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Isolation and Functional Analysis of Mitochondria from Cultured Cells and Mouse Tissue
09:27

Isolation and Functional Analysis of Mitochondria from Cultured Cells and Mouse Tissue

Published on: March 23, 2015

Reduction in mitochondrial membrane potential is an early event in Fas-independent CTL-mediated apoptosis

D Hu1, T J Kipps

  • 1Department of Medicine, University of California at San Diego School of Medicine, La Jolla, California, 92093-0663, USA.

Cellular Immunology
|August 6, 1999
PubMed

Insights

Cytotoxic T lymphocytes (CTL) kill target cells through granule-mediated pathways. This study shows mitochondrial membrane potential (DeltaPsi(m)) reduction is an early step in this Fas-independent killing process.

Area of Science:

  • Immunology
  • Cell Biology

Background:

  • Cytotoxic T lymphocytes (CTL) eliminate target cells via Fas-mediated or granule-mediated pathways.
  • Understanding the early events in CTL-mediated cytotoxicity is crucial for immune response research.

Purpose of the Study:

  • To investigate the role of mitochondrial membrane potential (DeltaPsi(m)) reduction in Fas-independent CTL killing.
  • To characterize the sequence of events in granule-mediated CTL cytotoxicity.

Main Methods:

  • Utilized Fas-negative target cells to isolate the granule-mediated pathway.
  • Monitored changes in target cell mitochondrial membrane potential (DeltaPsi(m)).
  • Assessed phosphatidyl serine translocation, cytosolic protein release, and plasma membrane integrity.

Main Results:

  • Reduction in DeltaPsi(m) is an early event in Fas-independent CTL killing, preceding other cell death markers.
  • This DeltaPsi(m) reduction and protein release were inhibited by N-carbobenzoxy-Ala-Pro-Phe chloromethyl ketone.
  • Caspase inhibitors (z-VAD-fmk, z-DEVD-fmk) did not inhibit Fas-independent CTL killing, unlike Fas-mediated apoptosis.

Conclusions:

  • Mitochondrial membrane potential (DeltaPsi(m)) reduction is a key early indicator of Fas-independent CTL-mediated target cell death.
  • The granule-mediated pathway involves distinct molecular mechanisms compared to Fas-mediated apoptosis.
  • Monitoring DeltaPsi(m) changes offers a sensitive method for assessing CTL activity.

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