Pure populations of transduced primary human cells can be produced using GFP expressing herpes virus vectors and flow cytometry.
Herpes simplex virus (HSV) has often been suggested as a vector for gene delivery to the nervous system although it is also capable of infecting many other cell types. HSV also has the ability to package large genetic insertions so the expression of multiple genes from a single virus is possible. Here we show that a green fluorescent protein (GFP) expressing HSV1 vector can transduce two primary human cell types--quiescent human CD34+ hematopoietic progenitor cells and dendritic cells--which are both hard to transduce by other means. We also show that GFP is an effective marker when expressed from an HSV vector in vivo in the mouse brain. When GFP is expressed together with a second gene (in this case lacZ) from a single virus, transduced GFP-positive CD34+ hematopoietic progenitor cells or dendritic cells can both be generated at an effective efficiency of 100% for the second gene. Here transduction with the vector is combined with flow cytometry allowing GFP-positive cells to be sorted from the untransduced population. Such completely transduced populations of quiescent CD34+ hematopoietic progenitor and dendritic cells cannot easily be achieved by other means, and might thus allow experimental or therapeutic protocols to be carried out requiring high-level transduction which would not otherwise be possible. Such an approach using HSV vectors might also be applicable to other cell types for which transduction is as yet unreliable or of low efficiency.
Coffin
R S
RS
Department of Molecular Pathology, University College London Medical School, UK.
Thomas
S K
SK
Thomas
N S
NS
Lilley
C E
CE
Pizzey
A R
AR
Griffiths
C H
CH
Gibb
B J
BJ
Wagstaff
M J
MJ
Inges
S J
SJ
Binks
M H
MH
Chain
B M
BM
Thrasher
A J
AJ
Rutault
K
K
Latchman
D S
DS
D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D002469:Cell Separation; D002478:Cells, Cultured; D006224:Cricetinae; D003713:Dendritic Cells / Q000166:cytology / Q000378:metabolism; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D005819:Genetic Markers; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism; D006651:Histocytochemistry; D006801:Humans; D007202:Indicators and Reagents / Q000378:metabolism; D007763:Lac Operon / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D018139:Simplexvirus / Q000235:genetics*
D018952:Antigens, CD34; D005819:Genetic Markers; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins