Nonviral gene targeting at rDNA locus of human mesenchymal stem cells.
Nonviral gene targeting at rDNA locus of human mesenchymal stem cells.
复制标题
靶向人类间充质干细胞 rDNA 位点的非病毒基因
DOI：
10.1155/2013/135189
复制
发表时间：
2013
影响因子：
--
通讯作者：
Liang D
中科院分区：
文献类型：
--
作者：
Hu Y;Liu X;Long P;Xiao D;Cun J;Li Z;Xue J;Wu Y;Luo S;Wu L;Liang D
背景遗传修饰，例如向MSC基因组中添加外源基因，对于它们作为细胞载体的用途至关重要。由于病毒载体存在插入突变等风险，更安全的非病毒载体引起了人们的广泛关注。方法.在MSC培养基中补充VEGF、bFGF、维生素C和胰岛素转铁蛋白硒X。MTT法、细胞集落形成率测定法、细胞累积数测定法检测细胞的增殖和存活能力。构建质粒pHr 2-NL，转染MSCs。用G418筛选重组子，并用PCR和Southern印迹法进行鉴定。结果碱性成纤维细胞生长因子对MSC生长至关重要，它与维生素C、血管内皮生长因子和ITS-X协同作用，使细胞显着扩增。使用非病毒人核糖体靶向载体将新霉素基因靶向至人MSC的rDNA位点。重组MSC保留了多潜能分化能力，hMSC表面标志物表达的典型水平，以及正常的核型，并且在裸鼠中没有致瘤性。结论.外源基因可以靶向人MSCs的rDNA位点，同时保持MSCs的特性。这是第一个非病毒基因靶向hMSCs。
Background. Genetic modification, such as the addition of exogenous genes to the MSC genome, is crucial to their use as cellular vehicles. Due to the risks associated with viral vectors such as insertional mutagenesis, the safer nonviral vectors have drawn a great deal of attention. Methods. VEGF, bFGF, vitamin C, and insulin-transferrin-selenium-X were supplemented in the MSC culture medium. The cells' proliferation and survival capacity was measured by MTT, determination of the cumulative number of cells, and a colony-forming efficiency assay. The plasmid pHr2-NL was constructed and nucleofected into MSCs. The recombinants were selected using G418 and characterized using PCR and Southern blotting. Results. BFGF is critical to MSC growth and it acted synergistically with vitamin C, VEGF, and ITS-X, causing the cells to expand significantly. The neomycin gene was targeted to the rDNA locus of human MSCs using a nonviral human ribosomal targeting vector. The recombinant MSCs retained multipotential differentiation capacity, typical levels of hMSC surface marker expression, and a normal karyotype, and none were tumorigenic in nude mice. Conclusions. Exogenous genes can be targeted to the rDNA locus of human MSCs while maintaining the characteristics of MSCs. This is the first nonviral gene targeting of hMSCs.
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DOI：
10.1007/s004290050237
发表时间：
1999-05-01
期刊：
ANATOMY AND EMBRYOLOGY
影响因子：
--
作者：
Bittner, RE;Schöfer, C;Wachtler, F
通讯作者：
Wachtler, F
影响因子：
4.5
作者：
Chieregato, Katia;Castegnaro, Silvia;Rodeghiero, Francesco
通讯作者：
Rodeghiero, Francesco
影响因子：
10.4
作者：
Liu, X.;Liu, M.;Xia, J.
通讯作者：
Xia, J.
影响因子：
2.9
作者：
Douglas, Kimberly L.
通讯作者：
Douglas, Kimberly L.
DOI：
10.1111/j.1365-2184.1970.tb00347.x
发表时间：
1970-01-01
期刊：
CELL AND TISSUE KINETICS
影响因子：
--
作者：
FRIEDENSTEIN, AJ;CHAILAKHJAN, RK;LALYKINA, KS
通讯作者：
LALYKINA, KS