BACKGROUNDThe release of Weibel-Palade Bodies (WPB) is a form of endothelial cell activation. But the signal transduction pathway leading to WPB release is not yet defined. We hypothesized that small G-protein rac1 and reactive oxygen species (ROS) mediate the ligand induced release of Weibel-Palade Bodies.

METHODSWe tested this hypothesis by using wild-type and mutant adenoviral rac1 expression vectors, and by manipulating the production and destruction of superoxide and hydrogen peroxide in human aortic endothelial cells (HAEC).

RESULTSThrombin (1.0 Unit, 30 min) induced the increase of WPB release by 3.7-fold in HAEC, and that H2O2 (0.1 mmol/L, 30 min) induced by 4.5-fold. These results correlated with thrombin-stimulated activation of rac-GTP binding activity by 3.5-fold, and increase of ROS production by 3.4-fold. The dominant negative adenoviral rac-N17 gene transfer dramatically inhibited the release of WPB by 64.2% (control) and 77.3% (thrombin-stimulation), and decreased ROS production by 65.5% (control) and 83.6% (thrombin-stimulation) compared with non-infected cells, respectively. Anti-oxidants, catalase and N-acetyl-cysteine significantly decreased the release of WPB by 34% and 79% in control cells, and further decreased by 63.6% and 46.7% in rac-N17 transferred cells compared with non-infected cells. We also confirmed that rac1 was located upstream of ROS in the WPB release pathway.

CONCLUSIONSSmall G-protein rac1 medicates ligand-induced release of Weibel-Palade Bodies in human aortic endothelial cells, and the signal pathway of WPB release is a rac1-dependent ROS regulating mechanism.

Aorta ; ultrastructure ; Endothelial Cells ; ultrastructure ; Humans ; Reactive Oxygen Species ; Signal Transduction ; Thrombin ; pharmacology ; Weibel-Palade Bodies ; physiology ; rac1 GTP-Binding Protein ; physiology

PURPOSE: Apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1/Ref-1) is a multifunctional protein that shows elevated expression in a number of cancers. We attempted to determine whether serum APE1/Ref-1 is elevated in patients with bladder cancer. MATERIALS AND METHODS: Serum APE1/Ref-1 levels were determined using enzyme-linked immunosorbent assay in serum from patients with bladder cancer who had not received chemotherapy or radiotherapy (n=51) and non-tumor controls (n=55). The area under the receiver operating characteristic area under the curve was applied to determine the correlation between clinical factors and the serum levels of APE1/Ref-1. RESULTS: Serum levels of APE1/Ref-1 in bladder cancer patients were significantly elevated compared to those of the control group (3.548+/-0.333 ng/100 muL [n=51] for bladder cancer vs. 1.547+/-0.319 ng/100 muL [n=55] for the control group), with a sensitivity and specificity of 93% and 59%, respectively. Serum APE1/Ref-1 levels are associated with tumor stage, grade, muscle invasion, and recurrence. CONCLUSION: Serum APE1/Ref-1 might be useful as a potential serologic biomarker for bladder cancer.
Biological Markers ; Drug Therapy ; Enzyme-Linked Immunosorbent Assay ; Humans ; Radiotherapy ; Recurrence ; ROC Curve ; Sensitivity and Specificity ; Urinary Bladder Neoplasms* ; Urinary Bladder*