Lentiviral transduction of CD34(+) cells induces genome-wide epigenetic modifications

PLoS One. 2012;7(11):e48943. doi: 10.1371/journal.pone.0048943. Epub 2012 Nov 7.

Abstract

Epigenetic modifications may occur during in vitro manipulations of stem cells but these effects have remained unexplored in the context of cell and gene therapy protocols. In an experimental model of ex vivo gene modification for hematopoietic gene therapy, human CD34(+) cells were cultured shortly in the presence of cytokines then with a gene transfer lentiviral vector (LV) expected to transduce cells but to have otherwise limited biological effects on the cells. At the end of the culture, the population of cells remained largely similar at the phenotypic level but some epigenetic changes were evident. Exposure of CD34(+) cells to cytokines increased nuclear expression of epigenetic regulators SIRT1 or DNMT1 and caused genome-wide DNA methylation changes. Surprisingly, the LV caused additional and distinct effects. Large-scale genomic DNA methylation analysis showed that balanced methylation changes occurred in about 200 genes following culture of CD34(+) cells in the presence of cytokines but 900 genes were modified following addition of the LV, predominantly increasing CpG methylation. Epigenetic effects resulting from ex vivo culture and from the use of LV may constitute previously unsuspected sources of biological effects in stem cells and may provide new biomarkers to rationally optimize gene and cell therapy protocols.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antigens, CD34 / genetics*
  • Antigens, CD34 / metabolism
  • Cells, Cultured
  • Cytokines / metabolism
  • DNA (Cytosine-5-)-Methyltransferase 1
  • DNA (Cytosine-5-)-Methyltransferases / genetics
  • DNA (Cytosine-5-)-Methyltransferases / metabolism
  • DNA Methylation / genetics
  • Epigenesis, Genetic
  • Epigenomics
  • Fetal Blood / metabolism
  • Genetic Therapy / methods*
  • Genetic Vectors / genetics
  • Genome / genetics
  • Hematopoietic Stem Cells / metabolism
  • Humans
  • Lentivirus / genetics*
  • Lentivirus / metabolism
  • Sirtuin 1 / genetics
  • Sirtuin 1 / metabolism
  • Transduction, Genetic / methods

Substances

  • Antigens, CD34
  • Cytokines
  • DNA (Cytosine-5-)-Methyltransferase 1
  • DNA (Cytosine-5-)-Methyltransferases
  • DNMT1 protein, human
  • SIRT1 protein, human
  • Sirtuin 1

Grant support

YY was supported by the Yamaguchi University in Japan and GC was a recipient of an AXA doctoral fellowship. This study was supported in part by the equipment funds from “Genopole Research-Evry, Conseil General de l’Esson and the University Evry Val d'Essone. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.