ABSTRACT
The rat aortic ring model is well utilized for evaluation of angiogenesis. We report here an alternative assay employing an ex vivo mouse aorta angiogenesis model that can be extensively manipulated and serially evaluated using digital-assisted image analysis. Mouse aortas were harvested, cut into 2-mm disks, and cultured in fibrin matrix with growth media. Radial vascular outgrowths arose from the cut edge of the aortic disk and were digitally photographed and serially quantified. A variety of culture conditions were evaluated to determine their ability to alter angiogenesis in this model. Vessel outgrowth became apparent on day 3 and continued through day 10 with linear growth occurring between days 3 and 6. Increasing concentrations of serum from 0% to 40% resulted in stimulation of angiogenesis after day 3. Suramin and endostatin dramatically inhibited angiogenesis, which was more profound when applied at day 0 than when linear growth could be identified (day 3). Cells isolated from vessel outgrowths were predominantly endothelial in origin by immunocytochemistry and FACS analysis. We demonstrate that angiogenesis in an ex vivo murine model can be easily quantified using digital image analysis, responds appropriately to stimulation and inhibition, and exhibits differential results based on time of inhibitor administration. Antiangiogenic agents may be most effective if administered before development of accelerated vessel growth.
Subject(s)
Endothelium, Vascular/physiology , Fibrin/chemistry , Neovascularization, Pathologic , Neovascularization, Physiologic , Physiology/methods , Animals , Aorta/cytology , Aorta/metabolism , Aorta/pathology , Cell Proliferation , Cell Separation , Endostatins/metabolism , Endothelium, Vascular/pathology , Flow Cytometry , Image Processing, Computer-Assisted , Immunohistochemistry , Mice , Microcirculation , Platelet Endothelial Cell Adhesion Molecule-1/biosynthesis , Suramin/chemistry , Time FactorsABSTRACT
Increased expression of vascular endothelial growth factor (VEGF) by pancreatic cancer correlates with poor survival. The significance of VEGF in biliary and pancreatic secretions in periampullary cancers is unknown. Bile and pancreatic juice samples were collected from patients undergoing pancreaticoduodenectomy (PD). All samples were frozen at -70 degrees C until subsequent analysis for VEGF concentration using enzyme-linked immunoabsorbent assay (ELISA). Plasma VEGF levels in pancreatic cancer patients were <10 pg/mL. The biliary VEGF concentration for patients with malignancy was significantly elevated compared to benign disease (P = 0.05). There was no difference in pancreatic VEGF concentrations between benign and malignant disease. Cancer patients undergoing preoperative chemoradiation (CRT) had lower biliary and pancreatic VEGF concentrations than those who did not. Preoperative biliary drainage (BD) was associated with decreased VEGF concentrations in bile (3500 pg/mL vs 7740 pg/mL, P = 0.027). Patients undergoing both CRT and BD had diminished biliary and pancreatic VEGF concentrations compared to those who had neither. This was statistically significant for pancreatic VEGF concentrations (917 pg/mL vs 4723 pg/mL, P = 0.044). VEGF is highly concentrated in bile and pancreatic juice compared to plasma. Preoperative CRT and BD significantly reduce these levels in patients with periampullary cancers. Antiangiogenic therapy aimed at interrupting the VEGF pathway appears to be a logical target in periampullary cancer.
