Abstract 4372722: Unveiling S1PR1 Internalization in Left-Sided Trabecular development

Abstract

Introduction: Congenital heart disease (CHD) is the most common birth defect and a major cause of neonatal mortality. Over half of CHD cases remain genetically unexplained. The lipid mediator sphingosine-1-phosphate (S1P) acts through the endothelial receptor S1PR1 to regulate cardiovascular development. While S1P–S1PR1 signaling has been studied in early cardiac morphogenesis, its role in later trabecular angiogenesis (E15.5–P7) is unknown. Hypothesis: Low S1PR1 levels in the left ventricular (LV) endocardium, achieved via spatially restricted internalization and degradation, are essential for proper coronary vessel development and trabecular compaction. Dysregulation of this pathway leads to structural heart defects. Methods: Reporter mice expressing GFP-tagged S1PR1 were used to track internalization. Immunohistochemistry and scRNA-seq were employed to analyze expression of S1PR1 and its degradation mediators (GRK2, ARRB2, WWP2) across ventricular regions. Functional studies in S1PR1 internalization-defective mice (S1PR1 S5A ), S1PR1 endothelial overexpression (ECOE) mice, and S1PR1 pharmacological inhibition models were conducted. Markers of angiogenesis (VEGFR2/3), EndoMT (SNAI2, TGFB2), and proliferation (Ki67, EdU) were used to evaluate ventricular development. Results: S1PR1 internalization was enriched in the LV and interventricular septum, aligning with coronary vessel formation. CHD patient data revealed elevated S1PR1 expression in endocardial cells, especially in LVNC and Tetralogy of Fallot cases. ECOE mice showed LV noncompaction, septal defects, and reduced trabecular angiogenesis. S1PR1 S5A mice exhibited similar phenotypes, confirming the importance of internalization. Pharmacological inhibition using AUY954 and W146 partially rescued these defects. Conclusions: Spatial suppression of S1PR1 via internalization is a crucial developmental mechanism in late-stage heart formation. Excess S1PR1 signaling disrupts endocardial-to-endothelial transition and coronary vessel development. These findings reveal a novel pathophysiological pathway for CHD and suggest therapeutic potential for S1PR1-targeting drugs in preventing structural heart defects.