Objective To investigate the role of vascular endothelial growth factor-C (VEGF-C) and its receptors in the formation of lymphatic vessels and lymphatic metastasis in gastric cancer. Methods By the domestic and overseas literatures review,the expressions of VEGF-C and its receptors in gastric cancer,their role in tumor lymphatic metastasis and prospect in treatment of gastric cancer were summarized. Results There was a significant correlation between VEGF-C and its receptors and the formation of lymphatic vessels and lymphatic metastasis in gastric cancer. VEGF-C high expression might be an early event in lymphatic metastasis and could be considered as an independent predictive factor of lymphaticmicrometastasis. By inhibition of gastric cancer cell from secrete VEGF-C or blockage of the interaction of VEGF-C with VEGFR-3,it was possible to inhibit tumor angiogenesis and the invasion and distant spread of cancer cells,thereby decreased mortality and improve survival. Conclusion VEGF-C and its receptors may promote the formation of lymphatic vessels and lymphatic metastasis in gastric cancer. It may be an effective way to gastric cancer for the treatments against VEGF-C and its receptors.

Objective The aim of the current study was to investigate the prognostic value of extracapsular lymph node spread in gastric cancer patients and to find correlations with clinicopathological parameters.Methods Clinicopathological data of 131 gastric cancer patients who underwent gastrectomy with lymphadenectomy were analyzed retrospectively. The number of metastatic lymph nodes with extracapsular spread were determined. Multivariate analysis was performed to find the clinical prognosis affecting extracapsular lymph node involvement. Results Seventy-eight patients (59.5%)had perigastric lymph node metastasis. Fortysix cases were detected extracapsular lymph node involvement. The 5-year cumulative survival rate for patients with extracapsular lymph node spread was 13. 5% , while 32 patients with lymph node metastasis but without extracapsular involvement had a 5-year survival rate of 39.3%. The survival rate decreased significantly with the increase of extracapsular lymph node involvement(P =0.001). Extracapsular lymph node involvement was significantly associated with the higher number of metastatic lymph nodes, the location of lymph node metastasis, tumor invasion depth and distant lymph node metastasis. In the multivariate analysis, extracapsular lymph node spread also remained as an independent prognostic factor(P =0.003). Conclusions Extracapsular lymph node involvement is a convenient and reliable prognostic index, and is an independent prognostic factor in gastric cancer patients. In future staging systems for gastric cancer, extracapsular lymph node involvement should be considered, be pathologically checked and reported in order to determine extracapsular spread status.

Objective To investigate the dynamic variation of flavonoids contents in the flowers of Citrus grandis at different flowering periods. Methods The content of total flavonoids in the flowers of Citrus grandis was determined by means of ultraviolet-visible absorption spectroscopy ,and the contents of naringin and rhoifolin were determined by HPLC. Results The content of total flavonoids in the flowers during the flower withering period ,the young fruit period ,the blossom period,and the budding period was 306.90 mg.g-1,277.93 mg.g-1,215.55 mg.g-1 and 162.74 mg.g-1 respectively. The naringin content during the above four different periods was 246.31 mg.g-1,213.93 mg.g-1,175.94 mg.g-1 and 130.90 mg.g-1 respectively. The rhoifolin content during the four periods was unchanged. Conclusion The contents of total flavonoids and naringin in flowers of Citrus grandis during flower withering period are the highest.

Objective To study the resistances of CDl33 + subset purified from gastric cancer cell line to chemotherapy drugs and the mechanism of this resistance regarding to the mRNA expressions of both Bcl-2 and BAX in relation to the relative apoptotic genes.Methods CD133 + subset and CD133-subset were purified from KATOⅢ cell linc by magnetic activated cell sorting.The proliferating ability of these two subsets resistantnt to 5-FU,Cisplatin(DDP,PDD) and Etoposide was checked and compared by CCK-8 test.The apoptotic changes of these two subsets regarding to the expression of mRNA of both Bcl-2 and BAX were also analized by RT-PCR.Results In CD133 + subset,the contant percentage of CD133 + expression rate was 90％ via analysis of flow cytometye.Twelve hours after treatment of5-FU,DDP and VP-16,the cells in both CD133 + subgroup and CD133-subgroup would gradually start to change in apoptotic morphology.The growth inhibiting rate by CCK-8 measurement for 5-FU,DDP and VP-16 groups in CD133 + subgroup was significantly lower than that in CD133-subgroup.The data under different treatment respectively was,5-FU:(30.56 ± 1.99) ％-(88.60 ± 1.95) ％ vs (32.81 ± 2.67) ％-(95.73±2.12)％,P=0.045,cisplatin:(45.89 ±3.64)％-(81.20 ± 1.18)％ vs (50.21 ±3.22)％-(90.46±1.89)％,P=0.043,VP-16:(37.21 ±3.80)％-(78.49 ±3.22)％ vs (35.55 ±3.23)％-(89.32 ±-3.54) ％,P =0.048).After treatment of these three kind of anti-tumour drugs,the expression level of Bcl-2 mR-NA decreased significantly and the expression level of BAX mRNA increased significantly in both CD133 + subgroup and CD133-subgroup.However,these changing ranges of Bcl-2 mRNA and BAX mRNA were more obvious in CD133 + subgroup in comparison with those in CD133-subgroup.Conclusions In some degree,resistent potentiality of CD133 + cells to 5-FU,DDP and VP-16 has been identified,which may probably be due to the up-regulation of the expression of BAX and down-regulation of the expression of Bcl-2.

Objective To detect the expression of c-met (hepatocyte growth factor receptor), e2f-1 (transcription factor) and Ki-67 in tissues of gastric cancer, explore the relationship among them, and investigate their correlationship with clinicopathological characteristics and prognosis. Methods The tissue samples of gastric cancer from 86 patients with radical resection were subjected to immunohistochemical staining, and the expression of c-met, e2f-1 and Ki-67 was detected. The relationship between the clinicopathological characteristics and the expression of c-met, e2f-1 and Ki-67, and that among the expression of c-met, e2f-1 and Ki-67 were explored by univariate and multivariate data analysis. And the relationship between prognosis and expression of c-met, e2f-1 and Ki-67 was analysed by Log rank test. Results c-met, e2f-1 and Ki-67 were widely expressed in tissues of gastric cancer. The higher expression of c-met was associated with higher expression of Ki-67, shorter survival time, upper tumor location, higher lymph node metastasis rate, higher N stage and TNM stage (P<0.05), and the lower expression of e2f-1 was associated with larger tumor diameter, higher TNM stage, deeper invasion, higher lymph node metastasis rate and higher N stage (P<0.05). The multivariate analysis revealed that depth of invasion, N stage, TNM stage and expression of Ki-67 were independent factors for positive expression of c-met, and age and survival time were independent factors for positive expression of e2f-1. Log rank test demonstrated that the factors related to survival included expression of c-met, e2f-1 and Ki-67, N stage, tumor diameter, tumor location, lymph node metastasis rate, depth of invasion and TNM stage. Conclusion c-met is an indicator for malignant behavior and poorer prognosis of gastric cancer. There is a higher expression of e2f-1 in early gastric cancer, while advanced gastric cancer may be associated with a lower expression of e2f-1.

Objective To investigate the expression of CD133 mRNA in tissues of gastric cancer,and explore its relationship with clinicopathological features. Methods The tissues of gastric cancer and normal tissues adjacent to gastric cancer were obtained from 31 patients.The expression of CD133 mRNA was detected by semi-quantitative RT-PCR,and its relationship with clinicopathological features such as sex,age,tumor diameter,infiltration depth,TNM staging,tumor differentiation,lymph node metastasis and Ki-67 proliferation index was analysed. Results The relative gray scale values of CD133 mRNA in tissues of gastric cancer and normal tissues adjacent to gastric cancer were 0.378 3±0.141 1 and 0.038 1 ±0.091 9,respectively(P=0.000).The relative gray scale values of CD133 mRNA in tissues of gastric cancer with tumor diameter>5 cm were significantly higher than those with tumor diameter≤5 cm[(0.439 3±0.148 4)vs(0.334 3±0.121 2)](P=0.041),and those in tissues with lymph node metastasis were significantly higher than those without lymph node metastasis[(0.426 6±0.132 0)vs(0.239 5±0.030 9)](P=0.004).The rate of lymph node metastasis and the number of metastatic lymph nodes were positively related to relative gray scale values of CD133 mRNA(r=0.466,P=0.008;r=0.464,P=0.009).The relative gray scale values of CD133 mRNA in those with low expression of Ki-67 were significantly higher than those with high expression of Ki-67[(0.436 4±0.139 8)vs(0.316 4±0.117 4)](P=0.02),and expression of Ki-67 were negatively related to relative gray scale values of CD133 mRNA(r=-0.461,P=0.009).Conclusion The expression of CD133 mRNA in tissues of gastric cancer was associated with the rate of lymph node metastasis,number of metastatic lymph nodes and expression of Ki-67,which reflect the status of lymph node metastasis and proliferation of gastric cancer.

Objective To compare the inhibition effects of three synthesized fragments used in small interfering RNA(siRNA) against CD133 gene in KATO-Ⅲ gastric cancer cells,and to study effects of suppressed CD133 on the proliferating ability of intervened cells.Methods Three fragments of siRNA were designed and synthesized targeted at the mRNA of CD133.Cell fluorescence counting under confocal laser scanning microscope was used to determine the transfection efficiency after transfection with the CD133FITC-siRNA.The knock-down effect of the CD133 gene was detected by RT-PCR and Western blotting.CCK-8 (cell counting kit-8 assay) was performed to measure the variation of the cell proliferative viability after the above-mentioned treatment.Results The transfection efficiency of siRNA was (85 ± 8) ％ in KATO Ⅲ Gastric cacer cell.All these three fragments of CD133 siRNA effectively inhibited the expression of CD133 gene,the inhibition rate being (11 ± 2) ％,(19 ± 2) ％,(24 ± 3) ％respectively.Compared with the control group,the cell proliferation viability was restrained (42 ± 4)％ in CD133siRNA-3 group (P ＜0.05).Conclusions CD133siRNAs were successfully transfected into KATO Ⅲ Gastric cacer cells and repressed the expression of CD133.Meanwhile,the CD133siRNA fragment 3 was screened from three CD133 siRNA,which has the best inhibition effect.The results provide preliminary evidence for the intereference of CD133+ gastric cancer cells subsequently.

Objective To investigate the biological effect of epithelial-to-mesenchymal transition (EMT) under the treatment of transforming growth factor (TGF)-β1 on the human gastric cancer cell line SGC7901 in vitro,and to observe whether the TGF-β1 can generate the tumor initiating cells ability in SGC7901 or not.Methods SGC7901 cells were cultured with TGF-β1.The morphological change was observed.The effect on proliferation of SGC7901 cells was detected by CCK-8.The invasion assay was used to investigate the motility and the invasion ability of SGC7901 cells.Immuofluorescence was used to detect the expression of E-cadherin and N-cadherin.The mRNA and protein's expression levels of EMT-related factors and CD44 were analyzed by RT-PCR and Western blotting respectively.Results TGF-β1 induced morphological alterations from epithelial to mesenchymal cells.The proliferation of SGC7901 cells was inhibited,and the ability of motility and invasion of SGC7901 cells were greatly enhanced after being treated with TGF-β1.RT-PCR and Western blotting showed that the expression of Snail (P ＜ 0.05),N-cadherin (P ＜ 0.05) and CD44 (P ＜ 0.05) were significantly increased while the expression of E-cadherin was decreased (P ＜ 0.05).Conclusions TGF-β1 can generate the EMT.The CD44 expression was up-regulated.TGF-β1 can inhibit the proliferation and promote the motility and invasion ability of SGC7901 cells.

Objective To investigate if TGF-β1 induces epithelial-mesenchymal transition (EMT) and promotes the obtaining of stemness characteristics in gastric cancer cell lines.Methods After KATO-Ⅲ cells were cultured with or without 5 ng/mL TGF-β1,the morphological change was observed and compared under phase-contrast microscopy.At the same time,the effect of TGF-β1 on the proliferation of KATO-Ⅲ cells was detected by CCK-8.On the other hand,the mRNA and protein' s expressions of EMT-related factors,ESC markers and TICs markers were analyzed by RT-PCR and Western blotting methods too.Results TGF-β1 induced morphological alterations from epithelial to mesenchymal cells.The proliferation of KATO-Ⅲ cells was inhibited after treated with TGF-β1 (P ＜ 0.05).After treated with TGF-β1,the relative mRNA expression levels of Snail (0.5219 ±0.0147) and N-cadherin(0.6640 ±0.0124) were higher than that in control group(0.2049 ±0.0214,P =0.004,0.2722 ± 0.0098,P =0.001),the relative protein expression levels of Snail (0.4769 ± 0.0234) and N-cadherin (0.5014 ± 0.0216) were higher than that in control group (0.2534 ± 0.0345,P =0.02,0.2026 ± 0.0268,P =0.009),while the relative E-cadherin mRNA and protein levels in TGF-β1 treated group (0.4701 ± 0.0215,0.1349 ± 0.0258) were lower than that in control group (0.6792 ± 0.0157,P =0.01 ; 0.6055 ± 0.0227,P =0.004),while the relative mRNA expressions of ESC markers such as Sox2,OCT4,Nanog in TGF-β1 treated group (0.594 ± 0.039、0.438 ± 0.033、0.489 ± 0.037) were higher than that in control group (0.143 ± 0.013,P =0.001,0.156 ± 0.025,P =0.001,0.325 ± 0.046,P =0.03),the relative mRNA expression levels of CD44 (0.437 ±0.037) and CD133(0.543 ±0.028) were higher than that in control group (0.247 ±0.024,P =0.000,0.139 ± 0.016,P =0.000),the relative protein expression levels of CD44 (0.429 ± 0.034) and CD133 (0.316 ±0.027) in TGF-β1 treated group were higher than that in control group (0.152 ± 0.014,P =0.000,0.110 ±0.010,P =0.000),cloning sphere-forming capacity was greatly enhanced after treated with TGF-β1 (P ＜ 0.01).Conclusion TGF-β1 can induce EMT in KATO-Ⅲ cells and promote the obtaining of stemness characteristics in gastric cancer cell lines.

Objective To investigate the clinicopathologic characters and prognostic value of epithelial mesenchymal transition (EMT) related proteins Snail and chemokine receptors 4(CXCR4) in gastric cancer.Methods The expressions of Snail and CXCR4 in the gastric tissues of 50 patients with gastric cancer were detected by Immunohistochemistry.The relation between the expressions of Snail and CXCR4 and the clinicopathologic characters were analyzed.The relative relationship between Snail expression and CXCR4 expression were identified by Spearman method.The correlation between the expressions of Snail and CXCR4 with the survival was analyzed by Kaplan-Meier method.Results The expression of Snail and CXCR4 expression were positive in 31 of 50 cases (62％) and 29 of 50 cases (58％) respectively.The expression levels of Snail and CXCR4 in the gastric cancer patients with diameter more than 5 cm,worse tissue differentiation,vascular invasion,lymphatic vessel invasion,lymph nodes metastasis,and T3 + T4 staging were significantly higher than those in the patients with diameter less than 5 cm,without vascular invasion,lymphatic vessel invasion,lymph nodes metastasis,and T1 + T2 staging (P ＜ 0.05).The Expression of Snail and CXCR4 was positively Correlated(r =0.330,P =0.014).The co-expression of Snail and CXCR4 was significantly associated with poorer prognosis and was applied as an independent prognostic factor for gastric cancer (P =0.001).Conclusions The co-expression of Snail and CXCR4 was the independent prognostic factor for gastric cancer.The combining effect of EMT and CXCR4 may promote the progressions and metastasis of gastric cancer.Therefore,it may be of an effective way for the metastasis of gastric cancer to adjust these targets of the Snail and CXCR4.