Proin¯ammatory cytokines, pathological iron deposition, and oxidative stress have been implicated in the pathogenesis of multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE). HO-1 mRNA levels and mitochondrial uptake of [

Bacterial superantigens, including the staphylococcal enterotoxins, are the most potent activators of T cells known and have been suggested as a causative factor in Gram‐positive shock in humans. Staphylococcal enterotoxin D (SED) is dependent upon Zn2+ for high affinity interactions with MHC class II molecules and thus SED was co‐crystallized with Zn2+. The crystal structure of SED has been determined in two different space groups, at 2.3 and 3.0 A resolution respectively. The three‐dimensional structure of SED is similar to structures of other bacterial superantigens, although this study has revealed that SED has the unique capability of forming dimers in the presence of Zn2+. The high affinity Zn2+ site used in dimer formation is located on the surface of the beta‐sheet in the C‐terminal domain. Two bound metal ions are coordinated by residues from both molecules in the dimer interface and thus contribute directly to formation of the dimer. A second Zn2+ site is located on the surface close to the domain interface of the molecule. The unique feature of SED in forming a Zn2+‐dependent homodimer seems to facilitate novel and biologically relevant multimeric interactions with MHC class II molecules, as shown by the induction of cytokine mRNA in human monocytes when exposed to SED and SED mutants.

SumnlaryStaphylococcal enterotoxin A (SEA) has two distinct binding sites for major histocompatibility complex (MHC) class II molecules. The aspartic acid located at position 227 (D227) in the COOH terminus of SEA is one of the three residues involved in its interaction with the DR[3 chain, whereas the phenylalanine 47 (F47) of the NH 2 terminus is critical for its binding to the DRot chain. Upon interaction with MHC class II molecules, SEA triggers several cellular events leading to cytokine gene expression. In the present study, we have demonstrated that, contrary to wild-type SEA, stimulation of the THP1 monocytic cell line with SEA mutated at position 47 (SEAF4v~ or at position 227 (SEAD227t0 failed to induce interleukin 113 and tumor necrosis factor-o~ messenger RNA expression. Pretreatment of the cells with a 10-fold excess of either SEAFa7A or SEAD227 A prevented the increase in cytokine messenger RNA induced by wild-type SEA. However, cross-linking of SEAF47A or SEAD227 A bound to MHC class II molecules with F(ab')2 anti-SEA mAb leads to cytokine gene expression, whereas cross-linking with F(ab) fragments had no effect. Taken together, these results indicate that cross-linking of two MHC class II molecules by one single SEA molecule is a requirement for cytokine gene expression.

Plasma and CSF HO-1 protein and lymphocyte HO-1 mRNA levels are decreased in subjects with sporadic AD. Quantitative assay for lymphocyte HO-1 mRNA expression may serve as a useful biologic marker in early sporadic AD.

Manganese superoxide dismutase (MnSOD) is an antioxidant enzyme that reduces superoxide anion to hydrogen peroxide in cell mitochondria. MnSOD is overexpressed in normal aging brain and in various central nervous system disorders; however, the mechanisms mediating the upregulation of MnSOD under these conditions remain poorly understood. We previously reported that cysteamine (CSH) and other pro-oxidants rapidly induce the heme oxygenase-1 (HO-1) gene in cultured rat astroglia followed by late upregulation of MnSOD in these cells. In the present study, we demonstrate that antecedent upregulation of HO-1 is necessary and sufficient for subsequent induction of the MnSOD gene in neonatal rat astroglia challenged with CSH or dopamine, and in astroglial cultures transiently transfected with full-length human HO-1 cDNA. Treatment with potent antioxidants attenuates MnSOD expression in HO-1-transfected astroglia, strongly suggesting that intracellular oxidative stress signals MnSOD gene induction in these cells. Activation of this HO-1-MnSOD axis may play an important role in the pathogenesis of Alzheimer disease, Parkinson disease and other free radical-related neurodegenerative disorders. In these conditions, compensatory upregulation of MnSOD may protect mitochondria from oxidative damage accruing from heme-derived free iron and carbon monoxide liberated by the activity of HO-1.

SummaryCells in the rheumatoid synovium express high levels of major histocompatibility complex (MHC) class II molecules in vivo. We have therefore examined the ability of engagement of MHC class II molecules by the superantigen Staphylococcal enterotoxin A (SEA) to activate interleukin 6 (IL-6) and ILo8 gene expression in type B synoviocytes isolated from patients with rheumatoid arthritis. SEA had a minimal or undetectable effect on the expression of either gene in resting synoviocytes, as determined by Northern blot and specific enzyme-linked immunosorbent assay. However, induction of MHC class II molecule expression after treatment of synoviocytes with interferon 3' (IFN-3') enabled the cells to respond to SEA in a dose-dependent manner, resulting in an increase in both the level of steady-state mRNA for IL-6 and IL-8, and the release of these cytokines into the supernatant. IFN-3' by itself had no effect on the expression of either cytokine. Pretreatment of the cells with the transcription inhibitor actinomycin D prevented the increase in cytokine mRNA induced by ~fEA, whereas cycloheximide superinduced mRNA for both cytokines after stimulation by SEA. Taken together, these results indicate that signaling through MHC class II molecules may represent a novel mechanism by which inflammatory cytokine production is regulated in type B rheumatoid synoviocytes, and potentially provides insight into the manner by which superantigens may initiate and/or propagate autoimmune diseases.heumatoid arthritis (RA) is a chronic articular inflammatory disorder in which the synovial environment is characterized by intense immunological activity (1). In particular, the macrophage-like (type A) and fibroblast-like (type B) synoviocytes of the hyperplastic lining layer exhibit an activated phenotype. Both cell types express a high level of MHC class II molecules (1), which are involved in the initial step of the immune response (2), and these cells are a major source of several inflammatory cytokines, which also are believed to play a crucial role in the pathogenesis of RA (1).Using superantigens such as Staphylococcal enterotoxins A or B (SEA or SEB), toxic shock syndrome toxin 1 (TSST-1), or mAbs that bind to various epitopes on the MHC class II molecule (3, 4), a number of reports have addressed the possibility that upon engagement, MHC class II molecules on APC (B cells and monocytes) may act as signal transduction molecules. For example, activation of B cells with an anti-MHC class II antibody stimulated translocation of protein kinase C (PKC) and a rise in intracellular cAMP levels (5). Binding of TSST-1 to MHC class II molecules also triggers human B cell proliferation and differentiation via MHC class II-unrestricted cognate T-B cell interaction (6). We have also reported that TSST-1 induces sustained LFA-l-dependent adhesion of MHC class II-positive cells (7) and delivers a comitogenic signal to human B cells primed with PMA or anti-IgM (8). In addition, it has recently been reported that treatment of human monocytes a...

Abstract:Little is currently known concerning the mechanisms responsible for the excessive deposition of redox-active iron in the substantia nigra of subjects with Parkinson's disease (PD). In the present study, we demonstrate that dopamine promotes the selective sequestration of non-transferrin-derived iron by the mitochondrial compartment of cultured rat astroglia and that the mechanism underlying this novel dopamine effect is oxidative in nature. We also provide evidence that up-regulation of the stress protein heme oxygenase-1 (HO-1) is both necessary and sufficient for mitochondrial iron trapping in dopamine-challenged astroglia. Finally, we show that opening of the mitochondrial transition pore (MTP) mediates the influx of non-transferrin-derived iron into mitochondria of dopamine-stimulated and HO-1 -transfected astroglia. Our findings provide an explanation for the pathological iron sequestration, mitochondrial insufficiency, and amplification of oxidative injury reported in the brains of PD subjects. Pharmacological blockade of transition metal trapping by "stressed" astroglial mitochondria (e.g., using HO-1 inhibitors or modulators of the MTP) may afford effective neuroprotection in patients with PD and other neurological afflictions.

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