1-palmitoyl-2-oleoylphosphatidylcholine and sphingosine-1-phosphate

Reconstituted high-density lipoprotein (rHDL) has been shown to produce a rapid regression of atherosclerosis in animal models and humans. Sphingosine-1-phosphate (S1P), which is a bioactive lipid in HDL, plays a role in mitogenesis, endothelial cell motility, and cell survival, as well as organization and differentiation into a vessel. In this study, we examined the direct role of a newly developed rHDL, [POPC(1-palmitoyl-2-oleoyl phosphatidylcholine)/S1P/apolipoproteinA-I(A-I)]rHDL containing S1P in tube formation in endothelial cells (ECs) as well as cholesterol efflux in macrophage. The effect of (POPC/S1P/A-I)rHDL on cholesterol efflux in macrophage was similar to that of conventional rHDL, (POPC/A-I)rHDL. In addition, (POPC/S1P/A-I)rHDL induced EC proliferation through the activation of phospho-Akt and phospho-extracellular-signal-regulated kinases (p-ERK) 1/2 and EC tube formation, and this effect was blocked by inhibitors of Akt, ERK and endothelial nitric-oxide synthase (eNOS). In addition, (POPC/S1P/A-I)rHDL-induced p-ERK1/2 activation and EC tube formation can be mainly attributed to S1P-stimulated signaling through S1P2 and S1P3 as determined by an anti-sense strategy. In conclusion, (POPC/S1P/A-I)rHDL induces cholesterol efflux independently of S1P but has additional S1P-mediated effects on EC tube formation mediated by Akt/ERK/NO through S1P2 and S1P3. In the future, these new discs may be useful for the treatment of atherosclerotic and ischemic cardiovascular disease, such as acute coronary syndrome and atherosclerosis obliterans.

Topics: Animals; Atherosclerosis; Cell Division; Cells, Cultured; CHO Cells; Cholesterol; Coronary Vessels; Cricetinae; Cricetulus; Endothelial Cells; Enzyme Inhibitors; Humans; In Vitro Techniques; Lipoproteins, HDL; Lysophospholipids; Macrophages; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase Type III; Oligonucleotides, Antisense; Phosphatidylcholines; Proto-Oncogene Proteins c-akt; ras Proteins; Sphingosine

Sphingosine 1-phosphate is a bioactive sphingolipid that regulates cell growth and suppresses programmed cell death. The biosynthesis of sphingosine 1-phosphate is catalyzed by sphingosine kinase (SK) but the mechanism by which the subcellular localization and activity of SK is regulated in response to various stimuli is not fully understood. To elucidate the origin and structural determinant of the specific subcellular localization of SK, we performed biophysical and cell studies of human SK1 (hSK1) and selected mutants. In vitro measurements showed that hSK1 selectively bound phosphatidylserine over other anionic phospholipids and strongly preferred the plasma membrane-mimicking membrane to other cellular membrane mimetics. Mutational analysis indicates that conserved Thr54 and Asn89 in the putative membrane-binding surface are essential for lipid selectivity and membrane targeting both in vitro and in the cell. Also, phosphorylation of Ser225 enhances the membrane affinity and plasma membrane selectivity of hSK1, presumably by modulating the interaction of Thr54 and Asn89 with the membrane. Collectively, these studies suggest that the specific plasma membrane localization and activation of SK1 is mediated largely by specific lipid-protein interactions.

Topics: Amino Acid Sequence; Asparagine; Cell Line; Cell Membrane; DNA Mutational Analysis; Green Fluorescent Proteins; Humans; Kinetics; Lipids; Lysophospholipids; Mass Spectrometry; Microscopy, Confocal; Microscopy, Fluorescence; Models, Biological; Molecular Sequence Data; Mutation; Peptides; Phosphatidylcholines; Phosphatidylethanolamines; Phosphatidylserines; Phospholipids; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Pressure; Protein Structure, Tertiary; Sequence Homology, Amino Acid; Serine; Sphingosine; Surface Plasmon Resonance; Threonine; Time Factors; Transfection