帕金森病细胞模型的构建及氨基酸表达观察

发布时间：2018-04-21 07:38

本文选题：帕金森病 + 鱼藤酮　；参考：《山东医药》2017年15期

【摘要】：目的建立帕金森病(PD)人神经母细胞瘤细胞模型,并筛选PD细胞中的氨基酸标志物。方法将人神经母细胞瘤细胞SH-SY5Y分为正常组、溶媒组、鱼藤酮组。鱼藤酮组分别加入0.15、0.25μmol/L的鱼藤酮,溶媒组加入二甲基亚砜(DMSO),正常组不加药物。分别于给药24、48 h后采用MTT法测算细胞存活率,倒置显微镜观察细胞形态,选择对细胞存活、形态影响较小的鱼藤酮作用浓度及作用时间(0.15μmol/L、24 h),采用Western blotting法检测SH-SY5Y细胞中的α-突触核蛋白以验证PD模型。采用高效液相色谱法检测PD细胞中的谷氨酸(Glu)、丝氨酸(Ser)、谷氨酰胺(Gln)、组氨酸(His)、甘氨酸(Gly)、丙氨酸(Ala)、脯氨酸(Pro)、色氨酸(Trp)、苯丙氨酸(Phe)、异亮氨酸(Ile)、亮氨酸(Leu)、赖氨酸(Lys),对鱼藤酮组和溶媒组差异有统计学意义的氨基酸进入偏最小二乘判别分析,选择P0.05且变量重要性投影值(VIP)1的氨基酸为PD的标志物。结果鱼藤酮组细胞中α-突触核蛋白相对表达量高于溶媒组(P均0.05),表明SH-SY5Y细胞PD模型制作成功。给药24 h时,鱼藤酮组细胞中Glu、Ser、Gln、His、Gly、Pro、Phe、Ile、Leu、Lys水平低于溶媒组(P均0.05);给药48 h时,鱼藤酮组除Gln外,其余11种氨基酸水平均低于溶媒组(P均0.05)。筛选氨基酸标志物为Glu、Ser、Gly、Pro(给药24 h时VIP值分别为1.016、1.016、1.016、1.009,给药48 h时VIP值均为1.001)。结论利用鱼藤酮诱导成功建立了PD细胞模型,筛选出Glu、Ser、Gly、Pro可能为PD的潜在标志物。
[Abstract]:Objective to establish a human neuroblastoma cell model with Parkinson's disease (PD) and to screen amino acid markers in PD cells. Methods Human neuroblastoma cells (SH-SY5Y) were divided into normal group, solvent group and rotenone group. The rotenone group was added with 0.150.25 渭 mol/L rotenone, the solvent group with dimethyl sulfoxide DMSOO, the normal group with no drug. The cell survival rate was calculated by MTT method 48 hours after 24 hours of administration. The morphology of cells was observed by inverted microscope and the survival rate of cells was selected. The rotenone concentration and the time of action were 0.15 渭 mol / L for 24 h. The 伪 -synaptophysin in SH-SY5Y cells was detected by Western blotting assay to verify the PD model. High performance liquid chromatography (HPLC) method for the detection of glutamate Glun, serine serine, glutamine glutamate, histidine Hisn, glycine Glycine, alanine Alaan, proprol, tryptophan TRP, phenylalanine Phehe, isoleucine Ilel, leucine leucine, lysine Lysn, Lysn in PD cells The difference between ketone group and solvent group was statistically significant. Amino acids entered partial least squares discriminant analysis. The amino acid of VIP1 was selected as the marker of PD. Results the relative expression of 伪 -synaptophysin in rotenone group was higher than that in solvent group (P 0.05), which indicated that the PD model of SH-SY5Y cells was made successfully. After 24 h of administration, the levels of Glutenone group, Glutenone group and Glutenone group were lower than those in the solvent group (P < 0.05) and the other 11 amino acids in the rotenone group were all lower than those in the solvent group (P < 0.05) at 48 h after the administration of rotenone, except for Gln, the levels of the other 11 amino acids in the rotenone group were all lower than those in the solvent group (P < 0.05). The amino acid markers were Glutene Seroglycoprol (VIP values were 1.016 ~ 1.016 ~ 1.016 ~ 1.009 at 24 h and VIP = 1.001 at 48 h) respectively. Conclusion the model of PD cells induced by rotenone was successfully established and the potential marker of PD was screened out.
【作者单位】：河北北方学院;
【基金】：河北省教育厅重大项目(ZD2014075) 河北北方学院博士科研启动基金项目
【分类号】：R742.5

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