title: Nonviral gene targeting at rDNA locus of human mesenchymal stem cells.
abstract: 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.
authors.affiliation: State Key Laboratory of Medical Genetics, Central South University, 110 Xiangya Road, Changsha, Hunan 410078, China.
authors.forename: Youjin
authors.initials: Y
authors.lastname: Hu
authors.forename: Xionghao
authors.initials: X
authors.lastname: Liu
authors.forename: Panpan
authors.initials: P
authors.lastname: Long
authors.forename: Di
authors.initials: D
authors.lastname: Xiao
authors.forename: Jintao
authors.initials: J
authors.lastname: Cun
authors.forename: Zhuo
authors.initials: Z
authors.lastname: Li
authors.forename: Jinfeng
authors.initials: J
authors.lastname: Xue
authors.forename: Yong
authors.initials: Y
authors.lastname: Wu
authors.forename: Sha
authors.initials: S
authors.lastname: Luo
authors.forename: Lingqian
authors.initials: L
authors.lastname: Wu
authors.forename: Desheng
authors.initials: D
authors.lastname: Liang
mesh_terms: D000818:Animals; D000954:Antigens, Surface / Q000378:metabolism; D001205:Ascorbic Acid / Q000494:pharmacology; D063646:Carcinogenesis / Q000187:drug effects / Q000473:pathology; D002454:Cell Differentiation / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004275:DNA, Ribosomal / Q000235:genetics*; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D018390:Gene Targeting; D056426:Genetic Loci / Q000235:genetics*; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D007621:Karyotyping; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D051379:Mice; D008819:Mice, Nude; D016297:Mutagenesis, Site-Directed; D012643:Selenium / Q000494:pharmacology; D014168:Transferrin / Q000494:pharmacology; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology; D014780:Viruses / Q000378:metabolism
chemical_list: D000954:Antigens, Surface; D004275:DNA, Ribosomal; D007328:Insulin; D036341:Intercellular Signaling Peptides and Proteins; D014168:Transferrin; D042461:Vascular Endothelial Growth Factor A; D016222:Fibroblast Growth Factor 2; D012643:Selenium; D001205:Ascorbic Acid