脂质体介导骨保护素基因转染大鼠骨髓基质干细胞及其表达

背景：基因修饰骨组织工程种子细胞,可提高对骨缺损的修复能力。 目的：构建含有骨保护素的真核表达载体并检测转染骨髓基质干细胞后的表达。 方法：取4周龄SD大鼠胫骨和股骨行骨髓基质干细胞的分离和培养,分为载体组、空载体组、对照组。空载体组转染plRES2-EGFP载体,载体组转染plRES2-EGFP-OPG。 结果与结论：转染后激光扫描共聚焦显微镜观察可见骨髓基质干细胞内绿色荧光蛋白表达;RT-PCR检测结果可见质粒载体组在1 200 bp有明显条带,空载体组及对照组未见表达;免疫细胞化学检测骨髓基质干细胞内骨保护素蛋白呈阳性;MTT法检测各组细胞增殖活力无明显改变(P > 0.05)。证实应用plRES2-EGFP-骨保护素质粒转染大鼠骨髓基质干细胞,可获得外源性骨保护素的基因及蛋白表达。

Expression of osteoprotegerin gene mediated with liposome in rat bone marrow stromal cells

BACKGROUND:Genetically modified seed cells for bone tissue engineering can improve the ability to repair bone defects. OBJECTIVE:To detect the expression of osteoprotegerin (OPG) in rat transfected bone marrow stromal cells (BMSCs). METHODS:BMSCs were aseptically obtained from the femur and tibia of 4-week-old SD rats and cultured in vitro. BMSCs were divided into three groups: plasmid group, blank plasmid group and non-transfection group. RESULTS AND CONCLUSION:After transfection, green fluorescent protein was observed in BMSCs under the laser scanning confocal microscope. There was obvious mark at 1 200 bp in plasmid group, but there was no expression of OPG mRNA in blank plasmid group and non-transfection group. The OPG protein was positive in BMSCs detected by immunocytochemistry examination. There were no significant difference in cell proliferation in each group (P > 0.05). plRES2- EGFP-OPG plasmid transfected BMSCs can express exogenous OPG and proteins.