EXPRESSION OF TYPE-II NITRIC-OXIDE SYNTHASE IN PRIMARY HUMAN ASTROCYTES AND MICROGLIA - ROLE OF IL-1-BETA AND IL-1 RECEPTOR ANTAGONISTS

In this work, we studied the expression of type II nitric oxide syntha se (NOS) in primary cultures of human astrocytes and microglia, Cytoki ne-activated human fetal astrocytes expressed a 4.5-kb type II NOS mRN A that was first evident at 8 h, steadily increased through 48 h, and persisted through 72 h, The inducing signals for astrocyte NOS II mRNA expression were in the order IL-1 beta + IFN-gamma > IL-1 beta + TNF- alpha > IL-1 beta, SDS-PAGE analysis of cytokine-stimulated astrocyte cultures revealed an approximately 130-kDa single NOS II band that was expressed strongly at 48 and 72 h (72 h > 48 h), Specific NOS II immu noreactivity was detected in cytokine-treated astrocytes, both in the cytosol and in a discrete paranuclear region, which corresponded to Go lgi-like membranes on immunoelectron microscopy, In human microglia, c ytokines and LPS failed to induce NOS II expression, while the same st imuli readily induced TNF-alpha expression, In cytokine-treated human astrocytes, neither NOS II mRNA/protein expression nor nitrite product ion was inhibited by TCF-beta, IL-4, or IL-10, In contrast, IL-1 recep tor antagonist exerted near complete inhibition of NOS II mRNA and nit rite induction, Monocyte chemoattractant peptide-1 mRNA was induced in TGF-beta-treated astrocytes, demonstrating the presence of receptors for TGF-beta in astrocytes. These results confirm that in humans, cyto kines stimulate astrocytes, but not microglia, to express NOS II belon ging to the high output nitric oxide system similar to that found in r odent macrophages. They also show that the regulation of type II NOS e xpression in human glia differs significantly from that in rodent glia , A crucial role for the IL-1 pathway in the regulation of human astro cyte NOS II is shown, suggesting a potential role for IL-1 as a regula tor of astrocyte activation in vivo.