Predictive value of vascular endothelial growth factor (VEGF) in metastasis and prognosis of human colorectal cancer

Vascular endothelial growth factor (VEGF) may affect the phenotype of cancer cells, such as growth velocity and metastatic potential, due to its probable multifunctional property including a mitogenic activity for vascular endothelial cells. The present study was designed to investigate the association of VEGF mRNA expression with progression and metastasis of human colorectal cancer. The level of VEGF mRNA expression was quantified by Northern blot hybridization in tumorous and non-tumorous tissues obtained from 60 primary colorectal cancer patients. The ratio of the former to the latter was defined as the VEGF T/N ratio, and the prognostic significance of this ratio, following surgery, in addition to the relationship to progression and metastatic potential, was evaluated. The value of the VEGF T/N ratio was significantly correlated with the depth of tumour infiltration (P=0.046), the incidence of liver metastasis (P < 0.0001) and lymph node metastasis (P=0.036). Patient prognosis was estimated by the Kaplan-Meier method and the log-rank test. When the VEGF T/N ratio was higher than 4.8 for which the chi2 value of the log-rank test was maximal, the tumour was defined as showing overexpression of VEGF mRNA. Patients with overexpression of VEGF mRNA demonstrated poorer survival than patients without overexpression of VEGF mRNA (P < 0.001). The overall estimated hazard ratio for death in patients with overexpression of VEGF mRNA was 1.94 according to a multivariate analysis (P=0.005). Thus, VEGF is associated with the progression, invasion and metastasis of colorectal cancer, and overexpression of VEGF mRNA in the primary tumour is assumed to be closely correlated with poor prognosis in colorectal cancer patients. Moreover, the VEGF T/N ratio may be used as an independent prognostic marker in colorectal cancer patients.     

Circulating platelet-derived endothelial cell growth factor increases in hepatocellular carcinoma patients

BACKGROUND: Platelet-derived endothelial cell growth factor (PD-ECGF) is an angiogenic factor that is expressed in various cancer tissues. Little is known regarding plasma PD-ECGF levels in patients with chronic liver disease such as chronic hepatitis (CH), cirrhosis, and hepatocellular carcinoma (HCC) with cirrhosis. The expression of PD-ECGF in HCC tissues also remains to be clarified. METHODS: Plasma PD-ECGF levels in patients with chronic liver disease were determined with an enzyme-linked immunoadsorbent assay system using the mouse monoclonal antibodies specific to PD-ECGF. These were cross-sectionally compared among groups of normal persons, CH, cirrhosis, and HCC patients. The HCC patients were classified into two groups based on TNM stage: early and advanced stage disease groups. PD-ECGF expressions in HCC tissues were immunohistologically examined. RESULTS: The plasma PD-ECGF levels from the normal individuals and those with CH, cirrhosis, and HCC specimens were 4.2+/-0.5, 4.3+/-0.6, 4.6+/-1.1, and 6.0 +/-2.5 U/mL, respectively. The plasma PD-ECGF concentration was highest in HCC (P < 0.05). No significant difference was found among the normal subjects, CH, and cirrhosis specimens. Plasma PD-ECGF concentrations were significantly higher in the advanced stage disease HCC group compared with the early stage disease group (6.75+/-2.62 U/mL vs. 4.19+/-0.34 U/mL) (P < 0.05). Immunohistochemical expression of PD-ECGF in HCC cells increased significantly compared with normal liver cells (P < 0.05). CONCLUSIONS: Circulating PD-ECGF plasma level might be a new tumor marker for progression in patients with HCC. Immunohistological findings correspond to elevation of the plasma PD-ECGF in HCC patients. It is possible that increased production of PD-ECGF in HCC cells causes abundant neovascularization.     

Expression of vascular endothelial growth factor (VEGF) in various compartments of the human hair follicle

Hair follicle vascularization appears to be closely related to the processes involved in hair cycle regulation, in which growth factors, cytokines and other bioactive molecules are involved. In particular, vascular endothelial growth factor (VEGF), essential for angiogenesis and vascular permeability, may be responsible for maintaining proper vasculature around the hair follicle during the anagen growth phase. The aim of our study was to compare the in vitro angiogenic capacity, i.e. the steady-state expression of the VEGF gene, of different cultured cell types derived from normal human hair follicles, corresponding to different follicular compartments. Human dermal papilla cells (DPC), fibrous sheath fibroblasts, dermal fibroblasts, and follicular and interfollicular keratinocytes were cultured and studied in vitro for VEGF expression at the mRNA level using RT-PCR, and for VEGF protein synthesis by radioimmunoprecipitation and immunocytochemistry. In vivo examination for VEGF expression in human terminal hair follicles was performed using immunohistochemical methods. In the present report the expression of four different VEGF molecular isoforms, differing in their angiogenic capacity, are described in different cultured follicular cell types for the first time. Cultured follicular cells strongly expressed mRNA of four VEGF molecular species identified as the 121-, 145-, 165- and 189-amino acid splice variants, the most prominent being the 121-amino acid molecule. DPC, and also other mesenchymal cells such as fibrous sheath fibroblasts and dermal fibroblasts, in vivo and in vitro strongly expressed VEGF mRNA and synthesized a 46-kDa VEGF protein, whereas follicular and interfollicular keratinocytes in vitro expressed lower levels of VEGF mRNA and proteins than mesenchymal cells. As the highest expression of VEGF was found in DPC, we suggest that DPC are mainly responsible for angiogenic processes possibly related to the human hair cycle.     

Vascular endothelial growth factor (VEGF) in endometriosis

Angiogenesis is likely to be involved in the pathogenesis of endometriosis. According to the transplantation theory, when the exfoliated endometrium is attached to the peritoneal layer, the establishment of a new blood supply is essential for the survival of the endometrial implant and development of endometriosis. From the known angiogenic factors, vascular endothelial growth factor (VEGF) has emerged as a pivotally important regulator of normal angiogenesis and pathological neovascularization. The VEGF protein was evaluated immunohistochemically in the eutopic endometrium of 10 women without endometriosis (group I) at laparoscopy and the eutopic endometrium and peritoneal endometriotic lesions of 43 women with endometriosis (group II). VEGF histological scores were 9.7 +/- 4.3 and 4.0 +/- 2.6 respectively in the epithelium and stroma of the eutopic endometrium of group I women, and 10.3 +/- 2.3 and 3.6 +/- 2.3 respectively in women of group II. In red lesions, the VEGF scores were 11.1 +/- 3.0 in the epithelium and 5.1 +/- 3.0 in the stroma, and in black lesions were 8.6 +/- 2.7 and 1.6 +/- 1.6, respectively. Significantly lower values were observed in black lesions as compared with eutopic endometrium and red lesions, the values of which were similar. Scores were also evaluated according to the phase of the cycle. In eutopic as well as ectopic endometrium, no significant cyclic variations were observed throughout the cycle. However, VEGF content was found to be higher in the eutopic glandular epithelium of women with endometriosis during the late secretory phase, possibly suggesting a more likely tendency to implant. In contrast, significantly higher VEGF content was noted in red lesions as compared with black lesions. During all phases of the cycle, the VEGF content in stromal cells of red lesions was higher than in black lesions. Similarities in VEGF content were observed in the glandular epithelium of the eutopic endometrium of women with endometriosis and red lesions, suggesting that endometriosis probably arises from the peritoneal seeding of viable endometrial cells during retrograde menstruation and that red lesions can be considered as the first stage of implantation. After the attachment phase, the high VEGF levels could provoke an increase in the subperitoneal vascular network and facilitate implantation and viability in the retroperitoneal space. Lower VEGF levels in black lesions explain the decrease in both stromal vascularization, followed by fibrosis and inactivation of the implant.     

Increased vascular endothelial growth factor (VEGF) and transforming growth factor beta (TGFbeta) in experimental autoimmune uveoretinitis: upregulation of VEGF without neovascularization

A combined two-step high-performance liquid chromatographic (HPLC) method was developed for the analysis of endogenous levels of cyclic adenosine diphosphoribose (cADPR) in cell extracts. The detection sensitivity for cADPR was about 10 pmol. Linearity of the HPLC detection system was demonstrated in the range of 10 pmol up to 2 nmol. The method was validated in terms of within-day and between-day reproducibility of retention times and peak areas of standard nucleotides. The method was applied to the analysis of endogenous cADPR in human T cell lines. Sequential separation of perchloric acid extracts from cells on strong anion-exchange and reversed-phase ion-pair HPLC resulted in a single symmetrical peak co-eluting with standard cADPR. The identity of this endogenous material was further confirmed by its ability to be converted to ADPR upon heating the cell samples at 80 degrees C for 2 h. Recoveries of the combined perchloric acid extraction-HPLC analysis procedures were 48.3 +/- 10.2%. The determined intracellular concentrations of cADPR in quiescent Jurkat and HPB. ALL human T cells were 198 +/- 41 and 28 +/- 9 pmol/10(8) cells, respectively. In conclusion, a non-radioactive HPLC method presenting a specificity and sensitivity suitable for precise quantification of cADPR in cell extracts was developed.     

Circulating vascular endothelial growth factor in patients with colorectal cancer

BACKGROUND: Venom immunotherapy (VIT) has proven to be safe and effective in wasp venom anaphylaxis. However, there are no good parameters to indicate when to stop venom immunotherapy. OBJECTIVE: To evaluate the relationship of the lymphocyte transformation test (LTT) to history and specific IgE determination, and to address the time course of lymphocyte transformation responses to wasp (Vespula) venom during VIT and the possible utility of LTT to determine the duration of therapy. METHODS: Peripheral blood mononuclear cells (PBMCs) of 18 individuals with a history of wasp sting anaphylaxis and a positive serum-venom-specific IgE, were stimulated with wasp venom before immunotherapy, at the end of a 5-day semi-rush immunotherapy and at 24 months during venom immunotherapy. Results, expressed as stimulation index (SI), were compared with the SI in seven asymptomatic stung controls. RESULTS: In controls the median (minimum-maximum) of the SI were 2.39 (0.52-3.39) before therapy and 2.39 (1.12-6.02) when repeated after 24 months. For patients the median (minimum-maximum) of the SI were 10.13 (1.19-44.88) before immunotherapy (d0), 2.73 (0.67-12.03) at the end of the build-up immunotherapy (d5) and 4.21 (0.88-14.66) at the end of 24 months of maintenance therapy (m24). The proliferation responses in vespid-allergic patients were significantly higher than in stung controls (P = 0.006) but only 13/18 patients showed a positive LTT result before the start of immunotherapy (sensitivity of the LTT 72%). When the LTT was repeated after a 5 day build-up hyposensitization course the SI significantly dropped as compared to the pre-treatment levels (P = 0.002). The SI of the LTT was negative in eight out of 18 patients at 24 months and the median values were significantly lower than before therapy (P = 0.03). CONCLUSIONS: Although, in the absence of sting challenge data it is not possible to draw conclusions about the predictive value of the LTT, our data may suggest that abolition of the LTT during VIT might indicate clinical insensitivity. Further studies, comparing the results of sting challenges, with the results of lymphocyte transformation will be necessary in order to evaluate the role of LTT in stopping immunotherapy.     

Overproduction of vascular endothelial growth factor/vascular permeability factor is causative in Crow-Fukase (POEMS) syndrome

Crow-Fukase or POEMS syndrome of polyneuropathy, organomegaly, endocrinopathy, M-protein, and skin changes is a rare multisystem disorder of obscure pathogenesis that is associated with microangiopathy, neovascularization, and accelerated vasopermeability. We examined the levels of the vascular endothelial growth factor/vascular permeability factor (VEGF) in the serum and cerebrospinal fluid (CSF) from 10 patients with this syndrome. Serum VEGF levels were about 15-30 times those in control subjects or patients with Guillain-Barré syndrome (GBS), chronic inflammatory demyelinating polyneuropathy (CIDP), and other neurological disorders. The CSF VEGF levels, however, were similar to those found in GBS and CIDP. Elevated VEGF levels in the serum decreased in 7 patients with Crow-Fukase syndrome after conventional therapy. The principal isoform of VEGF in Crow-Fukase syndrome was VEGF165. Elevated VEGF was independent of M-protein. Our results suggest that the overproduction of VEGF is important in the pathogenesis of this disorder.     

Pulsatile stretch stimulates vascular endothelial growth factor (VEGF) secretion by cultured rat cardiac myocytes

Evidence has accumulated that vascular endothelial growth factor (VEGF) is expressed in the heart, and its expression is markedly increased in response to hypoxia. Recently, it was shown that pulsatile myocardial stretch in vivo markedly enhanced VEGF mRNA level in the heart. To investigate whether pulsatile mechanical stretch really stimulates VEGF expression by cardiac myocytes, using an in vitro preparation, we examined the secretion of VEGF into the culture media from cardiac myocytes subjected to pulsatile stretch. We found that pulsatile mechanical stretch induced rapid secretion of VEGF by cultured rat cardiac myocytes and mRNA expression of VEGF and VEGF receptors in the cardiac myocytes. We also found that the stretch-induced secretion of VEGF was at least in part mediated by TGF-beta. These data provide the direct evidence that mechanical overload itself can induce VEGF secretion by cardiac myocytes, which may play a role in ameliorating the relative myocardial hypoxia.     

Sinusoidal capillarization of human hepatocellular carcinoma: possible promotion by fibroblast growth factor

Expression of acidic and basic fibroblast growth factors (aFGF and bFGF) and von Willebrand factor (vWF) was immunohistochemically investigated in 55 nodules of human hepatocellular carcinoma (HCC), 15 nodules of adenomatous hyperplasia (AH) of the liver and 10 cirrhotic livers (LC). AH, a putative preneoplastic lesion in the cirrhotic liver, was subdivided into ordinary and atypical types: the former was characterized by little cellular and structural atypia and the latter had some atypia equivocal as to benignity and malignancy. The positive rates of FGF (aFGF and/or bFGF) were as follows: 0% in LC, 20% in AH (ordinary and atypical) and 42% in HCC, whereas the positivity of vWF was 10% in LC, 20% in ordinary AH, 30% in atypical AH and 40% in HCC. There was no correlation between the expression of FGF or vWF and the size of HCC. No correlation was also found between the positivity of FGF and that of vWF in HCC and atypical AH. While vWF was not constantly expressed in the vicinity of FGF-positive HCC cells, capillarized sinusoids were significantly more numerous in FGF-positive cases than in FGF-negative cases (p < 0.01). These data indicate that FGF may be pathogenetically linked to the multistep development of HCC in relation to sinusoidal capillarization.     

Paracrine expression of a native soluble vascular endothelial growth factor receptor inhibits tumor growth, metastasis, and mortality rate

Vascular endothelial growth factor (VEGF) is a potent and selective vascular endothelial cell mitogen and angiogenic factor. VEGF expression is elevated in a wide variety of solid tumors and is thought to support their growth by enhancing tumor neovascularization. To block VEGF-dependent angiogenesis, tumor cells were transfected with cDNA encoding the native soluble FLT-1 (sFLT-1) truncated VEGF receptor which can function both by sequestering VEGF and, in a dominant negative fashion, by forming inactive heterodimers with membrane-spanning VEGF receptors. Transient transfection of HT-1080 human fibrosarcoma cells with a gene encoding sFLT-1 significantly inhibited their implantation and growth in the lungs of nude mice following i.v. injection and their growth as nodules from cells injected s.c. High sFLT-1 expressing stably transfected HT-1080 clones grew even slower as s.c. tumors. Finally, survival was significantly prolonged in mice injected intracranially with human glioblastoma cells stably transfected with the sflt-1 gene. The ability of sFLT-1 protein to inhibit tumor growth is presumably attributable to its paracrine inhibition of tumor angiogenesis in vivo, since it did not affect tumor cell mitogenesis in vitro. These results not only support VEGF receptors as antiangiogenic targets but also demonstrate that sflt-1 gene therapy might be a feasible approach for inhibiting tumor angiogenesis and growth.     

Expression of vascular endothelial growth factor in normal liver and hepatocellular carcinoma: an immunohistochemical study

Angiogenesis is of vital importance during the development and progression of solid tumors. To examine the role of vascular endothelial growth factor (VEGF) in hepatocarcinogenesis, we evaluated the expression of peptide in normal human liver (n = 6) and in 36 cases of hepatocellular carcinoma (HCC). Immunoreactivity for VEGF was present in the extracellular matrix of the portal tracts in the normal and nontumor part of liver, but not in hepatocytes and bile duct epithelium. For HCC, variable amounts of VEGF were expressed in 13 cases (36.1%) of tumor cells. Using a logistic regression model, expression of VEGF was significantly associated with a higher proliferative index (P = .01) and sonographic portal vein thrombosis (P = .05). However, VEGF expression did not correlate with a biochemical liver profile, alpha-fetoprotein levels, histological grading, gender, or clinical stage of cirrhosis (P > 0.1, respectively). Log-rank test showed that evaluation of VEGF did not provide more prognostic information (P > .5) than that from tumor volume and portal vein thrombosis (P < .01, respectively). In addition, VEGF was always present in the fibrovascular stroma or pericellular matrix of HCC, although no strong relationship was observed with the expression of VEGF in tumor cells (P > .5). Our data suggested that expression of VEGF may characterize a progression toward higher proliferation in hepatocarcinogenesis in vivo. The relevance of VEGF existing in the extracellular matrix of the normal liver and HCC remains to be clarified.     

Serum levels of vascular endothelial growth factor (VEGF) are markedly elevated in patients with Wegener's granulomatosis

OBJECTIVES: Necrotizing vasculitis and granuloma formation are the predominant features of Wegener's granulomatosis (WG). We have investigated the importance of vascular endothelial growth factor (VEGF) in monitoring disease activity in WG. METHODS: Serum VEGF levels were determined in 23 patients with active WG, 21 healthy controls and 25 patients with urinary infection, by ELISA using commercially available antibodies to VEGF. RESULTS: VEGF levels were enormously elevated in patients with WG compared to both controls and patients with urinary infection (P < 0.0001). Of the 23 patients, 21 (91.3%) had VEGF levels above the cut-off value (3.3 ng/ml, calculated as the mean of the controls + 2 S.D.). Further analysis of the data showed that VEGF levels did not correlate with age, sex, incidence of classic antineutrophil cytoplasmic antibodies (c-ANCA) or duration of the disease (P > 0.05), but there was correlation with disease activity (r = 0.51, P < 0.01). VEGF levels were higher in patients with major compared to those with minor disease activity (P < 0.01). However, there was no significant correlation between VEGF levels and the Birmingham scores for vascular activity and damage. CONCLUSION: VEGF levels are raised in WG patients compared to normal controls and may be a marker of disease activity. Further studies on serial blood samples from a large cohort of patients with WG and other systemic vasculitides are needed to evaluate the specificity and usefulness of VEGF levels in monitoring disease activity.     

Hypoxia and vascular endothelial growth factor expression in human squamous cell carcinomas using pimonidazole as a hypoxia marker

Hypoxia in human tumors is associated with poor prognosis, but the molecular mechanisms underlying this association are poorly understood. One possibility is that hypoxia is linked to malignant progression through vascular endothelial growth factor (VEGF) induction and the associated angiogenesis and metastasis. The present clinical study measures hypoxia and VEGF expression on a cell-by-cell basis in human squamous cell carcinomas to test the hypothesis that hypoxia and VEGF protein expression are coupled in human tumors. Eighteen patients with invasive squamous cell carcinoma of the uterine cervix and head and neck have been investigated by a quantitative image analysis of immunostained sections from their tumors. The hypoxia marker pimonidazole was used to measure tumor hypoxia, and a commercially available antibody was used to measure VEGF protein expression. A quantitative immunohistochemical comparison of hypoxia and VEGF protein expression revealed no correlation between the two factors.     

Concentration of vascular endothelial growth factor (VEGF) in the serum of patients with suspected ovarian cancer

As a promoter of angiogenesis, vascular endothelial growth factor (VEGF) is believed to play a pivotal role in tumour growth and metastasis. The aim of this study was to determine the value of preoperative serum VEGF levels in the early diagnosis of ovarian cancer and in the differential diagnosis of adnexal masses. We examined preoperative serum VEGF levels in healthy women (n = 131), patients with benign ovarian cysts (n = 81) and in ovarian cancer patients (n = 44) by using an ELISA (R&D Systems, Minneapolis, MN, USA). A logistic regression model was carried out to determine the influence of VEGF and CA 125 on the probability of malignancy. VEGF revealed a significant influence on the odds of presenting with malignancy vs healthy women (P = 0.001). At 363.7 pg ml(-1), VEGF achieved a sensitivity of 54% and a specificity of 77%. With respect to the differentiation between benign cysts and ovarian cancer, CA 125 (P < 0.0001) but not VEGF (P = 0.229) predicts the presence of malignancy in a multivariate model. In conclusion, VEGF does not appear to be a useful tool in the early diagnosis of ovarian cancer or for indicating the absence or presence of malignancy in patients with an adnexal mass.     

Regulation of vascular endothelial growth factor (VEGF) expression is mediated by internal initiation of translation and alternative initiation of transcription

The growing amount of high quality molecular dynamics simulations generated using the latest methodological developments and force fields has led to a sharper understanding of the forces underlying the dynamics of biomolecular systems, as well as to stimulating insights into the structure and catalysis of nucleic acids. It is now clear that inclusion of long-range electrostatic interactions and of the aqueous and ionic environment is necessary for producing realistic and accurate simulations. Yet, many papers hint at a force field and protocol dependence of the results and thus contain the seeds for the future improvements that will be necessary for deepening our understanding of recognition phenomena and folding of nucleic acids.