Molecular structure and organization of crustacean hyperglycemic hormone genes of Penaeus monodon.
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The Crustacean hyperglycemic hormone (CHH) has been shown to exist as multiple molecular forms in several crustacean species. In Penaeus monodon, a gene encoding CHH (so-called Pem-CHH1) was recently described. In this study, the molecular structures of two other CHH genes (Pem-CHH2 and Pem-CHH3) are reported. Both the Pem-CHH2 and Pem-CHH3 genes contain three exons that are separated by two introns that are similar to the structure of other genes in the same family. An analysis of the upstream nucleotide sequences of each Pem-CHH gene has identified the putative promoter element (TATA box) and putative binding sites for several transcription factors. The binding sites for CREB, Pit-1, and AP-1 were found upstream of all three Pem-CHH genes. A Southern blot analysis showed that at least one copy of each Pem-CHH gene was located within the same 10 kb genomic DNA fragment. These results suggest that the CHH genes are arranged in a cluster in the genome of P. monodon, and that their expression may be modulated by similar mechanisms. (+info)
Intranasal exposure of mice to house dust mite elicits allergic airway inflammation via a GM-CSF-mediated mechanism.
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It is now well established that passive exposure to inhaled OVA leads to a state of immunological tolerance. Therefore, to elicit allergic sensitization, researchers have been compelled to devise alternative strategies, such as the systemic delivery of OVA in the context of powerful adjuvants, which are alien to the way humans are exposed and sensitized to allergens. The objectives of these studies were to investigate immune-inflammatory responses to intranasal delivery of a purified house dust mite (HDM) extract and to evaluate the role of GM-CSF in this process. HDM was delivered to BALB/c mice daily for 10 days. After the last exposure, mice were killed, bronchoalveolar lavage was performed, and samples were obtained. Expression/production of Th2-associated molecules in the lymph nodes, lung, and spleen were evaluated by real-time quantitative PCR and ELISA, respectively. Using this exposure protocol, exposure to HDM alone generated Th2 sensitization based on the expression/production of Th2 effector molecules and airway eosinophilic inflammation. Flow cytometric analysis demonstrated expansion and activation of APCs in the lung and an influx of activated Th2 effector cells. Moreover, this inflammation was accompanied by airways hyper-responsiveness and a robust memory-driven immune response. Finally, administration of anti-GM-CSF-neutralizing Abs markedly reduced immune-inflammatory responses in both lung and spleen. Thus, intranasal delivery of HDM results in Th2 sensitization and airway eosinophilic inflammation that appear to be mediated, at least in part, by endogenous GM-CSF production. (+info)
Variation of crustacean hyperglycemic hormone (cHH) level in the eyestalk and haemolymph of the shrimp Palaemon elegans following stress.
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This study investigates (by means of bioassays and ELISA using an antibody against recombinant cHH) the variation of cHH levels in the eyestalks and haemolymph of Palaemon elegans (Decapoda, Caridea) following exposure to various stresses (heavy metals and lipopolysaccharide), and correlates them with the variation in amount and time course of blood glucose. The dose-relationship between exposure to copper and quick release of cHH from the eyestalk into haemolymph was confirmed by variation of blood glucose with a dose-related hyperglycaemia, that peaked 2 h after immersion in contaminated seawater. Animals exposed to a sublethal concentration of mercury showed the same dose relation between toxicant, release of cHH from the eyestalk, increment of circulating hormone level and subsequent hyperglycaemia as observed for copper contamination. It is of note that although the highest lethal mercury concentration induced the release of cHH from the eyestalk into the haemolymph, it was not followed by a significant variation of blood glucose. Step doses of a bacterial contaminant [such as lipopolysaccharide (LPS) from E. coli injected into shrimps] confirmed the dose-relationship and convergent chain of events that bring about hyperglycaemia. These are the first data that relate the release of cHH from the eyestalk, the circulating hormone level and the consequent glycaemic response to stress. Moreover, they confirm the dose-related pathway that leads to variation of blood glucose as a quantitative biomarker of environmental quality, even at sublethal toxicant concentrations. (+info)
Der f 2 activates phospholipase D in human T lymphocytes from Dermatophagoides farinae specific allergic individuals: involvement of protein kinase C-alpha.
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The major house-dust mite allergen, Der f 2, stimulates the phospholipase D (PLD) in T lymphocytes from Dermatophagoides farinae specific allergic individuals. PLD activity increased more than two-fold in T cells from allergic patients compared with those cells from normal controls with maximal responses within 30 min after exposure of Der f 2. A well-known PLD activator PKC-alpha was found to be translocated to membrane from cytosol in Der f 2-treated T cells from Dermatophagoides farinae specific allergic individuals. Down-regulation of PKC-alpha with phorbol myristate acetate pretreatment for 24 h abolished Der f 2-induced PLD activation. Ro 320432, PKC inhibitor also reduced the effects of Der f 2-induced PLD activation suggesting that PKC-alpha acts as upstream activator of PLD in Der f 2-treated T cells. Taken together, the present data suggest that Der f 2 can stimulate PLD activity through the PKC-alpha activation in T cells from Dermatophagoides farinae allergic individuals. (+info)
Habitat diversity and adaptation to environmental stress in encysted embryos of the crustacean Artemia.
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Encysted embryos (cysts) of the brine shrimp, Artemia, provide excellent opportunities for the study of biochemical and biophysical adaptation to extremes of environmental stress in animals. Among other virtues, this organism is found in a wide variety of hypersaline habitats, ranging from deserts, to tropics, to mountains. One adaptation implicated in the ecological success of Artemia is p26, a small heat shock protein that previous evidence indicates plays the role of a molecular chaperone in these embryos. We add to that evidence here. We summarize recently published work on thermal tolerance and stress protein levels in embryos from the San Francisco Bay (SFB) of California inoculated into experimental ponds in southern Vietnam where water temperatures are much higher. New results on the relative contents of three stress proteins (hsp70, artemin and p26) will be presented along with data on cysts of A. tibetiana collected from the high plateau of Tibet about 4.5 km above sea level. Unpublished results on the stress protein artemin are discussed briefly in the context of this paper, and its potential role as an RNA chaperone. Interestingly, we show that the substantial tolerance of A. franciscana embryos to ultraviolet (UV) light does not seem to result from intracellular biochemistry but, rather, from their surrounding thick shell, a biophysical adaptation of considerable importance since these embryos receive heavy doses of UV in nature. (+info)
Structure of the house dust mite allergen Der f 2: implications for function and molecular basis of IgE cross-reactivity.
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The X-ray structure of the group 2 major allergen from Dermatophagoides farinae (Der f 2) was determined to 1.83 A resolution. The overall Der f 2 structure comprises a single domain of immunoglobulin fold with two anti-parallel beta-sheets. A large hydrophobic cavity is formed in the interior of Der f 2. Structural comparisons to distantly related proteins suggest a role in lipid binding. Immunoglobulin E (IgE) cross-reactivity between group 2 house dust mite major allergens can be explained by conserved surface areas representing IgE binding epitopes. (+info)
Multiple-mutation at a potential ligand-binding region decreased allergenicity of a mite allergen Der f 2 without disrupting global structure.
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We assessed the effect of multiple-mutations within one IgE-binding area on allergenicity of Der f 2. The triple-mutant of Der f 2, P34/95/99A, exhibited the most significant reduction of allergenicity and circular dichroism analysis showed that the global structure of Der f 2 was maintained in P34/95/99A. These results indicate that such a strategy is effective when designing allergen-vaccines, which achieve less allergenicity for a broad population of patients without disrupting the global structure. Structurally, Der f 2 is a member of the MD-2 related lipid-recognition proteins. The sites for the triple-mutation located on the characteristically charged entrance of a cavity and corresponded to the regions critical to ligand-binding in the Niemann-Pick type 2 disease protein and MD-2. (+info)
DNA vaccine encoding Der p 2 allergen generates immunologic protection in recombinant Der p 2 allergen-induced allergic airway inflammation mice model.
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BACKGROUND: DNA immunization is a promising novel type of immunotherapy against allergy. An estimated 79.2% patients with asthma, wheezing and/or rhinitis suffer from Dermatophagoides pteronyssinus group 2 (Der p 2) allegen. The aim of the present study was to determine whether DNA vaccine encoding Der p 2 could generate immunologic protection in recombinant Der p 2 (rDer p 2) allergen-induced allergic airway inflammation mice model and to understand the role of DNA vaccination in specific-allergen immunotherapy for asthma. METHODS: After DNA vaccination, BALB/c mice were sensitized by intraperitoneal injection (i.p) and challenged by intranasal instillation of rDer p 2. The lung tissues were assessed using hematoxylin and eosin. Mucus-producing goblet cells were identifed using periodic acid-Schiff (PAS)/alcian blue. The total cell number and composition of bronchoalveolar lavage samples were determined. The levels of the cytokines IL-4 and IFN-gamma, as well as IgE and IgG2a in the serum were determined by enzyme-linked immunosorbent assay. Allergen-specific IL-4 and IFN-gamma production by spleen cells were also measured by enzyme-linked immunosorbent assay. Expression of signal transducer and activator of transcription 6 (STAT6) in splenocytes were determined by Western blot. RESULTS: DNA vaccine encoding Der p 2 allergen inhibited extensive infiltration of inflammatory cells and production of mucin induced by allergen. The influx of eosinophils into the lung interstitium was significantly reduced after administration of DNA vaccine. Significant reductions of IL-4 and increase in levels of IFN-gamma in bronchoalveolar lavage fluid were observed. The allergen-specific IgE was markedly decreased in mice receiving DNA vaccination. Allergen could induce higher IFN-gamma, weaker IL-4 in cultured spleen cells from mice receiving DNA vaccine. DNA vaccination inhibited STAT6 expression of spleen cells induced by allergen. CONCLUSION: These results indicated that DNA vaccine encoding Der p 2 allergen generates immunologic protection in recombinant Der p 2 allergen-induced allergic airway inflammation mice model with regulating the immune response towards a Th1-type reaction. (+info)