Lipidomic Analysis of Retinal Ganglion Cell Axons in Xenopus Laevis Ocular Explants

Purpose : Xenopus laevis serves as an exceptional model for studying retinal ganglion cell (RGC) regeneration due to its lifelong ability to regenerate optic nerve axons following injury, unlike the irreversible neurodegeneration seen in mammals. This study provides a comprehensive lipidomic profile of RGC axons isolated from stage 37/38 Xenopus eye explants, using untargeted LC-MS/MS analysis. The findings highlight lipid alterations associated with axonal regeneration, offering molecular insights with potential implications for therapeutic strategies in vision restoration. Methods : Xenopus laevis embryos were obtained via in vitro fertilization, staged to developmental stage 37/38, and maintained in 0.1× MMR at 14–22°C. Retinal ganglion cell (RGC) axons were isolated from cultured eye explants on laminin-coated dishes after 20–24 hours, using manual dissection and a mild detergent to preserve lipid integrity. Lipids were extracted via a modified Bligh and Dyer method, dried under vacuum, and stored at -80°C. Samples were analyzed using high-resolution LC-MS/MS with a Q Exactive Orbitrap, followed by lipid species identification with LipidSearch 5.0 and statistical analysis in MetaboAnalyst 5.0. Results : Preliminary analysis shows that lipidomic profiling of isolated Xenopus laevis retinal ganglion cell (RGC) axons identified 1,261 lipids, with 53 statistically significant lipids across eight LIPIDMAPS subclasses, including 12 triacylglycerols (TG), 16 phosphatidylethanolamines (PE), and 13 phosphatidylcholines (PC). Principal component analysis (PCA) revealed distinct clustering patterns, reflecting variations in lipid subclasses such as phosphatidylinositols (PI), lysophosphatidylcholines (LPC), and ceramides (Cer). Scatterplot visualization categorized these lipids, emphasizing key roles of cholesteryl esters (ChE) and lysophosphatidylethanolamines (LPE), offering insights into molecular changes relevant to axonal growth and regeneration. Conclusions : This study profiles the lipid composition of retinal ganglion cell (RGC) axons from Xenopus laevis, identifying 53 significant lipids across various subclasses such as TG, PC, and PE. These lipids are crucial for membrane remodeling and signaling in axonal repair. Findings validate Xenopus as a model for studying regeneration and suggest lipid-targeted therapies for optic nerve injuries and neurodegenerative diseases.