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Highly efficient transduction of the green fluorescent protein gene in human umbilical cord blood stem cells capable of cobblestone formation in long-term cultures and multilineage engraftment of immunodeficient mice.
Blood. 1998 Dec 01; 92(11):4013-22.Blood

Abstract

Purified CD34(+) and CD34(+)CD38(-) human umbilical cord blood (UCB) cells were transduced with the recombinant variant of Moloney murine leukemia virus (MoMLV) MFG-EGFP or with SF-EGFP, in which EGFP expression is driven by a hybrid promoter of the spleen focus-forming virus (SFFV) and the murine embryonic stem cell virus (MESV). Infectious MFG-EGFP virus was produced by an amphotropic virus producer cell line (GP+envAm12). SF-EGFP was produced in the PG13 cell line pseudotyped for the gibbon ape leukemia virus (GaLV) envelope proteins. Using a 2-day growth factor prestimulation, followed by a 2-day, fibronectin fragment CH-296-supported transduction, CD34(+) and CD34(+)CD38(-) UCB subsets were efficiently transduced using either vector. The use of the SF-EGFP/PG13 retroviral packaging cell combination consistently resulted in twofold higher levels of EGFP-expressing cells than the MFG-EGFP/Am12 combination. Transplantation of 10(5) input equivalent transduced CD34(+) or 5 x 10(3) input equivalent CD34(+)CD38(-) UCB cells in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice resulted in median engraftment percentages of 8% and 5%, respectively, which showed that the in vivo repopulating ability of the cells had been retained. In addition, mice engrafted after transplantation of transduced CD34(+) cells using the MFG-EGFP/Am12 or the SF-EGFP/PG13 combination expressed EGFP with median values of 2% and 23% of human CD45(+) cells, respectively, which showed that the NOD/SCID repopulating cells were successfully transduced. EGFP+ cells were found in all human hematopoietic lineages produced in NOD/SCID mice including human progenitors with in vitro clonogenic ability. EGFP-expressing cells were also detected in the human cobblestone area-forming cell (CAFC) assay at 2 to 6 weeks of culture on the murine stromal cell line FBMD-1. During the transduction procedure the absolute numbers of CAFC week 6 increased 5- to 10-fold. The transduction efficiency of this progenitor cell subset was similar to the fraction of EGFP+ human cells in the bone marrow of the NOD/SCID mice transplanted with MFG-EGFP/Am12 or SF-EGFP/PG13 transduced CD34(+) cells, ie, 6% and 27%, respectively. The study thus shows that purified CD34(+) and highly purified CD34(+)CD38(-) UCB cells can be transduced efficiently with preservation of repopulating ability. The SF-EGFP/PG13 vector/packaging cell combination was much more effective in transducing repopulating cells than the MFG-EGFP/Am12 combination.

Authors+Show Affiliations

Institute of Hematology, Erasmus University Rotterdam, The Netherlands; and the Department of Cryobiology and Cell Therapy, Institut de Recerca Oncologica, Barcelona, Spain.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

9834203

Citation

van Hennik, P B., et al. "Highly Efficient Transduction of the Green Fluorescent Protein Gene in Human Umbilical Cord Blood Stem Cells Capable of Cobblestone Formation in Long-term Cultures and Multilineage Engraftment of Immunodeficient Mice." Blood, vol. 92, no. 11, 1998, pp. 4013-22.
van Hennik PB, Verstegen MM, Bierhuizen MF, et al. Highly efficient transduction of the green fluorescent protein gene in human umbilical cord blood stem cells capable of cobblestone formation in long-term cultures and multilineage engraftment of immunodeficient mice. Blood. 1998;92(11):4013-22.
van Hennik, P. B., Verstegen, M. M., Bierhuizen, M. F., Limón, A., Wognum, A. W., Cancelas, J. A., Barquinero, J., Ploemacher, R. E., & Wagemaker, G. (1998). Highly efficient transduction of the green fluorescent protein gene in human umbilical cord blood stem cells capable of cobblestone formation in long-term cultures and multilineage engraftment of immunodeficient mice. Blood, 92(11), 4013-22.
van Hennik PB, et al. Highly Efficient Transduction of the Green Fluorescent Protein Gene in Human Umbilical Cord Blood Stem Cells Capable of Cobblestone Formation in Long-term Cultures and Multilineage Engraftment of Immunodeficient Mice. Blood. 1998 Dec 1;92(11):4013-22. PubMed PMID: 9834203.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Highly efficient transduction of the green fluorescent protein gene in human umbilical cord blood stem cells capable of cobblestone formation in long-term cultures and multilineage engraftment of immunodeficient mice. AU - van Hennik,P B, AU - Verstegen,M M, AU - Bierhuizen,M F, AU - Limón,A, AU - Wognum,A W, AU - Cancelas,J A, AU - Barquinero,J, AU - Ploemacher,R E, AU - Wagemaker,G, PY - 1998/12/3/pubmed PY - 1998/12/3/medline PY - 1998/12/3/entrez SP - 4013 EP - 22 JF - Blood JO - Blood VL - 92 IS - 11 N2 - Purified CD34(+) and CD34(+)CD38(-) human umbilical cord blood (UCB) cells were transduced with the recombinant variant of Moloney murine leukemia virus (MoMLV) MFG-EGFP or with SF-EGFP, in which EGFP expression is driven by a hybrid promoter of the spleen focus-forming virus (SFFV) and the murine embryonic stem cell virus (MESV). Infectious MFG-EGFP virus was produced by an amphotropic virus producer cell line (GP+envAm12). SF-EGFP was produced in the PG13 cell line pseudotyped for the gibbon ape leukemia virus (GaLV) envelope proteins. Using a 2-day growth factor prestimulation, followed by a 2-day, fibronectin fragment CH-296-supported transduction, CD34(+) and CD34(+)CD38(-) UCB subsets were efficiently transduced using either vector. The use of the SF-EGFP/PG13 retroviral packaging cell combination consistently resulted in twofold higher levels of EGFP-expressing cells than the MFG-EGFP/Am12 combination. Transplantation of 10(5) input equivalent transduced CD34(+) or 5 x 10(3) input equivalent CD34(+)CD38(-) UCB cells in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice resulted in median engraftment percentages of 8% and 5%, respectively, which showed that the in vivo repopulating ability of the cells had been retained. In addition, mice engrafted after transplantation of transduced CD34(+) cells using the MFG-EGFP/Am12 or the SF-EGFP/PG13 combination expressed EGFP with median values of 2% and 23% of human CD45(+) cells, respectively, which showed that the NOD/SCID repopulating cells were successfully transduced. EGFP+ cells were found in all human hematopoietic lineages produced in NOD/SCID mice including human progenitors with in vitro clonogenic ability. EGFP-expressing cells were also detected in the human cobblestone area-forming cell (CAFC) assay at 2 to 6 weeks of culture on the murine stromal cell line FBMD-1. During the transduction procedure the absolute numbers of CAFC week 6 increased 5- to 10-fold. The transduction efficiency of this progenitor cell subset was similar to the fraction of EGFP+ human cells in the bone marrow of the NOD/SCID mice transplanted with MFG-EGFP/Am12 or SF-EGFP/PG13 transduced CD34(+) cells, ie, 6% and 27%, respectively. The study thus shows that purified CD34(+) and highly purified CD34(+)CD38(-) UCB cells can be transduced efficiently with preservation of repopulating ability. The SF-EGFP/PG13 vector/packaging cell combination was much more effective in transducing repopulating cells than the MFG-EGFP/Am12 combination. SN - 0006-4971 UR - https://www.unboundmedicine.com/medline/citation/9834203/Highly_efficient_transduction_of_the_green_fluorescent_protein_gene_in_human_umbilical_cord_blood_stem_cells_capable_of_cobblestone_formation_in_long_term_cultures_and_multilineage_engraftment_of_immunodeficient_mice_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0006-4971(20)57729-X DB - PRIME DP - Unbound Medicine ER -