Publications by authors named "Zhong-Ying Shen"

51 Publications

Prevalence and association of human papillomavirus 16, Epstein-Barr virus, herpes simplex virus-1 and cytomegalovirus infection with human esophageal carcinoma: a case-control study.

Oncol Rep 2011 Jun 23;25(6):1731-8. Epub 2011 Mar 23.

Cardiovascular Research Center, Shantou University Medical College, Shantou 515041, PR China.

Recent research shows esophageal carcinoma (EC) as the ninth most common malignancy in the world. The association of viral infection and EC has been reported in the last 30 years. However, geographic variation in infection rates and the key mechanisms of the viral action have yet to be resolved. This study aimed to determine the prevalence and association of human papillomavirus 16 (HPV-16), herpes simplex virus 1 (HSV-1), Epstein-Barr virus (EBV) and cytomegalovirus (CMV) infection in the etiology of EC in the area of Shantou, Guangdong, China. Nested PCR was used to detect viral DNA in the mucosa of 70 cases of EC and in paracancerous tissues, as well as 100 cases of normal esophagus mucosa. Data were analyzed by χ2 test, Fisher's exact test and bivariate correlation analysis. The infection rates of HPV-16, HSV-1 and EBV were 40.0, 30.0 and 30.0%, respectively, in EC mucosa, and were significantly higher than those in normal mucosa. However, no CMV DNA was detected in either EC or normal mucosa. HPV-16 or EBV infection was mainly detected in EC patients 48-58 years old, and the infection rate was positively associated with pathological grade of EC (P<0.05). Tobacco smoking and alcohol consuption were high risk factors for HPV-16 infection for male patients [odds ratio (OR), 5.9; 95% confidence interval (CI), 1.4-24.6; OR = 3.8; 95% CI, 1.1-13.8]. Rates of infection with a mixture of these 3 viruses were all more than 10.0% in cancerous mucosa and closely related to the pathological grade of EC (P = 0.001). Infection with HPV-16, HSV-1 or EBV may be an important etiological factor in EC.
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http://dx.doi.org/10.3892/or.2011.1234DOI Listing
June 2011

NGAL and NGALR are frequently overexpressed in human gliomas and are associated with clinical prognosis.

J Neurooncol 2011 Aug 24;104(1):119-27. Epub 2010 Dec 24.

Department of Neurosurgery, Affiliated Shantou Hospital of Sun Yat-Sen University, Shantou, China.

Recently, neutrophil gelatinase-associated lipocalin (NGAL) and its cell surface receptor, NGALR, have been shown to have critical roles in the biology of various tumors. Therefore, we investigated the expression of NGAL and NGALR in tumor sections obtained from patients with gliomas, and compared these results with the clinical characteristics of the patients. Using immunohistochemical assays, the expression levels of NGAL and NGALR were found to be up-regulated in tumor tissues, and to be related to tumor grade (p < 0.001). A positive correlation between expression of the two markers was also observed in these assays (r = 0.849; p < 0.001). Overexpression of NGAL and NGALR in glioma tissues was also confirmed in western blot analysis and real-time quantitative RT-PCR assays. Furthermore, overexpression of NGAL and NGALR was found to be significantly associated with poor prognosis (p < 0.001 in each case). Multivariate analysis identified patient age, tumor grade, and expression levels of NGAL and NGALR to be independent prognostic factors. In particular, NGAL(2+)/NGALR(2+) tissues were associated with lower rates of survival (risk ratio, 1.378; 95% CI, 1.102-1.724; p = 0.005). These findings suggest that NGAL and NGALR expression are frequently up-regulated in gliomas, and are closely associated with poor clinical outcome.
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http://dx.doi.org/10.1007/s11060-010-0486-0DOI Listing
August 2011

Infection and integration of human papillomavirus in esophageal carcinoma.

Int J Hyg Environ Health 2011 Mar 3;214(2):156-61. Epub 2010 Dec 3.

Department of Preventive Medicine, Shantou University Medical College, Shantou, China.

Infection with human papillomavirus (HPV) could be a suspected or potential modifiable risk factor in esophageal carcinoma (EC) but findings have not been consistent. We therefore investigated the epidemiology of HPV infection and integration in the pathogenesis of esophageal carcinoma (EC) in the Shantou region, China. This was a retrospective study involving nested PCR to evaluate HPV presence, HPV genotyping, and analyzing HPV-16 integration status in esophageal tumor tissues (n=106) and paired tumor-adjacent normal tissues, as well as normal esophagus tissue from control subjects (n=100). The detection rates of HPV DNA in EC and tumor-adjacent tissue were significantly higher than that in normal controls (77.4% and 80.2% vs. 33.0%). HPV infection was mainly found in adults, ages 35-47 years old, and the infection rate was negatively associated with the age of EC patients (P-trend<0.05). In addition, the HPV infection rates in patients who smoked was 3.27 times higher than in non-smoking patients (84.9% vs. 67.4%, P<0.05) but was not associated with gender, alcohol consumption, tumor grade or lymph-node metastasis of EC patients. The distribution of HPV genotypes in patients from high to low proportion was HPV-16, -58, -18, -33, -31 and -11. Infection with multiple HPV genotypes mainly included HPV-16/-18 and HPV-16/-33. The integration rate of HPV-16 in EC tissue was higher than that in tumor-adjacent and control tissues (93.4% vs. 50.9% and 45.5%). Our findings indicate that infection with HPV, especially the high-risk HPV, and their integration suggest an association in malignant transformation of EC in the high-incidence EC region in Shantou, China.
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http://dx.doi.org/10.1016/j.ijheh.2010.11.001DOI Listing
March 2011

Genome-wide association study of esophageal squamous cell carcinoma in Chinese subjects identifies susceptibility loci at PLCE1 and C20orf54.

Nat Genet 2010 Sep 22;42(9):759-63. Epub 2010 Aug 22.

Cancer Research Center, Xinxiang Medical University, Xinxiang, Henan, China.

We performed a genome-wide association study of esophageal squamous cell carcinoma (ESCC) by genotyping 1,077 individuals with ESCC and 1,733 control subjects of Chinese Han descent. We selected 18 promising SNPs for replication in an additional 7,673 cases of ESCC and 11,013 control subjects of Chinese Han descent and 303 cases of ESCC and 537 control subjects of Chinese Uygur-Kazakh descent. We identified two previously unknown susceptibility loci for ESCC: PLCE1 at 10q23 (P(Han combined for ESCC) = 7.46 x 10(-56), odds ratio (OR) = 1.43; P(Uygur-Kazakh for ESCC) = 5.70 x 10(-4), OR = 1.53) and C20orf54 at 20p13 (P(Han combined for ESCC) = 1.21 x 10(-11), OR = 0.86; P(Uygur-Kazakh for ESCC) = 7.88 x 10(-3), OR = 0.66). We also confirmed association in 2,766 cases of gastric cardia adenocarcinoma cases and the same 11,013 control subjects (PLCE1, P(Han for GCA) = 1.74 x 10(-39), OR = 1.55 and C20orf54, P(Han for GCA) = 3.02 x 10(-3), OR = 0.91). PLCE1 and C20orf54 have important biological implications for both ESCC and GCA. PLCE1 might regulate cell growth, differentiation, apoptosis and angiogenesis. C20orf54 is responsible for transporting riboflavin, and deficiency of riboflavin has been documented as a risk factor for ESCC and GCA.
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http://dx.doi.org/10.1038/ng.648DOI Listing
September 2010

Phosphorylation of fascin decreases the risk of poor survival in patients with esophageal squamous cell carcinoma.

J Histochem Cytochem 2010 Nov 16;58(11):979-88. Epub 2010 Aug 16.

Institute of Oncologic Pathology, Medical College of Shantou University, Shantou, Guangdong, China.

Phosphorylation of fascin at serine 39 (phospho-S39-fascin) could inhibit its actin-binding and actin-bundling activities and decrease filopodia formation. However, the relationship between phospho-S39-fascin expression and clinicopathological parameters in tumors is still unknown. Here, Western blot analysis and IHC applied to tissue microarray technology were performed to examine the expression status of non-phosphorylated fascin (fascin) and phospho-S39-fascin and their impacts on the prognosis of patients with esophageal squamous cell carcinoma (ESCC). Fascin and phospho-S39-fascin expressions were tested by cytoplasmic staining. Among the 254 patients, 90 cases showed high expression of fascin and 87 cases showed high expression of phospho-S39-fascin. Survival analysis showed that high expression of fascin was significantly associated with a poor prognosis of the patients with ESCC (p=0.004). In contrast, high expression of phospho-S39-fascin correlated significantly with an improved outcome of patients (p=0.020). Multivariate analysis showed that both fascin and phospho-S39-fascin were independent prognostic factors. In a combined analysis, the patients with high expression of fascin and low expression of phospho-S39-fascin tumors had a shorter overall survival than those with high expression of both fascin and phospho-S39-fascin tumors (5-year overall survival rate: 28.7% vs 48.3%, p=0.068). Our results suggest that high expression of fascin correlates with poor outcome and that high expression of phospho-S39-fascin decreases the risk of poor prognosis in ESCC. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.
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http://dx.doi.org/10.1369/jhc.2010.955765DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2958140PMC
November 2010

Bronchogenic and alveologenic tumors in mice induced by N-nitrosopiperidine.

Biochem Cell Biol 2010 Aug;88(4):775-82

Department of Tumor Medicine, The First Affiliated Hospital, Shantou University Medical College, Shantou, P.R. China.

The aim of this study was to explore the histogenesis and carcinogenesis of pulmonary cancer induced by N-nitrosopiperidine (NPIP) in mice. NPIP is a form of N-nitrosamine found in tobacco smoke, which has been shown to be a genotoxic chemical as well as a mutagenic compound for inducing chromosome aberrations and severe clastogenicity. In this study, 80 BALB/C strain mice were injected with 0.2 mmol/kg NPIP intraperitoneally for 8 weeks, and experiments were conducted for a further 16 weeks. For the control group, 40 mice were injected with an equal volume of 0.9% NaCl. Pulmonary tissues and tumors in the NPIP-treated group were examined by light microscopy and transmission electron microscopy and compared with the control group at 4-week intervals. The mRNA levels of p53 (mutant), bcl-2, c-myc, ras, and subunits of telomerase - telomerase reverse transcriptase (TERT) and an RNA component, TR - were assayed by mPCR or RT-PCR. Twenty-two mice in the experimental group were found to develop pulmonary tumors, but none in the control group. All tumors found in the experimental group originated from alveolar type II epithelial cells. In addition, 6 of the 22 mice also developed tumors of bronchogenic origin. The expression of p53, bcl-2, c-myc, ras, and the subunits of telomerase were found to increase in all pulmonary tissues and tumors formed thereafter upon NPIP treatment. In summary, NPIP-induced mouse lung tumors exhibited morphological changes during carcinogenesis, which may be the consequence of overexpression of some genes associated with the development of carcinoma and changes in subunits of telomerase. This mouse model of lung tumor formation may be a useful tool to delineate the histogenesis and carcinogenesis of human pulmonary cancer.
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http://dx.doi.org/10.1139/O10-019DOI Listing
August 2010

Reduced membranous and ectopic cytoplasmic expression of DSC2 in esophageal squamous cell carcinoma: an independent prognostic factor.

Hum Pathol 2010 Oct;41(10):1456-65

The Key Immunopathology Laboratory of Guangdong Province, Institute of Oncologic Pathology, Medical College of Shantou University, Shantou, China.

Desmocollin 2, a desmosomal component, is a key membrane glycoprotein critically involved in cell-cell adhesion and the maintenance of normal tissue architectures in epithelia. Reports exploring the link of desmocollin expression to cancers are limited. The aim of this study was to investigate the expression of desmocollin 2 in esophageal squamous cell carcinoma and, in particular, to determine the extent to which the patterns of desmocollin 2 expression correlated with the clinical parameters. Desmocollin 2 expression was evaluated in 308 cases of esophageal squamous cell carcinoma using immunohistochemistry. Western blotting and reverse transcriptase polymerase chain reaction were performed to characterize the relative expression levels of desmocollin 2 isoforms. The results indicated that desmocollin 2 expression was reduced significantly in esophageal cancer in both protein and messenger RNA levels and that this reduction was associated with poor survival (P = .011). The expression of desmocollin 2 was prominent in normal esophageal epithelia and highly differentiated esophageal tumors, but was reduced or absent in poorly differentiated tumor specimens. Furthermore, in 74.7% of tumor tissues, desmocollin 2 immunoreactivity displayed an abnormal cytoplasmic localization that was correlated with poor tumor differentiation (P < .001), regional lymph node metastasis (P < .001), pathologic tumor-node-metastasis stages (P < .001), and poor prognosis (P = .048). Multivariate analysis showed that desmocollin 2 expression level was an independent prognostic factor for esophageal squamous cell carcinoma. These data suggest that desmocollin 2 is involved in the transformation and development of esophageal tumors and that desmocollin 2 expression level and intracellular localization may serve as a predictor for patient outcomes.
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http://dx.doi.org/10.1016/j.humpath.2010.04.003DOI Listing
October 2010

Specificity protein 1 regulates fascin expression in esophageal squamous cell carcinoma as the result of the epidermal growth factor/extracellular signal-regulated kinase signaling pathway activation.

Cell Mol Life Sci 2010 Oct 26;67(19):3313-29. Epub 2010 May 26.

Institute of Oncologic Pathology, The Key Immunopathology Laboratory of Guangdong Province, Shantou University Medical College, No. 22, Xinling Road, Shantou 515041, People's Republic of China.

The overexpression of fascin in human carcinomas is associated with aggressive clinical phenotypes and poor prognosis. However, the molecular mechanism underlying the increased expression of fascin in cancer cells is largely unknown. Here, we identified a Sp1 binding element located at -70 to -60 nts of the FSCN1 promoter and validated that Sp1 specifically bound to this element in esophageal carcinoma cells. Fascin expression was enhanced by Sp1 overexpression and blocked by Sp1 RNAi knockdown. Specific inhibition of ERK1/2 decreased phosphorylation levels of Sp1, and thus suppressed the transcription of the FSCN1, resulting in the down-regulation of fascin. Stimulation with EGF could enhance fascin expression via activating the ERK1/2 pathway and increasing phosphorylation levels of Sp1. These data suggest that FSCN1 transcription may be subjected to the regulation of the EGF/EGFR signaling pathway and can be used as a viable biomarker to predict the efficacy of EGFR inhibitors in cancer therapies.
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http://dx.doi.org/10.1007/s00018-010-0382-yDOI Listing
October 2010

The expression of CFL1 and N-WASP in esophageal squamous cell carcinoma and its correlation with clinicopathological features.

Dis Esophagus 2010 Aug 20;23(6):512-21. Epub 2010 Jan 20.

Department of Cardiothoracic Surgery of First Affiliated Hospital, Medical College of Shantou University, Shantou, China.

Cofilin1 (CFL1) is an actin-modulating protein, which belongs to the ADF/Cofilin family. Neural Wiskott-Aldrich syndrome protein (N-WASP) is the key regulator of the actin cytoskeleton, a member of Wiskott-Aldrich syndrome protein family. They have been suggested to be involved in cancer cell invasion and metastasis. In this study, the expression patterns of CFL1 and N-WASP in normal esophageal mucosa and esophageal squamous cell carcinoma (ESCC) and their correlation with clinical characteristics were investigated. Immunohistochemical staining showed that CFL1 was expressed in nuclear and cytoplasm of cancer cells. However, N-WASP was mainly found in the cytoplasm of the cancer cells. There were significant evidences that proved that CFL1 is correlated with clinicopathological factors in ESCC, such as infiltration depth, lymph node metastasis and pathological staging (P < 0.05). It is also proved that N-WASP is related to lymph node metastasis and pathological staging in ESCC (P < 0.05). Kaplan-Meier analysis showed that there was no correlation between CFL1 and N-WASP protein expression and survival (P > 0.05). Moreover, the mRNA expression of CFL1 and N-WASP was detected by quantitative real time PCR in 70 tissue specimens. The results showed that CFL1 mRNA level was over-expressed in ESCC tissue (P < 0.05), while N-WASP mRNA expression level was not different between cancerous tissues and adjacent normal esophageal mucosa (P > 0.05). Also, CFL1 mRNA expression was significantly associated with regional lymph node metastasis and pathological staging (P < 0.05). Kaplan-Meier analysis showed that there was no correlation between CFL1 and N-WASP mRNA expression and survival (P > 0.05). Our findings suggested that CFL1 and N-WASP may play an important role in the tumorigenesis of ESCC, and to be the candidate novel biomarkers for the diagnosis and prognosis of ESCC. These findings may have implications for targeted therapies in patients with ESCC.
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http://dx.doi.org/10.1111/j.1442-2050.2009.01035.xDOI Listing
August 2010

Involvement of CYR61 and CTGF in the fascin-mediated proliferation and invasiveness of esophageal squamous cell carcinomas cells.

Am J Pathol 2010 Feb 7;176(2):939-51. Epub 2010 Jan 7.

Department of Biochemistry and Molecular Biology, the Key Immunopathology Laboratory of Guangdong Province, Medical College of Shantou University, Shantou 515041, China.

Fascin is overexpressed in esophageal squamous cell [corrected] carcinoma (ESCC) and involved in the proliferation and invasiveness of ESCC cells. In this study, we retrospectively examined the expression of fascin in ESCC samples by immunohistochemistry and revealed that overexpression of fascin was related to poor patient survival. RNAi-mediated knockdown of fascin in ESCC cells significantly inhibited cell proliferation and invasiveness, whereas forced expression of fascin in immortalized esophageal epithelial cells accelerated cell proliferation and invasiveness. To explore the underlying mechanism, cDNA microarray was performed to identify the differential gene expression profiles between a fascin-depleted cell line by RNAi and the corresponding control ESCC cells. Results showed that 296 genes were differentially expressed on fascin depletion. In this study, we focused on two down-regulated genes: CYR61 and CTGF. We found that restored expression of either CYR61 or CTGF led to a recovery of the suppression of cellular proliferation and invasiveness induced by down-regulation of fascin expression; the protein level of CYR61 and CTGF were up-regulated in ESCCs and their expression pattern correlated with fascin overexpression. Finally, analysis of signal transduction revealed that fascin affected the expressions of CYR61 and CTGF through transforming growth factor (TGF)-beta pathway. Taken together, we propose that fascin regulates the proliferation and invasiveness of ESCC cells by modulating the expression of CTGF and CYR61 via TGF-beta pathway.
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http://dx.doi.org/10.2353/ajpath.2010.090118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2808098PMC
February 2010

Overexpression of neutrophil gelatinase-associated lipocalin and its receptor in colorectal carcinoma: Significant correlation with cell differentiation and tumour invasion.

Oncol Lett 2010 Jan 1;1(1):103-108. Epub 2010 Jan 1.

Department of Biochemistry and Molecular Biology, Medical College of Shantou University, Shantou 515041, P.R. China.

Neutrophil gelatinase-associated lipocalin (NGAL), a member of the lipocalin family, is related to imflammation and tumour. Recently, a specific cell-surface receptor (24p3R/NGALR) for lipocalin 24p3 was reported. However, the characteristics of NGALR expression in colorectal carcinoma (CRC) are not known. The objectives of this study were to investigate the expression of NGAL and NGALR in CRC specimens, and determine any relationship between the expression of these proteins and tumour progression. In the present study, CRC specimens of 102 patients were obtained, and the expression of NGAL, NGALR, ferritin and Ki67 was analyzed in paraffin sections by immunohistochemistry. Statistical analyses of the data collected were performed with SPSS software. We found that the cytoplasmic staining of NGAL, NGALR and ferritin, as well as the nuclear staining of Ki67 were significantly up-regulated in CRC tissues compared with normal colorectal tissues. Expression of NGAL was related to the deeper invasion of CRC (P=0.026), while NGALR was significantly associated with a deeper invasion (P=0.018) and a high degree of Tumor, Node and Metastasis stages (P=0.042) in CRC. The NGAL/NGALR co-expression was associated with poor cellular differentiation (P=0.004). Positive correlations between NGAL and NGALR (r=0.432, P<0.01), NGAL and ferritin (r=0.374, P<0.001), NGALR and Ki67 (r=0.228, P<0.05), NGAL/NGALR co-expression and ferritin (r=0.349, P<0.001), as well as NGAL/NGALR co-expression and Ki67 (r=0.205, P<0.05) were observed. However, the expression of NGAL or NGALR was not significantly associated with patient survival. These findings detected an elevated expression of NGAL and NGALR resulting in poor cellular differentiation and a deeper invasion of CRC. Thus, NGALR may be a novel target for the treatment of CRC.
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http://dx.doi.org/10.3892/ol_00000019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3436261PMC
January 2010

Expression and prognostic significance of THBS1, Cyr61 and CTGF in esophageal squamous cell carcinoma.

BMC Cancer 2009 Aug 22;9:291. Epub 2009 Aug 22.

Department of Biochemistry and Molecular Biology, Shantou University, Shantou, PR China.

Background: Thrombospondin1 (THBS1), cystene-rich protein 61 (Cyr61) and connective tissue growth factor (CTGF) are all involved in the transforming growth factor-beta (TGF-beta) signal pathway, which plays an important role in the tumorigenesis. The purpose of this study is to explore the expression and prognostic significance of these proteins in esophageal squamous cell carcinoma (ESCC).

Methods: We used immunohistochemistry and western blotting to examine the expression status of THBS1, Cyr61 and CTGF in ESCC. Correlations of THBS1, Cyr61 and CTGF over-expressions with various clinicopathologic factors were also determined by using the Chi-square test or Fisher's exact probability test. Survival analysis was assessed by the Kaplan-Meier analysis and the log-rank test. Relative risk was evaluated by the multivariate Cox proportional hazards model.

Results: THBS1, Cyr61 and CTGF were all over-expressed in ESCC. THBS1 over-expression was significantly associated with TNM stage (P = 0.029) and regional lymph node involvement (P = 0.026). Kaplan-Meier survival analysis showed that over-expression of THBS1, Cyr61 or CTGF was related to poor survival of ESCC patients (P = 0.042, P = 0.020, P = 0.018, respectively). Multivariate Cox analysis demonstrated that Cyr61 and CTGF were independent factors in prognosis of ESCC.

Conclusion: Cyr61, CTGF and THBS1 were all over-expressed in ESCC and might be new molecular markers to predict the prognosis of ESCC patients.
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http://dx.doi.org/10.1186/1471-2407-9-291DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2748090PMC
August 2009

Prognostic significance of perigastric lymph nodes metastases on survival in patients with thoracic esophageal cancer.

Dis Esophagus 2010 Jan 15;23(1):40-5. Epub 2009 Apr 15.

Department of Oncology Surgery, Affiliated Shantou Hospital of Sun Yat-sen University, and Department of Anatomy, Medical College of Shantou University, Shantou, China.

Several publications have showed that the number of metastatic lymph node (LN) should be taken into consideration in nodal category of esophageal cancer, but seldom considered extent of involved regional LNs. The aim of this study is to evaluate the significance of the extent of regional LN metastasis on survival in patients with esophageal cancer. A total of 245 thoracic esophageal cancer patients underwent transthoracic esophagectomy with standard lymphadenectomy between January 2000 and December 2006 were included in the study. Data including demographic factors, pathologic findings, LN parameters and survival outcomes were collected. The survival experience was depicted using Kaplan-Meier method. A multivariate Cox proportional hazard model was used to screen the significant prognostic factors. The univariate analysis to further explore the significant prognostic factor was done by log-rank test. After a median follow-up of 53.2 months, the 5-year survival rate was 46.3% for the entire cohort. Cox model regression indicated that the LN status and perigastric nodal status, aside from residual tumor status, histological tumor type and depth of invasion, were the independent prognostic factors. Patients without LN metastasis had better 5-year survival than those with positive nodes (64.2% vs. 18.9%, X2=35.875, P<0.001). However, For those patients with nodal involvement, there was no difference in 5-year survival between patients with involved nodes<3 and >or=3 (27.8% vs. 0%, X2=0.925, P=0.336). When considering the location of LN metastasis, patients could be further stratified according to whether the perigastric nodes were involved or not (37.5% vs. 10.0%, X2=4.295, P=0.038). In conclusion, involved LN number had no prognostic implication in nodal involved patients based on our data. Whereas, perigastric nodal involvement should be used to refine the N category (N0, no nodal metastasis, N1, non-perigastric node metastasis, N2, perigastric node metastasis) for the future esophageal cancer staging criteria.
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http://dx.doi.org/10.1111/j.1442-2050.2009.00964.xDOI Listing
January 2010

Roles of ezrin in the growth and invasiveness of esophageal squamous carcinoma cells.

Int J Cancer 2009 Jun;124(11):2549-58

Department of Biochemistry and Molecular Biology, Medical College of Shantou University, Shantou, People's Republic of China.

Ezrin, which crosslinks the cytoskeleton and plasma membrane, is involved in the growth and metastatic potential of cancer cells. Ezrin expression in esophageal squamous cell carcinoma (ESCC) was described recently, but its roles and the underlying mechanism(s) remain unclear. In our study, we first showed that ezrin in ESCC cell is expressed in the nucleus as well as in the cytoplasm and plasma membrane. Then, by using RNAi, we revealed that interference of ezrin expression suppressed the growth, adhesion and invasiveness of ESCC cells. Tumorigenesis experiments revealed that ezrin may directly regulate tumor formation in vivo. To explore the molecular mechanisms through which ezrin contributes to the proliferation and invasiveness of ESCC cells, we used cDNA microarrays to analyze ezrin knockdown cells and the control cells; of 39,000 genes examined, 297 were differentially expressed upon ezrin knockdown, including some proliferation- and invasiveness-related genes such as ATF3, CTGF and CYR61. Furthermore, pathway analysis showed that ezrin knockdown led to decreased activation of the TGF-beta and MAPK pathways, and ezrin-mediated cell invasiveness alteration was dependent on the activation of these pathways. Finally, immunohistochemical staining on 80 ESCC specimens and 50 normal esophageal mucosae revealed that the expression levels of 3 altered genes involved in the regulation of cell proliferation and tumor metastasis, including CTGF, CYR61 and ATF3, were altered in ESCCs, and their expression pattern correlated with ezrin expression. Taken together, we propose that ezrin might function in the growth and invasiveness of ESCC cells through the MAPK and TGF-beta pathways.
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http://dx.doi.org/10.1002/ijc.24216DOI Listing
June 2009

NGALR is overexpressed and regulated by hypomethylation in esophageal squamous cell carcinoma.

Clin Cancer Res 2008 Dec;14(23):7674-81

Department of Biochemistry and Molecular Biology, The Key Immunopathology Laboratory of Guangdong Province, Medical College of Shantou University, Shantou, People's Republic of China.

Purpose: Neutrophil gelatinase-associated lipocalin receptor (NGALR) mRNA level is reduced in isolated chronic myelogenous leukemia blasts but up-regulated in esophageal squamous cell carcinoma (ESCC). The mechanism of NGALR regulation is unknown. Here, we show the expression pattern of NGALR and examine the aberrant methylation of its gene in ESCC and esophageal carcinoma cell lines.

Experimental Design: The expression pattern of NGALR was analyzed by immunohistochemistry in 59 ESCCs and compared with noncancerous tissues. The DNA methylation status was investigated by methylation-specific PCR and by bisulfite genomic sequencing in esophageal carcinoma cell lines and surgically resected samples. Methylated cell lines were treated with a methylation inhibitor to restore NGALR expression.

Results: The expression of NGALR in ESCC was significantly higher in tumor cell membrane and cytoplasm than in normal esophageal epithelium (P < 0.01). Methylated alleles were detected in three NGALR-nonexpressing cell lines but were not detected in three NGALR-expressing cell lines. Treatment of methylated cell lines with 5-aza-2'-deoxycytidine, a methylation inhibitor, restored NGALR expression. In surgically resected samples, 31 of 77 (40.3%) primary esophageal carcinomas and 46 of 77 (59.7%) paired normal tissues contained methylated NGALR alleles (P < 0.05).

Conclusions: Our results suggest that NGALR hypomethylation contributes to its expression in esophageal carcinomas and that this overexpression may play a role in the pathogenesis of esophageal carcinomas.
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http://dx.doi.org/10.1158/1078-0432.CCR-08-0420DOI Listing
December 2008

Comparative proteomic analysis reveals differentially expressed proteins regulated by a potential tumor promoter, BRE, in human esophageal carcinoma cells.

Biochem Cell Biol 2008 Aug;86(4):302-11

Department of Histology and Embryology, Shantou University Medical College, Shantou, China.

Esophageal tumorigenesis is a complex and cascading process, involving the interaction of many genes and proteins. In this study, we have used the comparative proteomic approach to identify tumor-associated proteins and explore the carcinogenic mechanisms. Two-dimensional electrophoresis (2-DE) and MALDI-TOF MS analysis of esophageal carcinoma and control cells revealed 10 proteins that were upregulated. A further 10 proteins were downregulated. Among these 20 differentially expressed proteins, brain and reproductive organ-expressed (BRE) protein was identified as a potential tumor promoter. It was high expressed by the esophageal carcinoma cells, as confirmed by RT-PCR and immunoblotting. BRE has been reported to be a stress-responsive protein. To gain further insight into its function, BRE expression was silenced in esophageal carcinoma cells using BRE-specific small interference RNA. It was discovered that silencing BRE expression downregulated prohibitin expression, but upregulated tumor-suppressor p53 expression. Furthermore, cyclin A and CDK2 expressions were suppressed suggesting that BRE inhibited cell proliferation. These results implied that BRE plays a significant role in mediating antiapoptotic and proliferative responses in esophageal carcinoma cells.
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http://dx.doi.org/10.1139/o08-069DOI Listing
August 2008

[Inhibitory effect of Oridonin injection on heterotransplanted gastric adenocarcinoma in nude mice and its mechanism].

Zhonghua Zhong Liu Za Zhi 2008 Feb;30(2):89-92

Department of Oncology, Peking University Shenzhen Hospital, Shenzhen 518036, China.

Objective: To investigate the inhibitory effect of Oridonin injection on heterotransplanted tumors of human gastric adenocarcinoma cell line BGC823 cells in nude mice and explore its mechanism.

Methods: Heterotransplanted models of human gastric adenocarcinoma cell line BGC823 cells in nude mice were established. They were divided at random into three groups as control group, low-dose group and high-dose group. The Oridonin solution at concentration of 37.5 mg x kg(-1 x d(-1) and 75 mg x kg(-1) x d(-1) were injected to the mice in low-dose group and high-dose group, respectively, and 0.9% sodium chloride was injected to the mice of control group per day for 10 days sequentially. The mice of the three groups were sacrificed at 11th day after the first injection of Oridonin. The tumor weight of the sacrificed mice was measured. Morphological and ultrastructural examinations of the tumors were carried out by light and electron microscopy. The expression of bcl-2, Bax, Fas and FasL was detected by immunohistochemistry.

Results: Oridonin injection showed a suppressive effect on the growth of heterotransplanted tumors in the nude mice. The tumor growth inhibition rates were 48.5% and 70.7% in the low-dose and high-dose groups, respectively. The morphological study demonstrated that tumor cells displayed a typical appearance of apoptosis. The expression of bcl-2 was down-regulated, while Bax, Fas and FasL were up-regulated.

Conclusion: Oridonin can markedly inhibit the growth of heterotransplanted human gastric adenocarcinoma in nude mice. It was due, at least in part, to the induction of apoptosis in cancer cells.
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February 2008

Autophagy and endocytosis in the amnion.

J Struct Biol 2008 May 24;162(2):197-204. Epub 2006 Oct 24.

Department of Pathology, Key Immunopathology Laboratory of Guangdong Province, Medical College of Shantou University, Shantou 515031, China.

To search for the origin of nutrition in the amnion, we focused attention on both endocytotic and autophagic pathways. Using ultrastructural and biochemical methods, we examined 20 human amnions at term gestation. The uptake of horseradish peroxidase (HRP) was used for the detection of endocytosis. Transfection of the LC3-GFP plasmid and staining with monodansylcadaverine (MDC) and LysoTracker red (LTR) were used to demonstrate the formation of autophagic vacuoles. In addition, two autophagic genes, beclin 1 and Atg5, were assayed by RT-PCR. Within the amniotic epithelial (AE) cells, autophagic vacuoles contained organelles and cytoplasmic components and were enclosed by a double membrane. They contained autophagosomes with transfected LC3-GFP that stained positive for MDC and autolysosomes that stained positive for LTR. Endocytosis was an extremely active process in the cellular uptake of fluid and fluid contents and led to formation of vesicles and endosomes, which were found to be positive by HRP test. Many uniform vesicles were collected in the multivesicular bodies (MVBs). Finally, both endosomes and autophagosomes were fused and degraded by lysosomes. The data also demonstrated that large autophagosomes engulfed some endosomes or MVBs. Transcription of beclin 1 and Atg5 occurred in the amnion at term gestation. Taken together, these results show that AE cells have active endocytotic and autophagic capacities and that lysosomes are involved in the intracellular degradation of endosomes and autophagosomes. Sometimes the autophagic and endocytotic pathways converge. This study suggests that of endocytosis and autophagy activities in AE cells can be induced by nutrient limitation and are probably also evoked in response to some hormones in the amniotic fluid. Activation of both endocytotic and autophagic pathways plays different roles in the ability of the cell to acquire nutrients needed for its survival.
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http://dx.doi.org/10.1016/j.jsb.2006.10.010DOI Listing
May 2008

Expression of fascin in thyroid neoplasms: a novel diagnostic marker.

J Cancer Res Clin Oncol 2008 Sep 15;134(9):947-51. Epub 2008 Mar 15.

Department of General Surgery, The First Teaching Hospital of Jilin University, Changchun, Jilin Province, People's Republic of China.

Purpose: Fascin, an actin-bundling protein, is markedly upregulated in several epithelial tumors and its expression often correlates with high-grade, extensive invasion, and distant metastasis. However, reports about fascin expression in endocrine tumors remain rare. The aim of the present study was to assess the diagnostic significance of fascin in thyroid neoplasms.

Methods: Thyroid samples from 177 cases were examined for fascin and Ki-67 expression by immunohistochemistry.

Results: Fascin immunoreactivity was negative in normal follicles and nodular goiter. Fascin immunostaining was positive in 62.1% (41/66) of thyroid carcinomas and 26.4% (19/72) of thyroid adenomas; the difference being significant (P < 0.0001). In thyroid papillary carcinoma, upregulation of fascin was associated with both the Ki-67 labeling index and the occurrence of lymph node metastasis.

Conclusion: Fascin may be a novel marker to distinguish thyroid carcinoma from benign lesions and may be involved in the proliferation and metastasis of papillary carcinoma.
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http://dx.doi.org/10.1007/s00432-008-0374-6DOI Listing
September 2008

Fascin expression in human embryonic, fetal, and normal adult tissue.

J Histochem Cytochem 2008 Feb 12;56(2):193-9. Epub 2007 Nov 12.

Department of Biochemistry and Molecular Biology, Medical College of Shantou University, 22 Xinling Road, Shantou City 515041, Guangdong Province, China.

This study investigates the distribution of fascin in human embryonic, fetal, and normal adult tissues. Tissue microarray technology was used to perform immunohistochemical experiments on human embryos and fetuses at 4-22 weeks of gestation and adult specimens. Fascin was widely expressed in the nervous system. At 4 weeks of gestation, fascin was present in the neural tube. At 8-12 weeks of gestation, homogenous gene expression was seen in cells of the cerebellum and gastrointestinal tract. In later developmental stages and in adults, Purkinje cells of the cerebellum and glandular epithelium of the gastrointestinal tract showed no expression. Fascin was expressed in the cortex and medulla of the adrenal gland at 8-12 weeks of gestation, whereas immunoreactivity decreased from the zona glomerulosa through the zona reticularis and was essentially negative in the adrenal medulla of adults. Significant expression of fascin was seen throughout development in neurons, follicular dendritic cells of lymphoid tissue, basal layer cells of stratified squamous epithelia, mesenchyme, and vascular endothelial cells. Simple columnar epithelia of the biliary duct, colon, ovary, pancreas, and stomach were all negative for fascin expression. These results show that expression of fascin is time specific and highly tissue specific. Parallels between fascin expression in embryogenesis and carcinogenesis are discussed.
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http://dx.doi.org/10.1369/jhc.7A7353.2007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2324166PMC
February 2008

Epigenetic silencing of a Ca(2+)-regulated Ras GTPase-activating protein RASAL defines a new mechanism of Ras activation in human cancers.

Proc Natl Acad Sci U S A 2007 Jul 18;104(30):12353-8. Epub 2007 Jul 18.

State Key Laboratory in Oncology in South China, Sir Y. K. Pao Center for Cancer, Department of Clinical Oncology, Hong Kong Cancer Institute and Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong, Hong Kong.

Ras has achieved notoriety as an oncogene aberrantly activated in multiple human tumors. Approximately 30% of all human tumors express an oncogenic form of this GTPase that is locked in an active conformation as a result of being insensitive to Ras GTPase-activating proteins (GAPs), proteins that normally regulate the inactivation of Ras by enhancing its intrinsic GTPase activity. Besides oncogenic mutations in Ras, signaling by wild-type Ras is also frequently deregulated in tumors through aberrant coupling to activated cell surface receptors. This indicates that alternative mechanisms of aberrant wild-type Ras activation may be involved in tumorigenesis. Here, we describe another mechanism through which aberrant Ras activation is achieved in human cancers. We have established that Ras GTPase-activating-like protein (RASAL), a Ca(2+)-regulated Ras GAP that decodes the frequency of Ca(2+) oscillations, is silenced through CpG methylation in multiple tumors. With the finding that ectopic expression of catalytically active RASAL leads to growth inhibition of these tumor cells by Ras inactivation, we have provided evidence that epigenetically silencing of this Ras GAP represents a mechanism of aberrant Ras activation in certain cancers. Our demonstration that RASAL constitutes a tumor suppressor gene has therefore further emphasized the importance of Ca(2+) in the regulation of Ras signaling and has established that deregulation of this pathway is an important step in Ras-mediated tumorigenesis.
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http://dx.doi.org/10.1073/pnas.0700153104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1941473PMC
July 2007

[Expression and significance of heat shock proteins in esophageal squamous cell carcinoma].

Zhonghua Zhong Liu Za Zhi 2006 Oct;28(10):758-61

Department of Oncology, First Affiliated Hospital of Shantou University Medical College, Guangdong Shantou 515041, China.

Objective: To investigate the expression and significance of HSP27, HSP60, HSP70 and HSP90 alpha in esophageal squamous cell carcinoma (ESCC) and tissues along the incision margin (TIM).

Methods: The presence and the level of expression of HSP27, HSP60, HSP70 and HSP90 alpha were determined in 168 specimens from ESCC and 42 from tissues along TIM by EnVision immunohistochemistry and Western blotting, to compare their positive staining rates and explore the correlation between their expressions and clinicopathologic features in ESCC.

Results: The positive staining rates of HSP27, HSP60, HSP70 and HSP90 alpha in ESCC and TIM were 62.0% and 42.1%, 92.7% and 63.2%, 57.9% and 22.2%, and 33.7% and 18.5%, respectively. There was very significant difference between the expression of HSP60 and HSP70 in ESCC and TIM (P < 0.01), but not significant about HSP27 and HSP90 alpha (P > 0.05). The positive staining rate of HSP27 declined with the lower grade of differentiation of ESCC (P < 0.05).

Conclusion: The present findings suggest that the expression of HSPs of different molecular weight in ESCC and TIM is a common event. The level of expressions of HSP60 and HSP70 are higher than those in TIM. HSP60 and HSP70 expression correlated with the biological behavior of ESCC. The expression of HSP27 was positively correlated to the grade of differentiation of ESCC. Overexpression of HSP27 may be associated to the differentiation of squamous cell carcinoma.
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October 2006

A novel alternative spliced variant of neutrophil gelatinase-associated lipocalin receptor in oesophageal carcinoma cells.

Biochem J 2007 Apr;403(2):297-303

Department of Biochemistry and Molecular Biology, Medical College of Shantou University, Shantou 515041, Guangdong Province, China.

Recent studies suggest that NGAL (neutrophil gelatinase-associated lipocalin) is a novel iron transporter with functions distinct from that of transferrin and mediates a new iron-delivery pathway. To get a better understanding of NGAL function in oesophageal carcinoma, we analysed the expression of NGAL receptors in oesophageal carcinoma cells and identified a novel spliced variant designated NgalR-3. When expressed in a heterologous system, the protein produced from this novel spliced variant exhibits the biochemical characteristics of interaction and co-localization with NGAL protein in vivo. This new finding suggests that NgalR-3 may act as a potential NGAL receptor and play a role in NGAL-mediated iron transport in oesophageal carcinoma.
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http://dx.doi.org/10.1042/BJ20060836DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1874233PMC
April 2007

[The promotive effects of N-nitrosopiperidine on the malignant transformation of the immortalized esophageal epithelium induced by human papillomavirus].

Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi 2006 Jun;20(2):81-3

Shantou University Medical College, Shantou 515031, China.

Background: Study on the promotive effects of N-nitrosopiperidine on carcinogenesis process was performed, based on the immortalization of human fetal esophageal epithelium induced by human papillomavirus (HPV) 18E6E7 genes.

Methods: The immortalized esophageal epithelium SHEE was induced by HPV18E6E7. The cells at 17th passages were cultured in 50 ml flasks. The N-nitrosopiperidine (NPIP) 0, 2, 4, 8 mmol/L added to the cultured medium of SHEE cells for 3 weeks. The morphology, proliferation and apoptosis of the cells were studied by phase contrast microscopy and flow cytometry. Modal number of chromosomes was analyzed by standard method. Tumorigenicity of the cells was assessed by soft agar colony formation and by transplantation of cells into nude mice. Expression of HPV was detected by Western blot.

Results: When cells were exposed to high concentration (8 mmol/L) of NPIP, cell death was increased, leaving a few live cells. In normal cultural medium instead of NPIP proliferative status of the cells restored after 4 weeks and the cells progressed to the proliferation stage with continuous replication and atypical hyperplasia. At the end of the 8th week, the cells appeared with large colonies in soft-agar and tumor formation in transplanted nude mice. When the cells were cultured in 2, 4 mmol/L NPIP the doubling passage was delayed and without tumor formation in transplanted nude mice. Modal number of chromosomes was 61-65, in 8 mmol/L NPIP group and control group, 56-61. Expression of HPV18 appeared in experimental and control groups.

Conclusion: NPIP promotes malignant change of the immortalized esophageal epithelial cells induced by HPV18E6E7. HPV18E6E7 synergy with NPIP will accelerate malignant transformation in esophageal epithelium.
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June 2006

[Differential expression of STRBP8, a new candidate oncogene, in cancerization of human immortalized esophageal epithelial cells].

Ai Zheng 2005 Apr;24(4):385-90

Department of Biochemistry and Molecular Biology, Guangdong Medical College, Zhanjiang, Guangdong, 524023, P. R. China.

Background & Objective: Recently, changes in composition, structure, and function of nuclear matrix proteins (NMPs) in generation and development of tumors evoked more and more attention. Separation and identification of tumor-related NMPs is a new way to search for tumor specific biomarkers, and to study tumor pathogenesis. This study was to analyze differential expression of STRBP8, one of esophageal carcinoma specific NMPs, in cancerization of immortalized human esophageal epithelial cells.

Methods: NMPs were extracted from immortalized human esophageal epithelial cell line SHEE and malignantly transformed esophageal carcinoma cell line SHEEC. Differential expression of STRBP8 was detected by two-dimensional electrophoresis (2-DE), matrix-assisted laser desorption/ionization time of flying mass spectrometry (MALDI-TOF-MS), and reverse transcription-polymerase chain reaction (RT-PCR). STRBP8 cDNA obtained by RT-PCR was linked to pGEM-T easy vector, and introduced into TOP10F' E.coli competent cells. Positive clones were sequenced and analyzed with BLAST.

Results: STRBP8 was only detected in SHEEC cells by 2-DE, MALDI-TOF-MS, and RT-PCR. The sequence of positive clones contained STRBP8 cDNA was identical to that in GenBank database.

Conclusion: STRBP8, as a candidate oncogene, might relate to cancerization of esophageal epithelial cells, which might be a specific biomarker of esophageal carcinoma, and probe into the pathogenesis of esophageal carcinoma.
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April 2005

[Determination of arsenic in As2O3-treated esophagoscope-transplant mouse tissue by graphite furnace atomic absorption spectrometry].

Guang Pu Xue Yu Guang Pu Fen Xi 2004 Apr;24(4):487-90

Central Laboratories, Shantou University Medical College, Shantou 515031, China.

As2O3 was injected into tumour tissue of esophagoscope-transplant mouse. The concentration of As diffusing into other tissues was investigated. A method was proposed for the determination of As in tissue samples of mouse by graphite furnace atomic absorption spectrometry. After wet digestion (water-bath at 80 degrees C) with 2:1:1 (psi) of HNO3-H2SO4-HClO4, the digested tissue samples of mouse were diluted with 0.2% Triton X-100-0.4% AgNO3. The matrix-matching calibration curve of non-interference was established with standard addition method. The relative standard deviation was 3.2%-8.7%. The limit of detection was 1.57 microg x L(-1). The recoveries were 81.7%-105% Arsenic concentrations in mouse liver, kidney, brain, left-chest, and right-chest were determined after injection of arsenic at different times.
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April 2004

[Studies on transformation of human esophageal fibroblasts mediated by a retroviral vector containing the E6E7 ORFs of human papillomavirus type 16].

Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi 2004 Sep;18(3):223-6

Department of Oncogenic Virus and HIV, National Institute for Viral Disease and Prevention, Chinese Center for Disease Control and Prevention, Beijing 100052, China.

Objective: To study the etiological role of human papillomavirus type 16 (HPV16) infection in the development of esophageal cancers.

Methods: A recombinant retrovirus containing the E6E7 ORFs of HPV16 was packaged and human fetal esophageal fibroblasts were infected. The tumorigenecity of the fibroblasts was tested in SCID mice in synergy with 12-O-tetradecanoylphorbol-13-acetate (TPA).

Results: Human esophageal fibroblasts infected with the recombinant retrovirus induced sarcomas in SCID mice, the existence and expression of E6E7 ORFs was confirmed in the sarcomas. Fibroblasts cultured from the sarcoma were demonstrated heteroploid by cytoflowmetry. However, tumors were not observed in human fetal esophagus infected with such virus.

Conclusions: These results revealed that the established recombinant retroviral system can successfully mediate the transference of HPV16 E6E7 genes, and such system is applicable to researches on tumorigenesis of HPV.
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September 2004

[Expression of fascin 1 gene in the process of the immortalized esophageal carcinoma carcinogenesis].

Ai Zheng 2004 Mar;23(3):243-8

Department of Biochemistry and Molecular Biology, Shantou University Medical College, Shantou, Guangdong, 515041 PR China.

Background & Objective: Fascin 1 is the 55kDa F-actin- binding cytoskeleton protein. Fascin 1 gene was cloned from a human teratocarcinoma. Up to now, the carcinogenesis mechanism of esophageal squamous cell carcinoma is unclear. The study was designed to identify the differentially expressed proteins and mRNAs between the human immortalized esophageal epithelial cell line (SHEE) transfected by human papillomavirus type 18 E6E7 and the malignant transformation cell line (SHEEmt), which is derivated from SHEE, and to further understand the carcinogenesis mechanisms of esophageal squamous cell carcinoma.

Methods: Cellular proteins were separated by two-dimensional electrophoresis and differentially expressed proteins were identified by matrix-assisted laser desorption/ionization time of flying mass spectrometry (MALDI-TOF-MS). The mRNA of fascin 1 gene was assayed by reverse transcription polymerase chain reaction (RT-PCR) and its product was analyzed by sequencing assay.

Results: The results manifested 9 genes expressed differently in the progress of malignant transformation of SHEE to SHEEmt, fascin 1 protein increased about 3.64 times and its mRNA increased about 16.17 times.

Conclusion: The upregulated expression of fascin 1 gene may be correlated with the malignant transformation of SHEE to SHEEmt.
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March 2004

The multistage process of carcinogenesis in human esophageal epithelial cells induced by human papillomavirus.

Oncol Rep 2004 Mar;11(3):647-54

Department of Pathology, Medicine College of Shantou University, Shantou 515031, PR China.

To investigate the multistage process of carcinogenesis, the progressive alteration of the morphology, telomerase, cytogenesis, oncogenes and tumorigenicity in the process of immortalization and malignant transformation of the human fetal esophageal epithelial cell (SHEE) was studied. The SHEE cells were immortalized by gene E6E7 of human papilloma virus (HPV) type 18 in our laboratory and continually cultivated over 100 passages, which had been malignantly transformed. Cells at the 11th, 35th, 65th and 100th passage were examined according to the following criteria: morphological changes of cell growth, contact-inhibition and anchorage-independent growth (AIG); the cell proliferative and apoptotic index; the modal number of chromosomes; c-myc, p53, bcl-2, ras; telomere length and activities of telomerase and tumorigenicity in nude mice or severe combined immunodeficient (SCID) mice. The cells of the 11th passage were well differentiated and the cells of 100th passage were relatively poorly differentiated with polymorphism, while the cells of 35th and 65th had two distinct differentiations. The proliferative indexes were 21.1%, 32.5%, 33.2%, and 40.9% and the apoptotic indexes were 3.3%, 2.7%, 3.5%, 2.7% in the 11th, 35th, 65th and 100th passage respectively. Karyotypes of four cell passages belonged to hyperdiploidy and hypotriploidy. C-myc, ras, p53 genes were low in the 10th and 35th, and high in the 65th and 100th passage, but bcl-2 was low in 4 passages. Telomere length sharply decreased from normal fetal esophagus cells until the 35th passage, but it was stably expressed in the 65th and 100th passage. The activities of telomerase were expressed in cells of the 35th, 65th and 100th passages. The efficiency of AIG varied in different passages of the SHEE cell and was absent in the 11th passage, low efficiency in the 35th passage and 65th passage, and high efficiency in the 100th passage. Transplanted cells of the 65th and 100th passage into SCID mice resulted in tumor formation, but only the 100th passage cells could grow in nude mice. All of these characteristic changes were in dynamic progressive process. These data demonstrate that carcinogenesis of esophageal epithelial cells induced by HPV is the multistage process, which goes through the initial, immortal, premalignant and malignant transformation stages. The generation of esophageal carcinoma is caused by the accumulation of cellular, genetic and molecular changes.
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March 2004