过表达Periostin蛋白对内皮前体细胞功能影响的实验研究

目的构建Periostin蛋白过表达慢病毒载体, 并检测其对内皮前体细胞(EPCs)功能的影响。方法从小鼠骨髓中获取EPCs, 并进行鉴定。采用Oligos设计获得目的基因, 构建慢病毒载体LV5/Periostin。慢病毒感染EPCs, 荧光显微镜下观察被感染细胞绿色荧光蛋白(GFP)的表达情况, Western blotting检测Periostin蛋白表达情况。CCK-8法和流式细胞计数检测被感染细胞的增殖和凋亡情况, 划痕法和Transwell小室法检测被感染细胞迁移和侵袭能力的改变情况。结果被重组慢病毒感染的EPCs明显表达GFP; 过表达慢病毒组细胞增殖活性较阴性对照组低, 而凋亡细胞较阴性对照组多(P < 0.05);过表达慢病毒组细胞的迁移率较空白对照组细胞高, 侵袭细胞数目较多(P < 0.05)。结论可以从小鼠骨髓中成功分离和培养EPCs, 过表达Periostin蛋白会降低EPCs体外增殖的活性, 促进其凋亡; 但会增加EPCs的体外迁移和侵袭能力。

Effect of over-expression of Periostin protein on the biological function of endothelial progenitor cells

Objective To construct the over-expression of Periostin protein lentiviral vector, and investigate its effects on the biological function of endothelial progenitor cells(EPCs).Methods The mouse EPCs were isolated from bone marrow, cultured and identified.The LV5/Periostin lentiviral vector was constructed, and transferred into the EPCs.The expression of green fluorescent protein(GFP) in transfected cells was observed under fluorescence microscope.The expression level of Periostin in EPCs was verified by Western blotting, the proliferation and apoptosis of infected cells were detected using the cell counting kit-8(CCK-8) assay and flow cytometry, and the migration ability and invasion ability of cells were detected using the scratch and Transwell assgy.Results The expression of GFP in EPCs was obvious.Compared with the control group, the proliferation activity of cells was lower, and the number of apoptosis cells were more in lentiviral vector group(P < 0.05).Compared with the control group, the migration rate of cells was higher, the number of invasion cells were more in lentiviral vector group(P < 0.05).Conclusions EPCs can be isolated from bone marrow, and cultured and identified.The over-expression lentiviral vector of Periostin protein can reduce the growth and proliferation, promote the apotosis, and enhance the migration and invasion ability of EPCs.