Suppression of Aurora-A by RNA interference inhibits laryngeal cancer Hep-2 cell growth / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To investigate the effects of knockdown of Aurora-A by RNA interference on laryngeal cancer Hep-2 cell growth in vitro and in vivo.</p><p><b>METHODS</b>A plasmid containing siRNA against Aurora-A was constructed and transfected into human laryngeal cancer cell line Hep-2. Measurements included the CCK-8 assay for viability and proliferation, Transwell assay for invasion, colony formation assay for cell anchorage-independent growth. Western blot and immunohistochemistry assay for protein expression. Tumorigenicity was observed in vivo.</p><p><b>RESULTS</b>In Hep-2 cells transfected by Aurora-A siRNA (designated as siRNA-3), protein expression of Aurora-A was suppressed by 52%. In CCK-8 assay, absorbance value of siRNA-3 cells (3.268 ± 0.106, (x(-) ± s)) was lower than that of Hep-2 cells (3.722 ± 0.152, F = 17.634, P < 0.001). In Transwell assay, the average invasive cells per field in siRNA-3 cells (110.0 ± 18.0) was less than that in Hep-2 cells (236.0 ± 26.0, F = 26.462, P < 0.01). In colony formation assay, the average colony number of siRNA-3 cells (31.0 ± 6.6) was lower than that of Hep-2 cells (104.0 ± 14.0). The average tumor size in siRNA-3 group was (127.77 ± 174.83) mm(3), which was less than Hep-2 cell group (837.26 ± 101.80) mm(3), (F = 28.187, P < 0.001). Silencing of Aurora-A decreased the expression of focal adhesion kinase (FAK) and matrix metalloproteinase-2 (MMP-2), key regulators in cell adhesion and invasion.</p><p><b>CONCLUSIONS</b>The knockdown of Aurora-A inhibits the growth and invasiveness of Hep-2 cells in vitro and in vivo, which may be a promising therapeutic strategy for LSCC.</p>

Expression of metastasis associated 1 family member 2 (MTA2) in gastric cancer and its relationship with transcription factor Sp1 / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To determine the expression patterns of metastasis associated 1 family member 2 (MTA2) in gastric cancer and non-cancerous gastric mucosa, and analyze its relationship with nuclear transcription factor Sp1 expression.</p><p><b>METHODS</b>Tissue samples and clinicopathological information from 83 gastric cancer patients, who underwent surgery, were collected from Shanghai Rui Jin Hospital. All samples included cancer tissue and non-cancerous mucosa which was 5 cm away from the tumor lesion. The expression of MTA2 and Sp1 were detected by immunohistochemistry (IHC) staining. The mRNA of MTA2 was also detected by reverse transcription-polymerase chain reaction (RT-PCR). SPSS software was used for statistical analysis.</p><p><b>RESULTS</b>The expression of MTA2 protein was significantly higher in primary lesions of the gastric cancer than that in non-cancerous mucosa by IHC (31.3% vs 12.0%, P < 0.01). MTA2 expression was closely related with tumor invasion or T staging (χ(2) = 5.677, P < 0.05). Yet, no significant relationship was observed between MTA2 expression and other clinicopathological parameters, including the age, sex, tumor differentiation, Lauren classification, lymph node metastasis, distant metastasis, as well as pathological staging. Furthermore, MTA2 expression was concomitant with Sp1 expression (r = 0.320, P < 0.05). Elevated MTA2 expression was observed in Sp1 positive cancer tissues (χ(2) = 9.565, P < 0.01). RT-PCR results also demonstrated that MTA2 mRNA was also highly expressed in the tissue samples with Sp1 expression.</p><p><b>CONCLUSIONS</b>MTA2 is highly expressed in the primary lesions of gastric cancer than that in adjacent non-cancerous tissues, and is closely related with tumor invasion. MTA2 expression is elevated in Sp1 positive gastric cancer.</p>

Interaction of SerpinB5 and MAFbx in gastric cancer cell and its action site / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To explore the interaction between SerpinB5 and MAFbx in gastric cancer cell and to identify the interaction sites.</p><p><b>METHODS</b>The interaction between SerpinB5 and MAFbx was screened and validated by yeast two-hybrid screening and co-immunoprecipitation. The expression of MAFbx was analyzed after SerpinB5 expression being modified by RNA interference and pGBKT7-SerpinB5 transfection. The impact of SerpinB5 on the expression of MAFbx was studied in gastric cancer cell line SUN-16. A model of MAFbx was constructed by homology modeling. The related residues for interaction were analyzed by Autodock4.0.</p><p><b>RESULTS</b>The interaction between SerpinB5 and MAFbx was validated. The expression of MAFbx changed along with SerpinB5 expression. Amino acids including PRO261, ASN361, and LYS362 were key residue in the interaction of SerpinB5 and MAFbx.</p><p><b>CONCLUSION</b>SerpinB5 interacts with MAFbx in gastric cancer cell. Amino acids including PRO261, ASN361, and LYS362 are potential binding sites.</p>

Impacts of Bevacizumab on vascular endothelial growth factor and Sp1 expression in gastric cancer xenografts / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To evaluate the effects of Bevacizumab on the tumor growth, proliferation and apoptosis of gastric cancer xenograft, and the impacts on the VEGF and Sp1 expression.</p><p><b>METHODS</b>Gastric cancer xenografts in nude mice were established using SGC-7901 gastric cancer cell line. The nude mice were randomly divided into two groups, Bevacizumab treatment group and PBS group. The tumor sizes were measured for tumor growth curve. The proliferation and angiogenesis were evaluated by immunohistochemistry (IHC) staining of Ki67 and CD34. TUNEL assay was used for apoptosis evaluation. The expression of VEGF and Sp1 in tumor cells were detected by IHC and Western blot.</p><p><b>RESULTS</b>Compared to the PBS group, the tumor growth decreased significantly (P<0.05), the proliferation of tumor cells and angiogenesis decreased, and apoptosis index increased significantly [(5.3 ± 1.8)% vs. (16.7 ± 6.7)%, P<0.01] in Bevacizumab group. The results of IHC and Western blot demonstrated that the expression of VEGF and the microvessel density (MVD) was decreased (4.0 ± 1.0 vs. 16.3 ± 1.5, P<0.001) in Bevacizumab treatment group. No obvious changes of Sp1 expression were observed in Bevacizumab treatment group.</p><p><b>CONCLUSIONS</b>Bevacizumab can inhibit the growth of gastric cancer xenografts in nude mice, decrease the VEGF expression and MVD. However, the compensatory up-regulation of transcription factor Sp1 is not affected by Bevacizumab.</p>

Screening and identification of genes associated with multi-drug resistance in colonic cancer / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To identify novel multi-drug resistance-related genes, and to explore the mechanisms of multi-drug resistance.</p><p><b>METHODS</b>Multi-drug resistant cell line Lovo/5-FU was established by incubation with increasing dose of 5-FU. The sensitivity to 5-FU and cis-diaminodichloroplatinum (CDDP) was measured by MTT assay. Two dimensional electrophoresis plus mass spectrum(2-DE/MS) was used to identify the differentially expressed protein between Lovo and Lovo/5-FU. The identified protein was then verified by Western blot analysis.</p><p><b>RESULTS</b>The IC50 concentrations of Lovo/5-FU to 5-FU and CDDP were increased by 31 and 3 times, compared with Lovo (both P<0.01). 2DE-MS showed that CAP-G and RhoGDI2 were up-regulated, whereas 6-PGL, DCI, Prdx-6 and Maspin were down-regulated in Lovo/5-FU. Western blot analysis confirmed that the expression levels of RhoGDI2 and CAP-G in Lovo/5-FU were increased by 6.14 and 2.98 fold respectively (both P<0.01), whereas Maspin was decreased to 5.2% of Lovo(P<0.01).</p><p><b>CONCLUSIONS</b>Multi-gene and multi-pathway are involved in the development of multi-drug resistance of colorectal cancer cells. CAP-G, RhoGDI2 and Maspin are potential multi-drug resistant genes.</p>

Discovery and verification of matrix gla protein, a TNM staging and prognosis-related biomarker for gastric cancer / 中华病理学杂志

<p><b>OBJECTIVE</b>To analyze microarray datasets deposited in the public database and to identify TNM associated genes in gastric cancers.</p><p><b>METHODS</b>Microarray datasets of gastric cancer were selected from GEO database. Differentially expressed genes related to TNM staging were evaluated by significant analysis of the microarray using MultiExperiment Viewer (MEV) platform. Candidate gene expressions in gastric cancer tissues and cell lines were verified by reverse transcriptase polymerase chain reaction (RT-PCR), quantitative RT-PCR, Western blot and immunohistochemistry.</p><p><b>RESULTS</b>GES4007 dataset was re-analyzed leading to the identification of 14 genes associated with TNM staging. Over-expression of matrix gla protein (MGP) was confirmed in gastric cancer cell lines and tumor tissues by quantitative RT-PCR, Western blot and immunohistochemistry. Increased MGP expression was found in 22 of 54 cases of (40.7%) gastric cancer specimens compared to the normal gastric tissues. The up-regulation of MGP mRNA expression closely correlated with TNM stage (P = 0.001) and prognosis (P = 0.006).</p><p><b>CONCLUSIONS</b>Public databases of microarray studies are the valuable resources for data mining. MGP has been identified and confirmed as a novel biomarker for the TNM stage and prognosis of gastric cancer.</p>

Silencing invariant chains of dendritic cells enhances anti-tumor immunity using small-interfering RNA / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Genetic modification of dendritic cells (DCs) has been used as an effective approach to enhance anti-tumor immunity. RNA interference (RNAi), which can cause the degradation of any RNA in a sequence-specific manner, is a post-transcriptional gene silencing mechanism. In this study, small-interfering RNA (siRNA) specific for the Ii gene was transfected into DCs, and the anti-tumor immunity of Ii-silenced DCs was assessed.</p><p><b>METHODS</b>The silencing effect of siRNA was evaluated by Western blotting and real-time PCR analyses. In vitro cytotoxic activity of T cells was evaluated using a Cytotox 96(®) non-radioactive cytotoxicity assay kit. The time to tumor onset and the tumor volumes were used as reliable indices to assess the anti-tumor immunity in vivo. To further examine the mechanisms underlying the anti-tumor immunity, flow cytometry analysis was used.</p><p><b>RESULTS</b>The Ii expression of DCs was significantly reduced after Ii siRNA transfection. Significant in vitro anti-tumor ability was exhibited when DCs were co-transfected with Ii siRNA plus endogenous tumor antigen (P < 0.05). Furthermore, tumor growth was greatly inhibited when mice were immunized with DCs transfected with Ii siRNA plus tumor antigen prior to or subsequent to tumor implantation. Flow cytometry analysis in vitro and in vivo indicated that both CD4(+) and CD8(+) T cells were significantly activated in the Ii siRNA group (P < 0.05).</p><p><b>CONCLUSION</b>Silencing of the Ii gene of DCs may offer a potential approach to enhance DC-based anti-tumor immunity.</p>

Value of multidetector-row CT in the preoperative prediction of peritoneal metastasis from gastric cancer: a single-center and large-scale study / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the value of multidetector-row computed tomography (MDCT) in preoperatively predicting peritoneal metastasis of gastric cancer and to evaluate the indication for laparoscopic staging of gastric cancer on the basis of MDCT features.</p><p><b>METHODS</b>Six hundred and forty gastric cancer patients underwent preoperative MDCT examination, and the results of MDCT were compared with surgical and pathological findings. In addition, the relationship between MDCT features (depth of invasion, lymph node metastasis status, tumor size, and thickness of tumor) and peritoneal metastasis of gastric cancer was analyzed.</p><p><b>RESULTS</b>The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of MDCT in predicting peritoneal metastasis of gastric cancer were 51.0% (25/49), 99.3% (587/591), 86.2% (25/29), 96.1% (587/611), and 95.6% (612/640), respectively. Univariable analysis showed that all the four MDCT features (depth of invasion, lymph node metastasis status, tumor size, and tumor thickness) of gastric cancer were significantly correlated with the peritoneal metastasis of gastric cancer. None of the patients diagnosed with stage T(0~2)N(x)M(0) or T(x)N(0)M(0) gastric cancer by MDCT were found to have peritoneal metastasis. Receiver operating characteristic (ROC) analysis showed that the accuracy of the tumor size and thickness of gastric cancer in determining peritoneal metastasis was high(area under ROC curve was 0.83 and 0.75, respectively). Multivariable analysis showed that only tumor size was significantly correlated with the peritoneal metastasis from gastric cancer.</p><p><b>CONCLUSIONS</b>The clinical value of MDCT in preoperative prediction of peritoneal metastasis from gastric cancer is favorable. Laparoscopy can be avoided in patients with small tumor size or stage T(0~2)N(x)M(0) or T(x)N(0)M(0) gastric cancer diagnosed by MDCT due to lower incidence of peritoneal metastasis.</p>

Demethylation of the gamma-synuclein gene CpG island in colorectal cancer and its clinical significance / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To explore the relationship between gamma-synuclein gene expression and CpG island demethylation in colorectal cancer(CRC), and the relationship between the demethylation and clinicopathological factors of CRC.</p><p><b>METHODS</b>The expression of gamma-synuclein mRNA was examined in 30 pairs of tumor tissues and tumor-matched non-neoplastic adjacent tissues(NNAT) by RT-PCR. CRC cell lines including COLO205, LoVo, and SW480 were used and treated with a demethylating agent, 5-aza-2'-deoxycytidine(5-aza-C). Before and after the treatment, the expression of gamma-synuclein mRNA in the cells was determined by RT-PCR, and bisulfite sequencing PCR was also used to analyze methylation status of CpG island. The methylation status of gamma-synuclein was then examined in 67 CRC samples and 30 NNAT samples by nested methylation-specific PCR (NMSP) and real time methylation-specific PCR(real-time MSP). The relationship between the demethylation of gamma-synuclein in CRC and clinicopathological factors was analyzed.</p><p><b>RESULTS</b>The mean gamma-synuclein mRNA expression was 0.66+/-0.34 in CRC samples, which was much higher than 0.45+/-0.26 in NNAT samples(P=0.011). 5-aza-C could induce expression and demethylation of gamma-synuclein in COLO205, LoVo and SW480 cells. gamma-Synuclein gene was demethylated in 80.0%(24/30) of the CRC samples and 50.0%(15/30) of the NNAT samples. The demethylated status of gamma-synuclein was much higher in CRC samples than that in NNAT samples(P=0.030), and was significantly correlated with clinical stage, lymph node involvement, and distant metastasis of CRC(P<0.05).</p><p><b>CONCLUSION</b>The upregulation of gamma-synuclein expression in CRC is primarily attributed to the demethylation of CpG island, which may be used as a marker for prognosis.</p>

Promoter cloning and activity analysis for iroquois homeobox gene 1 in gastric mucosa cell line / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To clone core promoter regions of iroquois homeobox gene 1 (IRX1) gene and evaluate the regulatory mechanism of IRX1 transcription.</p><p><b>METHODS</b>Upstream sequence from transcriptional start site was predicted using bioinformatics methods. Serial deleted fragments from IRX1 promoter sequences were amplified by PCR and luciferase reporter plasmids were constructed. The luciferase intensity was analyzed after transferring reporters into GES-1 gastric mucosa cell line.</p><p><b>RESULTS</b>The promoter of IRX1 was predicted to be within 700 bp from the 5'-flanking region of IRX1 gene. Eight serial deleted luciferase reporter plasmids were constructed. The transcriptional activity of different fragments was expressed as following: p-416>p-584>p-715>p-350>p-687>p-320>p-188>p-92. Except p-320 and p-188, the transcriptional activity of other 6 fragments was higher than that of PGL3-basic plasmid. The transcriptional activity was the highest in p-416 and decreased sharply from p-320 to p-188.</p><p><b>CONCLUSIONS</b>The fragment p-416 shows the strongest promoter activity. The sequence from -320 bp to -188 bp is identified as core promoter region, which is focused as key sequence for further regulatory analysis, since some binding sites for important transcriptional factors such as Sp1 and TFII D are predicted.</p>

Analysis of human gastric cancer by transcriptome and proteome profiling / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To screen differential expression genes and proteins at transcriptome and proteome levels between human gastric cancer tissue and corresponding normal mucosa.</p><p><b>METHODS</b>Fresh-frozen gastric cancers were collected from patients treated at Ruijin Hospital. A total of 22 pairs of gastric cancer tissues and the corresponding noncancerous mucosa were analyzed. Commercially available cDNA microarray with 14 592 genes/ESTs was used. Genes were considered to be up-or down-regulated when the intensity ratio Cy3/Cy5 was > or = 2 or < or = 0.5 in over 50% samples (P<0.05). Immobilized pH gradient(IPG)-based 2-DE was applied to separate the total proteins of gastric cancer tissue and paired normal tissue. After staining and analysis by software,the differential expression proteins were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS) or MALDI-TOF-TOF-MS.</p><p><b>RESULTS</b>As compared with corresponding noncancerous tissue, there were totally 149 up-regulating genes/ESTs and 238 down-regulating genes/ESTs in gastric cancer, including 29 genes with 3-fold over-expression ratio and 21 genes with 5-fold under-expression. Fifteen protein spots were identified successfully, among whom there were ten over-expressed and five under-expressed proteins in gastric cancer tissue compared with normal tissue. Most of over-expressed genes and proteins were related to cell motility, cell proliferation, signal transduction, while those under-expressed genes and proteins were related to defense response, toxoid metabolism.</p><p><b>CONCLUSION</b>Studying gastric cancer at transcriptome and proteome levels can help demonstrate tumorigenesis and biological characteristics of gastric cancer comprehensively and provide powerful tools to find new biomarkers associated with gastric cancer and therapy targets.</p>

EGFR-blockade by antibody Cetuximab inhibits the growth of human gastric cancer xenograft in nude mice and its possible mechanism / 中华肿瘤杂志

<p><b>OBJECTIVE</b>EGFR-mediated tumor proliferation plays an important role in the development of cancer, and is a key candidate for targeted therapy. The aim of this study is to evaluate the impact of EGFR monoclonal antibody Cetuximab (C225) on the growth, proliferation and apoptsis of gastric cancer xenograft in nude mice, and its possible mechanisms.</p><p><b>METHODS</b>A gastric cancer cell line SGC-7901 with high EGFR expression level was screened from 7 gastric cancer cell lines. Gastric cancer xenografts in nude mice were established, and randomly divided into C225 treatment group and PBS control group. Tumor growth curves were calculated, the impact of C225 on the tumor growth, proliferation and angiogenesis was evaluated by immunohistochemical (IHC) staining Ki67 and CD34, respectively. The effect of C225 on apoptosis in the gastric cancer cells was evaluated by TUNEL assay. The expression levels of EGFR and its transcription factor Sp1 were detected by IHC staining and Western blot.</p><p><b>RESULTS</b>After C225 treatment, the proliferation and growth of gastric cancer xenograft in nude mice were significantly decreased. In the contrast, the apopotic indexes in C225 treatment group and PBS control group were (16.4% +/- 0.3%) and (3.1% +/- 0.9%), respectively, with a significant difference (P < 0.001). There was no significant difference of the densities of CD34-positive microvessels between C225 treatment group and control group. Elevated expression of EGFR and Sp1 after C225 treatment was observed by IHC staining and Western blot assay.</p><p><b>CONCLUSION</b>EGFR monoclonal antibody cetuximab (C225) can effectively inhibit the growth of gastric cancer xenografts in nude mice, and trigger its apoptosis. Yet, C225 treatment may upregulate the expression of EGFR and its transcription factor Sp1. A "block-transcription activation-compensation" mechanism may exist to explain the molecular mechanism of acquired resistance of a single target blockade treatment.</p>

Galectin-9 isoforms influence the adhesion between colon carcinoma LoVo cells and human umbilical vein endothelial cells in vitro by regulating the expression of E-selectin in LoVo cells / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the regulatory effect of galectin-9 isoforms on some molecules involved in cell adhesion/invasion, and the influence of this regulation action on the adhesion between colon carcinoma LoVo cells and human umbilical vein endothelial cells (HUVECs) in vitro.</p><p><b>METHODS</b>Various expression vectors of galectin-9 isoforms were transfected into LoVo cells. 24 h after transfection, the expression of integrin-beta1, E-cadherin, E-selectin, ICAM-1, CD44 and MMP-9 was detected by RT-PCR and Western blot analysis. LoVo cell-HUVEC adhesion assay was performed under conditions of galectin-9 transfection, galectin-9 transfection + galectin-9 antibody, galectin-9 transfection + E-selectin antibody and galectin-9 transfection + beta-lactose, respectively.</p><p><b>RESULTS</b>Galectin-9L down-regulates the mRNA and protein levels of E-selectin while galectin-9M and galectin-9S up-regulate the expression of E-selectin. In LoVo cell-HUVEC adhesion assay, the average fluorescence intensity of vector transfection group, galectin-9L transfection group, galectin-9M transfection group and galectin-9S transfection group was 0.90 +/- 0.20, 0.94 +/- 0.24, 1.60 +/- 0.11 and 1.45 +/- 0.13, respectively, indicating that galectin-9M and galectin-9S facilitated the adherence of LoVo cells to HUVECs (P < 0.05). E-selectin antibody, galectin-9 antibody or beta-lactose inhibited that effect.</p><p><b>CONCLUSION</b>Galectin-9 isoforms regulate the E-selectin expression in LoVo cells differently and thus influence the adhesion between LoVo cells and HUVECs in vitro in different modes. The mechanisms through which galectin-9 isoforms participate in the metastasis process of colon cancer may not be the same.</p>

Hedgehog signaling pathway activates in gastric carcinoma and promotes the proliferation through GLI1 in MKN28 cell / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the effect of Hedgehog (HH) pathway on proliferation and in vitro tumorigenicity of gastric cancer cell lines.</p><p><b>METHODS</b>The expression of SHH, PTCH, SMO, SUFU and GLI1 in seven cell lines were tested by RT-PCR. siRNA targeting GLI1 mRNA was transfected into MKN28 cells. Cell proliferation and in vitro tumorigenicity were examined by CCK8 and soft agar colony formation test.</p><p><b>RESULTS</b>SHH in six gastric cancer cell lines was up-regulated. Expression of PTCH in KATOIII cell lines and expression of SUFU in MKN28 and KATOIII were reduced. GLI1 siRNA significantly inhibited the expression of GLI1 in MKN28 cell line. Growth rate and colony formation rate of MKN28 cells treated with GLI1 siRNA were significantly lower than those of control cells (all P <0.001).</p><p><b>CONCLUSIONS</b>HH signaling pathway is widely activated in gastric cancer cell lines. The activation of HH signaling pathway promotes the growth of MKN28 cells.</p>

Expression and intracellular localization of FRZB gene in gastric cancer and its significance / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To study the expression and intracellular localization of FRZB gene in gastric cancer tissue, and to explore its significance in gastric cancer.</p><p><b>METHODS</b>The expression of FRZB in tumor tissues from 90 patients with gastric cancer and in normal gastric mucous as control were analyzed by immunohistochemistry in tissue array. FRZB expression in gastric cancer cell lines and immortalized gastric epithelial cell line GES-1 were detected by quantitative real-time PCR(Q-PCR) and Western blot. The intracellular localization of FRZB was observed by immunofluorescence staining.</p><p><b>RESULTS</b>The positive expression rate of FRZB in gastric cancer was 92.2%. FRZB expressed in gastric cancer with well differentiation was higher than that with poor differentiation.The positive rate in normal gastric mucous was 10.0% (one out of ten). By confocal microscope, FRZB localized both in cytoplasma and nucleus, especially on the nuclear membrane. The Q-PCR and Western blot results also showed that the expression of FRZB in gastric cancer cell lines was higher than that in GES-1.</p><p><b>CONCLUSIONS</b>The expression of FRZB in gastric cancer is correlated with tumor cell differentiation and tumor Lauren classification. The nuclear localization of FRZB may contribute to its function in gastric cancer formation and progression.</p>

Vascular endothelial growth factor D promoting the growth of lymphatic vessel in gastric carcinoma / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the relationship between vascular endothelial growth factor D (VEGF-D) and metastasis of lymphatic vessel in gastric carcinoma.</p><p><b>METHODS</b>The human VEGF-D cDNA was amplified from total RNA isolated from human normal gastric tissue, then it was inserted into T-A clone plasmid and subcloned into pEGFP eukaryotic expression vector. After the full-length sequence expected was confirmed by enzymatic digestion and sequencing,the human gastric carcinoma cell line SGC7901, which expressed a low level of VEGF-D, was transfected with the pEGFP/VEGF-D expression vector. Stable SGC7901 clones with high expression of VEGF-D were selected in vitro with G418, which were then combined and subcutaneously injected into nude mice to observe the density and morphology of lymphatic vessel. The outcomes were later compared with those of SGC7901 cells transfected with null vector(pEGFP) by immunostaining with a specific antibody LYVE-1.</p><p><b>RESULTS</b>The average weight of tumors in the pEGFP group (1.13+/-0.40) g at day 35, was significantly lower than that in the pEGFP/VEGF-D group (2.24+/-0.82)g (P<0.05). The lymphatic vessel density (LVD) in the pEGFP group (2.89+/-1.32) was significantly lower than that in the pEGFP/VEGF-D group (5.74+/-1.30)(P<0.01). There were dilated functional lymphatic vessels around the tumor margin.</p><p><b>CONCLUSION</b>VEGF-D may promote the growth and metastasis of tumor in gastric carcinoma by increasing the growth of lymphatic vessels.</p>

Cloning and functional analysis of the tumor metastasis related gene tropomyosin2 / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To construct an eukaryotic expression vector of tumor associated gene tropomyosin 2 (TM2) tagged with green fluorescence protein and study its biological role in tumorigenesis. METHODS; Whole length of TM2 cDNA sequence was amplified from human gastric cancer cell line AGS by reverse transcript PCR. The recombinant TM2 was inserted into eukaryotic expression vector pIRES2 tagged with enhanced green fluorescence protein. pIRES2-EGFP was transfected into human hepatoma cell line QGY-7703 cells by liposome technique and detected by fluorescence microscopy and flow cytometry. TM2 protein level was determined by Western blot. The in vitro invasion and motility were further studied. Its tumorigenesis was assessed on nude mice.</p><p><b>RESULTS</b>The recombinant TM2 was shown to be expressed in QGY-7703 cells by fluorescence microscopy and flow cytometry. The protein level of TM2 was increased. The invasion ability of TM2- transfected cells was increased(45.6 +/- 9.9) than that in controls(21.6 +/- 3.3, P < 0.05), as well as mobility(41.4 +/- 11.8 vs. 16.7 +/- 3.7). The tumor volume was (241.5 +/- 95.1) mm3, significantly higher than that in blank-vector controls (100.6 +/- 85.4) mm3 and negative-controls (123.7 +/- 92.3) mm3.</p><p><b>CONCLUSION</b>TM2 plays a role in growth and metastasis of hepatic tumor.</p>

Inhibition of the proliferation of human gastric cancer cells SGC-7901 in vitro and in vivo using Bcl-2 siRNA / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Bcl-2, the anti-apoptotic protein is overexpressed in the majority of gastric cancers and associated with its pathogenesis. To better understanding of the role of Bcl-2, RNA interference (RNAi) was used to inhibit Bcl-2 expression in the human gastric cancer cells in vitro and in vivo.</p><p><b>METHODS</b>Bcl-2 small interfering RNA (siRNA) was transfected into human gastric cancer cells SGC-7901, and Bcl-2 expression was monitored by real-time polymerase chain reaction (PCR) and Western blot. Cell proliferation, apoptosis, and telomerase activity were examined by MTT, flow cytometry, and TRAP assay, respectively. Gastric cancer cells treated with 100 nmol/L Bcl-2 siRNA were subcutaneously transplanted into nude mice and tumor growth was assessed.</p><p><b>RESULTS</b>Bcl-2 siRNA significantly inhibited the expression of Bcl-2 in human gastric cancer cells at both mRNA and protein levels in a time- and dose-dependent manner. Bcl-2 siRNA also decreased telomerase activity (by 78.76%) and increased the rate of apoptosis (by 37.47%). SGC-7901 cell growth was also significantly suppressed in vivo and in vitro.</p><p><b>CONCLUSIONS</b>Bcl-2 expression knockdown suppressed the growth of gastric cancer cells. Thus, Bcl-2 may play a very important role in carcinogenesis of gastric cancer and its knockdown may offer a new potential gene therapy approach for human gastric cancer in future.</p>

Chromosomal alterations analyzed by comparative genomic hybridization in primary gastric carcinoma / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To identify genetic abnormalities in primary gastric carcinoma.</p><p><b>METHODS</b>Comparative genomic hybridization (CGH) was used in screening DNA copy number changes along all chromosomes in 23 cases of primary gastric cancer.</p><p><b>RESULTS</b>Twenty-one out of 23 cases showed chromosomal losses and gains for at least one of the chromosomal arms in primary gastric cancer. The mean number of chromosomal alterations was 7.52. Chromosomal gains predominated over chromosomal losses in a ratio of 5.38:2.14. The most often involved chromosomal gains were observed in 8q (9/21, 42.9%), 20q (9/21, 42.9%), 17q (8/21, 38.1%), 3q (7/21, 33.3%), 7q (7/21, 33.3%), 11q (6/21, 28.6%), 13q (6/21, 28.6%), 1q (5/21, 23.8%) and 20p (5/21, 23.8%). The chromosomal arms with frequent losses were 17p (7/21, 33.3%), 18q (6/21, 28.6%), 5q (5/21, 23.8%), 8p (5/21, 23.8%), and 9p (5/21, 23.8%).</p><p><b>CONCLUSIONS</b>The phenomenon of gain and loss of chromosomal regions is observed in primary gastric cancer, which may induce the amplification of oncogenes and the loss of tumor suppressor genes to regulate the development and progression of gastric cancer.</p>

Expression and single nucleotide polymorphisms of kallikrein 10 in colorectal cancer / 中华外科杂志

<p><b>OBJECTIVE</b>To demonstrate expression and single nucleotide polymorphisms (SNP) of human kallikrein 10 (KLK 10) in colorectal cancer (CRC) and to correlate the KLK 10 expression level with clinicopathological factors of CRC.</p><p><b>METHODS</b>KLK 10 expression in 63 cases of tumoral and nontumoral colorectal tissues at the mRNA and protein levels were evaluated by quantitative real-time RT-PCR (qRT) and Western blot methods. KLK 10 protein was localized by immunohistochemistry. The KLK 10 genomic DNA from 16 cases of paired normal and cancerous colorectal tissues was PCR-amplified and examined for SNP by direct sequencing.</p><p><b>RESULTS</b>The KLK 10 mRNA expression was detected by qRT in 61 of 63 (97%) CRC specimens. The KLK 10 expression was much higher in tumor tissue than in the corresponding normal mucosal tissue at the mRNA and protein levels. The KLK 10 mRNA expression level significantly correlated with the lymphatic invasion (P < 0.05) and clinical stage of CRC (P < 0.05). No mutations or polymorphisms were detected in exon 1, 2 and 5 of KLK 10 gene in CRC. A SNP in codon 50 of exon 3, GCC (alanine) to TCC (serine) was identified. The genetic changes of exon 4 were located at codon 106 [GGC (glycine) to GGA (glycine)], codon 112 [ACG (threonine) to ACC (threonine)], codon 141 [CTA (leucine) to CTG (leucine)], and codon 149 [CCG (proline) to CTG (leucine)]. All these SNP were identical in tumor as well as the corresponding normal tissue DNA from the same individuals.</p><p><b>CONCLUSIONS</b>The KLK 10 expression is up-regulated in CRC and higher expression of KLK 10 closely correlate with advanced disease stage, which predicts a poorer prognosis, however, further follow-up study is needed.</p>