Methods for Lymphatic Vessel Culture and Gene Transfection
Identifieur interne : 002929 ( Pmc/Curation ); précédent : 002928; suivant : 002930Methods for Lymphatic Vessel Culture and Gene Transfection
Auteurs : A. A. Gashev [États-Unis] ; M. J. Davis [États-Unis] ; O. Yu. Gasheva [États-Unis] ; Z. V. Nepiushchikh [États-Unis] ; W. Wang [États-Unis] ; P. Dougherty [États-Unis] ; K. A. Kelly [États-Unis] ; S. Cai [Royaume-Uni] ; P.-Y. Von Der Weid [Canada] ; M. Muthuchamy [États-Unis] ; C. J. Meininger [États-Unis] ; D. C. Zawieja [États-Unis]Source :
- Microcirculation (New York, N.Y. : 1994) [ 1073-9688 ] ; 2009.
Abstract
To develop the techniques needed for the specific gene/protein targeting transfection experiments in isolated lymphatic vessels, we completed two major tasks: 1) optimize the experimental conditions to maintain the viability of isolated rat lymphatic vessels in culture for sufficiently long periods of time to permit knockdown or overexpression of selected proteins/genes; 2) develop effective transfection protocols for lymphatic muscle and endothelial cells in intact lymphatic vessels without nonspecific impairment of lymphatic contractile function due to the transfection protocol itself.
Experimental protocols were developed for the maintenance of isolated lymphatic vessels under non-pressurized and pressurized conditions for 3-12 days in culture and for adenoviral gene transfection of the lymphatic muscle and endothelial cells.
The data demonstrate the effectiveness of the newly developed experimental protocols for the maintenance of isolated rat mesenteric lymphatic vessels and thoracic duct in culture up to 3-12 days without significant impairment of the parameters of their pumping and effective adenoviral/GFP transfection of lymphatic endothelial and muscle cells in isolated rat mesenteric lymphatic vessels.
These experimental techniques will extend the set of the modern experimental tools available to researchers investigating the physiology of lymphatic function.
Url:
DOI: 10.1080/10739680903120778
PubMed: 19626551
PubMed Central: 3042427
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Kelly</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Cai, S" sort="Cai, S" uniqKey="Cai S" first="S." last="Cai">S. 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Muthuchamy</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Meininger, C J" sort="Meininger, C J" uniqKey="Meininger C" first="C. J." last="Meininger">C. J. 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A." last="Gashev">A. A. Gashev</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Davis, M J" sort="Davis, M J" uniqKey="Davis M" first="M. J." last="Davis">M. J. Davis</name><affiliation wicri:level="1"><nlm:aff id="A2">Department of Medical Pharmacology & Physiology, University of Missouri, Columbia, Missouri, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Medical Pharmacology & Physiology, University of Missouri, Columbia, Missouri</wicri:regionArea></affiliation></author><author><name sortKey="Gasheva, O Yu" sort="Gasheva, O Yu" uniqKey="Gasheva O" first="O. Yu." last="Gasheva">O. Yu. Gasheva</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Nepiushchikh, Z V" sort="Nepiushchikh, Z V" uniqKey="Nepiushchikh Z" first="Z. V." last="Nepiushchikh">Z. V. Nepiushchikh</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Wang, W" sort="Wang, W" uniqKey="Wang W" first="W." last="Wang">W. Wang</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Dougherty, P" sort="Dougherty, P" uniqKey="Dougherty P" first="P." last="Dougherty">P. Dougherty</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Kelly, K A" sort="Kelly, K A" uniqKey="Kelly K" first="K. A." last="Kelly">K. A. Kelly</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Cai, S" sort="Cai, S" uniqKey="Cai S" first="S." last="Cai">S. Cai</name><affiliation wicri:level="1"><nlm:aff id="A3">Department of Clinical Pharmacology, University of Oxford, Oxford, UK</nlm:aff><country xml:lang="fr">Royaume-Uni</country><wicri:regionArea>Department of Clinical Pharmacology, University of Oxford, Oxford</wicri:regionArea></affiliation></author><author><name sortKey="Von Der Weid, P Y" sort="Von Der Weid, P Y" uniqKey="Von Der Weid P" first="P.-Y." last="Von Der Weid">P.-Y. Von Der Weid</name><affiliation wicri:level="1"><nlm:aff id="A4">Department of Physiology & Biophysics, University of Calgary, Calgary, Alberta, Canada.</nlm:aff><country xml:lang="fr" wicri:curation="lc">Canada</country><wicri:regionArea>Department of Physiology & Biophysics, University of Calgary, Calgary, Alberta</wicri:regionArea></affiliation></author><author><name sortKey="Muthuchamy, M" sort="Muthuchamy, M" uniqKey="Muthuchamy M" first="M." last="Muthuchamy">M. Muthuchamy</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Meininger, C J" sort="Meininger, C J" uniqKey="Meininger C" first="C. J." last="Meininger">C. J. Meininger</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author><author><name sortKey="Zawieja, D C" sort="Zawieja, D C" uniqKey="Zawieja D" first="D. C." last="Zawieja">D. C. Zawieja</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas</wicri:regionArea></affiliation></author></analytic><series><title level="j">Microcirculation (New York, N.Y. : 1994)</title><idno type="ISSN">1073-9688</idno><idno type="eISSN">1549-8719</idno><imprint><date when="2009">2009</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><sec id="S1"><title>Objective</title><p id="P1">To develop the techniques needed for the specific gene/protein targeting transfection experiments in isolated lymphatic vessels, we completed two major tasks: 1) optimize the experimental conditions to maintain the viability of isolated rat lymphatic vessels in culture for sufficiently long periods of time to permit knockdown or overexpression of selected proteins/genes; 2) develop effective transfection protocols for lymphatic muscle and endothelial cells in intact lymphatic vessels without nonspecific impairment of lymphatic contractile function due to the transfection protocol itself.</p></sec><sec sec-type="methods" id="S2"><title>Methods</title><p id="P2">Experimental protocols were developed for the maintenance of isolated lymphatic vessels under non-pressurized and pressurized conditions for 3-12 days in culture and for adenoviral gene transfection of the lymphatic muscle and endothelial cells.</p></sec><sec id="S3"><title>Results</title><p id="P3">The data demonstrate the effectiveness of the newly developed experimental protocols for the maintenance of isolated rat mesenteric lymphatic vessels and thoracic duct in culture up to 3-12 days without significant impairment of the parameters of their pumping and effective adenoviral/GFP transfection of lymphatic endothelial and muscle cells in isolated rat mesenteric lymphatic vessels.</p></sec><sec id="S4"><title>Conclusions</title><p id="P4">These experimental techniques will extend the set of the modern experimental tools available to researchers investigating the physiology of lymphatic function.</p></sec></div></front></TEI><pmc article-type="research-article" xml:lang="EN"><pmc-comment>The publisher of this article does not allow downloading of the full text in XML form.</pmc-comment>
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<front><journal-meta><journal-id journal-id-type="nlm-journal-id">9434935</journal-id><journal-id journal-id-type="pubmed-jr-id">8801</journal-id><journal-id journal-id-type="nlm-ta">Microcirculation</journal-id><journal-title>Microcirculation (New York, N.Y. : 1994)</journal-title><issn pub-type="ppub">1073-9688</issn><issn pub-type="epub">1549-8719</issn></journal-meta><article-meta><article-id pub-id-type="pmid">19626551</article-id><article-id pub-id-type="pmc">3042427</article-id><article-id pub-id-type="doi">10.1080/10739680903120778</article-id><article-id pub-id-type="manuscript">NIHMS271473</article-id><article-categories><subj-group subj-group-type="heading"><subject>Article</subject></subj-group></article-categories><title-group><article-title>Methods for Lymphatic Vessel Culture and Gene Transfection</article-title></title-group><contrib-group><contrib contrib-type="author"><name><surname>Gashev</surname><given-names>A.A.</given-names></name><xref ref-type="aff" rid="A1">1</xref><xref ref-type="corresp" rid="CR1">*</xref></contrib><contrib contrib-type="author"><name><surname>Davis</surname><given-names>M.J.</given-names></name><xref ref-type="aff" rid="A2">2</xref></contrib><contrib contrib-type="author"><name><surname>Gasheva</surname><given-names>O.Yu.</given-names></name><xref ref-type="aff" rid="A1">1</xref></contrib><contrib contrib-type="author"><name><surname>Nepiushchikh</surname><given-names>Z.V.</given-names></name><xref ref-type="aff" rid="A1">1</xref></contrib><contrib contrib-type="author"><name><surname>Wang</surname><given-names>W.</given-names></name><xref ref-type="aff" rid="A1">1</xref></contrib><contrib contrib-type="author"><name><surname>Dougherty</surname><given-names>P.</given-names></name><xref ref-type="aff" rid="A1">1</xref></contrib><contrib contrib-type="author"><name><surname>Kelly</surname><given-names>K.A.</given-names></name><xref ref-type="aff" rid="A1">1</xref></contrib><contrib contrib-type="author"><name><surname>Cai</surname><given-names>S.</given-names></name><xref ref-type="aff" rid="A3">3</xref></contrib><contrib contrib-type="author"><name><surname>von der Weid</surname><given-names>P.-Y.</given-names></name><xref ref-type="aff" rid="A4">4</xref></contrib><contrib contrib-type="author"><name><surname>Muthuchamy</surname><given-names>M.</given-names></name><xref ref-type="aff" rid="A1">1</xref></contrib><contrib contrib-type="author"><name><surname>Meininger</surname><given-names>C.J.</given-names></name><xref ref-type="aff" rid="A1">1</xref></contrib><contrib contrib-type="author"><name><surname>Zawieja</surname><given-names>D.C.</given-names></name><xref ref-type="aff" rid="A1">1</xref></contrib></contrib-group><aff id="A1"><label>1</label>Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, Temple/College Station, Texas, USA</aff><aff id="A2"><label>2</label>Department of Medical Pharmacology & Physiology, University of Missouri, Columbia, Missouri, USA</aff><aff id="A3"><label>3</label>Department of Clinical Pharmacology, University of Oxford, Oxford, UK</aff><aff id="A4"><label>4</label>Department of Physiology & Biophysics, University of Calgary, Calgary, Alberta, Canada.</aff><author-notes><corresp id="CR1"><label>*</label><bold>Corresponding Author:</bold> Department of Systems Biology and Translational Medicine, College of Medicine, Cardiovascular Research Institute Division of Lymphatic Biology, Texas A&M Health Science Center, 702 SW H.K. Dodgen Loop, Temple, TX 76504 USA, Phone: 1-254-742-7147; Fax: 1-254-742-7145; <email>gashev@tamu.edu</email></corresp></author-notes><pub-date pub-type="nihms-submitted"><day>9</day><month>2</month><year>2011</year></pub-date><pub-date pub-type="ppub"><month>10</month><year>2009</year></pub-date><pub-date pub-type="pmc-release"><day>21</day><month>2</month><year>2011</year></pub-date><volume>16</volume><issue>7</issue><fpage>615</fpage><lpage>628</lpage><abstract><sec id="S1"><title>Objective</title><p id="P1">To develop the techniques needed for the specific gene/protein targeting transfection experiments in isolated lymphatic vessels, we completed two major tasks: 1) optimize the experimental conditions to maintain the viability of isolated rat lymphatic vessels in culture for sufficiently long periods of time to permit knockdown or overexpression of selected proteins/genes; 2) develop effective transfection protocols for lymphatic muscle and endothelial cells in intact lymphatic vessels without nonspecific impairment of lymphatic contractile function due to the transfection protocol itself.</p></sec><sec sec-type="methods" id="S2"><title>Methods</title><p id="P2">Experimental protocols were developed for the maintenance of isolated lymphatic vessels under non-pressurized and pressurized conditions for 3-12 days in culture and for adenoviral gene transfection of the lymphatic muscle and endothelial cells.</p></sec><sec id="S3"><title>Results</title><p id="P3">The data demonstrate the effectiveness of the newly developed experimental protocols for the maintenance of isolated rat mesenteric lymphatic vessels and thoracic duct in culture up to 3-12 days without significant impairment of the parameters of their pumping and effective adenoviral/GFP transfection of lymphatic endothelial and muscle cells in isolated rat mesenteric lymphatic vessels.</p></sec><sec id="S4"><title>Conclusions</title><p id="P4">These experimental techniques will extend the set of the modern experimental tools available to researchers investigating the physiology of lymphatic function.</p></sec></abstract><kwd-group><kwd>lymphatic vessel</kwd><kwd>tissue culture</kwd><kwd>transportation</kwd><kwd>adenoviral transfection</kwd></kwd-group><contract-num rid="HL1">R21 HL085659-01A2
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