冰岛刺参缩醛磷脂和醚磷脂对神经生长因子诱导PC12细胞的促神经分化作用

目的：分离纯化冰岛刺参（Cucumaria frondosa）中的缩醛型磷脂酰乙醇胺（plasmenylethonoamine，pPE）和烷氧型磷脂酰胆碱（plasmanylcholine，aPC），并对其促进神经生长因子（nerve growth factor，NGF）诱导的大鼠肾上腺嗜铬细胞瘤（pheochromocytoma，PC12）细胞分化的神经营养作用进行探究。方法：以冰岛刺参为原料，通过柱层析分离法提取pPE和aPC，并通过液相色谱-质谱联用法测定其纯度；分析冰岛刺参pPE和aPC对PC12细胞存活率和分化率的影响，并统计分析细胞轴突长度和数目的变化；进一步通过实时荧光定量聚合酶链式反应、免疫荧光技术等方法检测PC12分化细胞中突触素和生长相关蛋白43（growth-associated protein 43，GAP-43）的表达量，明确冰岛刺参pPE和aPC对PC12细胞突触生长及分化的影响。结果：制备获得的冰岛刺参pPE和aPC纯度分别为91.0%和89.1%，两种不同结构的磷脂均可显著提高PC12细胞的分化率（P＜0.05），通过促进突触素和GAP-43的表达促进细胞轴突生长，pPE的作用效果优于aPC。结论：体外细胞实验结果表明冰岛刺参pPE和aPC能显著促进PC12神经细胞分化及突触生长，且具有浓度依赖性和构效性。但详细的作用机制仍需进一步研究。

Effects of Plasmenylethonoamine and Plasmanylcholine from Cucumaria frondosa on Promoting Nerve Growth Factor-Induced Neuronal Differentiation of PC12 Cells

The objective of this study was to investigate the neurotrophic effects of plasmenylethonoamine(pPE)and plasmanylcholine(aPC)from Cucumaria frondosa on the nerve growth factor(NGF)-induced differentiation of pheochromocytoma(PC12)cells.We extracted and purified pPE and aPC by column chromatography and their purity were determined by liquid chromatography-mass spectrometry(LC-MS).The effects of pPE and aPC on the survival and differentiation of PC12 cells were analyzed by morphological examination,and changes in the length and number of neurites were statistically analyzed.Furthermore,the expression of synaptophysin and growth-associated protein 43(GAP-43)in differentiating PC12 cells were detected by real-time fluorescence quantitative polymerase chain reaction(qPCR)and immunofluorescence to evaluate the effects of pPE and aPC on the growth and differentiation of synapses in PC12 cells.Results showed that the purities of pPE and aPC were 91.0%and 89.1%,respectively,and both phospholipids,with different structures,could significantly improve the differentiation of PC12 cells(P<0.05)and promote the growth of neurites by promoting the expression of synaptophysin and GAP-43.The effect of pPE was better than that of aPC.In conclusion,pPE and aPC could significantly promote the differentiation and synaptic growth of PC12 cells in a concentration-dependent and structure-specific manner,but the exact underlying mechanism still needs further study.