دورية أكاديمية
Generation and customization of biosynthetic excitable tissues for electrophysiological studies and cell-based therapies.
العنوان: | Generation and customization of biosynthetic excitable tissues for electrophysiological studies and cell-based therapies. |
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المؤلفون: | Nguyen HX; Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA., Kirkton RD; Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA., Bursac N; Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA. |
المصدر: | Nature protocols [Nat Protoc] 2018 May; Vol. 13 (5), pp. 927-945. Date of Electronic Publication: 2018 Apr 05. |
نوع المنشور: | Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S. |
اللغة: | English |
بيانات الدورية: | Publisher: Nature Pub. Group Country of Publication: England NLM ID: 101284307 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1750-2799 (Electronic) Linking ISSN: 17502799 NLM ISO Abbreviation: Nat Protoc Subsets: MEDLINE |
أسماء مطبوعة: | Original Publication: London, UK : Nature Pub. Group, 2006- |
مواضيع طبية MeSH: | Action Potentials* , Electric Stimulation*, Cell Engineering/*methods , Ion Channels/*metabolism , Mutant Proteins/*metabolism, Cells, Cultured ; Fibroblasts/physiology ; Humans ; Ion Channels/genetics ; Mutant Proteins/genetics ; Patch-Clamp Techniques |
مستخلص: | We describe a two-stage protocol to generate electrically excitable and actively conducting cell networks with stable and customizable electrophysiological phenotypes. Using this method, we have engineered monoclonally derived excitable tissues as a robust and reproducible platform to investigate how specific ion channels and mutations affect action potential (AP) shape and conduction. In the first stage of the protocol, we combine computational modeling, site-directed mutagenesis, and electrophysiological techniques to derive optimal sets of mammalian and/or prokaryotic ion channels that produce specific AP shape and conduction characteristics. In the second stage of the protocol, selected ion channels are stably expressed in unexcitable human cells by means of viral or nonviral delivery, followed by flow cytometry or antibiotic selection to purify the desired phenotype. This protocol can be used with traditional heterologous expression systems or primary excitable cells, and application of this method to primary fibroblasts may enable an alternative approach to cardiac cell therapy. Compared with existing methods, this protocol generates a well-defined, relatively homogeneous electrophysiological phenotype of excitable cells that facilitates experimental and computational studies of AP conduction and can decrease arrhythmogenic risk upon cell transplantation. Although basic cell culture and molecular biology techniques are sufficient to generate excitable tissues using the described protocol, experience with patch-clamp techniques is required to characterize and optimize derived cell populations. |
References: | Nature. 2001 Feb 22;409(6823):1043-7. (PMID: 11234013) J Exp Med. 1953 May;97(5):695-710. (PMID: 13052828) PLoS One. 2013;8(2):e55400. (PMID: 23393574) Nat Protoc. 2006;1(3):1235-47. (PMID: 17406407) Circ Res. 2015 Jun 5;116(12):1989-2004. (PMID: 26044252) PLoS One. 2011;6(4):e18556. (PMID: 21602908) PLoS One. 2013 Dec 31;8(12):e85221. (PMID: 24391999) Curr Top Membr. 2016;78:39-64. (PMID: 27586280) J Neurosci. 2013 May 1;33(18):7928-40. (PMID: 23637184) Nature. 2007 Dec 6;450(7171):819-24. (PMID: 18064002) Physiol Rev. 2004 Apr;84(2):431-88. (PMID: 15044680) J Physiol. 1962 Feb;160:317-52. (PMID: 14480151) Nat Protoc. 2007;2(11):2826-41. (PMID: 18007618) Elife. 2016 May 24;5:. (PMID: 27215841) J Theor Biol. 2008 Aug 7;253(3):544-60. (PMID: 18495166) Cell. 2010 Apr 2;141(1):154-165. (PMID: 20303157) PLoS Comput Biol. 2017 Jan 20;13(1):e1005342. (PMID: 28107358) J Physiol. 1996 Nov 1;496 ( Pt 3):647-62. (PMID: 8930833) Prog Biophys Mol Biol. 2006 Jan-Apr;90(1-3):26-37. (PMID: 15979128) Biophys J. 2004 Sep;87(3):1507-25. (PMID: 15345532) Methods Mol Biol. 2001;154:269-92. (PMID: 11218653) Nature. 2016 Oct 20;538(7625):388-391. (PMID: 27723741) Nat Commun. 2011;2:300. (PMID: 21556054) Proc Natl Acad Sci U S A. 2008 Feb 26;105(8):2883-8. (PMID: 18287077) Nature. 2006 Mar 23;440(7083):440-7. (PMID: 16554803) Nature. 2011 Mar 10;471(7337):225-9. (PMID: 21240260) Chaos. 2002 Sep;12(3):852-892. (PMID: 12779613) Biophys J. 2009 May 6;96(9):3873-85. (PMID: 19413993) J Physiol. 1991 May;436:579-601. (PMID: 1712040) Proc R Soc Med. 1946 Dec 3;134(873):444-79. (PMID: 20281590) J Biol Chem. 2004 Mar 5;279(10):9532-8. (PMID: 14665618) Circ Res. 2003 Sep 5;93(5):421-8. (PMID: 12893743) Am J Physiol Heart Circ Physiol. 2004 Apr;286(4):H1573-89. (PMID: 14656705) Science. 1995 Jul 7;269(5220):92-5. (PMID: 7604285) Physiol Rev. 2005 Oct;85(4):1205-53. (PMID: 16183911) Europace. 2012 Nov;14 Suppl 5:v40-v49. (PMID: 23104914) Science. 2016 Dec 9;354(6317):1296-1301. (PMID: 27940875) Biotechnol Lett. 2009 Mar;31(3):353-9. (PMID: 19034387) Ann Biomed Eng. 2012 May;40(5):1006-17. (PMID: 22124794) Circ Arrhythm Electrophysiol. 2013 Dec;6(6):1200-7. (PMID: 24235268) Nat Commun. 2016 Oct 18;7:13132. (PMID: 27752065) Circulation. 2008 Feb 12;117(6):720-31. (PMID: 18212286) Proc R Soc Lond B Biol Sci. 1952 Oct 16;140(899):177-83. (PMID: 13003922) Biotechnol J. 2015 Apr;10(4):647-53. (PMID: 25650551) Circ Res. 1998 Jan 9-23;82(1):63-81. (PMID: 9440706) Science. 2001 Dec 14;294(5550):2372-5. (PMID: 11743207) J Gen Virol. 1977 Jul;36(1):59-74. (PMID: 886304) Nature. 2011 Mar 10;471(7337):230-4. (PMID: 21307850) J Mol Cell Cardiol. 2010 Jan;48(1):2-11. (PMID: 19744495) Hum Gene Ther. 2015 Jul;26(7):432-42. (PMID: 26086867) IEEE Eng Med Biol Mag. 2008 Jan-Feb;27(1):72-80. (PMID: 18270054) FASEB J. 2002 Jun;16(8):869-71. (PMID: 11967234) Cardiovasc Res. 2012 Feb 1;93(2):263-71. (PMID: 22072633) Cardiovasc Res. 2011 Mar 1;89(4):776-85. (PMID: 21159668) Cell. 2007 Nov 30;131(5):861-72. (PMID: 18035408) Med Biol Eng Comput. 2006 Jul;44(7):537-42. (PMID: 16937189) Nat Genet. 2009 Jun;41(6):753-61. (PMID: 19412179) Circ Arrhythm Electrophysiol. 2009 Apr;2(2):185-94. (PMID: 19808464) Biochem J. 1999 Oct 1;343 Pt 1:39-44. (PMID: 10493909) Mol Ther. 2011 Jul;19(7):1220-9. (PMID: 21505421) Nat Biotechnol. 2011 Jul 31;29(8):735-41. (PMID: 21804562) Neuron. 2009 Apr 30;62(2):191-8. (PMID: 19409264) Circ Res. 2005 Jul 22;97(2):159-67. (PMID: 15976318) Biophys J. 2001 May;80(5):2231-47. (PMID: 11325726) J Pharmacol Toxicol Methods. 2005 May-Jun;51(3):187-200. (PMID: 15862464) J Biol Chem. 2010 Feb 5;285(6):3685-3694. (PMID: 19959480) Mol Cell Biochem. 2002 Sep;238(1-2):69-79. (PMID: 12349911) Am J Physiol Heart Circ Physiol. 2007 Jan;292(1):H399-407. (PMID: 16877553) J Neurosci. 2006 Mar 8;26(10):2714-23. (PMID: 16525050) Chaos. 1998 Dec;8(4):879. (PMID: 12779795) Br J Pharmacol. 1996 Jun;118(3):748-54. (PMID: 8762103) Methods Mol Biol. 2014;1181:249-62. (PMID: 25070342) Neuron. 1994 Jun;12(6):1183-94. (PMID: 7516685) Nature. 2014 Jun 12;510(7504):273-7. (PMID: 24776797) |
معلومات مُعتمدة: | R01 HL132389 United States HL NHLBI NIH HHS; U01 HL134764 United States HL NHLBI NIH HHS; R21 HL095069 United States HL NHLBI NIH HHS; R21 HL126193 United States HL NHLBI NIH HHS; R21 HL106203 United States HL NHLBI NIH HHS; R21 HL083342 United States HL NHLBI NIH HHS; R01 HL126524 United States HL NHLBI NIH HHS; R01 HL104326 United States HL NHLBI NIH HHS |
المشرفين على المادة: | 0 (Ion Channels) 0 (Mutant Proteins) |
تواريخ الأحداث: | Date Created: 20180407 Date Completed: 20190415 Latest Revision: 20240610 |
رمز التحديث: | 20240610 |
مُعرف محوري في PubMed: | PMC6050172 |
DOI: | 10.1038/nprot.2018.016 |
PMID: | 29622805 |
قاعدة البيانات: | MEDLINE |
تدمد: | 1750-2799 |
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DOI: | 10.1038/nprot.2018.016 |