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Low level of gene transfer to and engraftment of murine bone marrow cells from long-term bone marrow cultures

Relander, Thomas LU ; Fahlman, Cecilia; Karlsson, Stefan LU and Richter, Johan LU (2000) In Experimental Hematology 28(4). p.373-381
Abstract
OBJECTIVE: We wanted to determine whether the long-term bone marrow culture (LTBMC) transduction system would lead to efficient gene transfer and engraftment of murine repopulating hematopoietic stem cells (HSC), particularly in nonablated recipients. MATERIALS AND METHODS: Congenic mouse strains expressing Ly 5.1 or Ly 5.2 and the GP+E86 cell line producing the MGirL22Y vector carrying the gene for enhanced GFP were used. Murine LTBMCs were established and demi-depopulated on days 7 and 14 with addition of vector supernatant on days 8 and 15. RESULTS: Cell recovery on day 21 was 21.3%+/-3.8% of input cells and CFU-C recovery was 9.7+/-3.4% as compared with CFU-C of input cells. In vitro transduction efficiency determined by CFU-C... (More)
OBJECTIVE: We wanted to determine whether the long-term bone marrow culture (LTBMC) transduction system would lead to efficient gene transfer and engraftment of murine repopulating hematopoietic stem cells (HSC), particularly in nonablated recipients. MATERIALS AND METHODS: Congenic mouse strains expressing Ly 5.1 or Ly 5.2 and the GP+E86 cell line producing the MGirL22Y vector carrying the gene for enhanced GFP were used. Murine LTBMCs were established and demi-depopulated on days 7 and 14 with addition of vector supernatant on days 8 and 15. RESULTS: Cell recovery on day 21 was 21.3%+/-3.8% of input cells and CFU-C recovery was 9.7+/-3.4% as compared with CFU-C of input cells. In vitro transduction efficiency determined by CFU-C expressing GFP was 22.2%+/-1.6%. In irradiated (950 cGy) mice transplanted with 2x10(6) LTBMC cells, 94% of nucleated cells in the blood at week 16 were of donor origin. However, GFP was only detected at low level in a few animals at week 4 and not later. Analysis of bone marrow from these mice at week 20 did not show any GFP expression and semiquantitative PCR revealed a transgene level of <1%. When 3.5-20.8x10(6) LTBMC cells (corresponding to 20-100x10(6) fresh cells) were transplanted to nonablated recipients, no engraftment or GFP expression were detected. Competitive repopulation experiments showed that the long-term repopulation ability (LTRA) of the LTMC cells was only 7% of fresh cells. CONCLUSION: These results indicate that LTBMC transduction of murine cells leads to low-level transduction of progenitors, no gene transfer to repopulating stem cells, and reduction in LTRA in ablated and nonablated recipients. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Gene transfer, Long-term culture, Engraftment, Bone marrow
in
Experimental Hematology
volume
28
issue
4
pages
373 - 381
publisher
Elsevier
external identifiers
  • pmid:10781895
  • scopus:0034058418
ISSN
1873-2399
DOI
10.1016/S0301-472X(00)00131-4
language
English
LU publication?
yes
id
9a3f7a1e-4554-468e-a6f5-542c613a232a (old id 1116586)
date added to LUP
2008-07-02 08:46:45
date last changed
2017-01-01 04:33:43
@article{9a3f7a1e-4554-468e-a6f5-542c613a232a,
  abstract     = {OBJECTIVE: We wanted to determine whether the long-term bone marrow culture (LTBMC) transduction system would lead to efficient gene transfer and engraftment of murine repopulating hematopoietic stem cells (HSC), particularly in nonablated recipients. MATERIALS AND METHODS: Congenic mouse strains expressing Ly 5.1 or Ly 5.2 and the GP+E86 cell line producing the MGirL22Y vector carrying the gene for enhanced GFP were used. Murine LTBMCs were established and demi-depopulated on days 7 and 14 with addition of vector supernatant on days 8 and 15. RESULTS: Cell recovery on day 21 was 21.3%+/-3.8% of input cells and CFU-C recovery was 9.7+/-3.4% as compared with CFU-C of input cells. In vitro transduction efficiency determined by CFU-C expressing GFP was 22.2%+/-1.6%. In irradiated (950 cGy) mice transplanted with 2x10(6) LTBMC cells, 94% of nucleated cells in the blood at week 16 were of donor origin. However, GFP was only detected at low level in a few animals at week 4 and not later. Analysis of bone marrow from these mice at week 20 did not show any GFP expression and semiquantitative PCR revealed a transgene level of &lt;1%. When 3.5-20.8x10(6) LTBMC cells (corresponding to 20-100x10(6) fresh cells) were transplanted to nonablated recipients, no engraftment or GFP expression were detected. Competitive repopulation experiments showed that the long-term repopulation ability (LTRA) of the LTMC cells was only 7% of fresh cells. CONCLUSION: These results indicate that LTBMC transduction of murine cells leads to low-level transduction of progenitors, no gene transfer to repopulating stem cells, and reduction in LTRA in ablated and nonablated recipients.},
  author       = {Relander, Thomas and Fahlman, Cecilia and Karlsson, Stefan and Richter, Johan},
  issn         = {1873-2399},
  keyword      = {Gene transfer,Long-term culture,Engraftment,Bone marrow},
  language     = {eng},
  number       = {4},
  pages        = {373--381},
  publisher    = {Elsevier},
  series       = {Experimental Hematology},
  title        = {Low level of gene transfer to and engraftment of murine bone marrow cells from long-term bone marrow cultures},
  url          = {http://dx.doi.org/10.1016/S0301-472X(00)00131-4},
  volume       = {28},
  year         = {2000},
}