Factors affecting retrovirus-mediated gene transfer to human CD34+ cells.
BACKGROUND
Retrovirus-mediated gene transfer is a useful technology in studying the biology of hematopoietic stem cells (HSCs) as well as in developing gene therapy products for a variety of human diseases. One of the most important factors determining the success of these studies is the number of HSCs receiving the gene of interest.


METHODS
We tested various parameters for their influences on gene transfer efficiency to CD34+ cells derived from bone marrow. Based on a literature survey, three medium formulations of CD34+ cells have been compared for their effects on gene delivery efficiency and differentiation of them. We also tested whether FBS, used in the medium formulation, could be replaced with human serum or synthetic material.


RESULTS
Formulation A, consisting of stem cell factor, Flt-3 ligand, thrombopoietin, and IL-3, provided optimum results in that it maintained the highest percentage of CD34+ cells during the culture as well as produced the highest gene delivery efficiency. It was found that the synthetic serum substitute containing bovine serum albumin, insulin and human transferrin could replace the fetal bovine serum present in the original formulation A without compromising gene transfer efficiency. When the transduction procedure was repeated three times, the gene could be delivered in up to 60% of the cell population. Gene delivery efficiency was comparable between CD34+ cells derived from bone marrow and mobilized peripheral blood.


CONCLUSIONS
Our data could be useful in designing a procedure for stem cell gene therapy and providing a basis for further improving the conditions for gene transfer to various HSCs.
Hong
Youngtae
Y
Institute of Molecular Biology and Genetics, Seoul National University, Kwan-Ak-Gu, Seoul 151-742, South Korea.
Lee
Karim
K
Yu
Seung Shin
SS
Kim
Sujeong
S
Kim
Joong-Gon
JG
Shin
Hee Young
HY
Kim
Sunyoung
S
D018952:Antigens, CD34 / Q000378:metabolism*; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D009000:Monocytes / Q000276:immunology / Q000378:metabolism*; D012190:Retroviridae / Q000235:genetics*
D018952:Antigens, CD34; D017931:DNA Primers