title: Comparison of Tetracycline-regulated Promoters in Lentiviral-based Vectors in Murine Transplantation Studies.
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.
authors.forename: Maike
authors.initials: M
authors.lastname: Stahlhut
authors.forename: Teng-Cheong
authors.initials: TC
authors.lastname: Ha
authors.forename: Michael
authors.initials: M
authors.lastname: Morgan
authors.forename: Axel
authors.initials: A
authors.lastname: Schambach
authors.affiliation: Institute of Experimental Hematology, OE6960 Hannover Medical School, Carl-Neuberg-Straße 1 Hannover 30625, Germany.
authors.forename: Olga S
authors.initials: OS
authors.lastname: Kustikova
mesh_terms: D000818:Animals; D019070:Cell Lineage; D017690:Cell Transplantation / Q000379:methods*; D004318:Doxycycline / Q000008:administration & dosage; D005786:Gene Expression Regulation / Q000187:drug effects; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D013752:Tetracycline / Q000494:pharmacology*; D014161:Transduction, Genetic; D019076:Transgenes
chemical_list: C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D013752:Tetracycline; D004318:Doxycycline
keywords: Doxycycline; Gene transfer; Hematopoietic; Lentiviral; Murine transplantation model; Tetracycline-regulated promoters