体外制备的晚期糖基化终产物对内皮祖细胞数量的影响
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The Depression in Number of Endothelial Progenitor Cells by Advanced Glycation End Products Prepared in Vitro
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    摘要:

    目的观察体外制备的晚期糖基化终产物对内皮祖细胞数量的影响。方法人血清白蛋白与葡萄糖共同孵育90d后,行Bradford检测法蛋白定量和荧光值测定。从正常成人外周血中分离提取单个核细胞,培养7d后,流式细胞仪鉴定细胞表型,分别加入浓度为2mg/L、20mg/L和200mg/L的晚期糖基化终产物并设置对照组,干预不同的时间(0h、24h、48h和72h),200倍光学显微镜下随机取10个视野计数。结果该方法制备的晚期糖基化终产物具有荧光性。细胞培养7d,CD34/CD133/KDR单克隆抗体流式细胞仪鉴定细胞,结果发现表达KDR达78.60%±2.20%、CD34为8.60%±2.00%和CD1332.80%±0.60%,提示大部分细胞为内皮祖细胞。以不同时间(0h、24h、48h和72h)和浓度(2mg/L、20mg/L和200mg/L)晚期糖基化终产物干预内皮祖细胞生长,对比0h组每个视野下细胞数量(186.0±6.7),干预24h细胞数量减少(146.67±4.98),72h降至最低(73.67±3.76)(P<0.001),呈现明显的时间效应(r=0.9658,P<0.01);终浓度为2mg/L或20mg/L晚期糖基化终产物干预内皮祖细胞72h后,与同时培养72h的对照组无显著差异(P>0.05),但随着浓度加大,浓度效明显(r=0.9988,P<0.01)。结论此方法制备的晚期糖基化终产物符合文献所述的特性,晚期糖基化终产物能抑制内皮祖细胞生长,具有时间效应和浓度效应。

    Abstract:

    Aim To investigate the effect of advanced glycation end products (AGE) on number of endothelial progenitor cells (EPC). Methods AGE was prepared by incubating D-glucose and human serum albumin (HSA) for 90 days, and then identified by fluorescence and SDS-PAGE.The mononuclear cells (MNCs) were isolated from peripheral blood of healthy adults,then were plated on fibronectin-coated culture 6-well plates respectively in EC basal medium-2.After cultured 7 days, attached cells were identified with antibodies recognizing human CD34,CD133 and KDR, several groups of attached cells plated in the absence or presence of AGE (2 mg/L, 20 mg/L, and 200 mg/L) for 0 h, 24 h, 48 h, and 72 h respectively,the cells were quantified by viewing 10 random microscopic fields. Results AGE showed the absorption and fluorescent spectra. After cultured 7 days, attached cells were identified with antibodies, the results demonstrated the positive expression of antibodies were KDR (78.60%±2.20%), CD34 (8.60%±2.00%) and CD133 (2.80%±0.60%). The decrease number of EPC per field was observed under 200 mg/L AGE at 24 h (146.67±4.98), while reached to the minimum (73.67±3.76) at 72 h. However, no statistical change in cell number was identified when cells were treated under low density AGE (2 mg/L and 20 mg/L). Conclusion High AGE concentration may inhibit EPC augmentation.

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李倩,沈成兴,刘乃丰.体外制备的晚期糖基化终产物对内皮祖细胞数量的影响[J].中国动脉硬化杂志,2008,16(4):284~286.

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  • 收稿日期:2007-12-17
  • 最后修改日期:2008-04-19
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