Whole animal in vivo imaging after transient, nonviral gene delivery to the rat central nervous system.
We previously showed that a vector:lipid delivery system, comprised of a plasmid DNA vector and cationic lipid (lipoplex), when injected into the cerebrospinal fluid (CSF) of rats can deliver reporter genes in vivo efficiently and with widespread expression to the Central Nervous System (CNS). To further characterize this delivery system, we now present experiments that demonstrate the in vivo time-to-peak expression of the reporter gene, firefly luciferase. We infused a formulated lipoplex containing the lipid MLRI [dissymmetric myristoyl (14:0) and lauroyl (12:1) rosenthal inhibitor-substituted compound formed from the tetraalkylammonium glycerol-based DORI] and pNDluc, a luciferase vector, into CSF in the cisterna magna (CM) of the rat. Luciferase activity was followed over time by bioluminescence imaging after injection of luciferin. Our results show that luciferase activity in the CNS of rats is widespread, peaks 72 hours after injection into CM and can be detected in vivo for at least 7-10 days after peak expression. We further show that in contrast to injection into CSF, enzyme activity is not widely distributed after injection of the vector into brain parenchyma, emphasizing the importance of CSF delivery to achieve widespread vector distribution. Finally, we confirm the distribution of firefly luciferase in brain by immunohistochemical staining from an animal that was euthanized at the peak of enzyme expression.
Hauck
Ellen S
ES
Department of Anesthesia, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6112, USA.
Zou
Shaomin
S
Scarfo
Keith
K
Nantz
Michael H
MH
Hecker
James G
JG
D000818:Animals; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D002490:Central Nervous System / Q000378:metabolism*; D002946:Cisterna Magna; D004337:Drug Carriers; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D008055:Lipids; D049409:Luciferases, Firefly / Q000096:biosynthesis / Q000134:cerebrospinal fluid / Q000235:genetics; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley
D004337:Drug Carriers; D008055:Lipids; D049409:Luciferases, Firefly