Name: anaerobic growth and maintenance of mammalian cell lines
Uploaded: 2018-07-21T15:00:00.000+00:00
Duration: 7 min 15 s
Description: Watch this Scientific Journal Video about Anaerobic Growth and Maintenance of Mammalian Cell Lines at JoVE.com

著者は、中西部大学研究局と主催のプログラムを彼らのサポートありがちましょう。

1. 様々 な哺乳類細胞の嫌気性培養メディアを準備します。
<ol><li>
無酸素細胞文化25の完全な PS 74656 培地を作る。
	<ol>
<li>50 mL の脱イオン水の亜硝酸イオンの 17.25 g を溶解して滅菌亜硝酸原液 (100 x; 5 M) をし、それを殺菌するためにフィルターします。</li>
		<li>110 mg/l L-グルタミン, ピルビン酸ナトリウムと 10% 牛胎児血清の 584 mg/L の低血糖 DMEM (ブドウ糖 1 g/L) の 50 mL あたり亜硝酸原液 0.5 mL を追加 (FBS; 熱不活化)。 注: FBS 濃度の有毒である 10% より大きいを使用します。テスト 1 つの癌細胞株は、注意、亜硝酸 (MDA MB 231、乳房癌細胞株) メディアの低い耐性を持っていたおよび、したがって、亜硝酸の不在で育った。さらに、PS 74656 媒体が調べたすべての細胞の成長をサポートされています (n = 9)、特定の細胞 (例えば、Vero 細胞) 細胞濃度が高いと高グルコース (4.5 g/L) DMEM で PS 74656 で高い生存率を展示25。</li>
		<li>滅菌 50 mL の円錐形遠心チューブに培地 20 mL を配置します。キャップを締めていません。それは緩いはずです。</li>
		<li>解消ガス媒体で真空ベルジャーのチューブを配置することによって、約 45 ° の角度、媒体の表面積を最大化します。</li>
		<li>部屋の温度で真空ポンプまたは同等のものを使用して真空を適用します。泡はもはや観察されない (24-48 h) までの真空を維持します。</li>
		<li>瓶からチューブを削除し、すぐに中への空気の導入を最小限に抑えるためにチューブをシール、キャップを閉じます。</li>
		<li>商業の嫌気性チャンバーにチューブを置きます。</li>
		<li>チューブ キャップを緩めます、少なくとも 24 のチャンバーで嫌気性ガスの混合物の平衡にチューブの大気を許可します。 注: それが尽きるまで媒体を商工会議所でなければなりません。チューブ フラッシュ プロセスをスピードアップするため無菌転送ピペットを使用して無酸素ガスの混合物で、少なくとも 3 倍は N、CO2H2から成っている無酸素ガスでいっぱい2.
</li>
		<li>ドガのすべてのチューブ (10-15 分) チューブをシールし、商工会議所に配置する前に上記のように嫌気性チャンバーに配置されます。使用前に 1.5-2 mL 転送ピペットを使用して嫌気性ガス混合物でそれをフラッシュまたは無酸素とフラッシュされているピペット チップ ガスの少なくとも 3 倍。</li>
		<li>嫌気性チャンバー中に無菌脱マイクロ遠心チューブに媒体の 100 μ L を取り外してチャンバ内酸素プローブを使用して溶存酸素のそれをテストします。 注: 使用する前に製造元のプロトコルあたり酸素電極を校正します。正しく脱と釣合い嫌気性ガス混合メディアの測定値は、0 です。 注意: 測定値は 0.3 ppm の酸素を超えている場合、は、媒体を平衡により多くの時間が必要なので、嫌気性チャンバー内のメディアをインキュベートに進みます。</li>
	</ol>
</li>
</ol>2. 無酸素栽培様々 な哺乳類細胞株の
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指定日-1
	<ol>
<li>3 24 ウェル プレート (80% 合流) の細胞の十分な数を提供するために、無酸素と比較文化コントロールの 90% 合流 T150 フラスコ細胞 (37 ° C、5% CO2) に拡張します。 注: この研究では、hela 細胞 229 および Vero 細胞が使用されました。このプロトコルはまたテストされ、成長と 7 他のセル行の25の維持の成功した証明しました。</li>
		<li>吸引フラスコからメディアを取り出して、HBSS の 10 mL で洗います。</li>
		<li>5 ml のトリプシンのメディアを交換してください。セルを視覚的にプラスチックからデタッチするまでフラスコを揺り動かしなさい。、トリプシンのほとんどを吸引し、付着性のセルを除去するフラスコをタップします。高グルコース DMEM の 10-15 mL を追加 (グルコースの 4.5 g/L、110 mg/L L-グルタミン、10% の FBS、ゲンタマイシンの 50 μ g/mL)、トリプシンを不活化します。</li>
		<li>すべての細胞塊が中断されるまでに 25 mL のピペットを使用してセルをカップ刻んだ。</li>
		<li>セルをカウントし、標準検定とトリパン ブルー色素排除法または自動化された同等を使用しての生存率を決定します。</li>
		<li>105セル/mL x 2.24 の細胞密度に前述高グルコース DMEM でセルを中断します。</li>
		<li>24 ウェル培養プレートの井戸に細胞懸濁液の 1 mL を追加 (2.24 105細胞/ウェル x; 80% 合流)。無酸素の文化とコントロールによる文化の種子 1 プレート。</li>
		<li>37 ° c 24 時間の培養条件 (5% CO2) で細胞を孵化させなさい。 注: セルは様々 な広い板にめっきすることが。ただし、フラスコで無酸素成長細胞を播種酸素導入のリスク システムにしない限り実行されます注意を払っても大気中のガスを完全に洗い流す。</li>
	</ol>
</li>
	<li>
指定日 0
	<ol>
<li>平地培養 24 時間後嫌気性チャンバー (商業嫌気性チャンバー; に無酸素の成長のためプレートを配置します。図 1)。</li>
		<li>嫌気性チャンバーに 24 h の順応されて抗生物質無料デ熟れた PS 74656 媒体媒体をすぐに変更します。湿ったタオルでコンテナーにプレートを置き、疎の水分を維持するためにそれを閉じます。 注: 平地成長板は、すべての治療法は、大気条件下で行われる例外と同じ処理されます。</li>
	</ol>
</li>
	<li>
指定日 1
	<ol>
<li>嫌気培養の日 1 は 80% N210% の CO210% H2の標準的な嫌気性ガスの混合物が含まれている商業嫌気性チャンバー内培養 24 時間後に開始します。</li>
		<li>さまざまな期間のセルを孵化させなさい。</li>
		<li>時間の経過とともに色の変化のための嫌気性媒体を監視します。 注: 赤オレンジからマゼンタの色シフト信号媒体を変更する時間です。発生するまで 2 週間かかるでしょう。赤オレンジ黄色に培地の色ずれは、細菌汚染の存在を示します。</li>
		<li>中は赤からマゼンタに変化したときは、吸引して無宿者セルを削除します。</li>
		<li>脱、嫌気性ガスの無宿者のセルと吸気の使用済みメディアの場所平衡 microfuge の管、シールが、安全性し、遠心分離 (326 gx; 10 分室温で) それらを確認しながらチューブを閉じます。すぐに井戸に吸気のメディアを新しい脱 PS 74656 培地 1 mL に置き換えます。</li>
		<li>それを開く前に嫌気性チャンバーに播種細胞および microfuge の管を配置します。使用済み培地を吸引し、新鮮な脱 PS 74656 媒体および microfuge の管に 1 ml の容量にそれを置き換えます。</li>
		<li>24 ウェル培養プレートのウェルにチューブからセルを返します。</li>
	</ol>
</li>
</ol>3. 嫌気培養細胞の顕微鏡観察による形質細胞分化の評価
<ol><li>
嫌気性チャンバー中に嫌気性ガス混合物でフラッシュ ジッパーのついた箱にプレートを置き、ボックスを閉じます。 注: ボックスは、水分を提供し乾燥からプレートを防ぐため少し湿らせたペーパー タオルを含める必要があります。顕微鏡、サンドイッチ袋は薄いのでベストを働かせます。<ol>
<li>顕微鏡下で細胞を観察するには、完全に袋を密封、商工会議所から取り外します。</li>
		<li>慎重に板の上にビニール袋を伸ばす、カメラの添付ファイルと逆の位相差顕微鏡を使用してセルを調べて、様々 な倍率での顕微鏡写真を取る。</li>
		<li>室に密封された袋のプレートを戻り、2.3.3 の手順で説明するように孵化を続けます。</li>
		<li>懸濁液にある無宿者セルを別々 に分析するには、手順 2.3.5 に示した手順に従います。 注: これらの細胞は好気性をテストするため使用できます。 または嫌気性チャンバーこの人口の特定のテストのために返すことができます。</li>
	</ol>
</li>
</ol>

<ol>
	<li>Cummins, J., Tangney, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bacteria+and+tumours:+causative+agents+or+opportunistic+inhabitants?.">Bacteria and tumours: causative agents or opportunistic inhabitants?.</a> Infectious Agents and Cancer. 8 (1), 11 (2013).</li><li>Liao, J., 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=Enhanced+efficiency+of+generating+induced+pluripotent+stem+(iPS)+cells+from+human+somatic+cells+by+a+combination+of+six+transcription+factors.">Enhanced efficiency of generating induced pluripotent stem (iPS) cells from human somatic cells by a combination of six transcription factors.</a> Cell Research. 18 (5), 600-603 (2008).</li><li>Park, I. H., Lerou, P. H., Zhao, R., Huo, H., Daley, G. Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+human-induced+pluripotent+stem+cells.">Generation of human-induced pluripotent stem cells.</a> Nature Protocols. 3 (7), 1180-1186 (2008).</li><li>Semenza, G. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=HIF-1:+mediator+of+physiological+and+pathophysiological+responses+to+hypoxia.">HIF-1: mediator of physiological and pathophysiological responses to hypoxia.</a> Journal of Applied Physiology. 88 (4), 1474-1480 (2000).</li><li>Biedermann, L., Rogler, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+intestinal+microbiota:+its+role+in+health+and+disease.">The intestinal microbiota: its role in health and disease.</a> European Journal of Pediatrics. 174 (2), 151-167 (2015).</li><li>Cui, X. 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=Hypoxia+influences+stem+cell-like+properties+in+multidrug+resistant+K562+leukemic+cells.">Hypoxia influences stem cell-like properties in multidrug resistant K562 leukemic cells.</a> Blood Cells, Molecules, and Diseases. 51 (3), 177-184 (2013).</li><li>Richter, A., Sanford, K. K., Evans, V. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+Oxygen+and+Culture+Media+on+Plating+Efficiency+of+Some+Mammalian+Tissue+Cells.">Influence of Oxygen and Culture Media on Plating Efficiency of Some Mammalian Tissue Cells.</a> JNCI: Journal of the National Cancer Institute. 49 (6), 1705-1712 (1972).</li><li>Packer, L., Fuehr, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Low+oxygen+concentration+extends+the+lifespan+of+cultured+human+diploid+cells.">Low oxygen concentration extends the lifespan of cultured human diploid cells.</a> Nature. 267, 423 (1977).</li><li>Warburg, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=On+the+origin+of+cancer.">On the origin of cancer.</a> Science. 123 (3191), 309-314 (1956).</li><li>D&#246;me, B., Hendrix, M. J. C., Paku, S., T&#243;v&#225;ri, J., T&#237;m&#225;r, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alternative+Vascularization+Mechanisms+in+Cancer.">Alternative Vascularization Mechanisms in Cancer.</a> The American Journal of Pathology. 170 (1), 1-15 (2007).</li><li>Jewell, U. 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=Induction+of+HIF-1&#945;+in+response+to+hypoxia+is+instantaneous.">Induction of HIF-1&#945; in response to hypoxia is instantaneous.</a> The FASEB Journal. 15 (7), 1312-1314 (2001).</li><li>Lee, J. W., Bae, S. H., Jeong, J. W., Kim, S. H., Kim, K. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hypoxia-inducible+factor+(HIF-1)alpha:+its+protein+stability+and+biological+functions.">Hypoxia-inducible factor (HIF-1)alpha: its protein stability and biological functions.</a> Experimental &amp; Molecular Medicine. 36 (1), 1-12 (2004).</li><li>Pagoulatos, D., Pharmakakis, N., Lakoumentas, J., Assimakopoulou, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Etaypoxia-inducible+factor-1alpha,+von+Hippel-Lindau+protein,+and+heat+shock+protein+expression+in+ophthalmic+pterygium+and+normal+conjunctiva.">Etaypoxia-inducible factor-1alpha, von Hippel-Lindau protein, and heat shock protein expression in ophthalmic pterygium and normal conjunctiva.</a> Molecular Vision. 20, 441-457 (2014).</li><li>Park, T. J., 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=Fructose-driven+glycolysis+supports+anoxia+resistance+in+the+naked+mole-rat.">Fructose-driven glycolysis supports anoxia resistance in the naked mole-rat.</a> Science. 356 (6335), 307 (2017).</li><li>Johnston, I. A., Bernard, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Utilization+of+the+Ethanol+Pathway+in+Carp+Following+Exposure+to+Anoxia.">Utilization of the Ethanol Pathway in Carp Following Exposure to Anoxia.</a> Journal of Experimental Biology. 104 (1), 73 (1983).</li><li>Takahashi, E., Sato, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anaerobic+respiration+sustains+mitochondrial+membrane+potential+in+a+prolyl+hydroxylase+pathway-activated+cancer+cell+line+in+a+hypoxic+microenvironment.">Anaerobic respiration sustains mitochondrial membrane potential in a prolyl hydroxylase pathway-activated cancer cell line in a hypoxic microenvironment.</a> American Journal of Physiology-Cell Physiology. 306 (4), C334-C342 (2014).</li><li>Mandziuk, S., 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=Effect+of+hypoxia+on+toxicity+induced+by+anticancer+agents+in+cardiomyocyte+culture.">Effect of hypoxia on toxicity induced by anticancer agents in cardiomyocyte culture.</a> Medycyna Weterynaryjna. 70 (5), 287-291 (2014).</li><li>Arai, H., Kodama, T., Igarashi, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+the+nirQOP+genes+in+energy+conservation+during+anaerobic+growth+of+Pseudomonas+aeruginosa.">The role of the nirQOP genes in energy conservation during anaerobic growth of Pseudomonas aeruginosa.</a> Bioscience, Biotechnology, and Biochemistry. 62 (10), 1995-1999 (1998).</li><li>M&#252;ller, 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=Biochemistry+and+Evolution+of+Anaerobic+Energy+Metabolism+in+Eukaryotes.">Biochemistry and Evolution of Anaerobic Energy Metabolism in Eukaryotes.</a> Microbiology and Molecular Biology Reviews. 76 (2), 444-495 (2012).</li><li>Gupta, K. J., Igamberdiev, A. U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+anoxic+plant+mitochondrion+as+a+nitrite:+NO+reductase.">The anoxic plant mitochondrion as a nitrite: NO reductase.</a> Mitochondrion. 11 (4), 537-543 (2011).</li><li>Gupta, K. J., Igamberdiev, A. U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reactive+Nitrogen+Species+in+Mitochondria+and+Their+Implications+in+Plant+Energy+Status+and+Hypoxic+Stress+Tolerance.">Reactive Nitrogen Species in Mitochondria and Their Implications in Plant Energy Status and Hypoxic Stress Tolerance.</a> Frontiers in Plant Science. 7 (369), (2016).</li><li>Kozlov, A. V., Staniek, K., Nohl, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nitrite+reductase+activity+is+a+novel+function+of+mammalian+mitochondria.">Nitrite reductase activity is a novel function of mammalian mitochondria.</a> FEBS letters. 454 (1-2), 127-130 (1999).</li><li>Nohl, H., Staniek, K., Kozlov, A. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+existence+and+significance+of+a+mitochondrial+nitrite+reductase.">The existence and significance of a mitochondrial nitrite reductase.</a> Redox Report. 10 (6), 281-286 (2005).</li><li>Lawson, J. C., Blatch, G. L., Edkins, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cancer+stem+cells+in+breast+cancer+and+metastasis.">Cancer stem cells in breast cancer and metastasis.</a> Breast Cancer Research and Treatment. 118 (2), 241-254 (2009).</li><li>Plotkin, B. J., 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+method+for+the+long-term+cultivation+of+mammalian+cells+in+the+absence+of+oxygen:+Characterization+of+cell+replication,+hypoxia-inducible+factor+expression+and+reactive+oxygen+species+production.">A method for the long-term cultivation of mammalian cells in the absence of oxygen: Characterization of cell replication, hypoxia-inducible factor expression and reactive oxygen species production.</a> Tissue and Cell. 50 (Supplement C), 59-68 (2018).</li><li>Sarmento, B., 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=Cell-based+in+vitro+models+for+predicting+drug+permeability.">Cell-based in vitro models for predicting drug permeability.</a> Expert Opinion on Drug Metabolism &amp; Toxicology. 8 (5), 607-621 (2012).</li><li>Collins, S. J., Gallo, R. C., Gallagher, R. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Continuous+growth+and+differentiation+of+human+myeloid+leukaemic+cells+in+suspension+culture.">Continuous growth and differentiation of human myeloid leukaemic cells in suspension culture.</a> Nature. 270 (5635), 347-349 (1977).</li><li>Plotkin, B. J., 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+method+for+the+long-term+cultivation+of+mammalian+cells+in+the+absence+of+oxygen:+Characterization+of+cell+replication,+hypoxia-inducible+factor+expression+and+reactive+oxygen+species+production.">A method for the long-term cultivation of mammalian cells in the absence of oxygen: Characterization of cell replication, hypoxia-inducible factor expression and reactive oxygen species production.</a> Tissue and Cell. 50, 59-68 (2018).</li></ol>

著者が明らかに何もありません。

このメソッドは、酸素のない状態で長期間培養初めて哺乳類セルを表します。現在の観測に基づいて、無酸素成長機能を介して非発酵的経路嫌気性成長が表現型の相違で起因した哺乳類細胞ラインの28日間で普遍的なようであります。これはテストしたすべての細胞の観察されました。嫌気培養細胞の比率を増やす丸くなった、懸濁液のような人口を開発し、レプリ?...

固形腫瘍、幹細胞ニッチと粘膜の表面を裏から細胞が低酸素レベル、無酸素1,2,3、4を含む環境で存在します。通常の生理学的状態の酸素は低酸素血症低酸素 (酸素の完全な欠如)5,6のそれを越える異なります。大気中の酸素に悪影響哺乳類細胞複製劣化酸素条件下での in vitro細胞成長を最適化できる実現は、1970 年代初頭に報告されました。リヒターら7は、酸素レベル (低酸素) の 1-3% が大気中の酸素 (20%) と比較してめっき効率を高めることを示した。ヒト二倍体細胞の寿命は、低酸素培養条件8にも拡張されます。
酸素店が劣化 (例えば、激しい運動中に)生体内で、低酸素の条件が発生する発酵 (嫌気呼吸) に好気性の呼吸からの骨格筋における ATP の生産をスイッチにより前記の乳酸9の最終製品。悪性腫瘍、内部の病理、腫瘍の質量は貧しい血管新生による無酸素に低酸素10。義務づける嫌気性菌1によって植民地化腫瘍インテリアで腫瘍内部に限られた血流の影響は独立して検証します。機械論的に、低酸素の腫瘍細胞の生存は、低酸素誘導因子 1 α 遺伝子の発現に依存すると考えられる (HIF1-アルファ)、低酸素4,11初期の自発的な応答であります。,12. HIF1-アルファ、 HIF1をバインド熱ショックタンパク質に起因する低酸素の条件の下で-アルファ プロモーターと遺伝子転写12活性を上昇させる。これら熱ショック蛋白質は腫瘍低酸素環境で見られる様々 な表現型の誘導を支援するためと考えられています。これらの表現型は、細胞膜糖輸送担体の発現増加と解糖系 (Warburg 効果)13の率を表わします。その結果、乳酸発酵するミトコンドリアの酸化的リン酸化からスイッチです。
無酸素の生存はまた生存現象14,15をサポートするグルコースに選択肢を利用できます。最高の学び哺乳類の例はほくろのラットは、果糖駆動糖発酵経路14を介して酸素なしにほぼ 20 分間生き残ることができます。特定の魚の代替の適応が発生します (例えば、大幅に長く生き残ることができる鯉 [フナsp] の期間にターミナルの副産物としてエタノールと解糖系を使用して)15。両方のケースで発酵は酸素のない状態での生存を可能に代謝を駆動します。無酸素の生存のための現在の仮説は、長い間HIF1として-16嫌気的条件下で発生する酸素の必要性なしに、低酸素症、ミトコンドリア呼吸時にアルファがアクティブします。さらに、細胞は細胞生存17に有害であることを証明できる酸化ストレスを避けるので発酵経路の低酸素・無酸素の生存のための使用が腫瘍の生存率を高めることを仮定します。この仮説は、心筋、低酸素血症が腫瘍細胞17は、酸化ストレスを減少させることを示す最近の研究によってサポートされます。
日には、無酸素の哺乳類細胞の生存のための発酵経路の本質染み付いている文献、ため、できないこと 3 日以上酸素の完全な欠如に哺乳類細胞には、大部分の。しかし、細菌の嫌気性の生存のためには、解糖系の代わりに発生します。ある特定の細菌の窒素または硫酸 (他の化合物) の間では、酸素18の不在のチトクロム酸化酵素システムのターミナル電子アクセプターとして使用できます。ミトコンドリアが細胞にエネルギーを提供する 154 万 2000 年19 海の酸素化前に、の最後の普遍的な共通の祖先から分岐後並行発生した細菌や真核生物の進化が推定されています。.研究者は、分離ミトコンドリアがターミナル電子アクセプターとして亜硝酸の酸素の存在下で ATP を作り出すことができることを示しているので細胞が酸素のない状態で 3 日間よりも長い期間にわたって機能すると仮定するが妥当です。20,21,22,23. 本研究の方法論は多数の細胞の嫌気性の哺乳類細胞の成長のためのユーティリティ。

<table><tbody><tr><td>Whitney A35 anaerobic chamber</td><td>Don Whitley Scientific&amp;nbsp;</td><td>Microbiology International</td><td>The ability to remove the front of the chamber makes it easy to put instruments inside without concern about getting them through the portals.</td></tr><tr><td>CO&lt;sub&gt;2&lt;/sub&gt; incubator</td><td>Fisher Scientific</td><td>3531</td><td></td></tr><tr><td>Tissue Culture Hood</td><td>Labconco</td><td>DO55735</td><td></td></tr><tr><td>pipet aid</td><td>Drummond</td><td>4-000-100</td><td></td></tr><tr><td>sterile transfer pipets</td><td>Santa Cruz</td><td>sc-358867</td><td></td></tr><tr><td>50 cc sterile conical centrifuge tubes</td><td>DOT</td><td>451-PG</td><td></td></tr><tr><td>vaccum jar</td><td>Nalgene 111410862889</td><td>BTA-Mall 5311-0250</td><td></td></tr><tr><td>DMEM high glucose (4.5 g/L)</td><td>CellGro</td><td>10-017-CM</td><td></td></tr><tr><td>DMEM low glucose (1 g/L)</td><td>CellGro</td><td>10-014-CV</td><td></td></tr><tr><td>FBS</td><td>VWR</td><td>1500-500</td><td></td></tr><tr><td>HBSS</td><td>VWR</td><td>20-021-CV</td><td></td></tr><tr><td>trypsin</td><td>VWR</td><td>25-052-CI</td><td></td></tr><tr><td>gentamicin</td><td>Gibco</td><td>15750-060</td><td></td></tr><tr><td>sterile pipets</td><td>CellTreat</td><td>229210B, 229205B</td><td></td></tr><tr><td>Tissue culture flask (T75 or T150)</td><td>Santa Cruz</td><td>sc-200263, sc-200264</td><td></td></tr><tr><td>24 well tissue culture treated dishes</td><td>DOT</td><td>667124</td><td></td></tr><tr><td>glad ziplock sandwich bags</td><td>Ziploc</td><td>Costco</td><td></td></tr><tr><td>inverted phase microscope (10, 20 40x objectives with camera mont)</td><td>Nikon Eclipse</td><td>TS100</td><td></td></tr><tr><td>trypan blue</td><td>Invitrogen</td><td>T10282</td><td></td></tr><tr><td>hemocytometer</td><td>Invitrogen</td><td>C10283</td><td></td></tr><tr><td>Countess Automated Cell Counter</td><td>Life Technologies</td><td>AMQAF1000</td><td></td></tr><tr><td>Rainin microtiter pipets</td><td>Mettler Toledo</td><td>Rainin Classic Pipette PR-100</td><td></td></tr><tr><td>microtiter tips</td><td>Santa Cruz Biotechnology</td><td>sc-213233 &amp;amp; sc-201717&amp;nbsp;</td><td></td></tr><tr><td>test tube rack (50cc tubes)</td><td>The Lab Depot</td><td>HS29050A</td><td></td></tr><tr><td>sodium nitrite</td><td>Sigma</td><td>https://www.sigmaaldrich.com/catalog/product/sigald/237213?lang=en&amp;amp;region=US</td><td></td></tr><tr><td>clear boxes with lids</td><td>Rosti Mepal</td><td>Rubber maid</td><td></td></tr><tr><td>paper towels</td><td>In House</td><td></td><td></td></tr><tr><td>cell scraper</td><td>CellTreat</td><td>229310</td><td></td></tr><tr><td>PBS</td><td>In house prepared</td><td></td><td></td></tr><tr><td>70% Ethanol</td><td>Fisher Scientific</td><td>64-17-5</td><td></td></tr><tr><td>microcentrifuge tubes sterile</td><td>Santa Cruz</td><td>sc-200273</td><td></td></tr><tr><td>biohazard bags with holder (desktop)</td><td>Heathrow Scientific</td><td>HS10320</td><td></td></tr><tr><td>Nitrile Gloves</td><td>VWR</td><td>89428</td><td></td></tr><tr><td>oxygen electrode</td><td>eDAQ</td><td>EPO354</td><td></td></tr><tr><td>pH meter</td><td>Jenway</td><td>3510</td><td></td></tr><tr><td>pH paper/ pHydrion</td><td>Sigma Aldich</td><td>Z111813</td><td></td></tr></tbody></table>

anaerobic growth and maintenance of mammalian cell lines

腫瘍と幹細胞ニッチにおける中点と共に最も粘膜表面が無酸素体の地理的な領域です。先行研究により平地 (空気 5% CO2 ) で孵化が示す低酸素または限られた可能性 (2-3 日) で無酸素インキュベーション (遊離酸素の不在) 結果続く条件 (低酸素のレベル)。小説の方法論は、(少なくとも 17 日; 最大時間テスト) の無酸素の培養を可能にする開発されました。この手法の最も重要な側面は、システムに酸素が生じないことを確認することです。これは、脱ガス、メディア、フラッシュ チューブや料理、フラスコ、ピペット、嫌気性ガスの混合物 (H2CO2N2) と無酸素 (無酸素) 環境に釣り合うために材料を許可することによって続いてによって得られる使用する前に。ときに得られる顕微鏡写真には、成果物が含まれていないことを確認するため顕微鏡写真を取得、追加のケアを行使しなければなりません。酸素のない場合は、細胞の形態を大幅に変更します。嫌気成長のすべてのセルの 2 つの異なる型が記載されています。成長し、酸素のない状態で哺乳類の細胞を維持する能力は、細胞生理学、モデルの相互作用、および小説がん新薬開発の生合成経路の特性の解析に適用できます。

このプロトコルの強さには、長寿と複数の細胞の成長のサポートと深遠な変化と細胞形態25の発散があることの認識があります。この研究の最も重要な要素は、転送と厳密な嫌気下で細胞の維持です。これは、嫌気性チャンバー構成プロトコル (図 1) を最大化する顕微鏡または他のテストのいずれかのセルが削除される保証は?...

Watch this Scientific Journal Video about Anaerobic Growth and Maintenance of Mammalian Cell Lines at JoVE.com

Anaerobic Growth and Maintenance of Mammalian Cell Lines

哺乳類セルラインの嫌気性成長と維持

ここでは、確立されたセルラインの嫌気の長期培養を可能にする新しい手法について述べる。テストされた最大生存期間は 17 日です。このメソッドは、細胞毒性薬のテストと、anoxically 複製細胞の生理機能の探索に適しています。

Most mucosal surfaces along with the midpoints in tumors and stem cell niches are geographic areas of the body that are anoxic. Previous studies show that ...

anaerobic-growth-and-maintenance-of-mammalian-cell-lines

Research

JoVE Journal

Biology

10.0K ビュー.  Midwestern University. ここでは、確立されたセルラインの嫌気の長期培養を可能にする新しい手法について述べる。テストされた最大生存期間は 17 日です。このメソッドは、細胞毒性薬のテストと、anoxically 複製細胞の生理機能の探索に適しています。

ビデオ: Anaerobic Growth and Maintenance of Mammalian Cell Lines

An In Vitro System to Study Tumor Dormancy and the Switch to Metastatic Growth

Recurrence of breast cancer often follows a long latent period in which there are no signs of cancer, and metastases may not become clinically apparent until many years after removal of the primary tumor and adjuvant therapy. A likely explanation of this phenomenon is that tumor cells have seeded metastatic sites, are resistant to conventional therapies, and remain dormant for long periods of time 1-4.
The existence of dormant cancer cells at secondary sites has been described previously as quiescent solitary cells that neither proliferate nor undergo apoptosis 5-7. Moreover, these solitary cells has been shown to disseminate from the primary tumor at an early stage of disease progression 8-10 and reside growth-arrested in the patients' bone marrow, blood and lymph nodes 1,4,11. Therefore, understanding mechanisms that regulate dormancy or the switch to a proliferative state is critical for discovering novel targets and interventions to prevent disease recurrence. However, unraveling the mechanisms regulating the switch from tumor dormancy to metastatic growth has been hampered by the lack of available model systems.
in vivo and ex vivo model systems to study metastatic progression of tumor cells have been described previously 1,12-14. However these model systems have not provided in real time and in a high throughput manner mechanistic insights into what triggers the emergence of solitary dormant tumor cells to proliferate as metastatic disease. We have recently developed a 3D in vitro system to model the in vivo growth characteristics of cells that exhibit either dormant (D2.OR, MCF7, K7M2-AS.46) or proliferative (D2A1, MDA-MB-231, K7M2) metastatic behavior in vivo . We demonstrated that tumor cells that exhibit dormancy in vivo at a metastatic site remain quiescent when cultured in a 3-dimension (3D) basement membrane extract (BME), whereas cells highly metastatic in vivo readily proliferate in 3D culture after variable, but relatively short periods of quiescence. Importantly by utilizing the 3D in vitro model system we demonstrated for the first time that the ECM composition plays an important role in regulating whether dormant tumor cells will switch to a proliferative state and have confirmed this in in vivo studies15-17. Hence, the model system described in this report provides an in vitro method to model tumor dormancy and study the transition to proliferative growth induced by the microenvironment.

An In Vitro System to Study Tumor Dormancy and the Switch to Metastatic Growth

A modified 3-D in vitro system is presented in which growth characteristics of several tumor cell lines in reconstituted basement membrane correlate with the dormant or proliferative behavior of the tumor cells at a metastatic secondary site in vivo.

アンインビトロ</em転移性増殖に腫瘍の休眠とスイッチを研究する>システム

Recurrence of breast cancer often follows a long latent period in which there are no signs of cancer, and metastases may not become clinically apparent ...

医学

FIBS-enabled Noninvasive Metabolic Profiling

In the era of computational biology, new high throughput experimental systems are necessary in order to populate and refine models so that they can be validated for predictive purposes. Ideally such systems would be low volume, which precludes sampling and destructive analyses when time course data are to be obtained. What is needed is an in situ monitoring tool which can report the necessary information in real-time and noninvasively. An interesting option is the use of fluorescent, protein-based in vivo biological sensors as reporters of intracellular concentrations. One particular class of in vivo biosensors that has found applications in metabolite quantification is based on F&#246;rster Resonance Energy Transfer (FRET) between two fluorescent proteins connected by a ligand binding domain. FRET integrated biological sensors (FIBS) are constitutively produced within the cell line, they have fast response times and their spectral characteristics change based on the concentration of metabolite within the cell. In this paper, the method for constructing Chinese hamster ovary (CHO) cell lines that constitutively express a FIBS for glucose and glutamine and calibrating the FIBS in vivo in batch cell culture in order to enable future quantification of intracellular metabolite concentration is described. Data from fed-batch CHO cell cultures demonstrates that the FIBS was able in each case to detect the resulting change in the intracellular concentration. Using the fluorescent signal from the FIBS and the previously constructed calibration curve, the intracellular concentration was accurately determined as confirmed by an independent enzymatic assay.

A description of how to calibrate F&#246;rster Resonance Energy Transfer integrated biological sensors (FIBS) for in situ metabolic profiling is presented.&#160;The FIBS can be used to measure intracellular levels of metabolites noninvasively aiding in the development of metabolic models and high throughput screening of bioprocess conditions.

FIBS対応の非侵襲メタボリック·プロファイリング

In the era of computational biology, new high throughput experimental systems are necessary in order to populate and refine models so that they can be ...

生物工学

Atomic Absorbance Spectroscopy to Measure Intracellular Zinc Pools in Mammalian Cells

Transition metals are essential micronutrients for organisms but can be toxic to cells at high concentrations by competing with physiological metals in proteins and generating redox stress. Pathological conditions that lead to metal depletion or accumulation are causal agents of different human diseases. Some examples include anemia, acrodermatitis enteropathica, and Wilson&#8217;s and Menkes&#8217; diseases. It is therefore important to be able to measure the levels and transport of transition metals in biological samples with high sensitivity and accuracy in order to facilitate research exploring how these elements contribute to normal physiological functions and toxicity. Zinc (Zn), for example, is a cofactor in many mammalian proteins, participates in signaling events, and is a secondary messenger in cells. In excess, Zn is toxic and can inhibit absorption of other metals, while in deficit, it can lead to a variety of potentially lethal conditions.

Graphite furnace atomic absorption spectroscopy (GF-AAS) provides a highly sensitive and effective method for determining Zn and other transition metal concentrations in diverse biological samples. Electrothermal atomization via GF-AAS quantifies metals by atomizing small volumes of samples for subsequent selective absorption analysis using wavelength of excitation of the element of interest. Within the limits of linearity of the Beer-Lambert Law, the absorbance of light by the metal is directly proportional to concentration of the analyte. Compared to other methods of determining Zn content, GF-AAS detects both free and complexed Zn in proteins and possibly in small intracellular molecules with high sensitivity in small sample volumes. Moreover, GF-AAS is also more readily accessible than inductively coupled plasma mass spectrometry (ICP-MS) or synchrotron-based X-ray fluorescence. In this method, the systematic sample preparation of different cultured cell lines for analyses in a GF-AAS is described. Variations in this trace element were compared in both whole cell lysates and subcellular fractions of proliferating and differentiated cells as proof of principle.

Cultured primary or established cell lines are commonly used to address fundamental biological and mechanistic questions as an initial approach before using animal models. This protocol describes how to prepare whole cell extracts and subcellular fractions for studies of zinc (Zn) and other trace elements with atomic absorbance spectroscopy.

哺乳類細胞内の細胞内亜鉛プールを測定するための原子吸光度分光法

Transition metals are essential micronutrients for organisms but can be toxic to cells at high concentrations by competing with physiological metals in ...

生化学

Cultivation of Mammalian Cells Using a Single-use Pneumatic Bioreactor System

Recent advances in mammalian, insect, and stem cell cultivation and scale-up have created tremendous opportunities for new therapeutics and personalized medicine innovations. However, translating these advances into therapeutic applications will require in vitro systems that allow for robust, flexible, and cost effective bioreactor systems. There are several bioreactor systems currently utilized in research and commercial settings; however, many of these systems are not optimal for establishing, expanding, and monitoring the growth of different cell types. The culture parameters most challenging to control in these systems include, minimizing hydrodynamic shear, preventing nutrient gradient formation, establishing uniform culture medium aeration, preventing microbial contamination, and monitoring and adjusting culture conditions in real-time. Using a pneumatic single-use bioreactor system, we demonstrate the assembly and operation of this novel bioreactor for mammalian cells grown on micro-carriers. This bioreactor system eliminates many of the challenges associated with currently available systems by minimizing hydrodynamic shear and nutrient gradient formation, and allowing for uniform culture medium aeration. Moreover, the bioreactor&#8217;s software allows for remote real-time monitoring and adjusting of the bioreactor run parameters. This bioreactor system also has tremendous potential for scale-up of adherent and suspension mammalian cells for production of a variety therapeutic proteins, monoclonal antibodies, stem cells, biosimilars, and vaccines.

Using a pneumatic bioreactor, we demonstrate the assembly, operation, and performance of this single-use bioreactor system for the growth of mammalian cells.

使い捨て空気圧バイオリアクターシステムを使用して、哺乳動物細胞の培養

Recent advances in mammalian, insect, and stem cell cultivation and scale-up have created tremendous opportunities for new therapeutics and personalized ...

Nutrient Regulation by Continuous Feeding for Large-scale Expansion of Mammalian Cells in Spheroids

In this demonstration, spheroids formed from the &#946;-TC6 insulinoma cell line were cultured as a model of manufacturing a mammalian islet cell product to demonstrate how regulating nutrient levels can improve cell yields. In previous studies, bioreactors facilitated increased culture volumes over static cultures, but no increase in cell yields were observed. Limitations in key nutrients such as glucose, which were consumed between batch feedings, can lead to limitations in cell expansion. Large fluctuations in glucose levels were observed, despite the increase in glucose concentrations in the media. The use of continuous feeding systems eliminated fluctuations in glucose levels, and improved cell growth rates when compared with batch fed static and SSB culture methods. Additional increases in growth rates were observed by adjusting the feed rate based on calculated nutrient consumption, which allowed the maintenance of physiological glucose over three weeks in culture. This method can also be adapted for other cell types.

Nutrient regulation using continuous growth adjusted feeding improves growth rates of mammalian cell spheroids compared to intermittent batch feeding for cultures in stirred suspension bioreactors. This study demonstrates the methods required for establishing simple adjusted rate fed cultures.

スフェロイドにおける哺乳動物細胞の大規模拡張のための連続供給することによって栄養規制

In this demonstration, spheroids formed from the &#946;-TC6 insulinoma cell line were cultured as a model of manufacturing a mammalian islet cell product ...

Propagation of Homalodisca coagulata virus-01 via Homalodisca vitripennis Cell Culture

The glassy-winged sharpshooter (Homalodisca vitripennis) is a highly vagile and polyphagous insect found throughout the southwestern United States. These insects are the predominant vectors of Xylella fastidiosa (X. fastidiosa), a xylem-limited bacterium that is the causal agent of Pierce&#39;s disease (PD) of grapevine. Pierce&#8217;s disease is economically damaging; thus, H. vitripennis have become a target for pathogen management strategies. A dicistrovirus identified as Homalodisca coagulata virus-01 (HoCV-01) has been associated with an increased mortality in H. vitripennis populations. Because a host cell is required for HoCV-01 replication, cell culture provides a uniform environment for targeted replication that is logistically and economically valuable for biopesticide production. In this study, a system for large-scale propagation of H. vitripennis cells via tissue culture was developed, providing a viral replication mechanism. HoCV-01 was extracted from whole body insects and used to inoculate cultured H. vitripennis cells at varying levels. The culture medium was removed every 24 hr for 168 hr, RNA extracted and analyzed with qRT-PCR. Cells were stained with trypan blue and counted to quantify cell survivability using light microscopy. Whole virus particles were extracted up to 96 hr after infection, which was the time point determined to be before total cell culture collapse occurred. Cells were also subjected to fluorescent staining and viewed using confocal microscopy to investigate viral activity on F-actin attachment and nuclei integrity. The conclusion of this study is that H. vitripennis cells are capable of being cultured and used for mass production of HoCV-01 at a suitable level to allow production of a biopesticide.

Propagation of Homalodisca coagulata virus-01 via Homalodisca vitripennis Cell Culture

Here we present a protocol to propagate Homalodisca vitripennis cells and HoCV-1 in vitro. Medium was removed from HoCV-1 positive cultures and RNA extracted every 24 hr for 168 hr. Cell survivability was quantified by trypan blue staining. Whole virus particles were extracted post-infection. Extracted RNA was quantified by qRT-PCR.

の伝播<em&gt; Homalodisca coagulataウイルス01</em&gt;経由<em&gt; Homalodisca vitripennis</em&gt;細胞培養

The glassy-winged sharpshooter (Homalodisca vitripennis) is a highly vagile and polyphagous insect found throughout the southwestern United States. These ...

免疫・感染学

A Two-Step Strategy that Combines Epigenetic Modification and Biomechanical Cues to Generate Mammalian Pluripotent Cells

Cell phenotype can be reversed or modified with different methods, with advantages and limitations that are specific for each technique. Here we describe a new strategy that combines the use of chemical epigenetic erasing with mechanosensing-related cues, to generate mammalian pluripotent cells. Two main steps are required. In the first step, adult mature (terminally differentiated) cells are exposed to the epigenetic eraser 5-aza-cytidine to drive them into a pluripotent state. This part of the protocol was developed, based on the increasing understanding of the epigenetic mechanisms controlling cell fate and differentiation, and involves the use of the epigenetic modifier to erase cell differentiated state and then drive into a transient high plasticity window.
In the second step, erased cells are encapsulated in polytetrafluoroethylene (PTFE) micro-bioreactors, also known as Liquid Marbles, to promote 3D cell rearrangement to extend and stably maintain the acquired high plasticity. PTFE is a non-reactive hydrophobic synthetic compound and its use permits the creation of a cellular microenvironment, which cannot be achieved in traditional 2D culture systems. This system encourages and boosts the maintenance of pluripotency though bio-mechanosensing-related cues.
The technical procedures described here are simple strategies to allow for the induction and maintenance of a high plasticity state in adult somatic cells. The protocol allowed the derivation of high plasticity cells in all mammalian species tested. Since it does not involve the use of gene transfection and is free of viral vectors, it may represent a notable technological advance for translational medicine applications. Furthermore, the micro-bioreactor system provides a notable advancement in stem cell organoid technology by in vitro re-creating a specific micro-environment that allows for the long-term culture of high plasticity cells, namely as ESCs, iPSCs, epigenetically erased cells and MSCs.

We here present a method that combines the use of chemical epigenetic erasing with mechanosensing-related cues to efficiently generate mammalian pluripotent cells, without the need of gene transfection or retroviral vectors. This strategy is, therefore, promising for translational medicine and represents a notable advancement in stem cell organoid technology.

エピジェネティック修飾と生体力学的手がかりを組み合わせて哺乳類多能性細胞を生成する2段階戦略

Cell phenotype can be reversed or modified with different methods, with advantages and limitations that are specific for each technique. Here we describe ...

生物学

マウス骨格筋を採取して幹細胞とそのニッチを培養

Unfractionated Bulk Culture of Mouse Skeletal Muscle to Recapitulate Niche and Stem Cell Quiescence

Skeletal muscle is the largest tissue of the body and performs multiple functions, from locomotion to body temperature control. Its functionality and recovery from injuries depend on a multitude of cell types and on molecular signals between the core muscle cells (myofibers, muscle stem cells) and their niche. Most experimental settings do not preserve this complex physiological microenvironment, and neither do they allow the ex vivo study of muscle stem cells in quiescence, a cell state that is crucial for them. Here, a protocol is outlined for the ex vivo culture of muscle stem cells with cellular components of their niche. Through the mechanical and enzymatic breakdown of muscles, a mixture of cell types is obtained, which is put in 2D culture. Immunostaining shows that within 1 week, multiple niche cells are present in culture alongside myofibers and, importantly, Pax7-positive cells that display the characteristics of quiescent muscle stem cells. These unique properties make this protocol a powerful tool for cell amplification and the generation of quiescent-like stem cells that can be used to address fundamental and translational questions.

著者スポットライト:ニッチ成分で静止筋幹細胞を培養するためのEx Vivoプロトコル 

Skeletal muscle comprises multiple cell types, including resident stem cells, each with a special contribution to muscle homeostasis and regeneration. Here, the 2D culture of muscle stem cells and the muscle cell niche in an ex vivo setting that preserves many of the physiological, in vivo, and environmental characteristics are described.

マウス骨格筋の未分画バルク培養によるニッチおよび幹細胞の静止を再現

Skeletal muscle is the largest tissue of the body and performs multiple functions, from locomotion to body temperature control. Its functionality and ...

哺乳類セルラインの嫌気性成長と維持

要約

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アンインビトロシステム

FIBS対応の非侵襲メタボリック·プロファイリング

哺乳類細胞内の細胞内亜鉛プールを測定するための原子吸光度分光法

使い捨て空気圧バイオリアクターシステムを使用して、哺乳動物細胞の培養

スフェロイドにおける哺乳動物細胞の大規模拡張のための連続供給することによって栄養規制

の伝播

エピジェネティック修飾と生体力学的手がかりを組み合わせて哺乳類多能性細胞を生成する2段階戦略

マウス骨格筋の未分画バルク培養によるニッチおよび幹細胞の静止を再現

マウス骨格筋の未分画バルク培養によるニッチおよび幹細胞の静止を再現

マウス骨格筋の未分画バルク培養によるニッチおよび幹細胞の静止を再現