Heme oxygenase-1 (HO-1), the rate-limiting enzyme in heme degradation, is an integral membrane protein of the smooth endoplasmic reticulum. However, we detected an HO-1 immunoreactive signal in the nucleus of cultured cells after exposure to hypoxia and heme or heme/hemopexin. Under these conditions, a faster migrating HO-1 immunoreactive band was enriched in nuclear extracts, suggesting that HO-1 was cleaved to allow nuclear entry. This was confirmed by the absence of immunoreactive signal with an antibody against the C terminus and the lack of a C-terminal sequence by gas chromatographymass spectrometry. Incubation with leptomycin B prior to hypoxia abolished nuclear HO-1 and the faster migrating band on Western analysis, suggesting that this process was facilitated by CRM1. Furthermore, preincubation with a cysteine protease inhibitor prevented nuclear entry of green fluorescent proteinlabeled HO-1, demonstrating that protease-mediated C-terminal cleavage was also necessary for nuclear transport of HO-1. Nuclear localization was also associated with reduction of HO activity. HO-1 protein, whether it was enzymatically active or not, mediated activation of oxidant-responsive transcription factors, including activator protein-1. Nevertheless, nuclear HO-1 protected cells against hydrogen peroxide-mediated injury equally as well as cytoplasmic HO-1. We speculate that nuclear localization of HO-1 protein may serve to up-regulate genes that promote cytoprotection against oxidative stress.Heme oxygenase (HO) 3 catalyzes the degradation of heme and the formation of biliverdin and carbon monoxide. It is highly inducible in response to various stimuli, including oxidative stress, heavy metals, UV radiation, and inflammation (1-4). Cytoprotective roles for HO have been demonstrated in many models; however, the mechanisms by which this occurs are still under intensive study. Many have speculated that either heme catabolites, such as biliverdin, or its derivative, bilirubin, and carbon monoxide or the degradation of the pro-oxidant heme results in cytoprotection against oxidative stress (5-7). Nevertheless, all of the by-products of the HO reaction, despite being potentially cytoprotective, are also cytotoxic. Bilirubin is a potent neurotoxin (8), as is carbon monoxide (9). Furthermore, the HO reaction releases iron, which could interact with cellular oxidants to generate the hydroxyl radical (10). Transfection with an inactive HO-1 mutant protein results in cytoprotection against chemically induced oxidative stress (11). Because this effect of the mutant HO-1 could not be attributable to changes in heme catabolites, it alludes to a role for the HO-1 protein itself. Furthermore, the inactive form of HO-1 increased catalase and glutathione content (11). This suggests that the HO-1 protein itself may play a role in cellular signaling. If this were true, HO-1 would need to migrate to the nucleus or produce nuclear changes that affect transcription. There are several examples of cytoplasmic enzymes serving in nuclear functi...

In the murine model of Cryptococcus neoformans infection Th1 (IL-12/IFN-γ) and Th17 (IL-23/IL-17) responses are associated with protection, whereas an IL-4-dependent Th2 response exacerbates disease. To investigate the role of the Th2 cytokine IL-13 during pulmonary infection with C. neoformans, IL-13-overexpressing transgenic (IL-13Tg+), IL-13-deficient (IL-13−/−), and wild-type (WT) mice were infected intranasally. Susceptibility to C. neoformans infection was found when IL-13 was induced in WT mice or overproduced in IL-13Tg+ mice. Infected IL-13Tg+ mice had a reduced survival time and higher pulmonary fungal load as compared with WT mice. In contrast, infected IL-13−/− mice were resistant and 89% of these mice survived the entire period of the experiment. Ag-specific production of IL-13 by susceptible WT and IL-13Tg+ mice was associated with a significant type 2 cytokine shift but only minor changes in IFN-γ production. Consistent with enhanced type 2 cytokine production, high levels of serum IgE and low ratios of serum IgG2a/IgG1 were detected in susceptible WT and IL-13Tg+ mice. Interestingly, expression of IL-13 by susceptible WT and IL-13Tg+ mice was associated with reduced IL-17 production. IL-13 was found to induce formation of alternatively activated macrophages expressing arginase-1, macrophage mannose receptor (CD206), and YM1. In addition, IL-13 production led to lung eosinophilia, goblet cell metaplasia and elevated mucus production, and enhanced airway hyperreactivity. This indicates that IL-13 contributes to fatal allergic inflammation during C. neoformans infection.

The prevalence of asthma continues to increase. Asthma is caused by a Th2 cell-driven immune response. Its optimal treatment remains a challenge, and a sufficient immunotherapeutic approach to treating asthma has yet to be found. Using a murine asthma model, we show that a single injection of an anti-CD137 (4-1BB) mAb prevents the development of airway hyperreactivity, eosinophilic airway inflammation, excessive mucus production, and elevated IgE during the observation period of 7 weeks. Most importantly, even established disease is completely reversed by anti-CD137 mAb administration. The protection is associated with markedly reduced Th2 cytokine production and increased secretion of the Th1 cytokine IFN-γ. While B lymphocytes are partly depleted, the number of CD8 + T cells is increased. Blockade of IFN-γ and depletion of CD8 + T cells during treatment with anti-CD137 mAb reduces in part but does not abrogate the protective effect of CD137 mAb. In contrast, CD137 mAb-mediated CD4 + T cell anergy is critical for the observed effects, since transfer of CD4 + T cells from CD137 mAb-treated mice conveyed protection. These data demonstrate, for the first time to our knowledge, the capacity of anti-CD137 mAb to ameliorate allergic asthma, and they indicate CD137 as a possible target for therapeutic intervention in this disease. IntroductionAsthma, which has increased substantially in prevalence in the last 2 decades, is characterized by airway hyperreactivity (AHR) to a variety of specific and nonspecific stimuli; chronic airway inflammation with pulmonary eosinophilia; mucus hypersecretion; and increased serum IgE levels (1). Asthma is believed to be a result of an inappropriate Th2 cell-mediated immune response to common aeroallergens in genetically susceptible individuals. Although the immunological mechanisms that induce asthma or allergies are relatively well characterized, the specific mechanisms that downmodulate Th2 cell-driven allergic inflammatory responses in the lung are poorly understood. Current therapies for asthma, such as inhaled corticosteroids and β 2 -agonists, relieve symptoms but do not reverse the progression of or cure this disease, and a sufficient immunotherapeutic approach to treating asthma has yet to be found. So far, it is not clear whether there are certain molecular interactions that could be targeted to either suppress ongoing lung inflammation or prevent the recurrence of asthmatic symptoms when airborne allergens are repeatedly encountered.One causal strategy to control asthma is to directly modify T cell activation by targeting costimulatory interactions that are involved in this process (2). Optimal T cell activation requires at least 2 signals, provided by recognition of peptide-MHC proteins by the TCR, and by interaction of T cell costimulatory receptors with their ligands on APCs. The ligation of the CD28 molecule on T cells to B7-1 or B7-2 on APCs is essential for naive T cell survival and differentiation. Memory T cell responses are often CD28

Epigenetic mechanisms have emerged as links between prenatal environmental exposure and disease risk later in life. Here, we studied epigenetic changes associated with maternal smoking at base pair resolution by mapping DNA methylation, histone modifications, and transcription in expectant mothers and their newborn children. We found extensive global differential methylation and carefully evaluated these changes to separate environment associated from genotype‐related DNA methylation changes. Differential methylation is enriched in enhancer elements and targets in particular “commuting” enhancers having multiple, regulatory interactions with distal genes. Longitudinal whole‐genome bisulfite sequencing revealed that DNA methylation changes associated with maternal smoking persist over years of life. Particularly in children prenatal environmental exposure leads to chromatin transitions into a hyperactive state. Combined DNA methylation, histone modification, and gene expression analyses indicate that differential methylation in enhancer regions is more often functionally translated than methylation changes in promoters or non‐regulatory elements. Finally, we show that epigenetic deregulation of a commuting enhancer targeting c‐Jun N‐terminal kinase 2 (JNK2) is linked to impaired lung function in early childhood.