Nrf2 is a key transcription factor that regulates antioxidant defense in macrophages and epithelial cells: protecting against the proinflammatory and oxidizing effects of diesel exhaust chemicals.

The proinflammatory effects of particulate pollutants, including diesel exhaust particles (DEP), are related to their content of redox cycling chemicals and their ability to generate oxidative stress in the respiratory tract. An antioxidant defense pathway, which involves phase II enzyme expression, protects against the pro-oxidative and proinflammatory effects of DEP. The expression of enzymes, including heme oxygenase-1 (HO-1) and GST, is dependent on the activity of a genetic antioxidant response element in their promoters. In this study we investigated the mechanism by which redox cycling organic chemicals, prepared from DEP, induce phase II enzyme expression as a protective response. We demonstrate that aromatic and polar DEP fractions, which are enriched in polycyclic aromatic hydrocarbons and quinones, respectively, induce the expression of HO-1, GST, and other phase II enzymes in macrophages and epithelial cells. We show that HO-1 expression is mediated through accumulation of the bZIP transcription factor, Nrf2, in the nucleus, and that Nrf2 gene targeting significantly weakens this response. Nrf2 accumulation and subsequent activation of the antioxidant response element is regulated by the proteasomal degradation of Nrf2. This pathway is sensitive to pro-oxidative and electrophilic DEP chemicals and is also activated by ambient ultrafine particles. We propose that Nrf2-mediated phase II enzyme expression protects against the proinflammatory effects of particulate pollutants in the setting of allergic inflammation and asthma.

Proteome analysis of lipid rafts in Jurkat cells characterizes a raft subset that is involved in NF-kappaB activation.

Lipid rafts are detergent-insoluble membrane domains that play a key role in signal transduction by the T-cell antigen receptor. Proteome analysis revealed the presence of amidosulfobetaine-soluble signal transducing, integral membrane, cytoskeletal, heat shock, and GTP-binding proteins in rafts prepared from Jurkat cells. Several of these proteins were recruited to rafts by CD3/CD28 costimulation. Of particular interest is the inducible association of activated IkappaB kinase complexes with raft vesicles that could be captured with anti-flotillin-1 antibodies. Following amidosulfobetaine solubilization, flotillin-beta and IKKbeta underwent reciprocal co-immunoprecipitation. Treatment of Jurkat cells with methyl-beta-cyclodextrin disrupted the assembly and activation of this raft complex and also interfered in CD3/ CD28-induced activation of a NF-kappaB response element in the IL-2 promoter.

The flotillins are integral membrane proteins in lipid rafts that contain TCR-associated signaling components: implications for T-cell activation.

Lipid rafts play an important role in signal integration and cellular activation by the T-cell antigen receptor (TCR). We demonstrate that flotillin-1 and flotillin-2 are important structural raft components, which redistribute to the site of TCR engagement. An antibody to flotillin-1 was able to immobilize other TCR-associated raft components. Although rafts purified from unstimulated cells demonstrated abundant Lck but inabundant LAT, rafts from stimulated cells include an abundance of both components. This suggests dynamic changes in lipid raft composition during CD3/CD28 costimulation. Stimulation of primary human CD4(+) T cells leads to increased GM1 and flotillin-1 expression in the surface membrane, where these components colocalize. This may reconstitute new signaling complexes required for T-cell activation. Altered lipid raft composition and function may play a role in the decline of antigen responsiveness in senescent T cells. In this regard, we observed an increase in the raft-associated gangliolipid, GM1, in resting human CD4(+) and CD8(+) lymphocytes with aging.

T-cell activation through the antigen receptor. Part 2: role of signaling cascades in T-cell differentiation, anergy, immune senescence, and development of immunotherapy.

Part 2 of this review on cellular activation by the T-cell antigen receptor (TCR) will highlight how TCR signaling pathways are adapted to achieve specific biologic outcomes, including different states of T-cell differentiation and the induction of T-cell tolerance. We will also explore how treatment with altered peptide ligands affects TCR signaling to change T-cell differentiation or to induce an anergy state. These changes are accomplished through alteration of protein tyrosine kinase activity, the stoichiometry of phosphorylation of immunoreceptor tyrosine-based activation motifs, intracellular free ionized calcium flux, mitogen-activated protein kinase activity, and transcriptional activation of key cytokine promoters. The CTLA-4 plays an important role in the induction and maintenance of anergy. The second theme will highlight how altered TCR signal transduction, including changes in the compartmentalization of signaling components at the TCR synapse, contributes to decreased T-cell activation during immune senescence. Finally, we will illustrate how the molecular details of TCR activation can be used to modify the function of the immune system. This includes a description of the mechanism of action of altered peptide ligands, CTLA-4Ig, and pharmacologic inhibitors of mitogen-activated protein kinases, nuclear factor kappaB, and protein kinase C cascades.