A novel approach to measure mitochondrial respiration in frozen biological samples.,The EMBO Journal

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A novel approach to measure mitochondrial respiration in frozen biological samples.
The EMBO Journal ( IF 11.4 ) Pub Date : 2020-05-20 , DOI: 10.15252/embj.2019104073
Rebeca Acin-Perez _{1,

2} , Ilan Y Benador _{1,

2,

3} , Anton Petcherski _{1,

2} , Michaela Veliova _{1,

2,

4} , Gloria A Benavides ₅ , Sylviane Lagarrigue ₆ , Arianne Caudal ₇ , Laurent Vergnes _{2,

8} , Anne N Murphy ₉ , Georgios Karamanlidis ₁₀ , Rong Tian ₇ , Karen Reue _{2,

8} , Jonathan Wanagat _{2,

11} , Harold Sacks _{1,

2} , Francesca Amati ₆ , Victor M Darley-Usmar ₅ , Marc Liesa _{1,

2,

4,

12} , Ajit S Divakaruni _{2,

4} , Linsey Stiles _{1,

2} , Orian S Shirihai _{1,

2,

3,

4,

12}

Affiliation

Department of Medicine, Endocrinology, David Geffen School of Medicine, University of California, Los Angeles, CA, USA.
Metabolism Theme, David Geffen School of Medicine, University of California, Los Angeles, CA, USA.
Nutrition and Metabolism, Graduate Medical Sciences, Boston University School of Medicine, Boston, MA, USA.
Department of Molecular and Medical Pharmacology, University of California, Los Angeles, CA, USA.
Department of Pathology and Mitochondrial Medicine Laboratory, University of Alabama at Birmingham, Birmingham, AL, USA.
Department of Biomedical Sciences, University of Lausanne, Lausanne, Switzerland.
Mitochondria and Metabolism Center, University of Washington, Seattle, WA, USA.
Department of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, CA, USA.
Department of Pharmacology, University of California, San Diego, CA, USA.
Cardiometabolic Disorders, Amgen Research, Thousand Oaks, CA, USA.
Department of Medicine, Division of Geriatrics, University of California, Los Angeles, CA, USA.
Molecular Biology Institute, UCLA, Los Angeles, CA, USA.

Respirometry is the gold standard measurement of mitochondrial oxidative function, as it reflects the activity of the electron transport chain complexes working together. However, the requirement for freshly isolated mitochondria hinders the feasibility of respirometry in multi‐site clinical studies and retrospective studies. Here, we describe a novel respirometry approach suited for frozen samples by restoring electron transfer components lost during freeze/thaw and correcting for variable permeabilization of mitochondrial membranes. This approach preserves 90–95% of the maximal respiratory capacity in frozen samples and can be applied to isolated mitochondria, permeabilized cells, and tissue homogenates with high sensitivity. We find that primary changes in mitochondrial function, detected in fresh tissue, are preserved in frozen samples years after collection. This approach will enable analysis of the integrated function of mitochondrial Complexes I to IV in one measurement, collected at remote sites or retrospectively in samples residing in tissue biobanks.

中文翻译：

一种测量冷冻生物样品中线粒体呼吸的新方法。

呼吸测量法是衡量线粒体氧化功能的黄金标准，因为它反映了电子传递链复合物协同工作的活性。然而，需要新鲜分离的线粒体阻碍了呼吸测量法在多中心临床研究和回顾性研究中的可行性。在这里，我们描述了一种适用于冷冻样品的新型呼吸测量方法，通过恢复冷冻/解冻过程中丢失的电子转移成分并校正线粒体膜的可变通透性。这种方法在冷冻样品中保留了 90-95% 的最大呼吸能力，并且可以高灵敏度地应用于分离的线粒体、透化细胞和组织匀浆。我们发现在新鲜组织中检测到的线粒体功能的主要变化，收集后保存在冷冻样品中。这种方法将能够在一次测量中分析线粒体复合物 I 至 IV 的综合功能，在远程站点收集或在组织生物库中的样本中回顾性分析。

更新日期：2020-07-01

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