Isolation, Culture, and Functional Analysis of Murine Thermogenic Adipocytes.
Isolation, Culture, and Functional Analysis of Murine Thermogenic Adipocytes.
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DOI：
10.1016/j.xpro.2020.100118
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发表时间：
2020-12-18
期刊：
影响因子：
--
通讯作者：
Li Y
中科院分区：
文献类型：
--
作者：
Oeckl J;Bast-Habersbrunner A;Fromme T;Klingenspor M;Li Y
研究棕色和脆性脂肪组织需要精确和可靠的细胞产热定量。该方案描述了原代小鼠前脂肪细胞的分离，分化为产热的棕色和褐色脂肪细胞，以及随后的耗氧量分析。通常应用的程序只测量基础和最大质子泄漏相关的氧气消耗，而不是显式解偶联蛋白1(UCP1)依赖的呼吸。有意义的耗氧量分析需要(1)激活UCP1，(2)控制细胞内游离脂肪酸水平，(3)抑制消耗ATP的无用循环。有关本议定书的使用和执行的完整细节，请参阅)和。表达UCP1的生热脂肪细胞的培养和生物能量表型失活UCP1蛋白不产生热有意义的分析需要点火，控制FFA水平，并且被阻断的ATP合成酶ECAR不能反映活跃脂解过程中脂肪细胞的糖酵解率研究棕色和棕色脂肪组织需要精确和可靠的细胞产热定量。该方案描述了原代小鼠前脂肪细胞的分离，分化为产热的棕色和褐色脂肪细胞，以及随后的耗氧量分析。通常应用的程序只测量基础和最大质子泄漏相关的氧气消耗，而不是显式解偶联蛋白1(UCP1)依赖的呼吸。有意义的耗氧量分析需要(1)激活UCP1，(2)控制细胞内游离脂肪酸水平，(3)抑制消耗ATP的无用循环。
Studying brown and brite adipose tissue requires precise and reliable quantification of cellular thermogenesis. This protocol describes the isolation of primary murine pre-adipocytes, differentiation into thermogenic brown and brite adipocytes, and subsequent oxygen consumption analysis. Commonly applied procedures only measure basal and maximal proton leak-linked oxygen consumption but not explicitly uncoupling protein 1 (UCP1)-dependent respiration. Meaningful oxygen consumption analyses require (1) the activation of UCP1, (2) control over intracellular free-fatty-acid levels, and (3) inhibition of ATP-consuming futile cycles. For complete details on the use and execution of this protocol, please refer to) and. Culture and bioenergetic phenotyping of UCP1-expressing thermogenic adipocytes Inactive UCP1 protein does not generate heat Meaningful assay requires ignition, control over FFA levels, and blocked ATP synthase ECAR does not reflect the glycolytic rate of adipocytes during active lipolysis Studying brown and brite adipose tissue requires precise and reliable quantification of cellular thermogenesis. This protocol describes the isolation of primary murine pre-adipocytes, differentiation into thermogenic brown and brite adipocytes, and subsequent oxygen consumption analysis. Commonly applied procedures only measure basal and maximal proton leak-linked oxygen consumption but not explicitly uncoupling protein 1 (UCP1)-dependent respiration. Meaningful oxygen consumption analyses require (1) the activation of UCP1, (2) control over intracellular free-fatty-acid levels, and (3) inhibition of ATP-consuming futile cycles.
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影响因子：
7.4
作者：
Gerencser, Akos A.;Neilson, Andy;Choi, Sung W.;Edman, Ursula;Yadava, Nagendra;Oh, Richard J.;Ferrick, David A.;Nicholls, David G.;Brand, Martin D.
通讯作者：
Brand, Martin D.
影响因子：
4.4
作者：
Schweizer S;Oeckl J;Klingenspor M;Fromme T
通讯作者：
Fromme T
影响因子：
8.8
作者：
Li, Yongguo;Schwalie, Petra C.;Klingenspor, Martin
通讯作者：
Klingenspor, Martin
影响因子：
7.7
作者：
Li, Yongguo;Fromme, Tobias;Klingenspor, Martin
通讯作者：
Klingenspor, Martin
影响因子：
3.7
作者：
Mookerjee SA;Nicholls DG;Brand MD
通讯作者：
Brand MD