title: A minimally invasive, lentiviral based method for the rapid and sustained genetic manipulation of renal tubules.
abstract: The accelerated discovery of disease-related genes emerging from genomic studies has strained the capacity of traditional genetically engineered mouse models (GEMMs) to provide in-vivo validation. Direct, somatic, genetic engineering approaches allow for accelerated and flexible genetic manipulation and represent an attractive alternative to GEMMs. In this study we investigated the feasibility, safety and efficiency of a minimally invasive, lentiviral based approach for the sustained in-vivo modification of renal tubular epithelial cells. Using ultrasound guidance, reporter vectors were directly injected into the mouse renal parenchyma. We observed transgene expression confined to the renal cortex (specifically proximal and distal tubules) and sustained beyond 2 months post injection. Furthermore, we demonstrate the ability of this methodology to induce long-term, in-vivo knockdown of candidate genes either through somatic recombination of floxed alleles or by direct delivery of specific shRNA sequences. This study demonstrates that ultrasound-guided injection of lentiviral vectors provides a safe and efficient method for the genetic manipulation of renal tubules, representing a quick and versatile alternative to GEMMs for the functional characterisation of disease-related genes.
authors.affiliation: Department of Oncology, University of Cambridge, CRUK Cambridge institute, Cambridge, CB2 0RE, UK.
authors.forename: Judit
authors.initials: J
authors.lastname: Espana-Agusti
authors.affiliation: 1] Department of Oncology, University of Cambridge, CRUK Cambridge institute, Cambridge, CB2 0RE, UK [2] Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Tuveson
authors.affiliation: Experimental Cancer Genetics, Wellcome Trust Sanger Institute, Hinxton, CB10 1SA, UK.
authors.forename: David J
authors.initials: DJ
authors.lastname: Adams
authors.affiliation: Department of Oncology, University of Cambridge, CRUK Cambridge institute, Cambridge, CB2 0RE, UK.
authors.forename: Athena
authors.initials: A
authors.lastname: Matakidou
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007684:Kidney Tubules / Q000000981:diagnostic imaging / Q000378:metabolism*; D016086:Lentivirus / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D041681:NIH 3T3 Cells; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D000077004:Tuberous Sclerosis Complex 1 Protein; D025521:Tumor Suppressor Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D014463:Ultrasonography
chemical_list: D034741:RNA, Small Interfering; D000077004:Tuberous Sclerosis Complex 1 Protein; D025521:Tumor Suppressor Proteins