Overexpression of endothelium derived nitric oxide synthase isoform 3 in the vasculature of human pancreatic tumor biopsies

Cellular nitric oxide (NO) synthesis determines whether NO has cytoprotecti ve or cytotoxic effects at anatomic sites; thus it is important to identify potential NO synthase isoforms in tumor tissue and tumor cell lines which might be involved in tumor development or destruction. Incubation of human pancreatic adenocarcinoma cell lines (AsPc-1, BxPc-3, CaPan-2) with cytokin es resulted in increased NO formation, indicating the existence of the NOS2 isoform. This was confirmed by reverse transcriptase-polymerase chain reac tion (RT-PCR) and western blot analysis. Furthermore, we identified the pre sence of the endothelium-derived NOS isoform 3 by RT-PCR analysis and immun ohistochemistry in normal and pancreatic tumor biopsies. NOS3 was markedly overexpressed in the vasculature of the tumor tissue. RT-PCR analysis of tu mor biopsies identified NOS isoform 2 mRNA in 60% of cases, but western blo t analysis or immunohistochemistry scored negative for this isoform. It is noteworthy that the NOS enzyme activity in pancreatic tumor cell lines and tumor biopsies was inhibited by EGTA by approximately 30% and 65%, respecti vely. Our results suggest that increased endothelium-derived NOS isoform 3 expression in pancreatic adenocarcinomas regulates blood flow and is theref ore involved in the vascularization and neovascularization of human pancrea tic tumors.