Quiescent Subpopulations of Human CD34-Positive Hematopoietic Stem Cells Are Preferred Targets for Stable Recombinant Adeno-Associated Virus Type 2 Transduction
| Autor: | Helicia Paz, Christie A. Wong, Wei Li, Leah Santat, K.K. Wong, Saswati Chatterjee |
|---|---|
| Rok vydání: | 2007 |
| Předmět: |
Stromal cell
viruses Genetic Vectors Population CD34 Gene Expression Antigens CD34 Bone Marrow Cells CD38 Biology Resting Phase Cell Cycle Transduction (genetics) Transduction Genetic Genetics Humans Transgenes Progenitor cell education Molecular Biology Cells Cultured Cell Proliferation education.field_of_study Cell Cycle Genetic Therapy Dependovirus Hematopoietic Stem Cells Virology Recombinant Proteins Cell biology Haematopoiesis Molecular Medicine Stem cell |
| Zdroj: | Human Gene Therapy. 18:614-626 |
| ISSN: | 1557-7422 1043-0342 |
| DOI: | 10.1089/hum.2006.188 |
| Popis: | We have previously demonstrated recombinant adeno-associated viral (rAAV) transduction of human CD34(+) hematopoietic stem cells (HSCs) capable of serial engraftment in vivo. Here we evaluated the capacity of rAAV2 to mediate gene transfer into nondividing, quiescent, primitive CD34(+) cells subdivided on the basis of metabolic, mitotic, and phenotypic properties. Results revealed that CD34(+)CD38() marrow cells are the most quiescent, exist primarily in G(0) at isolation and are the only population to remain nondividing during the entire exposure to free rAAV. Despite significant differences in the extended clonogenic capacities of CD34(+) subsets in stromal cultures, the frequency of rAAV marking of colonies derived from primitive progenitors was similar in all three populations, suggesting that both primitive and more differentiated progenitors were initially transduced at equal levels. After transduction, episomal and integrated rAAV genomes were detected in all CD34(+) subsets. However, the more quiescent cells displayed higher levels of integrated rAAV than did rapidly dividing cells. Importantly, stable long-term integration was observed only in the most primitive, quiescent CD34(+)CD38(-) subset, indicating that this HSC compartment comprises the preferred substrate for stable rAAV2 transduction. Previously described rate limitations to transgene expression were observed in transduced CD34(+) cells and could be overcome by tyrphostin pretreatment, which resulted in augmented second-strand synthesis. These results represent the first demonstration of rAAV-mediated gene transfer to primitive, quiescent human CD34(+)CD38(-) stem cells and reveal that nondividing CD34(+)CD38(-) HSCs are the optimal CD34(+) targets for rAAV transduction. |
| Databáze: | OpenAIRE |
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