匹伐他汀对体外培养人脐静脉血内皮祖细胞数量及功能的影响

Effects of Pitavastatin on Number and Activity of Endothelial Progenitor Cells from Umbilical Vein Blood

目的:通过体外培养人脐静脉血内皮祖细胞(endothelial progenitor cells,EPCs),观察他汀类新药(匹伐他汀)对EPCs数量及增殖、迁移和粘附功能的影响。方法:采用密度梯度离心法分离培养人脐静脉血单个核细胞,将其接种在包被有人纤维连接蛋白培养板上,培养7 d后,收集贴壁细胞,加入不同浓度匹伐他汀(分别为0.001μmol/L、0.01μmol/L、0.1μmol/L、1.0μmol/L)培养24h,用免疫荧光法观察EPCs吸收FITC-UEA-I和Dil-acLDL情况对EPCs进行鉴定,然后分别采用MTT比色法、改良的Boyden小室、粘附能力测定实验对各实验组测定,来观察匹伐他汀对EPCs数量及增殖、迁移和粘附功能影响。结果:匹伐他汀组与对照组相比,匹伐他汀显著提高了体外培养EPCs的数量及增殖、迁移与粘附能力(P<0.05)。匹伐他汀浓度在0.1μmol/L时对EPCs影响达到最大。随着药物浓度的继续增大,EPCs的上述功能反呈下降趋势,但1.0μmol/L组仍高于对照组。结论:匹伐他汀能增加体外培养EPCs的数量及增殖、迁移和粘附能力,可作为EPCs培养的一种改良方法,为其更好的应用于临床具有重要的意义。

Objective： To investigate influences of Pitavastatin on adhesion, spreading, migration of endothelial progenitor cells （EPCs）. Methods： Total mononuclear cells （MNCs） were isolated from peripheral blood by Ficoll density gradient centrifugation, and then the cells were plated on fibronectin-Coated culture dishes. After culture for7 days, attached cells were stimulated with Pitavastatin （to make a series of final concentrations： 0.001 ixmol/L, 0.01 ixmol/L, 0.1 txmol/L, 1.0 ixmol/L） for 24 h. EPCs were characterized as adher- ent cells double positive for DiLDL2 uptake and lectin binding by direct fluorescent staining. EPCs proliferation, migration were assayed with MTT assay and modified Boyden chamber assay, respectively. EPCs adhesion assay was performed by replating those on fibronectin 2 coated dishes, and then adherent cells were counted. Results： Incubation of isolated human MNCs with Pitavastatin dose dependently increased the number of EPCs, maximum at 0.1 txmol/L, （P〈0.05）. In addition, Pitavastatin also promotes EPCs proliferation, migratory and adhesive capacity in a concentration dependent manner. When the concentration is higher than 1.0 ixmol/L, the function of EPCs will be inhibited. Conclusion： The results of the present study defined that Pitavastatin can promote EPCs proliferative, migratory and adhe- sive capacity, and can be a new way for culture EPCs. All of these can provide more advise for clinical.