Vascular Endothelial Growth factor-C Protein Research Articles

VEGF-C: An Indicator of Lymph Node Metastasis for Patients that Underwent Esophagectomy

To investigate the expression of vascular endothelial growth factor-C (VEGF-C) and its relationship to lymph node metastasis in esophageal squamous cell cancer after esophagectomy. The expression of VEGF-C protein was detected by immunohistochemistry. Kaplan-Meier method was performed to calculate the survival rate. Cox regression multivariate analysis was performed to determine independent prognostic factors. The difference of VEGF-C protein expression in Tl, T2 and T3 patients was statistically significant (p=0.031). The expression of VEGF-C protein in patients with lymph node metastasis was significantly higher than those without metastasis (p=0.004). VEGF-C protein expression correlated with significantly worsened 5-year survival for all patients as well as those with T3 tumors and N1 nodal status. The result of Cox analysis demonstrated that N stage and VEGF-C protein expression were independent prognostic factors; the multivariate analyses indicated that the N classification is the independent pathological risk factor of VEGF-C expression. VEGF-C expression was found to be significantly associated with T stage and N stage. The patients with VEGF-C protein expression had a significantly lower 5-year survival rate than without VEGF-C protein expression. Lymph node metastasis and VEGF-C protein expression were independent prognostic factors.

Dynamic aspects of corneal lymphatic vessels after keratoplasty in rats

To study the development of lymphatic vessels after keratoplasty and to explore the molecular mechanisms of corneal lymphangiogenesis in transplanted corneas. Experimental research. The development of corneal lymphangiogenesis was examined by LYVE-1 immunohistochemistry and whole mount immunofluorescence 1, 3, 7, 10, 14, 30 and 60 days after corneal transplantation, then lymphatic vessels counting (LVC)was evaluated. The expression of vascular endothelial growth factor-C (VEGF-C) in transplanted corneas was examined by immunohistochemistry, enzyme-linked immunosorbent assay (ELISA), and real time-PCR at same time. In addition, the inflammatory index (IF) was recorded at each time point. The association of VEGF-C and IF with LVC in transplanted corneas was examined. Analysis of the significance of differences between two groups was performed using paired Student's t-test. Pearson's analysis was used to analyze the correlation between VEGF-C, IF and LVC. Corneal lymphangiogenesis occurred in the stroma with LVC (1.8 ± 0.3) on Day 3, then developed and reached the peak with LVC (9.1 ± 1.5) on Day 14 after corneal transplantation. Both VEGF-C protein and mRNA up-regulated dramatically in rat transplanted corneas. The immunoreactivity reached the peak on the 3(rd) day and the 14(th) day after keratoplasty. Compared with the expression of VEGF-C mRNA (1.62 ± 0.08 copies/g) on Day 3, the expression of VEGF-C mRNA (2.48 ± 0.03 copies/g) significantly increased 14 days after the transplantation (t = 4.296, P = 0.02). LVC was strongly and positively correlated with IF (r = 0.55, P = 0.003) and the expression of VEGF-C mRNA (r = 0.51, P = 0.003). Corneal lymphangiogenesis correlates closely with corneal inflammation. The increased expression of VEGF-C in the cornea may be one of the important molecular mechanisms in the occurrence of corneal lymphangiogenesis after keratoplasty.

Expression and clinical significance of extracellular matrix protein 1 and vascular endothelial growth factor-C in lymphatic metastasis of human breast cancer.

BackgroundExtracellular matrix protein 1 (ECM1) and vascular endothelial growth factor-C (VEGF-C) are secretory glycoproteins that are associated with lymphangiogenesis; these proteins could, therefore, play important roles in the lymphatic dissemination of tumors. However, very little is known about their potential roles in lymphangiogenesis. The aim of this study was to investigate whether correlations exist between ECM1 and VEGF-C in human breast cancer, lymphangiogenesis, and the clinicopathological characteristics of the disease.MethodsECM1 and VEGF-C mRNA and protein expression levels in 41 patients were investigated using real-time reverse transcriptase polymerase chain reaction (RT-PCR), or immunohistochemical (IHC) staining of breast cancer tissue, matched noncancerous breast epithelial tissues, and suspicious metastatic axillary lymph nodes. D2-40 labelled lymph vessels and lymphatic microvessel density (LMVD) were counted. Correlations between ECM1 or VEGF-C protein expression levels, LMVD, and clinicopathological parameters were statistically tested.ResultsThe rate of ECM1 positive staining in breast cancer tissues was higher (31/41, 75.6%) than that in the corresponding epithelial tissues (4/41, 9.8%, P < 0.001) and lymph nodes (13/41, 31.7%, P < 0.001). Similarly, the VEGF-C expression rate in cancer specimens was higher (33/41, 80.5%) than in epithelial tissues (19/41, 46.3%, P < 0.01) or lymph nodes (15/41, 36.6%, P < 0.01). Higher ECM1 and VEGF-C mRNA expression levels were also detected in the tumor tissues, compared to the non-cancerous tissue types or lymph nodes (P < 0.05). ECM1 protein expression was positively correlated with the estrogen receptor status (P < 0.05) and LMVD (P < 0.05). LMVD in the ECM1- and VEGF-C-positive tumor specimens was higher than that in the tissue types with negative staining (P < 0.05).ConclusionsBoth ECM1 and VEGF-C were overexpressed in breast cancer tissue samples. ECM1 expression was positively correlated with estrogen responsiveness and the metastatic properties of breast cancer. We conclude, therefore, that ECM1 and VEGF-C may have a synergistic effect on lymphangiogenesis to facilitate lymphatic metastasis of breast cancer.

Expression of vascular endothelial growth factor C in esophageal carcinoma and its clinical significance

Objective To investigate the expression of vascular endothelial growth factor C (VEGF-C) in human esophageal squamous cell carcinoma (ESCC) and its clinical significance.Methods The expression of VEGF-C in ESCC and its correlation with lymph node metastasis were detected by using immunohistochemistry (IHC) and in situ hybridization (ISH).Three small interfering RNA (siRNA) expression vectors targeting VEGF-C were constructed and transformed into EC9706 cells.The inhibitory effect of siRNA on the expression of VEGF-C protein and mRNA in tranfected cells was analyzed by using Western blotting and reverse transcription-polymerase chain reaction (RT-PCR).Stably transfected cells were transplanted in nude mice later.The expression of VEGF-C in transplant tumor was detected by using IHC and ISH.Results VEGF-C was up-expressed in ESCC.Its expression was correlated significantly with lymph node metastasis.The positive expression rate of VEGF-C in lymph node metastasis group and non-lymph node metastasis group was 90.0％ and 62.1％ respectively (P ＜0.01 ).VEGF-C-targeted siRNA could decrease the expression of VEGF-C mRNA and protein in EC9706 cells effectively.There was significant difference between transfected group (0.32 ± 0.02) and un-transfected group (0.91 ± 0.09,P ＜0.01 ).Correspondingly,the expression of VEGF-C was 9.26 ±0.95,8.94 ±0.81,9.19 ±0.92 and 32.47 ±3.25 respectively.There was significant difference among them (P ＜0.01 ).Conclusion Targeting VEGF-C siRNA can inhibit VEGF-C expression of EC9706 in vivo and in vitro effectively. Key words: Esophageal neoplasm; Vascular endothelial growth factor C

The effects of vascular endothelial growth factor C knockdown in esophageal squamous cell carcinoma

We investigated the role of vascular endothelial growth factor C (VEGF-C) in esophageal squamous cell carcinoma (ESCC) by knocking down VEGF-C expression in the ESCC cell line EC9706. Immunohistochemistry and in situ hybridization techniques were used to detect the expression of VEGF-C expression in ESCC tissues. We also investigated the relationship between VEGF-C expression and lymph node metastasis. We designed a siRNA expression plasmid for VEGF-C and transfected it into EC9706 cells. Stable clones were selected, and VEGF-C expression was analyzed by RT-PCR and western blotting. Cells were inoculated into nude mice. The expression of VEGF-C in the resulting tumors was analyzed by immunohistochemistry and in situ hybridization. VEGF-C is highly expressed in ESCC and correlated with lymph node metastasis, as high levels were observed in patients presenting with lymph node metastases relative to those who did not (P<0.01). Transfection with VEGF-C-siRNA decreased the expression of VEGF-C mRNA and protein. ESCC cells stably transfected with VEGF-C-siRNA expressed very low levels of VEGF-C (P<0.01 compared with control). This knockdown effect persisted when the cells were inoculated into nude mice and allowed to form tumors. The siRNA-targeted knockdown of VEGF-C led to a significant reduction in VEGF-C expression. This siRNA technique could be used for gene therapy in ESCC.

CYR61 Modulates the Vascular Endothelial Growth Factor C Expression of Decidual NK Cells Via PI3K/AKT Pathway

Either vascular endothelial growth factor C (VEGFC) or CYR61 plays an important role in placental development and may be involved in pre-eclampsia. Decidual natural killer (dNK) cells are the main source of VEGFC in the maternal-fetal interface. However, it is unclear about CYR61 on the regulation of VEGFC secretion in dNK cells. Decidual natural killer cells were isolated from decidual tissues of first trimester of pregnancy with anti-human CD56-conjugated microbeads. Integrin αvβ3 was detected using immunofluorescent staining. dNK cells were cultured in the presence of CYR61, anti-human αvβ3 integrin antibody (LM609), PI3K inhibitor (LY294002), or MEK inhibitor (U0126). VEGFC mRNA and protein were evaluated by real-time PCR and ELISA, respectively. Exogenous CYR61 induced the expression of VEGFC in dNK cells in both mRNA and protein levels. Integrin αvβ3 was strongly expressed on dNK cell surface. Anti-αvβ3 integrin antibody inhibited the effect of CYR61 on VEGFC expression. LY294002, but not U0126, significantly reduced this promotion effect of CYR61 on dNK cells. The upregulation of VEGFC secretion mainly depends on CYR61 binding with integrin αvβ3 on the surface of dNK cells. PI3K/AKT, rather than the ERK/MAPK signal, is involved in the regulation.

RNA Interference-Mediated Vascular Endothelial Growth Factor-C Reduction Suppresses Malignant Progression and Enhances Mitomycin C Sensitivity of Bladder Cancer T24 Cells

Vascular endothelial growth factor-C (VEGF-C) has been found to be significantly associated with lymphangiogenesis and regional lymph node metastasis in various human tumors. The present work was aimed to explore the role of VEGF-C in malignant progression of human bladder cancer T24 cell line. First, the expression of VEGF-C in T24 cells was detected by western blotting. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was employed to measure the cellular proliferation after treatment with various concentrations of recombinant human VEGF-C (rhVEGF-C). Then, lentivirus vector-based RNA interference (RNAi) was used to inhibit VEGF-C expression of T24 cells. The alterations of T24 cells regarding proliferation, invasiveness, and the apoptosis induced by mitomycin C (MMC) were evaluated. The results showed that the proliferation rate of T24 cells rose from 27.3% to 65.0%, with increasing rhVEGF-C concentration. T24 cells stably transfected with VEGF-C small interference RNA showed 85% reduction in VEGF-C mRNA expression (p < 0.05). The VEGF-C protein level was significantly downregulated (p < 0.05) and the growth and invasiveness were also inhibited (p < 0.05) compared with the control group. Further, the inhibition of VEGF-C expression markedly enhanced the apoptosis of T24 cells induced by MMC (p < 0.05). These were associated with the decreased ratio of Bcl-2/Bax, activation of Caspase-3, decreased expression of MMP-9, as well as the downregulation of phosphorylated p38 MAPK and Akt. The present study suggests that VEGF-C can enhance the proliferation and invasiveness of bladder cancer T24 cells, which is due to suppression of apoptosis and facilitation of migration, accompanied with upregulation of p38 MAPK and Akt phosphorylation. RNAi targeting VEGF-C could effectively suppress malignant progression and enhance chemosensitivity of T24 cells. Thus, inhibition of VEGF-C expression is a potential and promising therapeutic strategy for bladder cancer.

Metastasis-associated protein 1 induces VEGF-C and facilitates lymphangiogenesis in colorectal cancer

To study the correlation between high metastasis-associated protein 1 (MTA1) expression and lymphangiogenesis in colorectal cancer (CRC) and its role in production of vascular endothelial growth factor-C(VEGF-C). Impact of high MTA1 and VEGF-C expression levels on disease progression and lymphovascular density (LVD, D2-40-immunolabeled) in 81 cases of human CRC was evaluated by immunohistochemistry. VEGF-C mRNA and protein expressions in human LoVo and HCT116 cell lines were detected by real-time polymerase chain reaction and Western blotting, respectively, with a stable expression vector or siRNA. The elevated MTA1 and VEGF-C expression levels were correlated with lymph node metastasis and Dukes stages (P < 0.05). Additionally, high MTA1 expression level was correlated with a large tumor size (P < 0.05). A significant correlation was found between MTA1 and VEGF-C protein expressions in tumor cells (r = 0.371, P < 0.05). Similar to the VEGF-C expression level, high MTA1 expression level was correlated with high LVD in CRC (P < 0.05). Furthermore, over-expression of MTA1 significantly enhanced the VEGF-C mRNA and protein expression levels, whereas siRNAs - knocked down MTA1 decreased the VEGF-C expression level. MTA1, as a regulator of tumor-associated lymphangiogenesis, promotes lymphangiogenesis in CRC by mediating the VEGF-C expression.

Vascular Endothelial Growth Factor-C Associated with Computed Tomography Used in the Diagnosis of Lymph Node Metastasis of Bladder Carcinoma

Vascular endothelial growth factor C (VEGF-C) has been demonstrated to stimulate the growth of lymphatic vascular endothelium. The purpose of this study is to determine whether VEGF-C associated with computed tomography (CT) has a relationship with lymph node metastasis in bladder transitional cell carcinoma (BTCC). One hundred twenty seven cases of BTCC were studied: first, both plain and enhanced CT scans of abdomen and pelvis were performed preoperatively; second, VEGF-C protein expressions in tumor cells were tested by immunohistochemistry postoperatively; and finally, detailed pathological findings for lymph node metastasis were recorded. VEGF-C expressions in tumor bladder cells were significantly higher than those in normal bladder epithelium. In the group of BTCC-positive, VEGF-C was significantly correlated with lymph node metastasis (p <0.01). Sensitivity, specificity and accuracy of VEGF-C in the diagnosis of lymph node metastasis of bladder carcinoma were 87.0, 67.9, and 74.8%, respectively. Sensitivity, specificity and accuracy of CT were 47.8, 80.2, and 68.5%, respectively. When VEGF-C visually correlated with CT scan in the detection of lymph node metastasis, sensitivity was 91.3%; specificity was 84.0% and accuracy was 86.6%. Positive VEGF-C was significantly correlated with lymph node metastasis of bladder carcinoma. VEGF-C expression in biopsy specimens may be beneficial in predicting pelvic lymph nodes. It can improve accuracy when combined with CT.

Organ‐specific lymphangiectasia, arrested lymphatic sprouting, and maturation defects resulting from gene‐targeting of the PI3K regulatory isoforms p85α, p55α, and p50α

The phosphoinositide 3-kinase (PI3K) family has multiple vascular functions, but the specific regulatory isoform supporting lymphangiogenesis remains unidentified. Here, we report that deletion of the Pik3r1 gene, encoding the regulatory subunits p85alpha, p55alpha, and p50alpha impairs lymphatic sprouting and maturation, and causes abnormal lymphatic morphology, without major impact on blood vessels. Pik3r1 deletion had the most severe consequences among gut and diaphragm lymphatics, which share the retroperitoneal anlage, initially suggesting that the Pik3r1 role in this vasculature is anlage-dependent. However, whereas lymphatic sprouting toward the diaphragm was arrested, lymphatics invaded the gut, where remodeling and valve formation were impaired. Thus, cell-origin fails to explain the phenotype. Only the gut showed lymphangiectasia, lymphatic up-regulation of the transforming growth factor-beta co-receptor endoglin, and reduced levels of mature vascular endothelial growth factor-C protein. Our data suggest that Pik3r1 isoforms are required for distinct steps of embryonic lymphangiogenesis in different organ microenvironments, whereas they are largely dispensable for hemangiogenesis.

Heparanase expression correlates with angiogenesis and lymphangiogenesis in human lung cancer.

Heparanase has been thought to be a good molecular marker of tumor, and the heparanase expression level was correlated closely with tumor metastasis. In this study, we investigate the effects of heparanase on angiogenesis and lymphangiogenesis of lung cancer and the relationship between heparanase expression and vascular endothelial growth factor (VEGF), vascular endothelial growth factor-C (VEGF-C). Immunohistochemistry was used to detect the expression of heparanase, VEGF, VEGF-C protein and microvascular density (MVD), lymphatic vessel density (LVD) in 115 cases of non-small cell lung cancer (NSCLC) and 45 cases of adjacent normal tissue samples. Our results showed that heparanase expression was significantly increased in 91 (79.13%) of the 115 cases and correlated with lymph node metastasis (node positive rate 87.0%; node negative rate 36.8%; P=0.003). Heparanase positive expression cases have significantly higher concentration of microvascular density (MVD) and lymphatic vessel density (LVD) as compared with heparanase negative expression cases (P<0.01, P<0.01, respectively), heparanase expression was significantly correlated with VEGF, VEGF-C expression in NSCLC. Heparanase overexpression was associated with angiogenesis and lymphangiogenesis of lung cancer, targeting of heparanase may represent a significant therapeutic potential for lung cancer.

Inhibition of Cyclooxygenase‐2 Suppresses Lymph Node Metastasis via VEGF‐C

Most experimental work addressing cyclooxygenase-2 (COX-2) inhibitor has focused on suppressing hematogenic spread. Little is known about the mechanism by which this inhibitor can also block lymphatic metastasis. Here, the effects of COX-2 inhibitor on vascular endothelial growth factor-C (VEGF-C) expression, lymphangiogenesis and lymph node metastasis were investigated. Utilizing the highly metastatic human lung adenocarcinoma cell line Anip973 and its parental line AGZY83-a, which has a low metastatic capacity, we found elevated VEGF-C and COX-2 immunoreactivity in Anip973 cells compared with AGZY83-a cells. Celecoxib down-regulated expression of VEGF-C mRNA and protein in Anip973 cells while PGE(2) up-regulated expression of VEGF-C mRNA and protein in AGZY83-a cells in a concentration-dependent manner. The expression of COX-2 and VEGF-C was significantly increased in xenografted Anip973 tumors compared with AGZY83-a tumors. The Anip973 tumors showed more lymphatic vessels and lymph node metastasis than the AGZY83-a tumors. In vivo, celecoxib decreased VEGF-C expression in Anip973 tumor-treated mice to a similar level to that in the AGZY83-a tumor-treated mice. Consistent with this decrease in VEGF-C expression, the density of lymphatic vessels and lymph node metastasis in Anip973 tumor-treated mice were suppressed to approximately that found in the AGZY83-a tumor-treated ones. Taken together, our results suggest that the differential expression of COX-2 and VEGF-C might help explain the different metastasis phenotype of lung adenocarcinoma cancer, and that COX-2 inhibitor mediates VEGF-C to block lymphangiogenesis and lymph node metastasis. Thus, COX-2 may be a potential therapeutic target for blocking lymph node metastasis in lung adenocarcinoma.

Role of short haipin RNA targeting vascular endothelial growth factor-C on biological characteristics of human breast cancer cell MCF-7

To study the effect of short haipin RNA (shRNA) on expression of vascular endothelial growth factor C (VEGF-C) and proliferation and invasion behavior of human breast cancer cell MCF-7. The recombinant vector (pSIREN-VEGF-C) was transfected into the human breast cancer cell MCF-7 by liposome and the positive transfected cell clones were screened with puromycin. Expression of VEGF-C in MCF-7 cells after gene transfer was detected by real-time quantitative PCR and Western blot assay, respectively. Proliferation and invasion ability of transfected cells were analyzed by MTT and Transwell filter. The expressions of VEGF-C mRNA and protein were decreased markedly compared with the control group after the transfection and the inhibitive ratio was 95% and 100% respectively (P<0.05). The proliferation of MCF-7 cells transfected by pSIREN-VEGF-C, measured with MTT assays, was significantly decended (P<0.05). The invasion ability of passing through the Transwell filter of MCF-7 cells transfected by pSIREN-VEGF-C were declined evidently (P<0.05). The recombinant vector (pSIREN-VEGF-C) have been proved not only to be effective and specific for down-regulation of VEGF-C, but also can inhibit the proliferation and invasion of MCF-7 cells significantly.

The Expression of RNA-Binding Protein HuR in Non-Small Cell Lung Cancer Correlates with Vascular Endothelial Growth Factor-C Expression and Lymph Node Metastasis

The expression levels of the RNA-binding protein Hu antigen (HuR) and vascular endothelial growth factor-C (VEGF-C) were examined immunohistochemically in 81 non-small cell lung cancers (NSCLC) and 15 benign human lung tissues. HuR showed a nuclear overexpression in 82.7% (67/81) of NSCLC specimens. Cytoplasmic immunoreactivity for HuR was observed in 45.7% (37/81) of NSCLC, while only nuclear expression of HuR was observed in 13.3% (2/15) of benign lung tissues. The expression of VEGF-C was present in a subgroup of 70.4% (57/81) of tumor cases. In the human NSCLC samples, cytoplasmic but not nuclear HuR expression was significantly associated with increased levels of VEGF-C and with clinicopathological variables, including high tumor grade, poor differentiation and lymph node metastasis. In vitro, HuR showed a predominantly nuclear staining in Lewis lung cancer cells, as seen by confocal microscopy. When lung cancer cells were treated with siRNA targeted against HuR, expression levels of the HuR and VEGF-C proteins were significantly reduced, as seen by Western blotting. Our findings indicate that there is a dysregulation of the cellular distribution of the mRNA stability factor HuR in a subset of NSCLC. Examination of cytoplasmic HuR in NSCLC tissues will allow for valuable prognostic diagnosis of lymph node metastasis, as HuR might be an important mediator regulating the expression of VEGF-C.

Expression of cyclooxygenase-2, vascular endothelial growth factor-c and CD44v6 proteins and its relation with lymphatic metastasis of pancreatic cancer

Objective To investigate the expression of cyclooxygenase-2 (COX-2), vascular en-dothelial growth factor-c (VEGF-C) , CD44v6 in pancreatic cancer tissue and its relation with the ctinico-pathologic features. Methods The expression of COX-2, VEGF-C and CD44v6 proteins in 48 cases of pancreatic cancer was detected by immnohistochemical staining. Results The positive rate of COX-2, VEGF-C and CD44v6 protein in pancreatic cancer tissue was 79.2% ,58.3%, and 62.5%, respectively, which was higher than in adjacent tissues (0/14) ( P 0.05 ), but that of VEGF-C and CD44V6 remarkably with lymph node metastasis ( P 0.05 ). Conclusion The expres-sion of COX-2, VEGF-C and CD44v6 in pancreatic cancer tissue might play a role in lymphatic metastasis via lymphangiogenesis. Key words: Pancreatic neoplasm; Cyclooxygenase-2; Vascular endothelial growth factor

The Expression of Vascular Endothelial Growth Factor C in Transplanted Corneas

Purpose: To discuss the expression of vascular endothelial growth factor C (VEGF-C) after keratoplasty in rats. Methods: The Fischer 344 rat corneas were transplanted into Lewis rat eyes. The expression of VEGF-C in cornea was examined by immunohistochemistry and real-time PCR. The association of corneal lymphangiogenesis, corneal inflammation, and rejection score (RS), with VEGF-C in transplanted corneas was also examined. Results: Both VEGF-C protein and mRNA upregulated dramatically in rat transplanted corneas. The immunoreactivity was at peak on the 3rd day and the 14th day after keratoplasty, respectively. VEGF-C mRNA was strongly and positively correlated with inflammatory index (IF) (r = 0.93; p < 0.01), lymphatic vessel counting (LVC) (r = 0.40; p < 0.05), and RS (r = 0.50; p < 0.05). The development of VEGF-C mRNA, IF, and RS was almost parallel in the second week after keratoplasty. Conclusions: VEGF-C was expressed in rat transplanted corneas. The expression of VEGF-C correlated closely with corneal inflammation, corneal lymphangiogenesis, and allograft rejection.

Vascular Endothelial Growth Factor-C and C-C Chemokine Receptor 7 in Tumor Cell–Lymphatic Cross-talk Promote Invasive Phenotype

Most carcinomas spread to distant sites through lymphatic vessels. Several preclinical and clinical studies have shown a positive correlation between the incidence of lymph node metastasis and secretion of the lymphatic growth factor vascular endothelial growth factor-C (VEGF-C) by tumor cells, suggesting tumor lymphangiogenesis as an escape mechanism. However, recent evidence has shown VEGF receptor-3 (VEGFR-3) expression on tumor cells and autocrine signaling, which increase metastatic potential. Furthermore, there is growing evidence implicating lymphatic-homing chemokine receptors, particularly C-C chemokine receptor 7 (CCR7), in lymph node metastasis. We report here that expressions of VEGF-C and CCR7 by tumor cells act synergistically to promote their invasion toward lymphatics. First, VEGF-C acts to increase lymphatic secretion of CCL21, which in turn drives CCR7-dependent tumor chemoinvasion toward lymphatics. Second, VEGF-C acts in an autocrine fashion to increase tumor invasiveness by increasing the proteolytic activity and motility of tumor cells in a three-dimensional matrix. Both of these effects are VEGFR-3 dependent and evident only in three-dimensional environments. We further verified that VEGF-C induces lymphatic CCL21 up-regulation in vivo by direct injection of VEGF-C protein intradermally in the mouse. Taken together, these results bridge the prometastatic functions of CCR7 and VEGF-C in tumors and show that, beyond lymphangiogenesis, VEGF-C promotes tumor invasion toward lymphatics by both autocrine and CCR7-dependent paracrine signaling mechanisms, which may be a significant cause of lymph node metastasis.

Relationship between vascular endothelial growth factor C expression level and lymph node metastasis in non small cell lung cancer

To investigate the relationship between vascular endothelial growth factor C (VEGF-C) expression level and lymph node metastasis in non small cell lung cancer (NSCLC). Fifty-two NSCLC patients, 38 males and 14 females, aged (59+/-11) (29-77), were divided into 2 groups based on the pathological examination: lymph node metastasis positive group (n=25) and lymph node metastasis negative group (n=27). RT-PCR and immunohistochemistry were used to detect the mRNA and protein expression of VEGF-C in the tumor tissues and lymph nodes resected during operation. The VEGF-C mRNA expression level in the lung tumor tissue of the lymph node metastasis positive group was 0.273+/-0.179, significantly higher than that in the lymph node metastasis negative group (0.089+/-0.087, P<0.01). The VEGF-C mRNA expression level in the positive lymph node of the lymph node metastasis positive group was 0.207+/-0.174, significantly higher than that in the lymph node metastasis negative group (0.114+/-0.107, P<0.01), but not significantly different from that in the negative lymph nodes of the same group(0.196+/-0.186, P>0.05). The VEGF-C protein positive rate of the lung cancer tissues of the lymph node metastasis positive group was 93.3% (14/15), significantly higher than that of the lymph node metastasis negative group (6.7%, 1/15, P<0.01). The VEGF-C protein positive rate of the metastasis positive lymph noses was 80.4% (37/46), and all 52 metastasis negative lymph nodes were VEGF-C protein negative. VEGF-C mRNA and protein expression levels predict lymph node metastasis and can be useful predictors of lymph node metastasis in NSCLC.

Artemisinin Inhibits Tumor Lymphangiogenesis by Suppression of Vascular Endothelial Growth Factor C

We have previously reported that dihydroartemisinin is found to have a potent ability in influencing lymphatic endothelial cell migration and tube formation. In this study, we investigated the effect of artemisinin on tumor growth, lymphangiogenesis, metastasis and survival in mouse Lewis lung carcinoma (LLC) models. We found that orally administered artemisinin inhibited lymph node and lung metastasis and prolonged survival without retarding tumor growth. Consistent with the decrease in lymph node metastasis, tumor lymphangiogenesis and expression of vascular endothelial growth factor C (VEGF-C) was significantly decreased in artemisinin-treated mice, as compared to control mice. Furthermore, IL-1β-induced p38 mitogen-activated protein kinase (MAPK) activation and upregulation of VEGF-C mRNA and protein in LLC cells was also suppressed by artemisinin or by the p38 MAPK inhibitor SB-203580, suggesting that p38 MAPK could serve as a mediator of proinflammatory cytokine-induced VEGF-C expression. These data indicate that artemisinin may be useful for the prevention of lymph node metastasis by downregulating VEGF-C and reducing tumor lymphangiogenesis.

Relationship between Protein Expression of VEGF-C, MMP-2 and Lymph Node Metastasis in Papillary Thyroid Cancer

Papillary thyroid cancer (PTC) is a common malignancy and has a good prognosis after appropriate treatment. PTC cells spread mainly by lymph node metastasis (LNM), but the mechanism is not well understood. Expression of vascular endothelial growth factor C (VEGF-C) and matrix metalloproteinase 2 (MMP-2) protein was studied immunohistochemically from the archived specimens of 65 PTC patients who initially presented without LNM. In this retrospective study, the frequency of expression differed significantly between thyroid cancer tissue and adjacent normal follicular epithelium (VEGF-C 78.5% and 20.0%, respectively; MMP-2 81.5% and 36.7%, respectively). LNM developed in 35 of the patients during 5 - 15 years of follow-up, by the end of which the frequencies of expression of VEGF-C and MMP-2 protein expression were 91.4% and 94.3%, respectively. Both VEGF-C and MMP-2 protein expression were significantly more frequent in PTC with LNM than without LNM. VEGF-C and MMP-2 protein expression levels were significantly correlated with LNM and it is, therefore, feasible that VEGF-C and MMP-2 may be useful as tumour markers of PTC with cervical LNM.