Myristic Acid Remodels Sphingolipid Metabolism via Dual Pathways: Canonical d18-Sphingolipid Regulation and Non-Canonical d16-Sphingolipid Synthesis

<b>Background</b>: Myristic acid (MA), a 14-carbon saturated fatty acid, serves as a precursor for the synthesis of non-canonical d16-sphingoid bases via its activated form, C14:0-CoA. However, its broader regulatory role in sphingolipid (SL) metabolism remains poorly defined. <b>Methods</b>: Using HepG2 cells treated with 50 μM MA, we found that sphingolipidomic analysis revealed reprogrammed sphingolipid metabolism. <b>Results</b>: In the canonical d18-SL pathway, MA directs its activated product C14:0-CoA into ceramide <i>N</i>-acyl chains and downstream metabolites-especially d18:1-C14:0 hexosylceramide. Concurrently, in the non-canonical d16-SL pathway, MA promotes d16-SL synthesis, especially d16:1-ceramides (Cer), d16:1-hexosylceramides (HexCer), and d16:1-C14:0 lactosylceramide. MA treatment further induced a coordinated shift in cellular sphingolipid pools, characterized by a significant increase in total ceramide levels (encompassing both d16- and d18-species) alongside concurrent reductions in total sphingomyelin (SM) contents. At the gene transcriptional level, MA significantly suppressed <i>SPTLC2</i> mRNA expression while markedly upregulating <i>SMPD2</i> and <i>SMPD3</i> mRNA levels. <b>Conclusions</b>: Collectively, these findings position MA as a potent regulator of sphingolipid homeostasis, orchestrating dual pathway modulation: disrupting canonical d18-SL equilibrium through the selective enrichment of <i>N</i>-acyl C14:0-containing SLs, and activating non-canonical d16-SL synthesis. This dual pathway regulation reveals that dietary saturated fatty acids exploit sphingolipid subnetworks to regulate lipid metabolism. The interplay between dietary fatty acids and sphingolipid metabolism still requires deeper exploration. Our findings offer preliminary insights into their roles in regulating both normal and disease-associated lipid metabolism, setting the stage for subsequent mechanistic investigations.