RRC ID 62158
Author Muramatsu M, Hanazono Y, Ogasawara Y, Okada T, Mizukami H, Kume A, Mizoguchi H, Ozawa K.
Title Reversible integration of the dominant negative retinoid receptor gene for ex vivo expansion of hematopoietic stem/progenitor cells.
Journal Biochem Biophys Res Commun
Abstract Since hematopoietic stem cells (HSCs) differentiate readily ex vivo resulting in the loss of self-renewal and engraftment abilities, the transient block of differentiation is essential to maintain those abilities during their ex vivo expansion culture. To this end, we developed a method of reversible integration of the dominant negative retinoic acid receptor (DN-RAR) gene, a differentiation-blocking gene, into cells utilizing the Cre/loxP-dependent gene recombination system. The murine immature hematopoietic 32D cells differentiate into mature neutrophils upon G-CSF treatment. However, 32D cells transduced with a retroviral vector expressing the DN-RAR gene put between two loxP sites continued to proliferate without showing differentiation even in the presence of G-CSF. After the cells were fully amplified, the cells were transduced with the Cre recombinase gene. The cells then restored the ability to differentiate into mature neutrophils upon G-CSF treatment. PCR analysis showed that the DN-RAR gene was efficiently removed from the genome by introduction of the Cre gene. This system may eventually be applicable to the ex vivo expansion of HSCs.
Volume 285(4)
Pages 891-6
Published 2001-7-27
DOI 10.1006/bbrc.2001.5264
PII S0006-291X(01)95264-7
PMID 11467834
MeSH Animals Cell Culture Techniques / methods Cell Differentiation Cell Division Hematopoietic Stem Cells / cytology* Integrases Mice Neutrophils / cytology* Receptors, Retinoic Acid / genetics* Recombination, Genetic Transformation, Genetic Viral Proteins
IF 2.985
Human and Animal Cells 10T1/2(RCB0247)