Apoptosis induced by oxidized lipids is associated with up-regulation of
p66Shc in intestinal Caco-2 cells: protective effects of
phenolic compounds
Claudio Giovannini
a,
4
, Beatrice Scazzocchio
a
, Paola Matarrese
b
, Rosaria Varı`
a
,
Massimo D’Archivio
a
, Roberta Di Benedetto
a
, Stefania Casciani
c
, Maria Rita Dessı`
c
,
Elisabetta Straface
b
, Walter Malorni
b,1
, Roberta Masella
a,1
a
National Centre for Food Quality and Risk Assessment, Istituto Superiore di Sanita` , 299-00161 Rome, Italy
b
Department of Drug Research and Evaluation, Istituto Superiore di Sanita` , 299-00161 Rome, Italy
c
Department of Internal Medicine, II University of Rome bTor VergataQ, 1-00133 Rome, Italy
Received 4 May 2006; received in revised form 22 December 2006; accepted 10 January 2007
Abstract
In this study, we investigated the alterations of the redox balance induced by the lipid fraction of oxLDL in Caco-2 intestinal cells, and the
effects of tyrosol and protocatechuic acid, two dietary phenolic compounds. We found that oxidized lipids extracted from oxLDL (LipE)
induced oxidative stress by determining, 6 h after treatment, ROS overproduction (about a 100% and a 43% increase of O
2
À
and H
2
O
2
production, respectively, P b.05: LipE vs. control) and, 12 h after treatment, GSH depletion (about a 26% decrease, P b.05: LipE vs. control),
and by impairing the activities of superoxide dismutase, catalase and glutathione peroxidase. In response to the induced oxidative stress, we
observed significant overexpression of glutathione peroxidase (6 h after treatment: P b.05), glutathione reductase and g-glutamylcysteine
synthetase (12 h after treatment: P b.05). Notably, when GSH depletion occurred, p66Shc protein expression increased by about 300% with
respect to control ( P b.001; LipE vs. control). These effects were fully counteracted by dietary phenolics which inhibited ROS
overproduction and GSH consumption, rendered the reactive transcription of glutathione-associated enzymes unnecessary and blocked the
intracellular signals leading to the overexpression and rearrangement of p66Shc signalling molecule. Altogether, these results suggest that the
impairment of the antioxidant system hijacks intestinal cells towards an apoptotic-prone phenotype via the activation of p66Shc molecule.
They also propose a reappraisal of dietary polyphenols as intestinal protecting agents, indicating the antiapoptotic effect as a further
mechanism of action of these antioxidant compounds.
D 2008 Elsevier Inc. All rights reserved.
Keywords: Apoptosis; oxLDL; Oxidized lipids; Caco-2; Glutathione; p66Shc; Polyphenols
1. Introduction
In the gastrointestinal tract, prooxidants such as endog-
enous and exogenous lipid peroxides have been demon-
strated to impact intestinal integrity. In fact, cellular and
molecular events involved in degenerative pathological
processes leading to intestinal disorders have been associ-
ated with redox alterations. Being the interface between the
organism and its luminal environment, the intestine is
constantly challenged by diet-derived oxidants as well as
by endogenously generated reactive species or oxidants. In
particular, a high intake of dietary polyunsaturated fatty
acids can contribute to the luminal accumulation of lipid
hydroperoxides, and the subsequent lipid peroxidation can
induce oxidative stress and redox imbalance, contributing to
the development of gut pathologies, such as inflammation
and cancer [1,2]. To preserve cellular integrity and
tissue homeostasis, the intestine possesses several defence
0955-2863/$ – see front matter D 2008 Elsevier Inc. All rights reserved.
doi:10.1016/j.jnutbio.2007.01.010
Abbreviations: CAT, catalase; DHE, dihydroethidium; DHR 123,
dihydrorhodamine 123; GPx, glutathione peroxidase; GSSGred, glutathione
reductase; g-GCS, g-glutamylcysteine synthetase; GSH, glutathione; LipE,
lipid extract from oxLDL; MBC, monochlorobimane; oxLDL, oxidized
low-density lipoprotein; ROS, reactive oxygen species; SOD, superoxide-
dismutase.
4 Corresponding author. Tel.: +39 06 49902589; fax: +39 06
49902763.
E-mail address: clagiovn@iss.it (C. Giovannini).
1
Both should be considered as senior author.
Journal of Nutritional Biochemistry 19 (2008) 118 – 128
Available online at www.sciencedirect.com