CAJ | 학술논문

　　背景：骨形态发生蛋白2及转化生长因子β是骨再生中重要的因子，提高其表达可促进骨髓间充质干细胞的成骨分化。目的：构建携带转化生长因子β3和骨形态发生蛋白2基因的慢病毒载体，观察其在骨髓间充质干细胞中的表达情况。方法：应用重组慢病毒技术构建同时携带转化生长因子β3、骨形态发生蛋白2和绿色荧光蛋白基因的重组慢病毒表达载体，并用其转染体外培养的第3代兔骨髓间充质干细胞，以转染携带转化生长因子β3或骨形态发生蛋白2单一基因的慢病毒或单独慢病毒的骨髓间充质干细胞作为对照。转染后1周分别提取各组细胞的总RNA 和蛋白进行检测。结果与结论：荧光显微镜下见转染转化生长因子β3和(或)骨形态发生蛋白2基因3 d 的骨髓间充质干细胞发绿色荧光，转染效率达90%以上。RT-PCR 和 Western blot 结果显示，转染转化生长因子β3和骨形态发生蛋白2基因的骨髓间充质干细胞转化生长因子β3和骨形态发生蛋白2 mRNA 和蛋白的表达均高于单一基因转染组及空白对照组。可见应用慢病毒可成功将转化生长因子β3和骨形态发生蛋白2基因转染至骨髓间充质干细胞并实现其高效表达，且两种基因具有协同促表达作用。
　　배경：골형태발생단백2급전화생장인자β시골재생중중요적인자，제고기표체가촉진골수간충질간세포적성골분화。목적：구건휴대전화생장인자β3화골형태발생단백2기인적만병독재체，관찰기재골수간충질간세포중적표체정황。방법：응용중조만병독기술구건동시휴대전화생장인자β3、골형태발생단백2화록색형광단백기인적중조만병독표체재체，병용기전염체외배양적제3대토골수간충질간세포，이전염휴대전화생장인자β3혹골형태발생단백2단일기인적만병독혹단독만병독적골수간충질간세포작위대조。전염후1주분별제취각조세포적총RNA 화단백진행검측。결과여결론：형광현미경하견전염전화생장인자β3화(혹)골형태발생단백2기인3 d 적골수간충질간세포발록색형광，전염효솔체90%이상。RT-PCR 화 Western blot 결과현시，전염전화생장인자β3화골형태발생단백2기인적골수간충질간세포전화생장인자β3화골형태발생단백2 mRNA 화단백적표체균고우단일기인전염조급공백대조조。가견응용만병독가성공장전화생장인자β3화골형태발생단백2기인전염지골수간충질간세포병실현기고효표체，차량충기인구유협동촉표체작용。
BACKGROUND: Bone morphogenetic protein 2 and transforming growth factor β are important factors in bone regeneration, increasing the expressions of bone morphogenetic protein 2 and transforming growth factor β can promote the osteogenic differentiation of bone marrow mesenchymal stem cells. OBJECTIVE: To construct the lentivirus vector carrying bone morphogenetic protein 2 and transforming growth factor β3, and to observe the expression of lentivirus vector in bone marrow mesenchymal stem cells. METHODS: The recombinant lentiviral vectors carrying transforming growth factor β3, bone morphogenetic protein 2 and green fluorescent protein were constructed with recombinant lentiviral technology, and then the recombinant lentiviral vectors were used to transfect the passage 3 rabbit bone marrow mesenchymal stem cells in vitro cultured (transfection group). The bone marrow mesenchymal stem cells transfected with single gene lentivirals (single gene transfection group) carrying transforming growth factor β3 and bone morphogenetic protein 2 or single lentivirals were as control (control group). At 1 week after trasfection, the total RNA and protein were extracted from each group for detection. RESULTS AND CONCLUSION: The green fluorescence bone marrow mesenchymal stem cells transfected with transforming growth factor β3 and bone morphogenetic protein 2 gene for 3 days could be observed under fluorescence microscope, and the transfection efficiency was over 90%. Reverse transcription-PCR and Western blot results showed the mRNA and protein expressions of transforming growth factor β3 and bone morphogenetic protein 2 in the transfection group were higher than those in the single gene transfection group and the control group. The results indicate that lentivirus can successful y transfect transforming growth factor β3 and bone morphogenetic protein 2 into the bone marrow msenchymal stem cells and achieve its high expression, and these two genes have the synergistic effect of promoting expression.