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Acidic extracellular pH shifts colorectal cancer cell death from apoptosis to necrosis upon exposure to propionate and acetate, major end-products of the human probiotic propionibacteria

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Abstract

The human probiotic Propionibacterium freudenreichii kills colorectal adenocarcinoma cells through apoptosis in vitro via its metabolites, the short chain fatty acids (SCFA), acetate and propionate. However, the precise mechanisms, the kinetics of cellular events and the impact of environmental factors such as pH remained to be specified. For the first time, this study demonstrates a major impact of a shift in extracellular pH on the mode of propionibacterial SCFA-induced cell death of HT-29 cells, in the pH range 5.5 to 7.5 prevailing within the colon. Propionibacterial SCFA triggered apoptosis in the pH range 6.0 to 7.5, a lethal process lasting more than 96 h. Indeed at pH 7.5, SCFA induced cell cycle arrest in the G2/M phase, followed by a sequence of cellular events characteristic of apoptosis. By contrast, at pH 5.5, the same SCFA triggered a more rapid and drastic lethal process in less than 24 h. This was characterised by sudden mitochondrial depolarisation, inner membrane permeabilisation, drastic depletion in ATP levels and ROS accumulation, suggesting death by necrosis. Thus, in digestive cancer prophylaxis, the observed pH-mediated switch between apoptosis and necrosis has to be taken into account in strategies involving SCFA production by propionibacteria to kill colon cancer cells.

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Abbreviations

AV:

annexinV-FITC

ΔΨm:

mitochondrial transmembrane potential

DHE:

dihydroethidium

DiOC6(3):

dihexyloxacarbocyanine iodide

Eth:

Ethidium

Etop.:

etoposide

FDA:

fluorescein diacetate

IM:

inner membrane

LND:

lonidamine

O2 .− :

superoxide anion

Men:

Menadione

PARP:

poly-ADP-ribose polymerase

PCD:

programmed cell death

PI:

propidium iodide

PTPC:

permeability transition pore complex

RIP:

receptor-interacting protein

ROS:

reactive oxygen species

SCFA:

short-chain fatty acid

TNF:

tumor necrosis factor

TRAIL:

TNF α related apoptosis inducing ligand

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Acknowledgments

The authors wish thank Dr. O. Meurette and Dr. L. Huc for their helpful advice on cytometric analysis, and C. Longin for technical assistance in TEM assay.

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Authors and Affiliations

  1. UMR 1253 INRA Agrocampus, Science & Technologie du Lait et de l’Oeuf, 65, rue de St Brieuc, 35042, Rennes cedex, France

    Annaïg Lan & Gwénaël Jan

  2. Laboratoires Standa, 68 rue R. Kaskoreff, 14050, Caen cedex 4, France

    Annaïg Lan

  3. INSERM U620, Université Rennes1, Faculté de Pharmacie, 2 av Prof Léon Bernard, 35043 Rennes cedex, France

    Dominique Lagadic-Gossmann

  4. CNRS UMR 8159, LGBC, Université de Versailles, 45 av. des Etats Unis, 78035, Versailles, France

    Christophe Lemaire & Catherine Brenner

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  1. Annaïg Lan
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  2. Dominique Lagadic-Gossmann
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  3. Christophe Lemaire
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  4. Catherine Brenner
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  5. Gwénaël Jan
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Correspondence to Gwénaël Jan.

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This work has been supported by a grant from CRITT santé Bretagne.

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Lan, A., Lagadic-Gossmann, D., Lemaire, C. et al. Acidic extracellular pH shifts colorectal cancer cell death from apoptosis to necrosis upon exposure to propionate and acetate, major end-products of the human probiotic propionibacteria. Apoptosis 12, 573–591 (2007). https://doi.org/10.1007/s10495-006-0010-3

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