Tumor Necrosis Factor-α

Abstract

Background and Purpose Tumor necrosis factor-α (TNF-α) is a pleiotropic cytokine that rapidly upregulates in the brain after injury. The present study was designed to explore the pathophysiological significance of brain TNF-α in the ischemic brain by systematically evaluating the effects of lateral cerebroventricular administration of exogenous TNF-α and agents that block the effects of TNF-α on focal stroke and by examining the potential direct toxic effects of TNF-α on cultured neurons to better understand how TNF-α might mediate stroke injury. Methods TNF-α (2.5 or 25 pmol) was administered intracerebroventricularly to spontaneously hypertensive rats 24 hours before permanent or transient (80 minutes and 160 minutes) middle cerebral artery occlusion (MCAO). Animals were examined 24 hours later for neurological deficits and ischemic hemisphere necrosis and swelling. In some of these studies, neutralizing anti–TNF-α monoclonal antibody (mAb) (60 pmol) was injected intracerebroventricularly 30 minutes before exogenous TNF-α (25 pmol). In addition, the effects of blocking endogenous TNF-α on permanent focal ischemic injury were determined with the use of either mAb (60 pmol) or soluble TNF receptor I (sTNF-RI) (0.3 or 0.7 nmol) administered intracerebroventricularly 30 minutes before and 3 and 6 hours after MCAO. Finally, the direct neurotoxic effects of TNF-α were studied in cultured rat cerebellar granule cells exposed to TNF-α (10 to 2000 U/mL for 6 to 24 hours), and neurotransmitter release, glutamate toxicity, and oxygen radical toxicity were studied. Results TNF-α increased the percent hemispheric infarct induced by permanent MCAO in a dose-related manner from 13.1±1.3% (vehicle) to 18.9±1.7% at 2.5 pmol ( P <.05) and 27.1±1.3% at 25 pmol ( P <.0001). The high dose of TNF-α increased ischemia-induced forelimb deficits from 1.6±0.2 to 2.3±0.2 ( P <.01). TNF-α (2.5 pmol) also increased the infarction induced by 80 or 160 minutes of transient MCAO from 1.9±0.9% to 4.3±0.4% ( P <.01) and from 14.2±1.3% to 21.6±2.2% ( P <.05), respectively. The exacerbation of infarct size, swelling, and neurological deficit after exogenous TNF-α was reversed by preinjection of 60 pmol mAb. Blocking endogenous TNF-α also significantly reduced focal ischemic brain injury. Treatment with 60 pmol mAb before and after permanent MCAO significantly reduced infarct size compared with control (nonimmune) antibody treatment by 20.2% ( P <.05). Reduced brain infarction also was produced by brain administration of 0.3 nmol (decreased 18.2%) or 0.7 nmol (decreased 26.1%; P <.05) sTNF-RI before and after focal stroke. The intracerebroventricular administration of TNF-α or sTNF-RI did not alter brain or body temperature, blood gases or pH, blood pressure, blood glucose, or general blood chemistry. In cultured cerebellar granule cells, the application of TNF-α did not directly affect neurotransmitter release or glutamate or oxygen free radical toxicity. Conclusions These studies demonstrate that exogenous TNF-α exacerbates focal ischemic injury and that blocking endogenous TNF-α is neuroprotective. The specificity of the action(s) of TNF-α was demonstrated by antagonism of its effects with specific anti–TNF-α tools (ie, mAb and sTNF-RI). TNF-α toxicity does not appear to be due to a direct effect on neurons or modulation of neuronal sensitivity to glutamate or oxygen radicals and apparently is mediated through nonneuronal cells. These data suggest that inhibiting TNF-α may represent a novel pharmacological strategy to treat ischemic stroke.