Journal of Cancer Therapy, 2013, 4, 43-48 Published Online January 2013 (
The Influence of Survival Analysis on Runx3 Gene
Expression in the Primary Tumor of Patients Suffering
from Stomach Carcinoma
Hongwei Xu1*, Guangda Xu1, Qingyao Zhang1, Gang Qu1, Yanmin Yu2, Yangsen Jiang1*
1Department of General Surgery and Minimally Invasive Gastrointestinal Surgery, Xinhua Hospital, Dalian University, Dalian, China;
2Huangpu Central Hospital of Shanghai, Shanghai, China.
Email: *
Received October 16th, 2012; revised November 18th, 2012; accepted November 27th, 2012
Background: Runt domain transcription factor 3 (Runx3) is a putative tumor suppressor in human neoplasia. Previous
researches suggested that a lack of Runx3 function contributed to human gastric carcinogenesis, however, it is not clear
whether Runx3 is closely associated with clinicopathological features of primary stomach tumor and survival rate of
patients. Aims: The article is to investigate the influence of survival analysis on Runx3 gene expression in the primary
stomach tumor. Methods: Runx3 mRNA expression was detected in 108 primary gastric tumors and non-tumor tissue
by semiquantitative reverse transcription-PCR (RT-PCR). All patients were followed up more than five years after
radical gastrectomy. Results: There was a loss or substantial decrease of Runx3 mRNA expression in 108 cases of
gastric tumors as compared with that in normal gastric mucosa (p < 0.001). According to the gray scale median of
Runx3 mRNA in primary tumors, the 108 cases were separated into two groups: The lower expressing group (0.403)
and the over one (>0.403). By comparing analysis of clinical information between two groups, it was found that the
lower expression of Runx3 mRNA in the primary tumor was not only associated with the poor clinicopathological fac-
tors, but also the inferior survival duration and cumulative survival rate of patients (p < 0.05). Conclusions: These re-
sults strongly suggest that Runx3 was an independent prognostic factor and a potential therapeutic target for gastric
Keywords: Stomach Neoplasms; Runx3; Prognosis
1. Introduction
Gastric cancer is one of the most common malignancies,
with an extremely poor prognosis, and is the second
leading cause of cancer death worldwide [1]. The preva-
lence of gastric cancer in China is among the highest in
the world, along with Japan and Korea. Despite advances
in its diagnosis and treatment, the prognosis for advanced
gastric cancer is still poor, with a five-year survival rate
less than 10% [2]. By the time the patient is clinically
diagnosed, the gastric cancer has often grown beyond the
limits of curative resection. This reality has raised the-
rapeutic problems, and new early diagnostic tools, thera-
peutic techniques and prognosis concerning this disease
is urgently needed.
Runt domain transcription factors (Runxs) are ho-
mologous to products encoded by the Drosophila seg-
mentation genes runt and lozenge. The Runx gene family
consists of three members, Runx1/AML1, Runx2, and
Runx3 [3]. All three Runx family members play impor-
tant roles in normal developmental processes and in car-
cinogenesis [3-5]. The Runx3 gene is located on human
chromosome 1p36, α region that has long been suspected
to harbor one or more suppressors of various tumors [6].
Previous studies suggest that lack of Runx3 function is
causally related to the genesis and progression of human
gastric cancer, indicating that Runx3 is a novel tumour
suppressor [7]. Runx3 shows remarkable down-regula-
tion in gastric cancers compared to the surrounding mu-
cosa, and the percentage of down-regulation increases as
the cancer stage progresses. Furthermore, Runx3 expres-
sion is reduced in intestinal metaplasia, a precancerous
state, compared with normal mucosa, suggesting that
down-regulation of Runx3 occurs at the early stages of
gastric carcinogenesis, and that loss of Runx3 expression
increases the potential for gastric carcinogenesis [7].
Whether Runx3 is a prognostic marker in gastric can-
cer is still unclear since to date no extensive study has
been performed correlating Runx3 expression with cli-
*Corresponding authors.
Copyright © 2013 SciRes. JCT
The Influence of Survival Analysis on Runx3 Gene Expression in the Primary Tumor of Patients Suffering
from Stomach Carcinoma
nicopathological features and prognosis. We performed
an RT-PCR study to investigate the possible role of the
expression of Runx3 gene in clinicopathology and pro-
gnosis. Our results suggest that there was not only a sig-
nificant correlation of reduced Runx3 mRNA expression
in the primary tumor with the poor clinicopathological
factors and prognosis of stomach carcinoma, but also the
lower five-year survival rate and cumulative survival rate
of patients suffering stomach cacinoma. It is suggested
that Runx3 may play an important role in the evolution
of gastric adenocarcinoma and should be considered as a
potential marker for the prognosis.
This template, created in MS Word 2003 and saved as
“Word 97-2003 & 6.0/95-RTF” for the PC, provides au-
thors with most of the formatting specifications needed
for preparing electronic versions of their papers. All
standard paper components have been specified for three
reasons: 1) ease of use when formatting individual pa-
pers; 2) automatic compliance to electronic requirements
that facilitate the concurrent or later production of elec-
tronic products; and 3) conformity of style throughout a
journal paper. Margins, column widths, line spacing, and
type styles are built-in; examples of the type styles are
provided throughout this document and are identified in
italic type, within parentheses, following the example.
Some components, such as multi-leveled equations, gra-
phics, and tables are not prescribed, although the vari-
ous table text styles are provided. The formatter will
need to create these components, incorporating the ap-
plicable criteria that follow.
2. Materials and Methods
2.1. Tissue Samples
In this study we enrolled 108 patients (74 males and 34
females; age range 28 - 90 yearsmedian age 64 ± 10.5)
who underwent surgery at the Tenth People’s Hospital of
Shanghai Tongji University, China during from Decem-
ber 2000 to December 2003. Tumor tissue and non-neo-
plastic tissue from all patients was collected from the
resected specimen in the operating room within 30 min-
utes after the removal of the stomach. Non-neoplastic
tissue was removed from the normal gastric tissue at a
distance of at least 5 cm from the tumor. Written in-
formed consent was obtained from each patient before
tissue acquisition and all patients had a record of clini-
copathological parameters. The study was approved by
the Ethics Committee of the Tenth People’s Hospital of
Shanghai in accordance with the current Chinese rules.
Tissue sections from all gastric cancer cases were re-
viewed and confirmed by a pathologist. We excluded
cases preoperatively treated with radiation and/or che-
motherapy. All specimens were rapidly frozen in liquid
nitrogen and stored at 80˚C until RNA extraction. All
patients were followed up more than five years after
radical gastrectomy until death or December 2009, which-
ever came first.
2.2. RNA Extraction and Real-Time PCR for
Runx3 mRNA
Total RNAs were obtained from various gastric tissues
including primary gastric cancer and corresponding non-
tumorous gastric mucosa by using the Trizol according to
the manufacture’s protocol (Invitrogen), cDNAs were
synthesized from 1 μg total RNA by random priming
using a First-Strand cDNA Synthesis Kit (Amersham
Biosciences, Buckinghamshire, United Kingdom). The
reaction mixture was incubated for 10 minutes at 25˚C,
then at 37˚C for 120 minutes. cDNA was stored at 80˚C
until use.
Runx3 cDNA was amplified by PCR with the sense
primer, Runx3 gene, sense 5’-ATGACGAGAACTAC-
TCCGCT-3’ and the antisense primer 5’-GGTCGGA-
GAATGGGTTCAGT-3’. As a control, β-actin cDNA
was amplified using the sense 5’-GACGAGGCCCAGA-
GCAAGAGAGG-3’ and the antisense primer (5’-GAT-
tive PCR amplification was carried out in a 10 μL PCR
mixture containing 2 μL of the cDNA template and 0.1
μL of TaKaRa hotstart Taq polymerase, 0.2 mM of de-
oxynucleotide triphosphates, 0.5 μM of each primer
(sense and antisense), 1 μL of the primer 10× PCR buffer,
and 5.6 μL of distilled water. The PCR mixture was am-
plified using GeneAmp PCR system 9600 (Perkin-Elmer,
Wellesley, MA, USA). The PCR conditions were as fol-
lows: preheating at 95˚C for 5 minutes followed by 35
cycles of denaturation for 30 seconds at 95˚C , annealing
for 1 minute at 55˚C, and extension for 1 minute at 72˚C,
with a final extension for 5 minutes at 72˚C. The PCR
products were loaded onto a 2% agarose gel containing
ethidium bromide and visualized under ultraviolet (UV)
2.3. All Patients Were Followed Up
And all patients were followed up more than five years
after radical gastrectomy. Patients were observed until
death or end of follow-up (October 31, 2007), whichever
came first. Ascertainment of deaths included reporting by
the family. In rare patients who died as a result of gastric
cancer not previously reported, we obtained medical re-
cords with permission from next of kin. More than 98%
of deaths in the cohorts were identified by these methods.
2.4. Statistical Analysis
Statistical analysis was performed using Student’s t-test,
χ2-test or two-tailed Fisher’s exact test. Continuous vari-
Copyright © 2013 SciRes. JCT
The Influence of Survival Analysis on Runx3 Gene Expression in the Primary Tumor of Patients Suffering
from Stomach Carcinoma
Copyright © 2013 SciRes. JCT
ables were determined as the mean ± standard deviation.
Survival was analyzed by the Kaplan-Meier method, and
differences in the distribution were evaluated using the
log-rank test. Data analyses were performed using the
SPSS for Windows (V.17.0, SPSS, Chicago, IL, USA),
and p < 0.05 was considered as significant.
adenocarcinoma with lower expression of Runx3 was
55.5% (15 out of 27) at stages I and II, 77.8% (63 out of
81) at stages III and IV, respectivelythus the associa-
tivity between the loss of Runx3 mRNA expression in
the primary tumor and the advanced clinical stage was
statistically significant (p = 0.026) (Table 1). Lower ex-
pression of Runx3 was significantly correlated with dis-
tant metastasis (p = 0.007), LN metastasis (p = 0.035),
infiltrative depth (p = 0.033), and histologic grade (p =
0.034), respectively. But no significant associations were
seen with patient gender (p = 0.628), age (p = 0.633), or
with the growth pattern (p = 0.791) of the tumour (Table 1).
3. Results
3.1. Expression of Runx3 in Gastric
RT-PCR detection of Runx3 expression in normal gastric
mucosa and gastric carcinoma tissues are shown in Fig-
ure 1. Runx3 gene was expressed positively in 21.3%
(23 out of 108) of gastric adenocarcinoma and 62.0% (67
out of 108) of distal normal mucosa. Furthermore very
strong band was identified for Runx3 in normal gastric
muc o sa but more indefinite in the gastric cancer. The
fragment in size is as same as to the design (i.e. 396 bp).
The arithmetic mean on gray scale bands of Runx3
(Runx3/β-anctin) in primary tumors was significantly
lower than in non-tumorous gastric mucosa (0.33 ± 0.12
versus 0.65 ± 0.21, p < 0.001, Figure 2). According to
the gray scale median (0.403, range from 0.068 to 1.211)
of Runx3 mRNA in primary tumors, the 108 cases were
separated into two groups: the lower expressing group
(0.403) and the over one (>0.403).
3.2. Survival Analysis
The median overall survival (OS) for Runx3 mRNA
higher expression (>0.403) and lowed expression (
0.403) was 1345 and 909 days, respectively (log-rank
Figure 1. RT-PCR analyses of Runx3 mRNA expression in
primary gastric cancer tissues and corresponding non-tu-
morous gastric mucosa. Weak or loss expression of Runx3
mRNA in the primary tumor (T). Positive for Runx3
mRNA expression in non-tumorous gastric mucosa (N).
Abbreviations: M, maker.
Association between Runx3 expression and clinicopa-
thologic characteristics of gastric carcinoma patients.
The associations between Runx3 expression in pri-
mary tumors and clinicopathologic features are shown in
Table 1. With respect to the TNM tumor staging gastric
T1 N1 T2 N2 T3 N3
The mean on gray scale bands
Runx3 mRNA
Figure 2. The arithmetic mean on gray scale bands of Runx3 mRNA in primary tumors and non-tumorous gastric mucosa
(semiquantitative RT-PCR; T: the primary tumor; N: the non-tumorous gastric mucosa).
The Influence of Survival Analysis on Runx3 Gene Expression in the Primary Tumor of Patients Suffering
from Stomach Carcinoma
Table 1. Associations betw ee n expr e ssion of Runx3 and clinicopathological features of gastric cancer.
Runx3 mRNA expression n = 108
Cinicopathological Parameters n
0.403 >0.403
male 73 41 32
female 35 19 16
Age (years)
<60 49 28 21
60 59 31 28
Growth pattern
expansive 42 24 18
Infiltrative 66 36 30
Histologic gradec
WD and MD pd 59 45 14
PD 49 28 21
Infiltrative depth
T1+T2 47 32 15
T3+T4 61 52 9
LN metastasis
Absence 34 18 16
Presence 74 54 20
Distant metastasis
Absence 77 64 13
Presence 31 18 13
TNM stage
I and II 27 15 12
III and IV 81 63 18
Abbreviations: LN: lymph node; WD: well differentiated; MD: moderately differentiated; PD: poorly differentiated.
test p = 0.000, Figure 3). By comparing analysis of cli-
nical information between two groups with two-tailed
Fisher’s exact test or Kaplan—Meier method, it was
found that, the loss of Runx3 mRNA expression in the
primary tumor was not only associated with the poor
clinicopathological factors, but also the lower five-year
survival rate and overall survival rate of patients suffer-
ing stomach carcinoma (p < 0.05). Follow-up data showed
that a significantly decreasing trend in 5-year survival
was observed in primary tumors patients with the lower
expression of Runx3 (from 25% of the higher one to 13%
of the lower one, p < 0.005).
4. Discussion
The aggressive nature of human metastatic gastric car-
cinoma is related to genetic instability, reactivation of
telomerase, mutations of various oncogenes and tumor
suppressor genes [8] and abnormalities in several growth
factors and their receptors. But no single factor that ac-
counts for the majority of cases has been identified [9].
These abnormalities affect the downstream signal trans-
duction pathways involved in the control of cell growth
and differentiation. Specifically, they confer a tremend-
ous survival and development and progression to gastric
The Runt domain transcription factors 3 have been re-
ported to play key roles in developmental pathways in
various types of cancers [3-5]. Both human and animal
studies in vivo and in vitro had indicated that Runx3 is
closely related to gastric carcinoma. Silencing of the
Runx3 gene may induce many epithelial malignancies,
including gastric cancer [10-12]. Enhanced expression of
Runx3 has been demonstrated to inhibit gastric carci-
noma cell growth in vitro, as well as tumorigenicity and
metastasis. Moreover, Runx3 is inactivated in more than
60% of human gastric cancers and various human gastric
cancer cell lines have decreased Runx3 expression [12,
13]. Li et al. found that Runx3 expression was reduced in
Copyright © 2013 SciRes. JCT
The Influence of Survival Analysis on Runx3 Gene Expression in the Primary Tumor of Patients Suffering
from Stomach Carcinoma
Figure 3. Kaplan-Meier survival analysis for gastric cancer patients according to the expression of Runx3. The y-axis repre-
sents the percentage of patients; the x-axis, their survival in days. The green line represents Runx3 mRNA lower expression
in gastric carcinoma patients with a trend of worse survival than the blue line representing Runx3 mRNA higher expression
in gastric carcinoma patients (Log-rank test: p = 0.000). Mean survival times were 909 days for the Runx3 mRNA lower ex-
pression group and 1345 days for the Runx3 mRNA higher expression group.
40% of early stage carcinomas, and the level increased to
nearly 90% with the advancement of the cancer stage
[12]. Wei et al. found that loss of Runx3 expression sig-
nificantly affected the clinical outcome of gastric can-
cer patients [13]. Even though accumulating data sug-
gests that Runx3 may also be a potential tumor suppres-
sor in several tumors including gastric cancer [12,14,15],
systematic study about its association with clinicopa-
thological features of primary stomach tumor and sur-
vival rate of patients are still lacking.
In this study, RT-PCR was used to confirm the expres-
sion level of Runx3 mRNA in paired samples of primary
gastric cancer and non-tumorous tissue, and research data
has shown that expression of the Runx3 gene was more
frequently in the non-tumorous gastric mucosa than in
the primary gastric cancer, suggesting the involvement of
the tumor suppressor gene Runx3 in the pathogenesis of
gastric carcinoma. Specifically, we found that the loss of
Runx3 mRNA expression in the primary tumor was not
only associated with the poor clinicopathological factors
such as deep infiltration, distant organ metastasis, poorly
differentiation, lymph metastasis and later clinicopa-
thological stages, etc, but also the lower five-year sur-
vival rate and cumulative survival rate of patients suffer-
ing stomach carcinoma. The prognosis data showed that
the decrease or loss of Runx3 expression was inversely
correlated with survival and could be an independent
predictor of poor patient outcome.
Recently, studies have reported that silencing a tumor
suppressor gene may result in loss of its function in tu-
morigenesis [16]. Inactivation mechanisms of the Runx3
gene are believed to be promoter hypermethylation and
homozygous deletion [16,17]. Genetic variants of the
Runx3 gene may also play an inactivation role. For ex-
ample, Kim et al. [17]. demonstrated Runx3 methylation
in 8.1% of chronic gastritis specimens, 28.1% of intesti-
nal metaplasia specimens, 27.3% of gastric adenoma spe-
cimens and 64% of gastric carcinoma specimens, sug-
gesting increase of Runx3 methylation with progression
of the lesion along the path of multistep gastric carcino-
genesis. Li et al. found that the Runx3 Arg122Cys muta-
tion within the Runx3 conserved Runt domain could af-
fect the tumorsuppressive activity of Runx3 [12]. Also,
Kim et al. examined mutations in the Runx3 coding re-
gions in 34 bladder tumors and found missense mutations
and single nucleotide deletion in the conserved Runt do-
main that abolished the DNA-binding ability of Runx3
and resulted in truncation of the protein [18]. This evi-
dence generated the hypothesis that Runx3 genetic vari-
ants may affect the functions of Runx3, and consequently
modulate the cell growth and apoptosis capacity of
TGF-β, and participate in the etiology of human cancers
In conclusion, we have examined the expression of
Runx3 mRNA in human gastric cancer and its prognostic
significance. This study has shown for several strengths
that 1) a low proportion of gastric carcinomas expressed
Runx3 and low expression of Runx3 was significantly
associated with unfavorable clinicopathologic variables
such as low clinical stage (stages III and IV), lymph node
Copyright © 2013 SciRes. JCT
The Influence of Survival Analysis on Runx3 Gene Expression in the Primary Tumor of Patients Suffering
from Stomach Carcinoma
metastasis, histologic grade and distant metastasis, how-
ever, there are a few different with previous studies. The
reasons for this result may be caused by the long-term
prognosis of patients with gastric cancer is influenced by
not only Runx3 but also other factors such as patient’s
performance, tumor stage, ethnic differences, and the
performance of gastrectomy; and 2) it is the first study to
investigated the association between the Runx3 mRNA
expression and survival rate in the primary tumor of pa-
tients suffering from stomach carcinoma, and low ex-
pression of Runx3 was associated with a significantly
shorter survival in gastric carcinoma patients. Taken to-
gether, our findings indicate that Runx3 may be involved
in the progression of gastric carcinomas and are signifi-
cant prognostic indicators for gastric carcinoma patients.
5. Acknowledgements
This study was supported by the National Natural Sci-
ence Foundation of China, No. 30171039; Our work was
finished in Shanghai Institute of Endocrine and Meta-
bolic Diseases, Rui-Jin Hospital, Shanghai Jiao Tong
University School of Medicine, China and thank Dr.
Xiaoying Li for the kind help.
[1] F. Pasini, A. P. Fraccon and G. D. E. Manzoni, “The Role
of Chemotherapy in Metastatic Gastric Cancer,” Anti-
cancer Research, Vol. 31, No. 10, 2011, pp. 3543-3554.
[2] V. Catalano, R. Labianca, G. D. Beretta, G. Gatta, F. de
Braud and E. Van Cutsem, “Gastric Cancer,” Critical Re-
views in Oncology/Hematology, Vol. 71, No. 2, 2009, pp.
127-164. doi:10.1016/j.critrevonc.2009.01.004
[3] Y. Ito, “Oncogenic Potential of the Runx Gene Family:
‘Overview’,” Oncogene, Vol. 23, No. 24, 2004, pp. 4198-
4208. doi:10.1038/sj.onc.1207755
[4] S. C. Bae and J. K. Choi, “Tumor Suppressor Activity of
Runx3,” Oncogene, Vol. 23, No. 24, 2004, pp. 4336-4340.
[5] K. Blyth, E. R. Cameron and J. C. Neil, “The Runx Genes:
Gain or Loss of Function in Cancer,” Nature Reviews
Cancer, Vol. 5, No. 5, 2005, pp. 376-387.
[6] Y. Ito, “Runx Genes in Development and Cancer: Regu-
lation of Viral Gene Expression and the Discovery of
Runx Family Genes,” Advances in Cancer Research, Vol.
99, 2008, pp. 33-76.
[7] Y. Nakase, C. Sakakura, K. Miyagawa, et al., “Frequent
Loss of Runx3 Gene Expression in Remnant Stomach
Cancer and Adjacent Mucosa with Special Reference to
Topography,” British Journal of Cancer, Vol. 92, No. 3,
2008, pp. 562-569.
[8] B. G. Jang and W. H. Kim, “Molecular Pathology of
Gastric Carcinoma,” Pathobiology, Vol. 78, No. 6, 2011,
pp. 302-310. doi:10.1159/000321703
[9] J. P. Hamilton, and S. J. Meltzer, “A Review of the Ge-
nomics of Gastric Cancer,” Clinical Gastroenterology
and Hepatology, Vol. 4, No. 4, 2006, pp. 416-425.
[10] H. Shiraha, S. Nishina and K. Yamamoto, “Loss of Runt-
Related Transcription Factor 3 Causes Development and
Progression of Hepatocellular Carcinoma,” Journal of
Cellular Biochemistry, Vol. 112, No. 3, 2011, pp. 745-
749. doi:10.1002/jcb.22973
[11] N. Yanagawa, G. Tamura, H. Oizumi, et al., “Promoter
Hypermethylation of RASSF1A and Runx3 Genes as an
Independent Prognostic Prediction Marker in Surgically
Resected Non-Small Cell Lung Cancers,” Lung Cancer,
Vol. 58, No. 1, 2007, pp. 131-138.
[12] E. J. Kim, Y. J. Kim, P. Jeong, Y. S. Ha, S. C. Bae and W.
J. Kim, “Methylation of the Runx3 Promoter as a Poten-
tial Prognostic Marker for Bladder Tumor,” Journal of
Urology, Vol. 180, No. 3, 2008, pp. 1141-1145.
[13] D. Wei, W. Gong, S. C. Oh, et al., “Loss of Runx3 Ex-
pression Significantly Affects the Clinical Outcome of
Gastric Cancer Patients and Its Restoration Causes Dras-
tic Suppression of Tumor Growth and Metastasis,” Can-
cer Research, Vol. 65, No. 11, 2005, pp. 4809-4816.
[14] Y. Tokumaru, S. Nomoto, C. Jeronimo, et al., “Biallelic
Inactivation of the RIZ1 Gene in Human Gastric Cancer,”
Oncogene, Vol. 22, No. 44, 2003, pp. 6954-6958.
[15] A. Goel, C. N. Arnold, P. Tassone, et al., “Epigenetic
Inactivation of Runx3 in Microsatellite Unstable Sporadic
Colon Cancers,” International Journal of Cancer, Vol.
112, No. 5, pp. 754-759. doi:10.1002/ijc.20472
[16] D. Levanon, Y. Bernstein, V. Negreanu, K. R. Bone, A.
Pozner, R. Eilam, J. Lotem, O. Brenner and Y. Groner,
“Absence of Runx3 Expression in Normal Gastroin-
testinal Epithelium Calls into Question Its Tumour Sup-
pressor Function,” EMBO Molecular Medicine, Vol. 3,
No. 10, 2011, pp. 593-604.
[17] T. Y. Kim, H. J. Lee, K. S. Hwang, et al., “Methylation of
Runx3 in Various Types of Human Cancers and Prema-
lignant Stages of Gastric Carcinoma,” Laboratory Inves-
tigation, Vol. 84, No. 4, 2004, pp. 479-484.
[18] W. J. Kim, “Runx3 Inactivation by Point Mutations and
Aberrant DNA Methylation in Bladder Tumors,” Cancer
Research, Vol. 65, No. 20, 2005, pp. 9347-9354.
[19] M. Suzuki, H. Suzuki, Y. Minegishi, K. Ito, T. Nishiza-
waand and T. Hibi, “H. Pylori-Eradication Therapy In-
creases Runx3 Expression in the Glandular Epithelial Cells
in Enlarged-Fold Gastritis,” Journal of Clinical Biochemis-
try and Nutrition, Vol. 46, No. 3, 2010, pp. 259-264.
Copyright © 2013 SciRes. JCT