DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers
Identifieur interne : 002726 ( Istex/Corpus ); précédent : 002725; suivant : 002727DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers
Auteurs : Dipen S. Shah ; T. Sakthivel ; Istvan Toth ; Alexander T. Florence ; Andy F. WilderspinSource :
- International Journal of Pharmaceutics [ 0378-5173 ] ; 2000.
English descriptors
- KwdEn :
- DODAC, dioleyldimethylammonium chloride, DOGS, dioctadecylamidoglycylspermime, DOPE, dioleoyl phosphatidylethanolamine, DOSPA, 2,3-dioleyloxy-N-[2(Spermine-carboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoracrtate, DOTMA, 2,3-diokeyloxypropyl-1-trimethyl ammonium chloride, Dendrimers, G9-EDA, 9th generation PAMAN with ethylenediamine core, Gene delivery, Gene expression, Non-viral vectors, PAMAM, polyamidoamine, Transfection.
- Teeft :
- Agarose, Amphipathic, Aqueous solution, Brunswick square, Cationic, Cationic lipids, Cell viability, Charge ratio, Complex formation, Complex stability, Dendrimer, Dendrimer concentration, Dendrimers, Error bars show, Erythrocyte, Gene delivery, Gene expression, Gene transfer, Hydrophobic tail, International journal, Linear position, Lipid, Loading buffer, London wc1n, Mammalian cells, Minimal effect, Molar, Molar charge ratio, Molecular weight, Optimally transfected, Optimum charge ratio, Pamam, Pamam dendrimers, Percent survival, Percentage transfection, Pharmaceutics, Phosphate saline buffer, Plasmid, Psvbgal, Sakthivel, Shah, Stock solution, Supercoiled position, Terminal amines, Toth, Transfected, Transfection.
Abstract
Abstract: Amphipathic asymmetric dendrimers have been investigated for use in delivery of genes into cells, with the objective of optimising transfection efficiency and maintaining cell viability. We have synthesised amphipathic asymmetric dendrimers by solid phase methods. The ability of two of these to transfect BHK cells in culture with β-galactosidase gene was determined by X-gal staining. Cell viability was measured by the MTT assay for BHK cells, and by spectroscopy for lysis of erythrocytes. Interactions between dendrimer and DNA were investigated by agarose gel electrophoresis. BHK cells were optimally transfected at 5:1 +/− charge ratio yielding 20% cells receiving at least one copy of the plasmid. Cell viability decreased when the dendrimer to DNA ratio exceeded 5:1. Raising the pH significantly affected the electrophoretic mobility of complexes of dendrimer and DNA. We conclude that amphipathic asymmetric dendrimers enable efficient plasmid DNA uptake into BHK cells. Cell viability is maintained at high concentrations of dendrimer when complexed with DNA at a 5:1 +/− charge ratio. Efficiency of transfection and cell viability suggest the system may be suitable for gene delivery in vivo.
Url:
DOI: 10.1016/S0378-5173(00)00534-2
Links to Exploration step
ISTEX:CD94E85ACB7135922B0A5CDE0F4AA6EF02D6BE35Le document en format XML
<record><TEI wicri:istexFullTextTei="biblStruct"><teiHeader><fileDesc><titleStmt><title xml:lang="en">DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers</title><author><name sortKey="Shah, Dipen S" sort="Shah, Dipen S" uniqKey="Shah D" first="Dipen S" last="Shah">Dipen S. Shah</name><affiliation><mods:affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation></author><author><name sortKey="Sakthivel, T" sort="Sakthivel, T" uniqKey="Sakthivel T" first="T" last="Sakthivel">T. Sakthivel</name><affiliation><mods:affiliation>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation></author><author><name sortKey="Toth, Istvan" sort="Toth, Istvan" uniqKey="Toth I" first="Istvan" last="Toth">Istvan Toth</name><affiliation><mods:affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation><affiliation><mods:affiliation>1 Present address: School of Pharmacy, Steele Building, University of Queensland, Brisbane, Qld 4072.</mods:affiliation></affiliation></author><author><name sortKey="Florence, Alexander T" sort="Florence, Alexander T" uniqKey="Florence A" first="Alexander T" last="Florence">Alexander T. Florence</name><affiliation><mods:affiliation>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation></author><author><name sortKey="Wilderspin, Andy F" sort="Wilderspin, Andy F" uniqKey="Wilderspin A" first="Andy F" last="Wilderspin">Andy F. Wilderspin</name><affiliation><mods:affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation><affiliation><mods:affiliation>Corresponding author. Tel.: +44-207-7535878; fax: +44-207-7531939</mods:affiliation></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">ISTEX</idno><idno type="RBID">ISTEX:CD94E85ACB7135922B0A5CDE0F4AA6EF02D6BE35</idno><date when="2000" year="2000">2000</date><idno type="doi">10.1016/S0378-5173(00)00534-2</idno><idno type="url">https://api.istex.fr/ark:/67375/6H6-G5R0LGHP-C/fulltext.pdf</idno><idno type="wicri:Area/Istex/Corpus">002726</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Corpus" wicri:corpus="ISTEX">002726</idno></publicationStmt><sourceDesc><biblStruct><analytic><title level="a" type="main" xml:lang="en">DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers</title><author><name sortKey="Shah, Dipen S" sort="Shah, Dipen S" uniqKey="Shah D" first="Dipen S" last="Shah">Dipen S. Shah</name><affiliation><mods:affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation></author><author><name sortKey="Sakthivel, T" sort="Sakthivel, T" uniqKey="Sakthivel T" first="T" last="Sakthivel">T. Sakthivel</name><affiliation><mods:affiliation>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation></author><author><name sortKey="Toth, Istvan" sort="Toth, Istvan" uniqKey="Toth I" first="Istvan" last="Toth">Istvan Toth</name><affiliation><mods:affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation><affiliation><mods:affiliation>1 Present address: School of Pharmacy, Steele Building, University of Queensland, Brisbane, Qld 4072.</mods:affiliation></affiliation></author><author><name sortKey="Florence, Alexander T" sort="Florence, Alexander T" uniqKey="Florence A" first="Alexander T" last="Florence">Alexander T. Florence</name><affiliation><mods:affiliation>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation></author><author><name sortKey="Wilderspin, Andy F" sort="Wilderspin, Andy F" uniqKey="Wilderspin A" first="Andy F" last="Wilderspin">Andy F. Wilderspin</name><affiliation><mods:affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</mods:affiliation></affiliation><affiliation><mods:affiliation>Corresponding author. Tel.: +44-207-7535878; fax: +44-207-7531939</mods:affiliation></affiliation></author></analytic><monogr></monogr><series><title level="j">International Journal of Pharmaceutics</title><title level="j" type="abbrev">IJP</title><idno type="ISSN">0378-5173</idno><imprint><publisher>ELSEVIER</publisher><date type="published" when="2000">2000</date><biblScope unit="volume">208</biblScope><biblScope unit="issue">1–2</biblScope><biblScope unit="page" from="41">41</biblScope><biblScope unit="page" to="48">48</biblScope></imprint><idno type="ISSN">0378-5173</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0378-5173</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>DODAC, dioleyldimethylammonium chloride</term><term>DOGS, dioctadecylamidoglycylspermime</term><term>DOPE, dioleoyl phosphatidylethanolamine</term><term>DOSPA, 2,3-dioleyloxy-N-[2(Spermine-carboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoracrtate</term><term>DOTMA, 2,3-diokeyloxypropyl-1-trimethyl ammonium chloride</term><term>Dendrimers</term><term>G9-EDA, 9th generation PAMAN with ethylenediamine core</term><term>Gene delivery</term><term>Gene expression</term><term>Non-viral vectors</term><term>PAMAM, polyamidoamine</term><term>Transfection</term></keywords><keywords scheme="Teeft" xml:lang="en"><term>Agarose</term><term>Amphipathic</term><term>Aqueous solution</term><term>Brunswick square</term><term>Cationic</term><term>Cationic lipids</term><term>Cell viability</term><term>Charge ratio</term><term>Complex formation</term><term>Complex stability</term><term>Dendrimer</term><term>Dendrimer concentration</term><term>Dendrimers</term><term>Error bars show</term><term>Erythrocyte</term><term>Gene delivery</term><term>Gene expression</term><term>Gene transfer</term><term>Hydrophobic tail</term><term>International journal</term><term>Linear position</term><term>Lipid</term><term>Loading buffer</term><term>London wc1n</term><term>Mammalian cells</term><term>Minimal effect</term><term>Molar</term><term>Molar charge ratio</term><term>Molecular weight</term><term>Optimally transfected</term><term>Optimum charge ratio</term><term>Pamam</term><term>Pamam dendrimers</term><term>Percent survival</term><term>Percentage transfection</term><term>Pharmaceutics</term><term>Phosphate saline buffer</term><term>Plasmid</term><term>Psvbgal</term><term>Sakthivel</term><term>Shah</term><term>Stock solution</term><term>Supercoiled position</term><term>Terminal amines</term><term>Toth</term><term>Transfected</term><term>Transfection</term></keywords></textClass><langUsage><language ident="en">en</language></langUsage></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Abstract: Amphipathic asymmetric dendrimers have been investigated for use in delivery of genes into cells, with the objective of optimising transfection efficiency and maintaining cell viability. We have synthesised amphipathic asymmetric dendrimers by solid phase methods. The ability of two of these to transfect BHK cells in culture with β-galactosidase gene was determined by X-gal staining. Cell viability was measured by the MTT assay for BHK cells, and by spectroscopy for lysis of erythrocytes. Interactions between dendrimer and DNA were investigated by agarose gel electrophoresis. BHK cells were optimally transfected at 5:1 +/− charge ratio yielding 20% cells receiving at least one copy of the plasmid. Cell viability decreased when the dendrimer to DNA ratio exceeded 5:1. Raising the pH significantly affected the electrophoretic mobility of complexes of dendrimer and DNA. We conclude that amphipathic asymmetric dendrimers enable efficient plasmid DNA uptake into BHK cells. Cell viability is maintained at high concentrations of dendrimer when complexed with DNA at a 5:1 +/− charge ratio. Efficiency of transfection and cell viability suggest the system may be suitable for gene delivery in vivo.</div></front></TEI><istex><corpusName>elsevier</corpusName><keywords><teeft><json:string>dendrimer</json:string><json:string>dendrimers</json:string><json:string>transfection</json:string><json:string>plasmid</json:string><json:string>cationic</json:string><json:string>amphipathic</json:string><json:string>agarose</json:string><json:string>pamam</json:string><json:string>international journal</json:string><json:string>psvbgal</json:string><json:string>pharmaceutics</json:string><json:string>lipid</json:string><json:string>toth</json:string><json:string>transfected</json:string><json:string>erythrocyte</json:string><json:string>sakthivel</json:string><json:string>cell viability</json:string><json:string>charge ratio</json:string><json:string>gene delivery</json:string><json:string>molar charge ratio</json:string><json:string>molar</json:string><json:string>pamam dendrimers</json:string><json:string>dendrimer concentration</json:string><json:string>terminal amines</json:string><json:string>molecular weight</json:string><json:string>error bars show</json:string><json:string>shah</json:string><json:string>optimally transfected</json:string><json:string>stock solution</json:string><json:string>mammalian cells</json:string><json:string>complex formation</json:string><json:string>brunswick square</json:string><json:string>london wc1n</json:string><json:string>complex stability</json:string><json:string>loading buffer</json:string><json:string>phosphate saline buffer</json:string><json:string>gene transfer</json:string><json:string>percentage transfection</json:string><json:string>gene expression</json:string><json:string>aqueous solution</json:string><json:string>linear position</json:string><json:string>supercoiled position</json:string><json:string>minimal effect</json:string><json:string>hydrophobic tail</json:string><json:string>percent survival</json:string><json:string>optimum charge ratio</json:string><json:string>cationic lipids</json:string></teeft></keywords><author><json:item><name>Dipen S Shah</name><affiliations><json:string>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</json:string></affiliations></json:item><json:item><name>T Sakthivel</name><affiliations><json:string>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</json:string></affiliations></json:item><json:item><name>Istvan Toth</name><affiliations><json:string>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</json:string><json:string>1 Present address: School of Pharmacy, Steele Building, University of Queensland, Brisbane, Qld 4072.</json:string></affiliations></json:item><json:item><name>Alexander T Florence</name><affiliations><json:string>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</json:string></affiliations></json:item><json:item><name>Andy F Wilderspin</name><affiliations><json:string>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</json:string><json:string>Corresponding author. Tel.: +44-207-7535878; fax: +44-207-7531939</json:string></affiliations></json:item></author><subject><json:item><lang><json:string>eng</json:string></lang><value>Gene delivery</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>Gene expression</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>Transfection</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>Dendrimers</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>Non-viral vectors</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>DOTMA, 2,3-diokeyloxypropyl-1-trimethyl ammonium chloride</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>DOSPA, 2,3-dioleyloxy-N-[2(Spermine-carboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoracrtate</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>DOGS, dioctadecylamidoglycylspermime</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>DOPE, dioleoyl phosphatidylethanolamine</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>DODAC, dioleyldimethylammonium chloride</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>PAMAM, polyamidoamine</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>G9-EDA, 9th generation PAMAN with ethylenediamine core</value></json:item></subject><arkIstex>ark:/67375/6H6-G5R0LGHP-C</arkIstex><language><json:string>eng</json:string></language><originalGenre><json:string>Full-length article</json:string></originalGenre><abstract>Abstract: Amphipathic asymmetric dendrimers have been investigated for use in delivery of genes into cells, with the objective of optimising transfection efficiency and maintaining cell viability. We have synthesised amphipathic asymmetric dendrimers by solid phase methods. The ability of two of these to transfect BHK cells in culture with β-galactosidase gene was determined by X-gal staining. Cell viability was measured by the MTT assay for BHK cells, and by spectroscopy for lysis of erythrocytes. Interactions between dendrimer and DNA were investigated by agarose gel electrophoresis. BHK cells were optimally transfected at 5:1 +/− charge ratio yielding 20% cells receiving at least one copy of the plasmid. Cell viability decreased when the dendrimer to DNA ratio exceeded 5:1. Raising the pH significantly affected the electrophoretic mobility of complexes of dendrimer and DNA. We conclude that amphipathic asymmetric dendrimers enable efficient plasmid DNA uptake into BHK cells. Cell viability is maintained at high concentrations of dendrimer when complexed with DNA at a 5:1 +/− charge ratio. Efficiency of transfection and cell viability suggest the system may be suitable for gene delivery in vivo.</abstract><qualityIndicators><score>7.449</score><pdfWordCount>3277</pdfWordCount><pdfCharCount>20337</pdfCharCount><pdfVersion>1.2</pdfVersion><pdfPageCount>8</pdfPageCount><pdfPageSize>527 x 670 pts</pdfPageSize><refBibsNative>true</refBibsNative><abstractWordCount>181</abstractWordCount><abstractCharCount>1216</abstractCharCount><keywordCount>12</keywordCount></qualityIndicators><title>DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers</title><pmid><json:string>11064210</json:string></pmid><pii><json:string>S0378-5173(00)00534-2</json:string></pii><genre><json:string>research-article</json:string></genre><host><title>International Journal of Pharmaceutics</title><language><json:string>unknown</json:string></language><publicationDate>2000</publicationDate><issn><json:string>0378-5173</json:string></issn><pii><json:string>S0378-5173(00)X0161-5</json:string></pii><volume>208</volume><issue>1–2</issue><pages><first>41</first><last>48</last></pages><genre><json:string>journal</json:string></genre></host><namedEntities><unitex><date><json:string>2000</json:string></date><geogName></geogName><orgName><json:string>Research, The School of Pharmacy, Uni</json:string><json:string>University of Queensland</json:string><json:string>University of London</json:string><json:string>UK Received</json:string></orgName><orgName_funder></orgName_funder><orgName_provider></orgName_provider><persName><json:string>D. Shah</json:string></persName><placeName><json:string>UK</json:string><json:string>Glasgow</json:string><json:string>Paisley</json:string></placeName><ref_url></ref_url><ref_bibl><json:string>Toth et al., 1999</json:string><json:string>Huddle et al., 1999</json:string><json:string>Schoen et al., 1999</json:string><json:string>Sakthivel et al., 1998</json:string><json:string>Kukowska-Latallo et al., 1996</json:string><json:string>Tang et al., 1996</json:string><json:string>D.S. Shah et al.</json:string><json:string>Kukowska-Latallo et al., 1999</json:string></ref_bibl><bibl></bibl></unitex></namedEntities><ark><json:string>ark:/67375/6H6-G5R0LGHP-C</json:string></ark><categories><wos><json:string>1 - science</json:string><json:string>2 - pharmacology & pharmacy</json:string></wos><scienceMetrix><json:string>1 - health sciences</json:string><json:string>2 - clinical medicine</json:string><json:string>3 - pharmacology & pharmacy</json:string></scienceMetrix><scopus><json:string>1 - Life Sciences</json:string><json:string>2 - Pharmacology, Toxicology and Pharmaceutics</json:string><json:string>3 - Pharmaceutical Science</json:string></scopus><inist><json:string>1 - sciences appliquees, technologies et medecines</json:string><json:string>2 - sciences biologiques et medicales</json:string><json:string>3 - sciences medicales</json:string><json:string>4 - pharmacologie. traitements medicamenteux</json:string></inist></categories><publicationDate>2000</publicationDate><copyrightDate>2000</copyrightDate><doi><json:string>10.1016/S0378-5173(00)00534-2</json:string></doi><id>CD94E85ACB7135922B0A5CDE0F4AA6EF02D6BE35</id><score>1</score><fulltext><json:item><extension>pdf</extension><original>true</original><mimetype>application/pdf</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-G5R0LGHP-C/fulltext.pdf</uri></json:item><json:item><extension>zip</extension><original>false</original><mimetype>application/zip</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-G5R0LGHP-C/bundle.zip</uri></json:item><istex:fulltextTEI uri="https://api.istex.fr/ark:/67375/6H6-G5R0LGHP-C/fulltext.tei"><teiHeader><fileDesc><titleStmt><title level="a" type="main" xml:lang="en">DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers</title></titleStmt><publicationStmt><authority>ISTEX</authority><publisher scheme="https://scientific-publisher.data.istex.fr">ELSEVIER</publisher><availability><licence><p>©2000 Elsevier Science B.V.</p></licence><p scheme="https://loaded-corpus.data.istex.fr/ark:/67375/XBH-HKKZVM7B-M">elsevier</p></availability><date>2000</date></publicationStmt><notesStmt><note type="research-article" scheme="https://content-type.data.istex.fr/ark:/67375/XTP-1JC4F85T-7">research-article</note><note type="journal" scheme="https://publication-type.data.istex.fr/ark:/67375/JMC-0GLKJH51-B">journal</note><note type="content">Fig. 1: Model of amphipathic asymmetric dendrimer structure. Black spheres are nitrogen atoms, terminal nitrogens are positively charged at neutral pH (a) (α/εLys)7(αAMA)3amide and (b) (α/εLys)15(αAMA)3amide.</note><note type="content">Fig. 2: pH dependence of complex stability. Identical samples of markers and plasmid pSVβgal (100 ng) were loaded onto both gels. (a) Agarose gel pH gradient 4–11 (b) Agarose gel pH 7–8. Lane numbers are as follows: lane 1, markers; lane 2, stock plasmid; lane 3, HindIII cut plasmid; lanes 4–7 are DNA/(α/εLys)7(αAMA)3amide complexes at 10:1, 5:1, 1:5, 1:10 +/− charge ratio, respectively.</note><note type="content">Fig. 3: (a) Red blood cell viability. Mean percent survival (triplicate) in the presence of (α/εLys)7(αAMA)3amide (A) and (α/εLys)15(αAMA)3amide (B). Values were scaled between 100% (addition of PBS) and 0% (addition of Triton X-100) by assaying haemoglobin release. The final concentration of pSVβgal DNA was 12.5 ng/μl. Error bars show standard deviation from mean. (b) BHK cell viability. Mean percent survival (triplicate) in the presence of (α/εLys)15(αAMA)3amide (compound B) Values were scaled between 100% (addition of PBS) and 0% (addition of Triton X-100) by MTT assay. The final concentration of pSVβgal DNA was 28 ng/l. Error bars show standard deviation (S.D.) from mean.</note><note type="content">Fig. 4: (a) Transfection efficiency: BHK cells transfected with (α/εLys)15(αAMA)3amide/ pSVβgal stained blue by incubation with β-galactosidase substrate X-gal. (b) Transfection efficiency vs concentration of (α/εLys)15(αAMA)3amide at three molar charge ratios. Mean percentage transfection (triplicate) measured as in (a) but with 2.5:1, 5:1, 10:1 +/− molar charge ratio. Error bars show S.D. from mean.</note></notesStmt><sourceDesc><biblStruct type="inbook"><analytic><title level="a" type="main" xml:lang="en">DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers</title><author xml:id="author-0000"><persName><forename type="first">Dipen S</forename><surname>Shah</surname></persName><affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation></author><author xml:id="author-0001"><persName><forename type="first">T</forename><surname>Sakthivel</surname></persName><affiliation>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation></author><author xml:id="author-0002"><persName><forename type="first">Istvan</forename><surname>Toth</surname></persName><affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation><affiliation>1 Present address: School of Pharmacy, Steele Building, University of Queensland, Brisbane, Qld 4072.</affiliation></author><author xml:id="author-0003"><persName><forename type="first">Alexander T</forename><surname>Florence</surname></persName><affiliation>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation></author><author xml:id="author-0004"><persName><forename type="first">Andy F</forename><surname>Wilderspin</surname></persName><affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation><affiliation>Corresponding author. Tel.: +44-207-7535878; fax: +44-207-7531939</affiliation></author><idno type="istex">CD94E85ACB7135922B0A5CDE0F4AA6EF02D6BE35</idno><idno type="ark">ark:/67375/6H6-G5R0LGHP-C</idno><idno type="DOI">10.1016/S0378-5173(00)00534-2</idno><idno type="PII">S0378-5173(00)00534-2</idno></analytic><monogr><title level="j">International Journal of Pharmaceutics</title><title level="j" type="abbrev">IJP</title><idno type="pISSN">0378-5173</idno><idno type="PII">S0378-5173(00)X0161-5</idno><imprint><publisher>ELSEVIER</publisher><date type="published" when="2000"></date><biblScope unit="volume">208</biblScope><biblScope unit="issue">1–2</biblScope><biblScope unit="page" from="41">41</biblScope><biblScope unit="page" to="48">48</biblScope></imprint></monogr></biblStruct></sourceDesc></fileDesc><profileDesc><creation><date>2000</date></creation><langUsage><language ident="en">en</language></langUsage><abstract xml:lang="en"><p>Abstract: Amphipathic asymmetric dendrimers have been investigated for use in delivery of genes into cells, with the objective of optimising transfection efficiency and maintaining cell viability. We have synthesised amphipathic asymmetric dendrimers by solid phase methods. The ability of two of these to transfect BHK cells in culture with β-galactosidase gene was determined by X-gal staining. Cell viability was measured by the MTT assay for BHK cells, and by spectroscopy for lysis of erythrocytes. Interactions between dendrimer and DNA were investigated by agarose gel electrophoresis. BHK cells were optimally transfected at 5:1 +/− charge ratio yielding 20% cells receiving at least one copy of the plasmid. Cell viability decreased when the dendrimer to DNA ratio exceeded 5:1. Raising the pH significantly affected the electrophoretic mobility of complexes of dendrimer and DNA. We conclude that amphipathic asymmetric dendrimers enable efficient plasmid DNA uptake into BHK cells. Cell viability is maintained at high concentrations of dendrimer when complexed with DNA at a 5:1 +/− charge ratio. Efficiency of transfection and cell viability suggest the system may be suitable for gene delivery in vivo.</p></abstract><textClass xml:lang="en"><keywords scheme="keyword"><list><head>Keywords</head><item><term>Gene delivery</term></item><item><term>Gene expression</term></item><item><term>Transfection</term></item><item><term>Dendrimers</term></item><item><term>Non-viral vectors</term></item></list></keywords></textClass><textClass xml:lang="en"><keywords scheme="keyword"><list><head>Abbreviations</head><item><term>DOTMA, 2,3-diokeyloxypropyl-1-trimethyl ammonium chloride</term></item><item><term>DOSPA, 2,3-dioleyloxy-N-[2(Spermine-carboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoracrtate</term></item><item><term>DOGS, dioctadecylamidoglycylspermime</term></item><item><term>DOPE, dioleoyl phosphatidylethanolamine</term></item><item><term>DODAC, dioleyldimethylammonium chloride</term></item><item><term>PAMAM, polyamidoamine</term></item><item><term>G9-EDA, 9th generation PAMAN with ethylenediamine core</term></item></list></keywords></textClass></profileDesc><revisionDesc><change when="2000-07-31">Modified</change><change when="2000">Published</change></revisionDesc></teiHeader></istex:fulltextTEI><json:item><extension>txt</extension><original>false</original><mimetype>text/plain</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-G5R0LGHP-C/fulltext.txt</uri></json:item></fulltext><metadata><istex:metadataXml wicri:clean="Elsevier, elements deleted: ce:floats; body; tail"><istex:xmlDeclaration>version="1.0" encoding="utf-8"</istex:xmlDeclaration><istex:docType PUBLIC="-//ES//DTD journal article DTD version 4.5.2//EN//XML" URI="art452.dtd" name="istex:docType"><istex:entity SYSTEM="gr1" NDATA="IMAGE" name="gr1"></istex:entity><istex:entity SYSTEM="gr2" NDATA="IMAGE" name="gr2"></istex:entity><istex:entity SYSTEM="gr3" NDATA="IMAGE" name="gr3"></istex:entity><istex:entity SYSTEM="gr4" NDATA="IMAGE" name="gr4"></istex:entity></istex:docType><istex:document><converted-article version="4.5.2" docsubtype="fla" xml:lang="en"><item-info><jid>IJP</jid><aid>6341</aid><ce:pii>S0378-5173(00)00534-2</ce:pii><ce:doi>10.1016/S0378-5173(00)00534-2</ce:doi><ce:copyright type="full-transfer" year="2000">Elsevier Science B.V.</ce:copyright></item-info><head><ce:title>DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers</ce:title><ce:author-group><ce:author><ce:given-name>Dipen S</ce:given-name><ce:surname>Shah</ce:surname><ce:cross-ref refid="AFF1"><ce:sup>a</ce:sup></ce:cross-ref></ce:author><ce:author><ce:given-name>T</ce:given-name><ce:surname>Sakthivel</ce:surname><ce:cross-ref refid="AFF2"><ce:sup>b</ce:sup></ce:cross-ref></ce:author><ce:author><ce:given-name>Istvan</ce:given-name><ce:surname>Toth</ce:surname><ce:cross-ref refid="FN1"><ce:sup>1</ce:sup></ce:cross-ref><ce:cross-ref refid="AFF1"><ce:sup>a</ce:sup></ce:cross-ref></ce:author><ce:author><ce:given-name>Alexander T</ce:given-name><ce:surname>Florence</ce:surname><ce:cross-ref refid="AFF2"><ce:sup>b</ce:sup></ce:cross-ref></ce:author><ce:author><ce:given-name>Andy F</ce:given-name><ce:surname>Wilderspin</ce:surname><ce:cross-ref refid="AFF1"><ce:sup>a</ce:sup></ce:cross-ref><ce:cross-ref refid="CORR1">*</ce:cross-ref></ce:author><ce:affiliation id="AFF1"><ce:label>a</ce:label><ce:textfn>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</ce:textfn></ce:affiliation><ce:affiliation id="AFF2"><ce:label>b</ce:label><ce:textfn>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</ce:textfn></ce:affiliation><ce:correspondence id="CORR1"><ce:label>*</ce:label><ce:text>Corresponding author. Tel.: +44-207-7535878; fax: +44-207-7531939</ce:text></ce:correspondence><ce:footnote id="FN1"><ce:label>1</ce:label><ce:note-para>Present address: School of Pharmacy, Steele Building, University of Queensland, Brisbane, Qld 4072.</ce:note-para></ce:footnote></ce:author-group><ce:date-received day="10" month="5" year="2000"></ce:date-received><ce:date-revised day="31" month="7" year="2000"></ce:date-revised><ce:date-accepted day="1" month="8" year="2000"></ce:date-accepted><ce:abstract><ce:section-title>Abstract</ce:section-title><ce:abstract-sec><ce:simple-para>Amphipathic asymmetric dendrimers have been investigated for use in delivery of genes into cells, with the objective of optimising transfection efficiency and maintaining cell viability. We have synthesised amphipathic asymmetric dendrimers by solid phase methods. The ability of two of these to transfect BHK cells in culture with β-galactosidase gene was determined by X-gal staining. Cell viability was measured by the MTT assay for BHK cells, and by spectroscopy for lysis of erythrocytes. Interactions between dendrimer and DNA were investigated by agarose gel electrophoresis. BHK cells were optimally transfected at 5:1 +/− charge ratio yielding 20% cells receiving at least one copy of the plasmid. Cell viability decreased when the dendrimer to DNA ratio exceeded 5:1. Raising the pH significantly affected the electrophoretic mobility of complexes of dendrimer and DNA. We conclude that amphipathic asymmetric dendrimers enable efficient plasmid DNA uptake into BHK cells. Cell viability is maintained at high concentrations of dendrimer when complexed with DNA at a 5:1 +/− charge ratio. Efficiency of transfection and cell viability suggest the system may be suitable for gene delivery in vivo.</ce:simple-para></ce:abstract-sec></ce:abstract><ce:keywords class="keyword"><ce:section-title>Keywords</ce:section-title><ce:keyword><ce:text>Gene delivery</ce:text></ce:keyword><ce:keyword><ce:text>Gene expression</ce:text></ce:keyword><ce:keyword><ce:text>Transfection</ce:text></ce:keyword><ce:keyword><ce:text>Dendrimers</ce:text></ce:keyword><ce:keyword><ce:text>Non-viral vectors</ce:text></ce:keyword></ce:keywords><ce:keywords class="abr"><ce:section-title>Abbreviations</ce:section-title><ce:keyword><ce:text>DOTMA, 2,3-diokeyloxypropyl-1-trimethyl ammonium chloride</ce:text></ce:keyword><ce:keyword><ce:text>DOSPA, 2,3-dioleyloxy-<ce:italic>N</ce:italic>-[2(Spermine-carboxamido)ethyl]-<ce:italic>N</ce:italic>,<ce:italic>N</ce:italic>-dimethyl-1-propanaminium trifluoracrtate</ce:text></ce:keyword><ce:keyword><ce:text>DOGS, dioctadecylamidoglycylspermime</ce:text></ce:keyword><ce:keyword><ce:text>DOPE, dioleoyl phosphatidylethanolamine</ce:text></ce:keyword><ce:keyword><ce:text>DODAC, dioleyldimethylammonium chloride</ce:text></ce:keyword><ce:keyword><ce:text>PAMAM, polyamidoamine</ce:text></ce:keyword><ce:keyword><ce:text>G9-EDA, 9th generation PAMAN with ethylenediamine core</ce:text></ce:keyword></ce:keywords></head></converted-article></istex:document></istex:metadataXml><mods version="3.6"><titleInfo lang="en"><title>DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers</title></titleInfo><titleInfo type="alternative" lang="en" contentType="CDATA"><title>DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers</title></titleInfo><name type="personal"><namePart type="given">Dipen S</namePart><namePart type="family">Shah</namePart><affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">T</namePart><namePart type="family">Sakthivel</namePart><affiliation>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Istvan</namePart><namePart type="family">Toth</namePart><affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation><affiliation>1 Present address: School of Pharmacy, Steele Building, University of Queensland, Brisbane, Qld 4072.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Alexander T</namePart><namePart type="family">Florence</namePart><affiliation>Centre for Drug Delivery Research, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Andy F</namePart><namePart type="family">Wilderspin</namePart><affiliation>Department of Pharmaceutical and Biological Chemistry, The School of Pharmacy, University of London, 29/39, Brunswick Square, London WC1N 1AX, UK</affiliation><affiliation>Corresponding author. Tel.: +44-207-7535878; fax: +44-207-7531939</affiliation><role><roleTerm type="text">author</roleTerm></role></name><typeOfResource>text</typeOfResource><genre type="research-article" displayLabel="Full-length article" authority="ISTEX" authorityURI="https://content-type.data.istex.fr" valueURI="https://content-type.data.istex.fr/ark:/67375/XTP-1JC4F85T-7">research-article</genre><originInfo><publisher>ELSEVIER</publisher><dateIssued encoding="w3cdtf">2000</dateIssued><dateModified encoding="w3cdtf">2000-07-31</dateModified><copyrightDate encoding="w3cdtf">2000</copyrightDate></originInfo><language><languageTerm type="code" authority="iso639-2b">eng</languageTerm><languageTerm type="code" authority="rfc3066">en</languageTerm></language><abstract lang="en">Abstract: Amphipathic asymmetric dendrimers have been investigated for use in delivery of genes into cells, with the objective of optimising transfection efficiency and maintaining cell viability. We have synthesised amphipathic asymmetric dendrimers by solid phase methods. The ability of two of these to transfect BHK cells in culture with β-galactosidase gene was determined by X-gal staining. Cell viability was measured by the MTT assay for BHK cells, and by spectroscopy for lysis of erythrocytes. Interactions between dendrimer and DNA were investigated by agarose gel electrophoresis. BHK cells were optimally transfected at 5:1 +/− charge ratio yielding 20% cells receiving at least one copy of the plasmid. Cell viability decreased when the dendrimer to DNA ratio exceeded 5:1. Raising the pH significantly affected the electrophoretic mobility of complexes of dendrimer and DNA. We conclude that amphipathic asymmetric dendrimers enable efficient plasmid DNA uptake into BHK cells. Cell viability is maintained at high concentrations of dendrimer when complexed with DNA at a 5:1 +/− charge ratio. Efficiency of transfection and cell viability suggest the system may be suitable for gene delivery in vivo.</abstract><note type="content">Fig. 1: Model of amphipathic asymmetric dendrimer structure. Black spheres are nitrogen atoms, terminal nitrogens are positively charged at neutral pH (a) (α/εLys)7(αAMA)3amide and (b) (α/εLys)15(αAMA)3amide.</note><note type="content">Fig. 2: pH dependence of complex stability. Identical samples of markers and plasmid pSVβgal (100 ng) were loaded onto both gels. (a) Agarose gel pH gradient 4–11 (b) Agarose gel pH 7–8. Lane numbers are as follows: lane 1, markers; lane 2, stock plasmid; lane 3, HindIII cut plasmid; lanes 4–7 are DNA/(α/εLys)7(αAMA)3amide complexes at 10:1, 5:1, 1:5, 1:10 +/− charge ratio, respectively.</note><note type="content">Fig. 3: (a) Red blood cell viability. Mean percent survival (triplicate) in the presence of (α/εLys)7(αAMA)3amide (A) and (α/εLys)15(αAMA)3amide (B). Values were scaled between 100% (addition of PBS) and 0% (addition of Triton X-100) by assaying haemoglobin release. The final concentration of pSVβgal DNA was 12.5 ng/μl. Error bars show standard deviation from mean. (b) BHK cell viability. Mean percent survival (triplicate) in the presence of (α/εLys)15(αAMA)3amide (compound B) Values were scaled between 100% (addition of PBS) and 0% (addition of Triton X-100) by MTT assay. The final concentration of pSVβgal DNA was 28 ng/l. Error bars show standard deviation (S.D.) from mean.</note><note type="content">Fig. 4: (a) Transfection efficiency: BHK cells transfected with (α/εLys)15(αAMA)3amide/ pSVβgal stained blue by incubation with β-galactosidase substrate X-gal. (b) Transfection efficiency vs concentration of (α/εLys)15(αAMA)3amide at three molar charge ratios. Mean percentage transfection (triplicate) measured as in (a) but with 2.5:1, 5:1, 10:1 +/− molar charge ratio. Error bars show S.D. from mean.</note><subject lang="en"><genre>Keywords</genre><topic>Gene delivery</topic><topic>Gene expression</topic><topic>Transfection</topic><topic>Dendrimers</topic><topic>Non-viral vectors</topic></subject><subject lang="en"><genre>Abbreviations</genre><topic>DOTMA, 2,3-diokeyloxypropyl-1-trimethyl ammonium chloride</topic><topic>DOSPA, 2,3-dioleyloxy-N-[2(Spermine-carboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoracrtate</topic><topic>DOGS, dioctadecylamidoglycylspermime</topic><topic>DOPE, dioleoyl phosphatidylethanolamine</topic><topic>DODAC, dioleyldimethylammonium chloride</topic><topic>PAMAM, polyamidoamine</topic><topic>G9-EDA, 9th generation PAMAN with ethylenediamine core</topic></subject><relatedItem type="host"><titleInfo><title>International Journal of Pharmaceutics</title></titleInfo><titleInfo type="abbreviated"><title>IJP</title></titleInfo><genre type="journal" authority="ISTEX" authorityURI="https://publication-type.data.istex.fr" valueURI="https://publication-type.data.istex.fr/ark:/67375/JMC-0GLKJH51-B">journal</genre><originInfo><publisher>ELSEVIER</publisher><dateIssued encoding="w3cdtf">2000</dateIssued></originInfo><identifier type="ISSN">0378-5173</identifier><identifier type="PII">S0378-5173(00)X0161-5</identifier><part><date>2000</date><detail type="volume"><number>208</number><caption>vol.</caption></detail><detail type="issue"><number>1–2</number><caption>no.</caption></detail><extent unit="issue-pages"><start>1</start><end>130</end></extent><extent unit="pages"><start>41</start><end>48</end></extent></part></relatedItem><identifier type="istex">CD94E85ACB7135922B0A5CDE0F4AA6EF02D6BE35</identifier><identifier type="ark">ark:/67375/6H6-G5R0LGHP-C</identifier><identifier type="DOI">10.1016/S0378-5173(00)00534-2</identifier><identifier type="PII">S0378-5173(00)00534-2</identifier><accessCondition type="use and reproduction" contentType="copyright">©2000 Elsevier Science B.V.</accessCondition><recordInfo><recordContentSource authority="ISTEX" authorityURI="https://loaded-corpus.data.istex.fr" valueURI="https://loaded-corpus.data.istex.fr/ark:/67375/XBH-HKKZVM7B-M">elsevier</recordContentSource><recordOrigin>Elsevier Science B.V., ©2000</recordOrigin></recordInfo></mods><json:item><extension>json</extension><original>false</original><mimetype>application/json</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-G5R0LGHP-C/record.json</uri></json:item></metadata><serie></serie></istex></record>Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Sante/explor/ChloroquineV1/Data/Istex/Corpus
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 002726 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Istex/Corpus/biblio.hfd -nk 002726 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien
|wiki= Sante
|area= ChloroquineV1
|flux= Istex
|étape= Corpus
|type= RBID
|clé= ISTEX:CD94E85ACB7135922B0A5CDE0F4AA6EF02D6BE35
|texte= DNA transfection and transfected cell viability using amphipathic asymmetric dendrimers
}}
| This area was generated with Dilib version V0.6.33. |  |