高效液相色谱分析氧化型低密度脂蛋白处理的U937细胞胞内胆固醇及胆固醇酯
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湖南省中青年基金


Analysis the Difference of Cholesterol and Cholesteryl Ester in Oxidized Low Density Lipoprotein Treated U937 Monocyte by High Performance Liquid Chromatography
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    摘要:

    为找到一种判断泡沫细胞与非泡沫细胞的简便而较为实用的方法,本文采用正己烷-异丙醇从U937单核细胞内提取胆固醇,并用醇溶性氢氧化钾除去其中的甘油三酯成分,用三氯乙酸去其中的蛋白成分。以异丙醇:正己烷:乙腈为溶剂洗脱系统,采用疏水性较弱的核酸分离柱,进行高效液相色谱分析,分离纯化胆固醇和胆固醇酯成分,并采用L-B(LiebermanBurchard)显色法结合波谱分析法对所胆固醇峰进行鉴定。结果表明胆固醇含量在0.05~1g/L之间与其峰面积有较好的线性关系,其最低检出量为4mg/g细胞蛋白(100mg细胞蛋白相当于1×108个细胞),通过定性与定量分析,结果表明,U937单核细胞株经氧化型低密度脂蛋白处理48h后,其胞内的胆固醇酯占胞内总胆固醇60%以上。

    Abstract:

    Aim To determine cholesterol and cholesteryl ester of U937 monocyte within it's foaming procedure,and to obtained a simple and useful method to identify foamcell.Methods Cholesterol and cholesteryl ester was ex-tracted from U937 monocyte by hexane-isopropanol,and it's triglycerides was hydrolyzed by ethanolic KOH,and it's protein contents was removed by trichloroacetic acid.Father separation of cholesterol and cholesteryl ester was accompolished by high per-formance liquid chromatography(HPLC),using an eluent of isopropanol-n-heptane-acetonitrice and a Gen-Pak FAX column(with a little hydrophobic prop-erty,using for nucleic acid separation)and it's choles-terol peak was identified by L-B(Lieberman burchard)reaction and spectrum index analysis.Results Cholesterol concentration has a high linear relationship with it's peak area within 0.05-1.0g/L,the limits of detection of cholesterol is 4mg/g cell pro-tein(100mg cell protein equivalent to 1×108 cells).The cholestelyl ester inside U937 amount to 62% to total cholesterol after cholesterol-loaded for 48 hour.Conclusion This method is useful and simple to i-dentify foam cell.

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王佐,李全忠,杨向东,肖尚志,杨永宗.高效液相色谱分析氧化型低密度脂蛋白处理的U937细胞胞内胆固醇及胆固醇酯[J].中国动脉硬化杂志,1998,6(4):317~320.

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  • 收稿日期:1998-09-18
  • 最后修改日期:1998-11-15
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