Gene transfer into human bone marrow hematopoietic cells mediated by adenovirus vectors

Tsutomu Watanabe, Charles Kuszynski, Kazuhiko Ino, Dean G. Heimann, H. Michael Shephard, Yoshinori Yasui, Daniel C. Maneval, James E. Talmadge

Research output: Contribution to journalArticlepeer-review

60 Scopus citations

Abstract

Human bone marrow mononuclear cells (BMMNCs) and enriched CD34 positive (CD34+) cells were transduced with adenovirus vectors encoding Escherichia coli β-galactosidase gene. Transductions 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, β-galactosidase) were studied using X-Gel histochemical staining and flow cytometric analysis. X-Gal staining demonstrated that the percentage of positive cells at regis 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 fluorescent di-β- 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 regis 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.

Original languageEnglish (US)
Pages (from-to)5032-5039
Number of pages8
JournalBlood
Volume87
Issue number12
DOIs
StatePublished - Jun 15 1996

ASJC Scopus subject areas

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology

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