NF-Ya protein delivery as a tool for hematopoietic progenitor cell expansion.
The clinical potential of therapeutic quantities of primary hematopoietic cells, either unmodified or altered via genetic modification, has stimulated the search for techniques that allow the production of large numbers of hematopoietic precursors, more primitive progenitors, and perhaps hematopoietic stem cells (HSC) themselves. Modifications of in vitro culture conditions to promote progenitor cell expansion have included combinations of polypeptide cytokines, small molecules, and transcription factors. Here we describe the methods for use of the transcription factor linked to a TAT-based protein transcription domain, in combination with cytokines and serum-free culture condition to stimulate the proliferation of primary cells. Human peripheral blood (PB) CD34(+) cells treated with TAT-NF-Ya fusion protein and grown in vitro for 1 month proliferate four times more than did cells in cultures that contained only cytokines, including increased production of hematopoietic cells of all maturities. These results and techniques should be suitable for multiple applications of ex vivo generation of hematopoietic cells using protein transduction.
Haverford College, Haverford, PA, USA.
Alevtina D
AD
Domashenko
Susan
S
Wiener
Stephen G
SG
Emerson
D000595:Amino Acid Sequence; D015153:Blotting, Western; D023081:CCAAT-Binding Factor / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology*; D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation / Q000187:drug effects; D003114:Colony-Forming Units Assay; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D020014:K562 Cells; D055550:Protein Stability; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology
D023081:CCAAT-Binding Factor; C492061:NFYA protein, human; D011993:Recombinant Fusion Proteins