Number Of Tumor Blood Vessels Research Articles

Effect of lincRNA-p21 targeting HIF-1α on biological functions of liver cancer cells.

The effects of long intergenic non-coding ribonucleic acid (lincRNA)-p21 targeting hypoxia-inducible factor-1α (HIF-1α) on proliferation, apoptosis and migration of liver cancer cells were investigated. MHCC97H liver cancer cells were infected with control lentivirus (control group) and lincRNA-p21 lentivirus (observation group), and control stable cell lines and lincRNA-p21 stable cell lines were screened and obtained by using puromycin. The expression levels of lincRNA-p21 messenger RNA (mRNA) in the control and observation groups were analyzed via reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Bioinformatics was used to search for the lincRNA-p21 target. The expression of target gene was analyzed via western blotting, and the proliferation, apoptosis, migration and in vivo tumor formation of MHCC97H cells in the control and observation groups were also analyzed. Compared with that in control group, the lincRNA-p21 mRNA level in observation group was increased significantly (P<0.05). It was found via bioinformatic comparison that HIF-1α was one of the targets of lincRNA-p21. Results of Western blotting revealed that the expression level of HIF-1α protein in cells in observation group was significantly downregulated (P<0.05). Besides, the level of vascular endothelial growth factor (VEGF) protein in cells in control group was obviously higher than that in observation group (P<0.05). Compared with those in control group, the cell proliferation and migration capacities in observation group were markedly reduced, but the apoptosis level was significantly increased (P<0.05). According to the in vivo tumor formation assay, the cell proliferation rate in control group was obviously higher than that in observation group (P<0.05). The number of tumor blood vessels in cells in control group was obviously reduced compared with that in observation group (P<0.05). lincRNA-p21 can significantly downregulate the level of HIF-1α, thus downregulating the expression of VEGF and affecting the cell proliferation, apoptosis and migration.

Abstract 732: The impact of aerobic exercise on the tumor microenvironment

Abstract Intratumoral hypoxia (pO2 &amp;lt; 10mmHg) is associated with aggressive tumor progression, radiation resistance, suppression of the immune anti-tumor response and poor patient outcome. It is often found in solid tumors, and can constitute a major obstacle to anticancer therapy. The primary cause of abnormal oxygen tensions in tumors is the aberrant tumor vasculature, characterized by tortuous, leaky and immature vessels. A number of intervention strategies designed to overcome tumor hypoxia and improve patient outcome have been investigated including high oxygen content breathing and the use of bioreductive prodrugs, but these have had only moderate success in the clinic. The goal of the present investigations was to determine whether aerobic exercise could be applied to improve tumor perfusion, oxygenation and the immune anti-tumor response in breast cancer and fibrosarcoma models. Such modulation of the tumor physiology and host environment would be expected to lead to enhanced antitumor efficacy when combined with radiotherapy or chemotherapy. The effects of mild and moderate intensity treadmill running were studied in mice bearing syngeneic murine mammary carcinomas (4T1 and EMT6) or fibrosarcomas (KHT). The exercise intensities were determined by measuring the anaerobic threshold, which was assessed by measuring the steady rise in blood lactate during an exercise bout. Mice were orthotopically injected with tumor cells and exercise commenced when tumors reached a size of ∼500 mm3 (single) or ∼200 mm3 (repeated bouts). Once size was attained, mice were exposed to either mild (12 m/min) or moderate (18 m/min) intensity exercise. Controls for each treatment consisted of sedentary mice exposed to a stationary treadmill for the equivalent amount of time. At the end of the exercise period tumors were analyzed by histology to assess for physiological changes. Specifically, blood was collected and tumors were harvested, sectioned and evaluated by immunofluorescence. The detection of open blood vessels (Hoechst-33342) was used as an indirect indicator of perfusion and while the hypoxia marker (EF5) was used to determine the level of tumor hypoxia. Both markers were quantified using a Chalkley counter and ImageJ NIH software. In addition, the evaluation of tumor infiltrating immune cells after exercise compared to rest is under active investigation by histological analysis. Furthermore, plasma samples from exercised mice are being tested for immune related cytokines, chemokines and growth factors and compared to those from sedentary controls. Results to date indicate no difference in tumor growth rate between sedentary and exercising mice. Mild daily bouts of aerobic exercise do not affect the total number of tumor blood vessels but do increase the number of blood vessels that are actively perfused. These findings suggest that exercise may have potential utility in overcoming the aberrant microenvironmental conditions associated in solid tumors with therapeutic resistance. Citation Format: Jennifer M. Wiggins, Jennifer Lee, Lori Rice, Sharon Lepler, Christine Pampo, Dietmar Siemann. The impact of aerobic exercise on the tumor microenvironment. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 732.

The imaging findings of CT and MRI in patients with abdominal and pelvic solitary fibrous tumor and its pathological features

Objective To investigate the imaging findings of CT and MRI in patients with abdominal and pelvic solitary fibrous tumor and its pathological features. Methods 30 patients with abdominal and pelvic solitary fibrous tumor were chosen. Patients with CT and MRI, and pathologic features control study were analyzed. Results In 30 patients, mesenchymal tumors,malignant fibrous histiocytoma and ovarian fibroma patients were 12 cases,12 cases,6 cases.CT showed that mass state clearly, smooth contours, the substance of the composition of two different densities, increase significantly enhanced scan, saw a number of tumor blood vessels within the shadow arterial aneurysm.MRI showed that internal tumors were low signal stripes appear. Tumor necrosis internal punctate liquefaction zone, after enhanced scan, the patient did not appear to strengthen.6 patients had resection specimens.4 cases had a more complete capsule, two cases with incomplete capsule. Conclusion SFF has certain characteristics in CT and MRI imaging screening, combined with its pathological features analysis will help improve the diagnosis. Key words: Solitary Fibrous Tumors; Fibromatosis, Abdominal; Tomography, X-ray Computed; Magnetic Resonance Imaging

Combined Tumor Cell-Based Vaccination and Interleukin-12 Gene Therapy Polarizes the Tumor Microenvironment in Mice.

Tumor progression depends on tumor milieu, which influences neovasculature formation and immunosuppression. Combining immunotherapy with antiangiogenic/antivascular therapy might be an effective therapeutic approach. The aim of our study was to elaborate an anticancer therapeutic strategy based on the induction of immune response which leads to polarization of tumor milieu. To achieve this, we developed a tumor cell-based vaccine. CAMEL peptide was used as a B16-F10 cell death-inducing agent. The lysates were used as a vaccine to immunize mice bearing B16-F10 melanoma tumors. To further improve the therapeutic effect of the vaccine, we combined it with interleukin (IL)-12 gene therapy. IL-12, a cytokine with antiangiogenic properties, activates nonspecific and specific immune responses. We observed that combined therapy is significantly more effective (as compared with monotherapies) in inhibiting tumor growth. Furthermore, the tested combination polarizes the tumor microenvironment, which results in a switch from a proangiogenic/immunosuppressive to an antiangiogenic/immunostimulatory one. The switch manifests itself as a decreased number of tumor blood vessels, increased levels of tumor-infiltrating CD4+, CD8+ and NK cells, as well as lower level of suppressor lymphocytes (Treg). Our results suggest that polarizing tumor milieu by such combined therapy does inhibit tumor growth and seems to be a promising therapeutic strategy.

Endoglin (CD105) Silencing Mediated by shRNA Under the Control of Endothelin-1 Promoter for Targeted Gene Therapy of Melanoma.

Endoglin (CD105), a transforming growth factor (TGF)-β coreceptor, and endothelin-1, a vasoconstrictor peptide, are both overexpressed in tumor endothelial and melanoma cells. Their targeting is therefore a promising therapeutic approach for melanoma tumors. The aim of our study was to construct a eukaryotic expression plasmid encoding the shRNA molecules against CD105 under the control of endothelin-1 promoter and to evaluate its therapeutic potential both in vitro in murine B16F10-luc melanoma and SVEC4-10 endothelial cells and in vivo in mice bearing highly metastatic B16F10-luc tumors. Plasmid encoding shRNA against CD105 under the control of the constitutive U6 promoter was used as a control. We demonstrated the antiproliferative and antiangiogenic effects of both plasmids in SVEC4-10 cells, as well as a moderate antitumor and pronounced antimetastatic effect in B16F10-luc tumors in vivo. Our results provide evidence that targeting melanoma with shRNA molecules against CD105 under the control of endothelin-1 promoter is a feasible and effective treatment, especially for the reduction of metastatic spread.

Combining virotherapy and angiotherapy for the treatment of breast cancer

A breast cancer-selective oncolytic adenovirus was engineered to express antagonists of vascular endothelial growth factor (VEGF) and Notch signaling to combine direct anticancer activity with disruption of tumor-associated angiogenesis. Replication of the parental virus, AdEHE2F, is stimulated by estrogen receptor (ER), E2F1 and hypoxia, and it mediates selective lysis of breast cancer cells in vitro and in vivo. Here, we encoded soluble Flt-1 (sFlt1) and soluble Dll4 (sDll4) under control of the E3 promoter. sFlt1 (the extra-cellular domain of VEGF receptor 1) binds VEGF-A and inhibits stimulation of VEGFR2, decreasing angiogenic stimulus. Conversely, sDll4 (the extracellular domain of Delta-like 4) antagonizes Notch signaling to prevent endothelial maturation. We hypothesized that these agents might show additive or synergistic activity. In vitro, sFlt1 inhibited endothelial cell proliferation and sprouting, whereas sDll4 increased the number of vascular branchpoints. In ER-positive ZR75.1 tumors in vivo AdEHE2F showed the potent direct virotherapy with no augmentation owing to sFlt1 or sDll4; however, in ER-negative MDA-231 tumors efficacy was enhanced by encoding sFlt1 or sDll4, with survival time extending to double that of controls. There was also a dramatic decrease in the total number of tumour blood vessels, as well as the number of perfused vessels, suggesting that improved efficacy reflects combined anti-tumour and anti-vascular effects.

Multiple Delivery of siRNA against Endoglin into Murine Mammary Adenocarcinoma Prevents Angiogenesis and Delays Tumor Growth

Endoglin is a transforming growth factor-β (TGF- β) co-receptor that participates in the activation of a signaling pathway that mediates endothelial cell proliferation and migration in angiogenic tumor vasculature. Therefore, silencing of endoglin expression is an attractive approach for antiangiogenic therapy of tumors. The aim of our study was to evaluate the therapeutic potential of small interfering RNA (siRNA) molecules against endoglin in vitro and in vivo. Therapeutic potential in vitro was assessed in human and murine endothelial cells (HMEC-1, 2H11) by determining endoglin expression level, cell proliferation and tube formation. In vivo, the therapeutic potential of siRNA molecules was evaluated in TS/A mammary adenocarcinoma growing in BALB/c mice. Results of our study showed that siRNA molecules against endoglin have a good antiangiogenic therapeutic potential in vitro, as expression of endoglin mRNA and protein levels in mouse and human microvascular endothelial cells after lipofection were efficiently reduced, which resulted in the inhibition of endothelial cell proliferation and tube formation. In vivo, silencing of endoglin with triple electrotransfer of siRNA molecules into TS/A mammary adenocarcinoma also significantly reduced the mRNA levels, number of tumor blood vessels and the growth of tumors. The obtained results demonstrate that silencing of endoglin is a promising antiangiogenic therapy of tumors that could not be used as single treatment, but as an adjunct to the established cytotoxic treatment approaches.

Preclinical evaluation of Sunitinib as a single agent in the prophylactic setting in a mouse model of bone metastases

BackgroundA substantial number of breast cancer patients are identified as being at high risk of developing metastatic disease. With increasing number of targeted therapeutics entering clinical trials, chronic administration of these agents may be a feasible approach for the prevention of metastases within this subgroup of patients. In this preclinical study we examined whether Sunitinib, a multi-tyrosine kinase inhibitor which has anti-angiogenic and anti-resorptive activity, is effective in the prevention of bone metastases.MethodSunitinib was administered daily with the first dose commencing prior to tumor cell inoculation. Intracardiac injection was performed with MDA-MB23 bone-seeking cells, which were stably transfected with DsRed2. In vivo plain radiography and fluorescent imaging (Berthold NightOwl) was used in the analysis of bone metastases. Histomorphometry was used for the quantification of TRAP+ cells from bone sections and immunohistochemistry was performed using an antibody reactive to CD34 for quantification of microvessel density.ResultsPreventive dosing administration of Sunitinib does not inhibit colonization of tumor cells to bone or reduce the size of osteolytic lesions. There was a decrease in the number of TRAP+ cells with Sunitinib treatment but this did not reach significance. Sunitinib inhibited tumor growth as determined by imaging of fluorescent tumor area. Immunohistochemical analyses of microvessel density revealed a concomitant decrease in the number of tumor blood vessels.ConclusionsThe findings suggest that Sunitinib can be used as a therapeutic agent for the treatment of bone metastases but as a single agent it is not effective in terms of prevention. Therefore a combination approach with other cytostatic drugs should be pursued.

Establishment and identification of an orthotopic transplanted model of gastric carcinoma with peritoneal carcinomatosis in nude mice

Objective To establish an orthotopic transplanted model of gastric cancer with peritoneal carcinomatosis in nude mice and identify the pathological features of the model.Methods The homogenate of MGC-803 gastric cancer tissue was planted in gastric antrum of 11 BALB/c nude mice to establish the model of gastric cancer with peritoneal carcinomatosis.The general status was recorded.The routine morphological examination and immunohistochemistry were performed to detect tumor biological features related to proliferation,apoptosis,invasion and metastasis [B lymphocytes/leukemia-2 (bcl-2),bcl-6,Ki-67,TdT-mediated dUTP nick end labeling (TUNEL),vascular endothelial growth factor (VEGF),matrix metalloproteinase (MMP)-9,CD34,and D2-40].Results The animal models were constructed successfully (11/11).The body weight of nude mice was slowly increased during the model development.The orthotopic tumor weight was (210.5-379.0) mg (median 267.5 mg).Cancer cell heterogeneity,and microvascular/lymphatic tumor thrombus were obviously observed as well as perigastric lymph nodes and liver metastases.Tumor proliferation-related molecule detection showed that bcl-2 positive rate was 0.0％-10.4％ (median 3.3％),bcl-6 positive rate was 0.0％-11.1％ (median 3.3％),Ki-67 positive rate was 40.5％-48.6％ (median 45.4％),and TUNEL positive rate was 0.2％-3.1％ (median 0.9％).The number of tumor blood vessels marked with CD34 was 1-11 (median 3)/400 × field of vision.The number of lymphatics marked with D2-40 was 0-5 (median 2)/400 × field of vision.VEGF positive rate was 68.5％-95.8％ (median 82.4％),and MMP-9 positive rate was 13.5％-50.0％ (median 23.5％).Conclusion The orthotopic transplanted model of gastric cancer with peritoneal carcinomatosis in nude mice was successfully established and reproduced the typical features of poorly differentiated adenocarcinoma. Key words: Gastric cancer; Peritoneal metastasis; Orthotopic transplantation; Model, animal

Analysis of ultrasonographic features and characteristics of acoustic contrast graphs of breast fibroadenoma and breast cancer

Objective To study the ultrasonographic and acoustic contrast features of breast fibroadenoma and breast cancer and explore the value of contrast enhanced ultrasonography in the differential diagnosis of benign and malignant breast tumors. Method A retrospective analysis of 18 cases breast fibroadenoma and 12 cases breast cancer in the performance of contrast-enhanced ultrasound was done. Results After contrast agent injection, tumor dimension increased in 4 cases with breast fibroadenoma and 7 cases with breast cancer. The ultrasonographic and acoustic contrast features of breast fibroadenoma showed no significant changes. They showed stenosis, perpendicular and circumambulates vascular shape. The breast cancers significantly increased the number of tumor blood vessels with the thicker,tortuous blood vessels into the tumor. Conclusions The contrast enhanced ultrasonography assessed more accurate size of tumors. The image displayed grown sizes, more vascularity and bending vascular shape in the breast cancer. It was more accurate than gray-scale ultrasound on diagnosing breast cancer. Key words: Breast Cancer; Breast fibroadenoma ultrasonography; Contrast agent

Combination of combretastatin A4 phosphate and doxorubicin-containing liposomes affects growth of B16-F10 tumors.

The study aimed to check the effectiveness of anticancer therapy combining a vascular-disruptive drug (combretastatin phosphate, CA4P) and a liposomal formulation of a chemotherapeutic (doxorubicin). CA4P was synthesized in our laboratory according to a previously described procedure. The antivascular drug and long-circulating doxorubicin-loaded liposomes were used to treat B16-F10 murine melanoma experimental tumors. Seventy-four hours after drug administration, a decrease in the number of tumor blood vessels was apparent and necrotic areas within tumors were visible. Combination therapy consisting of alternate administrations of CA4P and liposomal doxorubicin yielded greater inhibition of tumor growth than monotherapies alone. The best therapeutic results were obtained with the antivascular drug administered intratumorally every second day at 50 mg/kg body mass. In the case of combined therapy, the best results were obtained when the vascular-disruptive agent (CA4P) and the antineoplastic agent (liposomal doxorubicin) were administered in alternation.

Effects of triple therapy with octreotide, galanin and serotonin on a human colon cancer cell line implanted in mice: comparison between different routes of administration.

A human colon cancer cell line was implanted subcutaneously in nude mice. After 7 days, the animals were divided into four groups. The first group received an intraperitoneal (i.p.) continuous infusion by an osmotic pump, the second was given i.p. bolus injections, the third received continuous subcutaneous (s.c.) infusion by an osmotic pump and the fourth group was given bolus s.c. injections. Each group was divided into 2 subgroups. The first subgroup received triple treatment with octreotide, galanin, and serotonin, 40 microg/kg body weight/day of each. The second subgroup was given sterile saline solution. Treatment lasted for 14 days. The volume and wet weight of the tumours in all treated groups tended to decrease, but was statistically significant only in the group with continuous i.p. infusion. The number of viable cells tended to decrease in all the treated groups, but was not statistically significant. Proliferation index was significantly reduced in mice given triple therapy i.p. as bolus injection and as continuous infusion, as compared with their respective controls. The apoptotic index increased significantly in mice receiving triple therapy as continuous i.p. infusion as revealed by both the TUNEL method and by poly (ADP-ribose) polymerase (PARP) expression. The number of tumour blood vessels was significantly reduced in the mice given triple therapy as continuous i.p. infusion, as compared with controls. There was no statistical difference between animals treated by different routes, regarding proliferation or apoptosis of the cancer cells, or the number or mean luminal area of tumour blood vessels. The present investigation showed that regardless of the route of administration, triple therapy with octreotide, galanin and serotonin generally reduced the volumes, weights, viable cells, vascularization and proliferation of the tumours, as well as inducing apoptosis. Continuous i.p. infusion appears, however, to be the most effective route of administration.

Effects of triple treatment with octreotide, galanin and serotonin on a human pancreas cancer cell line in xenografts.

Human pancreas cancer cells were implanted s.c. in nude mice. After 11 days, the mice were divided into two groups of 13. The first group received sterile saline solution and the second received triple therapy containing octreotide, galanin and serotonin, 40 microg/kg/day as a continuous i.p. infusion via an implanted osmotic pump for 14 days. Triple therapy prolonged the survival rate of the mice bearing human pancreatic carcinoma. Both the volume and weight of tumours in mice given triple therapy were less than in controls (not statistically significant). The proliferation index and the labelling index for epidermal growth factor (EGF) increased significantly in mice given triple therapy vis-a-vis controls. There was no statistically significant difference between control and treated tumours as regards, apoptotic index, necrosis, or number of tumour blood vessels. The increased survival rate was attributed to the reduced tumour load, since both weight and volume were reduced. It is most probable that octreotide was the responsible agent. Further investigation with single and double combinations of octreotide, galanin and serotonin are needed to identify the cause of increased cell proliferation in tumours subjected to these bioactive substances. Identifying the agent(s) inducing pancreatic cancer cell proliferation may be useful in combining a new treatment, as antagonists to these bioactive substances are available.

045 Optical imaging of lung cancer using near-infrared fluorescence

Introduction We report here the in vitro and in vivo characteristics of a new molecular conjugate vector for targeting and imaging of tumors. Its core is a cyclo-decapeptide plateform named RAFT, onto which two spatially independent functional domains can be cova-lently and stereo-specifically linked : a cell targeting domain for tumor targeting and a labeling domain able to carry two drugs and/or labeling agent. In order to prove the interest of this carrier, we used a well known cRGD cyclopeptide, a ligand for the α v β 3 integrin. Methods Using competition assays and confocal microscopy in vitro , as well as in vivo tumor formation and near-infrared optical imaging in mice, we analyzed the characteristics of the RAFT(cRGD)4 vector. Results We demonstrate that this vector presenting 4 cRGD motifs very efficiently prevents (α v β 3 -mediated cell adhesion to vit-ronectin. Furthermore, it is actively endocytosed because of the multivalent cRGD presentation, a major advantage for drug delivery. In vivo experiments in nude mice reveal that repeated intratu-moral injections of low doses of RAFT(cRGD)4 reduce tumor growth by reducing the number of tumor blood vessels. Furthermore, RAFT(cRGD)4 significantly improves the targeting specifi-city of subcutaneous tumor masses as well as disseminated metastasis after intravenous injection. Conclusion Thus, RAFT (cRGD)4 is specific, internalized, perfectly controlled and can carry multiple biological functions on a single, spatially defined backbone, making of it a powerful and versatile synthetic vector for drug delivery, molecular imaging or both. This versatility is a major advantage since we can load it with multiple functions. This was demonstrated by the addition of a cytotoxic KL peptide on the cargo domain of the RAFT. RAFT-RGD-KL shows a very promissing specific antitumoral activity when injected intravenously in tumors bearing mice. Finally, recent developments in the optical System allowed us to follow these optical probes using a new 3D optical tomographic system in lung metastasis. This allowed us to detect lung metastasis in 3D for the first time to our knowledge using optical NIR probes.

Effects of triple therapy with octreotide, galanin and serotonin on a human colon cancer cell line

Human colon cancer cells were implanted subcutaneously into nude mice. After 12 days, the animals were divided into two groups. The first group received 40 microg/kg body weight of octreotide, galanin and serotonin via an intraperitoneally implanted pump. The second group received sterile saline only. Treatment lasted for 14 days. The volume and weight of the tumours in treated mice tended to decrease, though not with statistical significance. The proliferation index and the number of tumour blood vessels was significantly reduced in the mice given triple therapy. The apoptotic index, as detected by TUNEL method and monoclonal anti-poly (ADP-ribose) polymerase, was significantly higher in the treated mice. Though the results of this investigation are promising, it is uncertain as to what use the present findings may imply for the treatment of patients with colorectal cancer.

Effects of triple therapy with octreotide, galanin and serotonin on liver metastasis of human colon cancer in xenografts

Human colon cancer cells (SW 620) were implanted under the capsule of the left liver lobe of female nude (C57BL/6JBom-nu) mice. After 7 days, relaparatomy was performed and an ALZET osmotic pump was implanted intraperitoneally and left in situ for 14 days. The mice were divided into 2 groups, 10 in each. The first group received 40 micro g/kg body weight of octreotide, galanin and serotonin, and the second group received sterile saline. The number of metastases in the liver, and to the intra-abdominal lymph nodes was significantly greater in the controls. The incidence of metastases to the peritoneal cavity was lower in the treated animals (though not statistically significantly). Tumour volume, wet weight, proliferation index and number of tumour blood vessels decreased significantly in the treated animals. The apoptotic index was significantly higher in the treated mice. The decrease in the volume and weight of tumours following the triple therapy seemed to be caused by low proliferation, and increased apoptosis, and reduced vascularization of the tumours. The low invasion of cancer cells observed following this treatment could have been due to the low tumour burden, and to the reduced number of the blood and lymph vessels.

Tumor angiogenesis in pheochromocytomas and paragangliomas

Angiogenesis correlates with growth and likely metastases in several tumors. To determine whether it has a similar role in pheochromocytomas, immunohistochemical staining of factor VIII was done on the tumor tissue of 42 patients. Formalin-fixed, paraffin-embedded tissue was obtained from 29 women and 13 men with 24 primary adrenal and 18 extraadrenal pheochromocytomas. Patients were divided into two groups. Group 1 included 32 patients with benign pheochromocytomas, and group 2 included 10 patients with malignant tumors evidenced by capsular or vascular invasion (six), liver metastases (three), or periaortic lymph node metastases (one). Blood vessels highlighted by factor VIII staining of endothelial cells with labeled streptavidin-biotin were counted under light microscopy. Mean vessel count within a 10 mm2 micrometer disk was calculated under x100, x200, and x400 magnification fields. There were no significant differences in patient age or clinical symptoms between the groups. The mean tumor size in group 2 of 8.8 +/- 5.3 cm was larger than the mean of 4.8 +/- 2.8 cm in group 1 (p < 0.005). The mean counts of vessels in the x100, x200, and x400 magnification fields were 102 +/- 48, 40 +/- 18, and 19 +/- 9 in group 1, and 203 +/- 77, 73 +/- 28, and 37 +/- 15 in group 2. The number of blood vessels in group 2 was significantly higher than in group 1 (p < 0.001) in each studied field. In this study the number of tumor blood vessels correlated with the invasive behavior of pheochromocytomas. Tumor angiogenesis may be useful in determining the likelihood of malignant behavior in pheochromocytomas.