低密度脂蛋白诱导Raw264.7巨噬细胞后脂质筏中脂类物质组分的变化

Severe alteration in lipid composition of lipid raft in Raw264.7 cell after LDL induction

目的研究Raw264.7细胞经低密度脂蛋白(LDL)诱导后脂质筏中脂类物质组分是否发生变化。方法蔗糖梯度超速离心得到对照组和实验组细胞中的脂质筏,气相色谱质谱(GC-MS)分析脂质筏中脂肪酸的变化;化学衍生脂质筏标志物-单唾液酸四己糖神经节苷脂(Ganglioside 1,GM1),高效液相色谱(HPLC)分析GM1的变化;胆固醇试剂盒检测脂质筏中胆固醇含量的变化。结果与对照组相比,经LDL诱导的Raw264.7细胞脂质筏中的单不饱和脂肪酸(MUFAs)组分中十六烯酸、油酸、二十四烯酸显著升高;多不饱和脂肪酸(PUFAs)组分中亚油酸、花生四烯酸和二十碳三烯酸显著降低;饱和脂肪酸组分中棕榈酸和硬脂酸显著升高,十四烷酸和二十烷酸含量无变化;GM1含量和胆固醇含量均显著增高,有统计学意义。说明LDL改变了细胞脂质筏的脂质微环境。结论 Raw264.7细胞脂质筏中的脂类物质可能与LDL氧化有关,有利于我们进一步研究LDL氧化过程中脂类物质的作用及其与蛋白质的相互作用。

Oxidized low-density lipoprotein （Ox-LDL） plays an important role in atherogenesis, and the mechanisms underlying cell-mediated LDL oxidation remain obscure. The study performed to detect the alteration in lipid composition of the lipid raft from LDL-induced Raw264.7 cell. We purified lipid rafts from normal and native-LDL induced Raw 264.7 cells by density gradient centrifugation and analyzed their lipid composition with Gas Chromatograph-Mass Spectrometer （GC-MS） and High performance liquid chromatograph （HPLC）. The cholesterol concentration was assayed by Amplex Red Cholesterol Assay Kit. Data showed that lipid rafts from native-LDL induced Raw264.7 cells exhibit dramatic increase of hexadecenoic acid, octadecenoic acid and tetracosenoic acid in their contents of monounsaturated fatty acid, while polyunsaturated fatty acids （PUFAs） including octadecadienoic acid, arachidonic acid and eicosatrienoic acid were reduced. In the contents of saturated fatty acids, hexadecanoic acid and octadecanoic acid were increased, others showed no changes. GM1 （the mark of lipid raft） and the cholesterol concentration in the lipid raft were significant up-regulated upon native-LDL stimulation. In conclusion, our data pinpoints a dramatic alteration in lipid composition of the lipid raft in LDL-induced cells and indicate that lipids of lipid rafts may be related to LDL oxidation, which will be helpful to exploring the role of lipids LDL oxidation and the protein-lipid interactions of lipid rafts to provide insights into the mechanism of LDL oxidation in the future.