title: Effective gene delivery to mesenchymal stem cells based on the reverse transfection and three-dimensional cell culture system.
abstract: OBJECTIVE
  To enhance the level and prolong the duration of gene expression for gene-engineered rat mesenchymal stem cells (MSCs) using non-viral vector.
  
  
  METHODS
  A novel transfection system based on reverse transfection method and three-dimensional (3D) scaffold was developed. The reverse gene transfection system was evaluated for transfection efficiency compared to conventional methods. Collagen sponge and polyethylene terephthalate non-woven fabric were introduced as scaffolds to perform 3D culture with reverse transfection. pDNA coding TGFβ-1 was delivered to MSCs to assess its ability in inducing chondrogenesis with the 3D non-viral reverse transfection system.
  
  
  RESULTS
  The reverse transfection method induced higher transgene levels than the conventional transfection in the presence of serum. The electric charge of the anionic gelatin plays an important role in this system by affecting the release pattern of the gene complexes and through the adsorption of serum protein to the substrate. During a long-time in vitro culture, MSCs cultured on 3D scaffolds exhibited a higher transgene expression level and more sustained transgene expression than those cultured and transfected on the two-dimensional substrate.
  
  
  CONCLUSIONS
  The combination of reverse transfection system with 3D cell culture scaffold benefits the cell proliferation and long-time gene transfection of MSCs.
authors.affiliation: Institute of Pharmaceutics, College of Pharmaceutical Sciences, Zhejiang University, 388 Yuhangtang Road, Hangzhou, 310058, People's Republic of China.
authors.forename: Cai-Xia
authors.initials: CX
authors.lastname: He
authors.forename: Ni
authors.initials: N
authors.lastname: Li
authors.forename: Yu-Lan
authors.initials: YL
authors.lastname: Hu
authors.forename: Xiu-Mei
authors.initials: XM
authors.lastname: Zhu
authors.forename: Hai-Jie
authors.initials: HJ
authors.lastname: Li
authors.forename: Min
authors.initials: M
authors.lastname: Han
authors.forename: Pei-Hong
authors.initials: PH
authors.lastname: Miao
authors.forename: Zhong-Jie
authors.initials: ZJ
authors.lastname: Hu
authors.forename: Gang
authors.initials: G
authors.lastname: Wang
authors.forename: Wen-Quan
authors.initials: WQ
authors.lastname: Liang
authors.forename: Yasuhiko
authors.initials: Y
authors.lastname: Tabata
authors.forename: Jian-Qing
authors.initials: JQ
authors.lastname: Gao
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D006367:HeLa Cells; D006801:Humans; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology; D008855:Microscopy, Electron, Scanning; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D004247:DNA