Intracellular Distribution and Mechanism of Delivery of Oligonucleotides Mediated by Cationic Lipids
Identifieur interne : 002A38 ( Main/Exploration ); précédent : 002A37; suivant : 002A39Intracellular Distribution and Mechanism of Delivery of Oligonucleotides Mediated by Cationic Lipids
Auteurs : Olivier Zelphati [États-Unis] ; Francis C. Szoka Jr [États-Unis]Source :
- Pharmaceutical Research [ 0724-8741 ] ; 1996-09-01.
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Abstract
Abstract: Purpose. To study the parameters influencing the intracellular trafficking of oligonucleotides delivered by cationic 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) lipids and to elucidate the mechanism of uptake. Methods. We have studied the intracellular localization of fluorescently labeled oligonucleotide (F-ODN) delivered by DOTAP using confocal microscopy and measured inhibition of luciferase synthesis. The delivery mechanism of ODN/DOTAP complexes was investigated using inhibitors of the endocytosis pathway. Results. F-ODN delivered by DOTAP liposomes redistribute from punctate cytoplasmic regions into the nucleus. The nuclear uptake of F-ODN depends on: charge ratio (+/–), time of incubation, temperature and presence of serum. A positively charged complex is required for enhanced uptake. The association of neutral lipids with DOTAP reduced the optimum charge ratio without altering the delivery efficiency. DOTAP lipids increased >100 fold the antisense activity of a specific anti-luciferase ODN. Inhibitors of the endocytosis pathway show that the majority of F-ODN are introduced through an endocytic pathway mainly involving uncoated vesicles. Nuclear accumulation of oligonucleotides can be decreased by inhibitors of actin microfilaments, energy metabolism and proteins implicated in the fusion of endosomes. Nuclear uptake is independent of acidification of the endosomal vesicles and unaffected by inhibitors of microtubules. Conclusions. Oligonucleotides are delivered by cationic lipids into the cytoplasm at an early stage of the endocytotic pathway which leads to a marked increase in antisense activity and oligonucleotide nuclear uptake.
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DOI: 10.1023/A:1016026101195
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Szoka Jr</name></author></titleStmt><publicationStmt><idno type="wicri:source">ISTEX</idno><idno type="RBID">ISTEX:F5BF4FE47CFEE33AE2495DBCCDC6097FA777667B</idno><date when="1996" year="1996">1996</date><idno type="doi">10.1023/A:1016026101195</idno><idno type="url">https://api.istex.fr/ark:/67375/1BB-QLG4XQVP-V/fulltext.pdf</idno><idno type="wicri:Area/Istex/Corpus">001162</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Corpus" wicri:corpus="ISTEX">001162</idno><idno type="wicri:Area/Istex/Curation">001162</idno><idno type="wicri:Area/Istex/Checkpoint">001828</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Checkpoint">001828</idno><idno type="wicri:doubleKey">0724-8741:1996:Zelphati O:intracellular:distribution:and</idno><idno type="wicri:Area/Main/Merge">002A90</idno><idno type="wicri:Area/Main/Curation">002A38</idno><idno type="wicri:Area/Main/Exploration">002A38</idno></publicationStmt><sourceDesc><biblStruct><analytic><title level="a" type="main" xml:lang="en">Intracellular Distribution and Mechanism of Delivery of Oligonucleotides Mediated by Cationic Lipids</title><author><name sortKey="Zelphati, Olivier" sort="Zelphati, Olivier" uniqKey="Zelphati O" first="Olivier" last="Zelphati">Olivier Zelphati</name><affiliation wicri:level="2"><country>États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Department of Biopharmaceutical Sciences and Pharmaceutical Chemistry, University of California, School of Pharmacy, 94143–04461, San Francisco</wicri:cityArea></affiliation></author><author><name sortKey="Szoka Jr, Francis C" sort="Szoka Jr, Francis C" uniqKey="Szoka Jr F" first="Francis C." last="Szoka Jr">Francis C. Szoka Jr</name><affiliation wicri:level="2"><country>États-Unis</country><placeName><region type="state">Californie</region></placeName><wicri:cityArea>Department of Biopharmaceutical Sciences and Pharmaceutical Chemistry, University of California, School of Pharmacy, 94143–04461, San Francisco</wicri:cityArea></affiliation></author></analytic><monogr></monogr><series><title level="j">Pharmaceutical Research</title><title level="j" type="sub">An Official Journal of the American Association of Pharmaceutical Scientists</title><title level="j" type="abbrev">Pharm Res</title><idno type="ISSN">0724-8741</idno><idno type="eISSN">1573-904X</idno><imprint><publisher>Kluwer Academic Publishers-Plenum Publishers</publisher><pubPlace>New York</pubPlace><date type="published" when="1996-09-01">1996-09-01</date><biblScope unit="volume">13</biblScope><biblScope unit="issue">9</biblScope><biblScope unit="page" from="1367">1367</biblScope><biblScope unit="page" to="1372">1372</biblScope></imprint><idno type="ISSN">0724-8741</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0724-8741</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>cationic liposomes</term><term>drug delivery</term><term>gene therapy</term><term>intracellular distribution</term><term>oligonucleotides</term></keywords></textClass><langUsage><language ident="en">en</language></langUsage></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Abstract: Purpose. To study the parameters influencing the intracellular trafficking of oligonucleotides delivered by cationic 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) lipids and to elucidate the mechanism of uptake. Methods. We have studied the intracellular localization of fluorescently labeled oligonucleotide (F-ODN) delivered by DOTAP using confocal microscopy and measured inhibition of luciferase synthesis. The delivery mechanism of ODN/DOTAP complexes was investigated using inhibitors of the endocytosis pathway. Results. F-ODN delivered by DOTAP liposomes redistribute from punctate cytoplasmic regions into the nucleus. The nuclear uptake of F-ODN depends on: charge ratio (+/–), time of incubation, temperature and presence of serum. A positively charged complex is required for enhanced uptake. The association of neutral lipids with DOTAP reduced the optimum charge ratio without altering the delivery efficiency. DOTAP lipids increased >100 fold the antisense activity of a specific anti-luciferase ODN. Inhibitors of the endocytosis pathway show that the majority of F-ODN are introduced through an endocytic pathway mainly involving uncoated vesicles. Nuclear accumulation of oligonucleotides can be decreased by inhibitors of actin microfilaments, energy metabolism and proteins implicated in the fusion of endosomes. Nuclear uptake is independent of acidification of the endosomal vesicles and unaffected by inhibitors of microtubules. Conclusions. Oligonucleotides are delivered by cationic lipids into the cytoplasm at an early stage of the endocytotic pathway which leads to a marked increase in antisense activity and oligonucleotide nuclear uptake.</div></front></TEI><affiliations><list><country><li>États-Unis</li></country><region><li>Californie</li></region></list><tree><country name="États-Unis"><region name="Californie"><name sortKey="Zelphati, Olivier" sort="Zelphati, Olivier" uniqKey="Zelphati O" first="Olivier" last="Zelphati">Olivier Zelphati</name></region><name sortKey="Szoka Jr, Francis C" sort="Szoka Jr, Francis C" uniqKey="Szoka Jr F" first="Francis C." last="Szoka Jr">Francis C. Szoka Jr</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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