[The role of NF-kappa B in the TNF-alpha-induced endothelial cell apoptosis]

Abstract

Objective: To investigate whether TNF-alpha could activate the signaling pathway-NF-kappa B/I-kappa B alpha required for the expression of inducible nitric oxide synthase (iNOS) to induce endothelial cell apoptosis by nitric oxide (NO).

Methods: Aminoguanidine (AG, an inhibitor of iNOS) and Pyrrolidine dithiocarbamate (PDTC, an inhibitor of NF-kappa B) were evaluated for their influence on the apoptosis of cultured bovine pulmonary artery endothelial cells (BPAEC) induced by TNF-alpha. Apoptosis was confirmed by morphology, ultrastructural observation, agarose gel electrophoresis of DNA, and percentage of DNA fragmentation. Western blot analyses were used to detect the expression of iNOS and the levels of I-kappa B alpha in BPAEC exposed to TNF-alpha. Electrophoretic mobility shift assays (EMSA) were used to determinate the activity of NF-kappa B in BPAEC.

Results: TNF-alpha induced BPAEC apoptosis (18.0 +/- 4.3)% in a concentration- and time-dependent manner. Both AG and PDTC attenuated the apoptosis of BPAEC induced by TNF-alpha, with peventages of DNA fragmentation in 2, 4, 10 mmol/L AG being (10.0 +/- 2.2)%, (7.8 +/- 1.2)% and (8.2 +/- 1.3)% vespectively. In BPAEC after TNF-alpha exposure, Western blot analyses revealed the expression of iNOS and the decrease in I-kappa B alpha. EMSA demonstrated an impermanency increase in NF-kappa B binding activity.

Conclusions: TNF-alpha can induce endothelial cell apoptosis by NO, which is produced by increasing iNOS expression and activating the signal pathway-NF-kappa B.