Regulation of astrocyte activation in inflammation: Roles of interferon-beta and mitogen-activated protein kinases

Understanding the regulation of inducible nitric oxide synthase (NOS) and cytokine (TNFα and IL-6) expression in glia is of great importance because of the critical roles these factors play in CNS inflammation. In diseases such as multiple sclerosis (MS) and HIV encephalitis (HIVE), these factors have been demonstrated to be important in disease pathology.; In our cell culture system of primary human fetal glia, we have shown that astrocytes but not microglia can be activated to express iNOS and cytokines. IL-1, a cytokine shown to be expressed in activated microglia in both MS and HIVE, is necessary to stimulate expression of these factors by astrocytes. Interferon (IFN)γ acts as a priming factor to boost expression of all these factors. We sought to determine what could inhibit astrocyte activation.; We therefore screened numerous soluble factors, including those known to have inhibitory effects on rodent iNOS expression. IFNβ was also tested because of its known therapeutic efficacy relapsing-remitting MS. Of these factors, only IFNβ was able to inhibit iNOS expression in astrocytes. IFNβ also inhibited expression of TNFα and IL-6. In comparing the effects of IFNβ on IL-1- and IL-1/IFNγ-induced iNOS and cytokine expression, we found that IFNβ only inhibited expression in the presence of IFNγ. IFNβ therefore appeared to mediate inhibition by antagonizing the IFNγ-, and not IL-1-, signaling pathway. We found that IFNβ inhibited IFNγ activity by sequestration of a common transcription factor in the IFN-mediated JAK/STAT signaling pathway.; IL-1 also stimulates mitogen-activated protein kinase pathways (MAPK), which are activated by pro-inflammatory cytokines and growth factors. The three main mammalian MAPK pathways are the p38, JNK/SAPK, and ERK pathways. Since IL-1 is necessary to activate human fetal astrocytes, we believed that MAPK might also be activated and contribute to NOS and cytokine expression in astrocytes. By Western blot analysis, IL-1-induced phosphorylation of p38 and JNK MAPK in a time-dependent manner.; Using specific pharmacological inhibitors of p38 and ERK MAP kinases, we found that SB203580 (p38 inhibitor) abolished IL-1 induced iNOS and TNFα expression, while PD98059 (ERK inhibitor) had no effect. PD98059 did, however, offer partial inhibition when iNOS or TNFα was induced by the combination of IL-1/IFNγ. These findings suggest that ERK kinases contribute to IFNγ-mediated induction. In addition, using dominant negative JNK plasmids, we also saw a decrease in nitrite production. Thus, these experiments demonstrated a critical role played by p38 and JNK MAPK in IL-1-induced inflammatory gene expression in human astrocytes.; In conclusion, we have examined the regulation of astrocyte activation under inflammatory conditions to better understand their contribution to inflammation in the CNS and the inflammatory process in general. Because of the relatively specific targeting of IL-1 activation by p38 and JNK inhibitors and IFNγ activation by IFNβ, it is feasible that combination treatment could lead to a more complete inhibition of inflammation.