A novel adenoviral vector-mediated neuronal selective gene expression in neonatal mouse brain in response to hypoxia.
Selective gene expression targeting neurons is a challenge, which, if successfully overcome, carries an enormous potential for clinical applications in therapeutics against neurodegenerative diseases. We have reported previously the construction of a series of adenoviral vectors capable of selectively expressing a reporter gene luciferase in cultured neurons [D. Huang, A. Desbois, S.T. Hou, A novel adenoviral vector which mediates hypoxia-inducible gene expression selectively in neurons, Gene Ther. 12 (2005) 1369-1376]. A combination of neuron restrictive silencer elements (NRSEs), hypoxia responsive elements (HREs) and CMV minimal promoter (CMVmp) was packaged into replication defective adenovirus to target gene expression selectively in neurons in a hypoxia-regulated manner. In the present study, we injected the adenoviral vectors into the neonatal mouse brain followed by treatment with hypoxia. The expression of the reporter luciferase gene was examined by luciferase assay and fluorescent immunostaining. Neurons and glial cells were identified by staining with antibodies against NeuN and GFAP, respectively. Remarkably, in response to hypoxia, Ad/5HRE-3NRSE showed strong hypoxia-inducible gene expression of the reporter luciferase selectively in neurons but not in glial cells. In contrast, brains infected with the control vector Ad/5HRE showed no selectivity in luciferase expression (in both neurons and glial cells) under the hypoxic condition. Taken together, these studies demonstrated that this vector (Ad/5HRE-3NRSE) can mediate gene expression selectively in neurons both in vitro and in vivo, supporting the suggestion that further refinement of this vector may lead to the development of a useful tool to investigate mechanisms of neuronal damage following cerebral ischemia and a possible effective gene therapy vector to stroke.
Hou
Sheng T
ST
Experimental Neurotherapeutics Laboratory, Institute for Biological Sciences, National Research Council Canada, 1200 Montreal Road, Bldg M54, Ottawa, Ontario, Canada K1A 0R6. sheng.hou@nrc-cnrc.gc.ca
Jiang
Susan X
SX
Huang
Deqi
D
Desbois
Angele
A
D000818:Animals; D000831:Animals, Newborn; D001921:Brain / Q000378:metabolism*; D002461:Cell Line, Transformed; D003587:Cytomegalovirus / Q000502:physiology*; D015870:Gene Expression / Q000502:physiology*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000235:genetics; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D006801:Humans; D000860:Hypoxia / Q000473:pathology* / Q000503:physiopathology; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009457:Neuroglia / Q000378:metabolism; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D000099229:RE1-Silencing Transcription Factor
D005904:Glial Fibrillary Acidic Protein; D008156:Luciferases; D010751:Phosphopyruvate Hydratase; D012097:Repressor Proteins; D014157:Transcription Factors; D000099229:RE1-Silencing Transcription Factor