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Gene transfer into human bone marrow hematopoietic cells mediated by
adenovirus vectors [see comments]
T Watanabe, C Kuszynski, K Ino, DG Heimann, HM Shepard, Y Yasui, DC Maneval and JE Talmadge
Department of Pathology, University of Nebraska Medical Center, NE
68198-5660.
Human bone marrow mononuclear cells (BMMNCs) and enriched CD34 positive
(CD34+) cells were transduced with adenovirus vectors encoding Escherichia
coli beta-galactosidase gene. Tranductions were carried out by 24-hour
coincubation with adenovirus vectors at different multiplicities of
infections (moi). Efficacy of gene transfer into BM cells and expression of
the gene product (ie, beta-galactosidase) were studied using X-Gal
histochemical staining and flow cytometric analysis. X-Gal staining
demonstrated that the percentage of positive cells at mois of 5 to 500 was
3.4% to 34.5% for BMMNCs and 6.0% to 20.0% for enriched CD34+ cells.
Similar results (1.5% to 35.7% for BMMNCs and 5.4% to 24.2% for enriched
CD34+ cells) were obtained with flow cytometric analysis using fluorescein
di-beta-D-galactopyranoside (FDG). Multicolor flow cytometry analysis,
which included FDG, demonstrated that BM progenitors (CD34+ or CD34+CD38-),
T cells (CD2+), B cells (CD19+), natural killer cells (CD56+),
granulocytes, and monocytes all expressed the adenovirus transgene. To
ascertain the effects of adenovirus vectors on normal BM progenitors, the
numbers of colony forming unit-granulocyte/macrophage (CFU-GM),
burst-forming unit- erythrocyte (BFU-E), and high-proliferative
potential-colony-forming cells (HPP-CFC) after 24-hour coincubation with
adenovirus vectors were determined. When BMMNCs or enriched CD34+ cells
were incubated with adenovirus vectors at mois of 5 and 50, no significant
differences in the numbers of CFU-GM, BFU-E, and HPP-CFC were observed
compared with the uninfected control cells. However, the numbers of CFU-GM
were significantly (P < .01) decreased when BMMNCs or enriched CD34+
cells were incubated with adenovirus vectors at a moi of 500, compared with
the uninfected control cells. The adenovirus infected cells, purified by
cell sorting for FDG expression, were capable of growing in culture and
gave rise to various colonies (ie, CFU-GM, BFU-E, and HPP-CFC). These data
indicate that recombinant adenovirus vectors can be used to transfer genes
to human BM hematopoietic cells with expression of the exogenous gene at a
high transduction efficiency.
Volume 87,
Issue 12,
pp. 5032-5039,
06/15/1996
Copyright © 1996 by The American Society of Hematology
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