Ferroptosis is a necrotic form of regulated cell death (RCD) mediated by phospholipid peroxidation in association with free iron-mediated Fenton reactions. Disrupted iron homeostasis resulting in excessive oxidative stress has been implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD). Here, we demonstrate the involvement of ferroptosis in COPD pathogenesis. Our in vivo and in vitro models show labile iron accumulation and enhanced lipid peroxidation with concomitant non-apoptotic cell death during cigarette smoke (CS) exposure, which are negatively regulated by GPx4 activity. Treatment with deferoxamine and ferrostatin-1, in addition to GPx4 knockdown, illuminate the role of ferroptosis in CS-treated lung epithelial cells. NCOA4-mediated ferritin selective autophagy (ferritinophagy) is initiated during ferritin degradation in response to CS treatment. CS exposure models, using both GPx4-deficient and overexpressing mice, clarify the pivotal role of GPx4-regulated cell death during COPD. These findings support a role for cigarette smoke-induced ferroptosis in the pathogenesis of COPD.

Gene products of hepatitis C virus (HCV), a possible major causative agent of posttransfusion non-A, non-B hepatitis, are considered to be produced from a precursor polyprotein via proteolytic processing mediated by either host cell or viral proteinases. The presence of HCV serine proteinase has been proposed from analyses of amino acid sequence homology. To examine the processing mechanism of the HCV precursor polyprotein, the amino-terminal region of the putative nonstructural protein region of the HCV genome, containing the serine proteinase motif, was expressed and analyzed by using an in vitro transcription/translation system and a transient expression system in cultured cells. Two distinct proteinase activities which function in the production of a 70-kDa protein (p70) from the precursor polyprotein were detected. One of these proteinase activities, which cleaved the carboxyl (C)-terminal side of p70, required the presence of the serine proteinase motif, which is located in the amino (N)-terminal region of p70. That suggested that the predicted HCV serine proteinase was functional. The other activity, which was responsible for the cleavage of the N-terminal side of p70, required the expression of the region upstream and downstream of that cleavage site, including the p70 serine proteinase domain. From the results of pulse-chase analysis, using proteinase inhibitors coupled with a point mutation analysis, the latter activity was proposed to be a novel zinc-dependent metalloproteinase.

Allergic rhinitis a chronic inflammatory disease of the upper airways that has a major impact on the quality of life of patients and is a socio-economic burden. Understanding the underlying immune mechanisms is central to developing better and more targeted therapies. The inflammatory response in the nasal mucosa includes an immediate IgE-mediated mast cell response as well as a latephase response characterized by recruitment of eosinophils, basophils, and T cells expressing Th2 cytokines including interleukin (IL)-4, a switch factor for IgE synthesis, and IL-5, an eosinophil growth factor and on-going allergic inflammation. Recent advances have suggested new pathways like local synthesis of IgE, the IgE-IgE receptor mast cell cascade in on-going allergic inflammation and the epithelial expression of cytokines that regulate Th2 cytokine responses (i.e., thymic stromal lymphopoietin, IL-25, and IL-33). In this review, we briefly review the conventional pathways in the pathophysiology of allergic rhinitis and then elaborate on the recent advances in the pathophysiology of allergic rhinitis. An improved understanding of the immune mechanisms of allergic rhinitis can provide a better insight on novel therapeutic targets.

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NAD kinase was purified to homogeneity from Escherichia coli MG1655. The enzyme was a hexamer consisting of 30 kDa subunits and utilized ATP or other nucleoside triphosphates as phosphoryl donors for the phosphorylation of NAD, most efficiently at pH 7.5 and 60 8C. The enzyme could not use inorganic polyphosphates as phosphoryl donors and was designated as ATP±NAD kinase. The N-terminal amino-acid sequence of the purified enzyme was encoded by yfjB, which had been deposited as a gene of unknown function in the E. coli whole genomic DNA sequence database. yfjB was cloned and expressed in E. coli BL21(DE3)pLysS. The purified product (YfjB) showed NAD kinase activity, and was identical to ATP±NAD kinase purified from E. coli MG1655 in molecular structure and other enzymatic properties. ) have been purified to homogeneity. Among them, the enzymes of M. flavus and M. tuberculosis have been shown to utilize inorganic polyphosphate [poly(P)] in addition to ATP, and were designated as poly(P)/ATP±NAD kinase' [7]. However, the gene for NAD kinase, to the best of our knowledge, had never been cloned from any organism before we isolated the gene for the enzyme from M. tuberculosis [7].The lack of information about the NAD kinase gene has hindered definative understanding of NADP metabolism in organisms. To overcome this, and to obtain new insights regarding the regulation of the cellular redox system, we have attempted to isolate the NAD kinase gene of Escherichia coli. NAD kinase has been partially purified from E. coli and some properties of the enzyme have been preliminarily reported [8]. In this study, we identified the gene for NAD kinase of E. coli through purification and analysis of the primary structure of the enzyme, and revealed that the E. coli NAD kinase can utilize ATP and other nucleoside triphosphates, but not poly(P), and designated the enzyme as ATP±NAD kinase to distinguish it from poly(P)/ATP±NAD kinase of M. flavus and M. tuberculosis. M A T E R I A L S A N D M E T H O D SBacterial strains E. coli MG1655 was cultured at 37 8C in YGD medium comprising 0.1% (NH4) 2 SO4, 0.05% MgSO 4 7H 2 O, 0.1% KH 2 PO4, 0.4% Na 2 HPO 4 , 0.5% yeast extract, and 0.5% glucose (pH 7.2). As a host for plasmid amplification, E. coli DH5a (Toyobo, Osaka, Japan) was routinely cultured at 37 8C in Luria±Bertani medium [9] supplemented with ampicillin (100 mg´mL 21 ). The growth conditions for the derivative strains of E. coli BL21(DE3) pLysS (Novagen, Darmstadt, Germany) are described below.Assays NAD kinase was assayed by means of a two-step method as described previously [10] in a reaction mixture (1.0 mL) consisting of 5.0 mm NAD, 5.0 mm MgCl 2 , 100 mm Tris/ HCl pH 7.0, and 5.0 mm ATP. NADP was determined enzymatically with isocitrate dehydrogenase [11]. One unit of enzyme activity was defined as 1.0 nmol of NADP produced in 1 min at 37 8C, and specific activity was expressed in U´mg protein 21 . K m , V max and Hill coefficients (h) were determined by means of Lineweaver±Burk and Hill plots, respectively. The rate constan...

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Anaplastic large cell lymphoma (ALCL) is a subtype of non-Hodgkin's lymphoma characterized by the CD30+ large neoplastic cells and sometimes carries a t(2;5)(p23;q35). Recently, we found a novel hyperphosphorylated 80-kD protein tyrosine kinase, p80, in ALCLs with t(2;5). Subsequent cDNA cloning showed p80 to be a fusion protein of two genes, the novel tyrosine kinase gene and the nucleophosmin gene, in accordance with the sequence of the NPM/ALK gene (Morris et al, Science 263:1281, 1994). Meanwhile, the clinicopathologic features of p80-carrying ALCLs have remained unclear. Paraffin sections of 105 cases of ALCL were immunostained using anti-p80 antibody, and 30 of them were shown to express p80. Clinicopathologic comparison between p80-positive and -negative ALCLs showed that p80-positive cases occurred in a far younger patient age group (16.2 +/- 12.9 years; p80- negative cases, 51.0 +/- 22.3 years; P < .0001) and the patients showed a far better 5-year survival rate (79.8%; p80-negative group, 32.9%; P < .01). These data showed that p80-positive ALCL is a distinct entity both clinically and pathogenetically and should be differentiated from p80-negative ALCL.

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