Efficient expansion and gene transduction of mouse neural stem/progenitor cells on recombinant fibronectin.
Neural stem/progenitor cells (NSCs) are commonly grown as floating neurospheres in medium containing basic fibroblast growth factor and epidermal growth factor. Under these conditions, about 1% of the cells retain multipotentiality. We developed a protocol based on culture of NSCs in adherence on recombinant fibronectin (rFN) to transduce up to 90% NSCs at a multiplicity of infection of 2 with no need for viral concentration or production of serum-free retroviral supernatants. NSCs grew faster on rFN than as neurospheres on tissue culture plastic and did not lose their stem cell nature or multipotentiality. Furthermore, retroviral-mediated transgene expression was sustained with time in culture and upon differentiation into neurons and astrocytes. These experimental conditions may be utilized to study the function of various genes in NSCs, and to manipulate NSCs for gene and cell therapy of several neurological diseases.
Rappa
G
G
Department of Tumor Biology, Norwegian Radium Hospital, Montebello, Oslo 0310, Norway.
Kunke
D
D
Holter
J
J
Diep
D B
DB
Meyer
J
J
Baum
C
C
Fodstad
O
O
Krauss
S
S
Lorico
A
A
D000818:Animals; D000831:Animals, Newborn; D001921:Brain / Q000166:cytology; D002448:Cell Adhesion; D002454:Cell Differentiation; D002478:Cells, Cultured; D005353:Fibronectins; D005822:Genetic Vectors; D051379:Mice; D008810:Mice, Inbred C57BL; D009474:Neurons / Q000166:cytology* / Q000502:physiology*; D020411:Oligonucleotide Array Sequence Analysis; D011994:Recombinant Proteins; D012190:Retroviridae / Q000235:genetics; D012867:Skin / Q000166:cytology; D013234:Stem Cells / Q000166:cytology* / Q000502:physiology*; D013997:Time Factors; D014161:Transduction, Genetic
D005353:Fibronectins; D011994:Recombinant Proteins