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Hong-Liang
Li, Xiao-Hong Li, Yan-Qing Lü, Chun-Ling Ye, Xian-Da Ren,
Department of Pharmacology, Jinan University Pharmacy College,
Guangzhou 510632, Guangdong, China
Dan-Dan Chen, Department of Cardiology, First Affiliated
Hospital, Zhongshan University, Guangzhou 510089, Guangdong, China
Hai-Wei Zhang, Department of Pathology, Jinan University
Medical College, Guangzhou 510632, Guangdong, China
Supported by National Natural Science Foundation of China,
No. 39770300, 30070873, and the Overseas Chinese Affairs Office of
the State Council Foundation, No. 98-33
Correspondence to: Prof. Xian-Da Ren, Department of
Pharmacology, Jinan University Pharmacy College, Guangzhou 510632,
Guangdong, China. tsam@jnu.edu.cn
Telephone: +86-20-85220261
Received 2002-01-28 Accepted 2002-03-05
Abstract
AIM: To identify whether JTE-522 can induce apoptosis in AGS
cells and ROS also involved in the process, and to investigate the
changes in NF-kB, p53, bcl-2 and caspase in the apoptosis process.
METHODS: Cell culture, MTT, Electromicroscopy, agarose gel
electrophoresis, lucigenin, Western blot and electrophoretic
mobility shift assay (EMSA) analysis were employed to investigate
the effect of JTE-522 on cell proliferation and apoptosis in AGS
cells and related molecular mechanisms.
RESULTS: JTE-522 inhibited the growth of AGS cells and
induced the apoptosis. Lucigenin assay showed the generation of ROS
in cells under incubation with JTE-522. The increased ROS generation
might contribute to the induction of AGS cells to apoptosis. EMSA
and Western blot revealed that NF-kB activity was almost completely
inhibited by preventing the degradation of IkBα. Additionally,
by using Western blot we confirmed that the level of bcl-2 was
decreased, whereas p53 showed a great increase following JTE-522
treatment. Their changes were in a dose-dependent manner.
CONCLUSION: These findings suggest that reactive oxygen
species, NF-kB, p53, bcl-2 and caspase-3 may play an important role
in the induction of apoptosis in AGS cells after treatment with
JTE-522.
Li HL, Chen DD, Li XH, Zhang HW, Lü YQ, Ye CL, Ren XD. Changes of
NF-kB, p53, Bcl-2 and caspase in apoptosis induced by JTE-522 in
human gastric adenocarcinoma cell line AGS cells: role of reactive
oxygen species.World J Gastroenterol 2002;8(3):431-435
INTRODUCTION
Apoptosis is an active cell death process, which requires
specific gene regulation. A critical role for p53 in the execution
of some forms of apoptosis has been suggested[1-6]. This
protein is a sequence-specific DNA-binding protein, active as a
transcription factor. It has been proposed that p53 may be involved
in the cellular response to DNA damage, producing arrest in the G1
phase of the cell cycle to allow efficient repair of DNA before
entry to S phase, or cell death if the damage is too large to be
repaired[7,8]. Another gene implicated in apoptosis is
bcl-2. The bcl-2 gene product functions as an anti-apoptotic signal,
suppressing apoptosis induced by a wide variety of stimuli,
including chemotherapeutic drugs and γ radiation[9-13].
The exact mechanism of bcl-2 in preventing apoptosis is still not
clear. However, bcl-2 has been implicated in cellular control of
their redox state[14].
Previous
studies have demonstrated that non-steroidal anti-inflammatory drugs
(NSAIDs) given in vivo to rodents and human can inhibit tumor growth[15,16].
JTE-522 is a novel NSAIDs, which is a specific inhibitor of
cyclooxygenase-2 (COX-2) with significant anti-inflammatory and
analgestic properties[17]. Some reported that JTE-522
possesses strong chemopreventive activity against colon
carcinogenesis[18], but the precise mechanism by which
JTE-522 inhibits colon carcinogenesis is not clear. It is often
attributed to specific inhibition of arachidonic acid metabolism via
coxenzymes. However, recent studies showed that the antitumor effect
had little connection with NSAIDs inhibitory activity against
cyclooxygenase, and was not prevented by exogenous supplementation
of 16,16-dimethyl prostaglandin E2. Several groups have
shown that certain NSAIDs induce apoptosis of tumor cell line, which
is associated with the generation of reactive oxygen species (ROS)[19,20].
However, the signaling pathway leading to apoptotic cell death
remains unclear.
ROS
can play a central role in regulating cell proliferation and cell
death. Evidence has been obtained that ROS such as superoxide and
hydrogen peroxide can influence cell death triggered by internal
cues (p53-mediated), external cues (TGF-beta-mediated) and
immunogenic signals (TNF-alpha)[21,22]. In other
instances, however, generation of ROS can inhibit apoptosis.
Although the mechanism involved is still controversial, redox status
and/or hydrogen peroxide have both been proposed as critical factors[19].
Therefore it is possible that ROS may play a role in regulating
apoptosis in gastric epithehum.
The
purpose of the present study is to identify whether JTE-522 can
induce apoptosis in AGS cells and ROS are also involved in the
process, and to investigate the changes in NF-kB, p53, bcl-2 and
caspase in the apoptosis process.
MATERIALS AND METHODS
Cell line and reagents
Human gastric adenocarcinoma cell line AGS was provided by
Cancer Institute, Zhongshan University. Cells were grown in
RPMI-1640 medium and supplemented with 10% new bovine serum,
penicillin G (100kU.L-1) and kanamycin (0.1g.L-1)
at 37℃
in a 5% CO2-95% air atmosphere. Antibodies used in this
study included p53, bcl-2, IkBα and Beta actin were obtained
from Santa Cruz. All other chemicals were purchased from Sigma
Chemical Co (St. Louis,MO,USA).
MTT assay
AGS cells growing on 96-well plates were treated with
JTE-522 (0.1mmol/L - 1mmol/L) for 72h, untreated cells served as a
control. 10μL of the 2.5g.L-1 stock solution of
3-[4, 5-dimethylthiaolyl]-2, 5-diphenyl-tetrazolium bromide (MTT)
was added to each well. After 1h of incubation at 37℃,
the medium was removed, 50μl of the extraction buffer (10%
Trition-X100; 0.1mol/L HCl) was added, and plates were gently shaken
for 30min at room temperature. The optical densities were measured
at 570nm.
Morphological and biochemical analysis of apoptosis
Morphological changes in the nuclear chromatin of cells
undergoing apoptosis were detected by electron microscopy (EM).
Cells were peleted and fixed with 30mL/L glutaraldehyde in PBS. EM
analysis was performed as described previously[23].
Oligonucleosomal cleavage of genomic DNA was detected by agarose gel
electrophoresis. In brief, genomic DNA isolated as previously
described[24] was subjected to 1.5% agarose gel
electrophoresis, followed by ethidium bromide staining.
Assay for reactive oxygen species production
Generation of ROS was assessed using lucigenin. AGS cells
grown in 75cm2 culture flasks were incubated for 6h with
JTE-522 (0.1-1mmol/L)in the presence or absence of 100μmol/L
pyrrolidine dithiocarbamate (PDTC). The cells were then scraped off
and washed in cold Hank's buffer. An aliquot containing 1×106
cells in 100μL of Hank's buffer was mixed in microtitrator
wells with 100μl of lucigenin prepared at a concentration of 40μmol/L.
Light emission was detected using a Berthold LB96V luminometer for
3min.
Assessment of caspase activity
Caspase-3 activity was measured using a caspase assay kit
according to the supplier's instruction. In brief, caspase-3
fluorogenic substrates (Ac-DEVD-AMC or Ac-IETD-AMC) were incubated
with JTE-522-treated with cell lysates for 1h at 37℃,
then AMC liberated from Ac-DEVD-AMC or Ac-IETD-AMC was measured
using a fluorometric plate reader with an excitation wavelength of
380nm and an emission wavelength of 420-460nm.
Western blot analysis
The cells were lysed in lysis buffer (25mmol/L hepes, 1.5%
Triton X-100,1% sodium deoxycholate,0.1% SDS, 0.5mol/L NaCl, 5mmol/L
EDTA, 50mmol/L NaF, 0.1mmol/L sodium vanadate,1mmol/L
phenylmethylsulfonyl fluoride(PMSF),and 0.1g.L-1
leupeptin(pH7.8) at 4℃
with sonication. The lysates were centrifuged at 15000g for 15min
and the concentration of the protein in each lysate was determined
with Coomassie brilliant blue G-250.Loading buffer (42mmol/L
Tris-HCl, 10% glycerol, 2.3% SDS, 5% 2-mercaptoethanol and 0.002%
bromophenol blue) was then added to each lysate,which was
subsequently boiled for 3min and then electrophoresed on a
SDS-polyacrylamidel gel. Proteins were transferred to nitrocellulose
and incubated sequentially with antibodies against IkBα, p53
and bcl-2 and then with peroxidase-conjugated secondary antibodies
in the second reaction. Detection was performed with enhanced
chemiluminescence reagent.
Electrophoretic mobility shift assay (EMSA)
Nuclear extracts were prepared from AGS cells treated with
JTE-522. Synthetic double-strand oligonucleotides of consensus NF-kB
binding sequence, GATCCCAACGGCAGGGGA, were end-labeled with [γ32]ATP
using T4 polynucleotide kinase. Nuclear extract was incubated with
the labeled probe in the presence of poly (dI-dC) in a binding
buffer containing 20mM
N-2-hydrocyethylpiperazine-N'-2-ethanesulfonic acid at room
temperature for 30min. For supershift assays, a total of 0.2μg
of antibodies against p65 subunit of NF-kB were included in the
reaction. DNA-protein complexes were resolved by electrophoresis in
a 5% non-denaturing polyacrylamide gel, which was dried and
visualized by autoradiography.
RESULTS
Effect of JTE-522 on cell proliferation and apoptosis
AGS cells were incubated with various does of JTE-522 for
72h. Analysis of cell viability using to MTT assay showed that
JTE-522 significantly inhibited cell viability. The inhibition of
cell viability was dependent on the dose of JTE-522 used
(Figure 1).
Figure 1(PDF)Effect
of JTE-522 on cell growth in AGS cells. The cells were treated with
various concentrations of JTE-522 for 72h. The antiproliferative
effect was measured by MTT assay. Results are the means±SD from
three independent determinations.
The
effect was due to apoptosis as demonstrated by EM and
electrophoresis of genomic DNA. JTE-522-treated cells showed
compacted nuclear chromatin with finely granular masses marginated
against the nuclear envelopeard condensed cytoplasm, the nuclear
outline was convoluted and the organelles were preserved (Figure 2)
and led to oligonucleosomal cleavage of genomic DNA (Figure 3),
which were hallmarks of apoptosis.
Figure 2(PDF)Electro
micrographs of JTE-522-treated AGS cells. Control AGS cells (A),
or treated with 1mmol/L (B)
JTE-522 for 72h, were examined by EM as in “Materials and
Methods”. Magnification: ×4000
We next investigated whether the activaton of caspase was involved
in JTE-522-induced apoptosis of AGS cells. JTE-522-induced apoptosis
of AGS cells was accompanied by the induction of caspases activity
as demonstrated by the cleavage of Ac-DEVD-AMC and Ac-IETD-AMC,
respectively (Figure 4). These results indicated that
JTE-522-induced cell death of AGS cells was a typical apoptosis
associated with caspase activation.
Figure 3 DNA
ladder pattern formation of AGS cells. Cells treated with different
concentrations of JTE-522 for 72h and uhe formation of
oligonucleosomal fragments was determined by 1.5% agarose gel
electrophoresis. M, DNA markers; lanes 1-4, AGS cells treated with
0, 0.25, 0.50, 1mmol/L of JTE-522
Figure 4(PDF)Activation
of caspase-3 activities by JTE-522 in AGS cells for indicated time
period. JTE-522 treatment (0.75mmol/L) induced cleavage of Ac-DEVD-AMC
and Ac-IETD-AMC, indicating activation of caspase-3 activity,
respectively
Effect of JTE-522 on the production of ROS in AGS cells
The effect of JTE-522 on the production of ROS in AGS cells,
as assessed with lucigenin chemiluminesecence, was shown in Figure
5. Chemiluminesecence was significantly enhanced by incubation with
various doses of JTE-522. This enhancement was prevented by
co-incubation with PDTC at 100μmol/L. These results
demonstrated the generation of reactive oxygen species in cells
under incubation with JTE-522.
Effect of JTE-522 on the expression of p53, bcl-2, IkBα
and the activation of NF-kB
NF-kB plays a complex role in regulating programmed cell
death. In many instances the inhibition of NF-kB activaty can
sensitize cells to death inducers[25]. In other
instances, however, NF-kB activation has been found to play an
important role in the induction of apoptosis. To determine whether
the treatment with JTE-522 have any effect in the NF-kB
transcriptional factors. We performed EMSA with nuclear extracts
prepared from control or treated cells exposed to JTE-522 for
various concentrations for 6h. The NF-kB specific complexes found in
this cell line were almost complete inhibited in comparision with
untreated cells (Figure 6A). In accordance with result, an analysis
of IkBα proteins level by Western blot demonstrated that the
degradation of this protein was greatly inhibited during the
apoptotic process(Figure 6B).
Additionally,
by using Western blot we confirmed that the level of bcl-2 was
decreased, whereas p53 showed a great increase following JTE-522
treatment. Their changes were in a dose-dependent manner (Figure 7).
Figure 5(PDF)Effect
of JTE-522 on the generation of ROS. AGS cells were incubated for 6h
with JTE-522 (0.25-1mmol/L) in the presence or absence of PDTC at
100μmol/L. Lucigenic-associated chemiluminescence was measured
for 3min with a lumirometer. (A). Lane 1: control; lane 2-5: AGS
cells treated with 0.25, 0.5, 0.75, 1mmol/L of JTE-522; lane 6:
JTE-522 (1mmol/L)+PDTC (100μmol/L) cP<0.01
vs control. Results are the means±SD from three independent
determinations.
Figure 6(PDF)Effect
of JTE-522 on NF-kB binding activity and IkBα degradation.
Cells were treated with JTE-522 for 6h. Cells were harvested and
EMSA was performed as described (A).
Lane 1: control; lane 2-5: AGS cells treated with 0.25, 0.50, 0.75,
1mmol/L of JTE-522. The identity of DNA-complexed proteins was
confirmed by supershift assays using antibodies against p65 subunit
of NF-kB (lane 6). Immunobolt analysis of IkBα of corresponding
cytosilic supernatant (B).
Representative results from four independent experiments.
Figure 7 P53,
bcl-2 protein levels in AGS cells treated with JTE-522. Cell lysates
were collected and processed at 6h. The whole cellular protein was
electrophoresed in SDS-PAGE gel, Western blot was performed using
antibodies against p53, bcl-2. Beta actin was used as a lane-loading
control. (1) control; (2) 0.25mmol/L; (3) 0.5mmol/L; (4) 0.75mmol/L;
(5) 1mmol/L. Representative results from three independent
experiments.
DISCUSSION
Using cultured AGS human gastric adenocarcinoma cells, the
observations described in this study demonstrate that JTE-522, a
novel NSAIDs inhibits the growth of AGS cells and induces apoptosis
in a concentration-dependent manner.
The
onset of apoptosis is associated with the proteolytic activation of
caspases. Caspases, a family of cysteine proteases, play a critical
role in the execution of apoptosis[26-31]. They are
synthesized as proenzymes that are processed by self-proteolysis
and/or cleavage by another protease to their active forms in cells
undergoing apoptosis Caspase-3 is a major executioner of apoptosis.
It is promoted during the early stage of apoptosis and the activated
form is a marker for cells undergoing apoptosis[32].
After activation by initiators, the proform(p32) is cleaved to the
active forms p20, p17, or p11, respectively[33].
Therefore, activation of caspase-3 in AGS cells in this study not
only indicated the occurrence of apoptosis, but also implied the
involvement of caspase-3 in JTE-522-induced apoptosis.
ROS
have been found to play a central role in regulating apoptosis in
numerous instances. Given that reduced rates of apoptosis may
contribute to carcinogenesis, the regulation of cellular ROS
production may be an important variable in the development of
neoplasias. Other studies have also suggested a potential role for
ROS in cancer suppression. For example, the p53 tumor suppressor
protein activates the expression of ROS-generating proteins that
increase cellular ROS production and eventually trigger apoptosis[34,35].
Increased ROS generation by chemopreventive agents may serve to
compensate the lower levels of ROS generation in p53 null cells (AGS
cells are p53 null)[36,37]. Animal studies have also
implicated ROS in regulating carcinogenesis. Mice with elevated
levels of glutathione peroxidase are more sensitive to skin
carcinogenesis than their wild type counterpart[32]. A
similar correlation has been made in the colon, where strains with
higher levels of glutathione peroxidase activity have a higher
cancer risk. The role of ROS in carcinogenesis is however likely to
be complex given the potential mutagenicity of ROS. For example, the
NSAIDs inhibition of cyclooxygenase has been proposed to suppress
carcinogenesis by suppressing the production of peroxyl radicals and
the subsequent formation of mutagenic lipid peroxidation breakdown
products[38]. The role of ROS in carcinogenesis may
depend on the relative levels of different ROS generated, and where
and when they are present. However, as demonstrated in this paper,
we showed that JTE-522 increased ROS generation in AGS cells, and
this increased ROS generation might contribute to the induction of
AGS cells to apoptosis.
The
bcl-2 protooncogene is unique among cellular genes for its ability
in many contexts to block apoptotic cell death. A mechanism has been
proposed in which bcl-2 regulates antioxidant pathways at sites of
free radical generation[39]. The protein of bcl-2 also
protects against apoptosis by blocking cytochrome C release hence
this protein may have an antioxidant function[40]. In our
experiment, the expression of bcl-2 decreased, and the p53 protein
upregulated following JTE-522 treatment, these changes may implies
that intracellular ROS may interfere with the expression of bcl-2
and p53, thereby contributing to inducing apoptosis in AGS cells.
In
studying the mechanism by which the NSAIDs influenced cell death,
common effects on the transcription factor NF-kB were noted. The NF-kB
transcription factor is ubiquitous and can be detected under its
inactive form in the cytoplasm of almost all cell types[41,42].
It supresses the expression of cytokines, chemokines, growth
factors, cell adhesion molecules, and some acute phase proteins in
health and various pathological states[43,44].
Experimental data clearly indicate that NF-kB is a major regulator
of the inflammatory reaction by controlling the expression of
pro-inflammatory molecules in response to cytokines oxidatives
stress and infectious agents[45,46]. NF-kB is maintained
under such an inactive cytoplasmic form by virtue of its association
with an inhibitory molecule named IkB. IkBα is the best
characterized member of this family. In our current work, EMSA
revealed that JTE-522 inhibited NF-kB activation. Western blot
experiments demonstrated that this effect was mediated by inhibition
of IkBα degradation. Determining the role of NF-kB in gastric
carcinogenesis could help to guide the development of improved
chemoprevention and treatment strategies.
In
summary, JTE-522 inhibited cell growth and induced apoptosis in AGS
cells. Increased ROS may play an important role in this caspase-3
mediated apoptotic process. The inhibition of NF-kB by JTE-522 may
be mediated by preventing IkBα degradation. The precise
relationship and importance of each of these factors in the
apoptotic process should be established by more direct and profound
analysis.
ACKNOWLEDGMENTS
We gratefully acknowledge Prof Geng-Tao Lui, Institute of
Material Medica, Chinese Academy of Medical Sciences for the many
helpful discussion and suggestions relating to this work; Special
thanks to Chris Simmet and Pasricha Jerriment for proofreading the
manuscript and for useful suggestions; Dr Cheng-Wei He for technical
advice and helpful discussion; Dr Gang-Fei Peng for FCM; Dr Tao Wang
for Western blot; Dr Guo-Qing Xie and Mr Hai-Nan Wang for photo
processing.
REFERENCES
1 Peng
XM, Peng MM, Chen Q, Yao JL. Apoptosis, Bcl-2 and p53 protein
expression in tissues from hepatocellular
carcinoma.
Huaren Xiaohua Zazhi 1998; 6: 834-836
2 Hua JS.
Effect of Hp: cell proliferation and apoptosis on stomach cancer.
Shijie Huaren Xiaohua Zazhi 1999;7: 647-648
3 Xue XC,
Fang GE, Hua JD. Gastric cancer and apoptosis. Shijie Huaren Xiaohua
Zazhi 1999;7: 359-361
4 Qin LF,
Wang RN. Prognostic significance of FCM DNA analysis in carcinoma of
stomach.
Shanghai
Dier Yike Daxue Xuebao 1992; 12: 198-202
5
Yu GQ, Zhou
Q, Ding Ivan, Gao SS, Zhang ZY, Zou JX, Li YX, Wang LD. Changes of
p53 protein blood level in esophageal
cancer
patients and nomal subjects from a high incidence area in Henan,
China. World J Gastroenterol 1998; 4: 218
6
Luo D, Liu
QF, Gove V, Namov NV, Su JJ, Williams R. Analysis of N-ras gene
mutation and p53 gene expression in human
hepatocellular
carcinomas. World J Gastroenterol 1998; 4: 97-99
7 Li HL,
Zhang HW, Ren XD. Synergism between heparin and adriamycin on cell
proliferation and apoptosis in human
nasopharyngeal
carcinoma CNE2 cells. Acta Pharmacol sin 2002;23:167-172
8 Li HL, Ye
KH, Ren XD. Heparin induced apoptosis in human nasopharyngeal
carcinoma CNE2 cells. Cell Research
2001;11:311-315
9 Qiao Q,
Wu JS, Zhang J, Ma QJ, Lai DN. Expression and significance of
apoptosis related gene bcl-2, bax in human large
intestine
adenocarcinoma. Shijie Huaren Xiaohua Zazhi 1999; 7: 936-938
10 Yuan RW, Ding Q,
Jiang HY, Qin XF, Zou SQ, Xia SS. Bcl-2, p53 protein expression and
apoptosis in pancreatic cancer.
Shijie
Huaren Xiaohua Zazhi 1999; 7: 851-854
11 Jiang YG, Li QF,
Wang YM, Gu CH. Bcl-2/bax expression and hepatocyte apoptosis on
liver tissue in tupaia with HDV/HBV
infection.
Shijie Huaren Xiaohua Zazhi 2000; 8: 625-628
12 Fan XQ, Ya JG.
Apoptosis in oncology. Cell Research 2001;11:1-7
13 Zhang MH, Zhang Q,
Shao BX. Effect of Bcl-2 and caspase-3 on calcium distribution in
apoptosis of HL-60 cell.
Cell
Research 2000; 10: 213-20
14 Kane DJ, Sarafian
TA, Anton R, Hahh H, Bntler E. Bcl-2, inhibition of neural death:
decreased generation of reactive
oxygen
species. Science 1993; 262: 1274-1277
15 Reddy BS, Rao CV,
Seibert K. Evaluation of COX-2 inhibitor for potential
chemopreventive properties in colon
carcinogenesis.
Cancer Res 1996; 56: 4566-4569
16 Shibata MA, Hasegawa
R, Imaida K, Hagiwara A. Chemprevention by dehydroepiandrosterone
and indomethacin in a
rat
model of multiorgan carcinogenesis model. Cancer Res 1995; 55:
4870-4874
17 Tomozawa S, Nagawa
H, Tsuno N. Inhibition of haematogenous metastasics of colon cancer
in mice by a selective
COX-2
inhibitor, JTE-522. Br J Cancer 1999; 81: 1274-1279
18 Yang ZY, Rorison KA.
Cyclooxygenase-2-selective antagonists do not inhibit growth of
colorectal carcinoma cell lines.
Cancer
letters 1998; 122: 25-30
19 Kusuhara H, Komatsu
H, Sugahara K. Reactive oxygen species are involved in the apoptosis
induced by NSAIDs in
cultured
gastric cells. Eur J Pharmacol 1999; 383: 331-337
20 Tanaka K, Pracyk JB,
Takeda K, Yu ZX, Finkel T. Expression of IdI results in apoptosis of
cardiac myocytes through a
redox-dependent
mechanism. J Biol Chem 1998; 273: 25922-25828
21 Fridovich I.
Superoxide radical and superoxide dismiutases. Annu Rev Biochem
1995; 64: 97-112
22 Manna SK, Zhang HJ,
Yan T, Oberley LW, Aggarwal BB. Overexpression of manganese
superoxide dismutase suppress
tumor
necrosis factor-induced apoptosis and activation of nuclear
transcription factor-b and AP-1. J Biol Chem 1998;
273:
13245-13254
23 Qiao L, Hanif R,
Sphical E, Steven J, Rigas B. Effect of aspirin on induction of
apoptosis in AGS human colon
adenocarcinoma
cells. Biochem Pharmacol 1998; 55: 53-64
24 J iang ZF, Zhao Y,
Hong X, Zhai ZH. Nuclear apoptosis induced by isolated mitochondria.
Cell Research 2000; 10:
221-232
25 Beg AA, Baltimore D.
An essential role for NF-kB in preventing TNF-alpha induced cell
death. Science 1996; 274:
787-789
26 Cohen GM. Caspases:
the executioners of apoptosis. Biochem J 1997; 326: 1-6
27 Kumar S, Lavin MF.
The ICE family of cysteine proteases as effectors of cell death.
Cell Death Differ 1996; 3: 255-267
28 Salvesen GS and
Dixit VM. Caspases: intracellular signaling by proteolysis. Cell
1997; 91: 443-446
29
Shen ZY, Shen J, Li QS,
Chen CY, Chen JY, Zeng Y. Morphological and functional changes of
mitochondria in apoptosis
esophaged
carcinoma cells induced by arsenic trioxide. World J Gastroenterol
2002;8:31-35
30 Du C, Fang M, Li Y,
Wang X. Smac A. Mitochondrial protein that promotes cytochrome c
dependent caspase activation
by
eliminating IAP inhibition. Cell 2000; 102: 43-53
31 Desagher S, OsenSand
A, Nichols A, Eskes R, Montessuit S, Lauper S, Maundrell K,
Antonsson B, Martinou JC.
Bid-induced
conformational cytochrome c release during aoptosis. J Cell Biol
1999; 144: 891-901
32 Schleggel J, Peters
I, Orrenius S, Miller DK. Cpp32/apopain is a key interleukin 1 beta
converting enzyme-like protease
involved
in Fas-mediated apoptosis. J Biol Chem 1996; 271: 1841-1844
33 Stennicke HR,
Salvesen GS. Properties of the caspases. Biochim Biophys Acta 1998;
1387: 17-31
34 Polyak K, Xia Y,
Zweier JL, Kinzler KW. A model for p53-mediated apoptosis. Nature
1997; 389: 300-305
35 Johnson TM, Yu ZX,
Ferrans RA. Relative oxygen species are downstream mediators of
p53-dependent apoptosis.
Proc
Natl Acad Sci USA 1996; 93: 11848-511852
36 Ossina NK, Cannas A,
Powers VC, Gilbert EM, Tomei SR. Interferon-gamma modulates a
p53-indendent apoptotic
pathway
and apoptosis-related gene expression. J Biol Chem 1997; 272:
16351-16357
37
Xu CT, Huang LT, Pan
BK. Current gene therapy for stomach carcinoma. World J
Gastroenterol 2001;7:752-759
38 Lu YP, Lou YR,
Newmark HL, Huang MT. Enhanced skin carcinogenesis in transgenic
mice with high expression of
glutathione
peroxidase or both glutathione peroxidase and superoxide dismutase.
Cancer Res 1997; 57: 1468-1474
39 Hockenbery OM,
Oltvai ZN, Yin XM, Korsmeyer SJ. Bcl-2 functions in an antioxidant
pathway to prevent apoptosis.
Cell
1993; 75: 241-251
40 Cai J, Jones DP.
Superoxide in apoptosis: mitochondrial generation triggered by
cytochrome C loss. J Biol Chem 1998;
273: 11401-1144
41 Barnes PJ, Karin M.
Nuclear factor-kB, a pivotal transcription factor in chronic
inflammatory disease. New Eng J Med
1997;
336: 1066-1071
42
Huang S, Li JY, Wu J,
Meng L, Shou CC. Mycoplasma infections and different human
carcinomas.
World
J Gastroenterol 2001;7:266-269
43 Baeuerle PA,
Baltimare D. NF-kB: ten years after. Cell 1996; 87: 13-20
44
Wu YL, Sun B, Zhang XJ,
Wang SN, He HY, Qiao MM, Zhong J,Xu JY. Growth inhibition and
apoptosis induction of
Sulindac
on human gastric cancer cells. World J Gastroenterol 2001;7:796-800
45 Kipp E, Ghosh S.
Inhibition of NF-kB by sodium salicylate and aspirin. Science 1994;
265: 956-959
46 Giardina C, Boulares
H, Inan MS. NSAIDs and butyrate sensitize a human colorectal cancer
cell line to TNF and Fas
ligation:
the role of reactive oxygen species. Biochim Biophys Acta 1999;
1448: 425-438
Edited
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