Ceramide structural features required to stimulate ABCA1-mediated cholesterol efflux to apolipoprotein A-I

Amy B. Ghering and W. Sean Davidson

Pathology and Laboratory Medicine, University of Cincinnati, Cincinnati, OH 45237

Corresponding Author: Sean.Davidson{at}uc.edu

Ceramide is a component of the sphingomyelin cycle and a well-established lipid signaling molecule. We recently reported that ceramide specifically increased ATP-Binding Cassette Transporter A1 (ABCA1)-mediated cholesterol efflux to apolipoprotein A-I (apoA-I), a critical process that leads to the formation of cardioprotective high density lipoprotein (HDL). In this report, we characterized the structural features of ceramide required for this effect. C2-dihydroceramide, which contains a fully saturated acyl chain and is commonly used as a negative control for ceramide apoptotic signaling, stimulated a 2-5 fold increase in ABCA1-mediated cholesterol efflux to apoA-I over a 0-60 M concentration range without the cell toxicity apparent with native C2 ceramide. In comparison to C2 ceramide, C6 and C8 ceramides with medium length N-acyl chains showed a similar extent of efflux stimulation (a 2-5 fold increase) but at a higher onset concentration than the less hydrophobic C2 ceramide. In contrast, the reduced and methylated ceramide analogs, dimethyl sphingosine and trimethyl sphingosine, failed to stimulate cholesterol efflux. We found that changes in the native spatial orientation at either of two chiral carbon centers (or both) resulted in ~50% decrease compared to native ceramide-stimulated cholesterol efflux. These data show that the overall ceramide shape and the amide bond and are critical for the cholesterol efflux effect and suggest that ceramide acts through a protein-mediated pathway to affect ABCA1 activity.

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