Histone H1-Mediated Transfection: Serum Inhibition Can Be Overcome by Ca2+ Ions
Identifieur interne : 000515 ( Istex/Corpus ); précédent : 000514; suivant : 000516Histone H1-Mediated Transfection: Serum Inhibition Can Be Overcome by Ca2+ Ions
Auteurs : Annekathrin Haberland ; Thomas Knaus ; Sergei V. Zaitsev ; Bernd Buchberger ; Andreas Lun ; Hermann Haller ; Michael BöttgerSource :
- Pharmaceutical Research [ 0724-8741 ] ; 2000-02-01.
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Abstract
Abstract: Purpose. One of the drawbacks of polycationic and cationic liposomalgene transfer is its sensitivity to serum. Gene therapy requires thetransfectant-DNA complex to be resistant to serum as well as blood.Since Ca2+ has proved to be an efficient cofactor of polycationic genetransfer, we decided to investigate its effects on transfection in thepresence of serum. Methods. We studied transgene expression of luciferase gene (pCMVLuc) on ECV 304 human endothelial cells using H1 histone andDOSPER as transfectants in the presence of 0-100% fetal calf serum. Results. H1-and DOSPER-mediated transfection was found to beinhibited by serum above the concentration of 10%. If 2 mM Ca2+ or2 mM Ca2+/0.1 mM chloroquine was included in the culture mediumwhich replace the transfection mixture and was left on the cells for24 hours postincubation, the inhibiting effect of even 100% serumwas overcome. Conclusions. A high serum level does not interfere with binding anduptake of H1- and DOSPER-DNA complexes, but inhibits subsequentsteps such as endosomal escape. Ca2+ in the form of nascent calciumphosphate microprecipitates and other lysosomolytical agents facilitateendosomal/lysosomal release by their fusigenic and membranolyticactivity.
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DOI: 10.1023/A:1007581700996
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<record><TEI wicri:istexFullTextTei="biblStruct"><teiHeader><fileDesc><titleStmt><title xml:lang="en">Histone H1-Mediated Transfection: Serum Inhibition Can Be Overcome by Ca2+ Ions</title><author><name sortKey="Haberland, Annekathrin" sort="Haberland, Annekathrin" uniqKey="Haberland A" first="Annekathrin" last="Haberland">Annekathrin Haberland</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation></author><author><name sortKey="Knaus, Thomas" sort="Knaus, Thomas" uniqKey="Knaus T" first="Thomas" last="Knaus">Thomas Knaus</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation></author><author><name sortKey="Zaitsev, Sergei V" sort="Zaitsev, Sergei V" uniqKey="Zaitsev S" first="Sergei V." last="Zaitsev">Sergei V. Zaitsev</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation><affiliation><mods:affiliation>Institute of Cytology of the Russian Academy of Sciences, St. Petersburg, Russia</mods:affiliation></affiliation></author><author><name sortKey="Buchberger, Bernd" sort="Buchberger, Bernd" uniqKey="Buchberger B" first="Bernd" last="Buchberger">Bernd Buchberger</name><affiliation><mods:affiliation>Roche Diagnostics GmbH, Penzberg, Germany</mods:affiliation></affiliation></author><author><name sortKey="Lun, Andreas" sort="Lun, Andreas" uniqKey="Lun A" first="Andreas" last="Lun">Andreas Lun</name><affiliation><mods:affiliation>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</mods:affiliation></affiliation></author><author><name sortKey="Haller, Hermann" sort="Haller, Hermann" uniqKey="Haller H" first="Hermann" last="Haller">Hermann Haller</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation></author><author><name sortKey="Bottger, Michael" sort="Bottger, Michael" uniqKey="Bottger M" first="Michael" last="Böttger">Michael Böttger</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation><affiliation><mods:affiliation>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</mods:affiliation></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">ISTEX</idno><idno type="RBID">ISTEX:5E39256AF6D067F01D2FA923B6BCC3574FA0A9AD</idno><date when="2000" year="2000">2000</date><idno type="doi">10.1023/A:1007581700996</idno><idno type="url">https://api.istex.fr/ark:/67375/VQC-Z8N2MRJ5-4/fulltext.pdf</idno><idno type="wicri:Area/Istex/Corpus">000515</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Corpus" wicri:corpus="ISTEX">000515</idno></publicationStmt><sourceDesc><biblStruct><analytic><title level="a" type="main" xml:lang="en">Histone H1-Mediated Transfection: Serum Inhibition Can Be Overcome by Ca2+ Ions</title><author><name sortKey="Haberland, Annekathrin" sort="Haberland, Annekathrin" uniqKey="Haberland A" first="Annekathrin" last="Haberland">Annekathrin Haberland</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation></author><author><name sortKey="Knaus, Thomas" sort="Knaus, Thomas" uniqKey="Knaus T" first="Thomas" last="Knaus">Thomas Knaus</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation></author><author><name sortKey="Zaitsev, Sergei V" sort="Zaitsev, Sergei V" uniqKey="Zaitsev S" first="Sergei V." last="Zaitsev">Sergei V. Zaitsev</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation><affiliation><mods:affiliation>Institute of Cytology of the Russian Academy of Sciences, St. Petersburg, Russia</mods:affiliation></affiliation></author><author><name sortKey="Buchberger, Bernd" sort="Buchberger, Bernd" uniqKey="Buchberger B" first="Bernd" last="Buchberger">Bernd Buchberger</name><affiliation><mods:affiliation>Roche Diagnostics GmbH, Penzberg, Germany</mods:affiliation></affiliation></author><author><name sortKey="Lun, Andreas" sort="Lun, Andreas" uniqKey="Lun A" first="Andreas" last="Lun">Andreas Lun</name><affiliation><mods:affiliation>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</mods:affiliation></affiliation></author><author><name sortKey="Haller, Hermann" sort="Haller, Hermann" uniqKey="Haller H" first="Hermann" last="Haller">Hermann Haller</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation></author><author><name sortKey="Bottger, Michael" sort="Bottger, Michael" uniqKey="Bottger M" first="Michael" last="Böttger">Michael Böttger</name><affiliation><mods:affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</mods:affiliation></affiliation><affiliation><mods:affiliation>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</mods:affiliation></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="2000-02-01">2000-02-01</date><biblScope unit="volume">17</biblScope><biblScope unit="issue">2</biblScope><biblScope unit="page" from="229">229</biblScope><biblScope unit="page" to="235">235</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>DOSPER</term><term>H1 histone</term><term>calcium</term><term>serum inhibition</term><term>transfection</term></keywords></textClass><langUsage><language ident="en">en</language></langUsage></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Abstract: Purpose. One of the drawbacks of polycationic and cationic liposomalgene transfer is its sensitivity to serum. Gene therapy requires thetransfectant-DNA complex to be resistant to serum as well as blood.Since Ca2+ has proved to be an efficient cofactor of polycationic genetransfer, we decided to investigate its effects on transfection in thepresence of serum. Methods. We studied transgene expression of luciferase gene (pCMVLuc) on ECV 304 human endothelial cells using H1 histone andDOSPER as transfectants in the presence of 0-100% fetal calf serum. Results. H1-and DOSPER-mediated transfection was found to beinhibited by serum above the concentration of 10%. If 2 mM Ca2+ or2 mM Ca2+/0.1 mM chloroquine was included in the culture mediumwhich replace the transfection mixture and was left on the cells for24 hours postincubation, the inhibiting effect of even 100% serumwas overcome. Conclusions. A high serum level does not interfere with binding anduptake of H1- and DOSPER-DNA complexes, but inhibits subsequentsteps such as endosomal escape. Ca2+ in the form of nascent calciumphosphate microprecipitates and other lysosomolytical agents facilitateendosomal/lysosomal release by their fusigenic and membranolyticactivity.</div></front></TEI><istex><corpusName>springer-journals</corpusName><author><json:item><name>Annekathrin Haberland</name><affiliations><json:string>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</json:string></affiliations></json:item><json:item><name>Thomas Knaus</name><affiliations><json:string>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</json:string></affiliations></json:item><json:item><name>Sergei V. Zaitsev</name><affiliations><json:string>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</json:string><json:string>Institute of Cytology of the Russian Academy of Sciences, St. Petersburg, Russia</json:string></affiliations></json:item><json:item><name>Bernd Buchberger</name><affiliations><json:string>Roche Diagnostics GmbH, Penzberg, Germany</json:string></affiliations></json:item><json:item><name>Andreas Lun</name><affiliations><json:string>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</json:string></affiliations></json:item><json:item><name>Hermann Haller</name><affiliations><json:string>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</json:string></affiliations></json:item><json:item><name>Michael Böttger</name><affiliations><json:string>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</json:string><json:string>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</json:string></affiliations></json:item></author><subject><json:item><lang><json:string>eng</json:string></lang><value>transfection</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>serum inhibition</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>H1 histone</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>DOSPER</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>calcium</value></json:item></subject><articleId><json:string>221578</json:string><json:string>Art19</json:string></articleId><arkIstex>ark:/67375/VQC-Z8N2MRJ5-4</arkIstex><language><json:string>eng</json:string></language><originalGenre><json:string>OriginalPaper</json:string></originalGenre><abstract>Abstract: Purpose. One of the drawbacks of polycationic and cationic liposomalgene transfer is its sensitivity to serum. Gene therapy requires thetransfectant-DNA complex to be resistant to serum as well as blood.Since Ca2+ has proved to be an efficient cofactor of polycationic genetransfer, we decided to investigate its effects on transfection in thepresence of serum. Methods. We studied transgene expression of luciferase gene (pCMVLuc) on ECV 304 human endothelial cells using H1 histone andDOSPER as transfectants in the presence of 0-100% fetal calf serum. Results. H1-and DOSPER-mediated transfection was found to beinhibited by serum above the concentration of 10%. If 2 mM Ca2+ or2 mM Ca2+/0.1 mM chloroquine was included in the culture mediumwhich replace the transfection mixture and was left on the cells for24 hours postincubation, the inhibiting effect of even 100% serumwas overcome. Conclusions. A high serum level does not interfere with binding anduptake of H1- and DOSPER-DNA complexes, but inhibits subsequentsteps such as endosomal escape. Ca2+ in the form of nascent calciumphosphate microprecipitates and other lysosomolytical agents facilitateendosomal/lysosomal release by their fusigenic and membranolyticactivity.</abstract><qualityIndicators><score>8.955</score><pdfWordCount>4843</pdfWordCount><pdfCharCount>29054</pdfCharCount><pdfVersion>1.3</pdfVersion><pdfPageCount>7</pdfPageCount><pdfPageSize>612 x 792 pts (letter)</pdfPageSize><refBibsNative>false</refBibsNative><abstractWordCount>176</abstractWordCount><abstractCharCount>1242</abstractCharCount><keywordCount>5</keywordCount></qualityIndicators><title>Histone H1-Mediated Transfection: Serum Inhibition Can Be Overcome by Ca2+ Ions</title><genre><json:string>research-article</json:string></genre><host><title>Pharmaceutical 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scheme="https://scientific-publisher.data.istex.fr">Kluwer Academic Publishers-Plenum Publishers</publisher><pubPlace>New York</pubPlace><availability><licence><p>Plenum Publishing Corporation, 2000</p></licence><p scheme="https://loaded-corpus.data.istex.fr/ark:/67375/XBH-3XSW68JL-F">springer</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></notesStmt><sourceDesc><biblStruct type="inbook"><analytic><title level="a" type="main" xml:lang="en">Histone H1-Mediated Transfection: Serum Inhibition Can Be Overcome by Ca2+ Ions</title><author xml:id="author-0000"><persName><forename type="first">Annekathrin</forename><surname>Haberland</surname></persName><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation></author><author xml:id="author-0001"><persName><forename type="first">Thomas</forename><surname>Knaus</surname></persName><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation></author><author xml:id="author-0002"><persName><forename type="first">Sergei</forename><surname>Zaitsev</surname></persName><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation><affiliation>Institute of Cytology of the Russian Academy of Sciences, St. Petersburg, Russia</affiliation></author><author xml:id="author-0003"><persName><forename type="first">Bernd</forename><surname>Buchberger</surname></persName><affiliation>Roche Diagnostics GmbH, Penzberg, Germany</affiliation></author><author xml:id="author-0004"><persName><forename type="first">Andreas</forename><surname>Lun</surname></persName><affiliation>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</affiliation></author><author xml:id="author-0005"><persName><forename type="first">Hermann</forename><surname>Haller</surname></persName><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation></author><author xml:id="author-0006"><persName><forename type="first">Michael</forename><surname>Böttger</surname></persName><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation><affiliation>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</affiliation></author><idno type="istex">5E39256AF6D067F01D2FA923B6BCC3574FA0A9AD</idno><idno type="ark">ark:/67375/VQC-Z8N2MRJ5-4</idno><idno type="DOI">10.1023/A:1007581700996</idno><idno type="article-id">221578</idno><idno type="article-id">Art19</idno></analytic><monogr><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="pISSN">0724-8741</idno><idno type="eISSN">1573-904X</idno><idno type="journal-ID">true</idno><idno type="issue-article-count">22</idno><idno type="volume-issue-count">12</idno><imprint><publisher>Kluwer Academic Publishers-Plenum Publishers</publisher><pubPlace>New York</pubPlace><date type="published" when="2000-02-01"></date><biblScope unit="volume">17</biblScope><biblScope unit="issue">2</biblScope><biblScope unit="page" from="229">229</biblScope><biblScope unit="page" to="235">235</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: Purpose. One of the drawbacks of polycationic and cationic liposomalgene transfer is its sensitivity to serum. Gene therapy requires thetransfectant-DNA complex to be resistant to serum as well as blood.Since Ca2+ has proved to be an efficient cofactor of polycationic genetransfer, we decided to investigate its effects on transfection in thepresence of serum. Methods. We studied transgene expression of luciferase gene (pCMVLuc) on ECV 304 human endothelial cells using H1 histone andDOSPER as transfectants in the presence of 0-100% fetal calf serum. Results. H1-and DOSPER-mediated transfection was found to beinhibited by serum above the concentration of 10%. If 2 mM Ca2+ or2 mM Ca2+/0.1 mM chloroquine was included in the culture mediumwhich replace the transfection mixture and was left on the cells for24 hours postincubation, the inhibiting effect of even 100% serumwas overcome. Conclusions. A high serum level does not interfere with binding anduptake of H1- and DOSPER-DNA complexes, but inhibits subsequentsteps such as endosomal escape. Ca2+ in the form of nascent calciumphosphate microprecipitates and other lysosomolytical agents facilitateendosomal/lysosomal release by their fusigenic and membranolyticactivity.</p></abstract><textClass xml:lang="en"><keywords scheme="keyword"><list><item><term>transfection</term></item><item><term>serum inhibition</term></item><item><term>H1 histone</term></item><item><term>DOSPER</term></item><item><term>calcium</term></item></list></keywords></textClass><textClass><keywords scheme="Journal Subject"><list><head>Biomedicine</head><item><term>Pharmacology/Toxicology</term></item><item><term>Pharmacy</term></item><item><term>Biochemistry, general</term></item><item><term>Medical Law</term></item><item><term>Biomedical Engineering</term></item></list></keywords></textClass></profileDesc><revisionDesc><change 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Ions</ArticleTitle><ArticleFirstPage>229</ArticleFirstPage><ArticleLastPage>235</ArticleLastPage><ArticleHistory><RegistrationDate><Year>2004</Year><Month>10</Month><Day>8</Day></RegistrationDate></ArticleHistory><ArticleCopyright><CopyrightHolderName>Plenum Publishing Corporation</CopyrightHolderName><CopyrightYear>2000</CopyrightYear></ArticleCopyright><ArticleGrants Type="Regular"><MetadataGrant Grant="OpenAccess"></MetadataGrant><AbstractGrant Grant="OpenAccess"></AbstractGrant><BodyPDFGrant Grant="Restricted"></BodyPDFGrant><BodyHTMLGrant Grant="Restricted"></BodyHTMLGrant><BibliographyGrant Grant="Restricted"></BibliographyGrant><ESMGrant Grant="Restricted"></ESMGrant></ArticleGrants><ArticleContext><JournalID>11095</JournalID><VolumeIDStart>17</VolumeIDStart><VolumeIDEnd>17</VolumeIDEnd><IssueIDStart>2</IssueIDStart><IssueIDEnd>2</IssueIDEnd></ArticleContext></ArticleInfo><ArticleHeader><AuthorGroup><Author AffiliationIDS="Aff1"><AuthorName DisplayOrder="Western"><GivenName>Annekathrin</GivenName><FamilyName>Haberland</FamilyName></AuthorName></Author><Author AffiliationIDS="Aff1"><AuthorName DisplayOrder="Western"><GivenName>Thomas</GivenName><FamilyName>Knaus</FamilyName></AuthorName></Author><Author AffiliationIDS="Aff1 Aff2"><AuthorName DisplayOrder="Western"><GivenName>Sergei</GivenName><GivenName>V.</GivenName><FamilyName>Zaitsev</FamilyName></AuthorName></Author><Author AffiliationIDS="Aff3"><AuthorName DisplayOrder="Western"><GivenName>Bernd</GivenName><FamilyName>Buchberger</FamilyName></AuthorName></Author><Author AffiliationIDS="Aff4"><AuthorName DisplayOrder="Western"><GivenName>Andreas</GivenName><FamilyName>Lun</FamilyName></AuthorName></Author><Author AffiliationIDS="Aff1"><AuthorName DisplayOrder="Western"><GivenName>Hermann</GivenName><FamilyName>Haller</FamilyName></AuthorName></Author><Author AffiliationIDS="Aff1 Aff4"><AuthorName DisplayOrder="Western"><GivenName>Michael</GivenName><FamilyName>Böttger</FamilyName></AuthorName></Author><Affiliation ID="Aff1"><OrgDivision>Max Delbrück Center for Molecular Medicine</OrgDivision><OrgName>Franz Volhard Clinic</OrgName><OrgAddress><City>Berlin-Buch</City><Country>Germany</Country></OrgAddress></Affiliation><Affiliation ID="Aff2"><OrgName>Institute of Cytology of the Russian Academy of Sciences</OrgName><OrgAddress><City>St. Petersburg</City><Country>Russia</Country></OrgAddress></Affiliation><Affiliation ID="Aff3"><OrgName>Roche Diagnostics GmbH</OrgName><OrgAddress><City>Penzberg</City><Country>Germany</Country></OrgAddress></Affiliation><Affiliation ID="Aff4"><OrgDivision>Institute of Laboratory Medicine and Pathobiochemistry, Charité</OrgDivision><OrgName>Humboldt University</OrgName><OrgAddress><City>Berlin</City><Country>Germany</Country></OrgAddress></Affiliation></AuthorGroup><Abstract ID="Abs1" Language="En"><Heading>Abstract</Heading><Para><Emphasis Type="BoldItalic">Purpose</Emphasis>. One of the drawbacks of polycationic and cationic liposomalgene transfer is its sensitivity to serum. Gene therapy requires thetransfectant-DNA complex to be resistant to serum as well as blood.Since Ca<Superscript>2+</Superscript> has proved to be an efficient cofactor of polycationic genetransfer, we decided to investigate its effects on transfection in thepresence of serum.</Para><Para><Emphasis Type="BoldItalic">Methods</Emphasis>. We studied transgene expression of luciferase gene (pCMVLuc) on ECV 304 human endothelial cells using H1 histone andDOSPER as transfectants in the presence of 0-100% fetal calf serum.</Para><Para><Emphasis Type="BoldItalic">Results</Emphasis>. H1-and DOSPER-mediated transfection was found to beinhibited by serum above the concentration of 10%. If 2 mM Ca<Superscript>2+</Superscript> or2 mM Ca<Superscript>2+</Superscript>/0.1 mM chloroquine was included in the culture mediumwhich replace the transfection mixture and was left on the cells for24 hours postincubation, the inhibiting effect of even 100% serumwas overcome.</Para><Para><Emphasis Type="BoldItalic">Conclusions</Emphasis>. A high serum level does not interfere with binding anduptake of H1- and DOSPER-DNA complexes, but inhibits subsequentsteps such as endosomal escape. Ca<Superscript>2+</Superscript> in the form of nascent calciumphosphate microprecipitates and other lysosomolytical agents facilitateendosomal/lysosomal release by their fusigenic and membranolyticactivity.</Para></Abstract><KeywordGroup Language="En"><Keyword>transfection</Keyword><Keyword>serum inhibition</Keyword><Keyword>H1 histone</Keyword><Keyword>DOSPER</Keyword><Keyword>calcium</Keyword></KeywordGroup></ArticleHeader><NoBody></NoBody></Article></Issue></Volume></Journal></Publisher></istex:document></istex:metadataXml><mods version="3.6"><titleInfo lang="en"><title>Histone H1-Mediated Transfection: Serum Inhibition Can Be Overcome by Ca2+ Ions</title></titleInfo><titleInfo type="alternative" contentType="CDATA"><title>Histone H1-Mediated Transfection: Serum Inhibition Can Be Overcome by Ca2+ Ions</title></titleInfo><name type="personal"><namePart type="given">Annekathrin</namePart><namePart type="family">Haberland</namePart><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Thomas</namePart><namePart type="family">Knaus</namePart><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Sergei</namePart><namePart type="given">V.</namePart><namePart type="family">Zaitsev</namePart><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation><affiliation>Institute of Cytology of the Russian Academy of Sciences, St. Petersburg, Russia</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Bernd</namePart><namePart type="family">Buchberger</namePart><affiliation>Roche Diagnostics GmbH, Penzberg, Germany</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Andreas</namePart><namePart type="family">Lun</namePart><affiliation>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Hermann</namePart><namePart type="family">Haller</namePart><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">Michael</namePart><namePart type="family">Böttger</namePart><affiliation>Max Delbrück Center for Molecular Medicine, Franz Volhard Clinic, Berlin-Buch, Germany</affiliation><affiliation>Institute of Laboratory Medicine and Pathobiochemistry, Charité, Humboldt University, Berlin, Germany</affiliation><role><roleTerm type="text">author</roleTerm></role></name><typeOfResource>text</typeOfResource><genre type="research-article" displayLabel="OriginalPaper" 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>Kluwer Academic Publishers-Plenum Publishers</publisher><place><placeTerm type="text">New York</placeTerm></place><dateIssued encoding="w3cdtf">2000-02-01</dateIssued><copyrightDate encoding="w3cdtf">2000</copyrightDate></originInfo><language><languageTerm type="code" authority="rfc3066">en</languageTerm><languageTerm type="code" authority="iso639-2b">eng</languageTerm></language><abstract lang="en">Abstract: Purpose. One of the drawbacks of polycationic and cationic liposomalgene transfer is its sensitivity to serum. Gene therapy requires thetransfectant-DNA complex to be resistant to serum as well as blood.Since Ca2+ has proved to be an efficient cofactor of polycationic genetransfer, we decided to investigate its effects on transfection in thepresence of serum. Methods. We studied transgene expression of luciferase gene (pCMVLuc) on ECV 304 human endothelial cells using H1 histone andDOSPER as transfectants in the presence of 0-100% fetal calf serum. Results. H1-and DOSPER-mediated transfection was found to beinhibited by serum above the concentration of 10%. If 2 mM Ca2+ or2 mM Ca2+/0.1 mM chloroquine was included in the culture mediumwhich replace the transfection mixture and was left on the cells for24 hours postincubation, the inhibiting effect of even 100% serumwas overcome. Conclusions. A high serum level does not interfere with binding anduptake of H1- and DOSPER-DNA complexes, but inhibits subsequentsteps such as endosomal escape. Ca2+ in the form of nascent calciumphosphate microprecipitates and other lysosomolytical agents facilitateendosomal/lysosomal release by their fusigenic and membranolyticactivity.</abstract><subject lang="en"><topic>transfection</topic><topic>serum inhibition</topic><topic>H1 histone</topic><topic>DOSPER</topic><topic>calcium</topic></subject><relatedItem type="host"><titleInfo><title>Pharmaceutical Research</title><subTitle>An Official Journal of the American Association of Pharmaceutical Scientists</subTitle></titleInfo><titleInfo type="abbreviated"><title>Pharm Res</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>Springer</publisher><dateIssued encoding="w3cdtf">2000-02-01</dateIssued><copyrightDate encoding="w3cdtf">2000</copyrightDate></originInfo><subject><genre>Biomedicine</genre><topic>Pharmacology/Toxicology</topic><topic>Pharmacy</topic><topic>Biochemistry, general</topic><topic>Medical Law</topic><topic>Biomedical Engineering</topic></subject><identifier type="ISSN">0724-8741</identifier><identifier type="eISSN">1573-904X</identifier><identifier type="JournalID">11095</identifier><identifier type="IssueArticleCount">22</identifier><identifier type="VolumeIssueCount">12</identifier><part><date>2000</date><detail type="volume"><number>17</number><caption>vol.</caption></detail><detail type="issue"><number>2</number><caption>no.</caption></detail><extent unit="pages"><start>229</start><end>235</end></extent></part><recordInfo><recordOrigin>Plenum Publishing Corporation, 2000</recordOrigin></recordInfo></relatedItem><identifier type="istex">5E39256AF6D067F01D2FA923B6BCC3574FA0A9AD</identifier><identifier type="ark">ark:/67375/VQC-Z8N2MRJ5-4</identifier><identifier type="DOI">10.1023/A:1007581700996</identifier><identifier type="ArticleID">221578</identifier><identifier type="ArticleID">Art19</identifier><accessCondition type="use and reproduction" contentType="copyright">Plenum Publishing Corporation, 2000</accessCondition><recordInfo><recordContentSource authority="ISTEX" authorityURI="https://loaded-corpus.data.istex.fr" valueURI="https://loaded-corpus.data.istex.fr/ark:/67375/XBH-3XSW68JL-F">springer</recordContentSource><recordOrigin>Plenum Publishing Corporation, 2000</recordOrigin></recordInfo></mods><json:item><extension>json</extension><original>false</original><mimetype>application/json</mimetype><uri>https://api.istex.fr/ark:/67375/VQC-Z8N2MRJ5-4/record.json</uri></json:item></metadata><serie></serie></istex></record>Pour manipuler ce document sous Unix (Dilib)
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