an endogenous anti-fibrotic factor that counteracts the pro-fibrotic actions of S1P and that the SPLR222Q mouse is a robust in vivo model in which to elucidate the role of the S1P/SPL axis in fibrosis and to test novel anti-fibrotic strategies. To confirm our hypothesis, we propose two Specific Aims: 1) Characterize the spectrum of organs affected by fibrosis and sphingolipid changes in the SPLR222Q mouse; 2) Establish the efficacy and mechanism of action of AAV-SPL anti-fibrotic therapy in the SPLR222Q mouse. Our published and preliminary findings implicate the S1P/SPL axis in fibrosis of kidneys liver, lungs and gut. Our experienced team will generate results that will establish SPL as a critical endogenous anti-fibrotic factor. We will identify pro-fibrotic sphingolipids. Lastly, we will validate AAV-SPL as a novel anti-fibrotic strategy with broad applications in many organ systems. In line with ORIP’s goals, our project will generate animal models, cryopreserved embryos, cell lines, lipidomics datasets, and viruses for sharing with the scientific community.
