The Role of Distinctive Sphingolipids in the Inflammatory and Apoptotic Effects of Electronegative LDL on Monocytes

Electronegative low-density lipoprotein (LDL(-)) is a minor LDL subfraction that is present in blood with inflammatory and apoptotic effects. We aimed to evaluate the role of sphingolipids ceramide (Cer), sphingosine (Sph), and sphingosine-1-phosphate (S1P) in the LDL(-)-induced effect on monocytes. Total LDL was subfractioned into native LDL and LDL(-) by anion-exchange chromatography and their sphingolipid content evaluated by mass spectrometry. LDL subfractions were incubated with monocytes in the presence or absence of enzyme inhibitors: chlorpromazine (CPZ), d-erythro-2-(N-myristoyl amino)-1-phenyl-1-propanol (MAPP), and N,N-dimethylsphingosine (DMS), which inhibit Cer, Sph, and S1P generation, respectively. After incubation, we evaluated cytokine release by enzyme-linked immunosorbent assay (ELISA) and apoptosis by flow cytometry. LDL(-) had an increased content in Cer and Sph compared to LDL(+). LDL(-)-induced cytokine release from cultured monocytes was inhibited by CPZ and MAPP, whereas DMS had no effect. LDL(-) promoted monocyte apoptosis, which was inhibited by CPZ, but increased with the addition of DMS. LDL enriched with Sph increased cytokine release in monocytes, and when enriched with Cer, reproduced both the apoptotic and inflammatory effects of LDL(-). These observations indicate that Cer content contributes to the inflammatory and apoptotic effects of LDL(-) on monocytes, whereas Sph plays a more important role in LDL(-)-induced inflammation, and S1P counteracts apoptosis.