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Poster Presentations
Day 2, June 23（Mon.）
Room P (Maesato East, Foyer, Ocean Wing)
- 2P-AM-34
Examining the subcellular localisation of ceramides in mouse tissue using targeted mass spectrometry
(1VCCRI, 2SoCM UNSW Sydney, 3SBMS UNSW Sydney, 4SCh UNSW Sydney)
oLaura Choong1,2, Sarah Hancock1,3, Amy Nguyen1, Iliya Dragutinovic4, Elysha Taylor4, Jonathan Morris4, Nigel Turner1,3
Ceramide, a sphingolipid, has a causal role in cardiometabolic disease. Those containing an acyl-tail of 16 or 18 carbons are considered deleterious due to frequent
association with cardiometabolic disease. Ceramide synthase (CerS) catalyses ceramide synthesis. The subcellular localisation of ceramides and CerS is important to understanding their role in insulin resistance, but this is not yet well defined. This study
aims to use subcellular fractionation paired with targeted liquid chromatography-mass spectrometry (LC-MS) to interrogate the subcellular ceramide distribution in key tissues.
Male C57BL/6 mice were provided chow, high-fat diet (HFD), or HFD + a CerS inhibitor, ET2.39 (~10mg/kg/day). Cellular fractions (plasma membrane, cytosol, nuclear and mitochondrial) from quadriceps and liver were extracted using an iodixanol gradient and ultracentrifugation. Enrichment was verified via western blotting of fraction markers. Sphingolipid profiles of fractions were analysed via targeted LC-MS which detected 62 different sphingolipid species.
Fractions demonstrated unique sphingolipid composition. Sphingolipid content was greatest in the plasma membrane and nucleus. HFD significantly increased deleterious ceramides across all cellular fractions. ET2.39 treatment significantly reduced C18
ceramides in quadriceps, with similar trends shown in cellular fractions. In conclusion, different sphingolipid distributions exist across cellular fractions, of which can altered
by diet and pharmacological inhibition.