This article focuses on the production of chemokines by resident glial cells of the nervous system. We describe studies in two distinct categories of inflammation within the nervous system: immune-mediated inflammation as seen in experimental autoimmune encephalomyelitis (EAE) or multiple sclerosis (MS) and post-traumatic inflammation. We provide evidence that chemokines play a role in amplifying the inflammatory reaction in EAE (and, probably, MS). In the context of neural trauma, chemokines appear to be primary stimuli for leukocyte recruitment. Strikingly, expression of monocyte chemoattractant protein (MCP)-1 and interferon-gamma-inducible protein-10 (IP-10) are largely restricted to astrocytes or other glial cells in these diverse pathological states. The remainder of the review focuses on studies that address the molecular mechanisms which underlie transcriptional regulation of three astrocyte-derived chemokines: MCP-1, IP-10 and beta-R1/interferon-gamma-inducible T-cell chemoattractant (I-TAC). Based on these studies, we propose that the complex promoters of these genes are marvelously organized for flexible and efficient response to challenge. In the case of MCP-1, several different stimuli can elicit gene transcription, acting through a conserved mechanism that includes binding of inducible transcription factors and recruitment of the constitutive factor Sp1. For IP-10 and beta-R1/I-TAC, it appears that efficient gene transcription occurs only in highly inflammatory circumstances that produce aggregates of simultaneous stimuli. These characteristics, in turn, mirror the expression patterns of the endogenous genes: MCP-1 is expressed under a variety of circumstances, while IP-10 appears primarily during immune-mediated processes that feature exposure of resident neuroglia to high levels of inflammatory cytokines.

We characterized regulation of the human monocyte chemoattractant protein-1 (hMCP-1) gene by IFN-γ in astrocytoma cells, because astroglial cells express chemokines in several central nervous system inflammatory states. It was found that IFN-γ-induced hMCP-1 transcription was rapid, transient, and mediated by a 213-bp promoter-proximal regulatory region of the gene. Our studies on both in vitro and in vivo states of the hMCP-1 regulatory region established requirement of an IFN-γ-activated site (GAS) and the presence of IFN-γ-inducible GAS-binding activity involving at least STAT-1α for IFN-γ-induced hMCP-1 expression. Unexpectedly, in vivo genomic footprinting of the proximal regulatory region of the IFN-γ-induced gene revealed protection of a GC-rich sequence (GC box) with the same temporal pattern as that seen at the GAS; in vitro, this GC-rich element is associated with nuclear factor Sp1. These observations suggested a cooperative interaction between the GAS and the GC box element. Interestingly, site-specific mutations that abolished GC-box or GAS-element function produced clearly disparate results. Disruption of the GC box did not affect fold induction by IFN-γ but reduced promoter-reporter expression by half. Conversely, GAS mutation abrogated induction but did not affect the magnitude of expression. These results establish the importance of the GAS element for induction of hMCP-1 and further our understanding of IFN-γ-mediated transcriptional induction by providing the first evidence in vivo for inducible signaling to the GC box by this cytokine.

Precise regulation of MHC class II gene expression is crucial for development and function of the immune system. Class II trans-activator (CIITA) has been shown to be required for constitutive and IFN-γ-induced MHC class II transcription. TNF-α is commonly coexpressed with IFN-γ during immune-mediated inflammatory responses and modulates IFN-γ-stimulated MHC class II expression. The effect of TNF-α on MHC class II expression depends on cell type and cellular differentiation state. We show here that TNF-α suppresses IFN-γ-induced CIITA mRNA accumulation, resulting in decreased MHC class II expression in human fibrosarcoma HT1080 cells. TNF-α also inhibits CIITA mRNA accumulation and protein expression in a tetracycline-regulated system without affecting promoter activity. CIITA mRNA, regulated by either IFN-γ or tetracycline, was destabilized in the presence of TNF-α, suggesting that TNF-α utilizes a distinct mechanism to suppress MHC class II expression in HT1080 cells. Consistent with this interpretation, TNF-α blocked IFN-γ-induced CIITA and MHC class II expression in mutant cells that are unresponsive to TGF-β or IFN-β. This is the first instance in which MHC class II expression is inhibited by destabilizing CIITA mRNA.