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Critical factors influencing stable transduction of human CD34(+) cells with HIV-1-derived lentiviral vectors.
Mol Ther. 2000 Jul; 2(1):71-80.MT

Abstract

Lentiviral vectors have been proposed as a more efficient alternative to Moloney murine leukemia virus-based retroviral vectors for transduction of human hematopoietic progenitors and stem cells. These studies were designed to evaluate the conditions that influence transduction frequency of CD34(+) progenitors, with the goal of optimizing efficiency of stable gene transfer with lentiviral vectors. CD34(+) human cord blood cells and 293 cells were transduced with a human immunodeficiency virus (HIV)-1 derived lentiviral vector pseudotyped with vesicular stomatitis virus glycoprotein and carrying an internal human cytomegalovirus promoter driving enhanced green fluorescent protein (eGFP) expression. Using fluorescence-activated cell sorting analysis of eGFP, we observed pseudotransduction beginning at the time of vector addition and lasting up to 24 h in CD34(+) cells and up to 72 h in 293 cells. Integrase-defective lentiviral vector caused transient eGFP expression for up to 10 days in CD34(+) cells and for up to 14 days in 293 cells. Protamine sulfate conferred no increase in transduction efficiency of CD34(+) cells on fibronectin-coated plates. Transduction frequency was related directly to vector concentration and not to multiplicity of infection across the ranges tested. First- and second-generation lentiviral vectors transduced CD34(+) cells equally, demonstrating a lack of dependence on HIV-1 accessory proteins. These findings will be useful for the optimal utilization of this new class of vectors for transduction of human hematopoietic stem cells.

Authors+Show Affiliations

Department of Molecular Microbiology and Immunology, University of Southern California Medical School, Los Angeles, CA 90033, USA.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

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

Language

eng

PubMed ID

10899830

Citation

Haas, D L., et al. "Critical Factors Influencing Stable Transduction of Human CD34(+) Cells With HIV-1-derived Lentiviral Vectors." Molecular Therapy : the Journal of the American Society of Gene Therapy, vol. 2, no. 1, 2000, pp. 71-80.
Haas DL, Case SS, Crooks GM, et al. Critical factors influencing stable transduction of human CD34(+) cells with HIV-1-derived lentiviral vectors. Mol Ther. 2000;2(1):71-80.
Haas, D. L., Case, S. S., Crooks, G. M., & Kohn, D. B. (2000). Critical factors influencing stable transduction of human CD34(+) cells with HIV-1-derived lentiviral vectors. Molecular Therapy : the Journal of the American Society of Gene Therapy, 2(1), 71-80.
Haas DL, et al. Critical Factors Influencing Stable Transduction of Human CD34(+) Cells With HIV-1-derived Lentiviral Vectors. Mol Ther. 2000;2(1):71-80. PubMed PMID: 10899830.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Critical factors influencing stable transduction of human CD34(+) cells with HIV-1-derived lentiviral vectors. AU - Haas,D L, AU - Case,S S, AU - Crooks,G M, AU - Kohn,D B, PY - 2000/7/19/pubmed PY - 2000/9/23/medline PY - 2000/7/19/entrez SP - 71 EP - 80 JF - Molecular therapy : the journal of the American Society of Gene Therapy JO - Mol Ther VL - 2 IS - 1 N2 - Lentiviral vectors have been proposed as a more efficient alternative to Moloney murine leukemia virus-based retroviral vectors for transduction of human hematopoietic progenitors and stem cells. These studies were designed to evaluate the conditions that influence transduction frequency of CD34(+) progenitors, with the goal of optimizing efficiency of stable gene transfer with lentiviral vectors. CD34(+) human cord blood cells and 293 cells were transduced with a human immunodeficiency virus (HIV)-1 derived lentiviral vector pseudotyped with vesicular stomatitis virus glycoprotein and carrying an internal human cytomegalovirus promoter driving enhanced green fluorescent protein (eGFP) expression. Using fluorescence-activated cell sorting analysis of eGFP, we observed pseudotransduction beginning at the time of vector addition and lasting up to 24 h in CD34(+) cells and up to 72 h in 293 cells. Integrase-defective lentiviral vector caused transient eGFP expression for up to 10 days in CD34(+) cells and for up to 14 days in 293 cells. Protamine sulfate conferred no increase in transduction efficiency of CD34(+) cells on fibronectin-coated plates. Transduction frequency was related directly to vector concentration and not to multiplicity of infection across the ranges tested. First- and second-generation lentiviral vectors transduced CD34(+) cells equally, demonstrating a lack of dependence on HIV-1 accessory proteins. These findings will be useful for the optimal utilization of this new class of vectors for transduction of human hematopoietic stem cells. SN - 1525-0016 UR - https://www.unboundmedicine.com/medline/citation/10899830/Critical_factors_influencing_stable_transduction_of_human_CD34_+__cells_with_HIV_1_derived_lentiviral_vectors_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1525-0016(00)90094-7 DB - PRIME DP - Unbound Medicine ER -