Cytokine-stimulated astrocytes damage human neurons via a nitric oxide mechanism

Astrocytes have been reported to play a neuropathogenic role within the brain, although little is known about the mechanism underlying astrocyte- mediated neuronal injury. We investigated the hypothesis that cytokine- stimulated astrocytes adversely affect neuronal cell survival via generation of the free radical nitric oxide (NO). Primary human astrocytes produced substantial amounts of NO in response to interleukin (IL)-1α or IL-1β, which was blocked by the NO synthase inhibitor N(G)-monomethyl-L- arginine (NMMA). IL-1β-induced NO production was markedly potentiated by interferon (IFN)-γ. IL-1 receptor agonist protein (IRAP) totally blocked NO generation by cytokine-stimulated astrocytes. Using reverse transcription- polymerase chain reaction and sequencing analyses of the astrocyte NO synthase gene, we found a single band encoding for a 615 bp product that was identical to the corresponding sequence reported for human hepatocytes. Treatment of human fetal brain cell cultures with IL-1β plus IFN-γ resulted in marked neuronal loss, as assessed by microscopic analysis and measurement of lactate dehydrogenase release. This cytokine-induced neuronal damage was blocked by simultaneous treatment of the brain cell cultures with NMMA or IRAP, suggesting a critical role of IL-1. These findings indicate that cytokine- stimulated astrocytes are neurotoxic via a NO-mediated mechanism and point to potential new therapies for neurodegenerative disorders that involve cytokines and reactive astrocytes.