A direct requirement of nuclear factor-kappa B for suppression of apoptosis in ventricular myocytes.

Nuclear factor-kappa B (NF-kappa B) is a ubiquitously expressed cellular factor regulated by the cytoplasmic factor inhibitor protein kappa B alpha (I kappa B alpha). Activation of NF-kappa B by cytokines, including tumor necrosis factor-alpha (TNF-alpha), requires the phosphorylation and degradation of I kappa B alpha. An anti-apoptotic role for NF-kappa B has recently been suggested. In the present study, we ascertained whether death-promoting signals and apoptosis mediated by TNF-alpha are suppressed by NF-kappa B in postnatal ventricular myocytes. Stimulation of myocytes with TNF-alpha resulted in a 12.1-fold increase (P < 0.01) in NF-kappa B-dependent gene transcription and DNA binding compared with controls. This was accompanied by a corresponding increase in the NF-kappa B target protein A20 as determined by Western blot analysis. Vital staining revealed that TNF-alpha was not cytotoxic to myocytes and did not provoke apoptosis. Adenovirus-mediated delivery of a nonphosphorylatable form of I kappa B alpha to inactivate NF-kappa B prevented TNF-alpha-stimulated NF-kappa B-dependent gene transcription and nuclear NF-kappa B DNA binding. Importantly, myocytes stimulated with TNF-alpha and defective for NF-kappa B activation resulted in a 2.2-fold increase (P < 0.001) in apoptosis. To our knowledge, the data provide the first indication that a functional NF-kappa B signaling pathway is crucial for suppressing death-promoting signals mediated by TNF-alpha in ventricular myocytes.