Loss of NF-kappa B activity during cerebral ischemia and TNF cytotoxicity

Recent evidence implicates tumor necrosis factor (TNF), a cytokine with bot h cytotoxic and cytoprotective activities, in the patho,oenesis of cerebral ischemia. The development of TNF cytotoxicity is dependent upon the balanc e between the activities of intracellular signaling pathways that mediate e ither apoptotic or anti-apoptotic effects. One critical protective signalin g mechanism is the activation of nuclear factor (NF)-kappa B, a ubiquitous transcription factor that regulates expression of anti-apoptotic gene produ cts. Here we show the distribution and kinetics of NF-kappa B activation an d the correlation between loss of NF-kappa B activity, TNF up-regulation, a nd apoptosis in a standardized rat model of focal cerebral ischemia. We obs erved a rapid and progressive ischemia-induced loss of p65 immunoreactivity within the ischemic core and nearby penumbra. These findings were confirme d by Western blot analysis of nuclear extracts and by electrophoretic mobil ity shift assay. The anatomical area of suppressed NF-kappa B activity over lapped significantly with the zones of TNF overexpression and apoptosis. Lo ss of NF-kappa B activity and increased TNF expression preceded the onset o f cell death. Direct evidence that loss of NF-kappa B activity can sensitiz e brain cells to TNF cytotoxicity was obtained in vitro by co-administratio n of MG-132, an inhibitor of NF-kappa B activation, and TNF to neuronal-lik e and glial-like cell cultures. Inhibition of NF-kappa B significantly incr eased the sensitivity of these cultures to TNF cytotoxicity, indicating tha t the observed loss of neuronal NF-kappa B activity during cerebral ischemi a can participate in the development of TNF-induced cytotoxicity.