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A whole cord model for the identification of mechanisms for the antivascular effects of DMXAA

Endothelial cells form the inner lining of a blood vessel and their structure and functional integrity are important in maintenance of the vessel wall and circulatory function. These cells play key roles in immune and inflammatory reactions by regulating lymphocyte and leukocyte movement into tissues; they are also main targets for antivascular agents in cancer therapy. Endothelial cell responses to different stimuli have been previously investigated using conventional approaches, 'HUVEC in vitro culture system'. In this study an ex vivo perfusion model was constructed and utilized using whole human umbilical cords, in attempts to replicate a more accurate in vivo microenvironment. Assessment of the proportion and duration of endothelial cell viability in the ex vivo model was undertaken using a MTT 3, (4,5-dimethylthiazol-2-yl) 2, 5-diphenyl-tetrazolium bromide viability assay. A time baseline was successfully established for all experimental perfusions. Endothelial cell immune response was assessed through intravenous perfusion of the endotoxin, Lipopolysaccharide (LPS). Gene expression profiles revealed a significant increase in expression levels of E-Selctin (E-Sel), Intracellular adhesion molecule (I-CAM) and Tissue Factor (TF) relative to the housekeeping gene Beta 2 Microglobulin. When LPS was administered in combination with Hypertonic Saline Solution (HSS), expression levels declined indicating HSS interferes with the activation pathway of LPS ultimately suppressing its effectiveness on endothelial cells. HSS impact was also recognized from perfusion experiments on resting endothelial cells. All identified genes were suppressed by HSS apart from inducible nitric oxide synthase (iNOS). As a potential target for antivascular agents, HUVEC were then stimulated with DMXAA and gene responses of Tumour Necrosis Factor-α (TNF-α) was analysed. DMXAA demonstrates excellent antivascular acivity in experimental tumours, so tumour conditioned media (TCM) was administered intravenously through umbilical cord segments to replicate a tumour microenvironment prior to DMXAA addition. When cells were stimulated with Tumour conditioned media then administered DMXAA, TNF-α expression was significantly upregulated; relative to the housekeeping gene Human Proteosome subunit Y, however, when the cells were exposed to tumour conditioned media in absence of DMXAA gene expression significantly decreased. Thus, the antitumour action of DMXAA is capable of inhibiting the gene response of TNF-α replicated in a tumour environment.
Type of thesis
Moses, K. M. (2007). A whole cord model for the identification of mechanisms for the antivascular effects of DMXAA (Thesis, Master of Science (MSc)). The University of Waikato, Hamilton, New Zealand. Retrieved from https://hdl.handle.net/10289/2488
The University of Waikato
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