IMR Press / FBL / Volume 14 / Issue 9 / DOI: 10.2741/3470

Frontiers in Bioscience-Landmark (FBL) is published by IMR Press from Volume 26 Issue 5 (2021). Previous articles were published by another publisher on a subscription basis, and they are hosted by IMR Press on imrpress.com as a courtesy and upon agreement with Frontiers in Bioscience.

Open Access Article
Pharmacological screening and enzymatic assays for apoptosis
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1 University of Versailles/SQY, PRES UniverSud Paris, CNRS UMR 8159, Versailles, France
2 Theraptosis R and D Laboratory, Theraptosis SA, Romainville, France
3 Inserm U602, PRES UniverSud Paris, Hopital Paul Brousse, Villejuif, France

Academic Editor: Catherine Brenner

Front. Biosci. (Landmark Ed) 2009, 14(9), 3550–3562; https://doi.org/10.2741/3470
Published: 1 January 2009
(This article belongs to the Special Issue Apoptosis and mitochondria)
Abstract

Mitochondria play a central role in the intrinsic pathway of apoptosis. In response to many pro-apoptotic stimuli, mitochondria undergo an irreversible process called mitochondrial membrane permeabilization (MMP). The detection of MMP in isolated mitochondria is most often based on assays that monitor either the loss of the inner transmembrane potential (∆Ψm; classically with Rhodamine 123), permeability transition (PT, cyclosporin A-sensitive matrix swelling), or the release of critical pro-apoptotic intermembrane space effectors. To gain complementary information on MMP mechanisms, we have systematically used three additional assays optimized for the 96-well microplate format: (1) inner membrane permeability, (2) VDAC-associated NADH reductase activity, and (3) ATP/ADP translocase activity. We report that ad hoc combinations of ANT and VDAC ligands, carbonyl cyanide m-chlorophenylhydrazone (CCCP), mastoparan and Vpr52-96 peptide and PT inhibitors, permit to explore relationships between enzymatic functions of sessile mitochondrial proteins (i.e. ANT, VDAC) and MMP. These assays should be useful tools to investigate mitochondrial apoptosis, decipher the implication of inner and outer membrane permeabilization and provide a multi-parametric approach for drug discovery.

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