摘要
目的 构建重组病毒载体,将较多细胞因子通过病毒载体转入种子细胞,提高细胞增殖及分泌基质能力,维持表型,延缓老化。方法 应用基因工程技术,构建缺陷型逆转录病毒重组载体和缺陷性腺病毒重组载体。分别通过PT67包装细胞筛选克隆及293细胞扩增,获得假病毒上清液,经浓缩纯化后转染靶细胞。结果 成功构建胶质细胞源性神经营养因子(GDNF)、Fas配体(Fas-Ligand)、绿色荧光蛋白(GFP)等逆转录病毒载体以及TGFβ1、人骨形成蛋白因子-7(hBMP7)、端粒酶、Smad3、Smad7等腺病毒载体,其中GFP-逆转录病毒重组载体在软骨细胞表达至2月。结论 通过有效的含目的基因重组载体转染种子细胞,延缓体外培养细胞的老化及促进细胞的增殖与基质分泌,获得组织工程所需要的大量种子细胞,为组织工程构建奠定了基础。
Objective The general principle of tissue engineering is the use of seed cells and biodegradable polymers to generate new tissue, in which seed cell is the key tissue. Construction of viral vector will provide a tool to transfect the genes of growth factors into the cells and thus promote cell proliferation and matrix production, maintain cell phenotype and delay cell aging. Methods Two types of viral vectors have been constructed. One is the retro-viral vector, and the other is the recombinant adenoviral vector. Both viral vectors were packaged in PT67 and 293 cells respectively to obtain cell clone and viral containing supernatant and concentrated virus. The obtained recombinant viruses were then used to infected cells. Results In two years, our laboratory have constructed GDNF, Fas-ligand and GFP retroviral vectors and TGF-β1 , hBMP7, human teromerase, human SmadS and Smad7 adeno Viral vectors . Among them, GFP expression mediated by retroviral vector could maintained up to 2 months. Conclusion Gene transfection of seed cells with various viral vectors containing specific genes can achieve the goals of delaying cell aging, prompting cell proliferation and matrix production and thus obtain enough seed cells for the tissue engineering of different types of tissues, which will further promote the development of tissue engineering.
出处
《上海第二医科大学学报》
CSCD
2004年第4期311-314,共4页
Acta Universitatis Medicinalis Secondae Shanghai
基金
国家重点基础研究发展规划资助项目("973"项目)(1999054305).
