Research Paper Volume 2, Issue 11 pp 791—803

Figure 5. Inhibition of in vivo angiogenesis by hUCBSC. (A) Glioma cells, from single and co-cultures with hUCBSC, were suspended in serum-free medium and injected into the diffusion chambers. The chambers were sealed with bone wax and placed in serum-free medium to keep until used. Animals were anesthetized and 5mL of air was injected to create a dorsal skin sac. A 1.5-2 cm incision was made along the edge of the dorsal air sac. The chambers were then carefully placed through the incision under the skin, and the incision was closed. The animals were sacrificed after 2 weeks. The skin around the implanted chambers were carefully removed and observed under the phase contrast microscope. The novel (delicate, irregularly arising) branches were recorded as tumor-induced neovascularization. These branches were then compared to the more organized, pre-existing vasculature. PV=Pre-existing vasculature; TN=Tumor induced neovasculature. Bar = 50μm. (B) Immunofluorescence analysis for the downregulation of FAK. Orthotopic intracranial tumors were established in nude mice by injecting the glioma cells (U251 and 5310) and then treating with hUCBSC. The paraffin-embedded brain sections were subjected to immunofluorescence detection for (B) FAK and CD81 and (D) VEGF. FAK (anti-rabbit) is conjugated with goat anti-rabbit secondary antibody. CD81 (anti-goat) with FITC and VEGF (anti-mouse) was conjugated with Texas-red secondary antibodies. Bar=100μm. (C) DAB immunohistochemistry of CD31 expression in control and treated brain sections. Bar = 200μm.