Hypoxia-inducible factor 1 regulates vascular endothelial growth factor expression in human pancreatic cancer

Introduction: The microenvironment of low oxygen that is present in human pancreatic cancer in vivo may actively influence tumor growth as well as neovascularization. Aims: To determine whether hypoxia-inducible factor 1 (HIF-1) is specifically activated by hypoxia in vitro in pancreatic cancer cells and correlated these findings with tumor specimens. Methodology: Hypoxic regulation of vascular endothelial growth factor (VEGF) was studied by northern blot analysis and enzyme-linked immunosorbent assay. Electrophoretic mobility shift assays and western blot analysis were used to demonstrate hypoxic activation of HIF-1. The relationship between HIF-1 and VEGF in human pancreatic cancer specimens was studied by immunohistochemical analysis, northern blot analysis, and in situ hybridization. Results: Studies in vivo of human pancreatic cancer tissue showed co-localization of VEGF mRNA, which is produced in ductal cancer cells, and HIF-1α protein, which was detectable in cell nuclei of the same cells. HIF-1α mRNA expression was dramatically upregulated (≈13-fold) in these specimens as well. In vitro, all pancreatic cancer cell lines increased VEGF production when exposed to low oxygen levels, by highly specific activation of HIF-1 DNA binding activity to the VEGF promoter. Cancer cell lines with high constitutive levels of HIF-1α protein were found to produce higher basal levels of VEGF. Conclusion: We conclude that HIF-1 is the regulatory link between tumor hypoxia and VEGF production in pancreatic cancer, thus establishing a biochemical pathway between tumor hypoxia and neoangiogenesis in this highly aggressive neoplasm.