Name: isolation of atrial cardiomyocytes from a rat model of metabolic syndrome-related heart failure with preserved ejection fraction
Uploaded: 2018-07-26T14:00:00
Description: Watch this Scientific Journal Video about Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction at JoVE.com

Denne forskning blev støttet af DZHK (tyske Center for hjerte-kar-forskning, D.B.), EKFS (Else-Kröner-Fresenius Stiftung, F.H.), og af BMBF (tysk Ministeriet for uddannelse og forskning) samt BIH-Charité kliniske videnskabsmand program finansieret ved Charité - Universitätsmedizin Berlin, og Berlin Institute of Health (F.H.).

Alle eksperimenter blev godkendt af det lokale etiske udvalg (TVA T0060/15 og T0003-15) og udført i overensstemmelse med retningslinjerne for pleje og anvendelse af forsøgsdyr (National Institute of Health, U.S.A.).
Bemærk: En forenklet rutediagram af proceduren er vist i figur 1.
1. prearrangements
<ol><li>Forberede buffere i henhold til tabel 1.</li>
</ol><table border="1" fo:keep-together.within-page="1" fo:keep...

<ol>
	<li>Alberti, K. G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Harmonizing+the+metabolic+syndrome:+a+joint+interim+statement+of+the+International+Diabetes+Federation+Task+Force+on+Epidemiology+and+Prevention;+National+Heart,+Lung,+and+Blood+Institute;+American+Heart+Association;+World+Heart+Federation;+International+Atherosclerosis+Society;+and+International+Association+for+the+Study+of+Obesity.">Harmonizing the metabolic syndrome: a joint interim statement of the International Diabetes Federation Task Force on Epidemiology and Prevention; National Heart, Lung, and Blood Institute; American Heart Association; World Heart Federation; International Atherosclerosis Society; and International Association for the Study of Obesity.</a> Circulation. 120 (16), 1640-1645 (2009).</li><li>. IDF Consensus Worldwide Definition of the Metabolic Syndrome Available from: <a target="_blank" href="https://www.idf.org/e-library/consensus-statements/60-idfconsensus-worldwide-definitionof-the-metabolic-syndrome.html">https://www.idf.org/e-library/consensus-statements/60-idfconsensus-worldwide-definitionof-the-metabolic-syndrome.html</a> (2006)</li><li>Melenovsky, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Left+atrial+remodeling+and+function+in+advanced+heart+failure+with+preserved+or+reduced+ejection+fraction.">Left atrial remodeling and function in advanced heart failure with preserved or reduced ejection fraction.</a> Circulation: Heart Failure. 8 (2), 295-303 (2015).</li><li>Goette, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=EHRA/HRS/APHRS/SOLAECE+expert+consensus+on+atrial+cardiomyopathies:+definition,+characterization,+and+clinical+implication.">EHRA/HRS/APHRS/SOLAECE expert consensus on atrial cardiomyopathies: definition, characterization, and clinical implication.</a> EP Europace. 18 (10), 1455-1490 (2016).</li><li>Schotten, U., Verheule, S., Kirchhof, P., Goette, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pathophysiological+mechanisms+of+atrial+fibrillation:+a+translational+appraisal.">Pathophysiological mechanisms of atrial fibrillation: a translational appraisal.</a> Physiological Reviews. 91 (1), 265-325 (2011).</li><li>Hohendanner, F., DeSantiago, J., Heinzel, F. R., Blatter, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dyssynchronous+calcium+removal+in+heart+failure-induced+atrial+remodeling.">Dyssynchronous calcium removal in heart failure-induced atrial remodeling.</a> American Journal of Physiology-Heart and Circulatory Physiology. 311 (6), H1352-H1359 (2016).</li><li>Hohendanner, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inositol-1,4,5-trisphosphate+induced+Ca2++release+and+excitation-contraction+coupling+in+atrial+myocytes+from+normal+and+failing+hearts.">Inositol-1,4,5-trisphosphate induced Ca2+ release and excitation-contraction coupling in atrial myocytes from normal and failing hearts.</a> The Journal of Physiology. 593 (6), 1459-1477 (2015).</li><li>Tada, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+mineralocorticoid+receptor+on+experimental+cerebral+aneurysms+in+rats.">Role of mineralocorticoid receptor on experimental cerebral aneurysms in rats.</a> Hypertension. 54 (3), 552-557 (2009).</li><li>Iwasaki, Y. 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E., Sheehan, K. A., Wolska, B. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Methods+in+cardiomyocyte+isolation,+culture,+and+gene+transfer.">Methods in cardiomyocyte isolation, culture, and gene transfer.</a> Journal of Molecular and Cellular Cardiology. 51 (3), 288-298 (2011).</li><li>Gunduz, D., Hamm, C. W., Aslam, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Simultaneous+Isolation+of+High+Quality+Cardiomyocytes,+Endothelial+Cells,+and+Fibroblasts+from+an+Adult+Rat+Heart.">Simultaneous Isolation of High Quality Cardiomyocytes, Endothelial Cells, and Fibroblasts from an Adult Rat Heart.</a> Journal of Visualized Experiments. (123), e55601 (2017).</li><li>Li, D., Wu, J., Bai, Y., Zhao, X., Liu, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+culture+of+adult+mouse+cardiomyocytes+for+cell+signaling+and+in+vitro+cardiac+hypertrophy.">Isolation and culture of adult mouse cardiomyocytes for cell signaling and in vitro cardiac hypertrophy.</a> Journal of Visualized Experiments. (87), e51357 (2014).</li><li>Graham, E. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation,+culture,+and+functional+characterization+of+adult+mouse+cardiomyoctyes.">Isolation, culture, and functional characterization of adult mouse cardiomyoctyes.</a> Journal of Visualized Experiments. (79), e50289 (2013).</li><li>Roth, G. M., Bader, D. M., Pfaltzgraff, E. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+physiological+analysis+of+mouse+cardiomyocytes.">Isolation and physiological analysis of mouse cardiomyocytes.</a> Journal of Visualized Experiments. (91), e51109 (2014).</li><li>Thum, T., Borlak, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+cultivation+of+Ca2++tolerant+cardiomyocytes+from+the+adult+rat:+improvements+and+applications.">Isolation and cultivation of Ca2+ tolerant cardiomyocytes from the adult rat: improvements and applications.</a> Xenobiotica. 30 (11), 1063-1077 (2000).</li><li>Egorova, M. V., Afanas'ev, S. A., Popov, S. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+method+for+isolation+of+cardiomyocytes+from+adult+rat+heart.">A simple method for isolation of cardiomyocytes from adult rat heart.</a> Bulletin of Experimental Biology and Medicine. 140 (3), 370-373 (2005).</li><li>Kohncke, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+Kv+channel+recordings+in+murine+atrial+and+ventricular+cardiomyocytes.">Isolation and Kv channel recordings in murine atrial and ventricular cardiomyocytes.</a> Journal of Visualized Experiments. (73), e50145 (2013).</li><li>Wagner, E., Brandenburg, S., Kohl, T., Lehnart, S. 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M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diastolic+dysfunction+and+left+atrial+volume:+a+population-based+study.">Diastolic dysfunction and left atrial volume: a population-based study.</a> Journal of the American College of Cardiology. 45 (1), 87-92 (2005).</li><li>Linz, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cathepsin+A+mediates+susceptibility+to+atrial+tachyarrhythmia+and+impairment+of+atrial+emptying+function+in+Zucker+diabetic+fatty+rats.">Cathepsin A mediates susceptibility to atrial tachyarrhythmia and impairment of atrial emptying function in Zucker diabetic fatty rats.</a> Cardiovascular Research. 110 (3), 371-380 (2016).</li><li>Ackers-Johnson, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Simplified,+Langendorff-Free+Method+for+Concomitant+Isolation+of+Viable+Cardiac+Myocytes+and+Nonmyocytes+From+the+Adult+Mouse+Heart.">A Simplified, Langendorff-Free Method for Concomitant Isolation of Viable Cardiac Myocytes and Nonmyocytes From the Adult Mouse Heart.</a> Circulation Research. 119 (8), 909-920 (2016).</li><li>Ramanathan, T., Skinner, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coronary+blood+flow.">Coronary blood flow.</a> Continuing Education in Anaesthesia Critical Care &amp; Pain. 5 (2), 61-64 (2005).</li><li>Bond, M. 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Her, beskrevet vi første gang en protokol til isolering af enkelt celle ACMs fra en rotte model af MetS-relaterede HFpEF der viser mærket atrieflimren remodeling22. Proceduren, der er entydigt udfordrende som overdreven fedtvæv kan gøre den kirurgiske forberedelse, samt cannulation af aorta, stadig vanskeligere. Guiden fejlfinding fastsat i tabel 2 adresser de mest almindelige spørgsmål af proceduren isolation.
<table border="1" fo:keep-together.within-page="1" fo:keep-w...

MetS beskriver en klynge af risikofaktorer for diabetes mellitus type-2 og hjerte-kar-sygdom og omfatter et øget arterielt blodtryk, dyslipidæmi (rejst triglycerider og sænket high-density lipoprotein kolesterol), øget fastende glukose, og centrale fedme1. Den verdensomspændende forekomsten af MetS skønnes for at være 25-30% og konstant stigende2. HFpEF er en heterogene klinisk syndrom ofte forbundet med MetS. Den kardiale remodeling under HFpEF og dens foregående faser (dvs., hypertensive hjertesygdomme) er også ledsaget af en ombygning af atria3. Reduceret kontraktile funktion og strukturelle ændringer af venstre atrium har været forbundet med øget dødelighed, atrieflimren og ny-debut hjertesvigt4. Atrieflimren remodellering er karakteriseret ved ændringer i ion-kanal funktion, Ca2 + homøostase, atrieflimren struktur, fibroblast aktivering og væv fibrose5. Venstre atriale remodeling i MetS-relaterede HFpEF og dens underliggende patologiske mekanismer er stadig dårligt forstået og kræver en yderligere grundig undersøgelse. Dyremodeller har vist sig at være et værdifuldt værktøj og føre til mange fremskridt inden for atrieflimren cardiomyopatier6,7,8,9.
Undersøgelser med isolerede encellede cardiomyocytes bruges ofte til at bekræfte og supplere i vivo resultater. En isolation, og den potentielle efterfølgende cellekultur, mulighed for undersøgelse af signalering veje, ionisk kanal strømninger og excitation sammentrækning kobling. Fysiologisk betingelser, cardiomyocytes ikke formere. Fusion mellem transkriptionel regulering sekvenser af en atrial natriuretic faktor og en simian 40 store T virusantigen i Transgene mus førte til oprettelsen af de første udødeliggjort ACMs, navnet på-110. Videreudvikling af i-1 celler gav anledning til HL-1 celler, der ikke kun være seriefremstillede passaged men også kontrakt spontant11. De viser imidlertid strukturelle og funktionelle forskelle i forhold til frisk isolerede celler, såsom en mindre organiseret ultrastruktur, en høj forekomst af udvikling af myofibrils11og en hyperpolarisering-aktiveret indad nuværende12. Isolering af ventrikulær cardiomyocytes (VCM) i rotter og mus fra en række forskellige modeller er veletableret13,14,15,16,17,18 , 19. generelt, skåret hjertet er monteret på en Langendorff apparatur og retrogradely perfunderet med en Ca2 +-gratis buffer indeholdende fordøjelsesenzymer, såsom collagenases og proteaser. Calcium er derefter genindført i en trinvis måde de fysiologiske forhold. Men selv om protokollerne dedikeret til isolering af ACMs er tilgængelige20,21, på grund af øget fibrose og pres-relaterede forskelle, deres anvendelighed i sygdomsmodeller med atrieflimren remodeling er begrænset.
I denne artikel, har vi implementeret en protokol til isolering af atrieflimren encellede cardiomyocytes fra dyr, der udviser atrieflimren remodeling (dvs, især for den ZFS1 rotte model for MetS-relaterede HFpEF)22. Eksisterende isolering protokoller blev optimeret og suppleret med en enkel, custom-made enhed til at kontrollere og ændre intraluminal presset af hjertets hulrum, fører til højere udbytter af morfologisk og funktionelt intakt cardiomyocytes. Følgende protokol giver forsker med en trinvis guide, en detaljeret beskrivelse af specialfremstillet udstyr, en liste over løsninger, samt en omfattende fejlfindingsvejledning.

<table border="0" cellpadding="0" cellspacing="0" width="935">
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">ZSF-1 Obese rat</td>
			<td style="width:292px;">Charles River Laboratories, Inc.</td>
			<td style="width:136px;"></td>
			<td style="width:227px;">21 weeks old</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Fine Iris Scissors</td>
			<td style="width:292px;">Fine Science Tools GmbH</td>
			<td style="width:136px;">14094-11</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Surgical Scissors</td>
			<td style="width:292px;">Fine Science Tools GmbH</td>
			<td style="width:136px;">14001-18</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Micro Dressing Forceps (curved, serrated)</td>
			<td style="width:292px;">Aesculap, Inc.</td>
			<td style="width:136px;">BD312R</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Tissue Forceps (straight, 1 x 2 teeth)</td>
			<td style="width:292px;">Aesculap, Inc.</td>
			<td style="width:136px;">BD537R</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Tying Forceps (angled)</td>
			<td style="width:292px;">Aesculap, Inc.</td>
			<td style="width:136px;">MA624R</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Rodent and Small Animal Guillotine</td>
			<td style="width:292px;">Kent Scientific Corp.</td>
			<td style="width:136px;">DCAP</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Low Cost Induction Chamber 3.0 L</td>
			<td style="width:292px;">Kent Scientific Corp.</td>
			<td style="width:136px;">SOMNO-0730&#160;</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Butterfly Winged Infusion Set 21 G</td>
			<td style="width:292px;">Hospira, Inc.</td>
			<td style="width:136px;">181106101</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Abbocath 16 G</td>
			<td style="width:292px;">Hospira, Inc.</td>
			<td>0G7149702</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Microlance Hypodermic Needle</td>
			<td style="width:292px;">Becton Dickinson GmbH</td>
			<td style="width:136px;">301300</td>
			<td>modify needle to make cannula</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Braun Original Perfusor Syringe 50 ml</td>
			<td style="width:292px;">B. Braun Melsungen AG</td>
			<td style="width:136px;">8728810F</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Braun Inject Solo Syringe 10 ml</td>
			<td style="width:292px;">B. Braun Melsungen AG</td>
			<td style="width:136px;">2057926</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Beaker 50ml</td>
			<td style="width:292px;">Duran Group (DWK Life Sciences GmbH)</td>
			<td>21 106 17</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Duroplan petri dish (100 x 20 mm)</td>
			<td style="width:292px;">Duran Group (DWK Life Sciences GmbH)</td>
			<td style="width:136px;">21 755 48</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Seraflex Suture USP 3/0</td>
			<td style="width:292px;">SERAG-WIESSNER GmbH &#38; Co. KG</td>
			<td style="width:136px;">IC208000</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">VWR disposable Square Weighin Boats 100ml</td>
			<td style="width:292px;">VWR, Inc.</td>
			<td style="width:136px;">10803-148</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Styrofoam surface</td>
			<td style="width:292px;"></td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Sodium chloride</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">71380</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Potassium chloride</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">P4504</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Potassium phosphate monobasic</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">P5379</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Sodium phosphate dibasic</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">S0876</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Magensium sulfate heptahydrate</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">230391</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Magensium chloride</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">M8266</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">HEPES</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">H3375</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Taurine</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">T0625</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Glucose</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">G7528</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">2,3-Butanedione monoxime</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">B0753</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Calcium chloride solution (1 M)</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">21115</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Bovine Serum Albumin</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">A9647</td>
			<td style="width:227px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Liberase</td>
			<td style="width:292px;">Roche (Sigma-Aldrich, Inc.)</td>
			<td style="width:136px;">LIBTM-RO</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Heparin</td>
			<td style="width:292px;">Rotexmedica GmbH</td>
			<td style="width:136px;">3862357</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Forene (Isoflurane)</td>
			<td style="width:292px;">Abbvie Deutschland GmbH &#38; Co. KG</td>
			<td style="width:136px;">10182054</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Laminin from Engelbreth-Holm-Swarm murine sarcoma basement membrane</td>
			<td style="width:292px;">Sigma-Aldrich, Inc.</td>
			<td style="width:136px;">L2020</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">WillCo glass-bottom dish 500&#181;l 0.005mm</td>
			<td style="width:292px;">WillCo Wells B.V.</td>
			<td style="width:136px;">HBST-3522</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Fluo4 AM</td>
			<td style="width:292px;">Invitrogen (Thermo Fisher Scientific, Inc.)</td>
			<td>F14201</td>
			<td>5&#181;M for 20min at RT</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Di-8-ANNEPS</td>
			<td style="width:292px;">Invitrogen (Thermo Fisher Scientific, Inc.)</td>
			<td>D3167</td>
			<td>10&#181;M for 45 min at 37&#176; C&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Mitotracker RED FM</td>
			<td style="width:292px;">Invitrogen (Thermo Fisher Scientific, Inc.)</td>
			<td style="width:136px;">M22425</td>
			<td>20nM for 30 min at 37&#176; C</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Jacketed reaction vessel 500 ml</td>
			<td style="width:292px;">Gebr. Rettberg GmbH</td>
			<td>107024414</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Jacketed reaction vessel 1000 ml</td>
			<td style="width:292px;">Gebr. Rettberg GmbH</td>
			<td>107025414</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Jacketed bubble trap</td>
			<td style="width:292px;">Gebr. Rettberg GmbH</td>
			<td>134720001</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">ED heating immersion circulator</td>
			<td style="width:292px;">Julabo GmbH</td>
			<td>9116000</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Reglo Digital MS-2/6 peristaltic pump</td>
			<td style="width:292px;">Ismatec (Cole-Parmer Gmbh)</td>
			<td style="width:136px;">ISM 831</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Voltcraft Thermometer 302 K/J</td>
			<td style="width:292px;">Conrad Electronic SE</td>
			<td style="width:136px;">030300546</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Tubing</td>
			<td style="width:292px;"></td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">LSM 700 microscope</td>
			<td style="width:292px;">Carl Zeiss, Inc.</td>
			<td style="width:136px;"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">ZEN 2.3 imaging software</td>
			<td style="width:292px;">Carl Zeiss, Inc.</td>
			<td style="width:136px;">410135-1011-240&#160;</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">Single channel heater controller TC-324B</td>
			<td style="width:292px;">Warner Instruments, LLC</td>
			<td style="width:136px;">64-2400</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">8 channel perfusion system</td>
			<td style="width:292px;">Warner Instruments, LLC</td>
			<td style="width:136px;">64-0185</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:281px;">8 channel Multi-Line In-Line Solution Heaters</td>
			<td style="width:292px;">Warner Instruments, LLC</td>
			<td style="width:136px;">64-0105</td>
			<td></td>
		</tr>
	</tbody>
</table>

isolation of atrial cardiomyocytes from a rat model of metabolic syndrome-related heart failure with preserved ejection fraction

I denne artikel vil vi beskrive en optimeret, Langendorff-baseret procedure til isolering af enkelt celle atrieflimren cardiomyocytes (ACMs) fra en rotte model af metabolisk syndrom (MetS)-relaterede hjertesvigt med bevarede uddrivningsfraktion (HFpEF). Forekomsten af MetS-relaterede HFpEF er stigende, og atrieflimren cardiomyopatier forbundet med atrieflimren remodellering og atrieflimren er klinisk meget relevante som atrial remodellering er en uafhængig prædiktor for dødelighed. Undersøgelser med isolerede encellede cardiomyocytes bruges ofte til at bekræfte og supplere i vivo resultater. Kredsløbssygdomme fartøj rarefication og interstitielle væv fibrose udgør et potentielt begrænsende faktor for succesfuld encellede isolation ACMs fra dyremodeller for denne sygdom.
Vi har behandlet dette spørgsmål ved at ansætte en enhed i stand til manuelt regulering intraluminal presset af hjertets hulrum under proceduren isolation væsentligt øge udbyttet af morfologisk og funktionelt intakt ACMs. De erhvervede celler kan bruges i en række forskellige eksperimenter, som cellekultur og funktionelle Calcium imaging (dvs., excitation sammentrækning kobling).
Vi give forskeren en trinvis protokol, en liste over optimerede løsninger, grundig vejledning til at forberede det nødvendige udstyr, og en omfattende fejlfindingsvejledning. Mens den indledende implementering af proceduren kan være temmelig vanskeligt, vil en vellykket tilpasning tillader læseren at udføre state-of-the-art ACM isolering i en rotte model af MetS-relaterede HFpEF for et bredt spektrum af eksperimenter.

21 uger gamle, kan 60 – 90% af levedygtige ACMs (anslået som beskrevet i trin 6.1), efter calcium fornyet tilpasning (trin 5.4-5.7), isoleres fra ZSF-1 fede rotter ved denne metode (figur 4A). Hos rotter, er ACMs karakteriseret ved en anden og mere heterogene fænotype i forhold til VCMs24,25. Figur 4B viser en individuel ACM med bevarede membraner og sarkomeret strukt...

Watch this Scientific Journal Video about Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction at JoVE.com

Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction

Isolering af atrieflimren Cardiomyocytes fra en rotte Model af metabolisk syndrom-relaterede hjertesvigt med bevarede uddrivningsfraktion

Her, beskriver vi en optimeret, Langendorff-baseret procedure til isolering af enkelt celle atrieflimren cardiomyocytes fra en rotte model af metabolisk syndrom-relaterede hjertesvigt med bevarede uddrivningsfraktion. En manuel regulering af intraluminal pres af hjertets hulrum er gennemført for at give funktionelt intakt myocytes egnet til magnetisering sammentrækning kobling undersøgelser.

In this article, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes (ACMs) from a rat model of ...

Research

JoVE Journal

Medicine

Gene Transfer for Ischemic Heart Failure in a Preclinical Model

Various emerging technologies are being developed for patients with heart failure. Well-established preclinical evaluations are necessary to determine ...

The WATCHMAN Left Atrial Appendage Closure Device for Atrial Fibrillation

Vagten venstre forkammer vedhæng lukkeanordning for atrieflimren

Atrial fibrillation (AF) is the most common cardiac arrhythmia, affecting an estimated 6 million people in the United States 1. Since AF affects primarily ...

A New Single Chamber Implantable Defibrillator with Atrial Sensing: A Practical Demonstration of Sensing and  Ease of Implantation

En ny single Afdeling implanterbar defibrillator med atrieflimren Sensing: en praktisk demonstration af Sensing og Brugervenlighed Implantation

Implantable cardioverter-defibrillators (ICDs) terminate ventricular tachycardia (VT) and ventricular fibrillation (VF) with high efficacy and can protect ...

Method of Isolated Ex Vivo Lung Perfusion in a Rat Model: Lessons Learned from Developing a Rat EVLP Program

Method of Isolated Ex Vivo Lung Perfusion in a Rat Model: Lessons Learned from Developing a Rat EVLP Program

Metode til isoleret<em&gt; Ex vivo</em&gt; Lung Perfusion i en rotte Model: Erfaringer fra at udvikle en rotte EVLP Program

The number of acceptable donor lungs available for lung transplantation is severely limited due to poor quality. Ex-Vivo Lung Perfusion (EVLP) has allowed ...

The Inverted Heart Model for Interstitial Transudate Collection from the Isolated Rat Heart

Den omvendte hjertemodel for interstitiell transudatsamling fra det isolerede rottehjerte

The present protocol describes a unique approach that enables the collection of cardiac transudate (CT) from the isolated, saline-perfused rat heart. ...

Isolation and Cultivation of Adult Rat Cardiomyocytes

Isolering og dyrkning af voksen rotte Cardiomyocytes

In an intact heart, adjacent cells influence adult cardiomyocytes. With the method of isolation and cultivation of adult cardiomyocytes, a precise ...

Tachycardia-Induced Cardiomyopathy As a Chronic Heart Failure Model in Swine

Takykardi-induceret kardiomyopati som kronisk hjertesvigt Model hos svin

A stable and reliable model of chronic heart failure is required for many experiments to understand hemodynamics or to test effects of new treatment ...

Isolation of Atrial Myocytes from Adult Mice

Isolering af atrial Myocytes fra voksne mus

The electrophysiological properties of atrial myocytes importantly affect overall cardiac function. Alterations in the underlying ionic currents ...

Generation of a Rat Model of Acute Liver Failure by Combining 70% Partial Hepatectomy and Acetaminophen

Generation af en rotte model af akut leversvigt ved at kombinere 70% partiel Hepatektomi og acetaminophen

Acute liver failure (ALF) is a clinical condition caused by various etiologies resulting in the loss of metabolic, biochemical, synthesizing, and ...

Establishing a Swine Model of Post-myocardial Infarction Heart Failure for Stem Cell Treatment

Etablering af en svin model af post-myokardieinfark hjertesvigt for stamcellebehandling

Although advances have been achieved in the treatment of heart failure (HF) following myocardial infarction (MI), HF following MI remains one of the major ...