title: [Experimental study on transfection of pleiotrophin gene into adipose-derived stem cells of mice].
abstract: OBJECTIVE
  To study the transfection and expression of pleiotrophin (Ptn) gene in mice adipose-derived stem cells (ADSCs) so as to provide a new approach for the treatment of ischemic injury.
  
  
  METHODS
  ADSCs from clean inbred C57BL/6W mice (weighing, 15-20 g) were isolated and cultured in vitro. The cell surface markers (CD29 and CD44) of ADSCs were identified by flow cytometry. The ADSCs were transfected with plasmid pIRES2-LEGFPN1 (containing Ptn gene coding sequence) as experimental group (group A) and with plasmid pLEGFP-N1 (containing GFP gene coding sequence) as control group (group B). After ADSCs were transfected by different plasmids respectively, the cells containing Ptn gene were selected by G418 (the best selected concentration was 200 microg/mL), and the immunophenotype of the cells was identified by flow cytometry after transfection. Meanwhile, real-time fluorescence quantitative PCR and Western blot were used to analyse the expression levels of Ptn mRNA and PTN protein in selected cells.
  
  
  RESULTS
  The mice ADSCs were isolated and cultured successfully in vitro. The positive rates of the cell surface markers CD29 and CD44 of ADSCs were 99.5% and 95.8%, respectively; the double positive rate of CD44 and CD29 was 93.6%. The positive rates of the cell surface markers CD29 and CD44 of ADSCs were 99.1% and 95.6%, respectively after transfection of Ptn gene; the double positive rate of CD44 and CD29 was 93.4%. The expression levels of Ptn gene and PTN protein in group A were significantly higher than those in group B (P < 0.05).
  
  
  CONCLUSIONS
  The ADSCs can be stablely transfected by Ptn gene, the transfected ADSCs can express PTN protein highly, which is a new idea for tissue engineering of vascular reconstruction.
authors.affiliation: Central Laboratory of Chengdu University of Traditional Chinese Medicine, Chengdu Sichuan, 610075, P R China. cjczhzha2002@yahoo.com
authors.forename: Changjin
authors.initials: C
authors.lastname: Chen
authors.forename: Hui
authors.initials: H
authors.lastname: Yuan
authors.forename: Haiying
authors.initials: H
authors.lastname: Gu
authors.forename: Xuemei
authors.initials: X
authors.lastname: Yang
authors.forename: Ziyi
authors.initials: Z
authors.lastname: Zhao
mesh_terms: D017667:Adipocytes / Q000166:cytology* / Q000378:metabolism; D000273:Adipose Tissue / Q000166:cytology; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D016207:Cytokines / Q000235:genetics* / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry; D005822:Genetic Vectors; D051379:Mice; D008810:Mice, Inbred C57BL; D010957:Plasmids / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D023822:Tissue Engineering / Q000379:methods; D014162:Transfection
chemical_list: D002352:Carrier Proteins; D016207:Cytokines; D012333:RNA, Messenger; C066845:pleiotrophin