<ol>
	<li>Chlopcikova, S., Psotov&#225;, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neonatal+rat+cardiomyocytes-a+model+for+the+study+of+morphological,+biochemical+and+electrophysiological+characteristics+of+the+heart.">Neonatal rat cardiomyocytes-a model for the study of morphological, biochemical and electrophysiological characteristics of the heart.</a> Biomedical Papers. , (2001).</li><li>Rosenthal, N., Brown, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+mouse+ascending:+perspectives+for+human-disease+models.">The mouse ascending: perspectives for human-disease models.</a> Nat. Cell Biol. 9, 993-999 (2007).</li><li>Ballou, L. M., Lin, R. Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapamycin+and+mTOR+kinase+inhibitors.">Rapamycin and mTOR kinase inhibitors.</a> J. Chem. Biol. 1, 27-36 (2008).</li><li>Brette, F., Orchard, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=T-Tubule+Function+in+Mammalian+Cardiac+Myocytes.">T-Tubule Function in Mammalian Cardiac Myocytes.</a> Circ Res. , (2003).</li><li>Sakata, Y., Hoit, B., Liggett, S., Walsh, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decompensation+of+pressure-overload+hypertrophy+in+G?q-overexpressing+mice.">Decompensation of pressure-overload hypertrophy in G?q-overexpressing mice.</a> Circulation. , (1998).</li><li>Lieu, D. K., 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=Absence+of+Transverse+Tubules+Contributes+to+Non-Uniform+Ca+2+Wavefronts+in+Mouse+and+Human+Embryonic+Stem+Cell-Derived+Cardiomyocytes.">Absence of Transverse Tubules Contributes to Non-Uniform Ca 2+Wavefronts in Mouse and Human Embryonic Stem Cell-Derived Cardiomyocytes.</a> Stem Cells and Development. 18, 1493-1500 (2009).</li><li>Lim, H., De Windt, L., Mante, J., Kimball, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reversal+of+cardiac+hypertrophy+in+transgenic+disease+models+by+calcineurin+inhibition.">Reversal of cardiac hypertrophy in transgenic disease models by calcineurin inhibition.</a> J. Mol. Cell Cardiol. 32 (4), 697-709 (2000).</li><li>Muthuchamy, 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=Mouse+model+of+a+familial+hypertrophic+cardiomyopathy+mutation+in+alpha-tropomyosin+manifests+cardiac+dysfunction.">Mouse model of a familial hypertrophic cardiomyopathy mutation in alpha-tropomyosin manifests cardiac dysfunction.</a> Circ. Res. 85, 47-56 (1999).</li><li>M&#252;ller, J. 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=Differential+regulation+of+the+cardiac+sodium+calcium+exchanger+promoter+in+adult+and+neonatal+cardiomyocytes+by+Nkx2.5+and+serum+response+factor.">Differential regulation of the cardiac sodium calcium exchanger promoter in adult and neonatal cardiomyocytes by Nkx2.5 and serum response factor.</a> J. Mol. Cell. Cardiol. 34, 807-821 (2002).</li><li>Epstein, F., Hunter, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Signaling+pathways+for+cardiac+hypertrophy+and+failure.">Signaling pathways for cardiac hypertrophy and failure.</a> New England Journal of Medicine. 341 (17), 1276-1283 (1999).</li><li>Snopko, R., Aromolaran, A., Karko, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell+culture+modifies+Ca+2++signaling+during+excitation-contraction+coupling+in+neonate+cardiac+myocytes.">Cell culture modifies Ca 2+ signaling during excitation-contraction coupling in neonate cardiac myocytes.</a> Cell Calcium. , (2007).</li><li>Ranu, H., Terracciano, C., Davia, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+Na+/Ca2+-exchanger+overexpression+on+excitation-contraction+coupling+in+adult+rabbit+ventricular+myocytes.">Effects of Na+/Ca2+-exchanger overexpression on excitation-contraction coupling in adult rabbit ventricular myocytes.</a> J. Mol. Cell Cardiol. 34 (4), 389-400 (2002).</li><li>Kaab, S., Nuss, H. B., Chiamvimonvat, N., O'Rourke, B., Pak, P. H., Kass, D. A., Marban, E., Tomaselli, G. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ionic+mechanism+of+action+potential+prolongation+in+ventricular+myocytes+from+dogs+with+pacing-induced+heart+failure.">Ionic mechanism of action potential prolongation in ventricular myocytes from dogs with pacing-induced heart failure.</a> Circ. Res. 78 (2), (1996).</li><li>Splawski, I., 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=Variant+of+SCN5A+sodium+channel+implicated+in+risk+of+cardiac+arrhythmia.">Variant of SCN5A sodium channel implicated in risk of cardiac arrhythmia.</a> Science. 297, 1333-1336 (2002).</li><li>Zhou, Y., Wang, S., Zhu, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Culture+and+adenoviral+infection+of+adult+mouse+cardiac+myocytes:+methods+for+cellular+genetic+physiology.">Culture and adenoviral infection of adult mouse cardiac myocytes: methods for cellular genetic physiology.</a> Am. J. Physiol. Heart Circ. Physiol. 279 (1), 429-436 (2000).</li><li>Sussman, M., Lim, H., Gude, N., Taigen, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prevention+of+cardiac+hypertrophy+in+mice+by+calcineurin+inhibition.">Prevention of cardiac hypertrophy in mice by calcineurin inhibition.</a> Science. , (1998).</li><li>Heinzel, F. R., 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=Impairment+of+diazoxide-induced+formation+of+reactive+oxygen+species+and+loss+of+cardioprotection+in+connexin+43+deficient+mice.">Impairment of diazoxide-induced formation of reactive oxygen species and loss of cardioprotection in connexin 43 deficient mice.</a> Circ. Res. 97, 583-586 (2005).</li><li>Li, X., Heinzel, F. R., Boengler, K., Schulz, R., Heusch, G. <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+connexin+43+in+ischemic+preconditioning+does+not+involve+intercellular+communication+through+gap+junctions.">Role of connexin 43 in ischemic preconditioning does not involve intercellular communication through gap junctions.</a> J. Mol. Cell. Cardiol. 36, 161-163 (2004).</li><li>Rockman, H. A., Ross, R. S., Harris, A. N., Knowlton, K. U., Steinhelper, M. E., Field, L. J., Ross, J., Chein, K. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Segregation+of+atrial-specific+and+inducible+expression+of+an+atrial+natriuretic+factor+transgene+in+an+in+vivo+murine+model+of+cardiac+hypertrophy.">Segregation of atrial-specific and inducible expression of an atrial natriuretic factor transgene in an in vivo murine model of cardiac hypertrophy.</a> Proc. Natl. Acad. Sci. USA. 88, 8277-8281 (1991).</li><li>Nakamura, A., Rokosh, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=LV+systolic+performance+improves+with+development+of+hypertrophy+after+transverse+aortic+constriction+in+mice.">LV systolic performance improves with development of hypertrophy after transverse aortic constriction in mice.</a> Am. J. Physiol. Heart Circ. Physiol. 281 (3), 1104-1112 (2001).</li><li>Bostr&#246;m, P., 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+PGC1-?-dependent+myokine+that+drives+brown-fat-like+development+of+white+fat+and+thermogenesis.">A PGC1-?-dependent myokine that drives brown-fat-like development of white fat and thermogenesis.</a> Nature. 481, 463-468 (2012).</li><li>Liao, R., Jain, M. <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+analysis+of+adult+mouse+cardiomyocytes.">Isolation, culture, and functional analysis of adult mouse cardiomyocytes.</a> Methods Mol. Med. 139, 251-262 (2007).</li><li>Volz, A., Piper, H. M., Siegmund, B., Schwartz, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Longevity+of+adult+ventricular+rat+heart+muscle+cells+in+serum-free+primary+culture.">Longevity of adult ventricular rat heart muscle cells in serum-free primary culture.</a> J. Mol. Cell. Cardiol. 23, 161-173 (1991).</li><li>Guatimosim, S., Guatimosim, C., Song, L. -. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Methods in Molecular Biology. 689, 205-214 (2010).</li><li>Yang, J., Delaloye, K., Lee, U. S., Cui, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patch+Clamp+and+Perfusion+Techniques+for+Studying+Ion+Channels+Expressed+in+Xenopus+oocytes.">Patch Clamp and Perfusion Techniques for Studying Ion Channels Expressed in Xenopus oocytes.</a> J. Vis. Exp. (47), e2269 (2011).</li><li>Brown, A. L., Johnson, B. E., Goodman, M. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patch+Clamp+Recording+of+Ion+Channels+Expressed+in+Xenopus+Oocytes.">Patch Clamp Recording of Ion Channels Expressed in Xenopus Oocytes.</a> J. Vis. Exp. (20), e936 (2008).</li><li>Arman, A. C., Sampath, A. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patch+Clamp+Recordings+from+Mouse+Retinal+Neurons+in+a+Dark-adapted+Slice+Preparation.">Patch Clamp Recordings from Mouse Retinal Neurons in a Dark-adapted Slice Preparation.</a> J. Vis. Exp. (43), e2107 (2010).</li><li>Wen, H., Brehm, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Paired+patch+clamp+recordings+from+motor-neuron+and+target+skeletal+muscle+in+zebrafish.">Paired patch clamp recordings from motor-neuron and target skeletal muscle in zebrafish.</a> J. Vis. Exp. (45), e2351 (2010).</li><li>Maier, S., Westenbroek, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+unexpected+role+for+brain-type+sodium+channels+in+coupling+of+cell+surface+depolarization+to+contraction+in+the+heart.">An unexpected role for brain-type sodium channels in coupling of cell surface depolarization to contraction in the heart.</a> Proc. Natl. Acad. Sci. U.S.A. 99 (6), 4073-4078 (2002).</li><li>Shroff, S. G., Saner, D. R., Lal, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamic+micromechanical+properties+of+cultured+rat+atrial+myocytes+measured+by+atomic+force+microscopy.">Dynamic micromechanical properties of cultured rat atrial myocytes measured by atomic force microscopy.</a> Am. J. Physiol. 269, 286-292 (1995).</li><li>Domke, J., Parak, W. J., George, M., Gaub, H. E., Radmacher, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mapping+the+mechanical+pulse+of+single+cardiomyocytes+with+the+atomic+force+microscope.">Mapping the mechanical pulse of single cardiomyocytes with the atomic force microscope.</a> European Biophysics Journal. 28, 179-186 (1999).</li><li>Smith, B. A., Tolloczko, B., Martin, J. G., Gr&#252;tter, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Probing+the+Viscoelastic+Behavior+of+Cultured+Airway+Smooth+Muscle+Cells+with+Atomic+Force+Microscopy:+Stiffening+Induced+by+Contractile+Agonist.">Probing the Viscoelastic Behavior of Cultured Airway Smooth Muscle Cells with Atomic Force Microscopy: Stiffening Induced by Contractile Agonist.</a> Biophysical Journal. 88, 2994-3007 (2005).</li></ol>

يعلن الكتاب أنه ليس لديهم مصالح مالية المتنافسة.

في هذا التقرير، وصفناها التقنيات اللازمة لعزل ناجحة وثقافة الكبار مضادة من القلب الماوس. أسلوبنا يسمح لدراسة لاحقة من وظيفة CM واستثارة باستخدام الأساليب المذكورة أعلاه. المعلمة الحرجة لدراسة وظيفة من الكبار مضادة هي الصحة ونوعية مضادة معزولة. كما هو موضح أعلاه، لدي...

خدموا نماذج الفئران من مرض القلب والأوعية الدموية كأدوات فعالة لتوضيح الآليات الأساسية المرض 1،2 فضلا عن تحديد الأهداف العلاجية المحتملة 1،3. على وجه الخصوص، واستخدام كلا النموذجين الفئران من أمراض القلب المكتسبة (مثل الضغط الزائد) 4،5 ونماذج الماوس المعدلة وراثيا قد تقدم فهمنا لمرض القلب 6-8. استخدام تقنيات زراعة الخلايا لدراسة الإشارات شلالات 3،9،10 وتغييرات في البروتينات الفردية التي تكمن وراء استثارة الخلوية والإثارة، تقلص اقتران في قلب 11-13 على مستوى خلية واحدة ويكمل في الجسم الحي نماذج الماوس. ومع ذلك، فإن عدم وجود خطوط الخلايا الكافية التي تعكس هيكل CM الكبار وظيفة جود قيود كبيرة. وقد سعى المحققون للتغلب على هذا من خلال دراسة البروتينات الفردية، مثل القنوات الأيونية، في أنظمة تعبير مغايرة <suص 14&gt;، وبينما قدمت هذه الدراسات لنا معلومات مفيدة من حيث الفيزياء الحيوية القناة الايونية أو الاتجار البروتين، والتمثيل غير الكافي للالمكروية مواليد مضادة وجود قيود كبيرة. وثانيا، لأن معظم هذه الخلايا مغاير لم يكن لديك جهاز مقلص ناضجة، لم يكن من الممكن لدراسة وظيفة مقلص والتفاعل المعقد بين استثارة الخلوية والانكماش. لهذا السبب، تحولت الباحثين لمزارع الخلايا القلبية الأساسية للعديد من الدراسات في المختبر وظيفية. أخيرا، والدراسات cardiomyocyte معزولة تسمح بتقييم وظيفة مقلص دون عوامل خارجية متعددة الخلايا من إعداد بما في ذلك أثر ندبة أو تليف واتجاه الألياف. وسهلة نسبيا للثقافة، يمكن أن يصاب الأطفال حديثي الولادة العضلية البطين الفئران الابتدائي (NRVMs) مع الفيروسات الغدية وlentiviruses للتلاعب التعبير الجيني 15، والثانية تم لذا استخدمت بنجاح 1، ولكن لديها قيود خاصة بهم. على الرغم من أنها توفر المكروية الفسيولوجية 1 وكانت العمود الفقري للحقل الإشارات، اختلافات جوهرية بين التشكل وتنظيم التحت خلوية من NRVMs وcardiomyoctyes الكبار جعلها نموذجا غير كافية للتحقيق في تدفقات الأيونية والإثارة، تقلص اقتران في قلب الكبار . أبرزها NRVMs تفتقر إلى نهائي تي أنبوبي الفرعي 4. منذ كا 2 + تدفق وديناميكية تعتمد بشكل حاسم على ناضجة تي أنبوبي وشبكية الهيولى العضلية (SR) هيكل 6، كا 2 + ديناميات والدراسات الفنية التابعة للانقباض القلب في NRVMs ليست انعكاسا دقيقا لهذه العمليات الحرجة في العضلية الكبار. علاوة على ذلك، بعض مكونات مسارات الإشارات تختلف بين الأطفال حديثي الولادة والكبار الفئران 9، وبالتالي توفير الحد أخرى لدراسة عمليات المرض وأنا علىMPACT على استثارة الخلوية وانقباض في NRVMs. أخيرا، وتوزيع آلات مقلص يؤدي إلى متعدد الاتجاهات وغير موحدة تقصير الخلايا مما يحد من دقة القياسات مقلص. وبالتالي فإن استخدام العضلية الكبار معزولة يوفر أكثر دقة في المختبر نظام النمذجة. نمو استثنائية من المعرفة الذي أصبح ممكنا بفضل التلاعب الجيني للفئران يؤكد أهمية الحصول العضلية المعزولة من الفئران وظيفية. في الواقع، فإن توصيف مضادة الكبار معزولة عن نماذج الماوس وتسليط الضوء على العديد من الأحداث البيولوجية والمرضية. وقد سمحت مضادة معزولة عن نماذج الماوس المعدلة وراثيا للدراسات من ربح أو خسارة وظيفة من البروتينات مقلص على خصائص الخلايا واحدة 2،16، وقدرتها على البقاء في نماذج الأمراض مثل نقص التروية / ضخه 17،18، وبالتالي استكمال المعلومات المكتسبة من في الدراسات المجراة علىالفئران حد ذاتها. استخدام الكبار معزولة مضادة من نماذج الفئران من أمراض القلب المكتسبة 3،19،20 (مثل عرضية الأبهر الناجم عن انقباض الضغط الزائد، الذي يحاكي ارتفاع ضغط الدم أو تضيق الأبهر صمام) أو ممارسة 5،21 (لنمذجة تضخم الفسيولوجية) يسمح للفحص تفاعل يشير شلالات المتورطين في هذه العمليات مع استثارة الخلوية والإثارة، تقلص اقتران على مستوى خلية واحدة. وعلاوة على ذلك، والقدرة على التعامل مع التعبير الجيني باستخدام يحركها الغدانية التعبير الجيني في الكبار مضادة يتيح لنا الفرصة لتشريح مكونات مسارات الإشارات المعقدة. من وجهة نظر الكهربية، وقد تم الجهد خلية كاملة والتجارب المشبك الحالي على الكبار معزولة مضادة حاسمة في توضيح طبيعة التدفقات الأيونية في الأساس وفي مختلف الحالات المرضية. بسبب بنية معقدة من الأغشية الخلوية والمتواجد التفاضليةفي هياكل السقالات بين الكبار مضادة وNRVMs أو خطوط الخلايا مغاير، والقدرة على التصحيح خلايا بالغة يعطي تمثيل أفضل بكثير من آثار بعض البروتينات الغشاء، والبروتينات الهيكلية، والقناة الايونية شركاء التفاعل على المكونات الكهربية للقلب الكبار. على الرغم من هذه المزايا البارزة في دراسة الكبار العضلية الفئران وعزل وزراعة العضلية الفئران تم تحدي، وحث على ضرورة وصف منظم ودقيق للمنهجية عزل العضلية الماوس قابلة للحياة والمحافظة عليها في الثقافة للسماح مزيد من التلاعب الجيني باستخدام الفيروسية ناقلات. وقد استخدمت الدراسات السابقة إما معزولة تماما الكبار الماوس مضادة أو مستنبت الكبار الفئران مضادة. هذه الأخيرة هي أسهل للثقافة من الماوس الكبار سم، واستخدمت معظم التجارب التلاعب التعبير الجيني في المختبر الفئران الكبار مضادة. غيرت بنجاح قليل من الدراسات والتحقيق الوظائالتعبير الجيني اونال في الماوس الكبار مضادة، وتقديم الحد الكبيرة في نطاق التجارب. وبالتالي، فإننا نقدم هنا في التفاصيل هذه المنهجيات، معدلة من التحقيقات السابقة، لعزل 7،8،22 والثقافة 3،10،15،23، عدوى الفيروسة الغدانية 11-13،15، والتحليل الوظيفي من الماوس الكبار cardiomyoctyes البطين. هذه النتائج بروتوكول العزلة في كا 2 +-تسامحا، منفعل العضلية التي لدينا مثقف بنجاح لمدة تصل إلى 72 ساعة وعابر مع اتش. وظيفة هذه الخلايا معزولة يمكن تقييمها باستخدام نظام التصوير MMSYS 14،24 والمشبك التصحيح، والتي ستناقش أيضا.

<table border="1">
<tbody>
 <tr>
 <td>Name of 			Reagent/Material</td>
 <td>Company</td>
 <td>Catalog Number</td>
 <td>Comments</td>
 </tr>
 <tr>
 <td>Sodium Chloride</td>
 <td>Sigma</td>
 <td>S7653</td>
 <td></td>
 </tr>
 <tr>
 <td>Potassium Chloride</td>
 <td>Sigma</td>
 <td>P9333</td>
 <td></td>
 </tr>
 <tr>
 <td>Magnesium Chloride</td>
 <td>Sigma</td>
 <td>M8266</td>
 <td></td>
 </tr>
 <tr>
 <td>HEPES</td>
 <td>Sigma</td>
 <td>H3375</td>
 <td></td>
 </tr>
 <tr>
 <td>Sodium Phosphate 			Monobasic</td>
 <td>Sigma</td>
 <td>S8282</td>
 <td></td>
 </tr>
 <tr>
 <td>D-glucose minimum</td>
 <td>Sigma</td>
 <td>G8270</td>
 <td></td>
 </tr>
 <tr>
 <td>Taurine</td>
 <td>Sigma</td>
 <td>T0625</td>
 <td></td>
 </tr>
 <tr>
 <td>2,3-Butanedione 			monoxime</td>
 <td>Sigma</td>
 <td>B0752</td>
 <td></td>
 </tr>
 <tr>
 <td>Collagenase B</td>
 <td>Roche Applied 			Science</td>
 <td>11088807001 </td>
 <td></td>
 </tr>
 <tr>
 <td>Collagenase D</td>
 <td>Roche Applied 			Science</td>
 <td>11088858001 </td>
 <td></td>
 </tr>
 <tr>
 <td>Protease 			XIV from Streptomyces griseus</td>
 <td>Sigma</td>
 <td>P5147</td>
 <td></td>
 </tr>
 <tr>
 <td>Albumin from Bovine 			Serum</td>
 <td>Sigma</td>
 <td>A2153</td>
 <td></td>
 </tr>
 <tr>
 <td>Calcium Chloride</td>
 <td>Sigma</td>
 <td>C8106</td>
 <td></td>
 </tr>
 <tr>
 <td>Minimum Essential 			Media</td>
 <td>Sigma</td>
 <td>51411C</td>
 <td></td>
 </tr>
 <tr>
 <td>Albumin solution 			from bovine serum</td>
 <td>Sigma</td>
 <td>A8412</td>
 <td></td>
 </tr>
 <tr>
 <td>L-glutamine</td>
 <td>Sigma</td>
 <td>G3126</td>
 <td></td>
 </tr>
 <tr>
 <td>Penicillin-Streptomycin</td>
 <td>Sigma</td>
 <td>P4333</td>
 <td></td>
 </tr>
 <tr>
 <td>Insulin-transferrin-sodium 			selenite media supplement</td>
 <td>Sigma</td>
 <td>I1884</td>
 <td></td>
 </tr>
 <tr>
 <td>Cesium Chloride</td>
 <td>Sigma</td>
 <td><a href="http://www.sigmaaldrich.com/catalog/product/sial/289329?lang=en&amp;region=US" target="_blank">289329</a></td>
 <td></td>
 </tr>
 <tr>
 <td>Glutamate</td>
 <td>Sigma</td>
 <td>G3291</td>
 <td></td>
 </tr>
 <tr>
 <td>Adenosine 			5'-triphosphate magnesium salt</td>
 <td>Sigma</td>
 <td>A9187</td>
 <td></td>
 </tr>
 <tr>
 <td>Ethylene 			glycol-bis(2-aminoethylether)-N,N,N',N'-tetraacetic 			acid</td>
 <td>Sigma</td>
 <td>E3889</td>
 <td></td>
 </tr>
 <tr>
 <td>Cesium Hydroxide 			Solution</td>
 <td>Sigma</td>
 <td>232041</td>
 <td></td>
 </tr>
 <tr>
 <td>Tetraethylammonium 			hydroxide solution</td>
 <td>Sigma</td>
 <td>86643</td>
 <td></td>
 </tr>
 <tr>
 <td>OptiVisor optical 			glass binocular 			visor</td>
 <td>Dohegan Optical 			Company Inc.</td>
 <td>N/A</td>
 <td></td>
 </tr>
 <tr>
 <td>Tissue 			forceps, 5.5", 1x2 teeth</td>
 <td>Roboz Scientific</td>
 <td>RS-8164</td>
 <td></td>
 </tr>
 <tr>
 <td>Moloney 			forceps - 4.5" (11.5 cm) long slight curve, serrated </td>
 <td>Roboz Scientific</td>
 <td>RS-8254</td>
 <td></td>
 </tr>
 <tr>
 <td>Dumont 			#3 Forceps, Dumostar, tip size 0.17 x 0.10mm</td>
 <td>Roboz Scientific</td>
 <td>RS-4966</td>
 <td></td>
 </tr>
 <tr>
 <td>Packer 			Mosquito Forceps 5" Straight Flat</td>
 <td>Roboz Scientific</td>
 <td>RS-7114</td>
 <td></td>
 </tr>
 <tr>
 <td>Micro 			Dissecting Scissors 4.5" Curved Sharp/Sharp </td>
 <td>Roboz Scientific</td>
 <td>RS-5917</td>
 <td></td>
 </tr>
 <tr>
 <td>Micro Dissecting Scissors 3.5" Straight Sharp/Sharp20mm</td>
 <td>Roboz Scientific</td>
 <td>RS-5907</td>
 <td></td>
 </tr>
 </tbody>
</table>

isolation, culture, and functional characterization of adult mouse cardiomyoctyes

وقد وفرت استخدام العضلية الأولية (CMS) في الثقافة تكملة قوية لنماذج الفئران من أمراض القلب في دفع فهمنا لمرض القلب. على وجه الخصوص، وقد أدت القدرة على دراسة توازن الأيوني، وظيفة القناة الايونية، استثارة الخلوية والإثارة، تقلص اقتران والتعديلات في ظروف المريضة والطفرات المسببة للمرض إلى رؤى كبيرة في أمراض القلب. علاوة على ذلك، فإن عدم وجود خط الخلية كافية لتقليد الكبار خلد مضادة، والقيود المفروضة على الأطفال حديثي الولادة مضادة (التي تفتقر إلى العديد من الخصائص الميكانيكا الحيوية الهيكلية والوظيفية من الكبار CMS) في الثقافة أعاقت فهمنا للتفاعل معقد بين مسارات الإشارات، القنوات الأيونية وخصائص مقلص في قلب الكبار تعزيز أهمية دراسة الكبار العضلية معزولة. هنا، فإننا نقدم طرق لعزل والثقافة، والتلاعب في التعبير الجيني عن طريق الغدانية-expreالبروتينات ssed، وتحليل وظيفي لاحقة من العضلية من الماوس الكبار. فإن استخدام هذه التقنيات تساعد على تطوير البصيرة الآلي في مسارات الإشارات التي تنظم استثارة الخلوية، كا 2 + ديناميات وانقباض وتوفير توصيف أكثر أهمية من الناحية الفسيولوجية الكثير من أمراض القلب والشرايين.

عزلة النتائج cardiomyoctyes الكبار في الخلايا على شكل قضيب، مخططة، وهادئة (لا ضرب عفويا) (الشكل 5A). والخلايا الميتة تبدو مدورة وسوف لا تكون موجودة التصدعات. خلايا هادئة يمكن أن يكون مثقف وtransfected مع اتش للتلاعب التعبير الجيني (أرقام 5B و5C). بعد 24 ساعة من الثقاف?...

Watch this Scientific Journal Video about Isolation, Culture, and Functional Characterization of Adult Mouse Cardiomyoctyes at JoVE.com

Isolation, Culture, and Functional Characterization of Adult Mouse Cardiomyoctyes

نحن هنا وصف عزلة cardiomyoctyes فأر بالغ باستخدام نظام نضح Langendorff. الخلايا الناتجة CA2 +-تسامحا، هادئة كهربائيا ويمكن أن يكون مثقف وtransfected مع-الغدة أو lentiviruses للتلاعب الجيني. ويمكن أيضا تحليل وظائفها باستخدام نظام MMSYS وتقنيات المشبك التصحيح.

العزلة والثقافة، وتوصيف وظيفي من الكبار ماوس Cardiomyoctyes

The use of primary cardiomyocytes (CMs) in culture has provided a powerful complement to murine models of heart disease in advancing our understanding of ...

isolation-culture-functional-characterization-adult-mouse

Research

JoVE Journal

Biology

24.2K Views.  Beth Israel Deaconess Medical Center, Harvard Medical School.  نحن هنا وصف عزلة cardiomyoctyes فأر بالغ باستخدام نظام نضح Langendorff. الخلايا الناتجة CA2 +-تسامحا، هادئة كهربائيا ويمكن أن يكون مثقف وtransfected مع-الغدة أو lentiviruses للتلاعب الجيني. ويمكن أيضا تحليل وظائفها باستخدام نظام MMSYS وتقنيات المشبك التصحيح. 

Video: Isolation, Culture, and Functional Characterization of Adult Mouse Cardiomyoctyes

Dissection of Hippocampal Dentate Gyrus from Adult Mouse

The hippocampus is one of the most widely studied areas in the brain because of its important functional role in memory processing and learning, its remarkable neuronal cell plasticity, and its involvement in epilepsy, neurodegenerative diseases, and psychiatric disorders. The hippocampus is composed of distinct regions; the dentate gyrus, which comprises mainly granule neurons, and Ammon's horn, which comprises mainly pyramidal neurons, and the two regions are connected by both anatomic and functional circuits. Many different mRNAs and proteins are selectively expressed in the dentate gyrus, and the dentate gyrus is a site of adult neurogenesis; that is, new neurons are continually generated in the adult dentate gyrus. To investigate mRNA and protein expression specific to the dentate gyrus, laser capture microdissection is often used. This method has some limitations, however, such as the need for special apparatuses and complicated handling procedures. In this video-recorded protocol, we demonstrate a dissection technique for removing the dentate gyrus from adult mouse under a stereomicroscope. Dentate gyrus samples prepared using this technique are suitable for any assay, including transcriptomic, proteomic, and cell biology analyses. We confirmed that the dissected tissue is dentate gyrus by conducting real-time PCR of dentate gyrus-specific genes, tryptophan 2,3-dioxygenase (TDO2) and desmoplakin (Dsp), and Ammon's horn enriched genes, Meis-related gene 1b (Mrg1b) and TYRO3 protein tyrosine kinase 3 (Tyro3). The mRNA expressions of TDO2 and Dsp in the dentate gyrus samples were detected at obviously higher levels, whereas Mrg1b and Tyro3 were lower levels, than those in the Ammon's horn samples. To demonstrate the advantage of this method, we performed DNA microarray analysis using samples of whole hippocampus and dentate gyrus. The mRNA expression of TDO2 and Dsp, which are expressed selectively in the dentate gyrus, in the whole hippocampus of alpha-CaMKII+/- mice, exhibited 0.037 and 0.10-fold changes compared to that of wild-type mice, respectively. In the isolated dentate gyrus, however, these expressions exhibited 0.011 and 0.021-fold changes compared to that of wild-type mice, demonstrating that gene expression changes in dentate gyrus can be detected with greater sensitivity. Taken together, this convenient and accurate dissection technique can be reliably used for studies focused on the dentate gyrus.

A dissection technique for removal of the dentate gyrus from adult mouse under a stereomicroscope was demonstrated in this video-recorded protocol.

تشريح التلفيف المسنن من الحصين ماوس الكبار

The hippocampus is one of the most widely studied areas in the brain because of its important functional role in memory processing and learning, its ...

Isolation and Culture of Adult Epithelial Stem Cells from Human Skin

The homeostasis of all self-renewing tissues is dependent on adult stem cells. As undifferentiated stem cells undergo asymmetric divisions, they generate daughter cells that retain the stem cell phenotype and transit-amplifying cells (TA cells) that migrate from the stem cell niche, undergo rapid proliferation and terminally differentiate to repopulate the tissue.
Epithelial stem cells have been identified in the epidermis, hair follicle, and intestine as cells with a high in vitro proliferative potential and as slow-cycling label-retaining cells in vivo 1-3. Adult, tissue-specific stem cells are responsible for the regeneration of the tissues in which they reside during normal physiologic turnover as well as during times of stress 4-5. Moreover, stem cells are generally considered to be multi-potent, possessing the capacity to give rise to multiple cell types within the tissue 6. For example, rodent hair follicle stem cells can generate epidermis, sebaceous glands, and hair follicles 7-9. We have shown that stem cells from the human hair follicle bulge region exhibit multi-potentiality 10.
Stem cells have become a valuable tool in biomedical research, due to their utility as an in vitro system for studying developmental biology, differentiation, tumorigenesis and for their possible therapeutic utility. It is likely that adult epithelial stem cells will be useful in the treatment of diseases such as ectodermal dysplasias, monilethrix, Netherton syndrome, Menkes disease, hereditary epidermolysis bullosa and alopecias 11-13. Additionally, other skin problems such as burn wounds, chronic wounds and ulcers will benefit from stem cell related therapies 14,15. Given the potential for reprogramming of adult cells into a pluripotent state (iPS cells)16,17, the readily accessible and expandable adult stem cells in human skin may provide a valuable source of cells for induction and downstream therapy for a wide range of disease including diabetes and Parkinson's disease.

A rapid, robust way of isolating viable adult epithelial stem cells from human skin is described. The method utilizes enzymatic digestion of skin collagen matrix , followed by plucking of hair follicles and isolation of single cell suspensions or tissue fragments for cell culture.

العزلة وثقافة الكبار الخلايا الجذعية الظهارية من الجلد البشري

The homeostasis of all self-renewing tissues is dependent on adult stem cells. As undifferentiated stem cells undergo asymmetric divisions, they generate ...

Isolation and Culture of Neonatal Mouse Cardiomyocytes

Cultured neonatal cardiomyocytes have long been used to study myofibrillogenesis and myofibrillar functions. Cultured cardiomyocytes allow for easy investigation and manipulation of biochemical pathways, and their effect on the biomechanical properties of spontaneously beating cardiomyocytes.	
The following 2-day protocol describes the isolation and culture of neonatal mouse cardiomyocytes. We show how to easily dissect hearts from neonates, dissociate the cardiac tissue and enrich cardiomyocytes from the cardiac cell-population. We discuss the usage of different enzyme mixes for cell-dissociation, and their effects on cell-viability. The isolated cardiomyocytes can be subsequently used for a variety of morphological, electrophysiological, biochemical, cell-biological or biomechanical assays. We optimized the protocol for robustness and reproducibility, by using only commercially available solutions and enzyme mixes that show little lot-to-lot variability. We also address common problems associated with the isolation and culture of cardiomyocytes, and offer a variety of options for the optimization of isolation and culture conditions.

Primary mouse cardiomyocyte cultures are one of the pivotal tools for the investigation of myofibrillar organization and function. The following protocol describes the isolation and culture of primary cardiomyocytes from neonatal mouse hearts. The resulting cardiomyocyte cultures may be subsequently used for a variety of biomechanical, biochemical and cell-biological assays.

العزلة وثقافة الأطفال حديثي الولادة ماوس العضلية

Cultured neonatal cardiomyocytes have long been used to study  myofibrillogenesis and myofibrillar functions. Cultured cardiomyocytes allow  for easy ...

Isolation and Culture of Dental Epithelial Stem Cells from the Adult Mouse Incisor

Understanding the cellular and molecular mechanisms that underlie tooth regeneration and renewal has become a topic of great interest1-4, and the mouse incisor provides a model for these processes. This remarkable organ grows continuously throughout the animal&#39;s life and generates all the necessary cell types from active pools of adult stem cells housed in the labial (toward the lip) and lingual (toward the tongue) cervical loop (CL) regions. Only the dental stem cells from the labial CL give rise to ameloblasts that generate enamel, the outer covering of teeth, on the labial surface. This asymmetric enamel formation allows abrasion at the incisor tip, and progenitors and stem cells in the proximal incisor ensure that the dental tissues are constantly replenished. The ability to isolate and grow these progenitor or stem cells in vitro allows their expansion and opens doors to numerous experiments not achievable in vivo, such as high throughput testing of potential stem cell regulatory factors. Here, we describe and demonstrate a reliable and consistent method to culture cells from the labial CL of the mouse incisor.

The continuously growing mouse incisor provides a model for studying renewal of dental tissues from dental epithelial stem cells (DESCs). A robust system for consistently and reliably obtaining these cells from the incisor and expanding them in vitro is reported here.

العزلة وثقافة الأسنان طلائي الخلايا الجذعية من الكبار ماوس القاطعة

Understanding the cellular and molecular mechanisms that underlie tooth regeneration and renewal has become a topic of great interest1-4, and the mouse ...

Isolation and Culture of Adult Mouse Cardiomyocytes for Cell Signaling and in vitro Cardiac Hypertrophy

Technological advances have made genetically modified mice, including transgenic and gene knockout mice, an essential tool in many research fields. Adult cardiomyocytes are widely accepted as a good model for cardiac cellular physiology and pathophysiology, as well as for pharmaceutical intervention. Genetically modified mice preclude the need for complicated cardiomyocyte infection processes to generate the desired genotype, which are inefficient due to cardiomyocytes&#8217; terminal differentiation. Isolation and culture of high quantity and quality functional cardiomyocytes will dramatically benefit cardiovascular research and provide an important tool for cell signaling transduction research and drug development. Here, we describe a well-established method for isolation of adult mouse cardiomyocytes that can be implemented with little training. The mouse heart is excised and cannulated to an isolated heart system, then perfused with a calcium-free and high potassium buffer followed by type II collagenase digestion in Langendorff retrograde perfusion mode. This protocol yields a consistent result for the collection of functional adult mouse cardiomyocytes from a variety of genetically modified mice.

Isolation and Culture of Adult Mouse Cardiomyocytes for Cell Signaling and in vitro Cardiac Hypertrophy

We describe a reliable method for isolation of adult mouse cardiomyocytes. This protocol yields a consistent result for the culture of functional adult cardiomyocytes from a variety of genetically modified mice.

العزلة وثقافة الكبار ماوس العضلية للاشارة الخليوي و<em&gt; في المختبر</em&gt; تضخم القلب

Technological advances have made genetically modified mice, including transgenic and gene knockout mice, an essential tool in many research fields. Adult ...

Isolation, Culture and Transduction of Adult Mouse Cardiomyocytes

Cultured cardiomyocytes can be used to study cardiomyocyte biology using techniques that are complementary to in vivo systems. For example, the purity and accessibility of in vitro culture enables fine control over biochemical analyses, live imaging, and electrophysiology. Long-term culture of cardiomyocytes offers access to additional experimental approaches that cannot be completed in short term cultures. For example, the in vitro investigation of dedifferentiation, cell cycle re-entry, and cell division has thus far largely been restricted to rat cardiomyocytes, which appear to be more robust in long-term culture. However, the availability of a rich toolset of transgenic mouse lines and well-developed disease models make mouse systems attractive for cardiac research. Although several reports exist of adult mouse cardiomyocyte isolation, few studies demonstrate their long-term culture. Presented here, is a step-by-step method for the isolation and long-term culture of adult mouse cardiomyocytes. First, retrograde Langendorff perfusion is used to efficiently digest the heart with proteases, followed by gravity sedimentation purification. After a period of dedifferentiation following isolation, the cells gradually attach to the culture and can be cultured for weeks. Adenovirus cell lysate is used to efficiently transduce the isolated cardiomyocytes. These methods provide a simple, yet powerful model system to study cardiac biology.

This protocol describes a step-by-step method for the reproducible isolation and long-term culture of adult mouse cardiomyocytes with high yield, purity, and viability.

العزلة والثقافة وتنبيغ الخلايا العضلية ماوس الكبار

Cultured cardiomyocytes can be used to study cardiomyocyte biology using techniques that are complementary to in vivo systems. For example, the purity and ...

Methods for the Isolation, Culture, and Functional Characterization of Sinoatrial Node Myocytes from Adult Mice

Sinoatrial node myocytes (SAMs) act as the natural pacemakers of the heart, initiating each heart beat by generating spontaneous action potentials (APs). These pacemaker APs reflect the coordinated activity of numerous membrane currents and intracellular calcium cycling. However the precise mechanisms that drive spontaneous pacemaker activity in SAMs remain elusive. Acutely isolated SAMs are an essential preparation for experiments to dissect the molecular basis of cardiac pacemaking. However, the indistinct anatomy, complex microdissection, and finicky enzymatic digestion conditions have prevented widespread use of acutely isolated SAMs. In addition, methods were not available until recently to permit longer-term culture of SAMs for protein expression studies. Here we provide a step-by-step protocol and video demonstration for the isolation of SAMs from adult mice. A method is also demonstrated for maintaining adult mouse SAMs in vitro and for expression of exogenous proteins via adenoviral infection. Acutely isolated and cultured SAMs prepared via these methods are suitable for a variety of electrophysiological and imaging studies.

Methods are demonstrated for the isolation of sinoatrial node myocytes (SAMs) from adult mice for patch clamp electrophysiology or imaging studies. Isolated cells can be used directly or can be maintained in culture to permit expression of proteins of interest, such as genetically encoded reporters.

طرق لعزل والثقافة، وتوصيف الوظيفي للالجيبية الأذينية عقدة Myocytes من الفئران الكبار

Sinoatrial node myocytes (SAMs) act as the natural pacemakers of the heart, initiating each heart beat by generating spontaneous action potentials (APs). ...

Isolation and Culture of Primary Mouse Keratinocytes from Neonatal and Adult Mouse Skin

The keratinocyte (KC) is the predominant cell type in the epidermis, the outermost layer of the skin. Epidermal KCs play a critical role in providing skin defense by forming an intact skin barrier against environmental insults, such as UVB irradiation or pathogens, and also by initiating an inflammatory response upon those insults. Here we describe methods to isolate KCs from neonatal mouse skin and from adult mouse tail skin. We also describe culturing conditions using defined growth supplements (dGS) in comparison to chelexed fetal bovine serum (cFBS). Functionally, we show that both neonatal and adult KCs are highly responsive to high calcium-induced terminal differentiation, tight junction formation and stratification. Additionally, cultured adult KCs are susceptible to UVB-triggered cell death and can release large amounts of TNF upon UVB irradiation. Together, the methods described here will be useful to researchers for the setup of in vitro models to study epidermal biology in the neonatal mouse and/or the adult mouse.

Epidermal keratinocytes form a functional skin barrier and are positioned at the front line of host defense against external environmental insults. Here we describe methods for isolation and primary culture of epidermal keratinocytes from neonatal and adult mouse skin, and induction of terminal differentiation and UVB-triggered inflammatory response from keratinocytes.

عزل وثقافة الكرياتينية الماوس الرئيسية من حديثي الولادة والفأر الكبار الجلد

The keratinocyte (KC) is the predominant cell type in the epidermis, the outermost layer of the skin. Epidermal KCs play a critical role in providing skin ...

Isolation, Characterization, and Differentiation of Cardiac Stem Cells from the Adult Mouse Heart

Myocardial infarction (MI) is a leading cause of morbidity and mortality around the world. A major goal of regenerative medicine is to replenish the dead myocardium after MI. Although several strategies have been used to regenerate myocardium, stem cell therapy remains a major approach to replenish the dead myocardium of an MI heart. Accumulating evidence suggests the presence of resident cardiac stem cells (CSCs) in the adult heart and their endocrine and/or paracrine effects on cardiac regeneration. However, CSC isolation and their characterization and differentiation toward myocardial cells, especially cardiomyocytes, remains a technical challenge. In the present study, we provided a simple method for the isolation, characterization, and differentiation of CSCs from the adult mouse heart. Here, we describe a density gradient method for the isolation of CSCs, where the heart is digested by a 0.2% collagenase II solution. To characterize the isolated CSCs, we evaluated the expression of CSCs/cardiac markers Sca-1, NKX2-5, and GATA4, and pluripotency/stemness markers OCT4, SOX2, and Nanog. We also determined the proliferation potential of isolated CSCs by culturing them in a Petri dish and assessing the expression of the proliferation marker Ki-67. For evaluating the differentiation potential of CSCs, we selected seven- to ten-days cultured CSCs. We transferred them to a new plate with a cardiomyocyte differentiation medium. They are incubated in a cell culture incubator for 12 days, while the differentiation medium is changed every three days. The differentiated CSCs express cardiomyocyte-specific markers: actinin and troponin I. Thus, CSCs isolated with this protocol have stemness and cardiac markers, and they have a potential for proliferation and differentiation toward cardiomyocyte lineage.

The overall goal of this article is to standardize the protocol for the isolation, characterization, and differentiation of cardiac stem cells (CSCs) from the adult mouse heart. Here, we describe a density gradient centrifugation method to isolate murine CSCs and elaborated methods for CSC culture, proliferation, and differentiation into cardiomyocytes.

عزل وتوصيف والتفريق في القلب من الخلايا الجذعية من قلب الماوس الكبار

Myocardial infarction (MI) is a leading cause of morbidity and mortality around the world. A major goal of regenerative medicine is to replenish the dead ...

Isolation and Characterization of Adult Cardiac Fibroblasts and Myofibroblasts

Cardiac fibrosis in response to injury is a physiological response to wound healing. Efforts have been made to study and target fibroblast subtypes that mitigate fibrosis. However, fibroblast research has been hindered due to the lack of universally acceptable fibroblast markers to identify quiescent as well as activated fibroblasts. Fibroblasts are a heterogenous cell population, making them difficult to isolate and characterize. The presented protocol describes three different methods to enrich fibroblasts and myofibroblasts from uninjured and injured mouse hearts. Using a standard and reliable protocol to isolate fibroblasts will enable the study of their roles in homeostasis as well as fibrosis modulation.

Obtaining a pure population of fibroblasts is crucial to studying their role in wound repair and fibrosis. Described here is a detailed method to isolate fibroblasts and myofibroblasts from uninjured and injured mouse hearts followed by characterization of their purity and functionality by immunofluorescence, RTPCR, fluorescence-assisted cell sorting, and collagen gel contraction.

عزل وتوصيف الخلايا الليفية القلبية للبالغين والخلايا الليفية الليفية

Cardiac fibrosis in response to injury is a physiological response to wound healing. Efforts have been made to study and target fibroblast subtypes that ...