PMID- 19399895
OWN - NLM
STAT- MEDLINE
DCOM- 20090728
LR  - 20211020
IS  - 1096-9861 (Electronic)
IS  - 0021-9967 (Print)
IS  - 0021-9967 (Linking)
VI  - 515
IP  - 1
DP  - 2009 Jul 1
TI  - The dynamics of long-term transgene expression in engrafted neural stem cells.
PG  - 83-92
LID - 10.1002/cne.21957 [doi]
AB  - To assess the dynamics and confounding variables that influence transgene 
      expression in neural stem cells (NSCs), we generated distinct NSC clones from the 
      same pool of cells, carrying the same reporter gene transcribed from the same 
      promoter, transduced by the same retroviral vector, and transplanted similarly at 
      the same differentiation state, at the same time and location, into the brains of 
      newborn mouse littermates, and monitored in parallel for over a year in vivo 
      (without immunosuppression). Therefore, the sole variables were transgene 
      chromosomal insertion site and copy number. We then adapted and optimized a 
      technique that tests, at the single cell level, persistence of stem cell-mediated 
      transgene expression in vivo based on correlating the presence of the transgene 
      in a given NSC's nucleus (by fluorescence in situ hybridization [FISH]) with the 
      frequency of that transgene's product within the same cell (by combined 
      immunohistochemistry [IHC]). Under the above-stated conditions, insertion site is 
      likely the most contributory variable dictating transgene downregulation in an 
      NSC after 3 months in vivo. We also observed that this obstacle could be 
      effectively and safely counteracted by simple serial infections (as few as three) 
      inserting redundant copies of the transgene into the prospective donor NSC. (The 
      preservation of normal growth control mechanisms and an absence of tumorigenic 
      potential can be readily screened and ensured ex vivo prior to transplantation.) 
      The combined FISH/IHC strategy employed here for monitoring the dynamics of 
      transgene expression at the single cell level in vivo may be used for other types 
      of therapeutic and housekeeping genes in endogenous and exogenous stem cells of 
      many organs and lineages.
CI  - Copyright 2009 Wiley-Liss, Inc.
FAU - Lee, Jean-Pyo
AU  - Lee JP
AD  - The Burnham Institute for Medical Research, La Jolla, California 92037, USA.
FAU - Tsai, David J
AU  - Tsai DJ
FAU - In Park, Kook
AU  - In Park K
FAU - Harvey, Alan R
AU  - Harvey AR
FAU - Snyder, Evan Y
AU  - Snyder EY
LA  - eng
GR  - P20 GM075059/GM/NIGMS NIH HHS/United States
GR  - P20 GM075059-01/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - J Comp Neurol
JT  - The Journal of comparative neurology
JID - 0406041
SB  - IM
MH  - Animals
MH  - Animals, Newborn
MH  - Brain Tissue Transplantation/*methods
MH  - Cell Culture Techniques
MH  - Cell Differentiation/genetics
MH  - Cell Proliferation
MH  - Clone Cells
MH  - Female
MH  - Gene Dosage/genetics
MH  - Gene Expression Regulation/*genetics
MH  - Genes, Reporter/genetics
MH  - Genetic Vectors/*genetics
MH  - Graft Survival/genetics
MH  - Male
MH  - Mice
MH  - Neurons/cytology/metabolism
MH  - Retroviridae/genetics
MH  - Stem Cell Transplantation/*methods
MH  - Stem Cells/cytology/metabolism
MH  - Time
MH  - Transfection/*methods
MH  - Transgenes/*genetics
PMC - PMC2703611
MID - NIHMS89633
EDAT- 2009/04/29 09:00
MHDA- 2009/07/29 09:00
PMCR- 2010/07/01
CRDT- 2009/04/29 09:00
PHST- 2009/04/29 09:00 [entrez]
PHST- 2009/04/29 09:00 [pubmed]
PHST- 2009/07/29 09:00 [medline]
PHST- 2010/07/01 00:00 [pmc-release]
AID - 10.1002/cne.21957 [doi]
PST - ppublish
SO  - J Comp Neurol. 2009 Jul 1;515(1):83-92. doi: 10.1002/cne.21957.