Simple generation of human induced pluripotent stem cells using poly-beta-amino esters as the non-viral gene delivery system

J Biol Chem. 2011 Apr 8;286(14):12417-28. doi: 10.1074/jbc.M110.168013. Epub 2011 Feb 1.

Abstract

Reprogramming of somatic cells to induced pluripotent stem (iPS) cells can be achieved by the delivery of a combination of transcription factors, including Oct4, Sox2, Klf4, and c-Myc. Retroviral and lentiviral vectors are commonly used to express these four reprogramming factors separately and obtain reprogrammed iPS cells. Although efficient and reproducible, these approaches involve the time-consuming and labor-intensive production of retroviral or lentiviral particles together with a high risk of working with potentially harmful viruses overexpressing potent oncogenes, such as c-Myc. Here, we describe a simple method to produce bona fide iPS cells from human fibroblasts using poly-β-amino esters as the transfection reagent for the delivery of a single CAG-driven polycistronic plasmid expressing Oct4, Sox2, Klf4, c-Myc, and a GFP reporter gene (OSKMG). We demonstrate for the first time that poly-β-amino esters can be used to deliver a single polycistronic reprogramming vector into human fibroblasts, achieving significantly higher transfection efficiency than with conventional transfection reagents. After a protocol of serial transfections using poly-β-amino esters, we report a simple methodology to generate human iPS cells from human fibroblasts avoiding the use of viral vectors.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Differentiation / genetics
  • Cell Differentiation / physiology
  • Cell Line
  • Cells, Cultured
  • Cellular Reprogramming / genetics
  • Cellular Reprogramming / physiology
  • Fibroblasts / cytology
  • Fibroblasts / metabolism
  • Fluorescent Antibody Technique
  • Gene Transfer Techniques*
  • Humans
  • Induced Pluripotent Stem Cells / cytology*
  • Induced Pluripotent Stem Cells / metabolism*
  • Karyotyping
  • Octamer Transcription Factor-3 / genetics
  • Octamer Transcription Factor-3 / physiology
  • Polymers / chemistry*
  • Proto-Oncogene Proteins c-myc / genetics
  • Proto-Oncogene Proteins c-myc / physiology
  • SOXB1 Transcription Factors / genetics
  • SOXB1 Transcription Factors / metabolism
  • Transfection

Substances

  • Octamer Transcription Factor-3
  • Polymers
  • Proto-Oncogene Proteins c-myc
  • SOXB1 Transcription Factors
  • poly(beta-amino ester)