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A Flow Cytometry-based Assay for Measuring Mitochondrial Membrane Potential in Cardiac Myocytes After Hypoxia/Reoxygenation
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A Flow Cytometry-based Assay for Measuring Mitochondrial Membrane Potential in Cardiac Myocytes After Hypoxia/Reoxygenation

Published on: July 13, 2018

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A Flow Cytometry-based Assay for Measuring Mitochondrial Membrane Potential in Cardiac Myocytes After

Guihao Chen1, Yuejin Yang2, Chuansheng Xu3

  • 1State Key Laboratory of Cardiovascular Disease, Department of Cardiology, Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College.

Journal of Visualized Experiments : Jove
|July 31, 2018
PubMed

Insights

Reperfusion injury can cause heart cell death. This study uses JC-1 dye to measure mitochondrial membrane potential changes, indicating cell damage after heart attack reperfusion.

Area of Science:

  • Cardiovascular Science
  • Cellular Biology
  • Biochemistry

Background:

  • ST-segment elevated myocardial infarction (STEMI) requires timely reperfusion to minimize heart damage.
  • Reperfusion, while necessary, can paradoxically cause further cardiomyocyte death (reperfusion injury).
  • Mitochondrial permeability transition pore (mPTP) opening and mitochondrial depolarization are key events in reperfusion injury, leading to cell death.

Purpose of the Study:

  • To investigate the role of mitochondrial permeability transition pore (mPTP) opening in reperfusion injury.
  • To utilize JC-1 dye as a sensitive indicator of mitochondrial membrane potential (MMP) changes.
  • To quantify mitochondrial depolarization in human cardiac myocytes following hypoxia/reoxygenation.

Main Methods:

  • Human cardiac myocytes were subjected to hypoxia/reoxygenation to simulate reperfusion injury.
  • JC-1, a fluorescent dye, was used to assess mitochondrial membrane potential (MMP).
  • Flow cytometry was employed to detect changes in JC-1 fluorescence, specifically the red/green emission ratio, indicating mitochondrial depolarization.

Main Results:

  • JC-1 accumulation in mitochondria is dependent on MMP; higher MMP leads to greater JC-1 uptake.
  • A shift in JC-1 fluorescence from green to red emission indicates increased mitochondrial accumulation.
  • A reduced red/green fluorescence intensity ratio signifies mitochondrial depolarization, a marker of mPTP opening.

Conclusions:

  • JC-1 fluorescence provides a reliable method to measure MMP and detect mPTP opening in cardiac myocytes.
  • This technique allows for the assessment of mitochondrial dysfunction during reperfusion injury.
  • Understanding mPTP dynamics is crucial for developing strategies to mitigate reperfusion injury in myocardial infarction.

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