Title:Comparison of Tetracycline-regulated Promoters in Lentiviral-based Vectors in Murine Transplantation Studies
Volume: 16
Issue: 4
Author(s): Maike Stahlhut, Teng-Cheong Ha, Michael Morgan, Axel Schambach and Olga S. Kustikova
Affiliation:
Keywords:
Tetracycline-regulated promoters, Lentiviral, Hematopoietic, Murine transplantation model, Gene transfer, Doxycycline.
Abstract: Tetracycline-regulated systems with efficient temporal and dose regulation of transgene expression
are useful for development of new physiologic/pathophysiologic experimental models and
gene therapy approaches. Lentiviral vectors with improved tetracycline-regulated promoters help to
overcome the existing limitations such as basal activity in the drug absence, poor inducibility or
unstable transgene expression. To compare conventional and improved tetracycline-regulated promoters
in lentiviral based vectors in vivo, we investigated doxycycline-regulated gene transfer/expression levels
in a long-term murine transplantation model and demonstrated that the lentiviral vector with the
improved T11 promoter exhibited more efficient inducibility and higher gene transfer level. The time
required to reverse transgene expression after doxycycline removal was increased for animals with
higher gene expression levels and vector copy numbers. Examination of peripheral blood leukocytes
and splenocytes revealed similar cell lineage distributions for transgene positive and negative cell
populations from experimental and control mice, but increased variability in the percentages of myeloid
and lymphoid cells was detected in transgene positive bone marrow cells. However, no indication of
lineage bias in total bone marrow cells and no signs of hematopoietic disease were observed seven
months after transplantation. Our results showed that the T11 tetracycline-regulated promoter enabled
improved transgene expression in a murine transplantation model. The established system allows further
development of tetracycline-regulated experimental models to investigate normal and malignant
hematopoiesis.
