Tryptophan metabolism by the kynurenine pathway (KP) is important to the pathogenesis of inflammatory, infectious, and degenerative diseases. The 3-hydroxykynurenine (3-HK) branch of the KP is activated in macrophages and microglia, leading to the generation of 3-HK, 3-hydroxyanthranilic acid (3-HAA), and quinolinic acid, which are considered neurotoxic owing to their free radical-generating and N-methyl-D-aspartic acid receptor agonist activities. We investigated the role of 3-HAA in inflammatory and antioxidant gene expression and neurotoxicity in primary human fetal central nervous system cultures treated with cytokines (IL-1 with or without interferon-␥) or with Toll-like receptor ligands mimicking the proinflammatory central nervous system environment. Results were analyzed by microarray, Western blot, immunostain, enzyme-linked immunosorbent assay, and neurotoxicity assays. 3-HAA suppressed glial cytokine and chemokine expression and reduced cytokine-induced neuronal death. 3-HK also suppressed cytokineinduced neuronal death. Unexpectedly, 3-HAA was highly effective in inducing in astrocytes the expression of hemeoxygenase-1 (HO-1), an antioxidant enzyme with anti-inflammatory and cytoprotective properties. Indoleamine-2,3-dioxygenase (IDO) is an interferon (IFN)-␥-inducible, rate-limiting enzyme in the kynurenine pathway (KP) of tryptophan metabolism generating various downstream metabolites collectively termed "kynurenines" 1 ( Figure 1). This process is compartmentalized due to cell-specific expression of the KP enzymes. For example, kynurenine monooxygenase (KMO) is expressed in macrophages and microglia, 2-4 whereas kynurenine aminotransferase II (KAT II) is present in astrocytes.5 A well-appreciated biological activity of IDO is T-cell suppression. IDO expressed in antigen-presenting cells (dendritic cells, macrophages, and microglia) can

Astrocytes, together with microglia and macrophages, participate in innate inflammatory responses in the CNS. While inflammatory mediators such as interferons generated by astrocytes may be critical in the defense of the CNS, sustained unopposed cytokine signaling could result in harmful consequences. Interferon regulatory factor 3 (IRF3) is a transcription factor required for IFNβ production and antiviral immunity. Most cells express low levels of IRF3 protein and the transcriptional mechanism that upregulates IRF3 expression is not known. In the current study, we explored the consequence of adenovirus-mediated IRF3 gene transfer (Ad-IRF3) in primary human astrocytes. We show that IRF3 transgene expression suppresses proinflammatory cytokine gene expression upon challenge with IL-1/IFNγ and alters astrocyte activation phenotype from a proinflammatory to an anti-inflammatory one, akin to an M1 to M2 switch in macrophages. This was accompanied by the rescue of neurons from cytokine-induced death in glial-neuronal cocultures. Furthermore, Ad-IRF3 suppressed the expression of microRNA-155 and its star-form partner miR-155*, immunoregulatory miRNAs highly expressed in multiple sclerosis lesions. Astrocyte miR-155/miR155* were induced by cytokines and TLR ligands with a distinct hierarchy, and were involved in proinflammatory cytokine gene induction by targeting suppressor of cytokine signaling 1 (SOCS1), a negative regulator of cytokine signaling and potentially other factors. Our results demonstrate a novel pro-inflammatory role for miR-155/miR-155* in human astrocytes, and suggest that IRF3 can suppress neuroinflammation through regulating immunomodulatory miRNA expression.

Aims-Cannabinoids have been proposed for treating various neurodegenerative disorders and as adjunct therapy for HIV+ patients with neurologic sequelae. The expression of cannabinoid receptors (CB1 and CB2) has been reported in neurodegenerative diseases and in SIV encephalitis, yet the receptor expression in the CNS of HIV+ individuals is not known. Methods-An anti-CB1 antibody and two anti-CB2 antibodies were employed for immunohistochemistry in the cerebral cortex and white matter of HIV encephalitis (HIVE) and HIV-associated comorbidities, as well as control brains (HIV− and HIV+).Results-By quantitative image analysis, we observed that CB1 was increased in HIVE brains and those with comorbidities, while CB2 was significantly increased in the white matter of HIVE. Morphologically, CB1 was present in neurons, and both CB1 and CB2 were present in meningeal macrophages and subpial glia in all brains. In HIVE, CB1 was found in white matter microglia and perivascular cells, while CB2 was increased in microglia, astrocytes and perivascular macrophages. Double immunofluorescence with cell-specific markers and immunoblots on primary cultured microglia and astrocytes substantiated the glial localization of the cannabinoid receptors and specificity of the antibodies.Conclusions-Our study indicates that cannabinoid receptor expression occurs in glia in HIVE brains, and this may have ramifications for the potential use of cannabinoid ligands in HIVinfected patients.

Interferon regulatory factor 3 (IRF3) is a transcription factor critical in the induction of antiviral immunity. IRF3 is activated following stimulation of cell membrane or cytosolic nucleic acid sensors and is essential in the induction of the IFNβ gene. Most cells constitutively express IRF3 in vitro, but little is known about the regulation of expression of IRF3 in vivo. Immunohistochemical analysis of selected human and mouse tissues demonstrated that IRF3 expression is highly organ- and cell-type specific, showing high expression in certain epithelial cells. In the CNS, while ependymal cells are strongly positive, brain parenchyma has little detectable IRF3 immunoreactivity. The importance of IRF3 in antiviral immunity has been demonstrated by the requirement for IRF3 in suppressing viral replication, but also by the demonstration that virus degrades IRF3 protein in infected cells. Furthermore, HIV-infected microglia in human CNS show abnormal IRF3+ aggregates, indicative of aberrant protein processing in vivo. In addition to antiviral immunity, IRF3 also plays a critical role in the modulation of neuroinflammation. A combination of dominant-negative and over-expression strategies in vitro as well as transgenic expression of IRF3 in vivo demonstrated that IRF3 plays a major role in modulating glial cytokine expression, i.e., suppression of proinflammatory cytokines and promotion of anti-inflammatory or immunoregulatory cytokines. These observations together suggest that IRF3 is a crucial regulator of immune responses against pathogen- and damage-associated molecules. We review recent literature on the molecular pathways of IRF3 activation and function of IRF3 and discuss their implications for CNS diseases.

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