Tightly Regulated and Homogeneous Transgene Expression in Human Adipose-Derived Mesenchymal Stem Cells by Lentivirus with Tet-Off System

Hiroyuki Moriyama, Mariko Moriyama, Kei Sawaragi, Hanayuki Okura, Akihiro Ichinose, Akifumi Matsuyama, Takao Hayakawa

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Genetic modification of human adipose tissue-derived multilineage progenitor cells (hADMPCs) is highly valuable for their exploitation in therapeutic applications. Here, we have developed a novel single tet-off lentiviral vector platform. This vector combines (1) a modified tetracycline (tet)-response element composite promoter, (2) a multi-cistronic strategy to express an improved version of the tet-controlled transactivator and the blasticidin resistance gene under the control of a ubiquitous promoter, and (3) acceptor sites for easy recombination cloning of the gene of interest. In the present study, we used the cytomegalovirus (CMV) or the elongation factor 1 α (EF-1α) promoter as the ubiquitous promoter, and EGFP was introduced as the gene of interest. hADMPCs transduced with a lentiviral vector carrying either the CMV promoter or the EF-1α promoter were effectively selected by blasticidin without affecting their stem cell properties, and EGFP expression was strictly regulated by doxycycline (Dox) treatment in these cells. However, the single tet-off lentiviral vector carrying the EF-1α promoter provided more homogenous expression of EGFP in hADMPCs. Intriguingly, differentiated cells from these Dox-responsive cell lines constitutively expressed EGFP only in the absence of Dox. This single tet-off lentiviral vector thus provides an important tool for applied research on hADMPCs.

Original languageEnglish
Article numbere66274
JournalPloS one
Volume8
Issue number6
DOIs
Publication statusPublished - 12-06-2013

Fingerprint

Lentivirus
Tetracycline
Stem cells
Mesenchymal Stromal Cells
Transgenes
transgenes
stem cells
Peptide Elongation Factor 1
Stem Cells
promoter regions
tetracycline
Adipose Tissue
Doxycycline
Tissue
adipose tissue
doxycycline
Genes
Cytomegalovirus
Trans-Activators
Cloning

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

Cite this

Moriyama, Hiroyuki ; Moriyama, Mariko ; Sawaragi, Kei ; Okura, Hanayuki ; Ichinose, Akihiro ; Matsuyama, Akifumi ; Hayakawa, Takao. / Tightly Regulated and Homogeneous Transgene Expression in Human Adipose-Derived Mesenchymal Stem Cells by Lentivirus with Tet-Off System. In: PloS one. 2013 ; Vol. 8, No. 6.
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Tightly Regulated and Homogeneous Transgene Expression in Human Adipose-Derived Mesenchymal Stem Cells by Lentivirus with Tet-Off System. / Moriyama, Hiroyuki; Moriyama, Mariko; Sawaragi, Kei; Okura, Hanayuki; Ichinose, Akihiro; Matsuyama, Akifumi; Hayakawa, Takao.

In: PloS one, Vol. 8, No. 6, e66274, 12.06.2013.

Research output: Contribution to journalArticle

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