Efficient Retroviral-Mediated Gene Transfer to Human Cord Blood Stem Cells With In Vivo Repopulating Potential

摘要: Recent studies have shown efficient gene transfer to primitive progenitors in human cord blood (CB) when the cells are incubated retrovirus-containing supernatants on fibronectin-coated dishes. We now used this approach achieve CB with capacity regenerate lymphoid and myeloid progeny nonobese diabetic (NOD)/severe combined immunodeficiency (SCID) mice. CD34(+) cell-enriched populations were first cultured for 3 days serum-free medium containing interleukin-3 (IL-3), IL-6, granulocyte colony-stimulating factor, Flt3-ligand, Steel factor followed by two 24-hour incubations a MSCV-NEO virus-containing obtained under either or serum-replete conditions. The presence of serum during latter 2 made no consistent difference total number cells, colony-forming (CFC), long-term culture-initiating (LTC-IC) recovered at end 5-day culture period, infected condition regenerated similar numbers (myeloid) CFC CD19(+) (B lymphoid) up 20 weeks NOD/SCID recipients. However, increased viral titer producer cell-conditioned was correlated twofold threefold higher efficiency all progenitor types. With supernatant, 17% +/- 3% 8%, G418-resistant vivo repopulating LTC-IC obtained. As expected, proportion NEO + determined polymerase chain reaction analysis generated even (32% 10%). There correlation between frequency unit-granulocyte-macrophage (CFU-GM), CFU-GM (r2 = 0.16 0.17, respectively), whereas values highly significantly 0.85). These findings provide further evidence close relationship (assessed using >/= 6-week output endpoint) indicate predictive value measurements such design clinical therapy protocols.