Interferon (IFN)-induced immunoproteasomes (i-proteasomes) have been associated with improved processing of major histocompatibility complex (MHC) class I antigens. Here, we show that i-proteasomes function to protect cell viability under conditions of IFN-induced oxidative stress. IFNs trigger the production of reactive oxygen species, which induce protein oxidation and the formation of nascent, oxidant-damaged proteins. We find that the ubiquitylation machinery is concomitantly upregulated in response to IFNs, functioning to target defective ribosomal products (DRiPs) for degradation by i-proteasomes. i-proteasome-deficiency in cells and in murine inflammation models results in the formation of aggresome-like induced structures and increased sensitivity to apoptosis. Efficient clearance of these aggregates by the enhanced proteolytic activity of the i-proteasome is important for the preservation of cell viability upon IFN-induced oxidative stress. Our findings suggest that rather than having a specific role in the production of class I antigens, i-proteasomes increase the peptide supply for antigen presentation as part of a more general role in the maintenance of protein homeostasis.

Short-chain dehydrogenases/reductases form a large, evolutionarily old family of NAD(P)(H)-dependent enzymes with over 60 genes found in the human genome. Despite low levels of sequence identity (often 10 -30%), the three-dimensional structures display a highly similar ␣/␤ folding pattern. We have analyzed the role of several conserved residues regarding folding, stability, steady-state kinetics, and coenzyme binding using bacterial 3␤/17␤-hydroxysteroid dehydrogenase and selected mutants. Structure determination of the wildtype enzyme at 1.2-Å resolution by x-ray crystallography and docking analysis was used to interpret the biochemical data. Enzyme kinetic data from mutagenetic replacements emphasize the critical role of residues Thr-12, Asp-60, Asn-86, Asn-87, and Ala-88 in coenzyme binding and catalysis. The data also demonstrate essential interactions of Asn-111 with active site residues. A general role of its side chain interactions for maintenance of the active site configuration to build up a proton relay system is proposed. This extends the previously recognized catalytic triad of Ser-Tyr-Lys residues to form a tetrad of Asn-Ser-Tyr-Lys in the majority of characterized short-chain dehydrogenases/reductase enzymes.

Repair processes that are activated in response to neuronal injury, be it inflammatory, ischaemic, metabolic, traumatic or other cause, are characterized by a failure to replenish neurons and by astrogliosis. The underlying molecular pathways, however, are poorly understood. Here, we show that subtle alterations of the redox state, found in different brain pathologies, regulate the fate of mouse neural progenitor cells (NPCs) through the histone deacetylase (HDAC) Sirt1. Mild oxidation or direct activation of Sirt1 suppressed proliferation of NPCs and directed their differentiation towards the astroglial lineage at the expense of the neuronal lineage, whereas reducing conditions had the opposite effect. Under oxidative conditions in vitro and in vivo, Sirt1 was upregulated in NPCs, bound to the transcription factor Hes1 and subsequently inhibited pro-neuronal Mash1. In utero shRNA-mediated knockdown of Sirt1 in NPCs prevented oxidation-mediated suppression of neurogenesis and caused upregulation of Mash1 in vivo. Our results provide evidence for an as yet unknown metabolic master switch that determines the fate of neural progenitors.

Statins, known as inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, exhibit numerous functions related to inflammation, such as MHC class II down-regulation, interference with T cell adhesion, and induction of apoptosis. Here we demonstrate that both subcutaneous and oral administration of atorvastatin inhibit the development of actively induced chronic experimental autoimmune encephalomyelitis in SJL/J mice and significantly reduce the inflammatory infiltration into the central nervous system (CNS). When treatment was started after disease onset, atorvastatin reduced the incidence of relapses and protected from the development of further disability. Both the reduced autoreactive T cell response measured by proliferation toward the encephalitogenic peptide PLP139–151 and the cytokine profile indicate a potent blockade of T helper cell type 1 immune response. In in vitro assays atorvastatin not only inhibited antigen-specific responses, but also decreased T cell proliferation mediated by direct TCR engagement independently of MHC class II and LFA-1. Inhibition of proliferation was not due to apoptosis induction, but linked to a negative regulation on cell cycle progression. However, early T cell activation was unaffected, as reflected by unaltered calcium fluxes. Thus, our results provide evidence for a beneficial role of statins in the treatment of autoimmune attack on the CNS.

Recent studies in multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE), point to the fact that even in the early phase of inflammation, neuronal pathology plays a pivotal role in the sustained disability of affected individuals. We show that the major green tea constituent, (−)-epigallocatechin-3-gallate (EGCG), dramatically suppresses EAE induced by proteolipid protein 139–151. EGCG reduced clinical severity when given at initiation or after the onset of EAE by both limiting brain inflammation and reducing neuronal damage. In orally treated mice, we found abrogated proliferation and TNF-α production of encephalitogenic T cells. In human myelin-specific CD4+ T cells, cell cycle arrest was induced, down-regulating the cyclin-dependent kinase 4. Interference with both T cell growth and effector function was mediated by blockade of the catalytic activities of the 20S/26S proteasome complex, resulting in intracellular accumulation of IκB-α and subsequent inhibition of NF-κB activation. Because its structure implicates additional antioxidative properties, EGCG was capable of protecting against neuronal injury in living brain tissue induced by N-methyl-d-aspartate or TRAIL and of directly blocking the formation of neurotoxic reactive oxygen species in neurons. Thus, a natural green tea constituent may open up a new therapeutic avenue for young disabled adults with inflammatory brain disease by combining, on one hand, anti-inflammatory and, on the other hand, neuroprotective capacities.

The detection of pathological tissue alterations by manual palpation is a simple but essential diagnostic tool, which has been applied by physicians since the beginnings of medicine. Recently, the virtual "palpation" of the brain has become feasible using magnetic resonance elastography, which quantifies biomechanical properties of the brain parenchyma by analyzing the propagation of externally elicited shear waves. However, the precise molecular and cellular patterns underlying changes of viscoelasticity measured by magnetic resonance elastography have not been investigated up to date. We assessed changes of viscoelasticity in a murine model of multiple sclerosis, inducing reversible demyelination by feeding the copper chelator cuprizone, and correlated our results with detailed histological analyses, comprising myelination, extracellular matrix alterations, immune cell infiltration and axonal damage. We show firstly that the magnitude of the complex shear modulus decreases with progressive demyelination and global extracellular matrix degradation, secondly that the loss modulus decreases faster than the dynamic modulus during the destruction of the corpus callosum, and finally that those processes are reversible after remyelination. magnetic resonance imaging | elasticity imaging | tissue integrity P alpation of the brain, a hands-on experience long exclusive to neurosurgeons and pathologists detecting brain pathology, has recently become a domain for physicists and radiologists: Using magnetic resonance elastography (MRE), it is possible today to noninvasively assess the biomechanical properties of brain parenchyma in vivo. In MRE, viscoelasticity describes the tendency of tissue to resist deformation, thus translating the subjective tactile information gained from palpation into a quantifiable objective measure. These properties can be acquired by analyzing the propagation of low-frequency shear waves, which are mechanically elicited in an organ of interest (1, 2).Recent preliminary studies described distinct viscoelastic characteristics of the brain parenchyma in healthy subjects as well as changes by aging and brain pathology, underlining the applicability and relevance of cerebral MRE (3, 4). During physiological aging, there was evidence for a brain parenchymal "liquification" reflected in the decrease of solid-fluid behavior of the tissue (5). In patients suffering from multiple sclerosis (MS), a significant decrease of cerebral viscoelasticity was noted already in early disease stages compared with healthy controls (6).However, despite a rising collection of in vivo viscoelasticity data, no study has yet directly correlated viscoelastic parameters assessed via MRE with histopathological analyses. Thus, the question on how in vivo mechanical properties translate into cellular and molecular conditions has remained open.Magnetic resonance imaging (MRI) has emerged as most important paraclinical tool for the diagnosis and monitoring of neuroinflammatory diseases like MS, as reflected by current diagnostic ...

Here, we provide evidence for a detrimental role of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in neural death in T cell-induced experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). Clinical severity and neuronal apoptosis in brainstem motor areas were substantially reduced upon brain-specific blockade of TRAIL after induction of EAE through adoptive transfer of encephalitogenic T cells. Furthermore, TRAIL-deficient myelin-specific lymphocytes showed reduced encephalitogenicity when transferred to wild-type mice. Conversely, intracerebral delivery of TRAIL to animals with EAE increased clinical deficits, while naive mice were not susceptible to TRAIL. Using organotypic slice cultures as a model for living brain tissue, we found that neurons were susceptible to TRAIL-mediated injury induced by encephalitogenic T cells. Thus, in addition to its known immunoregulatory effects, the death ligand TRAIL contributes to neural damage in the inflamed brain.

Previous proteomic and transcriptional analyses of multiple sclerosis lesions1, 2, 3 revealed modulation of the renin-angiotensin and the opposing kallikrein-kinin pathways. Here we identify kinin receptor B1 (Bdkrb1) as a specific modulator of immune cell entry into the central nervous system (CNS). We demonstrate that the Bdkrb1 agonist R838 (Sar-[d-Phe]des-Arg9-bradykinin) markedly decreases the clinical symptoms of experimental autoimmune encephalomyelitis (EAE) in SJL mice4, 5, 6, whereas the Bdkrb1 antagonist R715 (Ac-Lys-[d-βNal7, Ile8]des-Arg9-bradykinin) resulted in earlier onset and greater severity of the disease. Bdkrb1-deficient (Bdkrb1−/−) C57BL/6 mice7 immunized with a myelin oligodendrocyte glycoprotein fragment, MOG35–55, showed more severe disease with enhanced CNS-immune cell infiltration. The same held true for mixed bone marrow–chimeric mice reconstituted with Bdkrb1−/− T lymphocytes, which showed enhanced T helper type 17 (TH17) cell invasion into the CNS. Pharmacological modulation of Bdkrb1 revealed that in vitro migration of human TH17 lymphocytes across blood-brain barrier endothelium is regulated by this receptor. Taken together, these results suggest that the kallikrein-kinin system is involved in the regulation of CNS inflammation, limiting encephalitogenic T lymphocyte infiltration into the CNS, and provide evidence that Bdkrb1 could be a new target for the treatment of chronic inflammatory diseases such as multiple sclerosis.