Inflammatory gene transcription in human astrocytes exposed to hypoxia: roles of the nuclear factor-κB and autocrine stimulation

Abstract

Mechanisms of hypoxia-induced activation of nuclear factor-κB (NF-κB) and inflammatory genes were investigated in fetal human astrocytes in culture. Astrocytes were subjected to interleukin-1β (IL-1β; 50–100 u/ml; 4–24 h), or to a 4-h hypoxia (<2% O 2) followed by a 4–24-h reoxygenation. NF-κB binding and transcriptional activity increased up to 10-fold in astrocytes exposed to IL-1β, and up to 3-fold in astrocytes subjected to hypoxia followed by reoxygenation. Both IL-1β- mRNAs and proteins hypoxia-induced NF-κB activation were blocked by the proteasome inhibitor, MG-132. MG-132 inhibited IL-1β-induced up-regulation of IL-1β and IL-8 mRNA and protein but increased hypoxia-stimulated expression/release of IL-1β and IL-8. IL-1 receptor antagonist (IL-1Ra) blocked both hypoxic astrocyte-conditioned media-induced NF-κB activation and the expression/release of IL-1β and IL-8. Astrocytes subjected to hypoxia in the presence of IL-1Ra failed to activate NF-κB, but expressed elevated levels of IL-1β and IL-8. The data suggest that hypoxia/reoxygenation-induced up-regulation of IL-1β and IL-8 in human astrocytes has two components, a NF-κB independent up-regulation during hypoxia, followed by amplification through autocrine IL-1β-induced NF-κB activation during reoxygenation.