In situ entry of oligonucleotides into brain cells can occur through a nucleic acid channel.
Brain tissue has become a challenging therapeutic target, in part because of failure of conventional treatments of brain tumors and a gradually increasing number of neurodegenerative diseases. Because antisense oligonucleotides are readily internalized by neuronal cells in culture, these compounds could possibly serve as novel therapeutic agents to meet such a challenge. In previous in vitro work using cell culture systems, we have demonstrated that intracellular delivery requires a vector such as cationic liposomes since free oligonucleotides remain largely trapped in the endocytic pathway following cellular uptake. Here we studied the cellular uptake properties of oligonucleotides by explants of rat brain (brain slices), and by in vivo brain tissue after administration of oligonucleotides by bolus injection. In contrast to in vitro uptake, we show that in brain slices oligonucleotides were taken up by neuronal and nonneuronal cells, irrespective of their assembly with cationic liposomes. In either case, a diffuse distribution of oligonucleotides was seen in the cytosol and/or nucleus. Uptake of oligonucleotides by brain slices as a result of membrane damage, potentially arising from the isolation procedure, could be excluded. Interestingly, internalization was inhibited following treatment of the tissue with antibody GN-2640, directed against a nucleic acid channel, present in rat kidney cells. Our data support the view that an analogous channel is present in brain tissue, allowing entry of free oligonucleotides but not plasmids. Indeed, for delivery of the latter and accomplishment of effective transfection, cationic lipids were needed for gene translocation into both brain slices and brain tissue in vivo. These data imply that for antisense therapy to become effective in brain, cationic lipid-mediated delivery will only be needed for specific cell targeting but not necessarily for delivery per se to accomplish nuclear deposition of oligonucleotides into brain cells and subsequent down-regulation of disease-related targets.
Shi
Fuxin
F
Department of Membrane Cell Biology, Faculty of Medical Sciences, University of Groningen, The Netherlands.
Gounko
Natasha V
NV
Wang
Xiaoqin
X
Ronken
Eric
E
Hoekstra
Dick
D
D021581:Active Transport, Cell Nucleus; D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D001921:Brain / Q000378:metabolism*; D002412:Cations / Q000737:chemistry; D002467:Cell Nucleus / Q000378:metabolism*; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008055:Lipids / Q000737:chemistry; D009696:Nucleic Acids / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009841:Oligonucleotides / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014018:Tissue Distribution
D000906:Antibodies; D002412:Cations; D008055:Lipids; D009696:Nucleic Acids; D009841:Oligonucleotides