| Blebbistatin extends culture life of adult mouse cardiac myocytes and allows efficient and stable transgene expression. | |
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MedLine Citation:
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PMID: 18296569 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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The characterization of cellular phenotypes of heart disorders can be achieved by isolating cardiac myocytes from mouse models or genetically modifying wild-type cells in culture. However, adult mouse cardiac myocytes show extremely low tolerance to isolation and primary culture conditions. Previous studies indicate that 2,3-butanedione monoximine (BDM), a nonspecific excitation-contraction coupling inhibitor, can improve the viability of isolated adult mouse cardiac myocytes. The mechanisms of the beneficial and unwanted nonspecific actions of BDM on cardiac myocytes are not understood. To understand what contributes to murine adult cardiac myocyte stability in primary culture and improve this model system for experimental use, the specific myosin II inhibitor blebbistatin was explored as a media supplement to inhibit mouse myocyte contraction. Enzymatically isolated adult mouse cardiac myocytes were cultured with blebbistatin or BDM as a media supplement. Micromolar concentrations of blebbistatin significantly increased the viability, membrane integrity, and morphology of adult cardiac myocytes compared with cells treated with previously described 10 mM BDM. Cells treated with blebbistatin also showed efficient adenovirus gene transfer and stable transgene expression, and unlike BDM, blebbistatin does not appear to interfere with cell adhesion. Higher concentrations of BDM actually worsened myocyte membrane integrity and transgene expression. Therefore, the specific inhibition of myosin II activity by blebbistatin has significant beneficial effects on the long-term viability of adult mouse cardiac myocytes. Furthermore, the unwanted effects of BDM on adult mouse cardiac myocytes, perhaps due to its nonspecific activities or action as a chemical phosphatase, can be avoided by using blebbistatin. |
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Authors:
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Zhyldyz Kabaeva; Mei Zhao; Daniel E Michele |
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Publication Detail:
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Type: Comparative Study; Journal Article Date: 2008-02-22 |
Journal Detail:
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Title: American journal of physiology. Heart and circulatory physiology Volume: 294 ISSN: 0363-6135 ISO Abbreviation: Am. J. Physiol. Heart Circ. Physiol. Publication Date: 2008 Apr |
Date Detail:
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Created Date: 2008-04-02 Completed Date: 2008-05-29 Revised Date: 2010-05-20 |
Medline Journal Info:
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Nlm Unique ID: 100901228 Medline TA: Am J Physiol Heart Circ Physiol Country: United States |
Other Details:
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Languages: eng Pagination: H1667-74 Citation Subset: IM |
Affiliation:
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Dept. of Molecular and Integrative Physiology, University of Michigan, 7623A Medical Science II, Ann Arbor, MI 48109-0622, USA. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Adenoviridae
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genetics Animals Cell Adhesion / drug effects Cell Culture Techniques Cell Membrane / drug effects Cell Shape / drug effects Cell Survival / drug effects Cells, Cultured Diacetyl / analogs & derivatives*, pharmacology, toxicity Dose-Response Relationship, Drug Enzyme Inhibitors / pharmacology*, toxicity Genetic Vectors Green Fluorescent Proteins / biosynthesis, genetics Heterocyclic Compounds with 4 or More Rings / pharmacology* Mice Mice, Inbred C57BL Myocardial Contraction / drug effects* Myocytes, Cardiac / drug effects*, enzymology, metabolism Myosin Type II / antagonists & inhibitors*, metabolism Time Factors Transfection* Transgenes* |
| Grant Support | |
ID/Acronym/Agency:
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R01 HL080388-05/HL/NHLBI NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Enzyme Inhibitors; 0/Heterocyclic Compounds with 4 or More Rings; 0/blebbistatin; 147336-22-9/Green Fluorescent Proteins; 431-03-8/Diacetyl; 57-71-6/diacetylmonoxime; EC 3.6.1.-/Myosin Type II |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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