Nuclear STAT3 translocation in guinea pig and rat brain endothelium during systemic challenge with lipopolysaccharide and interleukin-6.

During systemic inflammation, cytokines are released by immune-competent cells into the circulation, which in turn signal the brain to mediate brain-controlled signs of illness. Cytokine-responsive brain cells can be mapped by histological analysis of cytokine-induced transcription factors or transcription factor-associated molecules revealing different cell phenotypes that respond to activation of the immune system. Critical sites mediating cytokine-dependent immuneffector functions can be divided into two groups, one group of responding cells situated along a tight blood-brain barrier (BBB), and a second cell group in structures with an open BBB, e.g., the sensory circumventricular organs (CVOs). Previous reports from our group suggest that activation of the signal transducer and activator of transcription factor 3 (STAT3) during lipopolysaccharide (LPS)-induced systemic inflammation is mediated by interleukin-6 (IL-6) and occurs in astrocytes of the rat CVOs. Here we show in the guinea pig a time-dependent marked LPS-induced STAT3 activation within astrocytes and endothelial cells of the CVOs, within astrocytes located in brain structures with a functional BBB and within the brain endothelium of the entire brain. In addition, systemic treatment of rats with either rat recombinant IL-6 or LPS induced STAT3 activation in brain endothelial cells in a similar way as observed in the guinea pig brain, stressing the involvement of IL-6 in this phenomenon in a more generalized way. The STAT3-activated brain cells are located in critical target structures mediating cytokine action during LPS-induced inflammation. STAT3-controlled transcriptional activation with yet unknown cell-specific functional consequences seems to be involved in this process.

STAT3 and COX-2 activation in the guinea-pig brain during fever induced by the Toll-like receptor-3 agonist polyinosinic:polycytidylic acid.

Intra-arterial injections of synthetic double-stranded RNA (polyinosinic:polycytidylic acid, PIPC) at a dose of 500 microg/kg evoked pronounced fever in guinea-pigs. PIPC-induced fever could be antagonized by treatment with the non-selective cyclooxygenase (COX) inhibitor diclofenac and was, in part, attenuated by the administration of the selective COX-2-inhibitor nimesulide (dose: 5 mg/kg for both COX inhibitors). We further investigated whether direct activation of brain cells during PIPC-induced fever could be demonstrated. Using radioactive in situ hybridization, we demonstrated that treatment with PIPC resulted in an upregulation of COX-2 and interleukin-1 beta mRNA in the guinea-pig brain. Thus, COX-2-specific hybridization signals seemed to be mainly associated with brain blood vessels. Intra-arterial injections of PIPC further induced the pronounced nuclear translocation of the transcription factor STAT3 in the endothelium of various fore- and hindbrain areas and in the meninges. In brain structures that lacked a tight blood-brain barrier, i.e. the sensory circumventricular organs (area postrema, vascular organ of laminae terminalis, subfornical organ), the astrocytes and a population of still undetermined cellular phenotype also showed marked STAT3 activation in response to PIPC. The Toll-like receptor-3 agonist PIPC therefore caused a similar activation of brain cells as that reported for other experimental models of systemic inflammation.

Localized vs. systemic inflammation in guinea pigs: a role for prostaglandins at distinct points of the fever induction pathways?

In guinea pigs, dose-dependent febrile responses were induced by injection of a high (100 microg/kg) or a low (10 microg/kg) dose of bacterial lipopolysaccharide (LPS) into artificial subcutaneously implanted Teflon chambers. Both LPS doses further induced a pronounced formation of prostaglandin E(2) (PGE(2)) at the site of localized subcutaneous inflammation. Administration of diclofenac, a nonselective cyclooxygenase (COX) inhibitor, at different doses (5, 50, 500, or 5,000 microg/kg) attenuated or abrogated LPS-induced fever and inhibited LPS-induced local PGE(2) formation (5 or 500 microg/kg diclofenac). Even the lowest dose of diclofenac (5 microg/kg) attenuated fever in response to 10 microg/kg LPS, but only when administered directly into the subcutaneous chamber, and not into the site contralateral to the chamber. This observation indicated that a localized formation of PGE(2) at the site of inflammation mediated a portion of the febrile response, which was induced by injection of 10 microg/kg LPS into the subcutaneous chamber. Further support for this hypothesis derived from the observation that we failed to detect elevated amounts of COX-2 mRNA in the brain of guinea pigs injected subcutaneously with 10 microg/kg LPS, whereas subcutaneous injections of 100 microg/kg LPS, as well as systemic injections of LPS (intra-arterial or intraperitoneal routes), readily caused expression of the COX-2 gene in the guinea pig brain, as demonstrated by in situ hybridization. Therefore, fever in response to subcutaneous injection of 10 microg/kg LPS may, in part, have been evoked by a neural, rather than a humoral, pathway from the local site of inflammation to the brain.