The Expression And The Association Of VEGF-C、VEGF-B And TIAM-1in Gastric Carcinoma

Background and objectiveGastric cancer is the forth most common malignancy in the world, and is the second leading cause of cancer death after lung cancer. Lymph node metastasis is the strongest prognostic factor, and the major cause of treatment failure and death of patients with gastric cancer. About60%of resecable gastric cancers have lymph node metastasis at the time of diagnosis, and so, the5-year overall survival of advanced gastric cancer is lower than20%. Thus, to predict Lymph node metastasis tendency, identify the prognosis markers, find the effective way for prevention and therapy and raise five-year survival rate of Gastric cancer patients have been the hot spots for studies, and also been an emphasis and difficult point in treatment and prevention of tumor.Growth and metastasis of tumor depend on angiogenesis; the most key factor is vascular endothelial growth factor family (VEGFs) in encouraging angiogenesis. VEGFs include VEGF-A、B、C、D、 and PIGF (placenta growth factor).Recently, many studies reveal that VEGF has abnormal expression in many malignant tumors. VEGF-B and its receptor Flt-1are found in human nervous system tumor, breast carcinoma, renal carcinoma, cancer of colon, lung cancer etc. They have relation to lymphatic vessel invasion and lymphatic metastasis of colon and lung cancer; Regulate formation of tumor vessel in renal carcinoma; they have correlation with lymphatic metastasis, not promote angiogenesis in breast carcinoma; in process of malignant neurospongi-oma vessel formation, they may play a role as a assistance factor. At present, the therapy with anti-VEGF receptor provides a new method for breast cancer. But, the expression and significance of VEGF-B and its receptor in gastric carcinoma haven’t been found in reports.Vascular endothelial growth factor C (VEGF-C) and its receptor VEGF receptor-3(VEGFR-3, also designated Flt-4) are recently identified lymphangiogenic stimulators that can selectively induce growth of the lymphatic vasculature. In recent years, the vascular endothelial growth factor C (VHGF-C) and its receptor VEGFR-3are found, and VEGF-C is known that it is a relatively specific lymphatic endothelial growth factor. The expansion of lymphatic endothelial to promote the formation of lymphatic vessels is occurred after VEGF-C and VEGFR-3combine together and thus to study the mechanism of lymph node metastasis has opened new avenues. They have strong special function of promoting the formation of lymphatic vessels after VEGF-C combined with VEGFR-3. And was known keep a close relationship with lymph node metastases.Tiam-1(T lymphoma invasion and metastasis1) was originally identified as the invasion-and metastasis-inducing gene by proviral tagging in combination with in vitro selection for invasiveness in T lymphoma cells. Tiam1is one of guanine nucleotide exchange factors (GEFs), which activate GTPases by promoting the exchange of their inactive GDP-bound forms to their active GTP-bound forms. Whereas Tiam-1displays GEF activity towards all three Rho-like GTPases Racl, Cdc42and RhoA in vitro, Tiaml specifically activates Rac in vivo. Recent evidence suggests that Tiam-1could influence Rac GTPases signaling specificity in addition to promoting their activation.Tiam1has been implicated to directly bind to many different cytoplasmic and membrane-associated proteins, which couples Tiam-1-Rac activity to specific signaling pathways. The role of Tiam-1in cellular migration, invasion and metastasis may not be limited to T lymphoma. Related studies on breast cancer, lung cancer and skin cancer induced by Ras confirmed that Tiam1could accelerate tumor progression and metastasis significantly. In the Tiam-1-knockout animal model, the incidence rate of mouse skin cancer obviously reduced and tumors were grown slowly. The above studies showed that Tiam1might accelerate carcinogenesis and metastasis in many tumors.Lymph node metastasis is one of the most important negative predictor to patients with gastric cancer. It’s especially important to be diagnosis for metastases of lymph node. The most common clinical device used to diagnosis cannot give us a little useful advice about lymph node metastases. There is no specificity or sensitivity to lymph node metastasis. To study the basic theory about lymph node metastases in gastric cancer can be helpful to look for the gold key to see the essential qualities of lymph metastases. With the study progresses on VEGF-C and Lymph node metastasis, we can gradually master the matter used on early diagnosis, correct stage, improve operation style, select better treatment plan after surgery and investigate new drugs resist tumor. It will be improve the treatment about gastric cancer and have great significance. In this study, the VEGF-C and Tiam-1expression level was detected by RT-PCR, immunohistochemical streptavidin peroxidase method, and ELISA method; And all the data was analyzed statistically with clinical and pathological features. So we can investigate the relationship among the expression of VEGF-C and Tiam-land the relationship between them and the lymphatic metastasis or micro metastasis. To study the correlation ship of the expression level of VEGF-C and Tiam-1between cancer tissue and peripheral blood serum, so identify if it can be taken the place by VEGF-C and Tiam-1protein detected in serum, so we can find a easier method detection way. The relationship between the VEGF-C and Tiam-1expression’s level and tumor treatment was selected. The clinical significance of VEGF-C and Tiam-1expression as a tumor marker was analyzed together.Methods1. Clinical-pathologic parameters:Study information of Age, gender, tumor size, Lauren classification, TNM stage, vascular invasion, serosal invasion, and lymphatic invasion is collected.2. Measurement of serum VEGF-B、VEGF-C level in patients with gastric carcinoma: Serum VEGF-B、VEGF-C level of normal people, preoperative and postoperative patients and followed up patients was examined by using ELISA.3. The expressions of VEGF-B、VEGF-C and Tiam-1gene/protein in tissue:Real time PCR, immunochemistry, Western blot were used to examine the expressions of VEGF-B、VEGF-C and Tiam-1in human gastric cancer tissue, surrounding lymph nodes and normal gastric tissue.4. Statistical analysis:SPSS13.0software for statistical applications the relationship of VEGF-C and Tiam-1protein expressions with clinical-pathologic parameters was analyzed with x2test. Kaplan-Meier method was used to retrospectively analyze the relationship of VEGF-C and Tiam-1protein expressions with life span of gastric carcinoma patients. The relationship of single factor of each index or multi-variable combination with life span was analyzed by COX regressionResults1. The serum VEGF-C level in patients with gastric cancer before surgery is higher than that in healthy controls. The concentrations were statistically significantly different in preoperation and postoperation, P<0.01; preoperation and health controls, P<0.01, respectively. But there is no statistically different in serum level between post-operation and healthy controls, P>0.05.There is no obvious difference of VEGF-B level in preoperation and postoperation2.The level of serum VEGF-C in group with metastatic cancer is different from that in group without it, There is a significantly statistically different, P<0.01. There is a significantly statistically different in serum VEGF-C levels between group with metastatic cancer and control healthes, P<0.01.The level is no different between group without metastatic cancer and control healthes, P>0.05. Serum VEGF-B is not significantly different among control healthes, metastatic group and nonmetastatic cancer group.3. The VEGF-C level of gastric cancer patients is related with the lymph node metastasis, serosal invasion and clinical stage, the VEGF-C level in patients without lymph node metastasis, no serosal invasion, TNM Ⅰ and well and moderate differentiated group phase was significantly lower than that suffering the lymph node metastasis, serosal invasion, TNM Ⅱ Ⅲ Ⅳ stage and poorly differentiated and undifferentiated group. The relationship between VEGF-B and them is not obvious4. The expression of VEGF-C and Tiam-1mainly focus on primary foci in stomach tissue and lymph node metastases. The positive rate of the VEGF-C and Tiam-1expression with lymph node metastasis was significantly higher than those without lymph node metastasis of gastric cancer, the difference was significant. The VEGF-C and Tiam-1expression of gastric cancer tissue is related with the lymph node metastasis, serosal invasion and clinical stage, the VEGF-C level in patients without lymph node metastasis, no serosal invasion and TNM Ⅰ Ⅱ was significantly lower than that suffering the lymph node metastasis, serosal invasion, TNMⅢ Ⅳ stage and poorly differentiated and undifferentiated group.5. Real-time quantitative PCR showed that the VEGF-C and Tiam-1gene level between normal gastric tissue and gastric cancer or lymph node tissue was significantly different (P<0.05); There was a close relationship between quantitaty of VEGF-C and Tiam-1mRNA and lymph node metastasis, quantitaty of VEGF-C and Tiam-1mRNA was higher in cases with lymph node metastasis than those without lymph node metastasis (P<0.01). With the degree of lymph node metastasis, the VEGF-C、TIAM-1mRNA got more obviously (p<0.01)6. Spearman correlation analysis showed that there was a line relationship between VEGF-C and Tiam-1expression in gastric cancer; the expression level of VEGF-C and TIAM-1increased With the lower degree of differentiation, increased depth of invasion, worse TNM stage and Wider lymph node metastasis7. Kaplan-Meier survival curves show the survival time had significant difference between patients with positive VEGF-C and Tiam-1expression and those without expression (P<0.05)..Multi-factor analysis showed that the high expression of VEGF-C in tumor tissue and Tiam-1in lymph node, TNM were independent factors affecting the survival prognosis of gastric carcinoma (P value was0.003、0.007、0.028respectively)Conclusion1. The serum VEGF-C rather than VEGF-B level is a reliable biologic marker in diagnosing gastric carcinoma, the VEGF-C levels up-regulation appears to be useful biologic marker for gastric cancer metastasis or recurrence.2. VEGF-B were expressed in some cases but it had no relation with clinic parameter2. VEGF-C and Tiam-1protein was up-regulated in gastric carcinoma tissues and metastatic lymph nodes, and had no expression in normal gastric tissue, indicating that VEGF-B、VEGF-C and Tiam-1may be regarded as the important marker for clinical detection of gastric carcinoma;3. As tight correlations existed between the expressions of VEGF-C, Tiam-1and the pathobiological behavior of gastric cancer, both of them maybe acted as reference indexes for judging the invasive and metastatic state of gastric cancer.4. Combined evaluation of VEGF-C and Tiam-1high expression and clinical follow-up data analysis showed that VEGF-C and Tiam-1had close relationship with prognosis of gastric carcinoma patients. VEGF-C and Tiam-1may act as the important marker for judging prognosis of gastric carcinoma;5. The VEGF-C and Tiam-1play an important role in the course of development, progression and lymphatic metastasis of gastric cancer. The agreed expression of VEGF-C and Tiam-1was related to lymph node metastasis, it indicated the cooperation effection of VEGF-C and Tiam-1may promote lymphatic metastasis in gastric cancer, which provides a new target for the Treatment of lymph node metastasis of gastric cancer...