Expression Of Vascular Endothelial Growth Factor In Tissues Research Articles

Estrogen-Related Receptor α Expression and Function Are Associated With Vascular Endothelial Growth Factor in Human Cervical Cancer

Estrogen-related receptor α (ERRα), one of orphan nuclear receptors with an unknown ligand, is expressed in various types of cancer. Increased ERRα levels are associated with a higher risk of recurrence and poor clinical outcome in breast cancer, suggesting that ERRα could be a negative prognostic factor. Recently, it has been suggested that vascular endothelial growth factor (VEGF) could be one of the transcriptional targets of ERRα in breast cancer. Here, we examined the expression of ERRα and the association of ERRα with VEGF in uterine cervical cancer cells and tissues. We evaluated the expression of ERRα and VEGF by immunohistologic analysis using specimens from 40 patients with invasive cervical cancer. We also evaluated the VEGF promoter activity of ERRα in cervical cancer cell lines by transfection and luciferase assay. We overexpressed or knocked down ERRα and examined VEGF expression by real-time polymerase chain reaction. Finally, cell proliferation assay was performed to examine whether ERRα affects tumor growth in cervical cancer. Immunohistologic analysis demonstrated that ERRα expression in cervical cancer tissues was higher than that in noncancerous tissues and that there was a positive association between ERRα and VEGF expression in cancer tissues (P < 0.05). We showed that ERRα stimulated the VEGF promoter activity in cervical cancer cell lines. We further showed the overexpression and knockdown of ERRα-regulated VEGF expression level by real-time polymerase chain reaction. Moreover, we showed that ERRα and VEGF knockdown by small interfering RNA or an inverse agonist of ERRα, XCT 790, could suppress cell growth compared with control cells in cervical cancer. We have provided compelling evidence that ERRα affects VEGF expression and tumor growth in cervical cancer. These results justify further investigation into the use of ERRα as a therapeutic target for patients with uterine cervical cancer.

Metronomic S-1 Chemotherapy and Vandetanib: An Efficacious and Nontoxic Treatment for Hepatocellular Carcinoma

Metronomic chemotherapy involves frequent, regular administration of cytotoxic drugs at nontoxic doses, usually without prolonged breaks. We investigated the therapeutic efficacies of metronomic S-1, an oral 5-fluorouracil prodrug, and vandetanib, an epidermal growth factor receptor and vascular endothelial growth factor (VEGF) receptor tyrosine kinase inhibitor, in models of hepatocellular carcinoma (HCC). We compared anti-HCC effects and toxicity in the six treatment groups: control (untreated), maximum tolerated dose (MTD) S-1, metronomic S-1, vandetanib, MTD S-1 with vandetanib, and metronomic S-1 with vandetanib. Tumor microvessel density (MVD) and tumor apoptosis were evaluated by immunohistochemistry. The expression of VEGF and thrombospondin-1, an endogenous inhibitor of angiogenesis, was analyzed by Western blot. Metronomic S-1 significantly inhibited tumor growth, which was enhanced by combination with vandetanib. With respect to toxicities, MTD S-1 caused severe body weight loss and myelosuppression, whereas metronomic S-1 did not cause any overt toxicities. Moreover, metronomic S-1 or metronomic S-1 with vandetanib prolonged survival, the latter treatment providing the greatest benefit. Metronomic S-1 and metronomic S-1 with vandetanib decreased MVDs and increased apoptosis in tumor tissues. The expression of VEGF in tumor tissues was upregulated by vandetanib and metronomic S-1 with vandetanib, whereas the expression of thrombospondin-1 was upregulated by metronomic S-1 and metronomic S-1 with vandetanib. Metronomic S-1 with an antiangiogenic agent seems to be an effective and safe therapeutic strategy for HCC.

Expression of carbonic anhydrase-IX in rectal cancer and its relationship with vascular endothelial growth factor and hepatic metastasis

Objective To explore the differential expression of carbonic anhydrases-Ⅸ ( CA-Ⅸ )in 128 patients with rectal cancer,and study its association with vascular endothelial growth factor (VEGF)and hepatic metastasis of rectal cancer.Methods Western blotting was used to examine the expression of CA-Ⅸ and VEGF in different tissues of 128 patients with rectal cancer and 30 cases of normal rectal mucosa and rectal adenoma,and the influence of anthdrases's inhibitor ( acetazolamide ) on the rectal cancer cell growth was studied.Results The positive expression rate of both CA-Ⅸ and VEGF in carcinoma tissues of patients with hepatic metastasis was 83.3%,which was higher than that in normal rectal tissue ( 16.7%,0% ) and rectal adenoma ( 16.7%,36.7% ) and carcinoma tissues of patients without hepatic metastasis (72.1%,65.4% ,P ＜0.01 ).The expression of VEGF and CA-Ⅸ in mesorectal tissues of the patients with hepatic metastasis was not higher than that of the patients without hepatic metastasis.There was a negative association between the expression of CA-Ⅸ with VEGF and hepatic metastasis of the patients with rectal cancer.Acetazolamide obviously enhanced the chemotaxymotor ability of fluorouracil to increase the apoptosis of recetal cancer cells.Conclusion The CA-Ⅸ is highly expressed in rectal cancer tissues and can promote the growth and proliferation of rectal cancer cells,but it cann't be used as an independent index to make the diagnosis of hepatic metastasis in rectal cancer patients. Key words: Rectal cancer; Hepatic metastasis; Carbonic anhydrase-Ⅸ; Vascular endothelial growth factor

Low temperature of radiofrequency ablation at the target sites can facilitate rapid progression of residual hepatic VX2 carcinoma

BackgroundRapid progression of residual tumor after radiofrequency ablation (RFA) of hepatocellular carcinoma has been observed increasingly. However, its underlying mechanisms remain to be clarified. The present study was designed to determine whether low temperature of RFA at the target sites facilitates rapid progression of residual hepatic VX2 carcinoma and to clarify the possible underlying mechanisms.MethodsThe residual VX2 hepatoma model in rabbits was established by using RFA at 55, 70 and 85°C. Rabbits that were implanted with VX2 hepatoma but did not receive RFA acted as a control group. The relationship between rapid progression of residual hepatic VX2 carcinoma and low temperature of RFA at the target sites was carefully evaluated. A number of potential contributing molecular factors, such as proliferating cell nuclear antigen (PCNA), matrix metalloproteinase 9 (MMP-9), vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF) and Interleukin-6 (IL-6) were measured.ResultsThe focal tumor volume and lung metastases of RFA-treated rabbits increased significantly compared with the control group (P < 0.05), and the greatest changes were seen in the 55°C group (P < 0.05). Expression of PCNA, MMP-9, VEGF, HGF and IL-6 in tumor tissues increased significantly in the RFA-treated groups compared with the control group, and of the increases were greatest in the 55°C group (P < 0.05). These results were consistent with gross pathological observation. Tumor re-inoculation experiments confirmed that low temperature of RFA at the target sites facilitated rapid progression of residual hepatic VX2 carcinoma.ConclusionsInsufficient RFA that is caused by low temperature at the target sites could be an important cause of rapid progression of residual hepatic VX2 carcinoma. Residual hepatic VX2 carcinoma could facilitate its rapid progression through inducing overexpression of several molecular factors, such as PCNA, MMP-9, VEGF, HGF and IL-6.

Relationship and prognostic significance of SPARC and VEGF protein expression in colon cancer

BackgroundSPARC (secreted protein, acidic and rich in cysteine) is closely related with the progress, invasion and metastasis of malignant tumor and angiogenesis.MethodsUsing human colon adenocarcinoma tissues (hereinafter referred to as colon cancer) and their corresponding non-diseased colon from 114 patients' biopsies, the expression of SPARC and vascular endothelial growth factor (VEGF) were investigated by immunohistochemistry staining to assessment the relationship between SPARC and VEGF, as well as their prognostic significance in patients. Evaluation of VEGF expression level with the same tissues was used to establish the antigenic profiles, and the marker of CD34 staining was used as an indicator of microvessel density (MVD).ResultsSPARC expression was mainly in the stromal cells surrounding the colon cancer, and was significant difference in those tissues with the lymph node metastasis and differentiation degree of tumor. Expression of SPARC was significantly correlated with the expression of VEGF and MVD in colon cancer tissues. Patients with low or absence expressing SPARC had significantly worse overall survival and disease-free survival in a Single Factor Analysis; Cox Regression Analysis, SPARC emerged as an overall survival and disease-free survival independent prognostic factor for colon cancer.ConclusionThe low expression or absence of stromal SPARC was an independent prognostic factor for poor prognosis of colon cancer. SPARC maybe involved in the regulation of anti-angiogenesis by which it may serve as a novel target for colon cancer treatment as well as a novel distinctive marker.

Relationships between levels of CXCR4 and VEGF and blood-borne metastasis and survival in patients with osteosarcoma

The present study was designed to determine whether the expression of chemokine receptor CXCR4 and vascular endothelial growth factor (VEGF) is correlated with the extent of metastasis and the survival of patients with osteosarcoma. Using tissue microarrays, we analyzed the expression of CXCR4 and VEGF in tumor tissues collected from 56 patients with osteosarcoma. A two-year follow-up was performed to evaluate tumor metastatic behavior and the overall survival of the patients. There was a significant correlation between the expression of CXCR4 and the expression of VEGF in tumor tissues of these patients (P = 0.002). Univariate analysis revealed that expression of these proteins was correlated with clinical stage, but not age, gender, or serum alkaline phosphatase levels. The patients with tumors expressing CXCR4 and VEGF had worse overall survival rates compared with the patients with tumors that did not express CXCR4 (P = 0.03) or VEGF (P = 0.04). These data indicate that CXCR4 and VEGF expression is highly correlated with metastatic progression in patients with osteosarcoma and had predictive value for the metastasis and survival of osteosarcoma patients.

Nerve Growth Factor and Vascular Endothelial Growth Factor: Retrospective Analysis of 63 Patients with Salivary Adenoid Cystic Carcinoma

AimTo detect the expression of nerve growth factor (NGF) and vascular endothelial growth factor (VEGF) in salivary adenoid cystic carcinoma (SACC) tissues, as well as to determine the correlation between growth factor expression and prognosis in SACC.MethodologyMedical records of 63 patients surgically treated for SACC between January 1988 and October 2005 were reviewed. Immunohistochemistry was performed to examine the expression of NGF and VEGF in tumor tissues. Kaplan-Meier analysis and Cox's proportional hazard regression model were applied to assess predictors of survival.ResultsNGF and VEGF were overexpressed in SACC tissues, compared with those in normal salivary tissues (P<0.05), and the staining intensity of these two factors was stronger in groups of solid subtype, advanced TNM stage, perineural invasion and recurrence. Patients with high-expression of NGF and VEGF, solid subtype, advanced stage, perineural invasion, recurrence and extended resection alone had worse survival rates (P<0.05).ConclusionNGF and VEGF are expressed increasingly in the tissues of SACC cases with invasion and metastasis. NGF expression and VEGF expression are independent prognosis factors for survival.

Expression of Efp, VEGF and bFGF in normal, hyperplastic and malignant endometrial tissue.

Vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) are two important angiogenic and prognostic factors for endometrial cancer. Estrogen is considered to be associated with increasing risk of endometrial adenocarcinoma. We investigated the expression of estrogen-responsive ring finger protein (Efp), VEGF and bFGF in endometrial cancer and correlated the results with clinicopathological features. We measured the expression of Efp, VEGF and bFGF in normal, hyperplastic and malignant endometrial tissues by quantitative RT-PCR, ELISA or Western blot analysis. The expression of Efp mRNA was significantly decreased in the groups of endometrial cancer (EC) in comparison with the groups of normal endometrium (NE) (P<0.05). Expression of VEGF mRNA in the groups of EC and atypical hyperplasia (AH) was significantly increased in comparison with the groups of NE (P<0.05). The expression of bFGF mRNA in groups of EC and AH was higher than that of NE groups (P<0.01 and P<0.05). There was positive relevance of bFGF expression and histologic grade and negative relevance of Efp expression and histologic grade. The expression of Efp, VEGF and bFGF protein was in agreement with the mRNA. The high expression of VEGF and bFGF indicate that both contribute in the angiogenesis of endometrial cancer. The regulation of Efp and bFGF expression during endometrial carcinogenesis suggests their potential utility as a prognostic biomarker.

Assessment of vascular endothelial growth factor and carbonic anhydrase-IX expression in rectal cancer

Objective To explore the expressions and role of carbonic anhydrases-Ⅸ (CA-Ⅸ) and vascular endothelial growth factor (VEGF) in rectal cancer. Methods The expression levels of CA-Ⅸ and VEGF in 104 pathologically diagnosed rectal cancer tissues, 30 normal tissues, and 30 cases of non-neoplastic rectal mucosa were detected by Western blotting. Results The overall positive expression rate of CA-Ⅸ and VEGF in rectal cancer was 72.1% and 65.4%, repeetively, which was higher than that in non-neoplastic tissue(P<0.05). The expression of CA-Ⅸ was related to Dukes' stage and status of lymph node metastasis. CA-Ⅸ expression positive rate was decreased with the progression of the Dukes' stage from A to C. CA-Ⅸ expression in patients with lymph node negative involvement was higher than that in lymph node involvement, 85.2% vs 58.8%, 56.3%, respectly, with the difference being statistically significant,P<0.05. The expression of VEGF was related to status of lymph node metastasis. VEGF expression in carcinoma tissues of patients with lymph node involvement (81.3%) was significandy higher than that in lymph node negative involvement (53.7%) ,X~2=6.88, P<0.05, and VEGF expression in mesorectal tissues with lymph node involvement (75.0%) was significantly higher than that in lymph node negative involvement (25.9%),X~2=14.52, P<0.05. There was no positive association between CA-Ⅸ and VEGF. Conclusion The hypoxia and angiogenesis are helpful to the judgment of prognosis. Key words: Rectal carcinoma; Hypoxia; Vascular endothelial growth factor; Carbonic anhydrase-9

Vascular Endothelial Growth Factor—A Positive and Negative Regulator of Tumor Growth

Over the past decade, the well-documented role of vascular endothelial growth factor (VEGF) in tumor angiogenesis has led it to become one of the leading therapeutic targets for the treatment of cancer. Emerging evidence from genetically modified animal models, however, suggests that elevated levels of VEGF, or a proangiogenic phenotype, may impede, rather than promote, early tumor development and progression. For example, hypermorph VEGF transgenic mice display delayed progression of a retroviral-induced murine leukemia, and knockdown of VEGF expression within the myeloid compartment accelerates tumor progression. Several mechanisms have been proposed to explain this paradox, whereby VEGF induces changes within the hematopoietic compartment and tumor microenvironment through recruitment of tumor inhibitory monocytic cells and the negative regulation of tumor angiogenesis. Thus, it is apparent that the levels of VEGF expression in both tumor and nontumor tissues, as well as the context and timing of its modulation relative to cancer induction, play an important role in determining the effects of VEGF expression on tumorigenicity. In light of these recent findings, the various mechanisms underlying the negative role of VEGF during early tumor development, progression, and metastasis will be discussed.

Constitutive activation of signal transducer and activator of transcription 3 regulates expression of vascular endothelial growth factor in human meningioma differentiation

PurposeJanus tyrosine kinases (JAKs) and signal transducer and activator of transcription factors (STATs), especially STAT3, are constitutively activated in human cancers. The function of STAT3 in the pathogenesis of meningioma remains unknown. In this study, we investigated the role of JAK1/STAT3 regulating vascular endothelial growth factor (VEGF) expression in the occurrence and progression of human meningioma.MethodsWe detected the expression of JAK1, p-JAK1, STAT3, p-STAT3, and VEGF in human meningioma and normal dura tissues by RT–PCR, Western blot analysis, and immunohistochemistry.ResultsJAK1, p-JAK1, STAT3, p-STAT3, and VEGF showed high expression in grade I and grade II meningioma. The level of STAT3 activation was associated with VEGF expression; all meningioma tumors that expressed p-STAT3 also expressed VEGF. Both frequency of positivity and expression were enhanced with increasing tumor grade; high frequencies and levels were found in grade II tumors, with no expression detected in normal dura tissues (P < 0.05).ConclusionsVEGF is directly regulated by constitutive STAT3 activity and associated with meningioma differentiation. STAT3 has an important role in the occurrence and development of human meningioma by regulating VEGF expression.

Effects of mild moxibustion on angiogenesis and microcirculation in wound repair after operation of anal fistula in rats

To observe the effects of mild moxibustion on angiogenesis and microcirculation in wound repair after operation of anal fistula, and to explore the mechanism of action. Seventy-two SD male rats were made to bear an "acute, open, bleeding, infected" wound surface and were divided randomly into mild moxibustion group, microwave group and control group (24 rats per group). The wound surfaces of the treatment groups were treated for 15 min with mild moxibustion or microwave medical instrument from the first day after operation, and the control group was not given any treatment. At the first, third, seventh and fourteenth days after operation (1, 3, 7 and 14 days of treatment), six rats in each group were sacrificed after examining the blood flow of the wound surface, and the samples of granulation tissues were collected. The numbers of CD34 and positive expression of vascular endothelial growth factor (VEGF) in granulation tissues were detected. In the early and middle stages of wound healing, mild moxibustion could obviously increase the expression of VEGF, the number of CD34, and the blood flow of the wound surface. And the expression of VEGF, the number of CD34, and the blood flow of the wound surface began to increase at the third day, and reached the peak level at the seventh day, and then began to decrease. In the late stage of wound healing (day 14), mild moxibustion could decrease the expression of VEGF, the number of CD34, and decrease the blood flow of the wound surface. There were significant differences between the mild moxibustion group and the control group in the blood flow of the wound surface, the expression of VEGF and the numbers of CD34 in granulation tissues (P<0.05) after treatment, and except the number of CD34 at 14-day treatment, there were also differences between the mild moxibustion group and the microwave group. Mild moxibustion has better regulative actions on blood flow of the wound surface and the number of CD34 and positive expression of VEGF in granulation tissues, and can regulate microcirculation in wound surface and promote the wound healing.

Expression of HIF-1alpha and VEGF in colorectal cancer: association with clinical outcomes and prognostic implications

BackgroundHypoxia-inducible factor 1 alpha (HIF-1α) and vascular endothelial growth factor (VEGF) are frequently overexpressed in numerous types of cancers and are known to be important regulators of angiogenesis. Until now, few studies have been carried out to investigate the prognostic role of these factors in solid tumors, especially in colorectal cancer (CRC). The purpose of this study was to evaluate the expression of HIF-1α and VEGF in CRC tissues, and to analyze the association of these two factors with several clinical and pathological characteristics, and patients' survival.MethodsParaffin-embedded tissue samples were retrospectively collected from 71 CRC patients, who received surgical resection between 2001 and 2002, with a median follow-up of 5 years. We examined the patterns of expression of HIF-1α and VEGF by immunohistochemistry method. Statistical analysis was performed with univariate tests and multivariate Cox proportional hazards model to evaluate the differences.ResultsExpression of HIF-1α and VEGF was positively observed in 54.93% and 56.34% among the patients, respectively. HIF-1α and VEGF status were significantly associated with tumor stage, lymph nodes and liver metastases (P < 0.05). Expression of both HIF-1α and VEGF remained significantly associated with overall survival (OS) (P < 0.01), and HIF-1α was positively correlative to VEGF in CRC (r = 0.72, P < 0.001).ConclusionsHIF-1α and VEGF could be used as biomarkers indicating tumors in advanced stage and independently implied poor prognosis in patients with CRC. Treatment that inhibits HIF-1α might be a promising targeted approach in CRC to exhibit its potential to improve outcomes in future perspective, just as VEGF targeting has proved to be.

Gene silencing of VEGF inhibits growth and angiogenesis of human glioma U251 implanted in nude mice

Objective To explore the inhibitory effect of silencing human vascular endothelial growth factor (VEGF) gene short hairpin RNA (shRNA) on the growth of human glioma in nude mice by RNA interference technique. Methods Human glioma U251 cells, cultured in vitro, were used to establish models of human glioma by subcutaneous injection into nude mice. The 30 mouse models were divided into 3 groups for varied purposes and treatment (n=10). The VEGF expression and microvessel density (MVD) in tumor tissues were measured by immunohistochemistry. Results The volume and inhibition rate of tumor in the therapy group (group A) was 654.0 mm~3 and 65.2% respectively,compared to 1790.4 mm~3,4.7% in the blank plasmid group (group B), and 1850.3 mm~3,0% in the PBS group (group C),with the difference being statistically significant (P<0.05). Moreover, VEGF and MVD in group A were 9.52, and 22.02,compared to 17.18, and 43.58 in group B, and 17.82, and 45.32 in group C,with the differ-ence being also statistically significant (P<0.05). There was a positive correlation between VEGF and MVD (r=0.721,P<0.01). Conclusion Silencing VEGF gene can decrease VEGF gene expressions,suppress tumor angiogenesis,and inhibit tumor growth in nude mice bearing human glioma cell hne U251. Key words: RNA interference; Vascular endothelial growth factor; Glioma; Gene therapy

Prognostic factors and recurrence of hepatitis B-related hepatocellular carcinoma after argon-helium cryoablation: a prospective study

To determine the long-term prognosis of hepatocellular carcinoma (HCC) after argon-helium cryoablation and identify the risk factors that predict metastasis and recurrence. A total of 156 patients with hepatitis B-related HCC less than 5 cm in diameter who underwent curative cryoablation were followed up prospectively for tumor metastasis and recurrence. Immunohistochemistry was used to analyze the expression of vascular endothelial growth factor (VEGF). HBV basal core promoter (BCP) and precore mutations were detected by DNA sequence analysis. Post-treatment prognostic factors influencing survival, tumor metastasis and recurrence were assessed by univariate and multivariate analyses. The variables included the expression of VEGF in HCC tissues, clinical and pathologic characteristics of patients, and HBV features (HBV DNA level, HBV genotype, BCP mutation). The median follow-up period of the 156 patients was 37 months (range 8-48 months). The 1-, 2-, and 3-year overall survival rates were 92, 82 and 64%, respectively. The 1-, 2-, and 3-year recurrence-free survival rates were 72, 56 and 43%, respectively. Eighty-five patients (54.5%) had tumor recurrence or metastasis. The multivariate analysis showed that Child-Pugh class and the expression of VEGF in HCC tissues could be used as independent prognostic factors for overall survival. Meanwhile, the expression of VEGF in HCC tissues and HBV BCP mutations were found to be independent prognostic factors for recurrence-free survival. Strong expression of VEGF in HCC tissues and HBV BCP mutations are important risk predictors for recurrence or metastasis of HCC smaller than 5 cm in diameter.

The molecular basis for impaired hypoxia-induced VEGF expression in diabetic tissues

Diabetes is associated with poor outcomes following acute vascular occlusive events. This results in part from a failure to form adequate compensatory microvasculature in response to ischemia. Since vascular endothelial growth factor (VEGF) is an essential mediator of neovascularization, we examined whether hypoxic up-regulation of VEGF was impaired in diabetes. Both fibroblasts isolated from type 2 diabetic patients, and normal fibroblasts exposed chronically to high glucose, were defective in their capacity to up-regulate VEGF in response to hypoxia. In vivo, diabetic animals demonstrated an impaired ability to increase VEGF production in response to soft tissue ischemia. This resulted from a high glucose-induced decrease in transactivation by the transcription factor hypoxia-inducible factor-1alpha (HIF-1alpha), which mediates hypoxia-stimulated VEGF expression. Decreased HIF-1alpha functional activity was specifically caused by impaired HIF-1alpha binding to the coactivator p300. We identify covalent modification of p300 by the dicarbonyl metabolite methylglyoxal as being responsible for this decreased association. Administration of deferoxamine abrogated methylglyoxal conjugation, normalizing both HIF-1alpha/p300 interaction and transactivation by HIF-1alpha. In diabetic mice, deferoxamine promoted neovascularization and enhanced wound healing. These findings define molecular defects that underlie impaired VEGF production in diabetic tissues and offer a promising direction for therapeutic intervention.

Treatment of hemangioma by transfection of antisense VEGF gene

The effect of transfection of antisense vascular endothelial growth factor (VEGF) gene on the growth of hemangioma was studied. A total of 49 cases of capillary hemangiomas of the skin were collected. Immunohistochemical method was used to detect the expression of PCNA in hemangioma tissues. According to the finding, 49 cases of hemangiomas fell into proliferating phase (27 cases) and involuting phase (22 cases) respectively. Another 5 cases of normal skin tissues adjacent to the tumor tissues served as control. Immunohistochemical staining was performed to detect the expression of VEGF in the tumor tissues and the normal tissues. The average absorbance (A) values and the average positive area rate of VEGF were measured by image analysis system (HPIAS-2000). Endothelial cells from the tumor tissues in proliferating phase were cultured. Eukaryotic expression vector was constructed by sub-cloning, and transfected into human hemangioma endothelial cells by using cation liposome as vector. The expression of VEGF mRNA and protein was detected by RT-PCR and indirect immunofluorescence assay (IFA), respectively, and the biological characteristics of the transfected endothelial cells were examined by MTT assay and flow cytometry (FCM) after transfection. Immunohistochemical results showed that the expression of VEGF in proliferating endothelial cells was remarkably higher than those in involuting endothelial cells and normal endothelial cells (P<0.01), but there was no significant difference in the expression of VEGF between involuting endothelial cells and normal ones (P>0.01). Electrophoresis and sequencing indicated that the eukaryotic expression vector containing antisense VEGF gene, i.e. pcDNA3.1-VEGF, was successfully constructed. After VEGF antisense RNA recombinant was transfected into hemangioma endothelial cells, RT-PCR revealed that the expression of VEGF mRNA in pcDNA-VEGF (V) group and blank group was obviously higher than that in pcDNA-VEGF (A) group, and that the expression of endogenous VEGF mRNA in pcDNA-VEGF (A) group was significantly inhibited. Immunohistochemical result demonstrated that, compared with blank group, there was statistically significant difference between pcDNA-VEGF (A) and pcDNA-VEGF (V) groups (P<0.01), but there was no significant difference between pcDNA-VEGF (V) group and blank group (P>0.05). The activity of endothelial cell proliferation was reduced significantly after transfection, and obvious apoptosis occurred in hemangioma endothelial cells after transfection of antisense VEGF. It was suggested that VEGF plays an important role in the pathological change of hemangiomas by promoting endothelial cell proliferation and angiogenesis. Antisense VEGF gene transfection could effectively inhibit the growth of hemanioma endothelial cells.

Tissue Factor and VEGF Expression in Prostate Carcinoma: A Tissue Microarray Study

Tissue factor (TF) is the principal physiologic initiator of coagulation. It also plays an important role in tumor growth and metastasis possibly by contributing to angiogenesis. We evaluated the expression of TF in benign and malignant prostate tissue and correlated it with the expression of the pro-angiogenic protein, vascular endothelial growth factor (VEGF). We used a tissue microarray (TMA) constructed from 80 archival prostatectomy specimens. Core samples were collected from benign prostate tissue (BP) (n= 77), high-grade prostatic intraepithelial neoplasia (PIN) (n= 26), and carcinoma (PCa) (n= 93). TMA sections were stained with an immunopurified polyclonal TF antibody and a rabbit polyclonal VEGF. Two pathologists manually scored staining in epithelial cells using the German Immunoreactive Score. Positive staining for TF was seen predominantly in PCa with rare positive glands in BP and PIN. TF expression was significantly lower in BP versus PCa specimens (p< .001) and in PIN versus PCa specimens (p< .001). Positive staining for VEGF was seen in PCa, BP, and PIN. Rates of VEGF expression were also significantly lower in BP versus PCa specimens (p= .003) but not in PCa versus PIN (p= .430). The majority of PCa samples positive for TF were also positive for VEGF (p< .001). Our findings reinforce the link between angiogenesis and TF expression in PCa. We suggest further exploration of TF-mediated pathways leading to increased tumor aggressiveness in PCa, and the possible use of anti-TF agents in PCa.

Effects of hyperthermia with dextran magnetic fluid on the growth of grafted H22 tumor in mice

Objective: To study the effect of dextran magnetic fluid hyperthermia on growth, vascular endothelial growth factor (VEGF) expression and apoptosis of the grafted H22 tumors in mice.Methods: Mice with grafted H22 tumor were randomly divided into four groups. The treatment group was intratumorally injected with dextran magnetic fluid and subsequently exposed to a 55 kHz 20 kA/m alternating magnetic field for 10 min. The three control groups were respectively injected with physiological saline and no magnetic field; with physiological saline and exposed to 55 kHz 20 kA/m alternating magnetic field for 10 min; and with dextran magnetic fluid only. The inhibitive ratios were calculated. Other mice with grafted tumor were randomly divided into two groups, respectively injected with dextran magnetic fluid or physiological saline and exposed to a 55 kHz 20 kA/m alternating magnetic field for 10 min. The VEGF expression and apoptosis in tumor tissues were analyzed at 1 h, 5 h, 24 h, 48 h and 336 h after treatment.Results: The inhibitive ratio of treated group was 59.74%, 52.51% and 40.35% respectively. Necrotic areas in tumor tissue were mainly distributing around the magnetic fluid and became larger with the lengthening of treatment. The VEGF expression and apoptosis of the treated group did not show obvious difference from that of the control group at different time points.Conclusion: Single time magnetic fluid hyperthermia significantly inhibited growth of grafted H22 tumor in mice, which mainly resulted from inducing necrosis while not inhibiting VEGF expression or inducing apoptosis.

Kidney-derived mesenchymal stem cells contribute to vasculogenesis, angiogenesis and endothelial repair

We isolated a clonal cell line (4E) from kidneys of mice expressing green fluorescent protein controlled by the endothelial-specific Tie2 promoter. When grown in a three-dimensional matrigel matrix they formed a fluorescent capillary network. In vivo angiogenesis assays using growth factor-depleted matrigel implanted plugs promoted a moderate angiogenesis of host endothelial cells. Using vascular endothelial growth factor (VEGF)-A and fibroblast growth factor-2 in the plugs containing 4E-cells resulted in a robust vasculogenesis. Transplantation of 4E cells into mice with acute renal ischemia showed selective engraftment in the ischemic kidney which promoted tubular regeneration by increasing epithelial proliferation and inhibiting apoptosis. This resulted in an accelerated functional recovery 3 days after ischemia. These mice showed a 5-fold increase in tissue VEGF expression compared to controls, but no difference in plasma VEGF level corresponding with better preservation of peritubular capillaries, perhaps due to a local paracrine effect following systemic 4E infusion. One month after ischemia, 9% of engrafted 4E cells expressed green fluorescent protein in the peritubular region while half of them expressed alpha-smooth muscle actin. Our study shows that kidney mesenchymal stem cells are capable of differentiation toward endothelial and smooth muscle cell lineages in vitro and in vivo, support new blood vessel formation in favorable conditions and promote functional recovery of an ischemic kidney.