Transgene expression in the brain stem effected by intramuscular injection of polyethylenimine/DNA complexes.
Gene delivery into the CNS without tissue destruction is challenging. As neurons are capable of taking up exogenous particulates from the muscles that they innervate, we investigated the feasibility of achieving gene transfer in CNS neurons by peripheral intramuscular injection of plasmid DNA complexed with the cationic polymer polyethylenimine (PEI) in the rat hypoglossal system. Using the luciferase reporter gene driven by a Rous sarcoma virus promoter, transgene expression of up to 4 x 10(6) RLU per brain stem at 20 microg of plasmid DNA was achieved after tongue injection. Using lacZ as a reporter gene, transgene expression in the brain stem was detected in hypoglossal motor neurons, a group of neurons that innervate tongue muscles. The plasmid DNA was detected by PCR analysis in the brain-stem samples, demonstrating that the PEI/DNA complexes had migrated by retrograde axonal transport to neuronal cell bodies in the brain stem after being internalized by nerve terminals in the tongue muscle. Using a therapeutic bcl-2 gene driven by a cytomegalovirus promoter and Western blotting, transgene expression was detectable in the brain stem as early as 18 h after tongue injection and lasted for at least 2 weeks. Two lipid transfection agents, GenePORTER and TransFast, mediated a weak gene expression in the hypoglossal system, but not two polymers, poly-l-lysine and chitosan. The nonviral neuronal gene delivery method established in this study bypasses the blood-brain barrier and suggests a possible therapeutic strategy for noninvasive CNS gene transfer.
Wang
S
S
Biomaterials/Tissue Engineering Program, Institute of Materials Research & Engineering, National University of Singapore, 3 Research Link, Singapore 117602. shu-wang@imre.org.sg
Ma
N
N
Gao
S J
SJ
Yu
H
H
Leong
K W
KW
D000818:Animals; D001812:Blood-Brain Barrier; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism*; D001933:Brain Stem / Q000378:metabolism*; D002490:Central Nervous System / Q000378:metabolism; D003587:Cytomegalovirus / Q000235:genetics; D004247:DNA / Q000378:metabolism*; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors; D007763:Lac Operon; D050356:Lipid Metabolism; D008156:Luciferases / Q000378:metabolism; D008297:Male; D009474:Neurons / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011094:Polyethyleneimine / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors; D014162:Transfection; D019076:Transgenes; D001616:beta-Galactosidase / Q000378:metabolism
D019253:Proto-Oncogene Proteins c-bcl-2; D011094:Polyethyleneimine; D004247:DNA; D008156:Luciferases; D001616:beta-Galactosidase