联合转染人VEGF165和ANG-1双基因对大鼠骨髓基质干细胞成骨分化的影响

目的]研究联合转染入血管内皮细胞生长因子165(VEGF165)和血管紧张素-1(ANG-1)双基因对大鼠骨髓基质干细胞(BMSCs)成骨分化潜能的影响.方法]将本室构建的编码人VEGF165和ANG-1双基因腺病毒质粒pAd-VIA在QBI-293A细胞内进行包装和扩增,获得腺病毒载体pAd-VIA,将其体外转染大鼠BMSCs,应用诱导培养液定向诱导向成骨细胞分化.实验分为4组:A.腺病毒载体pAd-VIA转染BMSCs并诱导组;B.BMSCs诱导组;C.腺病毒载体pAd-VIA转染BMSCs组;D.单纯BMSCs组.通过碱性磷酸酶(ALP)染色和活性的测定、Ⅰ型胶原免疫荧光染色、茜素红染色来评价联合转染双基因对BMSCs成骨分化潜能的影响.结果]重组腺病毒质粒pAdVIA在QBI-293A细胞内成功包装和扩增,有大量的绿色荧光蛋白表达,体外转染大鼠BMSCs后,A、B组的ALP染色、Ⅰ型胶原免疫荧光染色、茜素红染色为阳性,而C组和D组为阴性.ALP活性表达具有时间相关性,3 d开始表达,7 d到达高峰,在7、14 d,A、B组与C、D组之间ALP表达均具有统计学差异(P＜0.01),A组和B组之间,在各个时间点均无统计学差异(P＞0.05).结论]腺病毒载体pAd-VIA转染大鼠BMSCs后,未明显影响其体外成骨分化潜能.

Effect of adenoviruses-mediated human angiopoietin-1 and VEGF_(165) gene over-expression on ex vivo osteogenic potential of rat bone marrow stromal cells

Objective]The purpose of this study was to investigate the effect of adenoviruses-mediated human angiopoietin-1(ANG-1) and VEGF165 gene over-expression on ex vivo osteogenic potential of rat bone narrowstromal cells.Method]The recombinant pAd-VIA plasmid constructed by our laboratory was packaged and amplified in QBI-293A cells.The pAd-VIA recombined adenovirus was obtained.Rat BMSCs were infected with adenoviruses encoding VEGF165 and ANG-1 at about 80% confluency.The osteogenic potential of transfected ...