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Sample records for bacteriocin-like immunity protein

  1. The structure of pyogenecin immunity protein, a novel bacteriocin-like immunity protein from Streptococcus pyogenes

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    Volkart Lour

    2009-12-01

    Full Text Available Abstract Background Many Gram-positive lactic acid bacteria (LAB produce anti-bacterial peptides and small proteins called bacteriocins, which enable them to compete against other bacteria in the environment. These peptides fall structurally into three different classes, I, II, III, with class IIa being pediocin-like single entities and class IIb being two-peptide bacteriocins. Self-protective cognate immunity proteins are usually co-transcribed with these toxins. Several examples of cognates for IIa have already been solved structurally. Streptococcus pyogenes, closely related to LAB, is one of the most common human pathogens, so knowledge of how it competes against other LAB species is likely to prove invaluable. Results We have solved the crystal structure of the gene-product of locus Spy_2152 from S. pyogenes, (PDB:2fu2, and found it to comprise an anti-parallel four-helix bundle that is structurally similar to other bacteriocin immunity proteins. Sequence analyses indicate this protein to be a possible immunity protein protective against class IIa or IIb bacteriocins. However, given that S. pyogenes appears to lack any IIa pediocin-like proteins but does possess class IIb bacteriocins, we suggest this protein confers immunity to IIb-like peptides. Conclusions Combined structural, genomic and proteomic analyses have allowed the identification and in silico characterization of a new putative immunity protein from S. pyogenes, possibly the first structure of an immunity protein protective against potential class IIb two-peptide bacteriocins. We have named the two pairs of putative bacteriocins found in S. pyogenes pyogenecin 1, 2, 3 and 4.

  2. The structure of pyogenecin immunity protein, a novel bacteriocin-like immunity protein from streptococcus pyogenes.

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    Chang, C.; Coggill, P.; Bateman, A.; Finn, R.; Cymborowski, M.; Otwinowski, Z.; Minor, W.; Volkart, L.; Joachimiak, A.; Wellcome Trust Sanger Inst.; Univ. of Virginia; UT Southwestern Medical Center

    2009-12-17

    Many Gram-positive lactic acid bacteria (LAB) produce anti-bacterial peptides and small proteins called bacteriocins, which enable them to compete against other bacteria in the environment. These peptides fall structurally into three different classes, I, II, III, with class IIa being pediocin-like single entities and class IIb being two-peptide bacteriocins. Self-protective cognate immunity proteins are usually co-transcribed with these toxins. Several examples of cognates for IIa have already been solved structurally. Streptococcus pyogenes, closely related to LAB, is one of the most common human pathogens, so knowledge of how it competes against other LAB species is likely to prove invaluable. We have solved the crystal structure of the gene-product of locus Spy-2152 from S. pyogenes, (PDB: 2fu2), and found it to comprise an anti-parallel four-helix bundle that is structurally similar to other bacteriocin immunity proteins. Sequence analyses indicate this protein to be a possible immunity protein protective against class IIa or IIb bacteriocins. However, given that S. pyogenes appears to lack any IIa pediocin-like proteins but does possess class IIb bacteriocins, we suggest this protein confers immunity to IIb-like peptides. Combined structural, genomic and proteomic analyses have allowed the identification and in silico characterization of a new putative immunity protein from S. pyogenes, possibly the first structure of an immunity protein protective against potential class IIb two-peptide bacteriocins. We have named the two pairs of putative bacteriocins found in S. pyogenes pyogenecin 1, 2, 3 and 4.

  3. Growth of Enterococcus durans E204 producing bacteriocin-like ...

    African Journals Online (AJOL)

    Bacteriocin-like substance E204 is an antimicrobial compound produced by Enterococcus durans E204 isolated from camel milk of Morocco that shows a broad spectrum of inhibitory activity against taxonomically related microorganisms. It is sensitive to digestive proteases. In the first study, de Man, Regosa and Sharpe ...

  4. Effect of Bacteriocin-like Inhibitory Substances Produced by Vaginal ...

    African Journals Online (AJOL)

    Reduction of vaginal Lactobacillus population leads to overgrowth of opportunistic organisms such as Streptococcus agalactiae (Group B Streptococcus, GBS), which causes life threatening neonatal infections. The activities of bacteriocin-like inhibitory substances (BLIS) produced by Lactobacillus species isolated from the ...

  5. Characterization of Partially Purified Bacteriocin Like Substance (BLIS Produced by Probiotic Lactobacillus Strains

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    Saeed Ismail Khanian

    2014-05-01

    Full Text Available Background: There is an increasing interest in search for antimicrobial peptides (bacteriocins and bacteriocin-like compounds produced by lactic acid bacteria (LAB because of their potential to be used as antimicrobial agents for improving the safety of food products. Objectives: The main objective of study was to evaluate the antibacterial potential of locally isolated Lactic Acid bacteria (LAB and determine their bacteriocin producing ability in in-vitro conditions. Materials and Methods: The antibacterial activity of 77 isolated LAB strains was tested against a number of pathogens by well-diffusion method. The isolates demonstrating antimicrobial potential were selected and tested for the production of bacteriocin or bacteriocin like substance. The bacteriocin produced by two of the isolates were partially purified and characterized. Results: The results indicated the neutralized supernatant fluid of two of the isolates identified as L. brevis LB32 and L. pentosus LP05, were active against the growth of Listeria monocytogenes, Salmonella enteritidis, Shigella dysenteriae, Staphylococcus aureus and Streptococcus pneumoniae. Additionally, L. brevis LB32 was able to inhibit the growth of Salmonella typhi and Klebsiella pneumoniae, while, S. pnuemoniae and L. monocytogenes appeared to be the most sensitive strain as apparent by highest zone of inhibition against these pathogens, respectively. The antimicrobial activity in the supernatant fluids of the mentioned strains remained unaffected after treating with enzymes catalase, lipase and lysozyme, while were strongly sensitive to the action of proteolytic enzymes, suggesting the presence of bacteriocin like inhibitory substance (BLIS in the two isolates. The inhibitory substance produced by the two isolates appeared heat resistant and tolerated 100˚C and 121˚C for 55 minutes and 20 minutes, respectively. Partial purification of the concentrated culture supernatant fluids of L. brevis LB32 and L

  6. Description of two Enterococcus strains isolated from traditional Peruvian artisanal-produced cheeses with a bacteriocin-like inhibitory activity

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    Aguilar Galvez A.

    2009-01-01

    Full Text Available The aim of this work was to isolate and to characterize strains of lactic acid bacteria (LAB with bacteriocin-like inhibitory activity from 27 traditional cheeses artisanal-produced obtained from different Peruvian regions. Twenty Gram+ and catalasenegative strains among 2,277 isolates exhibited bacteriocin-like inhibitory activity against Listeria monocytogenes CWBIB2232 as target strain. No change in inhibitory activity was observed after organic acid neutralization and treatment with catalase of the cell-free supernatant (CFS. The proteinic nature of the antimicrobial activity was confirmed for the twenty LAB strains by proteolytic digestion of the CFS. Two strains, CWBI-B1431 and CWBI-B1430, with the best antimicrobial activity were selected for further researches. These strains were taxonomically identified by phenotypic and genotypic analyses as Enterococcus mundtii (CWBI-B1431 and Enterococcus faecium (CWBI-B1430. The two strains were sensitive to vancomycin (MIC 2 μg.ml-1 and showed absence of haemolysis.

  7. Characterization of a bacteriocin-like substance produced from a novel isolated strain of Bacillus subtilis SLYY-3

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    Li, Junfeng; Li, Hongfang; Zhang, Yuanyuan; Duan, Xiaohui; Liu, Jie

    2014-12-01

    In the present research, the strain SLYY-3 was isolated from sediments of Jiaozhou Bay, Qingdao, China. The strain SLYY-3, which produced a bacteriocin-like substance (BLS), was characterized to be a strain of Bacillus subtillis by biochemical profiling and 16S rDNA sequence analysis. It is the first time to report that Bacillus subtilis from Jiaozhou Bay sediments could produce a BLS. The BLS of B. subtillis SLYY-3 exhibited strong inhibitory activity against gram-positive bacteria (including Staphylococcus aureus and B. subtillis) and some fungi (including Penicillium glaucum, Aspergillus niger and Aspergillus flavus). The antimicrobial activity was detected from culture in the exponential growth phase and reached its maximum when culture entered into stationary growth phase. It was thermo-tolerant even when being kept at 100°C for 60 min without losing any activity and stable over a wide pH range from 1.0 to 12.0 while being inactivated by proteolytic enzyme and trypsin, indicating the proteinaceous nature of the BLS. The BLS was purified by precipitation with hydrochloric acid (HCl) and gel filteration (Sephadex G-100). SDS-PAGE analysis of the extracellular peptides of SLYY-3 revealed a bacteriocin-like protein with a molecular mass of 66 kDa. Altogether, these characteristics indicate the potential of the BLS for food industry as a protection against pathogenic and spoilage microorganisms.

  8. Production by Streptococcus sanguis of Bacteriocin-like Inhibitory Substances (BLIS) with Activity Against Streptococcus rattus

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    Skilton, C. J.; Tagg, J. R.

    2011-01-01

    A collection of 340 Streptoccus sanguis strains from 19 plaque specimens was isolated from cultures on tryptone, yeast extract, cystine (TYC) agar supplemented with sucrose (20 per cent) plus bacitracin (0.05 units/ml). By application of a deferred antagonism test procedure it was shown that 183 (53.8 per cent) of these relatively sucrose- and bacitracinresistant S. sanguis strains produced bacteriocin-like inhibitory substances (BLIS) having activity against strains of S. rattus, but apparcn...

  9. Streptococcal Bacteriocin-Like Inhibitory Substances: Some Personal Insights into the Bacteriocin-Like Activities Produced by Streptococci Good and Bad.

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    Tagg, John Robert

    2009-06-01

    The background to the discovery and commercial development of the first Streptococcus salivarius probiotic is documented. A 40-year search of the genus Streptococcus for a harmless natural antagonist of Streptococcus pyogenes had as its operational basis a simple deferred antagonism "fingerprinting" procedure, the application of which results in each tested strain being accorded an inhibitor production (P)-type and inhibitor sensitivity (S)-type profile. Systematic application of this schema has opened a "Pandora's Box" of novel streptococcal bacteriocin-like inhibitory substances (BLIS). The numerically prominent commensal S. salivarius is proposed to have a pivotal population-modulating role within the oral microbiota of humans. The probiotic strain S. salivarius K12 produces several megaplasmid-encoded BLIS including the lantibiotics salivaricin A and salivaricin B. Strain K12 and other BLIS-producing S. salivarius are currently in use or under development for application to the control of a variety of common maladies and infections of the human oral cavity.

  10. In vitro evaluation of bacteriocin-like inhibitory substances produced by lactic acid bacteria isolated during traditional Sicilian cheese making

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    Giusi Macaluso

    2016-02-01

    Full Text Available Bacteriocins are antimicrobial proteins produced by bacteria that inhibit the growth of other bacteria with a bactericidal or bacteriostatic mode of action. Many lactic acid bacteria (LAB produce a high diversity of different bacteriocins. Bacteriocinogenic LAB are generally recognised as safe (GRAS and useful to control the frequent development of pathogens and spoilage microorganisms. For this reason they are commonly used as starter cultures in food fermentations. In this study, the authors describe the results of a screening on 699 LAB isolated from wooden vat surfaces, raw milk and traditional Sicilian cheeses, for the production of bacteriocin-like inhibitory substances, by comparing two alternative methods. The antagonistic activity of LAB and its proteinaceous nature were evaluated using the spot-on-the-lawn and the well-diffusion assay (WDA and the sensitivity to proteolytic (proteinase K, protease B and trypsin, amylolytic (α-amylase and lipolytic (lipase enzymes. The indicator strains used were: Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, Salmonella enteritidis. A total of 223 strains (belonging to the species Enterococcus spp., Lactobacillus spp., Pediococcus spp., Streptococcus spp., Leuconostoc spp. and Lactococcus lactis were found to inhibit the growth of Listeria monocytogenes by using the spot-on-the-lawn method; only 37 of these were confirmed by using the WDA. The direct addition of bacteriocin-producing cultures into dairy products can be a more practical and economic option for the improvement of the safety and quality of the final product.

  11. Antibacterial activity of bacteriocin-like substance P34 on Listeria monocytogenes in chicken sausage

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    Voltaire Sant'Anna

    2013-12-01

    Full Text Available The antimicrobial activity of the bacteriocin-like substance (BLS P34 against Listeria monocytogenes was investigated in chicken sausage. The BLS was applied to chicken sausages (256 AU g-1 previously inoculated with a suspension of 10² cfu g-1 of L. monocytogenes. BLS P34 inhibited the indicator microorganism in situ in all incubation times for up to 10 days at 5 °C. The effectiveness of BLS P34 was increased when it was added in combination with nisin. The bacteriocin was also tested in natural eatable natural bovine wrapping (salty semi-dried tripe against the same indicator microorganism, also showing inhibitory capability in vitro. BLS P34 showed potential to control L. monocytogenes in refrigerated meat products.

  12. Isolation of Lactic Acid Bacteria That Produce Protease and Bacteriocin-Like Substance From Mud Crab (Scylla sp. Digestive Tract (Isolasi Bakteri Asam Laktat yang Menghasilkan Protease dan Senyawa Bacteriocin-Like dari Saluran Pencernaan Kepiting

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    Heru Pramono

    2015-03-01

    Kata kunci: Bakteri Asam Laktat, Bakteriosin-like substance, Protease, Scylla  sp. Digestive tract is complex environment consist of large amount of bacteria’s species. Fish intestine bacteria consist of aerobic or facultative anaerob bacteria which can produce antibacterial and enzym. The objectives of this research were to isolated lactic acid bacteria that produce bacteriocin-like and protease from mud crab digestive tract. Isolation and characterization of isolates were conducted employing media MRS.  Neutralized cell free supernatant of isolates were tested using disc diffusion agar of against pathogenic and spoilage bacteria to indicate bacteriocin-like-producing lactic acid bacteria. Protease-producing isolate was tested using disc diffusion method in casein agar. Among a hundred isolates, 96 isolates were showed clear zone in MRS+CaCO3,, catalase negative, and Gram positive bacteria. Thirty four isolates produced protease and only four isolates (i.e. IKP29, IKP30, IKP52, and IKP94 showed strong inhibition against pathogenic and spoilage bacteria. There were three patterns of inhibition among three isolates against Bacillus subtilis, Staphylococcus aureus, Eschericia coli, and Salmonella sp. All three isolates showed potential uses for produce starter culture for fishery product fermentation purpose. This is the first report of isolation lactic acid bacteria that produced protease and bacteriocin-like from digestive tract of mud crab. Keywords: Lactic acid bacteria, Bacteriocin-like substance, Protease, Scylla  sp.

  13. Characterization of Bacteriocin like inhibitory substance produced by a new Strain Brevibacillus borstelensis AG1 Isolated from 'Marcha'

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    Nivedita Sharma

    2014-09-01

    Full Text Available In the present study, a bacterium isolated from Marcha- a herbal cake used as traditional starter culture to ferment local wine in North East India, was evaluated for bacteriocin like inhibitory substance production and was tested against six food borne/spoilage causing pathogens viz. Listeria monocytogenes MTCC 839, Bacillus subtilis MTCC 121, Clostridium perfringens MTCC 450, Staphylococcus aureus, Lactobacillus plantarum and Leuconostoc mesenteroides MTCC 107 by using bit/disc method followed by well diffusion method. The bacterial isolate was identified as Brevibacillus borstelensis on the basis of phenotypic, biochemical and molecular characteristics using 16Sr RNA gene technique. Bacteriocin like inhibitory substance produced by Brevibacillus borstelensis AG1 was purified by gel exclusion chromatography. The molecular mass of the Brevibacillus borstelensis AG1 was found to be 12 kDa. Purified bacteriocin like inhibitory substance of Brevibacillus borstelensis was further characterized by studying the effect of temperature, pH, proteolytic enzyme and stability. Bacteriocin like inhibitory substance was found to be thermostable upto 100 °C, active at neutral pH, sensitive to trypsin, and partially stable till third week of storage thus showing a bright prospective to be used as a potential food biopreservative.

  14. Bacteriocin-like substance (BLS) production in Aeromonas hydrophila water isolates.

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    Messi, Patrizia; Guerrieri, Elisa; Bondi, Moreno

    2003-03-14

    30 Aeromonas hydrophila water isolates were tested for bacteriocin-like substance (BLS) production using a target panel of closely related microorganisms and other Gram-positive and Gram-negative bacteria, including food-borne pathogens. A. hydrophila showed antibacterial activity against one or more indicator microorganisms, but the activity emerged only with non-phylogenetically related genera or species. In particular all A. hydrophila showed antibacterial activity against one or more of the tested Staphylococcus strains, five against Listeria spp. (Listeria seeligeri, Listeria welshimeri and Listeria ivanovii), and eight presented a weak antagonistic activity towards Streptococcus agalactiae and Lactobacillus spp. Inhibitory activity was not observed against the other Gram-positive (Listeria monocytogenes, Listeria innocua and Enterococcus spp.) and Gram-negative tested strains, including Aeromonas sobria, Aeromonas caviae and the same A. hydrophila, when used as indicator. Anti-staphylococcal activity was observed with a gradual increase of the inhibition zone during incubation and seemed to be influenced by A. hydrophila hemolytic expression. Extrachromosomal analysis showed the presence, in 70% of the strains, of one to five plasmids with molecular masses ranging from 2.1 to 41.5 MDa, but it was not possible to relate this result with BLS production.

  15. Identification of a new Bacillus licheniformis strain producing a bacteriocin-like substance.

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    Guo, Yaoqi; Yu, Zhanqiao; Xie, Jianhua; Zhang, Rijun

    2012-06-01

    The emergence of antibiotic resistance has spurred a great number of studies for development of new antimicrobials in the past decade. The purpose of this study was to screen environmental samples for Bacillus strains producing potent antimicrobial agents. A new strain, which showed strong antimicrobial activity against Staphylococcus aureus and Salmonella enterica ser. Pullorum, was isolated from soil and designated as B116. This new isolate was identified as Bacillus licheniformis by morphological, biochemical and genetic analyses. The production of bacteriocin-like substance (BLS) started at early exponential phase and achieved highest level at early stationary phase. The BLS was precipitated by ammonium sulfate and its molecular mass was determined as ∼4 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Culture supernatant of the new isolate exhibited antimicrobial activity against both Gram-positive and Gram-negative bacteria, including Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Micrococcus luteus, Escherichia coli, and Salmonella spp. The BLS was resistant to heat, acid and alkaline treatment. Activity of the BLS was totally lost after digestion by pronase and partially lost after digestion by papain and lipase. The new isolate and relevant BLS are potentially useful in food and feed applications.

  16. Bacteriocin-like inhibitory activities of seven Lactobacillus delbrueckii subsp. bulgaricus strains against antibiotic susceptible and resistant Helicobacter pylori strains.

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    Boyanova, L; Gergova, G; Markovska, R; Yordanov, D; Mitov, I

    2017-12-01

    The aim of the study was to detect anti-Helicobacter pylori activity of seven Lactobacillus delbrueckii subsp. bulgaricus (GLB) strains by four cell-free supernatant (CFS) types. Activity of non-neutralized and non-heat-treated (CFSs1), non-neutralized and heat-treated (CFSs2), pH neutralized, catalase-treated and non-heat-treated (CFSs3), or neutralized, catalase- and heat-treated (CFSs4) CFSs against 18 H. pylori strains (11 of which with antibiotic resistance) was evaluated. All GLB strains produced bacteriocin-like inhibitory substances (BLISs), the neutralized CFSs of two GLB strains inhibited >81% of test strains and those of four GLB strains were active against >71% of antibiotic resistant strains. Two H. pylori strains were BLIS resistant. The heating did not reduce the CFS activity. Briefly, all GLB strains evaluated produced heat-stable BLISs, although GLB and H. pylori strain susceptibility patterns exhibited differences. Bacteriocin-like inhibitory substance activity can be an advantage for the probiotic choice for H. pylori infection control. In this study, anti-Helicobacter pylori activity of seven Lactobacillus delbrueckii subsp. bulgaricus (GLB) strains was evaluated by four cell-free supernatant (CFS) types. The GLB strains produced heat-stable bacteriocin-like inhibitory substances (BLISs) with a strong anti-H. pylori activity and some neutralized, catalase- and heat-treated CFSs inhibited >83% of the test strains. Bacteriocin-like inhibitory substance production of GLB strains can render them valuable probiotics in the control of H. pylori infection. © 2017 The Society for Applied Microbiology.

  17. Antimicrobial activity and partial characterization of bacteriocin-like inhibitory substances produced by Lactobacillus spp. isolated from artisanal Mexican cheese.

    Science.gov (United States)

    Heredia-Castro, Priscilia Y; Méndez-Romero, José I; Hernández-Mendoza, Adrián; Acedo-Félix, Evelia; González-Córdova, Aarón F; Vallejo-Cordoba, Belinda

    2015-12-01

    Lactobacillus spp. from Mexican Cocido cheese were shown to produce bacteriocin-like substances (BLS) active against Staphylococcus aureus,Listeria innocua,Escherichia coli, andSalmonella typhimurium by using the disk diffusion method. Crude extracts of Lactobacillus fermentum showed strong inhibitory activity against Staph. aureus, L. innocua, E. coli, and Salmonella cholerae. Complete inactivation of antimicrobial activity was observed after treatment of crude extracts with proteinase K, pronase, papain, trypsin, and lysozyme, confirming their proteinaceous nature. However, antimicrobial activity was partly lost for some of the crude extracts when treated with α-amylase, indicating that carbohydrate moieties were involved. The antimicrobial activity of the crude extracts was stable at 65°C for 30min over a wide pH range (2-8), and addition of potassium chloride, sodium citrate, ethanol, and butanol did not affect antibacterial activity. However, antimicrobial activity was lost after heating at 121°C for 15min, addition of methanol or Tween 80. Fourteen out of 18 Lactobacillus spp. showed antimicrobial activity against different test microorganisms, and 12 presented bacteriocin-like substances. Generation time and growth rate parameters indicated that the antimicrobial activity of crude extracts from 3 different strains was effective against the 4 indicator microorganisms. One of the crude extracts showed inhibition not only against gram-positive but also against gram-negative bacteria. Bacteriocin-like substances produced by this specific Lactobacillus strain showed potential for application as a food biopreservative. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  18. Purification and characterization of bacteriocin like substance produced from bacillus lentus with perspective of a new biopreservative for food preservation

    International Nuclear Information System (INIS)

    Sharma, N.; Gautam, N.

    2009-01-01

    Molecular weight of bacteriocin like substance (BLIS) of a new strain of Bacillus lentus 121 was found to be approximately 11 kDa. Purification of BLIS was attained by single step gel exclusion chromatography. BLIS was characterized by studying the inhibitory spectrum. It was active at broad pH range, high temperature and high NaCl concentration and showed sensitivity to proteolytic enzymes like trypsin, alpha-chymotrypsin and papain, the characters desirable for food preservation. BLIS extended the shelf stability of milk upto 21 days as a biopreservative. (author)

  19. Detection and partial characterization of a bacteriocin-like substance produced by Lactobacillus fermentum CS57 isolated from human vaginal secretions.

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    Sabia, Carla; Anacarso, Immacolata; Bergonzini, Alberto; Gargiulo, Raffaele; Sarti, Mario; Condò, Carla; Messi, Patrizia; de Niederhausern, Simona; Iseppi, Ramona; Bondi, Moreno

    2014-04-01

    Lactobacilli (150) from human vaginal secretions were tested for the production of antimicrobial substances which can provide a physiological defense against the pathogenic microorganisms in the vaginal area. Sixteen of the isolates (10.6%) showed antibacterial activity against one or several closely related microorganisms used as indicators. Lactobacillus fermentum CS57 was the best producer and secretes a bacteriocin-like substance (BLS) with antagonistic activity against Streptococcus agalactiae and Candida albicans. The compound was susceptible to the proteolytic enzymes and was heat labile. The mode of action was identified as bactericidal. The crude activity of the L. fermentum CS57 BLS was linked to a substance with a molecular weight larger than 30 kDa. Plasmid analysis of L. fermentum CS57 revealed the presence of a plasmid band with molecular weight of 54.7 kb. All L. fermentum CS57 non-producer variants (BLS-) obtained by curing experiments, showed loss of plasmid band and were susceptible to the BLS of the original strain. Therefore antimicrobial activity and immunity production seem to be linked to genes located on that same plasmid. Taking into account our results, L. fermentum CS57 could be considered a candidate for potential use as probiotic for the prophylaxis of vaginal human infections. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. POTENTIAL IN VITRO ANTI-HELICOBACTER ACTIVITY OF BACTERIOCIN AND BACTERIOCIN-LIKE COMPOUNDS PRODUCED BY LACTOBACILLI

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    Mohammed A. Ramadan

    2014-10-01

    Full Text Available The study was designed for screening of the potential activity of lactic acid bacteria against Helicobacter pylori and other enteropathogenic organisms. A total of 40 samples including natural cow milk and fresh infant stools were tested for the presence of lactic acid bacteria. Of these samples, 73 lactic acid bacterial isolates were recovered on MRS agar medium using the streak-plate method. Isolates inducing probiotic effect were tested under microaerophilic conditions against standard cultures of H. pylori, Esherichia coli and Salmonella enteritidis. The data obtained showed that five isolates of lactic acid bacteria were able to produce bacteriocin or bacteriocin-like compounds. Sequencing of 16S rRNA gene revealed that five isolates belonged to Lactobacillus rhamnosus and Lactobacillus plantarum in addition to other lactic acid bacteria. The most effective isolate (LAB1 showed a marked large inhibition zone against H. pylori. The bacteriocin or bacteriocin like compound(s produced by lactobacilli were further analyzed and characterized. We can conclude that probiotics might be useful in the prophylaxis or as co-therapy for treatment of H. pylori infections.

  1. Mutacins and bacteriocins like genes in Streptococcus mutans isolated from participants with high, moderate, and low salivary count.

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    Soto, Carolina; Padilla, Carlos; Lobos, Olga

    2017-02-01

    To detect S. mutans producers of mutacins and bacteriocins like substances (BLIS) from saliva of participants with low, moderate, and high salivary counts. 123 strains of S. mutans were obtained from participants with low, moderate, and high salivary counts (age 18 and 20 years old) and their antibacterial capacity analyzed. By using PCR amplification, the expression levels of mutacins and BLIS genes were studied (expressed in arbitrary units/ml) in all three levels. S. mutans strains from participants with low salivary counts show high production of mutacins (63%). In contrast, participants with moderate and high salivary counts depict relatively low levels of mutacins (22 and 15%, respectively). Moreover, participants with low salivary counts showed high expression levels of genes encoding mutacins, a result that correlates with the strong antimicrobial activity of the group. Participants with moderate and high salivary counts however depict low expression levels of mutacin related genes, and little antimicrobial activity. No BLIS were detected in any of the groups studied. S. mutans isolated from the saliva of participants with low bacterial counts have significant antibacterial capacity compared to that of participants with moderate and high salivary counts. The superior lethality of S. mutans in participants with low salivary counts is likely due to the augmented expression of mutacin- related genes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Characterisation of two novel bacteriocin-like substances produced by Bacillus amyloliquefaciens ELI149 with broad-spectrum antimicrobial activity.

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    Salazar, Francisco; Ortiz, Aurelio; Sansinenea, Estibaliz

    2017-12-01

    The aim of this study was to isolate and characterise antifungal and bactericidal compounds from Bacillus amyloliquefaciens strain ELI149. An absorbent resin (Amberlite ® XAD-16) and silica gel column chromatography were used for isolation and purification purposes, respectively. Antibacterial and antifungal assays were performed by the well diffusion method to demonstrate the biological activity of each compound. Cell damage of the tested fungi was evaluated for fengycin under phase-contrast microscopy. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectroscopy techniques were performed to estimate the approximate molecular mass of each compound. Two bacteriocin-like substances (BLSs) with different physical properties and inhibitory activities were isolated along with two known antifungal compounds. The two BLSs were heat stable and were not sensitive to acid or alkaline conditions (pH 2-10), with broad-spectrum antimicrobial activity. The antifungal compounds were identified as surfactin and fengycin. Only fengycin showed marked antifungal properties against several phytopathogens. The two isolated BLSs were partially characterised and their bactericidal properties were analysed. The antifungals compounds were identified as surfactin and fengycin, this latter being mainly responsible for the antifungal activity. Copyright © 2017 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

  3. Identification and partial characterization of a bacteriocin-like inhibitory substance (BLIS) from Lb. Bulgaricus K41 isolated from indigenous yogurts.

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    Zaeim, Davood; Soleimanian-Zad, Sabihe; Sheikh-Zeinoddin, Mahmoud

    2014-01-01

    Forty-two strains of Lactobacillus bulgaricus isolated from locally made yogurts were examined and compared for bacteriocin producing ability using spot on lawn assay which improved by taking photo and image processing. Lb. bulgaricus K41 exhibited the highest inhibition level against indicators. K41 Bacteriocin-like inhibitory substance is sensitive to proteolytic enzymes (proteinase K, pepsin, and trypsin) but α-amylase makes slight reduction in its activity and it is resistant to lipase. This antibacterial peptide is extremely heat-stable (121 °C for 15 min) and remains active over a wide pH range (pH = 2 to 10); also nonionic detergents (Tween-20, Tween-80, and Triton X100) showed no effect on its activity. The inhibitory spectrum is against Gram-positive bacteria (except Staphylococcus aureus) with extremely antilisterial activity and it is almost ineffective against Gram-negative bacteria. The mode of its action was identified as bactericidal against Listeria monocytogenes. The properties of K41 bacteriocin-like inhibitory substance add to its safety as a biopreservative produced by a generally recognized as safe (GRAS) bacterium suggesting it can be used in hurdle technology for ready-to-eat foods as one of the main sources of Listeria contaminations. © 2013 Institute of Food Technologists®

  4. Improving microphage innate immunity by modulating protein ...

    International Development Research Centre (IDRC) Digital Library (Canada)

    Improving microphage innate immunity by modulating protein tyrosine phosphatases: The complete mouse and human PTPomes. Diseases that result from an infection are most often resolved by cells that use an immune response to clear foreign agents. These cells include macrophages, which are the predominant type of ...

  5. Partial characterization of bacteriocin-like compounds from two strains of Bacillus cereus with biological activity against Paenibacillus larvae, the causal agent of American Foulbrood disease.

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    Minnaard, J; Alippi, A M

    2016-12-01

    American Foulbrood (AFB), caused by the spore-forming Gram-positive bacterium Paenibacillus larvae, is the most severe bacterial disease affecting honeybees worldwide. Two bacterial isolates showing specific inhibitory activity against P. larvae were identified as Bacillus cereus by 16S rDNA sequencing. Antagonistic compounds were obtained from cell-free supernatants of strains m6c and m387 growing on Trypticase Soy Broth and concentrated by NH 4 SO 4 precipitation, ultrafiltration and butanol extraction. Both compounds were characterized as bacteriocin-like inhibitory substances (BLIS). BLISm6c and BLISm387 were stable at 70°C for 30 min and active in the pH range from 3 to 7. The antibacterial activity was completely lost at pH values higher than 8 or temperatures >80°C. Both BLIS have a narrow activity range and highly inhibit the growth of P. larvae. BLISm6c and BLISm387 differ from each other and other BLIS reportedly produced by B. cereus with regard to their molecular weights, antibacterial activity, minimal inhibitory concentration values and sensitivity to degradative enzymes. The findings of this study suggest that BLISm6c and BLISm387 can potentially be used to control AFB. An Integrated Pest Management (IPM) approach is needed to ensure the sustainability of the beekeeping industry due to the increasing demand for organic honey and the reduction of dependence on antibiotics. Biocontrol agents produced by bacteria isolated from apiarian sources seem promising and able to combine with an IPM strategy. The most significant findings of this study are the characterization of bacteriocin-like compounds (BLIS) obtained from two strains of Bacillus cereus isolated from honey. Both BLIS have a narrow activity range and highly inhibit the growth of Paenibacillus larvae, the causal agent of American Foulbrood disease of honey bees. © 2016 The Society for Applied Microbiology.

  6. Immune evasion by gammaherpesvirus genome maintenance proteins.

    Science.gov (United States)

    Blake, Neil

    2010-04-01

    Viruses that establish lifelong latent infections must ensure that the viral genome is maintained within the latently infected cell throughout the life of the host, yet at the same time must also be capable of avoiding elimination by the immune surveillance system. Gammaherpesviruses, which include the human viruses Epstein-Barr virus and Kaposi's sarcoma-associated herpesvirus, establish latent infections in lymphocytes. Infection of this dynamic host-cell population requires that the viruses have appropriate strategies for enabling the viral genome to persist while these cells go through rounds of mitosis, but at the same time must avoid detection by host CD8(+) cytotoxic T lymphocytes (CTLs). The majority of gammaherpesviruses studied have been found to encode a specific protein that is critical for maintenance of the viral genome within latently infected cells. This protein is termed the genome maintenance protein (GMP). Due to its vital role in long-term latency, this offers the immune system a crucial target for detection and elimination of virus-infected cells. GMPs from different gammaherpesviruses have evolved related strategies that allow the protein to be present within latently infected cells, but to remain effectively hidden from circulating CD8(+) CTLs. In this review, I will summarize the role of the GMPs and highlight the available data describing the immune-evasion properties of these proteins.

  7. Effect of liposome-encapsulated nisin and bacteriocin-like substance P34 on Listeria monocytogenes growth in Minas frescal cheese.

    Science.gov (United States)

    Malheiros, Patrícia da Silva; Sant'Anna, Voltaire; Barbosa, Matheus de Souza; Brandelli, Adriano; Franco, Bernadette Dora Gombossy de Melo

    2012-06-01

    The efficacy of liposome-encapsulated nisin and bacteriocin-like substance (BLS) P34 to control growth of Listeria monocytogenes in Minas frescal cheese was investigated. Nisin and BLS P34 were encapsulated in partially purified soybean phosphatidylcholine (PC-1) and PC-1-cholesterol (7:3) liposomes. PC-1 nanovesicles were previously characterized. PC-1-cholesterol encapsulated nisin and BLS P34 presented, respectively, 218 nm and 158 nm diameters, zeta potential of -64 mV and -53 mV, and entrapment efficiency of 88.9% and 100%. All treatments reduced the population of L. monocytogenes compared to the control during 21 days of storage of Minas frescal cheese at 7°C. However, nisin and BLS P34 encapsulated in PC-1-cholesterol liposomes were less efficient in controlling L. monocytogenes growth in comparison with free and PC-1 liposome-encapsulated bacteriocins. The highest inhibitory effect was observed for nisin and BLS P34 encapsulated in PC-1 liposomes after 10 days of storage of the product. The encapsulation of bacteriocins in liposomes of partially purified soybean phosphatidylcholine may be a promising technology for the control of foodborne pathogens in cheeses. Copyright © 2012 Elsevier B.V. All rights reserved.

  8. Isolation of Pediococcus acidilactici Kp10 with ability to secrete bacteriocin-like inhibitory substance from milk products for applications in food industry

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    Abbasiliasi Sahar

    2012-11-01

    Full Text Available Abstract Background Lactic acid bacteria (LAB can be isolated from traditional milk products. LAB that secrete substances that inhibit pathogenic bacteria and are resistant to acid, bile, and pepsin but not vancomycin may have potential in food applications. Results LAB isolated from a range of traditional fermented products were screened for the production of bacteriocin-like inhibitory substances. A total of 222 LAB strains were isolated from fermented milk products in the form of fresh curds, dried curds, and ghara (a traditional flavor enhancer prepared from whey, and fermented cocoa bean. Eleven LAB isolates that produced antimicrobial substances were identified as Lactococcus lactis, Lactobacillus plantarum, and Pediococcus acidilactici strains by biochemical methods and 16S rDNA gene sequencing. Of these, the cell-free supernatant of Kp10 (P. acidilactici most strongly inhibited Listeria monocytogenes. Further analysis identified the antimicrobial substance produced by Kp10 as proteinaceous in nature and active over a wide pH range. Kp10 (P. acidilactici was found to be catalase-negative, able to produce β-galactosidase, resistant to bile salts (0.3% and acidic conditions (pH 3, and susceptible to most antibiotics. Conclusion Traditionally prepared fermented milk products are good sources of LAB with characteristics suitable for industrial applications. The isolate Kp10 (P. acidilactici shows potential for the production of probiotic and functional foods.

  9. Functional Classification of Immune Regulatory Proteins

    Energy Technology Data Exchange (ETDEWEB)

    Rubinstein, Rotem [Albert Einstein College of Medicine, Bronx, NY (United States); Ramagopal, Udupi A. [Albert Einstein College of Medicine, Bronx, NY (United States); Nathenson, Stanley G. [Albert Einstein College of Medicine, Bronx, NY (United States); Almo, Steven C. [Albert Einstein College of Medicine, Bronx, NY (United States); Fiser, Andras [Albert Einstein College of Medicine, Bronx, NY (United States)

    2013-05-01

    Members of the immunoglobulin superfamily (IgSF) control innate and adaptive immunity and are prime targets for the treatment of autoimmune diseases, infectious diseases, and malignancies. We describe a computational method, termed the Brotherhood algorithm, which utilizes intermediate sequence information to classify proteins into functionally related families. This approach identifies functional relationships within the IgSF and predicts additional receptor-ligand interactions. As a specific example, we examine the nectin/nectin-like family of cell adhesion and signaling proteins and propose receptor-ligand interactions within this family. We were guided by the Brotherhood approach and present the high-resolution structural characterization of a homophilic interaction involving the class-I MHC-restricted T-cell-associated molecule, which we now classify as a nectin-like family member. The Brotherhood algorithm is likely to have a significant impact on structural immunology by identifying those proteins and complexes for which structural characterization will be particularly informative.

  10. Identifying Bacterial Immune Evasion Proteins Using Phage Display.

    Science.gov (United States)

    Fevre, Cindy; Scheepmaker, Lisette; Haas, Pieter-Jan

    2017-01-01

    Methods aimed at identification of immune evasion proteins are mainly rely on in silico prediction of sequence, structural homology to known evasion proteins or use a proteomics driven approach. Although proven successful these methods are limited by a low efficiency and or lack of functional identification. Here we describe a high-throughput genomic strategy to functionally identify bacterial immune evasion proteins using phage display technology. Genomic bacterial DNA is randomly fragmented and ligated into a phage display vector that is used to create a phage display library expressing bacterial secreted and membrane bound proteins. This library is used to select displayed bacterial secretome proteins that interact with host immune components.

  11. [The role of protein glycosylation in immune system].

    Science.gov (United States)

    Ząbczyńska, Marta; Pocheć, Ewa

    2015-01-01

    Glycosylation is one of the most frequent post-translational modifications of proteins. The majority of cell surface and secreted proteins involved in immune response is glycosylated. The structural diversity of glycans depends on monosaccharide composition, type of glycosidic linkage and branching. These structural modifications determine a great variability of glycoproteins. The oligosaccharide components of proteins are regulated mostly by expression of glycosyltransferases and glycosidases and many environmental factors. Glycosylation influences the function of all immune cells. Glycans play a crucial role in intercellular contacts and leukocytes migration. These interactions are important in activation and proliferation of leukocytes and during immune response. The key immune proteins, such as TCR, MHC, TLR and antibodies are glycosylated. Sugars on the surface of pathogens and self-surface glycoproteins are recognized by special carbohydrate binding proteins called lectins. Changes of glycan structure are common in many pathological processes occurring in immune system, therefore they are used as molecular markers of different diseases.

  12. Circumvention of Immunity to the Adenovirus Major Coat Protein Hexon

    Science.gov (United States)

    Roy, Soumitra; Shirley, Pamela S.; McClelland, Alan; Kaleko, Michael

    1998-01-01

    Immunity to adenoviruses is an important hurdle to be overcome for successful gene therapy. The presence of antibodies to the capsid proteins prevents efficacious adenovirus vector administration in vivo. We tested whether immunity to a particular serotype of adenovirus (Ad5) may be overcome with a vector that encodes the hexon sequences from a different adenovirus serotype (Ad12). We successfully constructed an adenovirus vector with a chimeric Ad5-Ad12 hexon which was not neutralized by plasma from C57BL/6 mice immunized with Ad5. The vector was also capable of transducing the livers of C57BL/6 mice previously immunized with Ad5. PMID:9658137

  13. Immune modulation by the hepatitis C virus core protein.

    Science.gov (United States)

    Fernández-Ponce, C; Dominguez-Villar, M; Muñoz-Miranda, J P; Arbulo-Echevarria, M M; Litrán, R; Aguado, E; García-Cozar, F

    2017-05-01

    Hepatitis C virus (HCV) infection is currently the most important cause of chronic viral hepatitis in the world and one of the most frequent indications for liver transplantation. HCV uses different strategies to evade the innate and adaptive immune response, and this evasion plays a key role in determining viral persistence. Several HCV viral proteins have been described as immune modulators. In this review, we will focus on the effect of HCV nucleocapsid core protein in the function of immune cells and its correlation with the findings observed in HCV chronically infected patients. Effects on immune cell function related to both extracellular and intracellular HCV core localization will be considered. This review provides an updated perspective on the mechanisms involved in HCV evasion related to one single HCV protein, which could become a key tool in the development of new antiviral strategies able to control and/or eradicate HCV infection. © 2017 John Wiley & Sons Ltd.

  14. Identifying bacterial immune evasion proteins using phage display

    NARCIS (Netherlands)

    Fevre, Cindy; Scheepmaker, Lisette; Haas, Pieter Jan

    2017-01-01

    Methods aimed at identification of immune evasion proteins are mainly rely on in silico prediction of sequence, structural homology to known evasion proteins or use a proteomics driven approach. Although proven successful these methods are limited by a low efficiency and or lack of functional

  15. Immune regulation of Rab proteins expression and intracellular transport.

    Science.gov (United States)

    Pei, Gang; Bronietzki, Marc; Gutierrez, Maximiliano Gabriel

    2012-07-01

    Compartmentalization in cells of the immune system, the focus of this review, facilitates the spatiotemporal organization of cellular responses essential for specialized immune functions. In this process of compartment maintenance, Rab proteins are central regulators of protein-mediated transport and fusion of intracellular structures. It is widely believed that the intracellular concentration of proteins that regulate intracellular transport, including Rab proteins, is constitutively mantained. However, there is a growing body of evidence indicating that transcriptional rates of Rab proteins can be modified. This process is especially evident during immune activation and argues that after activation, these cells require higher levels of Rab proteins. The aim of this review is to discuss evidence showing the increasing links between Rab protein expression and intracellular transport, particularly in monocytes and macrophages. We highlight here biological processes in which the expression of Rab GTPases is selectively regulated, leading to the activation of specific intracellular routes. Further, we focus on the immune regulation of intracellular transport after cytokine activation and microbial infection, with an emphasis in mycobacterial infection.

  16. The immune response to sand fly salivary proteins and its influence on Leishmania immunity

    Directory of Open Access Journals (Sweden)

    Regis eGomes

    2012-05-01

    Full Text Available Leishmaniasis is a vector-borne disease transmitted by bites of phlebotomine sand flies. During Leishmania transmission, sand fly saliva is co-inoculated with parasites into the skin of the mammalian host. Sand fly saliva consists of roughly thirty different salivary proteins, many with known roles linked to blood feeding facilitation. Apart from the anti-hemostatic capacity of saliva, several sand fly salivary proteins have been shown to be immunogenic upon multiple contacts with a mammalian host. Immunization with single immunogenic salivary proteins or exposure to uninfected bites can produce protective immune responses against leishmaniasis. These sand fly salivary proteins induce cellular immune responses and/or antibodies. Antibodies to saliva are not required for protection in a mouse model against leishmaniasis. A strong body of evidence points to the role for saliva-specific T cells producing IFN-γ in the form of a delayed-type hypersensitivity reaction at the bite site as the main protective response. Herein, we review immunity to sand fly salivary proteins in the context of its vector-parasite-host combinations and vaccine potential, as well as some recent advances to shed light on the mechanism of how an immune response to sand fly saliva protects against leishmaniasis.

  17. Cooperative Immune Suppression by Escherichia coli and Shigella Effector Proteins.

    Science.gov (United States)

    de Jong, Maarten F; Alto, Neal M

    2018-04-01

    The enteric attaching and effacing (A/E) pathogens enterohemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC) and the invasive pathogens enteroinvasive E. coli (EIEC) and Shigella encode type III secretion systems (T3SS) used to inject effector proteins into human host cells during infection. Among these are a group of effectors required for NF-κB-mediated host immune evasion. Recent studies have identified several effector proteins from A/E pathogens and EIEC/ Shigella that are involved in suppression of NF-κB and have uncovered their cellular and molecular functions. A novel mechanism among these effectors from both groups of pathogens is to coordinate effector function during infection. This cooperativity among effector proteins explains how bacterial pathogens are able to effectively suppress innate immune defense mechanisms in response to diverse classes of immune receptor signaling complexes (RSCs) stimulated during infection. Copyright © 2018 American Society for Microbiology.

  18. Innate Immune Evasion Mediated by Flaviviridae Non-Structural Proteins.

    Science.gov (United States)

    Chen, Shun; Wu, Zhen; Wang, Mingshu; Cheng, Anchun

    2017-10-07

    Flaviviridae-caused diseases are a critical, emerging public health problem worldwide. Flaviviridae infections usually cause severe, acute or chronic diseases, such as liver damage and liver cancer resulting from a hepatitis C virus (HCV) infection and high fever and shock caused by yellow fever. Many researchers worldwide are investigating the mechanisms by which Flaviviridae cause severe diseases. Flaviviridae can interfere with the host's innate immunity to achieve their purpose of proliferation. For instance, dengue virus (DENV) NS2A, NS2B3, NS4A, NS4B and NS5; HCV NS2, NS3, NS3/4A, NS4B and NS5A; and West Nile virus (WNV) NS1 and NS4B proteins are involved in immune evasion. This review discusses the interplay between viral non-structural Flaviviridae proteins and relevant host proteins, which leads to the suppression of the host's innate antiviral immunity.

  19. Polyclonal Antibody Production for Membrane Proteins via Genetic Immunization.

    Science.gov (United States)

    Hansen, Debra T; Robida, Mark D; Craciunescu, Felicia M; Loskutov, Andrey V; Dörner, Katerina; Rodenberry, John-Charles; Wang, Xiao; Olson, Tien L; Patel, Hetal; Fromme, Petra; Sykes, Kathryn F

    2016-02-24

    Antibodies are essential for structural determinations and functional studies of membrane proteins, but antibody generation is limited by the availability of properly-folded and purified antigen. We describe the first application of genetic immunization to a structurally diverse set of membrane proteins to show that immunization of mice with DNA alone produced antibodies against 71% (n = 17) of the bacterial and viral targets. Antibody production correlated with prior reports of target immunogenicity in host organisms, underscoring the efficiency of this DNA-gold micronanoplex approach. To generate each antigen for antibody characterization, we also developed a simple in vitro membrane protein expression and capture method. Antibody specificity was demonstrated upon identifying, for the first time, membrane-directed heterologous expression of the native sequences of the FopA and FTT1525 virulence determinants from the select agent Francisella tularensis SCHU S4. These approaches will accelerate future structural and functional investigations of therapeutically-relevant membrane proteins.

  20. Staphylococcal Immune Evasion Proteins: Structure, Function, and Host Adaptation.

    Science.gov (United States)

    Koymans, Kirsten J; Vrieling, Manouk; Gorham, Ronald D; van Strijp, Jos A G

    2017-01-01

    Staphylococcus aureus is a successful human and animal pathogen. Its pathogenicity is linked to its ability to secrete a large amount of virulence factors. These secreted proteins interfere with many critical components of the immune system, both innate and adaptive, and hamper proper immune functioning. In recent years, numerous studies have been conducted in order to understand the molecular mechanism underlying the interaction of evasion molecules with the host immune system. Structural studies have fundamentally contributed to our understanding of the mechanisms of action of the individual factors. Furthermore, such studies revealed one of the most striking characteristics of the secreted immune evasion molecules: their conserved structure. Despite high-sequence variability, most immune evasion molecules belong to a small number of structural categories. Another remarkable characteristic is that S. aureus carries most of these virulence factors on mobile genetic elements (MGE) or ex-MGE in its accessory genome. Coevolution of pathogen and host has resulted in immune evasion molecules with a highly host-specific function and prevalence. In this review, we explore how these shared structures and genomic locations relate to function and host specificity. This is discussed in the context of therapeutic options for these immune evasion molecules in infectious as well as in inflammatory diseases.

  1. Bacteriocin-like activity of oral Fusobacterium nucleatum isolated from human and non-human primates Atividade semelhante a bacteriocina de Fusobacterium nucleatum orais isolados de primatas humanos e não-humanos

    Directory of Open Access Journals (Sweden)

    Elerson Gaetti-Jardim Júnior

    1999-12-01

    Full Text Available Fusobacterium nucleatum is indigenous of the human oral cavity and has been involved in different infectious processes. The production of bacteriocin-like substances may be important in regulation of bacterial microbiota in oral cavity. The ability to produce bacteriocin-like substances by 80 oral F. nucleatum isolates obtained from periodontal patients, healthy individuals and Cebus apella monkeys, was examinated. 17.5% of all tested isolates showed auto-antagonism and 78.8% iso- or hetero-antagonism. No isolate from monkey was capable to produce auto-inhibition. In this study, the antagonistic substances production was variable in all tested isolates. Most of the F. nucleatum showed antagonistic activity against tested reference strains. These data suggest a possible participation of these substances on the oral microbial ecology in humans and animals. However, the role of bacteriocins in regulating dental plaque microbiota in vivo is discussed.Fusobacterium nucleatum é indígena da cavidade oral humana e tem sido envolvido em diferentes processos infecciosos. A produção de substâncias semelhantes a bacteriocinas pode ser importante na regulação da microbiota bacteriana da cavidade oral. A capacidade de produzir substâncias tipo bacteriocina de 80 isolados de F. nucleatum orais, obtidos de pacientes com doença periodontal, indivíduos sadios e macaco Cebus apella, foi avaliada. 17,5% de todos os isolados mostrou auto-antagonismo e 78,8% iso- ou hetero-antagonismo. Nenhum isolado de macaco foi capaz de produzir auto-inibição. Neste estudo, a produção de substâncias antagonístas foi variável em todos os isolados testados. A maioria dos F. nucleatum mostrou atividade antagonísta para as cepas de referência testadas. Esses dados sugerem a possível participação dessas substâncias sobre a ecologia microbiana em humanos e animais. Entretanto, o papel das bacteriocinas na regulação da microbiota da placa dental in vivo

  2. Alkaline Phosphatase, an Unconventional Immune Protein

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    Bethany A. Rader

    2017-08-01

    Full Text Available Recent years have seen an increase in the number of studies focusing on alkaline phosphatases (APs, revealing an expanding complexity of function of these enzymes. Of the four human AP (hAP proteins, most is known about tissue non-specific AP (TNAP and intestinal AP (IAP. This review highlights current understanding of TNAP and IAP in relation to human health and disease. TNAP plays a role in multiple processes, including bone mineralization, vitamin B6 metabolism, and neurogenesis, is the genetic cause of hypophosphatasia, influences inflammation through regulation of purinergic signaling, and has been implicated in Alzheimer’s disease. IAP regulates fatty acid absorption and has been implicated in the regulation of diet-induced obesity and metabolic syndrome. IAP and TNAP can dephosphorylate bacterial-derived lipopolysaccharide, and IAP has been identified as a potential regulator of the composition of the intestinal microbiome, an evolutionarily conserved function. Endogenous and recombinant bovine APs and recombinant hAPs are currently being explored for their potential as pharmacological agents to treat AP-associated diseases and mitigate multiple sources of inflammation. Continued research on these versatile proteins will undoubtedly provide insight into human pathophysiology, biochemistry, and the human holobiont.

  3. Emerging Roles of Regulators of G Protein Signaling (RGS) Proteins in the Immune System.

    Science.gov (United States)

    Druey, Kirk M

    2017-01-01

    The regulators of G protein signaling (RGS) proteins are a large, evolutionarily conserved group of intracellular proteins expressed in every cell type and tissue throughout the body including the immune system. Through their signature GTPase-activating protein (GAP) activity on heterotrimeric G proteins and interactions with signaling complexes and membrane constituents (e.g., lipids), RGS proteins determine the intensity and duration of G protein-coupled receptor-induced responses. They may also have a function in generating intracellular signaling gradients necessary for the directional migration of leukocytes to inflamed tissues containing local accumulations of chemoattractants. Although physiological functions of most RGS proteins in leukocytes and lymphoid organs are largely unknown, it appears thus far that deficiency of individual RGS proteins in mice does not affect homeostatic immune responses in the absence of immunogenic challenge and/or microbial infection. Although aberrant expression of some RGS proteins has been linked to dysregulated immunity and/or neoplasia in humans, there are no human diseases attributed to specific RGS dysfunction. Here, we highlight mostly published work describing expression and functions of the core group of RGS proteins that were among the first discovered, in both innate and adaptive immune processes, with particular emphasis on cell trafficking. © 2017 Elsevier Inc. All rights reserved.

  4. Cross-serotype immunity induced by immunization with a conserved rhinovirus capsid protein.

    Directory of Open Access Journals (Sweden)

    Nicholas Glanville

    Full Text Available Human rhinovirus (RV infections are the principle cause of common colds and precipitate asthma and COPD exacerbations. There is currently no RV vaccine, largely due to the existence of ∼150 strains. We aimed to define highly conserved areas of the RV proteome and test their usefulness as candidate antigens for a broadly cross-reactive vaccine, using a mouse infection model. Regions of the VP0 (VP4+VP2 capsid protein were identified as having high homology across RVs. Immunization with a recombinant VP0 combined with a Th1 promoting adjuvant induced systemic, antigen specific, cross-serotype, cellular and humoral immune responses. Similar cross-reactive responses were observed in the lungs of immunized mice after infection with heterologous RV strains. Immunization enhanced the generation of heterosubtypic neutralizing antibodies and lung memory T cells, and caused more rapid virus clearance. Conserved domains of the RV capsid therefore induce cross-reactive immune responses and represent candidates for a subunit RV vaccine.

  5. Protein Kinase C δ: a Gatekeeper of Immune Homeostasis.

    Science.gov (United States)

    Salzer, Elisabeth; Santos-Valente, Elisangela; Keller, Bärbel; Warnatz, Klaus; Boztug, Kaan

    2016-10-01

    Human autoimmune disorders present in various forms and are associated with a life-long burden of high morbidity and mortality. Many different circumstances lead to the loss of immune tolerance and often the origin is suspected to be multifactorial. Recently, patients with autosomal recessive mutations in PRKCD encoding protein kinase c delta (PKCδ) have been identified, representing a monogenic prototype for one of the most prominent forms of humoral systemic autoimmune diseases, systemic lupus erythematosus (SLE). PKCδ is a signaling kinase with multiple downstream target proteins and with functions in various signaling pathways. Interestingly, mouse models have indicated a special role of the ubiquitously expressed protein in the control of B-cell tolerance revealed by the severe autoimmunity in Prkcd (-/-) knockout mice as the major phenotype. As such, the study of PKCδ deficiency in humans has tremendous potential in enhancing our knowledge on the mechanisms of B-cell tolerance.

  6. Immunization with truncated envelope protein of Zika virus induces protective immune response in mice.

    Science.gov (United States)

    Han, Jian-Feng; Qiu, Yang; Yu, Jiu-Yang; Wang, Hong-Jiang; Deng, Yong-Qiang; Li, Xiao-Feng; Zhao, Hui; Sun, Han-Xiao; Qin, Cheng-Feng

    2017-08-30

    The global spread of Zika virus (ZIKV) as well as its unexpected link to infant microcephaly have resulted in serious public health concerns. No antiviral drugs against ZIKV is currently available, and vaccine development is of high priority to prepare for potential ZIKV pandemic. In the present study, a truncated E protein with the N-terminal 90% region reserved (E90) from a contemporary ZIKV strain was cloned and expressed in Escherichia coli, purified by a Ni-NTA column, and characterized by Western blotting assays. Immunization with recombinant E90 induced robust ZIKV-specific humoral response in adult BALB/c mice. Passive transfer of the antisera from E90-immunized mice conferred full protection against lethal ZIKV challenge in a neonatal mice model. Our results indicate that recombinant ZIKV E90 described here represents as a promising ZIKV subunit vaccine that deserves further clinical development.

  7. Alphacoronavirus Protein 7 Modulates Host Innate Immune Response

    Science.gov (United States)

    Cruz, Jazmina L. G.; Becares, Martina; Sola, Isabel; Oliveros, Juan Carlos; Zúñiga, Sonia

    2013-01-01

    Innate immune response is the first line of antiviral defense resulting, in most cases, in pathogen clearance with minimal clinical consequences. Viruses have developed diverse strategies to subvert host defense mechanisms and increase their survival. In the transmissible gastroenteritis virus (TGEV) as a model, we previously reported that accessory gene 7 counteracts the host antiviral response by associating with the catalytic subunit of protein phosphatase 1 (PP1c). In the present work, the effect of the absence of gene 7 on the host cell, during infection, was further analyzed by transcriptomic analysis. The pattern of gene expression of cells infected with a recombinant mutant TGEV, lacking gene 7 expression (rTGEV-Δ7), was compared to that of cells infected with the parental virus (rTGEV-wt). Genes involved in the immune response, the interferon response, and inflammation were upregulated during TGEV infection in the absence of gene 7. An exacerbated innate immune response during infection with rTGEV-Δ7 virus was observed both in vitro and in vivo. An increase in macrophage recruitment and activation in lung tissues infected with rTGEV-Δ7 virus was observed compared to cells infected with the parental virus. In summary, the absence of protein 7 both in vitro and in vivo led to increased proinflammatory responses and acute tissue damage after infection. In a porcine animal model, which is immunologically similar to humans, we present a novel example of how viral proteins counteract host antiviral pathways to determine the infection outcome and pathogenesis. PMID:23824792

  8. R4 Regulator of G Protein Signaling (RGS) Proteins in Inflammation and Immunity.

    Science.gov (United States)

    Xie, Zhihui; Chan, Eunice C; Druey, Kirk M

    2016-03-01

    G protein-coupled receptors (GPCRs) have important functions in both innate and adaptive immunity, with the capacity to bridge interactions between the two arms of the host responses to pathogens through direct recognition of secreted microbial products or the by-products of host cells damaged by pathogen exposure. In the mid-1990s, a large group of intracellular proteins was discovered, the regulator of G protein signaling (RGS) family, whose main, but not exclusive, function appears to be to constrain the intensity and duration of GPCR signaling. The R4/B subfamily--the focus of this review--includes RGS1-5, 8, 13, 16, 18, and 21, which are the smallest RGS proteins in size, with the exception of RGS3. Prominent roles in the trafficking of B and T lymphocytes and macrophages have been described for RGS1, RGS13, and RGS16, while RGS18 appears to control platelet and osteoclast functions. Additional G protein independent functions of RGS13 have been uncovered in gene expression in B lymphocytes and mast cell-mediated allergic reactions. In this review, we discuss potential physiological roles of this RGS protein subfamily, primarily in leukocytes having central roles in immune and inflammatory responses. We also discuss approaches to target RGS proteins therapeutically, which represents a virtually untapped strategy to combat exaggerated immune responses leading to inflammation.

  9. Antagonism of Innate Immunity by Paramyxovirus Accessory Proteins

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    Raychel Chambers

    2009-10-01

    Full Text Available Paramyxovirinae, a subfamily of Paramyxoviridae, are negative strand RNA viruses comprised of many important human and animal pathogens, which share a high degree of genetic and structural homology. The accessory proteins expressed from the P/V/C gene are major factors in the pathogenicity of the viruses, because of their ability to abrogate various facets of type I interferon (IFN induction and signaling. Most of the paramyxoviruses exhibit a commonality in their ability to antagonize innate immunity by blocking IFN induction and the Jak/STAT pathway. However, the manner in which the accessory proteins inhibit the pathway differs among viruses. Similarly, there are variations in the capability of the viruses to counteract intracellular detectors (RNA helicases, mda-5 and RIG-I. Furthermore, a functional specificity in the antagonism of the IFN response has been reported, suggesting that specificity in the circumvention of innate immunity restricts viral host range. Available evidence indicates that paramyxoviruses employ specific strategies to antagonize the IFN response of their specific hosts, which is one of the major factors that determine viral pathogenicity and host range.

  10. SAPOSIN-LIKE PROTEINS IN ANTI-INFECTIOUS IMMUNE RESPONSE

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    V. V. Yeremeev

    2012-01-01

    Full Text Available Abstract. Besides the multiple hydrolytic enzymes, lysosomes are equipped with proteins apt to activate sphyngo-lipids — saposins (SAP. SAP belong to a broad and diverse family of moderate-size (~80 AA saposin-like proteins (SAPLIP containing specific domains with three disulfid e bonds bridging six cysteine residues. The diversity of SAPLIPS is likely explained by their involvement in distinct phases of engulfed bacteria digesting. Functionally similar SAPLIP were identified in a wide range of species — from amoeba to mammals, including humans. Saposins per se form a subfamily with six members: saposins A-D and the protein GM2 which possesses activatory functions. SAP do not have enzymatic activity, are heat-stable and protease resistant. The major in vivo function of SAP is released via participation in sphyngolipid catabolism and membrane digestion. In addition, complex association of SAP with membrane bi-layer and CD1 glycolipids is essential for loading lipid antigens onto antigen-presenting CD1 molecules for subsequent activation of lipid-specific T-cells. Of particular interest is participation of SAP in cross-presentation of bacterial antigens to CD8+ T-cells. A broad spectrum of SAP and SAPLIP involvement in the reactions of innate and adaptive immunity indicates their evolutionary conserved role in host defense.

  11. Immune-responsive gene 1 protein links metabolism to immunity by catalyzing itaconic acid production

    Science.gov (United States)

    Michelucci, Alessandro; Cordes, Thekla; Ghelfi, Jenny; Pailot, Arnaud; Reiling, Norbert; Goldmann, Oliver; Binz, Tina; Wegner, André; Tallam, Aravind; Rausell, Antonio; Buttini, Manuel; Linster, Carole L.; Medina, Eva; Balling, Rudi; Hiller, Karsten

    2013-01-01

    Immunoresponsive gene 1 (Irg1) is highly expressed in mammalian macrophages during inflammation, but its biological function has not yet been elucidated. Here, we identify Irg1 as the gene coding for an enzyme producing itaconic acid (also known as methylenesuccinic acid) through the decarboxylation of cis-aconitate, a tricarboxylic acid cycle intermediate. Using a gain-and-loss-of-function approach in both mouse and human immune cells, we found Irg1 expression levels correlating with the amounts of itaconic acid, a metabolite previously proposed to have an antimicrobial effect. We purified IRG1 protein and identified its cis-aconitate decarboxylating activity in an enzymatic assay. Itaconic acid is an organic compound that inhibits isocitrate lyase, the key enzyme of the glyoxylate shunt, a pathway essential for bacterial growth under specific conditions. Here we show that itaconic acid inhibits the growth of bacteria expressing isocitrate lyase, such as Salmonella enterica and Mycobacterium tuberculosis. Furthermore, Irg1 gene silencing in macrophages resulted in significantly decreased intracellular itaconic acid levels as well as significantly reduced antimicrobial activity during bacterial infections. Taken together, our results demonstrate that IRG1 links cellular metabolism with immune defense by catalyzing itaconic acid production. PMID:23610393

  12. Immunization with Brucella VirB proteins reduces organ colonization in mice through a Th1-type immune response and elicits a similar immune response in dogs.

    Science.gov (United States)

    Pollak, Cora N; Wanke, María Magdalena; Estein, Silvia M; Delpino, M Victoria; Monachesi, Norma E; Comercio, Elida A; Fossati, Carlos A; Baldi, Pablo C

    2015-03-01

    VirB proteins from Brucella spp. constitute the type IV secretion system, a key virulence factor mediating the intracellular survival of these bacteria. Here, we assessed whether a Th1-type immune response against VirB proteins may protect mice from Brucella infection and whether this response can be induced in the dog, a natural host for Brucella. Splenocytes from mice immunized with VirB7 or VirB9 responded to their respective antigens with significant and specific production of gamma interferon (IFN-γ), whereas interleukin-4 (IL-4) was not detected. Thirty days after an intraperitoneal challenge with live Brucella abortus, the spleen load of bacteria was almost 1 log lower in mice immunized with VirB proteins than in unvaccinated animals. As colonization reduction seemed to correlate with a Th1-type immune response against VirB proteins, we decided to assess whether such a response could be elicited in the dog. Peripheral blood mononuclear cells (PBMCs) from dogs immunized with VirB proteins (three subcutaneous doses in QuilA adjuvant) produced significantly higher levels of IFN-γ than cells from control animals upon in vitro stimulation with VirB proteins. A skin test to assess specific delayed-type hypersensitivity was positive in 4 out of 5 dogs immunized with either VirB7 or VirB9. As both proteins are predicted to locate in the outer membrane of Brucella organisms, the ability of anti-VirB antibodies to mediate complement-dependent bacteriolysis of B. canis was assessed in vitro. Sera from dogs immunized with either VirB7 or VirB9, but not from those receiving phosphate-buffered saline (PBS), produced significant bacteriolysis. These results suggest that VirB-specific responses that reduce organ colonization by Brucella in mice can be also elicited in dogs. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  13. Protective immunity against Naegleria fowleri infection on mice immunized with the rNfa1 protein using mucosal adjuvants.

    Science.gov (United States)

    Lee, Jinyoung; Yoo, Jong-Kyun; Sohn, Hae-Jin; Kang, Hee-kyoung; Kim, Daesik; Shin, Ho-Joon; Kim, Jong-Hyun

    2015-04-01

    The free-living amoeba, Naegleria fowleri, causes a fatal disease called primary amoebic meningoencephalitis (PAM) in humans and experimental animals. Of the pathogenic mechanism of N. fowleri concerning host tissue invasion, the adherence of amoeba to hose cells is the most important. We previously cloned the nfa1 gene from N. fowleri. The protein displayed immunolocalization in the pseudopodia, especially the food-cups structure, and was related to the contact-dependent mechanism of the amoebic pathogenicity in N. fowleri infection. The cholera toxin B subunit (CTB) and Escherichia coli heat-labile enterotoxin B subunit (LTB) have been used as potent mucosal adjuvants via the parenteral route of immunization in most cases. In this study, to examine the effect of protective immunity of the Nfa1 protein for N. fowleri infection with enhancement by CTB or LTB adjuvants, intranasally immunized BALB/c mice were infected with N. fowleri trophozoites for the development of PAM. The mean time to death of mice immunized with the Nfa1 protein using LTB or CTB adjuvant was prolonged by 5 or 8 days in comparison with that of the control mice. In particular, the survival rate of mice immunized with Nfa1 plus CTB was 100% during the experimental period. The serum IgG levels were significantly increased in mice immunized with Nfa1 protein plus CTB or LTB adjuvants. These results suggest that the Nfa1 protein, with CTB or LTB adjuvants, induces strong protective immunity in mice with PAM due to N. fowleri infection.

  14. Different protein of Echinococcus granulosus stimulates dendritic induced immune response.

    Science.gov (United States)

    Wang, Yana; Wang, Qiang; Lv, Shiyu; Zhang, Shengxiang

    2015-06-01

    Cystic echinococcosis is a chronic infectious disease that results from a host/parasite interaction. Vaccination with ferritin derived from Echinococcus granulosus is a potential preventative treatment. To understand whether ferritin is capable of inducing a host immune response, we investigated the response of dendritic cells (DCs) to both recombinant ferritin protein and the hydatid fluid (HF) of E. granulosus. We evaluated the immunomodulatory potential of these antigens by performing, immunocytochemistry, electron microscopy and in vivo imaging of monocyte-derived murine DCs. During antigen stimulation of DCs, ferritin cause DCs maturation and induced higher levels of surface marker expression and activated T-cell proliferation and migration. On contrary, HF failed to induce surface marker expression and to stimulate T-cell proliferation. In response to HF, DCs produced interleukin-6 (IL-6), but no IL-12 and IL-10. DCs stimulated with ferritin produced high levels of cytokines. Overall, HF appears to induce host immunosuppression in order to ensure parasite survival via inhibits DC maturation and promotes Th2-dependent secretion of cytokines. Although ferritin also promoted DC maturation and cytokine release, it also activates CD4+T-cell proliferation, but regard of the mechanism of the Eg.ferritin induce host to eradicate E. granulosus were not clear.

  15. Immune Response of Multiparous Hyper-Immunized Sows against Peptides from Non-Structural and Structural Proteins of PRRSV

    Directory of Open Access Journals (Sweden)

    Edgar Rascón-Castelo

    2015-11-01

    Full Text Available The purpose of this study was to evaluate the humoral and cellular responses of commercial multiparous and hyper-immunized sows against peptides from non-structural (nsp and structural proteins of porcine reproductive and respiratory syndrome virus (PRRSV. We selected sows with different numbers of parities from a commercial farm. Management practices on this farm include the use of the MLV commercial vaccine four times per year, plus two vaccinations during the acclimation period. The humoral response was evaluated via the antibody recognition of peptides from nsp and structural proteins, and the cellular response was assessed by measuring the frequency of peptide and PRRSV-specific IFN-gamma-secreting cells (IFNγ-SC. Our results show that sows with six parities have more antibodies against peptides from structural proteins than against peptides from nsp. The analysis of the cellular response revealed that the number of immunizations did not affect the frequency of IFNγ-SC and that the response was stronger against peptides from structural proteins (M protein than against nsp (nsp2. In summary, these results demonstrate that multiparous, hyper-immunized sows have a stronger immune humoral response to PRRSV structural peptides than nsp, but no differences in IFNγ-SC against the same peptides were observed.

  16. Immune responses against SARS-coronavirus nucleocapsid protein induced by DNA vaccine

    International Nuclear Information System (INIS)

    Zhao Ping; Cao Jie; Zhao Lanjuan; Qin Zhaolin; Ke Jinshan; Pan Wei; Ren Hao; Yu Jianguo; Qi Zhongtian

    2005-01-01

    The nucleocapsid (N) protein of SARS-coronavirus (SARS-CoV) is the key protein for the formation of the helical nucleocapsid during virion assembly. This protein is believed to be more conserved than other proteins of the virus, such as spike and membrane glycoprotein. In this study, the N protein of SARS-CoV was expressed in Escherichia coli DH5α and identified with pooled sera from patients in the convalescence phase of SARS. A plasmid pCI-N, encoding the full-length N gene of SARS-CoV, was constructed. Expression of the N protein was observed in COS1 cells following transfection with pCI-N. The immune responses induced by intramuscular immunization with pCI-N were evaluated in a murine model. Serum anti-N immunoglobulins and splenocytes proliferative responses against N protein were observed in immunized BALB/c mice. The major immunoglobulin G subclass recognizing N protein was immunoglobulin G2a, and stimulated splenocytes secreted high levels of gamma interferon and IL-2 in response to N protein. More importantly, the immunized mice produced strong delayed-type hypersensitivity (DTH) and CD8 + CTL responses to N protein. The study shows that N protein of SARS-CoV not only is an important B cell immunogen, but also can elicit broad-based cellular immune responses. The results indicate that the N protein may be of potential value in vaccine development for specific prophylaxis and treatment against SARS

  17. Sublingual immunization with a live attenuated influenza a virus lacking the nonstructural protein 1 induces broad protective immunity in mice.

    Directory of Open Access Journals (Sweden)

    Hae-Jung Park

    Full Text Available The nonstructural protein 1 (NS1 of influenza A virus (IAV enables the virus to disarm the host cell type 1 IFN defense system. Mutation or deletion of the NS1 gene leads to attenuation of the virus and enhances host antiviral response making such live-attenuated influenza viruses attractive vaccine candidates. Sublingual (SL immunization with live influenza virus has been found to be safe and effective for inducing protective immune responses in mucosal and systemic compartments. Here we demonstrate that SL immunization with NS1 deleted IAV (DeltaNS1 H1N1 or DeltaNS1 H5N1 induced protection against challenge with homologous as well as heterosubtypic influenza viruses. Protection was comparable with that induced by intranasal (IN immunization and was associated with high levels of virus-specific antibodies (Abs. SL immunization with DeltaNS1 virus induced broad Ab responses in mucosal and systemic compartments and stimulated immune cells in mucosa-associated and systemic lymphoid organs. Thus, SL immunization with DeltaNS1 offers a novel potential vaccination strategy for the control of influenza outbreaks including pandemics.

  18. Sublingual immunization with a live attenuated influenza a virus lacking the nonstructural protein 1 induces broad protective immunity in mice.

    Science.gov (United States)

    Park, Hae-Jung; Ferko, Boris; Byun, Young-Ho; Song, Joo-Hye; Han, Gye-Yeong; Roethl, Elisabeth; Egorov, Andrej; Muster, Thomas; Seong, Baiklin; Kweon, Mi-Na; Song, Manki; Czerkinsky, Cecil; Nguyen, Huan H

    2012-01-01

    The nonstructural protein 1 (NS1) of influenza A virus (IAV) enables the virus to disarm the host cell type 1 IFN defense system. Mutation or deletion of the NS1 gene leads to attenuation of the virus and enhances host antiviral response making such live-attenuated influenza viruses attractive vaccine candidates. Sublingual (SL) immunization with live influenza virus has been found to be safe and effective for inducing protective immune responses in mucosal and systemic compartments. Here we demonstrate that SL immunization with NS1 deleted IAV (DeltaNS1 H1N1 or DeltaNS1 H5N1) induced protection against challenge with homologous as well as heterosubtypic influenza viruses. Protection was comparable with that induced by intranasal (IN) immunization and was associated with high levels of virus-specific antibodies (Abs). SL immunization with DeltaNS1 virus induced broad Ab responses in mucosal and systemic compartments and stimulated immune cells in mucosa-associated and systemic lymphoid organs. Thus, SL immunization with DeltaNS1 offers a novel potential vaccination strategy for the control of influenza outbreaks including pandemics.

  19. Impact of heat shock protein 60KD in combination with outer membrane proteins on immune response against Brucella melitensis.

    Science.gov (United States)

    Abbassi-Daloii, Tooba; Yousefi, Soheil; Sekhavati, Mohammad Hadi; Tahmoorespur, Mojtaba

    2018-01-01

    Brucellosis caused by the bacterium Brucella affects various domestic and wild species. The outer membrane proteins 25 and 31 play key roles on stimulation of cell-mediated immune response against Brucella. GroEL as one of the major Brucella antigens stimulates the immune system and increases intracellular survival of bacteria. In the present study, we assumed injection of GroEL in combination with OMP25 and OMP31 would offer higher immunity levels. So, the impact of GroEL with different concentrations of recombinant outer membrane proteins emulsified in Chitosan Nanoparticles on immune responses was evaluated in mice model. Results showed both univalent (except rGroEL) and divalent immunized groups induced higher IFN-γ, TNF-α, and IL-4 titers in comparison to negative control groups. While GroEL showed negative effect on TNF-α titer, there were positive increase trends in IFN-γ in some treatments. Analysis of humoral antibody response revealed both univalent and divalent immunized groups induced higher IgG2a titer than IgG1 titer, indicating strong bent of Th1 immune response. Also, results showed GroEL can have positive impact on lymphocyte proliferation response. Overall, mice immunization using individual OMP25 or OMP31 demonstrated more effective cell-mediated immunity, although some combinations of rGroEL and rOMP31 vaccines were more efficient than other divalent ones. © 2017 APMIS. Published by John Wiley & Sons Ltd.

  20. Protein-Free Hapten-Carbon Nanotube Constructs Induce the Secondary Immune Response.

    Science.gov (United States)

    Ceballos-Alcantarilla, Eric; Abad-Somovilla, Antonio; Agulló, Consuelo; Abad-Fuentes, Antonio; Mercader, Josep V

    2017-06-21

    Carbon nanotubes are novel technological tools with multiple applications. The interaction between such nanoparticles and living organisms is nowadays a matter of keen research by academic and private institutions. In this study, carbon nanotube constructs were investigated as delivery vehicles for immunostimulation and induction of the secondary immune response to a small organic molecule, namely, a hapten. Two types of nanoconstructs were prepared: on one hand, carbon nanotubes carrying a protein bioconjugate of a hapten covalently linked to the carbon surface, and on the other hand, covalent carbon nanotube constructs of the same model chemical compound without the carrier protein. Nanotube vehicles carrying a hapten-protein bioconjugate were demonstrated to stimulate the immune system and to induce a strong primary immune response against the hapten with as low as 0.1 μg of the model chemical. The influence of the different elements of those nanoconstructs over the immune response was investigated to better understand the molecular mechanisms that are involved. As expected, the presence of the carrier protein was shown to be necessary in order to trigger the immune response. Interestingly, we found that a remarkable secondary immune response to the model organic compound occurred in the absence of a carrier protein. Additionally, a satisfactory adjuvant effect of carbon nanotubes was observed and a potent immune response was elicited without employing an oil-based adjuvant.

  1. Production of bacteriocin-like inhibitory substances (BLIS by Streptococcus salivarius strains isolated from the tongue and throat of children with and without sore throat Produção de substâncias inibidoras semelhantes à bacteriocina por cepas de Streptococcus salivarius, isoladas da língua e garganta de crianças com e sem dor de garganta

    Directory of Open Access Journals (Sweden)

    Vera Fantinato

    1999-12-01

    Full Text Available Streptococcus salivarius strains, isolated from children with and without sore throat, were tested for bacteriocin production against Streptococcus pyogenes. S. salivarius strains producing bacteriocin-like inhibitory substances (BLIS against S. pyogenes were more frequently found in children without sore throat. These results suggest that these children may be protected against sore throat by the presence of BLIS-positive S. salivarius strains.Cepas de Streptococcus salivarius, isoladas de crianças com e sem dor de garganta, foram testadas quanto à produção de bacteriocina contra Streptococcus pyogenes. Os resultados mostraram que as crianças que não tinham dor de garganta possuiam, na boca, cepas de bactérias produtoras de substâncias inibidoras semelhantes à bacteriocina contra S. pyogenes.

  2. Immunization

    Science.gov (United States)

    ... a lot worse. Some are even life-threatening. Immunization shots, or vaccinations, are essential. They protect against things like measles, ... B, polio, tetanus, diphtheria, and pertussis (whooping cough). Immunizations are important for adults as well as children. ...

  3. The Nanoscience of Polyvalent Binding by Proteins in the Immune Response

    DEFF Research Database (Denmark)

    Vorup-Jensen, Thomas

    2016-01-01

    clear or in other way incapacitate the function of these drugs. To rationalize, why and how, the innate immune system especially interacts with nanomedicines, this chapter points to the prominent role of polyvalent interactions by large, immunoactive proteins with the surfaces of nanoparticles. From......Recent research has demonstrated that the successful use of nanometer-scaled material, such as nanoparticles, as medicines is often challenged by the host immune system. Mechanisms of the innate immunity seem to provide a swift response to administration of particulate nanomedicines, which may...... important ways of regulating the interactions, wanted or unwanted, with the innate immune system....

  4. Helminth Protein Vaccine Induced Follicular T Helper Cell for Enhancement of Humoral Immunity against Schistosoma japonicum

    Directory of Open Access Journals (Sweden)

    Jingyao Zhang

    2013-01-01

    Full Text Available Protein vaccines combined with adjuvants have been widely used to induce immune responses, especially the humoral immune response, against molecular targets including parasites. Follicular T helper (Tfh cells are the specialized providers of B-cell help, however, the induction of Tfh cells in protein vaccination has been rarely studied. Here, we report that the Schistosoma japonicum recombinant protein (SjGST-32 combined with tacrolimus (FK506 augmented the induction of Tfh cells, which expressed the canonical markers CXCR5, BCL6, and IL-21, and enhanced the humoral immune responses in BALB/c mice. Furthermore, the expression of IL-21R on germinal center (GC B cells and memory B cells increased in immunized mice, which indicated that IL-21 from the induced Tfh cells interacted with IL-21R for activation of B cells and maintenance of long-lived humoral immunity. Our results suggest that helminth protein vaccine combined with FK506 induces Tfh cell for stimulating humoral immune responses and inducing long-lived humoral immunity.

  5. Protein A Suppresses Immune Responses during Staphylococcus aureus Bloodstream Infection in Guinea Pigs

    Science.gov (United States)

    Kim, Hwan Keun; Falugi, Fabiana; Thomer, Lena; Missiakas, Dominique M.

    2015-01-01

    ABSTRACT   Staphylococcus aureus infection is not associated with the development of protective immunity, and disease relapses occur frequently. We hypothesize that protein A, a factor that binds immunoglobulin Fcγ and cross-links VH3 clan B cell receptors (IgM), is the staphylococcal determinant for host immune suppression. To test this, vertebrate IgM was examined for protein A cross-linking. High VH3 binding activity occurred with human and guinea immunoglobulin, whereas mouse and rabbit immunoglobulins displayed little and no binding, respectively. Establishing a guinea pig model of S. aureus bloodstream infection, we show that protein A functions as a virulence determinant and suppresses host B cell responses. Immunization with SpAKKAA, which cannot bind immunoglobulin, elicits neutralizing antibodies that enable guinea pigs to develop protective immunity. Importance  Staphylococcus aureus is the leading cause of soft tissue and bloodstream infections; however, a vaccine with clinical efficacy is not available. Using mice to model staphylococcal infection, earlier work identified protective antigens; however, corresponding human clinical trials did not reach their endpoints. We show that B cell receptor (IgM) cross-linking by protein A is an important immune evasion strategy of S. aureus that can be monitored in a guinea pig model of bloodstream infection. Further, immunization with nontoxigenic protein A enables infected guinea pigs to elicit antibody responses that are protective against S. aureus. Thus, the guinea pig model may support preclinical development of staphylococcal vaccines. PMID:25564466

  6. Immunization of Zika virus envelope protein domain III induces specific and neutralizing immune responses against Zika virus.

    Science.gov (United States)

    Yang, Ming; Dent, Matthew; Lai, Huafang; Sun, Haiyan; Chen, Qiang

    2017-07-24

    In this study, we described the generation and immunogenicity of the Zika Virus (ZIKV) envelope protein (E) domain III (DIII) as a protein subunit vaccine candidate. ZIKV EDIII (zEDIII) was rapidly produced in E. coli in inclusion bodies. ZIKV EDIII was solubilized, refolded and purified to >95% homogeneity with a one-step Ni 2+ affinity chromatography process. Further analysis revealed that zEDIII was refolded properly and demonstrated specific binding to an anti-zEDIII monoclonal antibody that recognizes a zEDIII conformational epitope. Subcutaneous immunization of mice with 25 and 50μg of zEDIII was performed over a period of 11weeks. zEDIII evoked ZIKV-specific and neutralizing antibody response with titers that exceed the threshold that correlates with protective immunity against ZIKV. The antigen-specific IgG isotypes were predominantly IgG1 and splenocyte cultures from immunized mice secreted IFN-gamma, IL-4 and IL-6. Notably, zEDIII-elicited antibodies did not enhance the infection of dengue virus in Fc gamma receptor (FcγR)-expressing cells. This study provided a proof of principle for the further development of recombinant protein-based subunit vaccines against ZIKV. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Low cost delivery of proteins bioencapsulated in plant cells to human non-immune or immune modulatory cells.

    Science.gov (United States)

    Xiao, Yuhong; Kwon, Kwang-Chul; Hoffman, Brad E; Kamesh, Aditya; Jones, Noah T; Herzog, Roland W; Daniell, Henry

    2016-02-01

    Targeted oral delivery of GFP fused with a GM1 receptor binding protein (CTB) or human cell penetrating peptide (PTD) or dendritic cell peptide (DCpep) was investigated. Presence of GFP(+) intact plant cells between villi of ileum confirm their protection in the digestive system from acids/enzymes. Efficient delivery of GFP to gut-epithelial cells by PTD or CTB and to M cells by all these fusion tags confirm uptake of GFP in the small intestine. PTD fusion delivered GFP more efficiently to most tissues or organs than the other two tags. GFP was efficiently delivered to the liver by all fusion tags, likely through the gut-liver axis. In confocal imaging studies of human cell lines using purified GFP fused with different tags, GFP signal of DCpep-GFP was only detected within dendritic cells. PTD-GFP was only detected within kidney or pancreatic cells but not in immune modulatory cells (macrophages, dendritic, T, B, or mast cells). In contrast, CTB-GFP was detected in all tested cell types, confirming ubiquitous presence of GM1 receptors. Such low-cost oral delivery of protein drugs to sera, immune system or non-immune cells should dramatically lower their cost by elimination of prohibitively expensive fermentation, protein purification cold storage/transportation and increase patient compliance. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  8. Ligand-triggered de-repression of Arabidopsis heterotrimeric G proteins coupled to immune receptor kinases.

    Science.gov (United States)

    Liang, Xiangxiu; Ma, Miaomiao; Zhou, Zhaoyang; Wang, Jinlong; Yang, Xinru; Rao, Shaofei; Bi, Guozhi; Li, Lin; Zhang, Xiaojuan; Chai, Jijie; Chen, She; Zhou, Jian-Min

    2018-03-15

    Arabidopsis heterotrimeric G proteins regulate diverse processes by coupling to single-transmembrane receptors. One such receptor is the FLS2 receptor kinase, which perceives bacterial flagellin epitope flg22 to activate immunity through a class of cytoplasmic kinases called BIK1/PBLs. Unlike animal and fungal heterotrimeric G proteins that are activated by a ligand-induced guanine nucleotide exchange activity of seven-transmembrane G protein-coupled receptors (GPCRs), plant heterotrimeric G proteins are self-activating. How plant receptors regulate heterotrimeric G proteins in response to external ligands remains unknown. Here we show that RGS1, a GTPase accelerating protein, maintains Arabidopsis G proteins in an inactive state in complex with FLS2. Activation of FLS2 by flg22 induces a BIK1/PBL-mediated phosphorylation of RGS1 at Ser428 and Ser431 and that promotes RGS1 dissociation from the FLS2-G protein complex. This relieves G proteins from the RGS1-mediated repression and enables positive regulation of immune signaling. We additionally show that RGS1 is similarly regulated by multiple immune receptors. Our results uncover ligand-induced de-repression as a mechanism for G protein signaling in plants that is distinct from previously reported mechanism underlying the activation of heterotrimeric G proteins in other systems.

  9. G protein coupled receptors signaling pathways implicate in inflammatory and immune response of rheumatoid arthritis.

    Science.gov (United States)

    Shu, Jinling; Zhang, Feng; Zhang, Lingling; Wei, Wei

    2017-05-01

    G protein-coupled receptors (GPCRs) are transmembrane receptor proteins, which allow the transfer of signals across the membrane. Rheumatoid arthritis (RA) is an autoimmune disease characterized by synovitis and accompanied with inflammatory and abnormal immune response. GPCRs signaling pathways play a significant role in inflammatory and immune response processes including RA. In this review, we have focused on the advances in GPCRs signaling pathway implicating the inflammatory and immune response of RA. The signaling pathways of GPCRs-adenylyl cyclase (AC)-cyclic adenosine 3', 5'-monophosphate (cAMP) include β 2 adrenergic receptors (β 2 -ARs)-AC-cAMP signaling pathways, E-prostanoid2/4 (EP2/4)-AC-cAMP signaling pathways and so on. Regulatory proteins, such as G protein-coupled receptor kinases (GRKs) and β-arrestins, play important modulatory roles in GPCRs signaling pathway. GPCRs signaling pathway and regulatory proteins implicate the pathogenesis process of inflammatory and immune response. GPCRs-AC-cAMP signal pathways involve in the inflammatory and immune response of RA. Different signaling pathways are mediated by different receptors, such as β 2 -AR, PGE2 receptor, chemokines receptor, and adenosine receptor. GRKs and β-arrestins are crucial proteins in the regulation of GPCRs signaling pathways. The potential therapeutic targets as well as strategies to modulate GPCRs signaling pathway are new development trends.

  10. Protein energy malnutrition decreases immunity and increases susceptibility to influenza infection in mice.

    Science.gov (United States)

    Taylor, Andrew K; Cao, Weiping; Vora, Keyur P; De La Cruz, Juan; Shieh, Wun-Ju; Zaki, Sherif R; Katz, Jacqueline M; Sambhara, Suryaprakash; Gangappa, Shivaprakash

    2013-02-01

    Protein energy malnutrition (PEM), a common cause of secondary immune deficiency in children, is associated with an increased risk of infections. Very few studies have addressed the relevance of PEM as a risk factor for influenza. We investigated the influence of PEM on susceptibility to, and immune responses following, influenza virus infection using isocaloric diets providing either adequate protein (AP; 18%) or very low protein (VLP; 2%) in a mouse model. We found that mice maintained on the VLP diet, when compared to mice fed with the AP diet, exhibited more severe disease following influenza infection based on virus persistence, trafficking of inflammatory cell types to the lung tissue, and virus-induced mortality. Furthermore, groups of mice maintained on the VLP diet showed significantly lower virus-specific antibody response and a reduction in influenza nuclear protein-specific CD8(+) T cells compared with mice fed on the AP diet. Importantly, switching diets for the group maintained on the VLP diet to the AP diet improved virus clearance, as well as protective immunity to viral challenge. Our results highlight the impact of protein energy on immunity to influenza infection and suggest that balanced protein energy replenishment may be one strategy to boost immunity against influenza viral infections.

  11. Functions of Calcium-Dependent Protein Kinases in Plant Innate Immunity

    Directory of Open Access Journals (Sweden)

    Xiquan Gao

    2014-03-01

    Full Text Available An increase of cytosolic Ca2+ is generated by diverse physiological stimuli and stresses, including pathogen attack. Plants have evolved two branches of the immune system to defend against pathogen infections. The primary innate immune response is triggered by the detection of evolutionarily conserved pathogen-associated molecular pattern (PAMP, which is called PAMP-triggered immunity (PTI. The second branch of plant innate immunity is triggered by the recognition of specific pathogen effector proteins and known as effector-triggered immunity (ETI. Calcium (Ca2+ signaling is essential in both plant PTI and ETI responses. Calcium-dependent protein kinases (CDPKs have emerged as important Ca2+ sensor proteins in transducing differential Ca2+ signatures, triggered by PAMPs or effectors and activating complex downstream responses. CDPKs directly transmit calcium signals by calcium binding to the elongation factor (EF-hand domain at the C-terminus and substrate phosphorylation by the catalytic kinase domain at the N-terminus. Emerging evidence suggests that specific and overlapping CDPKs phosphorylate distinct substrates in PTI and ETI to regulate diverse plant immune responses, including production of reactive oxygen species, transcriptional reprogramming of immune genes, and the hypersensitive response.

  12. Analysis of the humoral immune response to Chlamydia outer membrane protein 2

    DEFF Research Database (Denmark)

    Mygind, P; Christiansen, Gunna; Persson, K

    1998-01-01

    The humoral immune response to Chlamydia outer membrane protein 2 (Omp2) was studied. Omp2 is a highly genus-conserved structural protein of all Chlamydia species, containing a variable N-terminal fragment. To analyze where the immunogenic parts were localized, seven highly purified truncated...... patient sera, Omp2 was found to be a major immunogen of both C. pneumoniae and C. trachomatis infections (P immune responses were not confined to any particular region of the Omp2 protein, and no species-specific anti-Omp2 immunoglobulins were detected....

  13. Effects of experimentally increased protein supply to postpartum dairy cows on plasma protein synthesis, rumen tissue proliferation, and immune homeostasis

    DEFF Research Database (Denmark)

    Larsen, Mogens; Røntved, Christine Maria; Theil, Peter Kappel

    2017-01-01

    The effect of experimentally increasing the postpartum protein supply on plasma protein synthesis, rumen tissue proliferation, and immune homeostasis was studied using 8 periparturient Holstein cows in a complete randomized design. At calving, cows were assigned to abomasal infusion of water (CTRL......]Phe isotopic enrichment, and mRNA expression of selected genes was measured by real-time qPCR. Total and differential leukocyte counts were performed and immune responsiveness of monocytes was evaluated by tumor necrosis factor ɑ (TNFɑ) concentration on ex vivo whole blood stimulation with Escherichia coli...

  14. Immunization against lysozyme-like proteins affect sperm function and fertility in the rat.

    Science.gov (United States)

    Narmadha, Ganapathy; Yenugu, Suresh

    2016-11-01

    Proteins of the epididymal and testicular mileu contribute to sperm maturation and a vast majority of them remain uncharacterised. In this study, the role of three Lysozyme-like (LYZL) proteins, namely LYZL1, LYZL4 and LYZL6 in sperm function was assessed using in vitro neutralization and auto antibodies generation model. Rats immunized with LYZL1, LYZL4 and LYZL6 proteins had a litter size of 5.93, 8.47 and 2.10 respectively compared to 9.96 in the control rats. The litter size was further reduced to 4.53, 7.67 and 1.23 for the corresponding proteins in the second mating conducted 14 weeks after immunization. Epididymal and testicular fluids obtained from the immunized rats displayed a very high antibody titre against all the three proteins. Sperm count was significantly reduced in rats immunized with LYZL1 or LYZL6 and to a lower extent in LYZL4 group. Acrosome reaction associated calcium release was inhibited in spermatozoa obtained from LYZL1 or LYZL4 or LYZL6 immunized rats as well as in spermatozoa incubated with antiserum against the three proteins. Impairment in path velocity, progressive velocity and track speed were observed in spermatozoa obtained from LYZL6 immunized rats. Treatment of spermatozoa with LYZL6 recombinant protein did not potentiate calcium release and acrosome reaction. Results of this study indicate a role for LYZL proteins in sperm function and further studies are warranted to explore them as potential contraceptive agents. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  15. Salivary Defense Proteins: Their Network and Role in Innate and Acquired Oral Immunity

    Directory of Open Access Journals (Sweden)

    Gábor Fábián

    2012-04-01

    Full Text Available There are numerous defense proteins present in the saliva. Although some of these molecules are present in rather low concentrations, their effects are additive and/or synergistic, resulting in an efficient molecular defense network of the oral cavity. Moreover, local concentrations of these proteins near the mucosal surfaces (mucosal transudate, periodontal sulcus (gingival crevicular fluid and oral wounds and ulcers (transudate may be much greater, and in many cases reinforced by immune and/or inflammatory reactions of the oral mucosa. Some defense proteins, like salivary immunoglobulins and salivary chaperokine HSP70/HSPAs (70 kDa heat shock proteins, are involved in both innate and acquired immunity. Cationic peptides and other defense proteins like lysozyme, bactericidal/permeability increasing protein (BPI, BPI-like proteins, PLUNC (palate lung and nasal epithelial clone proteins, salivary amylase, cystatins, prolin-rich proteins, mucins, peroxidases, statherin and others are primarily responsible for innate immunity. In this paper, this complex system and function of the salivary defense proteins will be reviewed.

  16. Comparisons of Allergenic and Metazoan Parasite Proteins: Allergy the Price of Immunity.

    Directory of Open Access Journals (Sweden)

    Nidhi Tyagi

    2015-10-01

    Full Text Available Allergic reactions can be considered as maladaptive IgE immune responses towards environmental antigens. Intriguingly, these mechanisms are observed to be very similar to those implicated in the acquisition of an important degree of immunity against metazoan parasites (helminths and arthropods in mammalian hosts. Based on the hypothesis that IgE-mediated immune responses evolved in mammals to provide extra protection against metazoan parasites rather than to cause allergy, we predict that the environmental allergens will share key properties with the metazoan parasite antigens that are specifically targeted by IgE in infected human populations. We seek to test this prediction by examining if significant similarity exists between molecular features of allergens and helminth proteins that induce an IgE response in the human host. By employing various computational approaches, 2712 unique protein molecules that are known IgE antigens were searched against a dataset of proteins from helminths and parasitic arthropods, resulting in a comprehensive list of 2445 parasite proteins that show significant similarity through sequence and structure with allergenic proteins. Nearly half of these parasite proteins from 31 species fall within the 10 most abundant allergenic protein domain families (EF-hand, Tropomyosin, CAP, Profilin, Lipocalin, Trypsin-like serine protease, Cupin, BetV1, Expansin and Prolamin. We identified epitopic-like regions in 206 parasite proteins and present the first example of a plant protein (BetV1 that is the commonest allergen in pollen in a worm, and confirming it as the target of IgE in schistosomiasis infected humans. The identification of significant similarity, inclusive of the epitopic regions, between allergens and helminth proteins against which IgE is an observed marker of protective immunity explains the 'off-target' effects of the IgE-mediated immune system in allergy. All these findings can impact the discovery and

  17. Immune-responsive gene 1 protein links metabolism to immunity by catalyzing itaconic acid production

    OpenAIRE

    Michelucci, Alessandro; Cordes, Thekla; Ghelfi, Jenny; Pailot, Arnaud; Reiling, Norbert; Goldmann, Oliver; Binz, Tina; Wegner, André; Tallam, Aravind; Rausell, Antonio; Buttini, Manuel; Linster, Carole L.; Medina, Eva; Balling, Rudi; Hiller, Karsten

    2013-01-01

    Immunoresponsive gene 1 (Irg1) is highly expressed in mammalian macrophages during inflammation, but its biological function has not yet been elucidated. Here, we identify Irg1 as the gene coding for an enzyme producing itaconic acid (also known as methylenesuccinic acid) through the decarboxylation of cis-aconitate, a tricarboxylic acid cycle intermediate. Using a gain-and-loss-of-function approach in both mouse and human immune cells, we found Irg1 expression levels correlating with the amo...

  18. The calcium-dependent protein kinase CPK28 buffers plant immunity and regulates BIK1 turnover

    DEFF Research Database (Denmark)

    Monaghan, Jacqueline; Matschi, Susanne; Shorinola, Oluwaseyi

    2014-01-01

    Plant perception of pathogen-associated molecular patterns (PAMPs) triggers a phosphorylation relay leading to PAMP-triggered immunity (PTI). Despite increasing knowledge of PTI signaling, how immune homeostasis is maintained remains largely unknown. Here we describe a forward-genetic screen...... to identify loci involved in PTI and characterize the Arabidopsis calcium-dependent protein kinase CPK28 as a negative regulator of immune signaling. Genetic analyses demonstrate that CPK28 attenuates PAMP-triggered immune responses and antibacterial immunity. CPK28 interacts with and phosphorylates...... the plasma-membrane-associated cytoplasmic kinase BIK1, an important convergent substrate of multiple pattern recognition receptor (PRR) complexes. We find that BIK1 is rate limiting in PTI signaling and that it is continuously turned over to maintain cellular homeostasis. We further show that CPK28...

  19. Role of macrophage inflammatory protein-1alpha in T-cell-mediated immunity to viral infection

    DEFF Research Database (Denmark)

    Madsen, Andreas N; Nansen, Anneline; Christensen, Jan P

    2003-01-01

    The immune response to lymphocytic choriomeningitis virus in mice lacking macrophage inflammatory protein-1alpha (MIP-1alpha) was evaluated. Generation of virus-specific effector T cells is unimpaired in MIP-1alpha-deficient mice. Furthermore, MIP-1alpha is not required for T-cell-mediated virus...... control or virus-induced T-cell-dependent inflammation. Thus, MIP-1alpha is not mandatory for T-cell-mediated antiviral immunity....

  20. Immune response to synthetic peptides of dengue prM protein.

    Science.gov (United States)

    Vázquez, Susana; Guzmán, María Guadalupe; Guillen, Gerardo; Chinea, Glay; Pérez, Ana Beatriz; Pupo, Maritza; Rodriguez, Rosmary; Reyes, Osvaldo; Garay, Hilda Elisa; Delgado, Iselys; García, Gissel; Alvarez, Mayling

    2002-03-15

    The immunological activities of five synthetic peptides of the prM protein of dengue-2 (DEN-2) virus containing B cell epitopes were evaluated in BALB/c mice. Two peptides elicited neutralizing antibodies against all four DEN serotypes. Virus-specific proliferative responses were demonstrated in mice immunized with four of the five peptides, demonstrating the presence of T cell epitopes. Mice immunized with three of the five peptides conjugated with bovine albumin showed statistically significant levels (Pvaccines.

  1. Inhibitor of apoptosis (IAP) proteins in regulation of inflammation and innate immunity

    DEFF Research Database (Denmark)

    Damgaard, Rune B; Gyrd-Hansen, Mads

    2011-01-01

    Inflammatory and innate immune signaling in response to recognition of pathogens is essential for immunity and host survival. However, deregulation may lead to detrimental pathologies including immunodeficiency, inflammatory diseases, and cancer. Inhibitor of apoptosis (IAP) proteins have emerged...... as important regulators of innate immune signaling downstream of pattern recognition receptors (PRRs) such as Toll-like receptor 4 (TLR4), the nucleotide-binding oligomerization domain 1 (NOD1) and NOD2 receptors, and the retinoic acid-inducible gene (RIG)-I receptor. Recent evidence suggests that cIAP1, cIAP2...

  2. A Probiotic Adjuvant Lactobacillus rhamnosus Enhances Specific Immune Responses after Ocular Mucosal Immunization with Chlamydial Polymorphic Membrane Protein C.

    Directory of Open Access Journals (Sweden)

    Aleksandra Inic-Kanada

    Full Text Available Recent advances in the development of chlamydia vaccines, using live-attenuated or ultraviolet light-inactivated chlamydia, are paving the way for new possibilities to oppose the societal challenges posed by chlamydia-related diseases, such as blinding trachoma. An effective subunit vaccine would mitigate the risks associated with the use of a whole-cell vaccine. Our rationale for the design of an efficient subunit vaccine against Chlamydia trachomatis (Ct is based on the membrane proteins involved in the initial Ct-host cell contact and on the route of immunization that mimics the natural infection process (i.e., via the ocular mucosa. The first aim of our study was to characterize the specific conjunctival and vaginal immune responses following eye drop immunization in BALB/c mice, using the N-terminal portion of the Ct serovar E polymorphic membrane protein C (N-PmpC as the subunit vaccine antigen. Second, we aimed to examine the adjuvant properties of the probiotic Lactobacillus rhamnosus (LB when formulated with N-PmpC. N-PmpC applied alone stimulated the production of N-PmpC- and Ct serovar B-specific antibodies in serum, tears and vaginal washes, whereas the combination with LB significantly enhanced these responses. The N-PmpC/LB combination initiated a T cell response characterized by an elevated percentage of CD25+ T cells and CD8+ effector T cells, enhanced CD4+ T-helper 1 skewing, and increased regulatory T cell responses. Together, these results show that eye drop vaccination with combined use of N-PmpC and a live probiotic LB stimulates specific cellular and humoral immune responses, not only locally in the conjunctiva but also in the vaginal mucosa, which could be a promising approach in Ct vaccine development.

  3. ImmTree: Database of evolutionary relationships of genes and proteins in the human immune system

    Science.gov (United States)

    Ortutay, Csaba; Siermala, Markku; Vihinen, Mauno

    2007-01-01

    Background The immune system, which is a complex machinery, is based on the highly coordinated expression of a wide array of genes and proteins. The evolutionary history of the human immune system is not well characterised. Although several studies related to the development and evolution of immunological processes have been published, a full-scale genome-based analysis is still missing. A database focused on the evolutionary relationships of immune related genes would contribute to and facilitate research on immunology and evolutionary biology. Results An Internet resource called ImmTree was constructed for studying the evolution and evolutionary trees of the human immune system. ImmTree contains information about orthologs in 80 species collected from the HomoloGene, OrthoMCL and EGO databases. In addition to phylogenetic trees, the service provides data for the comparison of human-mouse ortholog pairs, including synonymous and non-synonymous mutation rates, Z values, and Ka/Ks quotients. A versatile search engine allows complex queries from the database. Currently, data is available for 847 human immune system related genes and proteins. Conclusion ImmTree provides a unique data set of genes and proteins from the human immune system, their phylogenetics, and information for comparisons of human-mouse ortholog pairs, synonymous and non-synonymous mutation rates, as well as other statistical information. PMID:17376226

  4. Studies on Shigella boydii infection in Caenorhabditis elegans and bioinformatics analysis of immune regulatory protein interactions.

    Science.gov (United States)

    Kesika, Periyanaina; Balamurugan, Krishnaswamy

    2012-12-01

    Shigella boydii causes bacillary dysentery or shigellosis and generates a significant burden in the developing nations. S. boydii-mediated infection assays were performed at both physiological and molecular levels using Caenorhabditis elegans as a host. Continuous exposure of worms to S. boydii showed a reduced life span indicating the pathogenicity of Shigella. Quantitative Real-Time PCR analysis was performed to analyze the expression and regulation of host specific candidate-antimicrobial genes (clec-60, clec-87, lys-7), which were expressed significantly during early infection, but weakened during the latter hours. Increased mortality of mutant RB1285 by S. boydii and Shigella flexneri indicated the role of lys-7 during Shigella infection. Protein-protein interactions (PPIs) database was used to analyze the interaction of immune proteins in both C. elegans and humans. In addition, the expression and regulation were revealed about immune genes (clec-61, clec-62, clec-63, F54D5.3 and ZK1320.2), which encode several intermediate immune protein partners (CLEC-61, CLEC-62, CLEC-63, F54D5.3, ZK1320.2, W03D2.6 and THN-2) that interact with LYS-7 and CLEC-60 and were found to play a role in C. elegans immune defense against S. boydii infections. Similarly, the immune genes that are specific to the human defense system, which encode IGHV4-39, A2M, LTF, and CD79A, were predicted to be expressed with LYZ and MBL2, thus indicating their regulation during Shigella infections. Our results using the lowest eukaryotic model system and human database indicated that the major players involved in immunity-related processes appear to be common in cases of Shigella sp. mediated immune responses. This article is part of a Special Issue entitled: Computational Methods for Protein Interaction and Structural Prediction. Copyright © 2012 Elsevier B.V. All rights reserved.

  5. Insight into bacterial virulence mechanisms against host immune response via the Yersinia pestis-human protein-protein interaction network.

    Science.gov (United States)

    Yang, Huiying; Ke, Yuehua; Wang, Jian; Tan, Yafang; Myeni, Sebenzile K; Li, Dong; Shi, Qinghai; Yan, Yanfeng; Chen, Hui; Guo, Zhaobiao; Yuan, Yanzhi; Yang, Xiaoming; Yang, Ruifu; Du, Zongmin

    2011-11-01

    A Yersinia pestis-human protein interaction network is reported here to improve our understanding of its pathogenesis. Up to 204 interactions between 66 Y. pestis bait proteins and 109 human proteins were identified by yeast two-hybrid assay and then combined with 23 previously published interactions to construct a protein-protein interaction network. Topological analysis of the interaction network revealed that human proteins targeted by Y. pestis were significantly enriched in the proteins that are central in the human protein-protein interaction network. Analysis of this network showed that signaling pathways important for host immune responses were preferentially targeted by Y. pestis, including the pathways involved in focal adhesion, regulation of cytoskeleton, leukocyte transendoepithelial migration, and Toll-like receptor (TLR) and mitogen-activated protein kinase (MAPK) signaling. Cellular pathways targeted by Y. pestis are highly relevant to its pathogenesis. Interactions with host proteins involved in focal adhesion and cytoskeketon regulation pathways could account for resistance of Y. pestis to phagocytosis. Interference with TLR and MAPK signaling pathways by Y. pestis reflects common characteristics of pathogen-host interaction that bacterial pathogens have evolved to evade host innate immune response by interacting with proteins in those signaling pathways. Interestingly, a large portion of human proteins interacting with Y. pestis (16/109) also interacted with viral proteins (Epstein-Barr virus [EBV] and hepatitis C virus [HCV]), suggesting that viral and bacterial pathogens attack common cellular functions to facilitate infections. In addition, we identified vasodilator-stimulated phosphoprotein (VASP) as a novel interaction partner of YpkA and showed that YpkA could inhibit in vitro actin assembly mediated by VASP.

  6. Immunity of replicating Mu to self-integration: a novel mechanism employing MuB protein

    Directory of Open Access Journals (Sweden)

    Ge Jun

    2010-02-01

    Full Text Available Abstract We describe a new immunity mechanism that protects actively replicating/transposing Mu from self-integration. We show that this mechanism is distinct from the established cis-immunity mechanism, which operates by removal of MuB protein from DNA adjacent to Mu ends. MuB normally promotes integration into DNA to which it is bound, hence its removal prevents use of this DNA as target. Contrary to what might be expected from a cis-immunity mechanism, strong binding of MuB was observed throughout the Mu genome. We also show that the cis-immunity mechanism is apparently functional outside Mu ends, but that the level of protection offered by this mechanism is insufficient to explain the protection seen inside Mu. Thus, both strong binding of MuB inside and poor immunity outside Mu testify to a mechanism of immunity distinct from cis-immunity, which we call 'Mu genome immunity'. MuB has the potential to coat the Mu genome and prevent auto-integration as previously observed in vitro on synthetic A/T-only DNA, where strong MuB binding occluded the entire bound region from Mu insertions. The existence of two rival immunity mechanisms within and outside the Mu genome, both employing MuB, suggests that the replicating Mu genome must be segregated into an independent chromosomal domain. We propose a model for how formation of a 'Mu domain' may be aided by specific Mu sequences and nucleoid-associated proteins, promoting polymerization of MuB on the genome to form a barrier against self-integration.

  7. Hijacking Complement Regulatory Proteins for Bacterial Immune Evasion.

    Science.gov (United States)

    Hovingh, Elise S; van den Broek, Bryan; Jongerius, Ilse

    2016-01-01

    The human complement system plays an important role in the defense against invading pathogens, inflammation and homeostasis. Invading microbes, such as bacteria, directly activate the complement system resulting in the formation of chemoattractants and in effective labeling of the bacteria for phagocytosis. In addition, formation of the membrane attack complex is responsible for direct killing of Gram-negative bacteria. In turn, bacteria have evolved several ways to evade complement activation on their surface in order to be able to colonize and invade the human host. One important mechanism of bacterial escape is attraction of complement regulatory proteins to the microbial surface. These molecules are present in the human body for tight regulation of the complement system to prevent damage to host self-surfaces. Therefore, recruitment of complement regulatory proteins to the bacterial surface results in decreased complement activation on the microbial surface which favors bacterial survival. This review will discuss recent advances in understanding the binding of complement regulatory proteins to the bacterial surface at the molecular level. This includes, new insights that have become available concerning specific conserved motives on complement regulatory proteins that are favorable for microbial binding. Finally, complement evasion molecules are of high importance for vaccine development due to their dominant role in bacterial survival, high immunogenicity and homology as well as their presence on the bacterial surface. Here, the use of complement evasion molecules for vaccine development will be discussed.

  8. Arabidopsis heterotrimeric G proteins regulate immunity by directly coupling to the FLS2 receptor.

    Science.gov (United States)

    Liang, Xiangxiu; Ding, Pingtao; Lian, Kehui; Wang, Jinlong; Ma, Miaomiao; Li, Lin; Li, Lei; Li, Meng; Zhang, Xiaojuan; Chen, She; Zhang, Yuelin; Zhou, Jian-Min

    2016-04-04

    The Arabidopsis immune receptor FLS2 perceives bacterial flagellin epitope flg22 to activate defenses through the central cytoplasmic kinase BIK1. The heterotrimeric G proteins composed of the non-canonical Gα protein XLG2, the Gβ protein AGB1, and the Gγ proteins AGG1 and AGG2 are required for FLS2-mediated immune responses through an unknown mechanism. Here we show that in the pre-activation state, XLG2 directly interacts with FLS2 and BIK1, and it functions together with AGB1 and AGG1/2 to attenuate proteasome-mediated degradation of BIK1, allowing optimum immune activation. Following the activation by flg22, XLG2 dissociates from AGB1 and is phosphorylated by BIK1 in the N terminus. The phosphorylated XLG2 enhances the production of reactive oxygen species (ROS) likely by modulating the NADPH oxidase RbohD. The study demonstrates that the G proteins are directly coupled to the FLS2 receptor complex and regulate immune signaling through both pre-activation and post-activation mechanisms.

  9. The Unfolded Protein Response in Homeostasis and Modulation of Mammalian Immune Cells.

    Science.gov (United States)

    Martins, Ana Sofia; Alves, Inês; Helguero, Luisa; Domingues, Maria Rosário; Neves, Bruno Miguel

    2016-11-01

    The endoplasmic reticulum (ER) plays important roles in eukaryotic protein folding and lipid biosynthesis. Several exogenous and endogenous cellular sources of stress can perturb ER homeostasis leading to the accumulation of unfolded proteins in the lumen. Unfolded protein accumulation triggers a signal-transduction cascade known as the unfolded protein response (UPR), an adaptive mechanism which aims to protect cells from protein aggregates and to restore ER functions. Further to this protective mechanism, in immune cells, UPR molecular effectors have been shown to participate in a wide range of biological processes such as cell differentiation, survival and immunoglobulin and cytokine production. Recent findings also highlight the involvement of the UPR machinery in the maturational program and antigen presentation capacities of dendritic cells. UPR is therefore a key element in immune system homeostasis with direct implications on both adaptive and innate immune responses. The present review summarizes the knowledge on the emerging roles of UPR signaling cascades in mammalian immune cells as well as the consequences of their dysregulation in relation to the pathogenesis of several diseases.

  10. Vaccine platforms combining circumsporozoite protein and potent immune modulators, rEA or EAT-2, paradoxically result in opposing immune responses.

    Directory of Open Access Journals (Sweden)

    Nathaniel J Schuldt

    Full Text Available Malaria greatly impacts the health and wellbeing of over half of the world's population. Promising malaria vaccine candidates have attempted to induce adaptive immune responses to Circumsporozoite (CS protein. Despite the inclusion of potent adjuvants, these vaccines have limited protective efficacy. Conventional recombinant adenovirus (rAd based vaccines expressing CS protein can induce CS protein specific immune responses, but these are essentially equivalent to those generated after use of the CS protein subunit based vaccines. In this study we combined the use of rAds expressing CS protein along with rAds expressing novel innate immune response modulating proteins in an attempt to significantly improve the induction of CS protein specific cell mediated immune (CMI responses.BALB/cJ mice were co-vaccinated with a rAd vectors expressing CS protein simultaneous with a rAd expressing either TLR agonist (rEA or SLAM receptors adaptor protein (EAT-2. Paradoxically, expression of the TLR agonist uncovered a potent immunosuppressive activity inherent to the combined expression of the CS protein and rEA. Fortunately, use of the rAd vaccine expressing EAT-2 circumvented CS protein's suppressive activity, and generated a fivefold increase in the number of CS protein responsive, IFNγ secreting splenocytes, as well as increased the breadth of T cells responsive to peptides present in the CS protein. These improvements were positively correlated with the induction of a fourfold improvement in CS protein specific CTL functional activity in vivo.Our results emphasize the need for caution when incorporating CS protein into malaria vaccine platforms expressing or containing other immunostimulatory compounds, as the immunological outcomes may be unanticipated and/or counter-productive. However, expressing the SLAM receptors derived signaling adaptor EAT-2 at the same time of vaccination with CS protein can overcome these concerns, as well as significantly

  11. Intranasal immunization with novel EspA-Tir-M fusion protein induces protective immunity against enterohemorrhagic Escherichia coli O157:H7 challenge in mice.

    Science.gov (United States)

    Lin, Ruqin; Zhu, Bo; Zhang, Yiduo; Bai, Yang; Zhi, Fachao; Long, Beiguo; Li, Yawen; Wu, Yuhua; Wu, Xianbo; Fan, Hongying

    2017-04-01

    Enterohemorrhagic Escherichia coli (EHEC) O157:H7 causes hemorrhagic colitis and hemolytic uremic syndrome in humans. Due to the risks associated with antibiotic treatment against EHEC O157:H7 infection, vaccines represent a promising method for prevention of EHEC O157:H7 infection. Therefore, we constructed the novel bivalent antigen EspA-Tir-M as a candidate EHEC O157:H7 subunit vaccine. We then evaluated the immunogenicity of this novel EHEC O157:H7 subunit vaccine. Immune responses to the fusion protein administered by intranasal and subcutaneous routes were compared in mice. Results showed higher levels of specific mucosal and systemic antibody responses induced by intranasal as compared to subcutaneous immunization. Intranasal immunization enhanced the concentration of interleukin-4, interleukin-10, and interferon-γ, while subcutaneous immunization enhanced only the latter two. In addition, intranasal immunization protected against EHEC O157:H7 colonization and infection in mice at a rate of 90%.Histopathological analysis revealed that vaccination reduced colon damage, especially when administered intranasally. In contrast, subcutaneous immunization elicited a weak immune response and exhibited a low protection rate. These findings demonstrate that intranasal immunization with the fusion protein induces both humoral and cellular immune (Th1/Th2) responses in mice. The novel EspA-Tir-M novel fusion protein therefore represents a promising subunit vaccine against EHEC O157:H7 infection. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Immunity of Surface Layer Protein of Aeromonas ‎hydrophila in Rabbits

    Directory of Open Access Journals (Sweden)

    Lobna Adil Al-Noori

    2017-11-01

    Full Text Available In this study the Surface layer (S-layer protein was extracted from Aeromonas hydrophila bacteria ,the humoral immune response that induced by S-layer protein only or as adjuvant was investigated  by using 16 males New Zealand rabbits and divided into four groups, each group contained four rabbits, the first group was immunized with  S-layer protein only, the second group was immunized with heated killed antigen(HKAof Sallmonella typhi only, the third group was immunized with mixed antigens (S-layer+ HKA,while the fourth group considered as control group and immunized with normal saline. The HKA of S. typhi  was used to evaluate the efficiency of S-layer protein as adjuvant. After the immunization period, the humoral immune response was investigated by several tests include, tube agglutination test and passive agglutination test that used to detect the antibody titer. Biuret method was used to determine the total protein concentration in serum  samples and total protein concentration of secretory immunoglobulin that extracted form appendix samples. In addition the Radical Immunodiffution (RID  method was used to detect the concentration level of the IgG in serum samples. Moreover the concentration level of the CD4 in the serum samples was determined by enzyme linked immunosorbent assay (ELISA method .In all these tests the result revealed, both of S-layer protein only , HKA of only and mixed antigens(S-layer+ HKA were given significantly increased in comparison with control group at P<0.05. The result showed that the  concentration level of IgG with mean values (2365.5 , 3505 and 2916 mg/dl respectively  while the control group with mean value (1662mg/dl. In addition the concentration  level of CD4 molecule with mean values (9.37, 11.77 and 17.36 ng/ml respectively while the control group with mean value (6.91 ng/ml .The results showed that these three types of antigens induced the humoral immune response

  13. The essential role of G protein-coupled receptor (GPCR) signaling in regulating T cell immunity.

    Science.gov (United States)

    Wang, Dashan

    2018-02-12

    The aim of this paper is to clarify the critical role of GPCR signaling in T cell immunity. The G protein-coupled receptors (GPCRs) are the most common targets in current pharmaceutical industry, and represent the largest and most versatile family of cell surface communicating molecules. GPCRs can be activated by a diverse array of ligands including neurotransmitters, chemokines as well as sensory stimuli. Therefore, GPCRs are involved in many key cellular and physiological processes, such as sense of light, taste and smell, neurotransmission, metabolism, endocrine and exocrine secretion. In recent years, GPCRs have been found to play an important role in immune system. T cell is an important type of immune cell, which plays a central role in cell-mediated immunity. A variety of GPCRs and their signaling mediators (RGS proteins, GRKs and β-arrestin) have been found to express in T cells and involved T cell-mediated immunity. We will summarize the role of GPCR signaling and their regulatory molecules in T cell activation, homeostasis and function in this article. GPCR signaling plays an important role in T cell activation, homeostasis and function. GPCR signaling is critical in regulating T cell immunity.

  14. NY-ESO-1 protein glycosylated by yeast induces enhanced immune responses.

    Science.gov (United States)

    Wadle, Andreas; Mischo, Axel; Strahl, Sabine; Nishikawa, Hiroyoshi; Held, Gerhard; Neumann, Frank; Wullner, Beate; Fischer, Eliane; Kleber, Sascha; Karbach, Julia; Jager, Elke; Shiku, Hiroshi; Odunsi, Kunle; Shrikant, Protul A; Knuth, Alexander; Cerundolo, Vincenzo; Renner, Christoph

    2010-11-01

    Vaccine strategies that target dendritic cells to elicit potent cellular immunity are the subject of intense research. Here we report that the genetically engineered yeast Saccharomyces cerevisiae, expressing the full-length tumour-associated antigen NY-ESO-1, is a versatile host for protein production. Exposing dendritic cells (DCs) to soluble NY-ESO-1 protein linked to the yeast a-agglutinin 2 protein (Aga2p) protein resulted in protein uptake, processing and MHC class I cross-presentation of NY-ESO-1-derived peptides. The process of antigen uptake and cross-presentation was dependent on the glycosylation pattern of NY-ESO-1-Aga2p protein and the presence of accessible mannose receptors. In addition, NY-ESO-1-Aga2p protein uptake by dendritic cells resulted in recognition by HLA-DP4 NY-ESO-1-specific CD4(+) T cells, indicating MHC class II presentation. Finally, vaccination of mice with yeast-derived NY-ESO-1-Aga2p protein led to an enhanced humoral and cellular immune response, when compared to the bacterially expressed NY-ESO-1 protein. Together, these data demonstrate that yeast-derived full-length NY-ESO-1-Aga2p protein is processed and presented efficiently by MHC class I and II complexes and warrants clinical trials to determine the potential value of S. cerevisiae as a host for cancer vaccine development. Copyright © 2010 John Wiley & Sons, Ltd.

  15. Antibody study in canine distemper virus nucleocapsid protein gene-immunized mice.

    Science.gov (United States)

    Yuan, B; Li, X Y; Zhu, T; Yuan, L; Hu, J P; Chen, J; Gao, W; Ren, W Z

    2015-04-10

    The gene for the nucleocapsid (N) protein of canine distemper virus was cloned into the pMD-18T vector, and positive recombinant plasmids were obtained by enzyme digestion and sequencing. After digestion by both EcoRI and KpnI, the plasmid was directionally cloned into the eukaryotic expression vector pcDNA; the positive clone pcDNA-N was screened by electrophoresis and then transfected into COS-7 cells. Immunofluorescence analysis results showed that the canine distemper virus N protein was expressed in the cytoplasm of transfected COS-7 cells. After emulsification in Freund's adjuvant, the recombinant plasmid pcDNA-N was injected into the abdominal cavity of 8-week-old BABL/c mice, with the pcDNA original vector used as a negative control. Mice were immunized 3 times every 2 weeks. The blood of immunized mice was drawn 2 weeks after completing the immunizations to measure titer levels. The antibody titer in the pcDNA-N test was 10(1.62 ± 0.164), while in the control group this value was 10(0.52 ± 0.56), indicating that specific humoral immunity was induced in canine distemper virus nucleocapsid protein-immunized mice.

  16. Mechanisms of Host-Pathogen Protein Complex Formation and Bacterial Immune Evasion of Streptococcus suis Protein Fhb.

    Science.gov (United States)

    Li, Xueqin; Liu, Peng; Gan, Shuzhen; Zhang, Chunmao; Zheng, Yuling; Jiang, Yongqiang; Yuan, Yuan

    2016-08-12

    Streptococcus suis serotype 2 (S. suis 2)-induced sepsis and meningitis are often accompanied by bacteremia. The evasion of polymorphonuclear leukocyte-mediated phagocytic clearance is central to the establishment of bacteremia caused by S. suis 2 and is facilitated by the ability of factor H (FH)-binding protein (Fhb) to bind FH on the bacterial surface, thereby impeding alternative pathway complement activation and phagocytic clearance. Here, C3b/C3d was found to bind to Fhb, along with FH, forming a large immune complex. The formation of this immune complex was mediated by domain II of Fhb via electrostatic and hydrophobic interactions, which, to our knowledge, is a new type of interaction. Interestingly, Fhb was found to be associated with the cell envelope and also present in the culture supernatant, where secreted Fhb inhibited complement activation via interactions with domain II, thereby enhancing antiphagocytic clearance by polymorphonuclear leukocytes. Thus, Fhb is a multifunctional bacterial protein, which binds host complement component C3 as well as FH and interferes with innate immune recognition in a secret protein manner. S. suis 2 therefore appears to have developed a new strategy to combat host innate immunity and enhance survival in host blood. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Mechanisms of Host-Pathogen Protein Complex Formation and Bacterial Immune Evasion of Streptococcus suis Protein Fhb*

    Science.gov (United States)

    Li, Xueqin; Liu, Peng; Gan, Shuzhen; Zhang, Chunmao; Zheng, Yuling; Jiang, Yongqiang; Yuan, Yuan

    2016-01-01

    Streptococcus suis serotype 2 (S. suis 2)-induced sepsis and meningitis are often accompanied by bacteremia. The evasion of polymorphonuclear leukocyte-mediated phagocytic clearance is central to the establishment of bacteremia caused by S. suis 2 and is facilitated by the ability of factor H (FH)-binding protein (Fhb) to bind FH on the bacterial surface, thereby impeding alternative pathway complement activation and phagocytic clearance. Here, C3b/C3d was found to bind to Fhb, along with FH, forming a large immune complex. The formation of this immune complex was mediated by domain II of Fhb via electrostatic and hydrophobic interactions, which, to our knowledge, is a new type of interaction. Interestingly, Fhb was found to be associated with the cell envelope and also present in the culture supernatant, where secreted Fhb inhibited complement activation via interactions with domain II, thereby enhancing antiphagocytic clearance by polymorphonuclear leukocytes. Thus, Fhb is a multifunctional bacterial protein, which binds host complement component C3 as well as FH and interferes with innate immune recognition in a secret protein manner. S. suis 2 therefore appears to have developed a new strategy to combat host innate immunity and enhance survival in host blood. PMID:27342778

  18. Fasciola gigantica: immunization of rabbits with proteins isolated from coproantigen.

    Science.gov (United States)

    Abdel-Rahman, Eman H; Abdel-Megeed, Kadria N

    2004-08-01

    Two fractions were isolated from coproantigen by ion-exchange chromatography in which DEAE cellulose was utilized. Both fractions and crude antigen were characterized by SDS-polyacrylamide gel electrophoresis which revealed 13 bands of molecular weight ranged from 205-31 in crude coproantigen. While fraction I resolved into six bands of molecular weight 198, 178, 148, 111, 101 & 45. Fraction II showed seven bands of 191 KDa, 178KDa, 166KDa, 118KDa, 98.5KDa, 72KDa & 32KDa. Fraction 1I was higher immunoreactivity than fraction by ELISA. Three immunoreactive bands of 191KDa, 118KDa & 98.5KDa were identified in fraction II using immunoblot assay. Five bands of 178KDa, 148KDa, 111KDa, 101KDa & 45KDa were detected in fraction I. Immunization of rabbits twice with fraction II in Freund's adjuvant with two weeks interval followed by challenge with F. gigantica metacercariae resulted in 66.6% protection from infection. The protection was assessed by detect-ion of hepatic damage, worm recoveries and antibody response. High level of IgG response in vaccinated rabbits than control infected ones occurred and being responsible for the recorded protection.

  19. Shigella Manipulates Host Immune Responses by Delivering Effector Proteins with Specific Roles

    Science.gov (United States)

    Ashida, Hiroshi; Mimuro, Hitomi; Sasakawa, Chihiro

    2015-01-01

    The intestinal epithelium deploys multiple defense systems against microbial infection to sense bacterial components and danger alarms, as well as to induce intracellular signal transduction cascades that trigger both the innate and the adaptive immune systems, which are pivotal for bacterial elimination. However, many enteric bacterial pathogens, including Shigella, deliver a subset of virulence proteins (effectors) via the type III secretion system (T3SS) that enable bacterial evasion from host immune systems; consequently, these pathogens are able to efficiently colonize the intestinal epithelium. In this review, we present and select recently discovered examples of interactions between Shigella and host immune responses, with particular emphasis on strategies that bacteria use to manipulate inflammatory outputs of host-cell responses such as cell death, membrane trafficking, and innate and adaptive immune responses. PMID:25999954

  20. Shigella manipulates host immune responses by delivering effector proteins with specific roles

    Directory of Open Access Journals (Sweden)

    Hiroshi eAshida

    2015-05-01

    Full Text Available The intestinal epithelium deploys multiple defense systems against microbial infection to sense bacterial components and danger alarms, as well as to induce intracellular signal transduction cascades that trigger both the innate and adaptive immune system, which are pivotal for bacterial elimination. However, many enteric bacterial pathogens, including Shigella, deliver a subset of virulence proteins (effectors via the type III secretion system (T3SS that enable bacterial evasion from host immune systems; consequently, these pathogens are able to efficiently colonize the intestinal epithelium. In this review, we present select recently discovered examples of interactions between Shigella and host immune responses, with particular emphasis on strategies that bacteria use to manipulate inflammatory outputs of host cell responses such as cell death, membrane trafficking, and innate and adaptive immune responses.

  1. Surfactant Protein-D Is Essential for Immunity to Helminth Infection.

    Science.gov (United States)

    Thawer, Sumaiyya; Auret, Jennifer; Schnoeller, Corinna; Chetty, Alisha; Smith, Katherine; Darby, Matthew; Roberts, Luke; Mackay, Rosie-Marie; Whitwell, Harry J; Timms, John F; Madsen, Jens; Selkirk, Murray E; Brombacher, Frank; Clark, Howard William; Horsnell, William G C

    2016-02-01

    Pulmonary epithelial cell responses can enhance type 2 immunity and contribute to control of nematode infections. An important epithelial product is the collectin Surfactant Protein D (SP-D). We found that SP-D concentrations increased in the lung following Nippostrongylus brasiliensis infection; this increase was dependent on key components of the type 2 immune response. We carried out loss and gain of function studies of SP-D to establish if SP-D was required for optimal immunity to the parasite. N. brasiliensis infection of SP-D-/- mice resulted in profound impairment of host innate immunity and ability to resolve infection. Raising pulmonary SP-D levels prior to infection enhanced parasite expulsion and type 2 immune responses, including increased numbers of IL-13 producing type 2 innate lymphoid cells (ILC2), elevated expression of markers of alternative activation by alveolar macrophages (alvM) and increased production of the type 2 cytokines IL-4 and IL-13. Adoptive transfer of alvM from SP-D-treated parasite infected mice into naïve recipients enhanced immunity to N. brasiliensis. Protection was associated with selective binding by the SP-D carbohydrate recognition domain (CRD) to L4 parasites to enhance their killing by alvM. These findings are the first demonstration that the collectin SP-D is an essential component of host innate immunity to helminths.

  2. Programming the composition of polymer blend particles for controlled immunity towards individual protein antigens.

    Science.gov (United States)

    Zhan, Xi; Shen, Hong

    2015-05-28

    In order for a more precise control over the quality and quantity of immune responses stimulated by synthetic particle-based vaccines, it is critical to control the colloidal stability of particles and the release of protein antigens in both extracellular space and intracellular compartments. Different proteins exhibit different sizes, charges and solubilities. This study focused on modulating the release and colloidal stability of proteins with varied isoelectric points. A polymer particle delivery platform made from the blend of three polymers, poly(lactic-co-glycolic acid) (PLGA) and two random pH-sensitive copolymers, were developed. Our study demonstrated its programmability with respective to individual proteins. We showed the colloidal stability of particles at neutral environment and the release of each individual protein at different pH environments were dependent on the ratio of two charge polymers. Subsequently, two antigenic proteins, ovalbumin (OVA) and Type 2 Herpes Simplex Virus (HSV-2) glycoprotein D (gD) protein, were incorporated into particles with systematically varied compositions. We demonstrated that the level of in vitro CD8(+) T cell and in vivo immune responses were dependent on the ratio of two charged polymers, which correlated well with the release of proteins. This study provided a promising design framework of pH-responsive synthetic vaccines for protein antigens of interest. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Identifying Schistosoma japonicum excretory/secretory proteins and their interactions with host immune system.

    Science.gov (United States)

    Liao, Qi; Yuan, Xiongying; Xiao, Hui; Liu, Changning; Lv, Zhiyue; Zhao, Yi; Wu, Zhongdao

    2011-01-01

    Schistosoma japonicum is a major infectious agent of schistosomiasis. It has been reported that large number of proteins excreted and secreted by S. japonicum during its life cycle are important for its infection and survival in definitive hosts. These proteins can be used as ideal candidates for vaccines or drug targets. In this work, we analyzed the protein sequences of S. japonicum and found that compared with other proteins in S. japonicum, excretory/secretory (ES) proteins are generally longer, more likely to be stable and enzyme, more likely to contain immune-related binding peptides and more likely to be involved in regulation and metabolism processes. Based on the sequence difference between ES and non-ES proteins, we trained a support vector machine (SVM) with much higher accuracy than existing approaches. Using this SVM, we identified 191 new ES proteins in S. japonicum, and further predicted 7 potential interactions between these ES proteins and human immune proteins. Our results are useful to understand the pathogenesis of schistosomiasis and can serve as a new resource for vaccine or drug targets discovery for anti-schistosome.

  4. Identifying Schistosoma japonicum excretory/secretory proteins and their interactions with host immune system.

    Directory of Open Access Journals (Sweden)

    Qi Liao

    Full Text Available Schistosoma japonicum is a major infectious agent of schistosomiasis. It has been reported that large number of proteins excreted and secreted by S. japonicum during its life cycle are important for its infection and survival in definitive hosts. These proteins can be used as ideal candidates for vaccines or drug targets. In this work, we analyzed the protein sequences of S. japonicum and found that compared with other proteins in S. japonicum, excretory/secretory (ES proteins are generally longer, more likely to be stable and enzyme, more likely to contain immune-related binding peptides and more likely to be involved in regulation and metabolism processes. Based on the sequence difference between ES and non-ES proteins, we trained a support vector machine (SVM with much higher accuracy than existing approaches. Using this SVM, we identified 191 new ES proteins in S. japonicum, and further predicted 7 potential interactions between these ES proteins and human immune proteins. Our results are useful to understand the pathogenesis of schistosomiasis and can serve as a new resource for vaccine or drug targets discovery for anti-schistosome.

  5. CBL-interacting protein kinase 6 negatively regulates immune response to Pseudomonas syringae in Arabidopsis.

    Science.gov (United States)

    Sardar, Atish; Nandi, Ashis Kumar; Chattopadhyay, Debasis

    2017-06-15

    Cytosolic calcium ion (Ca2+) is an essential mediator of the plant innate immune response. Here, we report that a calcium-regulated protein kinase Calcineurin B-like protein (CBL)-interacting protein kinase 6 (CIPK6) functions as a negative regulator of immunity against the bacterial pathogen Pseudomonas syringae in Arabidopsis thaliana. Arabidopsis lines with compromised expression of CIPK6 exhibited enhanced disease resistance to the bacterial pathogen and to P. syringae harboring certain but not all avirulent effectors, while restoration of CIPK6 expression resulted in abolition of resistance. Plants overexpressing CIPK6 were more susceptible to P. syringae. Enhanced resistance in the absence of CIPK6 was accompanied by increased accumulation of salicylic acid and elevated expression of defense marker genes. Salicylic acid accumulation was essential for improved immunity in the absence of CIPK6. CIPK6 negatively regulated the oxidative burst associated with perception of pathogen-associated microbial patterns (PAMPs) and bacterial effectors. Accelerated and enhanced activation of the mitogen-activated protein kinase cascade in response to bacterial and fungal elicitors was observed in the absence of CIPK6. The results of this study suggested that CIPK6 negatively regulates effector-triggered and PAMP-triggered immunity in Arabidopsis. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  6. Opa proteins and CEACAMs: pathways of immune engagement for pathogenic Neisseria.

    Science.gov (United States)

    Sadarangani, Manish; Pollard, Andrew J; Gray-Owen, Scott D

    2011-05-01

    Neisseria meningitidis and Neisseria gonorrhoeae are globally important pathogens, which in part owe their success to their ability to successfully evade human immune responses over long periods. The phase-variable opacity-associated (Opa) adhesin proteins are a major surface component of these organisms, and are responsible for bacterial adherence and entry into host cells and interactions with the immune system. Most immune interactions are mediated via binding to members of the carcinoembryonic antigen cell adhesion molecule (CEACAM) family. These Opa variants are able to bind to different receptors of the CEACAM family on epithelial cells, neutrophils, and T and B lymphocytes, influencing the innate and adaptive immune responses. Increased epithelial cell adhesion creates the potential for prolonged infection, invasion and dissemination. Furthermore, Opa proteins may inhibit T-lymphocyte activation and proliferation, B-cell antibody production, and innate inflammatory responses by infected epithelia, in addition to conferring increased resistance to antibody-dependent, complement-mediated killing. While vaccines containing Opa proteins could induce adhesion-blocking and bactericidal antibodies, the consequence of CEACAM binding by a candidate Opa-containing vaccine requires further investigation. This review summarizes current knowledge of the immunological consequences of the interaction between meningococcal and gonococcal Opa proteins and human CEACAMs, considering the implications for pathogenesis and vaccine development. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  7. Naturally Acquired Human Immunity to Pneumococcus Is Dependent on Antibody to Protein Antigens

    NARCIS (Netherlands)

    Wilson, R. (Robert); J. Cohen (Jonathan); Reglinski, M. (Mark); R.J. Jose; Chan, W.Y. (Win Yan); Marshall, H. (Helina); C.P. de Vogel (Corné); S.B. Gordon (Stephen); Goldblatt, D. (David); Petersen, F.C. (Fernanda C.); H. Baxendale (Helen); Brown, J.S. (Jeremy S.)

    2017-01-01

    textabstractNaturally acquired immunity against invasive pneumococcal disease (IPD) is thought to be dependent on anti-capsular antibody. However nasopharyngeal colonisation by Streptococcus pneumoniae also induces antibody to protein antigens that could be protective. We have used human intravenous

  8. Characterization of protective immune responses induced by pneumococcal surface protein A in fusion with pneumolysin derivatives.

    Directory of Open Access Journals (Sweden)

    Cibelly Goulart

    Full Text Available Pneumococcal surface protein A (PspA and Pneumolysin derivatives (Pds are important vaccine candidates, which can confer protection in different models of pneumococcal infection. Furthermore, the combination of these two proteins was able to increase protection against pneumococcal sepsis in mice. The present study investigated the potential of hybrid proteins generated by genetic fusion of PspA fragments to Pds to increase cross-protection against fatal pneumococcal infection. Pneumolisoids were fused to the N-terminus of clade 1 or clade 2 pspA gene fragments. Mouse immunization with the fusion proteins induced high levels of antibodies against PspA and Pds, able to bind to intact pneumococci expressing a homologous PspA with the same intensity as antibodies to rPspA alone or the co-administered proteins. However, when antibody binding to pneumococci with heterologous PspAs was examined, antisera to the PspA-Pds fusion molecules showed stronger antibody binding and C3 deposition than antisera to co-administered proteins. In agreement with these results, antisera against the hybrid proteins were more effective in promoting the phagocytosis of bacteria bearing heterologous PspAs in vitro, leading to a significant reduction in the number of bacteria when compared to co-administered proteins. The respective antisera were also capable of neutralizing the lytic activity of Pneumolysin on sheep red blood cells. Finally, mice immunized with fusion proteins were protected against fatal challenge with pneumococcal strains expressing heterologous PspAs. Taken together, the results suggest that PspA-Pd fusion proteins comprise a promising vaccine strategy, able to increase the immune response mediated by cross-reactive antibodies and complement deposition to heterologous strains, and to confer protection against fatal challenge.

  9. Stealth proteins: in silico identification of a novel protein family rendering bacterial pathogens invisible to host immune defense.

    Directory of Open Access Journals (Sweden)

    Peter Sperisen

    2005-11-01

    Full Text Available There are a variety of bacterial defense strategies to survive in a hostile environment. Generation of extracellular polysaccharides has proved to be a simple but effective strategy against the host's innate immune system. A comparative genomics approach led us to identify a new protein family termed Stealth, most likely involved in the synthesis of extracellular polysaccharides. This protein family is characterized by a series of domains conserved across phylogeny from bacteria to eukaryotes. In bacteria, Stealth (previously characterized as SacB, XcbA, or WefC is encoded by subsets of strains mainly colonizing multicellular organisms, with evidence for a protective effect against the host innate immune defense. More specifically, integrating all the available information about Stealth proteins in bacteria, we propose that Stealth is a D-hexose-1-phosphoryl transferase involved in the synthesis of polysaccharides. In the animal kingdom, Stealth is strongly conserved across evolution from social amoebas to simple and complex multicellular organisms, such as Dictyostelium discoideum, hydra, and human. Based on the occurrence of Stealth in most Eukaryotes and a subset of Prokaryotes together with its potential role in extracellular polysaccharide synthesis, we propose that metazoan Stealth functions to regulate the innate immune system. Moreover, there is good reason to speculate that the acquisition and spread of Stealth could be responsible for future epidemic outbreaks of infectious diseases caused by a large variety of eubacterial pathogens. Our in silico identification of a homologous protein in the human host will help to elucidate the causes of Stealth-dependent virulence. At a more basic level, the characterization of the molecular and cellular function of Stealth proteins may shed light on fundamental mechanisms of innate immune defense against microbial invasion.

  10. Stealth Proteins: In Silico Identification of a Novel Protein Family Rendering Bacterial Pathogens Invisible to Host Immune Defense.

    Directory of Open Access Journals (Sweden)

    2005-11-01

    Full Text Available There are a variety of bacterial defense strategies to survive in a hostile environment. Generation of extracellular polysaccharides has proved to be a simple but effective strategy against the host's innate immune system. A comparative genomics approach led us to identify a new protein family termed Stealth, most likely involved in the synthesis of extracellular polysaccharides. This protein family is characterized by a series of domains conserved across phylogeny from bacteria to eukaryotes. In bacteria, Stealth (previously characterized as SacB, XcbA, or WefC is encoded by subsets of strains mainly colonizing multicellular organisms, with evidence for a protective effect against the host innate immune defense. More specifically, integrating all the available information about Stealth proteins in bacteria, we propose that Stealth is a D-hexose-1-phosphoryl transferase involved in the synthesis of polysaccharides. In the animal kingdom, Stealth is strongly conserved across evolution from social amoebas to simple and complex multicellular organisms, such as Dictyostelium discoideum, hydra, and human. Based on the occurrence of Stealth in most Eukaryotes and a subset of Prokaryotes together with its potential role in extracellular polysaccharide synthesis, we propose that metazoan Stealth functions to regulate the innate immune system. Moreover, there is good reason to speculate that the acquisition and spread of Stealth could be responsible for future epidemic outbreaks of infectious diseases caused by a large variety of eubacterial pathogens. Our in silico identification of a homologous protein in the human host will help to elucidate the causes of Stealth-dependent virulence. At a more basic level, the characterization of the molecular and cellular function of Stealth proteins may shed light on fundamental mechanisms of innate immune defense against microbial invasion.

  11. Immune responsiveness of Japanese quail selected for egg yolk testosterone content under severe protein restriction.

    Science.gov (United States)

    Kankova, Zuzana; Okuliarova, Monika; Zeman, Michal

    2014-11-01

    Yolk testosterone concentrations vary in response to environmental conditions and different testosterone contents can subsequently modify the phenotypic traits of offspring. Apart from effects on growth, proactive behaviour and secondary sexual characteristics, the possible negative impacts of maternal testosterone on the immune system are often considered a limitation for its deposition. The effects of maternal testosterone can be modulated by postnatal environmental conditions, such as the availability of food resources. However, the majority of studies considering the effects of maternal testosterone on the immune system have been conducted under optimum conditions. We evaluated the influence of genetic selection for high (HET) and low (LET) egg testosterone content in Japanese quail on immune responsiveness of offspring to phytohaemagglutinin (PHA) and lipopolysaccharide (LPS) stimulation under severe protein restriction. Protein restriction negatively influenced body weight and performance in the PHA-test. We observed an increase in Cort (corticosterone) and He/Ly (heterophil/lymphocyte ratio) after LPS, while no changes occurred in total IgY levels in the protein-restricted group. HET quails showed higher body mass and total IgY levels and lower He/Ly ratio than LET quails, while the PHA index and Cort concentration did not differ between lines. No interactions were found between protein restriction and genetic line. In conclusion, the immune response was not compromised under conditions of severe protein restriction in the faster growing HET line compared with the LET line. We hypothesise that the immune responsiveness of birds with higher yolk testosterone may be linked with other maternally-derived substances in a context-dependent manner. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. Sequence-structure-function relations of the mosquito leucine-rich repeat immune proteins

    Directory of Open Access Journals (Sweden)

    Povelones Michael

    2010-09-01

    Full Text Available Abstract Background The discovery and characterisation of factors governing innate immune responses in insects has driven the elucidation of many immune system components in mammals and other organisms. Focusing on the immune system responses of the malaria mosquito, Anopheles gambiae, has uncovered an array of components and mechanisms involved in defence against pathogen infections. Two of these immune factors are LRIM1 and APL1C, which are leucine-rich repeat (LRR containing proteins that activate complement-like defence responses against malaria parasites. In addition to their LRR domains, these leucine-rich repeat immune (LRIM proteins share several structural features including signal peptides, patterns of cysteine residues, and coiled-coil domains. Results The identification and characterisation of genes related to LRIM1 and APL1C revealed putatively novel innate immune factors and furthered the understanding of their likely molecular functions. Genomic scans using the shared features of LRIM1 and APL1C identified more than 20 LRIM-like genes exhibiting all or most of their sequence features in each of three disease-vector mosquitoes with sequenced genomes: An. gambiae, Aedes aegypti, and Culex quinquefasciatus. Comparative sequence analyses revealed that this family of mosquito LRIM-like genes is characterised by a variable number of 6 to 14 LRRs of different lengths. The "Long" LRIM subfamily, with 10 or more LRRs, and the "Short" LRIMs, with 6 or 7 LRRs, also share the signal peptide, cysteine residue patterning, and coiled-coil sequence features of LRIM1 and APL1C. The "TM" LRIMs have a predicted C-terminal transmembrane region, and the "Coil-less" LRIMs exhibit the characteristic LRIM sequence signatures but lack the C-terminal coiled-coil domains. Conclusions The evolutionary plasticity of the LRIM LRR domains may provide templates for diverse recognition properties, while their coiled-coil domains could be involved in the formation

  13. Tumor necrosis factor-α-induced protein 1 and immunity to hepatitis B virus

    Science.gov (United States)

    Lin, Marie C; Lee, Nikki P; Zheng, Ning; Yang, Pai-Hao; Wong, Oscar G; Kung, Hsiang-Fu; Hui, Chee-Kin; Luk, John M; Lau, George Ka-Kit

    2005-01-01

    AIM: To compare the gene expression profile in a pair of HBV-infected twins. METHODS: The gene expression profile was compared in a pair of HBV-infected twins. RESULTS: The twins displayed different disease outcomes. One acquired natural immunity against HBV, whereas the other became a chronic HBV carrier. Eighty-eight and forty-six genes were found to be up- or down-regulated in their PBMCs, respectively. Tumor necrosis factor-alpha-induced protein 1 (TNF-αIP1) that expressed at a higher level in the HBV-immune twins was identified and four pairs of siblings with HBV immunity by RT-PCR. However, upon HBV core antigen stimulation, TNF-αIP1 was downregulated in PBMCs from subjects with immunity, whereas it was slightly upregulated in HBV carriers. Bioinformatics analysis revealed a K+ channel tetramerization domain in TNF-αIP1 that shares a significant homology with some human, mouse, and C elegan proteins. CONCLUSION: TNF-αIP1 may play a role in the innate immunity against HBV. PMID:16437679

  14. Detecting Immune System Response Proteins in a 500 Year-old Inca Mummy

    Science.gov (United States)

    Corthals, A.; Davalos, L.; Martin, D.W.; Rieger, R.; Chen, E.I.; Koller, A.

    2011-01-01

    Disease detection in ancient human samples currently relies on genomic-based assays, which are error prone due to contamination and cannot distinguish between active and latent pathogenic infection. On the other hand, protein-based assays such as global protein profiling offer complementary alternatives for the pathological diagnosis of archeological specimen. The discovery of three Inca mummies in 1998, perfectly preserved in the permafrost of the high Andes, allowed us to analyze mummy samples by protein-based and genomic-based assay. A buccal swab from one of the 500 year old mummy was analyzed by shotgun proteomics to detect the protein profile. Among the identified proteins, we found a signature of proteins indicating an immune response to a bacterial infection at the time of the mummy's death. Based on the external visible symptoms and the gamut of immune response proteins obtained from the mouth swab, we suspected that the pulmonary infection was caused by Mycobacterium. PCR assay followed by direct sequencing of the PCR products confirmed the presence of Mycobacterium sp. in the mouth swab. Until now, immunoassays have been the only way to detect an active immune response and infer infection in historical samples, but these were plagued by low specificity and sensitivity. However, we demonstrate here the feasibility of incorporating global protein profiling in the diagnosis of infection from archeological samples. Protein signatures obtained from these samples could be extremely useful in determining the status of infection while genomic-based assays can be used to detect the identity of the pathogen.

  15. A cDNA Immunization Strategy to Generate Nanobodies against Membrane Proteins in Native Conformation

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    Thomas Eden

    2018-01-01

    Full Text Available Nanobodies (Nbs are soluble, versatile, single-domain binding modules derived from the VHH variable domain of heavy-chain antibodies naturally occurring in camelids. Nbs hold huge promise as novel therapeutic biologics. Membrane proteins are among the most interesting targets for therapeutic Nbs because they are accessible to systemically injected biologics. In order to be effective, therapeutic Nbs must recognize their target membrane protein in native conformation. However, raising Nbs against membrane proteins in native conformation can pose a formidable challenge since membrane proteins typically contain one or more hydrophobic transmembrane regions and, therefore, are difficult to purify in native conformation. Here, we describe a highly efficient genetic immunization strategy that circumvents these difficulties by driving expression of the target membrane protein in native conformation by cells of the immunized camelid. The strategy encompasses ballistic transfection of skin cells with cDNA expression plasmids encoding one or more orthologs of the membrane protein of interest and, optionally, other costimulatory proteins. The plasmid is coated onto 1 µm gold particles that are then injected into the shaved and depilated skin of the camelid. A gene gun delivers a helium pulse that accelerates the DNA-coated particles to a velocity sufficient to penetrate through multiple layers of cells in the skin. This results in the exposure of the extracellular domains of the membrane protein on the cell surface of transfected cells. Repeated immunization drives somatic hypermutation and affinity maturation of target-specific heavy-chain antibodies. The VHH/Nb coding region is PCR-amplified from B cells obtained from peripheral blood or a lymph node biopsy. Specific Nbs are selected by phage display or by screening of Nb-based heavy-chain antibodies expressed as secretory proteins in transfected HEK cells. Using this strategy, we have successfully

  16. A cDNA Immunization Strategy to Generate Nanobodies against Membrane Proteins in Native Conformation

    Science.gov (United States)

    Eden, Thomas; Menzel, Stephan; Wesolowski, Janusz; Bergmann, Philine; Nissen, Marion; Dubberke, Gudrun; Seyfried, Fabienne; Albrecht, Birte; Haag, Friedrich; Koch-Nolte, Friedrich

    2018-01-01

    Nanobodies (Nbs) are soluble, versatile, single-domain binding modules derived from the VHH variable domain of heavy-chain antibodies naturally occurring in camelids. Nbs hold huge promise as novel therapeutic biologics. Membrane proteins are among the most interesting targets for therapeutic Nbs because they are accessible to systemically injected biologics. In order to be effective, therapeutic Nbs must recognize their target membrane protein in native conformation. However, raising Nbs against membrane proteins in native conformation can pose a formidable challenge since membrane proteins typically contain one or more hydrophobic transmembrane regions and, therefore, are difficult to purify in native conformation. Here, we describe a highly efficient genetic immunization strategy that circumvents these difficulties by driving expression of the target membrane protein in native conformation by cells of the immunized camelid. The strategy encompasses ballistic transfection of skin cells with cDNA expression plasmids encoding one or more orthologs of the membrane protein of interest and, optionally, other costimulatory proteins. The plasmid is coated onto 1 µm gold particles that are then injected into the shaved and depilated skin of the camelid. A gene gun delivers a helium pulse that accelerates the DNA-coated particles to a velocity sufficient to penetrate through multiple layers of cells in the skin. This results in the exposure of the extracellular domains of the membrane protein on the cell surface of transfected cells. Repeated immunization drives somatic hypermutation and affinity maturation of target-specific heavy-chain antibodies. The VHH/Nb coding region is PCR-amplified from B cells obtained from peripheral blood or a lymph node biopsy. Specific Nbs are selected by phage display or by screening of Nb-based heavy-chain antibodies expressed as secretory proteins in transfected HEK cells. Using this strategy, we have successfully generated agonistic

  17. Effects of inadequate maternal dietary protein:carbohydrate ratios during pregnancy on offspring immunity in pigs

    Directory of Open Access Journals (Sweden)

    Tuchscherer Margret

    2012-11-01

    Full Text Available Abstract Background Inadequate nutrition in utero may retard foetal growth and alter physiological development of offspring. This study investigated the effects of low and high protein diets fed to primiparous German Landrace sows throughout pregnancy on the immune function of their offspring at different ages. Sows were fed diets with adequate (AP, 12.1%; n = 13, low (LP, 6.5%; n = 15, or high (HP, 30%; n = 14 protein content, made isoenergetic by varying carbohydrate levels. Cortisol, total protein and immunoglobulin (IgG, IgM, IgA concentrations were measured in the blood of sows over the course of pregnancy. Cortisol, total protein, immunoglobulins, lymphocyte proliferation, immune cell counts, and cytokines were assessed in the blood of offspring at baseline and under challenging conditions (weaning; lipopolysaccharide (LPS administration. Results In sows, the LP diet increased cortisol (P P P P + cell percentage and the CD4+/CD8+ ratio increased after weaning (P P = 0.09 and HP (P P  Conclusions Our results indicate that both low and high protein:carbohydrate ratios in the diet of pregnant sows can induce short-term as well as long-lasting effects on immune competence in piglets that may have serious consequences for host defence against bacterial pathogens.

  18. Protein Kinase G Induces an Immune Response in Cows Exposed to Mycobacterium avium Subsp. paratuberculosis

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    Horacio Bach

    2018-01-01

    Full Text Available To establish infection, pathogens secrete virulence factors, such as protein kinases and phosphatases, to modulate the signal transduction pathways used by host cells to initiate immune response. The protein MAP3893c is annotated in the genome sequence of Mycobacterium avium subspecies paratuberculosis (MAP, the causative agent of Johne’s disease, as the serine/threonine protein kinase G (PknG. In this work, we report that PknG is a functional kinase that is secreted within macrophages at early stages of infection. The antigen is able to induce an immune response from cattle exposed to MAP in the form of interferon gamma production after stimulation of whole blood with PknG. These findings suggest that PknG may contribute to the pathogenesis of MAP by phosphorylating macrophage signalling and/or adaptor molecules as observed with other pathogenic mycobacterial species.

  19. Immune response in mice to ingested soya protein: antibody production, oral tolerance and maternal transfer

    DEFF Research Database (Denmark)

    Christensen, Hanne Risager; Pedersen, Susanne Brix; Frøkiær, Hanne

    2004-01-01

    by ELISA, and to the presence of oral tolerance detected as a suppressed antibody and cell-proliferation response upon immunisation with soya protein. F0 mice generated soya-specific antibodies, while oral tolerance to the same soya proteins was also clearly induced. When F0 dams were transferred to soya...... antibody response in the offspring, bat in this case in the absence of oral tolerance. This indicates that, under certain conditions, factors involved in spontaneous antibody production can be transmitted from mother to offspring. Understanding the immune response to soya protein ingested under healthy...

  20. Quality of colostral passive immunity and pattern of serum protein fluctuation in newborn calves

    Directory of Open Access Journals (Sweden)

    Pauletti Patricia

    2003-01-01

    Full Text Available Immunity acquired by newborn animals is known as passive immunity, and for ruminants, antibody acquisition depends on the ingestion and absorption of adequate amounts of immunoglobulins from colostrum. This study relates different initial levels of acquired passive protection and serum total protein (TP and immunoglobulin G (IgG. Serum immunoglobulin concentration and total protein were evaluated for female Holstein calves in the first sixty days of life. Animals were separated into three groups according to their initial level of passive immunity: group 1- animals with a low level of passive immunity (below 20 mg mL-1; group 2- animals with a medium level (between 20 and 30 mg mL-1, and group 3- animals with a high level (above 30 mg mL-1. Serum total protein was determined through the biuret method and IgG was determined by radial immunodiffusion. Data were analyzed as a completely randomized, split-plot statistical design. Fluctuation of the variables along the experimental period was determined through non-linear regression by the DUD method (PROC NLIN - Non Linear SAS. Animals with low antibody acquisition started to produce antibodies earlier, reflecting a compensatory synthesis. On the other hand, animals having adequate levels exhibited an extended period of immunoglobulin catabolism and the beginning of the endogenous phase was delayed. Regardless initial levels, the fluctuations in IgG contents occurred around adequate physiological concentrations, ranging from 20 to 25 mg mL-1.

  1. The innate immune protein Nod2 binds directly to MDP, a bacterial cell wall fragment.

    Science.gov (United States)

    Grimes, Catherine Leimkuhler; Ariyananda, Lushanti De Zoysa; Melnyk, James E; O'Shea, Erin K

    2012-08-22

    Mammalian Nod2 is an intracellular protein that is implicated in the innate immune response to the bacterial cell wall and is associated with the development of Crohn's disease, Blau syndrome, and gastrointestinal cancers. Nod2 is required for an immune response to muramyl dipeptide (MDP), an immunostimulatory fragment of bacterial cell wall, but it is not known whether MDP binds directly to Nod2. We report the expression and purification of human Nod2 from insect cells. Using novel MDP self-assembled monolayers (SAMs), we provide the first biochemical evidence for a direct, high-affinity interaction between Nod2 and MDP.

  2. The immunoreceptor adapter protein DAP12 suppresses B lymphocyte?driven adaptive immune responses

    OpenAIRE

    Nakano-Yokomizo, Takako; Tahara-Hanaoka, Satoko; Nakahashi-Oda, Chigusa; Nabekura, Tsukasa; Tchao, Nadia K.; Kadosaki, Momoko; Totsuka, Naoya; Kurita, Naoki; Nakamagoe, Kiyotaka; Tamaoka, Akira; Takai, Toshiyuki; Yasui, Teruhito; Kikutani, Hitoshi; Honda, Shin-ichiro; Shibuya, Kazuko

    2011-01-01

    DAP12, an immunoreceptor tyrosine-based activation motif?bearing adapter protein, is involved in innate immunity mediated by natural killer cells and myeloid cells. We show that DAP12-deficient mouse B cells and B cells from a patient with Nasu-Hakola disease, a recessive genetic disorder resulting from loss of DAP12, showed enhanced proliferation after stimulation with anti-IgM or CpG. Myeloid-associated immunoglobulin-like receptor (MAIR) II (Cd300d) is a DAP12-associated immune receptor. L...

  3. Circumsporozoite Protein-Specific Kd-Restricted CD8+ T Cells Mediate Protective Antimalaria Immunity in Sporozoite-Immunized MHC-I-Kd Transgenic Mice

    Directory of Open Access Journals (Sweden)

    Jing Huang

    2014-01-01

    Full Text Available Although the roles of CD8+ T cells and a major preerythrocytic antigen, the circumsporozoite (CS protein, in contributing protective antimalaria immunity induced by radiation-attenuated sporozoites, have been shown by a number of studies, the extent to which these players contribute to antimalaria immunity is still unknown. To address this question, we have generated C57BL/6 (B6 transgenic (Tg mice, expressing Kd molecules under the MHC-I promoter, called MHC-I-Kd-Tg mice. In this study, we first determined that a single immunizing dose of IrPySpz induced a significant level of antimalaria protective immunity in MHC-I-Kd-Tg mice but not in B6 mice. Then, by depleting various T-cell subsets in vivo, we determined that CD8+ T cells are the main mediator of the protective immunity induced by IrPySpz. Furthermore, when we immunized (MHC-I-Kd-Tg × CS-Tg F1 mice with IrPySpz after crossing MHC-I-Kd-Tg mice with PyCS-transgenic mice (CS-Tg, which are unable to mount PyCS-specific immunity, we found that IrPySpz immunization failed to induce protective antimalaria immunity in (MHC-I-Kd-Tg × CS-Tg F1 mice, thus indicating the absence of PyCS antigen-dependent immunity in these mice. These results indicate that protective antimalaria immunity induced by IrPySpz in MHC-I-Kd-Tg mice is mediated by CS protein-specific, Kd-restricted CD8+ T cells.

  4. Systemic Immunization with Papillomavirus L1 Protein Completely Prevents the Development of Viral Mucosal Papillomas

    Science.gov (United States)

    Suzich, Joann A.; Ghim, Shin-Je; Palmer-Hill, Frances J.; White, Wendy I.; Tamura, James K.; Bell, Judith A.; Newsome, Joseph A.; Bennett Jenson, A.; Schlegel, Richard

    1995-12-01

    Infection of mucosal epithelium by papillomaviruses is responsible for the induction of genital and oral warts and plays a critical role in the development of human cervical and oropharyngeal cancer. We have employed a canine model to develop a systemic vaccine that completely protects against experimentally induced oral mucosal papillomas. The major capsid protein, L1, of canine oral papillomavirus (COPV) was expressed in Sf9 insect cells in native conformation. L1 protein, which self-assembled into virus-like particles, was purified on CsCl gradients and injected intradermally into the foot pad of beagles. Vaccinated animals developed circulating antibodies against COPV and became completely resistant to experimental challenge with COPV. Successful immunization was strictly dependent upon native L1 protein conformation and L1 type. Partial protection was achieved with as little as 0.125 ng of L1 protein, and adjuvants appeared useful for prolonging the host immune response. Serum immunoglobulins passively transferred from COPV L1-immunized beagles to naive beagles conferred protection from experimental infection with COPV. Our results indicate the feasibility of developing a human vaccine to prevent mucosal papillomas, which can progress to malignancy.

  5. Induction of Mucosal and Systemic Immunity to a Recombinant Simian Immunodeficiency Viral Protein

    Science.gov (United States)

    Lehner, T.; Bergmeier, L. A.; Panagiotidi, C.; Tao, L.; Brookes, R.; Klavinskis, L. S.; Walker, P.; Walker, J.; Ward, R. G.; Hussain, L.; Gearing, A. J. H.; Adams, S. E.

    1992-11-01

    Heterosexual transmission through the cervico-vaginal mucosa is the principal route of human immunodeficiency virus (HIV) infection in Africa and is increasing in the United States and Europe. Vaginal immunization with simian immunodeficiency virus (SIV) had not yet been studied in nonhuman primates. Immune responses in macaques were investigated by stimulation of the genital and gut-associated lymphoid tissue with a recombinant, particulate SIV antigen. Vaginal, followed by oral, administration of the vaccine elicited three types of immunity: (i) gag protein p27-specific, secretory immunoglobulin A (IgA) and immunoglobulin G (IgG) in the vaginal fluid, (ii) specific CD4^+ T cell proliferation and helper function in B cell p27-specific IgA synthesis in the genital lymph nodes, and (iii) specific serum IgA and IgG, with CD4^+ T cell proliferative and helper functions in the circulating blood.

  6. Immunization with H1, HASPB1 and MML Leishmania proteins in a vaccine trial against experimental canine leishmaniasis

    OpenAIRE

    Moreno, J.; Nieto, J.; Masina, S.; Ca?avate, C.; Cruz, I.; Chicharro, C.; Carrillo, E.; Napp, S.; Reymond, C.; Kaye, P.M.; Smith, D.F.; Fasel, N.; Alvar, J.

    2007-01-01

    The protective capabilities of three Leishmania recombinant proteins - histone 1 (H1) and hydrophilic acylated surface protein B1 (HASPB1) immunized singly, or together as a protein cocktail vaccine with Montanide, and the polyprotein MML immunized with MPL-SE adjuvant - were assessed in beagle dogs. Clinical examination of the dogs was carried out periodically under blinded conditions and the condition of the dogs defined as asymptomatic or symptomatic. At the end of the trial, we were able ...

  7. Immunization against Rumen Methanogenesis by Vaccination with a New Recombinant Protein.

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    Litai Zhang

    Full Text Available Vaccination through recombinant proteins against rumen methanogenesis provides a mitigation approach to reduce enteric methane (CH4 emissions in ruminants. The objective of present study was to evaluate the in vivo efficacy of a new vaccine candidate protein (EhaF on methanogenesis and microbial population in the rumen of goats. We amplified the gene mru 1407 encoding protein EhaF using fresh rumen fluid samples of mature goats and successfully expressed recombinant protein (EhaF in Escherichia coli Rosetta. This product was evaluated using 12 mature goats with half for control and other half injected with 400ug/goat the purified recombinant protein in day 1 and two subsequent booster immunizations in day 35 and 49. All measurements were undertaken from 63 to 68 days after the initial vaccination, with CH4 emissions determined using respiration calorimeter chambers. The results showed that the vaccination caused intensive immune responses in serum and saliva, although it had no significant effect on total enteric CH4 emissions and methanogen population in the rumen, when compared with the control goats. However, the vaccination altered the composition of rumen bacteria, especially the abundance of main phylum Firmicutes and genus Prevotella. The results indicate that protein EhaF might not be an effective vaccine to reduce enteric CH4 emissions but our vaccine have potential to influence the rumen ecosystem of goats.

  8. Oral and parenteral immunization of chickens (Gallus gallus) against West Nile virus with recombinant envelope protein

    Science.gov (United States)

    Fassbinder-Orth, C. A.; Hofmeister, Erik K.; Weeks-Levy, C.; Karasov, W.H.

    2009-01-01

    West Nile virus (WNV) causes morbidity and mortality in humans, horses, and in more than 315 bird species in North America. Currently approved WNV vaccines are designed for parenteral administration and, as yet, no effective oral WNV vaccines have been developed. WNV envelope (E) protein is a highly antigenic protein that elicits the majority of virus-neutralizing antibodies during a WNV immune response. Leghorn chickens were given three vaccinations (each 2 wk apart) of E protein orally (20 ??g or 100 ??g/dose), of E protein intramuscularly (IM, 20 ??g/dose), or of adjuvant only (control group) followed by a WNV challenge. Viremias were measured post-WNV infection, and three new enzyme-linked immunosorbent assays were developed for quantifying IgM, IgY, and IgA-mediated immune response of birds following WNV infection. WNV viremia levels were significantly lower in the IM group than in both oral groups and the control group. Total WNV E protein-specific IgY production was significantly greater, and WNV nonstructural 1-specific IgY was significantly less, in the IM group compared to all other treatment groups. The results of this study indicate that IM vaccination of chickens with E protein is protective against WNV infection and results in a significantly different antibody production profile as compared to both orally vaccinated and nonvaccinated birds. ?? 2009 American Association of Avian Pathologists.

  9. Functional analysis of the human cytomegalovirus immune evasion protein, pUS322kDa

    International Nuclear Information System (INIS)

    Zhao Yiqiang; Biegalke, Bonita J.

    2003-01-01

    Human cytomegalovirus (HCMV) is an important opportunistic pathogen that infrequently causes disease in individuals with mature immune systems. The HCMV US3 gene encodes a 22-kDa protein that interferes with immune recognition of virally infected cells. The 22-kDa US3 protein binds to major histocompatibility complex (MHC) class I complexes, retaining them in the endoplasmic reticulum (ER), thereby decreasing the presentation of viral antigen to cytotoxic T cells. Our studies demonstrate that correct folding of the ER lumenal domain of the US3 protein is essential, but insufficient for interactions with MHC class I complexes. We demonstrate a requirement for the transmembrane domain of the 22-kDa US3 protein, confirming the results of others, and also show that the cytosolic carboxyl-terminal tail influences the function of the protein. Anchoring of the ER-lumenal immunoglobulin-like fold of the US3 protein to the membrane of the endoplasmic reticulum is critical for the binding and retention of MHC class I complexes

  10. Determinants of antigenicity and specificity in immune response for protein sequences

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    Li Cheng

    2011-06-01

    Full Text Available Abstract Background Target specific antibodies are pivotal for the design of vaccines, immunodiagnostic tests, studies on proteomics for cancer biomarker discovery, identification of protein-DNA and other interactions, and small and large biochemical assays. Therefore, it is important to understand the properties of protein sequences that are important for antigenicity and to identify small peptide epitopes and large regions in the linear sequence of the proteins whose utilization result in specific antibodies. Results Our analysis using protein properties suggested that sequence composition combined with evolutionary information and predicted secondary structure, as well as solvent accessibility is sufficient to predict successful peptide epitopes. The antigenicity and the specificity in immune response were also found to depend on the epitope length. We trained the B-Cell Epitope Oracle (BEOracle, a support vector machine (SVM classifier, for the identification of continuous B-Cell epitopes with these protein properties as learning features. The BEOracle achieved an F1-measure of 81.37% on a large validation set. The BEOracle classifier outperformed the classical methods based on propensity and sophisticated methods like BCPred and Bepipred for B-Cell epitope prediction. The BEOracle classifier also identified peptides for the ChIP-grade antibodies from the modENCODE/ENCODE projects with 96.88% accuracy. High BEOracle score for peptides showed some correlation with the antibody intensity on Immunofluorescence studies done on fly embryos. Finally, a second SVM classifier, the B-Cell Region Oracle (BROracle was trained with the BEOracle scores as features to predict the performance of antibodies generated with large protein regions with high accuracy. The BROracle classifier achieved accuracies of 75.26-63.88% on a validation set with immunofluorescence, immunohistochemistry, protein arrays and western blot results from Protein Atlas database

  11. Innovative immunization protocols using chimeric recombinant protein for the production of polyspecific loxoscelic antivenom in horses.

    Science.gov (United States)

    Figueiredo, Luís F M; Dias-Lopes, Camila; Alvarenga, Larissa M; Mendes, Thais M; Machado-de-Ávila, Ricardo A; McCormack, Jessica; Minozzo, João C; Kalapothakis, Evanguedes; Chávez-Olórtegui, Carlos

    2014-08-01

    A chimeric protein (rCpLi) was constructed expressing three epitopes of rLiD1, a dermonecrotic toxin from the venom of Loxosceles intermedia spider. We have analyzed the neutralization potential of sera obtained by immunization of horses with rCpLi and rCpLi combined with initial doses of venoms and compared these with antivenom traditionally produced in horses using crude Loxosceles gaucho, Loxosceles laeta and L. intermedia venoms as antigens. We have demonstrated by ELISA that horses immunized with three initial doses of crude venom containing mixtures of L. intermedia, L. gaucho and L. laeta followed by nine doses of rCpLi generate antibodies with the same reactivity as those produced following immunization with traditional antivenom, towards the venoms of the three Loxosceles sp. species. Results from in vivo and in vitro neutralization assays showed that the new horse sera are able to neutralize the dermonecrotic activity of Loxosceles venoms, which are of medical importance in Brazil and some of these sera are capable of meeting the necessary potency requirements that could allow for their therapeutic use in humans. This immunization strategy combining both antigens used approximately 67% less crude Loxosceles venoms compared to traditional immunization protocol and can mean the development of Loxosceles antivenoms with the consequent reduction of devastation of arachnid fauna. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. HDAC inhibition induces HIV-1 protein and enables immune-based clearance following latency reversal

    DEFF Research Database (Denmark)

    Wu, Guoxin; Swanson, Michael; Talla, Aarthi

    2017-01-01

    Promising therapeutic approaches for eradicating HIV include transcriptional activation of provirus from latently infected cells using latency-reversing agents (LRAs) and immune-mediated clearance to purge reservoirs. Accurate detection of cells capable of producing viral antigens and virions......, and the measurement of clearance of infected cells, is essential to assessing therapeutic efficacy. Here, we apply enhanced methodology extending the sensitivity limits for the rapid detection of subfemtomolar HIV gag p24 capsid protein in CD4+ T cells from ART-suppressed HIV+ individuals, and we show viral protein...... induction following treatment with LRAs. Importantly, we demonstrate that clinical administration of histone deacetylase inhibitors (HDACis; vorinostat and panobinostat) induced HIV gag p24, and ex vivo stimulation produced sufficient viral antigen to elicit immune-mediated cell killing using anti-gp120/CD3...

  13. Human papillomavirus type 16 E6 and NFX1-123 mislocalize immune signaling proteins and downregulate immune gene expression in keratinocytes.

    Directory of Open Access Journals (Sweden)

    Justine Levan

    Full Text Available Human papillomavirus (HPV is the most prevalent sexually transmitted infection, affecting an estimated 11% of the world's population. The high-risk HPV types (HR HPV account for approximately 5% of the global burden of cancer and thus cause high morbidity and mortality. Although it is known that persistent infection with HR HPV is the greatest risk factor for developing HPV-associated cancer, and that the HPV early proteins E6 and E7 dysregulate immune detection by its host cells, the mechanisms of immune evasion by HR HPV are not well understood. Previous work in the laboratory identified the endogenous cytoplasmic host protein NFX1-123 as a binding partner of the HR HPV type 16 oncoprotein E6 (16E6. Together NFX1-123 and 16E6 affect cellular growth, differentiation, and immortalization genes and pathways. In a whole genome microarray, human foreskin keratinocytes (HFKs stably expressing 16E6 and overexpressing NFX1-123 showed a diverse set of innate immune genes downregulated two-fold or more when compared to 16E6 cells with endogenous NFX1-123. We demonstrated that 16E6 and NFX1-123 decreased expression of pro-inflammatory cytokines and interferon-stimulated genes (ISGs in 16E6 HFKs at the mRNA and protein level. Knock down of NFX1-123 in 16E6 HFKs resulted in a derepression of innate immune genes, pointing to the requirement of NFX1-123 for immune regulation in the context of 16E6. Studies using immunofluorescent microscopy revealed that 16E6 and NFX1-123 disturbed the normal localization of signaling proteins involved in initiating the immune response. This study identifies NFX1-123 as a critical host protein partner through which 16E6 is able to subvert the immune response and in turn permit a long-lived HR HPV infection.

  14. Chlamydial Type III Secretion System Needle Protein Induces Protective Immunity against Chlamydia muridarum Intravaginal Infection

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    Ekaterina A. Koroleva

    2017-01-01

    Full Text Available Chlamydia trachomatis imposes serious health problems and causes infertility. Because of asymptomatic onset, it often escapes antibiotic treatment. Therefore, vaccines offer a better option for the prevention of unwanted inflammatory sequelae. The existence of serologically distinct serovars of C. trachomatis suggests that a vaccine will need to provide protection against multiple serovars. Chlamydia spp. use a highly conserved type III secretion system (T3SS composed of structural and effector proteins which is an essential virulence factor. In this study, we expressed the T3SS needle protein of Chlamydia muridarum, TC_0037, an ortholog of C. trachomatis CdsF, in a replication-defective adenoviral vector (AdTC_0037 and evaluated its protective efficacy in an intravaginal Chlamydia muridarum model. For better immune responses, we employed a heterologous prime-boost immunization protocol in which mice were intranasally primed with AdTC_0037 and subcutaneously boosted with recombinant TC_0037 and Toll-like receptor 4 agonist monophosphoryl lipid A mixed in a squalene nanoscale emulsion. We found that immunization with TC_0037 antigen induced specific humoral and T cell responses, decreased Chlamydia loads in the genital tract, and abrogated pathology of upper genital organs. Together, our results suggest that TC_0037, a highly conserved chlamydial T3SS protein, is a good candidate for inclusion in a Chlamydia vaccine.

  15. Autophagy Proteins in Viral Exocytosis and Anti-Viral Immune Responses

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    Christian Münz

    2017-10-01

    Full Text Available Abstract: Autophagy-related (Atg gene-encoded proteins were originally described for their crucial role in macroautophagy, a catabolic pathway for cytoplasmic constituent degradation in lysosomes. Recently it has become clear that modules of this machinery can also be used to influence endo- and exocytosis. This mini review discusses how these alternative Atg functions support virus replication and viral antigen presentation on major histocompatibility (MHC class I and II molecules. A better understanding of the modular use of the macroautophagy machinery might enable us to manipulate these alternative functions of Atg proteins during anti-viral therapies and to attenuate virus-induced immune pathologies.

  16. Naturally Acquired Human Immunity to Pneumococcus Is Dependent on Antibody to Protein Antigens.

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    Robert Wilson

    2017-01-01

    Full Text Available Naturally acquired immunity against invasive pneumococcal disease (IPD is thought to be dependent on anti-capsular antibody. However nasopharyngeal colonisation by Streptococcus pneumoniae also induces antibody to protein antigens that could be protective. We have used human intravenous immunoglobulin preparation (IVIG, representing natural IgG responses to S. pneumoniae, to identify the classes of antigens that are functionally relevant for immunity to IPD. IgG in IVIG recognised capsular antigen and multiple S. pneumoniae protein antigens, with highly conserved patterns between different geographical sources of pooled human IgG. Incubation of S. pneumoniae in IVIG resulted in IgG binding to the bacteria, formation of bacterial aggregates, and enhanced phagocytosis even for unencapsulated S. pneumoniae strains, demonstrating the capsule was unlikely to be the dominant protective antigen. IgG binding to S. pneumoniae incubated in IVIG was reduced after partial chemical or genetic removal of bacterial surface proteins, and increased against a Streptococcus mitis strain expressing the S. pneumoniae protein PspC. In contrast, depletion of type-specific capsular antibody from IVIG did not affect IgG binding, opsonophagocytosis, or protection by passive vaccination against IPD in murine models. These results demonstrate that naturally acquired protection against IPD largely depends on antibody to protein antigens rather than the capsule.

  17. Transgenic Carrot Expressing Fusion Protein Comprising M. tuberculosis Antigens Induces Immune Response in Mice

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    Natalia V. Permyakova

    2015-01-01

    Full Text Available Tuberculosis remains one of the major infectious diseases, which continues to pose a major global health problem. Transgenic plants may serve as bioreactors to produce heterologous proteins including antibodies, antigens, and hormones. In the present study, a genetic construct has been designed that comprises the Mycobacterium tuberculosis genes cfp10, esat6 and dIFN gene, which encode deltaferon, a recombinant analog of the human γ-interferon designed for expression in plant tissues. This construct was transferred to the carrot (Daucus carota L. genome by Agrobacterium-mediated transformation. This study demonstrates that the fusion protein CFP10-ESAT6-dIFN is synthesized in the transgenic carrot storage roots. The protein is able to induce both humoral and cell-mediated immune responses in laboratory animals (mice when administered either orally or by injection. It should be emphasized that M. tuberculosis antigens contained in the fusion protein have no cytotoxic effect on peripheral blood mononuclear cells.

  18. Role of DAF-21protein in Caenorhabditis elegans immunity against Proteus mirabilis infection.

    Science.gov (United States)

    JebaMercy, Gnanasekaran; Durai, Sellegounder; Prithika, Udayakumar; Marudhupandiyan, Shanmugam; Dasauni, Pushpanjali; Kundu, Suman; Balamurugan, Krishnaswamy

    2016-08-11

    Caenorhabditis elegans is emerging as one of the handy model for proteome related studies due to its simplest system biology. The present study, deals with changes in protein expression in C. elegans infected with Proteus mirabilis. Proteins were separated using two-dimensional differential gel electrophoresis (2D-DIGE) and identified using MALDI-TOF. Twelve distinctly regulated proteins identified in the infected worms, included heat shock proteins involved stress pathway (HSP-1 and HSP-6), proteins involved in immune response pathway (DAF-21), enzymes involved in normal cellular process (Eukaryotic translation Elongation Factor, actin family member, S-adenosyl homocysteine hydrolase ortholog, glutamate dehydrogenase and Vacuolar H ATPase family member) and few least characterized proteins (H28O16.1 and H08J11.2). The regulation of selected players at the transcriptional level during Proteus mirabilis infection was analyzed using qPCR. Physiological experiments revealed the ability of P. mirabilis to kill daf-21 mutant C. elegans significantly compared with the wild type. This is the first report studying proteome changes in C. elegans and exploring the involvement of MAP Kinase pathway during P. mirabilis infection. This is the first report studying proteome changes in C. elegans during P. mirabilis infection. The present study explores the role and contribution of MAP Kinase pathway and its regulator protein DAF-21 involvement in the immunity against opportunistic pathogen P. mirabilis infection. Manipulation of this DAF-21 protein in host, may pave the way for new drug development or disease control strategy during opportunistic pathogen infections. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Serum immune-related proteins are differentially expressed during hibernation in the American black bear.

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    Brian A Chow

    Full Text Available Hibernation is an adaptation to conserve energy in the face of extreme environmental conditions and low food availability that has risen in several animal phyla. This phenomenon is characterized by reduced metabolic rate (∼25% of the active basal metabolic rate in hibernating bears and energy demand, while other physiological adjustments are far from clear. The profiling of the serum proteome of the American black bear (Ursus americanus may reveal specific proteins that are differentially modulated by hibernation, and provide insight into the remarkable physiological adaptations that characterize ursid hibernation. In this study, we used differential gel electrophoresis (DIGE analysis, liquid chromatography coupled to tandem mass spectrometry, and subsequent MASCOT analysis of the mass spectra to identify candidate proteins that are differentially expressed during hibernation in captive black bears. Seventy serum proteins were identified as changing by ±1.5 fold or more, out of which 34 proteins increased expression during hibernation. The majority of identified proteins are involved in immune system processes. These included α2-macroglobulin, complement components C1s and C4, immunoglobulin μ and J chains, clusterin, haptoglobin, C4b binding protein, kininogen 1, α2-HS-glycoprotein, and apoplipoproteins A-I and A-IV. Differential expression of a subset of these proteins identified by proteomic analysis was also confirmed by immunodetection. We propose that the observed serum protein changes contribute to the maintenance of the hibernation phenotype and health, including increased capacities for bone maintenance and wound healing during hibernation in bears.

  20. Serum immune-related proteins are differentially expressed during hibernation in the American black bear.

    Science.gov (United States)

    Chow, Brian A; Donahue, Seth W; Vaughan, Michael R; McConkey, Brendan; Vijayan, Mathilakath M

    2013-01-01

    Hibernation is an adaptation to conserve energy in the face of extreme environmental conditions and low food availability that has risen in several animal phyla. This phenomenon is characterized by reduced metabolic rate (∼25% of the active basal metabolic rate in hibernating bears) and energy demand, while other physiological adjustments are far from clear. The profiling of the serum proteome of the American black bear (Ursus americanus) may reveal specific proteins that are differentially modulated by hibernation, and provide insight into the remarkable physiological adaptations that characterize ursid hibernation. In this study, we used differential gel electrophoresis (DIGE) analysis, liquid chromatography coupled to tandem mass spectrometry, and subsequent MASCOT analysis of the mass spectra to identify candidate proteins that are differentially expressed during hibernation in captive black bears. Seventy serum proteins were identified as changing by ±1.5 fold or more, out of which 34 proteins increased expression during hibernation. The majority of identified proteins are involved in immune system processes. These included α2-macroglobulin, complement components C1s and C4, immunoglobulin μ and J chains, clusterin, haptoglobin, C4b binding protein, kininogen 1, α2-HS-glycoprotein, and apoplipoproteins A-I and A-IV. Differential expression of a subset of these proteins identified by proteomic analysis was also confirmed by immunodetection. We propose that the observed serum protein changes contribute to the maintenance of the hibernation phenotype and health, including increased capacities for bone maintenance and wound healing during hibernation in bears.

  1. Nanodiamond enhances immune responses in mice against recombinant HA/H7N9 protein.

    Science.gov (United States)

    Pham, Ngoc Bich; Ho, Thuong Thi; Nguyen, Giang Thu; Le, Thuy Thi; Le, Ngoc Thu; Chang, Huan-Cheng; Pham, Minh Dinh; Conrad, Udo; Chu, Ha Hoang

    2017-10-05

    The continuing spread of the newly emerged H7N9 virus among poultry in China, as well as the possibility of human-to-human transmission, has attracted numerous efforts to develop an effective vaccine against H7N9. The use of nanoparticles in vaccinology is inspired by the fact that most pathogens have a dimension within the nano-size range and therefore can be processed efficiently by the immune system, which leads to a potent immune response. Herein, we report a facile approach to increase antigen size to achieve not only fast but also effective responses against the recombinant HA/H7N9 protein via a simple conjugation of the protein onto the surface of nanodiamond particles. In this study, trimeric Haemagglutinin (H7) that is transiently expressed in N. benthamiana was purified using affinity chromatography, and its trimeric state was revealed successfully by the cross-linking reaction. The trimeric H7 solution was subsequently mixed with a nanodiamond suspension in different ratios. The successful conjugation of the trimeric H7 onto the surface of nanodiamond particles was demonstrated by the changes in size and Zeta-potential of the particles before and after protein coating, Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and Western-blot analysis. Next, biofunction of the protein-nanodiamond conjugates was screened using a haemagglutination assay. A mixture containing 5 µg of trimeric H7 and 60 µg of nanodiamond corresponds to a ratio of 1:12 (w/w) of agglutinated chicken red blood cells at HA titer of 1024, which is 512-fold higher than the HA titer of free trimeric H7. After the 2nd and 3rd immunization in mice, ELISA and Western blot analyses demonstrated that the physical mixture of trimeric H7 protein and nanodiamond (1:12, w/w) elicited statistically significant stronger H7-specific-IgG response demonstrated by higher amounts of H7N9-specific IgG (over 15.4-fold with P < 0.05 after the second immunization). These results

  2. No immune responses by the expression of the yeast Ndi1 protein in rats.

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    Mathieu Marella

    Full Text Available BACKGROUND: The rotenone-insensitive internal NADH-quinone oxidoreductase from yeast, Ndi1, has been shown to work as a replacement molecule for complex I in the respiratory chain of mammalian mitochondria. In the so-called transkingdom gene therapy, one major concern is the fact that the yeast protein is foreign in mammals. Long term expression of Ndi1 observed in rodents with no apparent damage to the target tissue was indicative of no action by the host's immune system. METHODOLOGY/PRINCIPAL FINDINGS: In the present study, we examined rat skeletal muscles expressing Ndi1 for possible signs of inflammatory or immune response. In parallel, we carried out delivery of the GFP gene using the same viral vector that was used for the NDI1 gene. The tissues were subjected to H&E staining and immunohistochemical analyses using antibodies specific for markers, CD11b, CD3, CD4, and CD8. The data showed no detectable signs of an immune response with the tissues expressing Ndi1. In contrast, mild but distinctive positive reactions were observed in the tissues expressing GFP. This clear difference most likely comes from the difference in the location of the expressed protein. Ndi1 was localized to the mitochondria whereas GFP was in the cytosol. CONCLUSIONS/SIGNIFICANCE: We demonstrated that Ndi1 expression did not trigger any inflammatory or immune response in rats. These results push forward the Ndi1-based molecular therapy and also expand the possibility of using foreign proteins that are directed to subcellular organelle such as mitochondria.

  3. Venom allergen-like proteins in secretions of plant-parasitic nematodes activate and suppress extracellular plant immune receptors

    NARCIS (Netherlands)

    Lozano Torres, J.L.

    2014-01-01

    Parasitic worms threaten human, animal and plant health by infecting people, livestock and crops worldwide. Animals and plants share an anciently evolved innate immune system. Parasites modulate this immune system by secreting proteins to maintain their parasitic lifestyle. This thesis

  4. Venom allergen-like proteins in secretions of plant-parasitic nematodes activate and suppress extracellular plant immune receptors

    NARCIS (Netherlands)

    Lozano Torres, J.L.

    2014-01-01

      Parasitic worms threaten human, animal and plant health by infecting people, livestock and crops worldwide. Animals and plants share an anciently evolved innate immune system. Parasites modulate this immune system by secreting proteins to maintain their parasitic lifestyle. This thesis

  5. Cell-mediated immunity against human retinal extract, S-antigen, and interphotoreceptor retinoid binding protein in onchocercal chorioretinopathy

    NARCIS (Netherlands)

    van der Lelij, A.; Rothova, A.; Stilma, J. S.; Hoekzema, R.; Kijlstra, A.

    1990-01-01

    Autoimmune mechanisms are thought to be involved in the pathogenesis of onchocercal chorioretinopathy. Cell-mediated immune responses to human retinal S-antigen, interphotoreceptor retinoid binding protein (IRBP), and crude retinal extract were investigated in patients with onchocerciasis from

  6. Serum protein changes in immune and nonimmune pigeons infected with various strains of Trichomonas gallinae.

    Science.gov (United States)

    Kocan, R M; Herman, C M

    1970-01-01

    Serum protein changes were studied in immune and nonimmune pigeons infected with three different strains of Trichomonas gallinae. Strain I (nonvirulent) produced no change in the relative concentration of serum components. Strains II (oral canker) and III (Jones' Barn) produced decreases in albumin and alpha globulins, and increases in beta and gamma globulins between the 7th and 20th days post infection. Birds infected with strain II began to return to normal by the 20th day, while all those infected with strain III were dead between 10 and 14 days post infection. Two serum protein patterns resulted from infection of immune birds with the Jones' Barn strain. One showed no change in relative protein concentrations and no tissue invasion by the parasite while the other was similar to that seen in nonimmune birds infected with a strain producing oral canker. These also showed evidence of tissue invasion by the parasite. It was concluded that tissue invasion was necessary to evoke a quantitative change in serum protein concentrations.

  7. Immune response to dna vaccine expressing transferrin binding protein a gene of Pasteurella multocida

    Directory of Open Access Journals (Sweden)

    Satparkash Singh

    2011-06-01

    Full Text Available Haemorrhagic Septicaemia (HS, an acute and fatal disease of cattle and buffalo is primarily caused by serotype B:2 or E:2 of Pasteurella multocida. The transferrin binding protein A (TbpA has been found to act as immunogen and potent vaccine candidate in various Gram negative bacteria including P. multocida. The present study was carried out to evaluate the potential of this antigen as a DNA vaccine against HS in mice model. The tbpA gene of P. multocida serotype B:2 was cloned in a mammalian expression vector alone and along with murine IL2 gene as immunological adjuvant to produce monocistronic and bicistronic DNA vaccine constructs, respectively. The immune response to DNA vaccines was evaluated based on serum antibody titres and lymphocyte proliferation assay. A significant increase in humoral and cell mediated immune responses was observed in mice vaccinated with DNA vaccines as compared to non immunized group. Additionally, the bicistronic DNA vaccine provided superior immune response and protection level following challenge as compared to monocistronic construct. The study revealed that DNA vaccine presents a promising approach for the prevention of HS.

  8. Immunization of stromal cell targeting fibroblast activation protein providing immunotherapy to breast cancer mouse model.

    Science.gov (United States)

    Meng, Mingyao; Wang, Wenju; Yan, Jun; Tan, Jing; Liao, Liwei; Shi, Jianlin; Wei, Chuanyu; Xie, Yanhua; Jin, Xingfang; Yang, Li; Jin, Qing; Zhu, Huirong; Tan, Weiwei; Yang, Fang; Hou, Zongliu

    2016-08-01

    Unlike heterogeneous tumor cells, cancer-associated fibroblasts (CAF) are genetically more stable which serve as a reliable target for tumor immunotherapy. Fibroblast activation protein (FAP) which is restrictively expressed in tumor cells and CAF in vivo and plays a prominent role in tumor initiation, progression, and metastasis can function as a tumor rejection antigen. In the current study, we have constructed artificial FAP(+) stromal cells which mimicked the FAP(+) CAF in vivo. We immunized a breast cancer mouse model with FAP(+) stromal cells to perform immunotherapy against FAP(+) cells in the tumor microenvironment. By forced expression of FAP, we have obtained FAP(+) stromal cells whose phenotype was CD11b(+)/CD34(+)/Sca-1(+)/FSP-1(+)/MHC class I(+). Interestingly, proliferation capacity of the fibroblasts was significantly enhanced by FAP. In the breast cancer-bearing mouse model, vaccination with FAP(+) stromal cells has significantly inhibited the growth of allograft tumor and reduced lung metastasis indeed. Depletion of T cell assays has suggested that both CD4(+) and CD8(+) T cells were involved in the tumor cytotoxic immune response. Furthermore, tumor tissue from FAP-immunized mice revealed that targeting FAP(+) CAF has induced apoptosis and decreased collagen type I and CD31 expression in the tumor microenvironment. These results implicated that immunization with FAP(+) stromal cells led to the disruption of the tumor microenvironment. Our study may provide a novel strategy for immunotherapy of a broad range of cancer.

  9. Hiding in plain sight: immune evasion by the staphylococcal protein SdrE.

    Science.gov (United States)

    Herr, Andrew B; Thorman, Alexander W

    2017-05-10

    The human immune system is responsible for identification and destruction of invader cells, such as the bacterial pathogen Staphylococcus aureus In response, S. aureus brings to the fight a large number of virulence factors, including several that allow it to evade the host immune response. The staphylococcal surface protein SdrE was recently reported to bind to complement Factor H, an important regulator of complement activation. Factor H attaches to the surface of host cells to inhibit complement activation and amplification, preventing the destruction of the host cell. SdrE binding to Factor H allows S. aureus to mimic a host cell and reduces bacterial killing by granulocytes. In a new study published in Biochemical Journal , Zhang et al. describe crystal structures of SdrE and its complex with the C-terminal portion of Factor H. The structure of SdrE and its interaction with the Factor H peptide closely resemble a family of surface proteins that recognize extracellular matrix components such as fibrinogen. However, unbound SdrE forms a novel 'Closed' conformation with an occluded peptide-binding groove. These structures reveal a fascinating mechanism for immune evasion and provide a potential avenue for the development of novel antimicrobial agents to target SdrE. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

  10. Analysis of Thioester-Containing Proteins during the Innate Immune Response of Drosophila melanogaster

    Science.gov (United States)

    Bou Aoun, Richard; Hetru, Charles; Troxler, Laurent; Doucet, Daniel; Ferrandon, Dominique; Matt, Nicolas

    2010-01-01

    Thioester-containing proteins (TEPs) are conserved proteins among insects that are thought to be involved in innate immunity. In Drosophila, the Tep family is composed of 6 genes named Tep1–Tep6. In this study, we investigated the phylogeny, expression pattern and roles of these genes in the host defense of Drosophila. Protostomian Tep genes are clustered in 3 distinct branches, 1 of which is specific to mosquitoes. Most D. melanogaster Tep genes are expressed in hemocytes, can be induced in the fat body, and are expressed in specific regions of the hypodermis. This expression pattern is consistent with a role in innate immunity. However, we find that TEP1, TEP2, and TEP4 are not strictly required in the body cavity to fight several bacterial and fungal infections. One possibility is that Drosophila TEPs act redundantly or that their absence can be compensated by other components of the immune response. TEPs may thus provide a subtle selective advantage during evolution. Alternatively, they may be required in host defense against specific as yet unidentified natural pathogens of Drosophila. PMID:21063077

  11. Isolation and Purification of an Antibacterial Protein from Immune Induced Haemolymph of American Cockroach, Periplaneta americana

    Directory of Open Access Journals (Sweden)

    Hamid Reza Basseri

    2016-10-01

    Full Text Available Background: Antimicrobial peptides play a role as effectors substances in the immunity of vertebrate and inverte­brate hosts. In the current study, antimicrobial peptide was isolated from the haemolymph of the American cock­roach, Periplaneta americana.Methods: Micrococcus luteus as Gram-positive bacteria and Escherichia coli as Gram-negative bacteria were candi­date for injection. Induction was done by injecting both bacteria into the abdominal cavity of two groups of cock­roaches separately. The haemolymphs were collected 24 hours after post injection and initially tested against both bacteria. Subsequently, the immune induced haemolymph was purified by high performance liquid chromatography (HPLC to separate the proteins responsible for the antibacterial activity.Results: The non-induced haemolymph did not show any activity against both bacteria whereas induced haemo­lymph exhibited high activity against M. luteus but did less against E. coli. Two fractions showed antibacterial activ­ity against M. luteus. Finally the molecular weight of the isolated antibacterial proteins were determined as 72 kDa and 62 kDa using SDS-PAGE.Conclusion: Induced haemolymph of American cockroaches has the ability to produce peptides to combat against Gram-positive bacteria when an immune challenge is mounted. Further work has to be done to sequence of the pro­tein, which it would be advantageous.

  12. Co-expression of Interleukin-15 Enhances the Protective Immune Responses Induced by Immunization with a Murine Malaria MVA-Based Vaccine Encoding the Circumsporozoite Protein.

    Science.gov (United States)

    Parra, Marcela; Liu, Xia; Derrick, Steven C; Yang, Amy; Molina-Cruz, Alvaro; Barillas-Mury, Carolina; Zheng, Hong; Thao Pham, Phuong; Sedegah, Martha; Belmonte, Arnel; Litilit, Dianne D; Waldmann, Thomas A; Kumar, Sanjai; Morris, Sheldon L; Perera, Liyanage P

    2015-01-01

    Malaria remains a major global public health problem with an estimated 200 million cases detected in 2012. Although the most advanced candidate malaria vaccine (RTS,S) has shown promise in clinical trials, its modest efficacy and durability have created uncertainty about the impact of RTS,S immunization (when used alone) on global malaria transmission. Here we describe the development and characterization of a novel modified vaccinia virus Ankara (MVA)-based malaria vaccine which co-expresses the Plasmodium yoelii circumsporozoite protein (CSP) and IL-15. Vaccination/challenge studies showed that C57BL/6 mice immunized with the MVA-CSP/IL15 vaccine were protected significantly better against a P. yoelii 17XNL sporozoite challenge than either mice immunized with an MVA vaccine expressing only CSP or naïve controls. Importantly, the levels of total anti-CSP IgG were elevated about 100-fold for the MVA-CSP/IL15 immunized group compared to mice immunized with the MVA-CSP construct that does not express IL-15. Among the IgG subtypes, the IL-15 expressing MVA-CSP vaccine induced levels of IgG1 (8 fold) and IgG2b (80 fold) higher than the MVA-CSP construct. The significantly enhanced humoral responses and protection detected after immunization with the MVA-CSP/IL15 vaccine suggest that this IL-15 expressing MVA construct could be considered in the development of future malaria immunization strategies.

  13. Human metapneumovirus M2-2 protein inhibits innate immune response in monocyte-derived dendritic cells.

    Directory of Open Access Journals (Sweden)

    Junping Ren

    Full Text Available Human metapneumovirus (hMPV is a leading cause of lower respiratory infection in young children, the elderly and immunocompromised patients. Repeated hMPV infections occur throughout life. However, immune evasion mechanisms of hMPV infection are largely unknown. Recently, our group has demonstrated that hMPV M2-2 protein, an important virulence factor, contributes to immune evasion in airway epithelial cells by targeting the mitochondrial antiviral-signaling protein (MAVS. Whether M2-2 regulates the innate immunity in human dendritic cells (DC, an important family of immune cells controlling antigen presenting, is currently unknown. We found that human DC infected with a virus lacking M2-2 protein expression (rhMPV-ΔM2-2 produced higher levels of cytokines, chemokines and IFNs, compared to cells infected with wild-type virus (rhMPV-WT, suggesting that M2-2 protein inhibits innate immunity in human DC. In parallel, we found that myeloid differentiation primary response gene 88 (MyD88, an essential adaptor for Toll-like receptors (TLRs, plays a critical role in inducing immune response of human DC, as downregulation of MyD88 by siRNA blocked the induction of immune regulatory molecules by hMPV. Since M2-2 is a cytoplasmic protein, we investigated whether M2-2 interferes with MyD88-mediated antiviral signaling. We found that indeed M2-2 protein associated with MyD88 and inhibited MyD88-dependent gene transcription. In this study, we also identified the domains of M2-2 responsible for its immune inhibitory function in human DC. In summary, our results demonstrate that M2-2 contributes to hMPV immune evasion by inhibiting MyD88-dependent cellular responses in human DC.

  14. Antimicrobial activities of the bacteriocin-like substances produced ...

    African Journals Online (AJOL)

    A total of 450 different colonies, isolated from 25 samples of dromedary milk collected from Laâyoune region of Morocco, were tested for antimicrobial compounds production. Out of these, 30 were determined to be lactic acid bacteria (LAB) and able to inhibit the growth of the indicator strain Listeria innocua CECT 4030.

  15. Antimicrobial activities of the bacteriocin-like substances produced ...

    African Journals Online (AJOL)

    Administrator

    2011-09-07

    Sep 7, 2011 ... activity from hydrogen peroxide was blocked by the addition of a sterile solution of catalase (2619 U/mg. sigma), dissolved in phosphate buffer at pH 7 and 1 .... arginine and esculin, the production of acetoin and CO2 and for growth at 40°C in the presence of 4% NaCl. The characteristics of strain R112 are ...

  16. Antimicrobial activities of the bacteriocin-like substances produced ...

    African Journals Online (AJOL)

    Administrator

    2011-09-07

    Sep 7, 2011 ... A total of 450 different colonies, isolated from 25 samples of dromedary milk collected from Laâyoune region of Morocco, were tested for antimicrobial compounds production. Out of these, 30 were determined to be lactic acid bacteria (LAB) and able to inhibit the growth of the indicator strain Listeria innocua ...

  17. FAM26F: An Enigmatic Protein Having a Complex Role in the Immune System.

    Science.gov (United States)

    Malik, Uzma; Javed, Aneela

    2016-09-19

    Mammalian immune system is a complex amalgam of diverse cellular and noncellular components such as cytokines, receptors and co-receptors. FAM26F (family with sequence similarity 26, member F) is a recently identified tetraspanin-like membrane glycoprotein which is predicted to make homophilic interactions and potential synapses between several immune cells including CD4 + , CD8 + , NK, dendritic cells and macrophages. Various whole transcriptome analyses have demonstrated the differential expression of FAM26F in several bacterial, viral and parasitic infections, in certain pathophysiological conditions such as liver and heart transplantation, and in various cancers. The complete understanding of transcriptional regulation of FAM26F is in its infancy however it is up regulated by various stimulants such as polyI:C, LPS, INF gamma and TNF alpha, and via various proposed pathways including TLR3, TLR4 IFN-β and Dectin-1. These pathways can merge in STAT1 activation. The synergistic expression of FAM26F on both NK-cells and myeloid dendritic cells is required to activate NK-cells against tumors via its cytoplasmic tail, thus emphasizing therapeutic potential of FAM26F for NK sensitive tumors. Current review provides a comprehensive basis to propose that FAM26F expression level is at least a hallmark for IFN-γ-lead immune responses and thus can proficiently be regarded as an early diagnostic marker. Future investigation dissecting the role of FAM26F in activation of various immune cell populations in local amplification by cell-cell contact is crucial to provide the missing link imperative for elucidating the relevance of this protein in immune responses.

  18. The tumor necrosis factor-alpha-induced protein 8 family in immune homeostasis and inflammatory cancer diseases.

    Science.gov (United States)

    Luan, Y Y; Yao, Y M; Sheng, Z Y

    2013-01-01

    Within the immune system homeostasis is maintained by a myriad of mechanisms that include the regulation of immune cell activation and programmed cell death. The breakdown of immune homeostasis may lead to fatal inflammatory diseases. We set out to identify genes of tumor necrosis factor-alpha-induced protein 8 (TNFAIP8) family that has a functional role in the process of immune homeostasis. Tumor necrosis factor-alpha-induced protein 8 (TNFAIP8), which functions as an oncogenic molecule, is also associated with enhanced cell survival and inhibition of apoptosis. Tumor necrosis factor-alpha-induced protein 8-like 2 (TIPE2) governs immune homeostasis in both the innate and adaptive immune system and prevents hyper-responsiveness by negatively regulating signaling via T cell receptors and Toll-like receptors (TLRs). There also exist two highly homologous but uncharacterized proteins, TIPE1 and TIPE3. This review is an attempt to provide a summary of TNFAIP8 family associated with immune homeostasis and inflammatory cancer diseases.

  19. Immunization of Mice with Recombinant Brucella abortus Organic Hydroperoxide Resistance (Ohr) Protein Protects Against a Virulent Brucella abortus 544 Infection.

    Science.gov (United States)

    Hop, Huynh Tan; Reyes, Alisha Wehdnesday Bernardo; Simborio, Hannah Leah Tadeja; Arayan, Lauren Togonon; Min, Won Gi; Lee, Hu Jang; Lee, Jin Ju; Chang, Hong Hee; Kim, Suk

    2016-01-01

    In this study, the Brucella abortus ohr gene coding for an organic hydroperoxide resistance protein (Ohr) was cloned into a maltose fusion protein expression system (pMAL), inserted into Escherichia coli, and purified, and its immunogenicity was evaluated by western blot analysis using Brucella-positive mouse sera. The purified recombinant Ohr (rOhr) was treated with adjuvant and injected intraperitoneally into BALB/c mice. A protective immune response analysis revealed that rOhr induced a significant increase in both the IgG1 and IgG2a titers, and IgG2a reached a higher level than IgG1 after the second and third immunizations. Additionally, immunization with rOhr induced high production of IFN-γ as well as proinflammatory cytokines such as TNF, MCP-1, IL-12p70, and IL-6, but a lesser amount of IL-10, suggesting that rOhr predominantly elicited a cell-mediated immune response. In addition, immunization with rOhr caused a significantly higher degree of protection against a virulent B. abortus infection compared with a positive control group consisting of mice immunized with maltose-binding protein. These findings showed that B. abortus rOhr was able to induce both humoral and cell-mediated immunity in mice, which suggested that this recombinant protein could be a potential vaccine candidate for animal brucellosis.

  20. Rotavirus nonstructural protein 1 antagonizes innate immune response by interacting with retinoic acid inducible gene I

    Directory of Open Access Journals (Sweden)

    Qin Lan

    2011-12-01

    Full Text Available Abstract Background The nonstructural protein 1 (NSP1 of rotavirus has been reported to block interferon (IFN signaling by mediating proteasome-dependent degradation of IFN-regulatory factors (IRFs and (or the β-transducin repeat containing protein (β-TrCP. However, in addition to these targets, NSP1 may subvert innate immune responses via other mechanisms. Results The NSP1 of rotavirus OSU strain as well as the IRF3 binding domain truncated NSP1 of rotavirus SA11 strain are unable to degrade IRFs, but can still inhibit host IFN response, indicating that NSP1 may target alternative host factor(s other than IRFs. Overexpression of NSP1 can block IFN-β promoter activation induced by the retinoic acid inducible gene I (RIG-I, but does not inhibit IFN-β activation induced by the mitochondrial antiviral-signaling protein (MAVS, indicating that NSP1 may target RIG-I. Immunoprecipitation experiments show that NSP1 interacts with RIG-I independent of IRF3 binding domain. In addition, NSP1 induces down-regulation of RIG-I in a proteasome-independent way. Conclusions Our findings demonstrate that inhibition of RIG-I mediated type I IFN responses by NSP1 may contribute to the immune evasion of rotavirus.

  1. Male homosexuality and maternal immune responsivity to the Y-linked protein NLGN4Y.

    Science.gov (United States)

    Bogaert, Anthony F; Skorska, Malvina N; Wang, Chao; Gabrie, José; MacNeil, Adam J; Hoffarth, Mark R; VanderLaan, Doug P; Zucker, Kenneth J; Blanchard, Ray

    2018-01-09

    We conducted a direct test of an immunological explanation of the finding that gay men have a greater number of older brothers than do heterosexual men. This explanation posits that some mothers develop antibodies against a Y-linked protein important in male brain development, and that this effect becomes increasingly likely with each male gestation, altering brain structures underlying sexual orientation in their later-born sons. Immune assays targeting two Y-linked proteins important in brain development-protocadherin 11 Y-linked (PCDH11Y) and neuroligin 4 Y-linked (NLGN4Y; isoforms 1 and 2)-were developed. Plasma from mothers of sons, about half of whom had a gay son, along with additional controls (women with no sons, men) was analyzed for male protein-specific antibodies. Results indicated women had significantly higher anti-NLGN4Y levels than men. In addition, after statistically controlling for number of pregnancies, mothers of gay sons, particularly those with older brothers, had significantly higher anti-NLGN4Y levels than did the control samples of women, including mothers of heterosexual sons. The results suggest an association between a maternal immune response to NLGN4Y and subsequent sexual orientation in male offspring. Copyright © 2018 the Author(s). Published by PNAS.

  2. Unfolded protein response (UPR) signaling regulates arsenic trioxide-mediated macrophage innate immune function disruption

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, Ritesh K.; Li, Changzhao; Chaudhary, Sandeep C. [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Ballestas, Mary E. [Department of Pediatrics Infectious Disease, Children' s of Alabama, School of Medicine, University of Alabama at Birmingham, AL (United States); Elmets, Craig A. [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Robbins, David J. [Department of Surgery, Molecular Oncology Program, Miller School of Medicine, University of Miami, Miami (United States); Matalon, Sadis [Department of Anesthesiology, University of Alabama at Birmingham, Birmingham, AL (United States); Deshane, Jessy S. [Department of Medicine, Division of Pulmonary, Allergy and Critical Care Medicine, University of Alabama at Birmingham, Birmingham, AL (United States); Afaq, Farrukh [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Bickers, David R. [Department of Dermatology, Columbia University Medical Center, New York (United States); Athar, Mohammad, E-mail: mathar@uab.edu [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States)

    2013-11-01

    Arsenic exposure is known to disrupt innate immune functions in humans and in experimental animals. In this study, we provide a mechanism by which arsenic trioxide (ATO) disrupts macrophage functions. ATO treatment of murine macrophage cells diminished internalization of FITC-labeled latex beads, impaired clearance of phagocytosed fluorescent bacteria and reduced secretion of pro-inflammatory cytokines. These impairments in macrophage functions are associated with ATO-induced unfolded protein response (UPR) signaling pathway characterized by the enhancement in proteins such as GRP78, p-PERK, p-eIF2α, ATF4 and CHOP. The expression of these proteins is altered both at transcriptional and translational levels. Pretreatment with chemical chaperon, 4-phenylbutyric acid (PBA) attenuated the ATO-induced activation in UPR signaling and afforded protection against ATO-induced disruption of macrophage functions. This treatment also reduced ATO-mediated reactive oxygen species (ROS) generation. Interestingly, treatment with antioxidant N-acetylcysteine (NAC) prior to ATO exposure, not only reduced ROS production and UPR signaling but also improved macrophage functions. These data demonstrate that UPR signaling and ROS generation are interdependent and are involved in the arsenic-induced pathobiology of macrophage. These data also provide a novel strategy to block the ATO-dependent impairment in innate immune responses. - Highlights: • Inorganic arsenic to humans and experimental animals disrupt innate immune responses. • The mechanism underlying arsenic impaired macrophage functions involves UPR signaling. • Chemical chaperon attenuates arsenic-mediated macrophage function impairment. • Antioxidant, NAC blocks impairment in arsenic-treated macrophage functions.

  3. Complex structure of type VI peptidoglycan muramidase effector and a cognate immunity protein

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Tianyu [Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Ding, Jinjing; Zhang, Ying; Wang, Da-Cheng, E-mail: dcwang@ibp.ac.cn [Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101 (China); Liu, Wei, E-mail: dcwang@ibp.ac.cn [The Third Military Medical University, Chongqing 400038 (China); Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101 (China)

    2013-10-01

    The structure of the Tse3–Tsi3 complex associated with the bacterial type VI secretion system of P. aeruginosa has been solved and refined at 1.9 Å resolution. The structural basis of the recognition of the muramidase effector and its inactivation by its cognate immunity protein is revealed. The type VI secretion system (T6SS) is a bacterial protein-export machine that is capable of delivering virulence effectors between Gram-negative bacteria. The T6SS of Pseudomonas aeruginosa transports two lytic enzymes, Tse1 and Tse3, to degrade cell-wall peptidoglycan in the periplasm of rival bacteria that are competing for niches via amidase and muramidase activities, respectively. Two cognate immunity proteins, Tsi1 and Tsi3, are produced by the bacterium to inactivate the two antibacterial effectors, thereby protecting its siblings from self-intoxication. Recently, Tse1–Tsi1 has been structurally characterized. Here, the structure of the Tse3–Tsi3 complex is reported at 1.9 Å resolution. The results reveal that Tse3 contains a C-terminal catalytic domain that adopts a soluble lytic transglycosylase (SLT) fold in which three calcium-binding sites were surprisingly observed close to the catalytic Glu residue. The electrostatic properties of the substrate-binding groove are also distinctive from those of known structures with a similar fold. All of these features imply that a unique catalytic mechanism is utilized by Tse3 in cleaving glycosidic bonds. Tsi3 comprises a single domain showing a β-sandwich architecture that is reminiscent of the immunoglobulin fold. Three loops of Tsi3 insert deeply into the groove of Tse3 and completely occlude its active site, which forms the structural basis of Tse3 inactivation. This work is the first crystallographic report describing the three-dimensional structure of the Tse3–Tsi3 effector–immunity pair.

  4. Unfolded protein response (UPR) signaling regulates arsenic trioxide-mediated macrophage innate immune function disruption

    International Nuclear Information System (INIS)

    Srivastava, Ritesh K.; Li, Changzhao; Chaudhary, Sandeep C.; Ballestas, Mary E.; Elmets, Craig A.; Robbins, David J.; Matalon, Sadis; Deshane, Jessy S.; Afaq, Farrukh; Bickers, David R.; Athar, Mohammad

    2013-01-01

    Arsenic exposure is known to disrupt innate immune functions in humans and in experimental animals. In this study, we provide a mechanism by which arsenic trioxide (ATO) disrupts macrophage functions. ATO treatment of murine macrophage cells diminished internalization of FITC-labeled latex beads, impaired clearance of phagocytosed fluorescent bacteria and reduced secretion of pro-inflammatory cytokines. These impairments in macrophage functions are associated with ATO-induced unfolded protein response (UPR) signaling pathway characterized by the enhancement in proteins such as GRP78, p-PERK, p-eIF2α, ATF4 and CHOP. The expression of these proteins is altered both at transcriptional and translational levels. Pretreatment with chemical chaperon, 4-phenylbutyric acid (PBA) attenuated the ATO-induced activation in UPR signaling and afforded protection against ATO-induced disruption of macrophage functions. This treatment also reduced ATO-mediated reactive oxygen species (ROS) generation. Interestingly, treatment with antioxidant N-acetylcysteine (NAC) prior to ATO exposure, not only reduced ROS production and UPR signaling but also improved macrophage functions. These data demonstrate that UPR signaling and ROS generation are interdependent and are involved in the arsenic-induced pathobiology of macrophage. These data also provide a novel strategy to block the ATO-dependent impairment in innate immune responses. - Highlights: • Inorganic arsenic to humans and experimental animals disrupt innate immune responses. • The mechanism underlying arsenic impaired macrophage functions involves UPR signaling. • Chemical chaperon attenuates arsenic-mediated macrophage function impairment. • Antioxidant, NAC blocks impairment in arsenic-treated macrophage functions

  5. Conserved hypothetical protein Rv1977 in Mycobacterium tuberculosis strains contains sequence polymorphisms and might be involved in ongoing immune evasion.

    Science.gov (United States)

    Jiang, Yi; Liu, Haican; Wang, Xuezhi; Li, Guilian; Qiu, Yan; Dou, Xiangfeng; Wan, Kanglin

    2015-01-01

    Host immune pressure and associated parasite immune evasion are key features of host-pathogen co-evolution. A previous study showed that human T cell epitopes of Mycobacterium tuberculosis are evolutionarily hyperconserved and thus it was deduced that M. tuberculosis lacks antigenic variation and immune evasion. Here, we selected 151 clinical Mycobacterium tuberculosis isolates from China, amplified gene encoding Rv1977 and compared the sequences. The results showed that Rv1977, a conserved hypothetical protein, is not conserved in M. tuberculosis strains and there are polymorphisms existed in the protein. Some mutations, especially one frameshift mutation, occurred in the antigen Rv1977, which is uncommon in M.tb strains and may lead to the protein function altering. Mutations and deletion in the gene all affect one of three T cell epitopes and the changed T cell epitope contained more than one variable position, which may suggest ongoing immune evasion.

  6. Extraordinary Diversity of Immune Response Proteins among Sea Urchins: Nickel-Isolated Sp185/333 Proteins Show Broad Variations in Size and Charge

    Science.gov (United States)

    Sherman, Lauren S.; Schrankel, Catherine S.; Brown, Kristy J.; Smith, L. Courtney

    2015-01-01

    Effective protection against pathogens requires the host to produce a wide range of immune effector proteins. The Sp185/333 gene family, which is expressed by the California purple sea urchin Strongylocentrotus purpuratus in response to bacterial infection, encodes a highly diverse repertoire of anti-pathogen proteins. A subset of these proteins can be isolated by affinity to metal ions based on multiple histidines, resulting in one to four bands of unique molecular weight on standard Western blots, which vary depending on the individual sea urchin. Two dimensional gel electrophoresis (2DE) of nickel-isolated protein samples followed by Western blot was employed to detect nickel-isolated Sp185/333 (Ni-Sp185/333) proteins and to evaluate protein diversity in animals before and after immune challenge with marine bacteria. Ni-Sp185/333 proteins of the same molecular weight on standard Western blots appear as a broad complex of variants that differ in pI on 2DE Western blots. The Ni-Sp185/333 protein repertoire is variable among animals, and shows a variety of changes among individual sea urchins in response to immune challenges with both the same and different species of bacteria. The extraordinary diversity of the Ni-Sp185/333 proteins may provide significant anti-pathogen capabilities for sea urchins that survive solely on innate immunity. PMID:26406912

  7. A cell wall protein-based vaccine candidate induce protective immune response against Sporothrix schenckii infection.

    Science.gov (United States)

    Portuondo, Deivys Leandro; Batista-Duharte, Alexander; Ferreira, Lucas Souza; Martínez, Damiana Téllez; Polesi, Marisa Campos; Duarte, Roberta Aparecida; de Paula E Silva, Ana Carolina Alves; Marcos, Caroline Maria; Almeida, Ana Marisa Fusco de; Carlos, Iracilda Zeppone

    2016-02-01

    Sporotrichosis is a subcutaneous mycosis caused by several closely related thermo-dimorphic fungi of the Sporothrix schenckii species complex, affecting humans and other mammals. In the last few years, new strategies have been proposed for controlling sporotrichosis owning to concerns about its growing incidence in humans, cats, and dogs in Brazil, as well as the toxicity and limited efficacy of conventional antifungal drugs. In this study, we assessed the immunogenicity and protective properties of two aluminum hydroxide (AH)-adsorbed S. schenckii cell wall protein (ssCWP)-based vaccine formulations in a mouse model of systemic S. schenckii infection. Fractioning by SDS-PAGE revealed nine protein bands, two of which were functionally characterized: a 44kDa peptide hydrolase and a 47kDa enolase, which was predicted to be an adhesin. Sera from immunized mice recognized the 47kDa enolase and another unidentified 71kDa protein, whereas serum from S. schenckii-infected mice recognized both these proteins plus another unidentified 9.4kDa protein. Furthermore, opsonization with the anti-ssCWP sera led to markedly increased phagocytosis and was able to strongly inhibit the fungus' adhesion to fibroblasts. Immunization with the higher-dose AH-adjuvanted formulation led to increased ex vivo release of IL-12, IFN-γ, IL-4, and IL-17, whereas only IL-12 and IFN-γ were induced by the higher-dose non-adjuvanted formulation. Lastly, passive transference of the higher-dose AH-adjuvanted formulation's anti-ssCWP serum was able to afford in vivo protection in a subsequent challenge with S. schenckii, becoming a viable vaccine candidate for further testing. Copyright © 2015 Elsevier GmbH. All rights reserved.

  8. Purification and characterization of the colicin A immunity protein in detergent micelles.

    Science.gov (United States)

    Metola, Ane; Bouchet, Ana M; Alonso-Mariño, Marian; Diercks, Tammo; Mäler, Lena; Goñi, Félix M; Viguera, Ana R

    2017-11-01

    The immunity proteins against pore-forming colicins represent a family of integral membrane proteins that reside in the inner membrane of producing cells. Cai, the colicin A immunity protein, was characterized here in detergent micelles by circular dichroism (CD), size exclusion chromatography, chemical cross-linking, nuclear magnetic resonance (NMR) spectroscopy, cysteine accessibility, and colicin A binding in detergent micelles. Bile-salt derivatives induced extensive protein polymerization that precluded further investigation. The physical characterization of detergent-solubilized protein indicates that phosphate-containing detergents are more efficient in extracting, solubilizing and maintaining Cai in a monomeric state. Yet, their capacity to ensure protein activity, reconstitution, helix packing, and high-quality NMR spectra was inferior to that of milder detergents. Solvent ionic strength and composition greatly modified the solubilizing capacity of milder detergents. Most importantly, binding to the colicin A pore-forming domain (pf-ColA) occurred almost exclusively in sugar-derived detergents. The relative performance of the different detergents in each experiment depends on their impact not only on Cai structure, solubility and oligomerization state, but also on other reaction components and technical aspects. Thus, proteoliposomes were best obtained from protein in LDAO micelles, possibly also due to indirect effects on the lipidic bilayer. The compatibility of a detergent with Cai/pf-ColA complex formation is influenced by its effect on the conformational landscape of each protein, where detergent-mediated pf-ColA denaturation could also lead to negative results. The NMR spectra were greatly affected by the solubility, monodispersity, fold and dynamics of the protein-detergent complexes, and none of those tested here provided NMR spectra of sufficient quality to allow for peak assignment. Cai function could be proven in alkyl glycosides and not in

  9. Cross-species Virus-host Protein-Protein Interactions Inhibiting Innate Immunity

    Science.gov (United States)

    2016-07-01

    joule (J) kiloton (kt) (TNT equivalent) 4.184 × 1012 joule (J) British thermal unit (Btu) (thermochemical) 1.054 350 × 10 3 joule (J) foot-pound...Orthomyxoviridae influenza A has an extensively tracked history of host species jumps, including birds, pigs and other mammals , and humans... biology , and protein chemistry, and he presented his research studies at the HWI-UB-RPCI Summer Undergraduate Biomedical Research Day (August 1, 2014

  10. Fibroblast activation protein is dispensable in the anti-influenza immune response in mice.

    Directory of Open Access Journals (Sweden)

    Sioh-Yang Tan

    Full Text Available Fibroblast activation protein alpha (FAP is a unique dual peptidase of the S9B serine protease family, being capable of both dipeptidyl peptidase and endopeptidase activities. FAP is expressed at low level in healthy adult organs including the pancreas, cervix, uterus, submaxillary gland and the skin, and highly upregulated in embryogenesis, chronic inflammation and tissue remodelling. It is also expressed by cancer-associated stromal fibroblasts in more than 90% of epithelial tumours. FAP has enzymatic and non-enzymatic functions in the growth, immunosuppression, invasion and cell signalling of tumour cells. FAP deficient mice are fertile and viable with no gross abnormality, but little data exist on the role of FAP in the immune system. FAP is upregulated in association with microbial stimulation and chronic inflammation, but its function in infection remains unknown. We showed that major populations of immune cells including CD4+ and CD8+ T cells, B cells, dendritic cells and neutrophils are generated and maintained normally in FAP knockout mice. Upon intranasal challenge with influenza virus, FAP mRNA was increased in the lungs and lung-draining lymph nodes. Nonetheless, FAP deficient mice showed similar pathologic kinetics to wildtype controls, and were capable of supporting normal anti-influenza T and B cell responses. There was no evidence of compensatory upregulation of other DPP4 family members in influenza-infected FAP-deficient mice. FAP appears to be dispensable in anti-influenza adaptive immunity.

  11. Isolation and partial purification of antimicrobial peptides/proteins from dung beetle, Onthophagus taurus immune hemolymph

    International Nuclear Information System (INIS)

    Vasanth Patil, H.B.; Sathish Kumar, B.Y.

    2012-01-01

    Antimicrobial peptides are important in the first line of the host defense system of all insect species. In the present study antimicrobial peptide(s) were isolated from the hemolymph of the dung beetle Onthophagus taurus. Both non induced and immune induced hemolymphs were tested for their antimicrobial activity against different bacterial strains and C. albicans. Induction was done by injecting E. coli into the abdominal cavity of the O. taurus. The non induced hemolymph did not show activity against any of the tested fungal and bacterial strains where as induced hemolymph showed activity against all tested bacterial strains but no activity against C. albicans. The induced hemolymph was subjected to non reducing SDS-PAGE and UV wavelength scan was performed to detect the presence of peptides. The immune induced hemolymph was purified by gel filtration chromatography to separate the proteins responsible for the antibacterial activity. The fractions within the peak were tested against those bacteria which previously showed sensitivity to the crude immune induced hemolymph. All fractions were found to be active against all tested bacteria with difference in zone of inhibition. The peptides are active against prokaryotes and not against eukaryotes. These properties reveal its unique characteristics and therapeutic application. (author)

  12. FINE SPECIFICITY OF CELLULAR IMMUNE-RESPONSES IN HUMANS TO HUMAN CYTOMEGALOVIRUS IMMEDIATE-EARLY 1-PROTEIN

    NARCIS (Netherlands)

    ALP, NJ; ALLPORT, TD; VANZANTEN, J; RODGERS, B; SISSONS, JGP; BORYSIEWICZ, LK

    Cell-mediated immunity is important in maintaining the virus-host equilibrium in persistent human cytomegalovirus (HCMV) infection. The HCMV 72-kDa major immediate early 1 protein (IE1) is a target for CD8+ cytotoxic T cells in humans, as is the equivalent 89-kDa protein in mouse. Less is known

  13. Protective Immunity and Reduced Renal Colonization Induced by Vaccines Containing Recombinant Leptospira interrogans Outer Membrane Proteins and Flagellin Adjuvant

    Science.gov (United States)

    Monaris, D.; Sbrogio-Almeida, M. E.; Dib, C. C.; Canhamero, T. A.; Souza, G. O.; Vasconcellos, S. A.; Ferreira, L. C. S.

    2015-01-01

    Leptospirosis is a global zoonotic disease caused by different Leptospira species, such as Leptospira interrogans, that colonize the renal tubules of wild and domestic animals. Thus far, attempts to develop effective leptospirosis vaccines, both for humans and animals, have failed to induce immune responses capable of conferring protection and simultaneously preventing renal colonization. In this study, we evaluated the protective immunity induced by subunit vaccines containing seven different recombinant Leptospira interrogans outer membrane proteins, including the carboxy-terminal portion of the immunoglobulinlike protein A (LigAC) and six novel antigens, combined with aluminum hydroxide (alum) or Salmonella flagellin (FliC) as adjuvants. Hamsters vaccinated with the different formulations elicited high antigen-specific antibody titers. Immunization with LigAC, either with alum or flagellin, conferred protective immunity but did not prevent renal colonization. Similarly, animals immunized with LigAC or LigAC coadministered with six leptospiral proteins with alum adjuvant conferred protection but did not reduce renal colonization. In contrast, immunizing animals with the pool of seven antigens in combination with flagellin conferred protection and significantly reduced renal colonization by the pathogen. The present study emphasizes the relevance of antigen composition and added adjuvant in the efficacy of antileptospirosis subunit vaccines and shows the complex relationship between immune responses and renal colonization by the pathogen. PMID:26108285

  14. Mycobacterium tuberculosis co-operonic PE32/PPE65 proteins alter host immune responses by hampering Th1 response

    Directory of Open Access Journals (Sweden)

    Mohd eKhubaib

    2016-05-01

    Full Text Available PE/PPE genes, present in cluster with ESAT-6 like genes, are suspected to have a role in antigenic variation and virulence of Mycobacterium tuberculosis. Their roles in immune evasion and immune modulation of host are also well documented. We present evidence that PE32/PPE65 present within the RD8 region are co-operonic, co-transcribed and co-translated, and play role in modulating host immune responses. Experiments with macrophage cell lines revealed that this protein complex suppresses pro-inflammatory cytokines such as TNF-α and IL-6 whereas also inducing high expression of anti-inflammatory IL-10. Immunization of mice with these recombinant proteins dampens an effective Th1 response as evident from reduced frequency of IFN-g and IL-2 producing CD4+ and CD8+ T cells. IgG sub-typing from serum of immunized mice revealed high levels of IgG1 when compared with IgG2a and IgG2b. Further IgG1/IgG2a ratio clearly demonstrated that the protein complex manipulates the host immune response favourable to the pathogen. Our results demonstrate that the co-transcribed and co-translated PE32 and PPE65 antigens are involved specifically in modulating anti-mycobacterial host immune response by hampering Th1 response.

  15. Humoral and cellular immune responses to synthetic peptides of the Leishmania donovani kinetoplastid membrane protein-11

    DEFF Research Database (Denmark)

    Jensen, A T; Gasim, S; Ismail, A

    1998-01-01

    Native kinetoplastid membrane protein-11 (KMP-11), purified from crude extracts of Leishmania donovani parasites, activates T cells from individuals who have recovered from visceral leishmaniasis. In this work we used three 38-mer peptides spanning the amino acid sequence of the L. donovani KMP-11...... as solid-phase ligands in enzyme-linked immunosorbent assays (ELISAs) and as stimulating antigens in lymphoproliferative assays in order to evaluate humoral and cellular immune responses to well-defined sequences of the protein. Antibody reactivity against the three peptides was measured in plasma from 63......-11 peptides was detected in plasma from Sudanese patients suffering from Leishmania major infections and in plasma from Sudanese and Danish patients infected with Plasmodium falciparum. In lymphoproliferative assays, 10 of 17 PBMC isolates from donors previously infected with L. donovani showed...

  16. HIV immune evasion disruption of antigen presentation by the HIV Nef protein.

    Science.gov (United States)

    Wonderlich, Elizabeth R; Leonard, Jolie A; Collins, Kathleen L

    2011-01-01

    The Human Immunodeficiency Virus (HIV) Nef protein is necessary for high viral loads and for timely progression to AIDS. Nef plays a number of roles, but its effect on antigen presentation and immune evasion are among the best characterized. Cytotoxic T lymphocytes (CTLs) recognize and lyse virally infected cells by detecting viral antigens in complex with host major histocompatibility complex class I (MHC-I) molecules on the infected cell surface. The HIV Nef protein disrupts antigen presentation at the cell surface by interfering with the normal trafficking pathway of MHC-I and thus reduces CTL recognition and lysis of infected cells. The molecular mechanism by which Nef causes MHC-I downmodulation is becoming more clear, but some questions remain. A better understanding of how Nef disrupts antigen presentation may lead to the development of drugs that enhance the ability of the anti-HIV CTLs to control HIV disease. Copyright © 2011 Elsevier Inc. All rights reserved.

  17. Differential humoral and cellular immunity induced by vaccination using plasmid DNA and protein recombinant expressing the NS3 protein of dengue virus type 3.

    Science.gov (United States)

    Hurtado-Melgoza, M L; Ramos-Ligonio, A; Álvarez-Rodríguez, L M; Meza-Menchaca, T; López-Monteon, A

    2016-12-01

    The dengue non-structural 3 (NS3) is a multifunctional protein, containing a serine-protease domain, located at the N-terminal portion, and helicase, NTPase and RTPase domains present in the C-terminal region. This protein is considered the main target for CD4+ and CD8+ T cell responses during dengue infection, which may be involved in protection. However, few studies have been undertaken evaluating the use of this protein as a protective antigen against dengue, as well as other flavivirus. In the present work we evaluated the potential of the NS3 (protease domain) as a protective antigen by comparing the administration of a recombinant protein versus a DNA vaccine in the mouse model. BALB/c mice were immunized with the recombinant protein NS3-DEN3 via intraperitoneal and with plasmid pcDNA3/NS3-DEN3 intramuscularly and the immune response was evaluated. The activity of T lymphocytes was analyzed by the MTT assay, and cells of mice immunized with the recombinant protein showed no activity when stimulated with the homologous protein. However, cells from mice immunized with DNA, responded to stimulation with the recombinant protein. When the expression (RT-PCR) and cytokine production (ELISA) was evaluated in the splenocytes, different behavior depending on the type of immunization was observed, splenocytes of mice immunized with the recombinant protein expressed cytokines such as IL-4, IL-10 and produced high concentrations of IL-1, IL-6 and TNFα. Splenocytes from mice immunized with DNA expressed IL-2 and IFNγ and did not produce IL-6. In addition, immunization with the recombinant protein induced the production of antibodies that are detected up to a dilution 1:3200 by ELISA and Western blot assays, however, the serum of mice immunized with DNA presented no detectable antibody titers. The results obtained in this study show that administration of pcDNA3/NS3-DEN3 induces a favorable response in the activation of T lymphocytes with low production of specific

  18. Protein corona-mediated targeting of nanocarriers to B cells allows redirection of allergic immune responses.

    Science.gov (United States)

    Shen, Limei; Tenzer, Stefan; Storck, Wiebke; Hobernik, Dominika; Raker, Verena Katherina; Fischer, Karl; Decker, Sandra; Dzionek, Andrzej; Krauthäuser, Susanne; Diken, Mustafa; Nikolaev, Alexej; Maxeiner, Joachim; Schuster, Petra; Kappel, Cinja; Verschoor, Admar; Schild, Hansjörg; Grabbe, Stephan; Bros, Matthias

    2018-01-31

    Nanoparticle (NP)-based vaccines are attractive immunotherapy tools because of their capability to codeliver antigen and adjuvant to antigen-presenting cells. Their cellular distribution and serum protein interaction ("protein corona") after systemic administration and their effect on the functional properties of NPs is poorly understood. We analyzed the relevance of the protein corona on cell type-selective uptake of dextran-coated NPs and determined the outcome of vaccination with NPs that codeliver antigen and adjuvant in disease models of allergy. The role of protein corona constituents for cellular binding/uptake of dextran-coated ferrous nanoparticles (DEX-NPs) was analyzed both in vitro and in vivo. DEX-NPs conjugated with the model antigen ovalbumin (OVA) and immunostimulatory CpG-rich oligodeoxynucleotides were administered to monitor the induction of cellular and humoral immune responses. Therapeutic effects of this DEX-NP vaccine in mouse models of OVA-induced anaphylaxis and allergic asthma were assessed. DEX-NPs triggered lectin-induced complement activation, yielding deposition of activated complement factor 3 on the DEX-NP surface. In the spleen DEX-NPs targeted predominantly B cells through complement receptors 1 and 2. The DEX-NP vaccine elicited much stronger OVA-specific IgG 2a production than coadministered soluble OVA plus CpG oligodeoxynucleotides. B-cell binding of the DEX-NP vaccine was critical for IgG 2a production. Treatment of OVA-sensitized mice with the DEX-NP vaccine prevented induction of anaphylactic shock and allergic asthma accompanied by IgE inhibition. Opsonization of lectin-coated NPs by activated complement components results in selective B-cell targeting. The intrinsic B-cell targeting property of lectin-coated NPs can be exploited for treatment of allergic immune responses. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

  19. Naturally-acquired cellular immune response against Plasmodium vivax merozoite surface protein-1 paralog antigen.

    Science.gov (United States)

    Changrob, Siriruk; Leepiyasakulchai, Chaniya; Tsuboi, Takafumi; Cheng, Yang; Lim, Chae Seung; Chootong, Patchanee; Han, Eun-Taek

    2015-04-15

    Plasmodium vivax merozoite surface protein-1 paralog (PvMSP1P) is a glycosylphosphatidylinositol-anchored protein expressed on the merozoite surface. This molecule is a target of natural immunity, as high anti-MSP1P-19 antibody levels were detected during P. vivax infection and the antibody inhibited PvMSP1P-erythrocyte binding. Recombinant PvMSP1P antigen results in production of a significant Th1 cytokine response in immunized mice. The present study was performed to characterize natural cellular immunity against PvMSP1P-19 and PvDBP region II in acute and recovery P. vivax infection. Peripheral blood mononuclear cells (PBMCs) from acute and recovery P. vivax infection were obtained for lymphocyte proliferation assay upon PvMSP1P-19 and PvDBP region II antigen stimulation. The culture supernatant was examined for the presence of the cytokines IL-2, TNF, IFN-γ and IL-10 by enzyme-linked immunosorbent assay (ELISA). To determine whether Th1 or Th2 have a memory response against PvMSP1P-19 and PvDBPII protein antigen, PBMCs from subjects who had recovered from P. vivax infection 8-10 weeks prior to the study were obtained for lymphocyte proliferation assay. Cytokine-producing cells were analysed by flow cytometry. IL-2 was detected at high levels in lymphocyte cultures from acutely infected P. vivax patients upon PvMSP1P-19 stimulation. Analysis of the Th1 or Th2 memory response in PBMC cultures from subjects who had recovered from P. vivax infection showed significantly elevated levels of PvMSP1P-19 and PvDBPII-specific IFN-γ-producing cells (P  response of IFN-γ-producing cells in PvMSP1P stimulation was fourfold greater in recovered subjects than that in acute-infection patients. CD4(+) T cells were the major cell phenotype involved in the response to PvMSP1P-19 and PvDBPII antigen. PvMSP1P-19 strongly induces a specific cellular immune response for protection against P. vivax compared with PvDBPII as the antigen induces activation of IFN

  20. Immune Abnormalities in Fontan Protein-Losing Enteropathy: A Case-Control Study.

    Science.gov (United States)

    Magdo, H Sonali; Stillwell, Terri L; Greenhawt, Matthew J; Stringer, Kathleen A; Yu, Sunkyung; Fifer, Carlen G; Russell, Mark W; Schumacher, Kurt R

    2015-08-01

    To comprehensively characterize the immunologic characteristics of patients with protein-losing enteropathy (PLE) post-Fontan and compare them with patients without PLE post-Fontan. Patients with PLE post-Fontan and age-matched controls post-Fontan were prospectively studied with laboratory markers of immune function. Infectious history was obtained by interview and chart review. The groups' demographics, cardiac history, immune characteristics, and infection history were compared using appropriate 2-group statistics. A total of 16 patients enrolled (8 patients with PLE and 8 controls). All patients with PLE had lymphopenia compared with 25% of controls (P = .01). All patients with PLE had markedly depressed CD4 T cell counts (median 58 cells/μL) compared with controls (median 450 cells/μL, P = .0002); CD4% was also low in the PLE group (12.3%) and normal in control (36.9%, P = .004). Both groups had mildly depressed CD8 T cells and normal to slightly elevated natural killer and B-cell subsets. A majority of patients with PLE (62.5%) had negative titers to measles, mumps, and rubella vaccination, compared with no control Fontan with a negative titer (P = .03). Despite profoundly low CD4 counts, the frequency of infection was not different between groups with no reported opportunistic infections. Patients with Fontan-associated PLE have extensive quantitative immune abnormalities, particularly CD4 deficiency. These immune abnormalities are similar to those found in non-Fontan patients with PLE caused by intestinal lymphangiectasia. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. An improved and robust DNA immunization method to develop antibodies against extra-cellular loops of multi-transmembrane proteins

    Science.gov (United States)

    Hazen, Meredith; Bhakta, Sunil; Vij, Rajesh; Randle, Steven; Kallop, Dara; Chiang, Vicki; Hötzel, Isidro; Jaiswal, Bijay S; Ervin, Karen E; Li, Bing; Weimer, Robby M; Polakis, Paul; Scheller, Richard H; Junutula, Jagath R; Hongo, Jo-Anne S

    2014-01-01

    Multi-transmembrane proteins are especially difficult targets for antibody generation largely due to the challenge of producing a protein that maintains its native conformation in the absence of a stabilizing membrane. Here, we describe an immunization strategy that successfully resulted in the identification of monoclonal antibodies that bind specifically to extracellular epitopes of a 12 transmembrane protein, multi-drug resistant protein 4 (MRP4). These monoclonal antibodies were developed following hydrodynamic tail vein immunization with a cytomegalovirus (CMV) promoter-based plasmid expressing MRP4 cDNA and were characterized by flow cytometry. As expected, the use of the immune modulators fetal liver tyrosine kinase 3 ligand (Flt3L) and granulocyte-macrophage colony-stimulating factor positively enhanced the immune response against MRP4. Imaging studies using CMV-based plasmids expressing luciferase showed that the in vivo half-life of the target antigen was less than 48 h using CMV-based plasmids, thus necessitating frequent boosting with DNA to achieve an adequate immune response. We also describe a comparison of plasmids, which contained MRP4 cDNA with either the CMV or CAG promoters, used for immunizations. The observed luciferase activity in this comparison demonstrated that the CAG promoter-containing plasmid pCAGGS induced prolonged constitutive expression of MRP4 and an increased anti-MRP4 specific immune response even when the plasmid was injected less frequently. The method described here is one that can be broadly applicable as a general immunization strategy to develop antibodies against multi-transmembrane proteins, as well as target antigens that are difficult to express or purify in native and functionally active conformation. PMID:24121517

  2. Immunization with heat shock protein 105-pulsed dendritic cells leads to tumor rejection in mice.

    Science.gov (United States)

    Yokomine, Kazunori; Nakatsura, Tetsuya; Minohara, Motozumi; Kira, Jun-ichi; Kubo, Tatsuko; Sasaki, Yutaka; Nishimura, Yasuharu

    2006-04-28

    Recently, we reported that heat shock protein 105 (HSP105) DNA vaccination induced anti-tumor immunity. In this study, we set up a preclinical study to investigate the usefulness of dendritic cells (DCs) pulsed with mouse HSP105 as a whole protein for cancer immunotherapy in vivo. The recombinant HSP105 did not induce DC maturation, and the mice vaccinated with HSP105-pulsed BM-DCs were markedly prevented from the growth of subcutaneous tumors, accompanied with a massive infiltration of both CD4+ T cells and CD8+ T cells into the tumors. In depletion experiments, we proved that both CD4+ T cells and CD8+ T cells play a crucial role in anti-tumor immunity. Both CD4+ T cells and CD8+ T cells specific to HSP105 were induced by stimulation with HSP105-pulsed DCs. As a result, vaccination of mice with BM-DCs pulsed with HSP105 itself could elicit a stronger tumor rejection in comparison to DNA vaccination.

  3. Ternary WD40 repeat-containing protein complexes: evolution, composition and roles in plant immunity

    Directory of Open Access Journals (Sweden)

    Jimi C. Miller

    2016-01-01

    Full Text Available Plants, like mammals, rely on their innate immune system to perceive and discriminate among the majority of their microbial pathogens. Unlike mammals, plants respond to this molecular dialogue by unleashing a complex chemical arsenal of defense metabolites to resist or evade pathogen infection. In basal or non-host resistance, plants utilize signal transduction pathways to detect non-self, damaged-self and altered-self-associated molecular patterns and translate these danger signals into largely inducible chemical defenses. The WD40 repeat (WDR-containing proteins Gβ and TTG1 are constituents of two independent ternary protein complexes functioning at opposite ends of a plant immune signaling pathway. Gβ and TTG1 are also encoded by single-copy genes that are ubiquitous in higher plants, implying the limited diversity and functional conservation of their respective complexes. In this review, we summarize what is currently known about the evolutionary history of these WDR-containing ternary complexes, their repertoire and combinatorial interactions, and their downstream effectors and pathways in plant defense.

  4. Identification of Vibrio harveyi proteins involved in the specific immune response of Senegalese sole (Solea senegalensis, Kaup).

    Science.gov (United States)

    Medina, A; Mancera, J M; Martínez-Manzanares, E; Moriñigo, M A; Arijo, S

    2015-11-01

    Senegalese sole cultures are frequently affected by Vibrio harveyi disease outbreaks. Vaccines in aquaculture are one of the most successful methods of preventing fish pathologies; however, these vaccines are usually composed of inactivated whole cells containing a wide pool of antigens, and some do not induce any protection against pathogens. Thus, the aim of this study was to identify immunogenic proteins of V. harveyi involved in the specific antibody production by Senegalese sole. S. senegalensis specimens were immunized, by intraperitoneal injection, with V. harveyi bacterin supplemented with inactivated extracellular polymeric substances (ECP) and Freund incomplete adjuvant to obtain polyclonal antiserum. One month later, specimens were re-inoculated with the same antigens. Sera from immunized fish were collected two months post first immunization. Strong specific immune response to V. harveyi antigens was detected by ELISA using bacterin (limit dilutions of sera were 1:64000), ECP (1:4000) and outer membrane proteins (OMP) (1:4000) as antigens. Presence of immunogenic proteins in V. harveyi ECP and OMP were determined by 2D-PAGE. For Western Blot analysis some gels were transferred onto nitrocellulose membranes and incubated with sera from S. senegalensis specimens immunized against V. harveyi. 2D-PAGE and Western Blot showed at least five reactive proteins in the ECP and two in the OMP fraction. The spots that clearly reacted with the sole antiserum were excised from stained gel, and analyzed by mass spectrometry (MALDI/TOFTOF). A database search was then performed, using MASCOT as the search method. According to the results, the five ECP spots were identified as Maltoporine, protein homologous to Metal dependent phosphohydrolase, two porins isoforms of V. harveyi and a protein homologous to the cell division protein FtsH. Reactive proteins in the OMP fraction were identified as the protein 3-hydroxyisobutyrate dehydrogenase and a protein homologous to acid

  5. Effects of immunization with the rNfa1 protein on experimental Naegleria fowleri-PAM mice.

    Science.gov (United States)

    Lee, Y J; Kim, J H; Sohn, H J; Lee, J; Jung, S Y; Chwae, Y J; Kim, K; Park, S; Shin, H J

    2011-07-01

    Free-living Naegleria fowleri causes primary amoebic meningoencephalitis (PAM) in humans and animals. To examine the effect of immunization with Nfa1 protein on experimental murine PAM because of N. fowleri, BALB/c mice were intra-peritoneally or intra-nasally immunized with a recombinant Nfa1 protein. We analysed Nfa1-specific antibody and cytokine induction, and the mean survival time of infected mice. Mice immunized intra-peritoneally or intra-nasally with rNfa1 protein developed specific IgG, IgA and IgE antibodies; the IgG response was dominated by IgG1, followed by IgG2b, IgG2a and IgG3. High levels of the Th1 cytokine, IFN-γ, and the regulatory cytokine, IL-10, were also induced. The mean survival time of mice immunized intra-peritoneally with rNfa1 protein was prolonged compared with controls, (25.0 and 15.5 days, respectively). Similarly, the mean survival time of mice immunized intra-nasally with rNfa1 protein was 24.7 days, compared with 15.0 days for controls. © 2011 Blackwell Publishing Ltd.

  6. Natural Resistance Associated Macrophage Protein Is Involved in Immune Response of Blunt Snout Bream, Megalobrama amblycephala.

    Science.gov (United States)

    Jiang, Yu-Hong; Mao, Ying; Lv, Yi-Na; Tang, Lei-Lei; Zhou, Yi; Zhong, Huan; Xiao, Jun; Yan, Jin-Peng

    2018-03-29

    The natural resistance-associated macrophage protein gene ( Nramp ), has been identified as one of the significant candidate genes responsible for modulating vertebrate natural resistance to intracellular pathogens. Here, we identified and characterized a new Nramp family member, named as maNramp , in the blunt snout bream. The full-length cDNA of maNramp consists of a 153 bp 5'UTR, a 1635 bp open reading frame encoding a protein with 544 amino acids, and a 1359 bp 3'UTR. The deduced protein (maNRAMP) possesses the typical structural features of NRAMP protein family, including 12 transmembrane domains, three N-linked glycosylation sites, and a conserved transport motif. Phylogenetic analysis revealed that maNRAMP shares the significant sequence consistency with other teleosts, and shows the higher sequence similarity to mammalian Nramp2 than Nramp1 . It was found that maNramp expressed ubiquitously in all normal tissues tested, with the highest abundance in the spleen, followed by the head kidney and intestine, and less abundance in the muscle, gill, and kidney. After lipopolysaccharide (LPS) stimulation, the mRNA level of maNramp was rapidly up-regulated, which reached a peak level at 6 h. Altogether, these results indicated that maNramp might be related to fish innate immunity and similar to mammalian Nramp1 in function.

  7. Evaluation of humoral, mucosal, and cellular immune responses following co-immunization of HIV-1 Gag and Env proteins expressed by Newcastle disease virus.

    Science.gov (United States)

    Khattar, Sunil K; Palaniyandi, Senthilkumar; Samal, Sweety; LaBranche, Celia C; Montefiori, David C; Zhu, Xiaoping; Samal, Siba K

    2015-01-01

    The combination of multiple HIV antigens in a vaccine can broaden antiviral immune responses. In this study, we used NDV vaccine strain LaSota to generate rNDV (rLaSota/optGag) expressing human codon optimized p55 Gag protein of HIV-1. We examined the effect of co-immunization of rLaSota/optGag with rNDVs expressing different forms of Env protein gp160, gp120, gp140L [a version of gp140 that lacked cytoplasmic tail and contained complete membrane-proximal external region (MPER)] and gp140S (a version of gp140 that lacked cytoplasmic tail and distal half of MPER) on magnitude and breadth of humoral, mucosal and cellular immune responses in guinea pigs and mice. Our results showed that inclusion of rLaSota/optGag with rNDVs expressing different forms of Env HIV Gag did not affect the Env-specific humoral and mucosal immune responses in guinea pigs and that the potent immune responses generated against Env persisted for at least 13 weeks post immunization. The highest Env-specific humoral and mucosal immune responses were observed with gp140S+optGag group. The neutralizing antibody responses against HIV strains BaL.26 and MN.3 induced by gp140S+optGag and gp160+optGag were higher than those elicited by other groups. Inclusion of Gag with gp160, gp140S and gp140L enhanced the level of Env-specific IFN-γ-producing CD8(+) T cells in mice. Inclusion of Gag with gp160 and gp140L also resulted in increased Env-specific CD4(+) T cells. The level of Gag-specific CD8(+) and CD4(+) T cells was also enhanced in mice immunized with Gag along with gp140S and gp120. These results indicate lack of antigen interference in a vaccine containing rNDVs expressing Env and Gag proteins.

  8. Towards development of novel immunization strategies against leishmaniasis using PLGA nanoparticles loaded with kinetoplastid membrane protein-11.

    Science.gov (United States)

    Santos, Diego M; Carneiro, Marcia W; de Moura, Tatiana R; Fukutani, Kiyoshi; Clarencio, Jorge; Soto, Manuel; Espuelas, Socorro; Brodskyn, Claudia; Barral, Aldina; Barral-Netto, Manoel; de Oliveira, Camila I

    2012-01-01

    Vaccine development has been a priority in the fight against leishmaniases, which are vector-borne diseases caused by Leishmania protozoa. Among the different immunization strategies employed to date is inoculation of plasmid DNA coding for parasite antigens, which has a demonstrated ability to induce humoral and cellular immune responses. In this sense, inoculation of plasmid DNA encoding Leishmania kinetoplasmid membrane protein-11 (KMP-11) was able to confer protection against visceral leishmaniasis. However, recently the use of antigen delivery systems such as poly(lactic-co-glycolic acid) (PLGA) nanoparticles has also proven effective for eliciting protective immune responses. In the present work, we tested two immunization strategies with the goal of obtaining protection, in terms of lesion development and parasite load, against cutaneous leishmaniasis caused by L. braziliensis. One strategy involved immunization with plasmid DNA encoding L. infantum chagasi KMP-11. Alternatively, mice were primed with PLGA nanoparticles loaded with the recombinant plasmid DNA and boosted using PLGA nanoparticles loaded with recombinant KMP-11. Both immunization strategies elicited detectable cellular immune responses with the presence of both proinflammatory and anti-inflammatory cytokines; mice receiving the recombinant PLGA nanoparticle formulations also demonstrated anti-KMP-11 IgG1 and IgG2a. Mice were then challenged with L. braziliensis, in the presence of sand fly saliva. Lesion development was not inhibited following either immunization strategy. However, immunization with PLGA nanoparticles resulted in a more prominent reduction in parasite load at the infection site when compared with immunization using plasmid DNA alone. This effect was associated with a local increase in interferon-gamma and in tumor necrosis factor-alpha. Both immunization strategies also resulted in a lower parasite load in the draining lymph nodes, albeit not significantly. Our results

  9. Immunization with LJM11 salivary protein protects against infection with Leishmania braziliensis in the presence of Lutzomyia longipalpis saliva.

    Science.gov (United States)

    Cunha, Jurema M; Abbehusen, Melissa; Suarez, Martha; Valenzuela, Jesus; Teixeira, Clarissa R; Brodskyn, Cláudia I

    2018-01-01

    Leishmania is transmitted in the presence of sand fly saliva. Protective immunity generated by saliva has encouraged identification of a vector salivary-based vaccine. Previous studies have shown that immunization with LJM11, a salivary protein from Lutzomyia longipalpis, is able to induce a Th1 immune response and protect mice against bites of Leishmania major-infected Lutzomyia longipalpis. Here, we further investigate if immunization with LJM11 recombinant protein is able to confer cross-protection against infection with Leishmania braziliensis associated with salivary gland sonicate (SGS) from Lutzomyia intermedia or Lu. longipalpis. Mice immunized with LJM11 protein exhibited an increased production of anti-LJM11 IgG, IgG1 and IgG2a and a DTH response characterized by an inflammatory infiltrate with the presence of CD4 + IFN-γ + T cells. LJM11-immunized mice were intradermally infected in the ear with L. braziliensis in the presence of Lu. longipalpis or Lu. intermedia SGS. A significant reduction of parasite numbers in the ear and lymph node in the group challenged with L. braziliensis plus Lu. longipalpis SGS was observed, but not when the challenge was performed with L. braziliensis plus Lu. intermedia SGS. A higher specific production of IFN-γ and absence of IL-10 by lymph node cells were only observed in LJM11 immunized mice after infection. After two weeks, a similar frequency of CD4 + IFN-γ + T cells was detected in LJM11 and BSA groups challenged with L. braziliensis plus Lu. longipalpis SGS, suggesting that early events possibly triggered by immunization are essential for protection against Leishmania infection. Our findings support the specificity of saliva-mediated immune responses and reinforce the importance of identifying cross-protective salivary antigens. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Protective value of immune responses developed in goats vaccinated with insoluble proteins from Sarcoptes Scabiei

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    Simson Tarigan

    2005-06-01

    Full Text Available Vaccines developed from certain membrane proteins lining the lumen of arthropod’s gut have been demonstrated effective in the control of some arthropod ectoparasites. A similar approach could also be applied to Sarcoptes scabiei since this parasite also ingests its host immunoglobulins. To evaluate immune protection of the membrane proteins, insoluble mite proteins were fractionated by successive treatment in the solutions of 1.14 M NaCl, 2% SB 3-14 Zwitterion detergent, 6 M urea, 6 M guanidine-HCl and 5% SDS. Five groups of goats (6 or 7 goats per group were immunised respectively with the protein fractions. Vaccination was performed 6 times, each with a dosage of 250 μg proteins, and 3 week intervals between vaccination. Group 6 (7 goats received PBS and adjuvant only, and served as an unvaccinated control. One week after the last vaccination, all goats were challenged with 2000 live mites on the auricles. The development of lesions were examined at 1 day, 2 days, and then every week from week 1 to 8. All animals were bled and weighed every week, and at the end of the experiment, skin scrapings were collected to determine the mite burden. Antibody responses induced by vaccination and challenge were examined by ELISA and Western blotting. This experiment showed that vaccination with the insoluble-protein fractions resulted in the development of high level of specific antibodies but the responses did not have any protective value. The severity of lesions and mite burden in the vaccinated animals were not different from those in the unvaccinated control.

  11. Effect of immunization against prostate- and testis-expressed (PATE) proteins on sperm function and fecundity in the rat.

    Science.gov (United States)

    Rajesh, Angireddy; Yenugu, Suresh

    2015-08-01

    Evaluating the immunocontraceptive potential of sperm-bound proteins is an active area of investigation. In this study, we analyzed the role of prostate- and testes-expressed (PATE) and PATE-F proteins in sperm function. Capacitation was measured as a function of tyrosine phosphorylation of sperm membrane proteins. Ionophore-induced acrosome reaction was assessed by measuring the fluorescence intensity of calcium-bound Fluo 3-AM and sperm-bound PNA-FITC in a flow cytometer. Rat spermatozoa subjected to capacitation and acrosome reaction in vitro displayed changes in the PATE and PATE-F protein localization on their surface, indicating the role of these proteins in sperm function. Capacitation and ionophore-induced acrosome reaction in vitro were inhibited in spermatozoa pre-incubated with antiserum raised in rabbit against PATE or PATE-F. Male rats were immunized with PATE proteins to assess their role in sperm function and fecundity. Antibody titer in the serum, testicular, and epididymal fluid was measured by ELISA. The motility parameters were recorded using CASA. High antibody titer was observed in serum, epididymal, and testicular fluid in rats immunized with PATE or PATE-F protein. Immunization did not cause any structural damage and inflammation in the testis and epididymis. PATE and PATE-F antisera obtained from the immunized rats inhibited acrosome reaction. Motility parameters, capacitation, acrosome reaction, and fecundity were compromised in PATE-F-immunized rats, whereas the same were not affected in rats immunized with PATE. These results suggest that PATE-F might play an important role in sperm function and fecundity and can be explored further to determine its immunocontraceptive potential. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  12. An immunoglobulin binding protein (antigen 5) of the stable fly (Diptera: Muscidae) salivary gland stimulates bovine immune responses.

    Science.gov (United States)

    Ameri, M; Wang, X; Wilkerson, M J; Kanost, M R; Broce, A B

    2008-01-01

    The stable fly, Stomoxys calcitrans (L.) (Diptera: Muscidae), is an economically important pest of livestock. Previous studies demonstrated lymphocyte suppression by crude salivary gland extract (SGE) of the stable fly. A dominant 27-kDa protein identified in the SGE was reported to stimulate immunodominant antibody responses in exposed cattle. The purpose of this study was to determine whether this protein, now identified as ahomolog of insect proteins named antigen 5 (Ag5), was responsible for the lymphocyte suppression and whether naive calves can mount an immune response to it. Calves raised in the winter were immunized with recombinant Ag5 (rAg5) expressed in Drosophila S2 cells or with "natural" Ag5 protein isolated by preparative gel electrophoresis of SGE. Control calves were immunized with adjuvant alone. Rising antibody concentrations to rAg5 were detected in two of three calves immunized with rAg5 and one of three calves immunized with natural Ag5. Recall lymphocyte responses to rAg5 were detected at 21 and 28 d postimmunization in calves immunized with rAg5 but not in calves immunized with the natural Ag5 or those exposed to adjuvant alone. Mitogen-stimulated bovine lymphocyte responses were not suppressed by rAg5. Further investigation using immunoblotting revealed that rAg5 binds to the Fc and F (ab')2 portions of bovine IgG, but not to an Fab fragment. These findings suggest that Ag5 of the stable fly salivary gland is not immunosuppressive but that it has immunoglobulin binding properties and can invoke specific antibody and memory lymphocyte responses in immunized calves.

  13. The human metapneumovirus matrix protein stimulates the inflammatory immune response in vitro.

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    Audrey Bagnaud-Baule

    Full Text Available Each year, during winter months, human Metapneumovirus (hMPV is associated with epidemics of bronchiolitis resulting in the hospitalization of many infants. Bronchiolitis is an acute illness of the lower respiratory tract with a consequent inflammation of the bronchioles. The rapid onset of inflammation suggests the innate immune response may have a role to play in the pathogenesis of this hMPV infection. Since, the matrix protein is one of the most abundant proteins in the Paramyxoviridae family virion, we hypothesized that the inflammatory modulation observed in hMPV infected patients may be partly associated with the matrix protein (M-hMPV response. By western blot analysis, we detected a soluble form of M-hMPV released from hMPV infected cell as well as from M-hMPV transfected HEK 293T cells suggesting that M-hMPV may be directly in contact with antigen presenting cells (APCs during the course of infection. Moreover, flow cytometry and confocal microscopy allowed determining that M-hMPV was taken up by dendritic cells (moDCs and macrophages inducing their activation. Furthermore, these moDCs enter into a maturation process inducing the secretion of a broad range of inflammatory cytokines when exposed to M-hMPV. Additionally, M-hMPV activated DCs were shown to stimulate IL-2 and IFN-γ production by allogeneic T lymphocytes. This M-hMPV-mediated activation and antigen presentation of APCs may in part explain the marked inflammatory immune response observed in pathology induced by hMPV in patients.

  14. A prokineticin-like protein responds to immune challenges in the gastropod pest Pomacea canaliculata.

    Science.gov (United States)

    Accorsi, Alice; Benatti, Stefania; Ross, Eric; Nasi, Milena; Malagoli, Davide

    2017-07-01

    The golden apple snail Pomacea canaliculata is an invasive pest originating from South America. It has already been found in Asia, the southern United States and more recently in the EU. Aiming to target the immune system of the snail as a way to control its spreading, we have developed organ-specific transcriptomes and looked for molecules controlling replication and differentiation of snail hemocytes. The prokineticin domain-containing protein Astakine 1 is the only cytokine known thus far capable of regulating invertebrate hematopoiesis, and we analyzed the transcriptomes looking for molecules containing a prokineticin domain. We have identified a prokineticin-like protein (PlP), that we called Pc-plp and we analyzed by real-time PCR (qPCR) its expression. In control snails, highest levels of Pc-plp were detected in the digestive gland, the ampulla (i.e., a hemocyte reservoir) and the pericardial fluid (i.e., the hematopoietic district). We tested Pc-plp expression after triggering hematopoiesis via multiple hemolymph withdrawals, or during bacterial challenge through LPS injection. In both cases a reduction of Pc-plp mRNA was observed. The multiple hemolymph withdrawals caused a significant decrease of Pc-plp mRNA in pericardial fluid and circulating hemocytes, while the LPS injection promoted the Pc-plp mRNA drop in anterior kidney, mantle and gills, organs that may act as immune barrier in molluscs. Our data indicate an important role for prokineticin domain-containing proteins as immunomodulators also in gastropods and their dynamic expression may serve as a biosensor to gauge the effectiveness of immunological interventions aimed at curtailing the spreading of the gastropod pest P. canaliculata. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Plasma immune protein analysis in the orange-spotted grouper Epinephelus coioides: Evidence for altered expressions of immune factors associated with a choline-supplemented diet.

    Science.gov (United States)

    Shiu, Ya-Li; Chiu, Kuo-Hsun; Huynh, Truong-Giang; Liu, Ping-Chung; Liu, Chun-Hung

    2017-06-01

    This study aimed to unravel the regulatory roles of choline in activating immune responses and disease resistance of the orange-spotted grouper Epinephelus coioides. Fish were fed a choline-supplemented diet at 1 g kg -1 of feed for 30 days. Fish fed a fish meal basal diet without choline-supplement served as controls. At the end of the feeding trial, fish were challenged with Vibrio alginolyticus. Meanwhile, plasma proteomics of fish in each group were also evaluated by two-dimensional gel electrophoresis (2-DE), and differentially expressed proteins were identified by tandem mass spectrophotometry (MS/MS), then a Western blot analysis or real-time polymerase chain reaction was used to confirm differential expressions of immune-enhancing proteins. Results showed that choline significantly increased survival of E. coioides 48 days after being injected with V. alginolyticus. From maps of plasma proteins, a comparative analysis between the control and choline groups revealed that 111 spots matched, with 26 altered expression spots in the choline group. Of these 26 spots, 16 were upregulated and 10 downregulated. After protein identification by reverse-phase nano-high-performance liquid chromatography-electrospray ionization MS/MS analysis, eight of 26 proteins were found to be immune-related proteins, all of which were upregulated, including complement 3 (C3), alpha-2-macroglobulin-P-like isoform (A2M), fibrinogen beta chain precursor (FBG), and immunoglobulin heavy constant mu (Ighm) proteins. Expression of the A2M protein and A2M enzyme activity in plasma of fish fed choline significantly increased compared to the control group. Additionally, A2M messenger (m)RNA transcripts were also upregulated in the liver and kidneys. Significantly higher C3 expressions at both the mRNA and protein levels were detected in the liver of fish in the choline group. Moreover, FBG gene expressions in the liver and kidneys significantly increased, while Ighm increased in the

  16. The hemochromatosis protein HFE 20 years later: An emerging role in antigen presentation and in the immune system.

    Science.gov (United States)

    Reuben, Alexandre; Chung, Jacqueline W; Lapointe, Réjean; Santos, Manuela M

    2017-09-01

    Since its discovery, the hemochromatosis protein HFE has been primarily defined by its role in iron metabolism and homeostasis, and its involvement in the genetic disease termed hereditary hemochromatosis (HH). While HH patients are typically afflicted by dysregulated iron levels, many are also affected by several immune defects and increased incidence of autoimmune diseases that have thereby implicated HFE in the immune response. Growing evidence has supported an immunological role for HFE with recent studies describing HFE specifically as it relates to MHC I antigen presentation. Here, we present a comprehensive overview of the relationship between iron metabolism, HFE, and the immune system to better understand the origin and cause of immune defects in HH patients. We further describe the role of HFE in MHC I antigen presentation and its potential to impair autoimmune responses in homeostatic conditions, a mechanism which may be exploited by tumors to evade immune surveillance. Overall, this increased understanding of the role of HFE in the immune response sets the stage for better treatment and management of HH and other iron-related diseases, as well as of the immune defects related to this condition. © 2017 The Authors. Immunity, Inflammation and Disease Published by John Wiley & Sons Ltd.

  17. Secretion of Rhoptry and Dense Granule Effector Proteins by Nonreplicating Toxoplasma gondii Uracil Auxotrophs Controls the Development of Antitumor Immunity

    Science.gov (United States)

    Fox, Barbara A.; Sanders, Kiah L.; Rommereim, Leah M.; Bzik, David J.

    2016-01-01

    Nonreplicating type I uracil auxotrophic mutants of Toxoplasma gondii possess a potent ability to activate therapeutic immunity to established solid tumors by reversing immune suppression in the tumor microenvironment. Here we engineered targeted deletions of parasite secreted effector proteins using a genetically tractable Δku80 vaccine strain to show that the secretion of specific rhoptry (ROP) and dense granule (GRA) proteins by uracil auxotrophic mutants of T. gondii in conjunction with host cell invasion activates antitumor immunity through host responses involving CD8α+ dendritic cells, the IL-12/interferon-gamma (IFN-γ) TH1 axis, as well as CD4+ and CD8+ T cells. Deletion of parasitophorous vacuole membrane (PVM) associated proteins ROP5, ROP17, ROP18, ROP35 or ROP38, intravacuolar network associated dense granule proteins GRA2 or GRA12, and GRA24 which traffics past the PVM to the host cell nucleus severely abrogated the antitumor response. In contrast, deletion of other secreted effector molecules such as GRA15, GRA16, or ROP16 that manipulate host cell signaling and transcriptional pathways, or deletion of PVM associated ROP21 or GRA3 molecules did not affect the antitumor activity. Association of ROP18 with the PVM was found to be essential for the development of the antitumor responses. Surprisingly, the ROP18 kinase activity required for resistance to IFN-γ activated host innate immunity related GTPases and virulence was not essential for the antitumor response. These data show that PVM functions of parasite secreted effector molecules, including ROP18, manipulate host cell responses through ROP18 kinase virulence independent mechanisms to activate potent antitumor responses. Our results demonstrate that PVM associated rhoptry effector proteins secreted prior to host cell invasion and dense granule effector proteins localized to the intravacuolar network and host nucleus that are secreted after host cell invasion coordinately control the

  18. The alternative complement pathway control protein H binds to immune complexes and serves their detection

    International Nuclear Information System (INIS)

    Nydegger, U.E.; Corvetta, A.; Spaeth, P.J.; Spycher, M.

    1983-01-01

    During solubilization of immune complexes C3b becomes fixed to the immunoglobulin part and serves as a receptor for the alternative complement pathway control protein H. The H-C3b immune complex interaction can be made detectable using 4% polyethyleneglycol to separate free from bound 125 I-H. Tetanus toxoid (Te)/anti-Te complexes kept soluble with fresh serum and containing 125 IU of specific antibody bound 18% of 125 I-H; when fresh serum was chelated with 10 mM EDTA, 125 I-H binding was only 5%. On sucrose density gradients, the H-binding material sedimented in the range of 12 to 30 S. In 36 serum samples from rheumatoid arthritis (RA) patients and in 12 serum samples from patients with systemic lupus erythematosus (SLE), 125 I-H binding was significantly elevated to 9.5 +/- 4.7% (mean +/- 1 SD) and 13.3 +/- 5.6%, respectively, while 125 I-H binding by 36 normal human sera was 4 +/- 2%. RA samples (17/36, 47%) and SLE samples (9/12, 75%) had H-binding values increased by more than 2 SD above the normal mean. The serum samples were also assessed for conglutinin- and C1q-binding activities; a significant correlation between H and C1q binding was observed (P less than 0.001); there was no correlation between H and conglutinin binding. Although binding to immune complexes through its interaction with C3b, H clearly detects a population of complexes other than conglutinin, thus expanding the possibilities of further characterizing pathological complexes

  19. Evasion of antiviral innate immunity by Theiler's virus L* protein through direct inhibition of RNase L.

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    Frédéric Sorgeloos

    Full Text Available Theiler's virus is a neurotropic picornavirus responsible for chronic infections of the central nervous system. The establishment of a persistent infection and the subsequent demyelinating disease triggered by the virus depend on the expression of L*, a viral accessory protein encoded by an alternative open reading frame of the virus. We discovered that L* potently inhibits the interferon-inducible OAS/RNase L pathway. The antagonism of RNase L by L* was particularly prominent in macrophages where baseline oligoadenylate synthetase (OAS and RNase L expression levels are elevated, but was detectable in fibroblasts after IFN pretreatment. L* mutations significantly affected Theiler's virus replication in primary macrophages derived from wild-type but not from RNase L-deficient mice. L* counteracted the OAS/RNase L pathway through direct interaction with the ankyrin domain of RNase L, resulting in the inhibition of this enzyme. Interestingly, RNase L inhibition was species-specific as Theiler's virus L* protein blocked murine RNase L but not human RNase L or RNase L of other mammals or birds. Direct RNase L inhibition by L* and species specificity were confirmed in an in vitro assay performed with purified proteins. These results demonstrate a novel viral mechanism to elude the antiviral OAS/RNase L pathway. By targeting the effector enzyme of this antiviral pathway, L* potently inhibits RNase L, underscoring the importance of this enzyme in innate immunity against Theiler's virus.

  20. Dengue Virus Targets the Adaptor Protein MITA to Subvert Host Innate Immunity

    Science.gov (United States)

    Yu, Chia-Yi; Chang, Tsung-Hsien; Liang, Jian-Jong; Chiang, Ruei-Lin; Lee, Yi-Ling; Liao, Ching-Len; Lin, Yi-Ling

    2012-01-01

    Dengue is one of the most important arboviral diseases caused by infection of four serotypes of dengue virus (DEN). We found that activation of interferon regulatory factor 3 (IRF3) triggered by viral infection and by foreign DNA and RNA stimulation was blocked by DEN-encoded NS2B3 through a protease-dependent mechanism. The key adaptor protein in type I interferon pathway, human mediator of IRF3 activation (MITA) but not the murine homologue MPYS, was cleaved in cells infected with DEN-1 or DEN-2 and with expression of the enzymatically active protease NS2B3. The cleavage site of MITA was mapped to LRR↓96G and the function of MITA was suppressed by dengue protease. DEN replication was reduced with overexpression of MPYS but not with MITA, while DEN replication was enhanced by MPYS knockdown, indicating an antiviral role of MITA/MPYS against DEN infection. The involvement of MITA in DEN-triggered innate immune response was evidenced by reduction of IRF3 activation and IFN induction in cells with MITA knockdown upon DEN-2 infection. NS2B3 physically interacted with MITA, and the interaction and cleavage of MITA could be further enhanced by poly(dA:dT) stimulation. Thus, we identified MITA as a novel host target of DEN protease and provide the molecular mechanism of how DEN subverts the host innate immunity. PMID:22761576

  1. Dengue virus targets the adaptor protein MITA to subvert host innate immunity.

    Science.gov (United States)

    Yu, Chia-Yi; Chang, Tsung-Hsien; Liang, Jian-Jong; Chiang, Ruei-Lin; Lee, Yi-Ling; Liao, Ching-Len; Lin, Yi-Ling

    2012-01-01

    Dengue is one of the most important arboviral diseases caused by infection of four serotypes of dengue virus (DEN). We found that activation of interferon regulatory factor 3 (IRF3) triggered by viral infection and by foreign DNA and RNA stimulation was blocked by DEN-encoded NS2B3 through a protease-dependent mechanism. The key adaptor protein in type I interferon pathway, human mediator of IRF3 activation (MITA) but not the murine homologue MPYS, was cleaved in cells infected with DEN-1 or DEN-2 and with expression of the enzymatically active protease NS2B3. The cleavage site of MITA was mapped to LRR↓(96)G and the function of MITA was suppressed by dengue protease. DEN replication was reduced with overexpression of MPYS but not with MITA, while DEN replication was enhanced by MPYS knockdown, indicating an antiviral role of MITA/MPYS against DEN infection. The involvement of MITA in DEN-triggered innate immune response was evidenced by reduction of IRF3 activation and IFN induction in cells with MITA knockdown upon DEN-2 infection. NS2B3 physically interacted with MITA, and the interaction and cleavage of MITA could be further enhanced by poly(dA:dT) stimulation. Thus, we identified MITA as a novel host target of DEN protease and provide the molecular mechanism of how DEN subverts the host innate immunity.

  2. Dengue virus targets the adaptor protein MITA to subvert host innate immunity.

    Directory of Open Access Journals (Sweden)

    Chia-Yi Yu

    Full Text Available Dengue is one of the most important arboviral diseases caused by infection of four serotypes of dengue virus (DEN. We found that activation of interferon regulatory factor 3 (IRF3 triggered by viral infection and by foreign DNA and RNA stimulation was blocked by DEN-encoded NS2B3 through a protease-dependent mechanism. The key adaptor protein in type I interferon pathway, human mediator of IRF3 activation (MITA but not the murine homologue MPYS, was cleaved in cells infected with DEN-1 or DEN-2 and with expression of the enzymatically active protease NS2B3. The cleavage site of MITA was mapped to LRR↓(96G and the function of MITA was suppressed by dengue protease. DEN replication was reduced with overexpression of MPYS but not with MITA, while DEN replication was enhanced by MPYS knockdown, indicating an antiviral role of MITA/MPYS against DEN infection. The involvement of MITA in DEN-triggered innate immune response was evidenced by reduction of IRF3 activation and IFN induction in cells with MITA knockdown upon DEN-2 infection. NS2B3 physically interacted with MITA, and the interaction and cleavage of MITA could be further enhanced by poly(dA:dT stimulation. Thus, we identified MITA as a novel host target of DEN protease and provide the molecular mechanism of how DEN subverts the host innate immunity.

  3. Induced prion protein controls immune-activated retroviruses in the mouse spleen.

    Directory of Open Access Journals (Sweden)

    Marius Lötscher

    Full Text Available The prion protein (PrP is crucially involved in transmissible spongiform encephalopathies (TSE, but neither its exact role in disease nor its physiological function are known. Here we show for mice, using histological, immunochemical and PCR-based methods, that stimulation of innate resistance was followed by appearance of numerous endogenous retroviruses and ensuing PrP up-regulation in germinal centers of the spleen. Subsequently, the activated retroviruses disappeared in a PrP-dependent manner. Our results reveal the regular involvement of endogenous retroviruses in murine immune responses and provide evidence for an essential function of PrP in the control of the retroviral activity. The interaction between PrP and ubiquitous endogenous retroviruses may allow new interpretations of TSE pathophysiology and explain the evolutionary conservation of PrP.

  4. Immune response of calves inoculated with proteins ofAnaplasma marginale bound to an immunostimulant complex

    Directory of Open Access Journals (Sweden)

    Marcela Ribeiro Gasparini

    Full Text Available Despite our current knowledge of the immunology, pathology, and genetics of Anaplasma marginale, prevention in cattle is currently based on old standbys, including live attenuated vaccines, antibiotic treatment, and maintaining enzootic stability in cattle herds. In the present study, we evaluated the use of an immunostimulant complex (ISCOMATRIX adjuvant, associated with a pool of recombinant major surface proteins (rMSP1a, rMSP1b, rMSP4 and rMSP5 to improve the humoral immune response triggered in calves mainly by IgG2. Ten calves were divided in three groups: 4 calves were inoculated with the ISCOMATRIX/rMSPs (G1; 2 calves were inoculated with ISCOMATRIX adjuvant (G2; and 4 calves received saline (G3. Three inoculations were administered at 21-day intervals. In G1, the calves showed significant increases in total IgG, IgG1 and IgG2 levels 21 days after the second inoculation, compared to the control group (p < 0.05, and G1 calves remained above the cut-off value 28 days after the third inoculation (p < 0.05. The post-immunized sera from calves in G1 reacted specifically for each of the rMSPs used. In conclusion, the ISCOMATRIX/rMSPs induced antigen-specific seroconversion in calves. Therefore, additional testing to explore the protection induced by rMSPs, both alone and in conjunction with proteins previously identified as subdominant epitopes, is warranted.

  5. Inhibition of immune responses and related proteins in Rhamdia quelen exposed to diclofenac.

    Science.gov (United States)

    Ribas, João L C; Sherry, James P; Zampronio, Aleksander R; Silva de Assis, Helena C; Simmons, Denina B D

    2017-08-01

    Nonsteroidal anti-inflammatory drugs are among the most widely detected pharmaceuticals in surface water worldwide. The nonsteroidal anti-inflammatory drug diclofenac is used to treat many types of pain and inflammation. Diclofenac's potential to cause adverse effects in exposed wildlife is a growing concern. To evaluate the effects of waterborne diclofenac on the immune response in Rhamdia quelen (South American catfish), fish were exposed to 3 concentrations of diclofenac (0.2, 2.0, and 20.0 μg/L) for 14 d. Some of the exposed fish were also given an intraperitoneal injection on day 14 of 1 mg/kg of carrageenan to evaluate cell migration to the peritoneum. Total blood leukocyte count and carrageenan-induced leukocyte migration to the peritoneal cavity, particularly of polymorphonuclear cells, were significantly affected for all diclofenac exposure groups. Nitric oxide production was significantly reduced in the diclofenac-treated fish. Plasma and kidney proteins were analyzed by means of liquid chromatography-tandem mass spectrometry in a shotgun proteomic approach. In both plasma and kidney of diclofenac-exposed R. quelen, the expression of 20 proteins related to the inflammatory process, nitric oxide production, leukocyte migration, and the complement cascade was significantly altered. In addition, class I major histocompatibility complex was significantly decreased in plasma of diclofenac-treated fish. Thus, waterborne exposure to diclofenac could lead to suppression of the innate immune system in R. quelen. Environ Toxicol Chem 2017;36:2092-2107. © 2017 SETAC. © 2017 SETAC.

  6. Microbiota-inducible Innate Immune, Siderophore Binding Protein Lipocalin 2 is Critical for Intestinal Homeostasis.

    Science.gov (United States)

    Singh, Vishal; Yeoh, Beng San; Chassaing, Benoit; Zhang, Benyue; Saha, Piu; Xiao, Xia; Awasthi, Deepika; Shashidharamurthy, Rangaiah; Dikshit, Madhu; Gewirtz, Andrew; Vijay-Kumar, Matam

    2016-07-01

    Lipocalin 2 (Lcn2) is a multifunctional innate immune protein whose expression closely correlates with extent of intestinal inflammation. However, whether Lcn2 plays a role in the pathogenesis of gut inflammation is unknown. Herein, we investigated the extent to which Lcn2 regulates inflammation and gut bacterial dysbiosis in mouse models of IBD. Lcn2 expression was monitored in murine colitis models and upon microbiota ablation/restoration. WT and Lcn2 knockout ( Lcn2 KO) mice were analyzed for gut bacterial load, composition by 16S rRNA gene pyrosequencing and, their colitogenic potential by co-housing with Il-10 KO mice. Acute (dextran sodium sulfate) and chronic (IL-10R neutralization and T-cell adoptive transfer) colitis was induced in WT and Lcn2 KO mice with or without antibiotics. Lcn2 expression was dramatically induced upon inflammation and was dependent upon presence of a gut microbiota and MyD88 signaling. Use of bone-marrow chimeric mice revealed non-immune cells are the major contributors of circulating Lcn2. Lcn2 KO mice exhibited elevated levels of entA -expressing gut bacteria burden and, moreover, a broadly distinct bacterial community relative to WT littermates. Lcn2 KO mice developed highly colitogenic T-cells and exhibited exacerbated colitis upon exposure to DSS or neutralization of IL-10. Such exacerbated colitis could be prevented by antibiotic treatment. Moreover, exposure to the microbiota of Lcn2 KO mice, via cohousing, resulted in severe colitis in Il-10 KO mice. Lcn2 is a bacterially-induced, MyD88-dependent, protein that play an important role in gut homeostasis and a pivotal role upon challenge. Hence, therapeutic manipulation of Lcn2 levels may provide a strategy to help manage diseases driven by alteration of the gut microbiota.

  7. Viral proteins expressed in the protozoan parasite Eimeria tenella are detected by the chicken immune system.

    Science.gov (United States)

    Marugan-Hernandez, Virginia; Cockle, Charlotte; Macdonald, Sarah; Pegg, Elaine; Crouch, Colin; Blake, Damer P; Tomley, Fiona M

    2016-08-23

    Eimeria species are parasitic protozoa that cause coccidiosis, an intestinal disease commonly characterised by malabsorption, diarrhoea and haemorrhage that is particularly important in chickens. Vaccination against chicken coccidiosis is effective using wild-type or attenuated live parasite lines. The development of protocols to express foreign proteins in Eimeria species has opened up the possibility of using Eimeria live vaccines to deliver heterologous antigens and function as multivalent vaccine vectors that could protect chickens against a range of pathogens. In this study, genetic complementation was used to express immunoprotective virus antigens in Eimeria tenella. Infectious bursal disease virus (IBDV) causes Gumboro, an immunosuppressive disease that affects productivity and can interfere with the efficacy of poultry vaccination programmes. Infectious laryngotracheitis virus (ILTV) causes a highly transmissible respiratory disease for which strong cellular immunity and antibody responses are required for effective vaccination. Genes encoding the VP2 protein from a very virulent strain of IBDV (vvVP2) and glycoprotein I from ILTV (gI) were cloned downstream of 5'Et-Actin or 5'Et-TIF promoter regions in plasmids that also contained a mCitrine fluorescent reporter cassette under control of the 5'Et-MIC1 promoter. The plasmids were introduced by nucleofection into E. tenella sporozoites, which were then used to infect chickens. Progeny oocysts were sorted by FACS and passaged several times in vivo until the proportion of fluorescent parasites in each transgenic population reached ~20 % and the number of transgene copies per parasite genome decreased to < 10. All populations were found to transcribe and express the transgene and induced the generation of low titre, transgene-specific antibodies when used to immunise chickens. E. tenella can express antigens of other poultry pathogens that are successfully recognised by the chicken immune system. Nonetheless

  8. Changes over lactation in breast milk serum proteins involved in the maturation of immune and digestive system of the infant.

    Science.gov (United States)

    Zhang, Lina; de Waard, Marita; Verheijen, Hester; Boeren, Sjef; Hageman, Jos A; van Hooijdonk, Toon; Vervoort, Jacques; van Goudoever, Johannes B; Hettinga, Kasper

    2016-09-16

    To objective of this study was to better understand the biological functions of breast milk proteins in relation to the growth and development of infants over the first six months of life. Breast milk samples from four individual women collected at seven time points in the first six months after delivery were analyzed by filter aided sample preparation and dimethyl labeling combined with liquid chromatography tandem mass spectrometry. A total of 247 and 200 milk serum proteins were identified and quantified, respectively. The milk serum proteome showed a high similarity (80% overlap) on the qualitative level between women and over lactation. The quantitative changes in milk serum proteins were mainly caused by three groups of proteins, enzymes, and transport and immunity proteins. Of these 21 significantly changed proteins, 30% were transport proteins, such as serum albumin and fatty acid binding protein, which are both involved in transporting nutrients to the infant. The decrease of the enzyme bile salt-activated lipase as well as the immunity proteins immunoglobulins and lactoferrin coincide with the gradual maturation of the digestive and immune system of infants. The human milk serum proteome didn't differ qualitatively but it did quantitatively, both between mothers and as lactation advanced. The changes of the breast milk serum proteome over lactation corresponded with the development of the digestive and immune system of infants. Breast milk proteins provide nutrition, but also contribute to healthy development of infants. Despite the previously reported large number of identified breast milk proteins and their changes over lactation, less is known on the changes of these proteins in individual mothers. This study is the first to determine the qualitative and quantitative changes of milk proteome over lactation between individual mothers. The results indicate that the differences in the milk proteome between individual mothers are more related to the

  9. Induction of Boosted Immune Response in Mice by Leptospiral Surface Proteins Expressed in Fusion with DnaK

    Directory of Open Access Journals (Sweden)

    Marina V. Atzingen

    2014-01-01

    Full Text Available Leptospirosis is an important global disease of human and veterinary concern. Caused by pathogenic Leptospira, the illness was recently classified as an emerging infectious disease. Currently available veterinarian vaccines do not induce long-term protection against infection and do not provide cross-protective immunity. Several studies have suggested the use of DnaK as an antigen in vaccine formulation, due to an exceptional degree of immunogenicity. We focused on four surface proteins: rLIC10368 (Lsa21, rLIC10494, rLIC12690 (Lp95, and rLIC12730, previously shown to be involved in host-pathogen interactions. Our goal was to evaluate the immunogenicity of the proteins genetically fused with DnaK in animal model. The chosen genes were amplified by PCR methodology and cloned into pAE, an E. coli vector. The recombinant proteins were expressed alone or in fusion with DnaK at the N-terminus. Our results demonstrate that leptospiral proteins fused with DnaK have elicited an enhanced immune response in mice when compared to the effect promoted by the individual proteins. The boosted immune effect was demonstrated by the production of total IgG, lymphocyte proliferation, and significant amounts of IL-10 in supernatant of splenocyte cell cultures. We believe that this approach could be employed in vaccines to enhance presentation of antigens of Leptospira to professional immune cells.

  10. Spodoptera frugiperda X-Tox Protein, an Immune Related Defensin Rosary, Has Lost the Function of Ancestral Defensins

    OpenAIRE

    Destoumieux-Garz?n, Delphine; Brehelin, Michel; Bulet, Philippe; Boublik, Yvan; Girard, Pierre-Alain; Baghdiguian, Stephen; Zumbihl, Robert; Escoubas, Jean-Michel

    2009-01-01

    BACKGROUND: X-tox proteins are a family of immune-related proteins only found in Lepidoptera and characterized by imperfectly conserved tandem repeats of several defensin-like motifs. Previous phylogenetic analysis of X-tox genes supported the hypothesis that X-tox have evolved from defensins in a lineage-specific gene evolution restricted to Lepidoptera. In this paper, we performed a protein study in which we asked whether X-tox proteins have conserved the antimicrobial functions of their an...

  11. Immune response and growth of Lates calcarifer fed on hydrolized protein-supplemented diet

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    Romi Novriadi

    2015-10-01

    Full Text Available ABSTRACT This experiment was aimed to investigate the effects of protein hydrolysates on growth performance and immune response of the Asian sea bass Lates calcarifer Bloch. The experiment was performed in two different rearing period, namely nursery and grow out by using completely randomized design. Experimental fish were fed with three types of diets and each treatment was repeated three times: a local commercial diet (control, coated or not, with 2%, and 3% of protein hydrolysates. Challenge test was performed with Vibrio parahaemolyticus at a density of 105 sel/mL by using immersion method. The results showed that the the neutrofil, leukocyte and monocyte of the fish fed on protein hydrolisates were significantly higher than those non-supplemented group (P<0.05. Meanwhile, the lymphocyte on the fish treated with hydrolisates showed no difference amongst treatments (P<0.05. The growth performance of Asian sea bass fed on protein hydrolisates significantly improved the total fish weight (g, relative weight gain, specific growth rate, final weight (g and final length (cm in comparison to the control (P<0.05 both at nursery and grow out phase. Higher survival both at the nursery and grow out phase were obtained by the group fed with 3% protein hydrolisates: 97.28±0.18% and 86.0±4.32%. Followed with 2% supplementation level 96.75±0.28% and 78.4±7.7%. The lowest survival was shown by the control group with 93.65±0.13% and 20.1±21.1% from nursery and grow-out phase respectively. Results of challenged test showed that the protein hydrolisates supplementation was able to improve the post-challenged survival and resistency of Asian sea bass against V. parahaemolyticus infection. Fish treated with 3% protein hydrolisates generated higher survival 78,33±2,89%, followed by treatment 2% 73,33±5,77%, and control 31,67±7,64% after five days post immersion (P<0.05. Keywords: Asian sea bass, protein hydrolisates, growth, immune system

  12. Immunization with H1, HASPB1 and MML Leishmania proteins in a vaccine trial against experimental canine leishmaniasis.

    Science.gov (United States)

    Moreno, J; Nieto, J; Masina, S; Cañavate, C; Cruz, I; Chicharro, C; Carrillo, E; Napp, S; Reymond, C; Kaye, P M; Smith, D F; Fasel, N; Alvar, J

    2007-07-20

    The protective capabilities of three Leishmania recombinant proteins - histone 1 (H1) and hydrophilic acylated surface protein B1 (HASPB1) immunized singly, or together as a protein cocktail vaccine with Montanide, and the polyprotein MML immunized with MPL-SE adjuvant - were assessed in beagle dogs. Clinical examination of the dogs was carried out periodically under blinded conditions and the condition of the dogs defined as asymptomatic or symptomatic. At the end of the trial, we were able to confirm that following infection with L. infantum promastigotes, five out of eight dogs immunized with H1 Montanide, and four out of eight dogs immunized with either the combination of HASPB1 with Montanide or the combination of H1+HASPB1 with Montanidetrade mark, remained free of clinical signs, compared with two out of seven dogs immunized with the polyprotein MML and adjuvant MPL-SE, and two out of eight dogs in the control group. The results demonstrate that HASPB1 and H1 antigens in combination with Montanide were able to induce partial protection against canine leishmaniasis, even under extreme experimental challenge conditions.

  13. Suppression or Activation of Immune Responses by Predicted Secreted Proteins of the Soybean Rust Pathogen Phakopsora pachyrhizi.

    Science.gov (United States)

    Qi, Mingsheng; Grayczyk, James P; Seitz, Janina M; Lee, Youngsill; Link, Tobias I; Choi, Doil; Pedley, Kerry F; Voegele, Ralf T; Baum, Thomas J; Whitham, Steven A

    2018-01-01

    Rust fungi, such as the soybean rust pathogen Phakopsora pachyrhizi, are major threats to crop production. They form specialized haustoria that are hyphal structures intimately associated with host-plant cell membranes. These haustoria have roles in acquiring nutrients and secreting effector proteins that manipulate host immune systems. Functional characterization of effector proteins of rust fungi is important for understanding mechanisms that underlie their virulence and pathogenicity. Hundreds of candidate effector proteins have been predicted for rust pathogens, but it is not clear how to prioritize these effector candidates for further characterization. There is a need for high-throughput approaches for screening effector candidates to obtain experimental evidence for effector-like functions, such as the manipulation of host immune systems. We have focused on identifying effector candidates with immune-related functions in the soybean rust fungus P. pachyrhizi. To facilitate the screening of many P. pachyrhizi effector candidates (named PpECs), we used heterologous expression systems, including the bacterial type III secretion system, Agrobacterium infiltration, a plant virus, and a yeast strain, to establish an experimental pipeline for identifying PpECs with immune-related functions and establishing their subcellular localizations. Several PpECs were identified that could suppress or activate immune responses in nonhost Nicotiana benthamiana, N. tabacum, Arabidopsis, tomato, or pepper plants.

  14. Immunization with recombinant enterovirus 71 viral capsid protein 1 fragment stimulated antibody responses in hamsters

    Directory of Open Access Journals (Sweden)

    Ch’ng Wei-Choong

    2012-08-01

    Full Text Available Abstract Enterovirus 71 (EV71 causes severe neurological diseases resulting in high mortality in young children worldwide. Development of an effective vaccine against EV71 infection is hampered by the lack of appropriate animal models for efficacy testing of candidate vaccines. Previously, we have successfully tested the immunogenicity and protectiveness of a candidate EV71 vaccine, containing recombinant Newcastle disease virus capsids that display an EV71 VP1 fragment (NPt-VP11-100 protein, in a mouse model of EV71 infection. A drawback of this system is its limited window of EV71 susceptibility period, 2 weeks after birth, leading to restricted options in the evaluation of optimal dosing regimens. To address this issue, we have assessed the NPt-VP11-100 candidate vaccine in a hamster system, which offers a 4-week susceptibility period to EV71 infection. Results obtained showed that the NPt-VP11-100 candidate vaccine stimulated excellent humoral immune response in the hamsters. Despite the high level of antibody production, they failed to neutralize EV71 viruses or protect vaccinated hamsters in viral challenge studies. Nevertheless, these findings have contributed towards a better understanding of the NPt-VP11-100 recombinant protein as a candidate vaccine in an alternative animal model system.

  15. Neospora caninum immune mapped protein 1 (NcIMP1 is a novel vaccine candidate against neosporosis

    Directory of Open Access Journals (Sweden)

    Xia CUI,Daoyu YANG,Tao LEI,Hui WANG,Pan HAO,Qun LIU

    2015-03-01

    Full Text Available The Neospora caninum immune mapped protein 1 (NcIMP1 was identified as a membrane protein, and a previous study indicated that NcIMP1 could be a promising vaccine candidate against neosporosis. In this study, the immune response and protection efficacy of NcIMP1 were evaluated. The coding sequence of NcIMP1 was inserted into the eukaryotic expression vector pcDNA 3.1(+, resulting in the recombination plasmid pcDNA-IMP1, which was used for the intramuscular immunization of BALB/c mice. After immunization, the immune response was evaluated using a lymphoproliferative assay and cytokine and antibody measurements. Quantification of the cerebral parasite burden of mice challenged with 2 × 106N. caninum was performed 14 days after the last immunization. The results showed that the mice immunized with pcDNA-IMP1 developed a high level of specific antibody responses against recombinant NcIMP1, with a mixed IgG1/IgG2a response and a predominance of IgG2a production. The cellular immune response was associated with the production of IFN-&Ggr;, IL-2, IL-4 and IL-10 cytokines. The experiment was terminated 30 days p.i., and the cerebral parasite burden in each mouse was assessed by quantitative PCR. The parasite burden was significantly reduced in the pcDNA-IMP1-vaccinated mice. These data suggest that IMP1 is a promising vaccine candidate against neosporosis.

  16. DNA repair proteins in cells of the human immune system; Le proteine della riparazione del DNA in cellule del sistema immunitario umano

    Energy Technology Data Exchange (ETDEWEB)

    Frasca, D.; Barattini, P.; Guidi, F.; Scarpaci, S. [ENEA, Sez. Tossicologia e Scienze Biomediche, Rome (Italy); Doria, G. [Rome Univ. Tor Vergata, Rome (Italy). Cattedra di Immunologia

    2001-02-01

    Human longevity depends on the efficiency of DNA repair mechanisms. In irradiated cells of the human immune system, the principal repair mechanism involves the DNA-Pk protein complex. [Italian] La durata della vita dipende dalla efficienza di meccanismi di riparazione del DNA. Nelle cellule del sistema immunitario umano danneggiate il principale meccanismo di riparazione coinvolge il complesso proteico DNA-PK.

  17. Humoral immune-response against human cytomegalovirus (hcmv)-specific proteins after hcmv infection in lung transplantation as detected with recombinant and naturally-occurring proteins

    NARCIS (Netherlands)

    van Zanten, J; Harmsen, M. C.; van der Giessen, M.; van der Bij, W; Prop, J.; de Leij, L; The, T. Hauw

    The humoral immune response to four intracellularly located cytomegalovirus (CMV) proteins was studied in 15 lung transplant recipients experiencing active CMV infections. Five patients had primary infections, and 10 had secondary infections. Antibodies of the immunoglobulin M (IgM) and IgG classes

  18. Suppression or activation of immune responses by predicted secreted proteins of the soybean rust pathogen Phakopsora pachyrhizi

    Science.gov (United States)

    Rust fungi, such as Phakopsora pachyrhizi, are major threats to crop production. They form specialized haustoria that are intimately associated with plant cells. These haustoria have roles in acquiring nutrients and secreting effector proteins that manipulate host immune systems. Functional characte...

  19. Immunization strategy against cervical cancer involving an alphavirus vector expressing high levels of a stable fusion protein of human papillomavirus 16 E6 and E7

    NARCIS (Netherlands)

    Daemen, T; Regts, J; Holtrop, M; Wilschut, J

    We are developing immunization strategies against cervical carcinoma and premalignant disease, based on the use of recombinant Semliki Forest virus (SFV) encoding the onco-proteins E6 and E7 from high-risk human papilloma viruses (HPV). Thus far, protein-based, as well as genetic immunization

  20. Receptor interacting protein kinase-2 inhibition by CYLD impairs anti-bacterial immune responses in macrophages

    Directory of Open Access Journals (Sweden)

    Katharina eWex

    2016-01-01

    Full Text Available Upon infection with intracellular bacteria, nucleotide oligomerization domain protein 2 (NOD2 recognizes bacterial muramyl dipeptide and binds, subsequently, to receptor-interacting serine/threonine kinase 2 (RIPK2. RIPK2 mediates the activation of immune responses via the nuclear factor-κB (NF-κB and extracellular-signal regulated kinase (ERK pathways. Previously, it has been shown that RIPK2 activation dependens on its K63-ubiquitination by the E3 ligases pellino-3 and ITCH, whereas the deubiquitinating enzyme A20 counter-regulates RIPK2 activity by cleaving K63-polyubiquitin chains from RIPK2. Here, we newly identify the deubiquitinating enzyme CYLD as a new interacting partner and inhibitor of RIPK2. We show that CYLD binds to and removes K63-polyubiquitin chains from RIPK2 in Listeria monocytogenes (Lm infected bone-marrow-derived macrophages (BMDM. CYLD-mediated K63-deubiquitination of RIPK2 resulted in an impaired activation of both NF-κB and ERK1/2 pathways, reduced production of proinflammatory cytokines (IL-6, IL-12, anti-listerial ROS and NO, and, finally, impaired pathogen control. In turn, RIPK2 inhibition by siRNA prevented activation of NF-κB and ERK1/2 and completely abolished the protective effect of CYLD-deficiency with respect to the production of IL-6, NO, ROS and pathogen control. Noteworthy, CYLD also inhibited autophagy of Listeria in a RIPK2-ERK1/2 dependent manner.The protective function of CYLD-deficiency was dependent on IFN-γ pre-stimulation of infected macrophages. Interestingly, the reduced NF-κB activation in CYLD-expressing macrophages limited the protective effect of IFN-γ by reducing NF-κB-dependent STAT1 activation. Taken together, our study identifies CYLD as an important inhibitor of RIPK2-dependent anti-bacterial immune responses in macrophages.

  1. Spirulina ameliorates methotrexate hepatotoxicity via antioxidant, immune stimulation, and proinflammatory cytokines and apoptotic proteins modulation.

    Science.gov (United States)

    Khafaga, Asmaa F; El-Sayed, Yasser S

    2018-03-01

    Methotrexate (MTX) is an efficient cytotoxic drug used against various carcinogenic, inflammatory and autoimmune diseases; however, the hepatotoxicity of MTX limits its use. Therefore, the present study aimed to evaluate the potential hepatoprotective and immune-stimulant effect of Spirulina platensis (SP) against MTX acute toxicity. Thirty-two male Wistar rats were randomly allocated into the following four groups (n = 8): control, SP (500 mg/kg bwt, oral gavage daily for 21 days), MTX (20 mg/kg bwt, single ip injection), and MTX+SP. Hepatic and splenic histoarchitecture, leukocyte counts and serum immunoglobulins were evaluated. Hepatic oxidant/antioxidant status, proinflammatory cytokines (tumor necrosis factor-α, and interleukin 6), and pro-apoptotic proteins (caspase 3 and Bax) immunoexpression were assessed. MTX induced extensive hepatic necrosis and vacuolation, and sever lymphoid depletion in splenic white pulp with increased levels of serum transaminases, lactate dehydrogenase, and hepatic malondialdehyde, tumor necrosis factor-α and interleukin 6; and number of caspase 3- and Bax-positive hepatocytes. A significant decrease in leukocyte counts, serum immunoglobulins (IgA, IgM and IgG) level, and hepatic antioxidant enzymes (GSH, GPx, SOD, and CAT) was also detected. Pretreatment with SP resulted in significant improvements in hepatic and splenic histologic architecture, as well as restoring liver enzymes and reduction of lipid peroxidation product, proinflammatory cytokines, and caspase 3 and Bax immunoexpression. Additionally, a significant increase in antioxidant enzymes, serum immunoglobulins, and total leukocyte counts was demonstrated. SP possesses promising antioxidant, anti-inflammatory, anti-apoptotic and immune stimulatory properties against MTX-induced hepatotoxicity and immunosuppression. Copyright © 2018 Elsevier Inc. All rights reserved.

  2. Immunization with a recombinant vaccinia virus that encodes nonstructural proteins of the hepatitis C virus suppresses viral protein levels in mouse liver.

    Directory of Open Access Journals (Sweden)

    Satoshi Sekiguchi

    Full Text Available Chronic hepatitis C, which is caused by infection with the hepatitis C virus (HCV, is a global health problem. Using a mouse model of hepatitis C, we examined the therapeutic effects of a recombinant vaccinia virus (rVV that encodes an HCV protein. We generated immunocompetent mice that each expressed multiple HCV proteins via a Cre/loxP switching system and established several distinct attenuated rVV strains. The HCV core protein was expressed consistently in the liver after polyinosinic acid-polycytidylic acid injection, and these mice showed chronic hepatitis C-related pathological findings (hepatocyte abnormalities, accumulation of glycogen, steatosis, liver fibrosis, and hepatocellular carcinoma. Immunization with one rVV strain (rVV-N25, which encoded nonstructural HCV proteins, suppressed serum inflammatory cytokine levels and alleviated the symptoms of pathological chronic hepatitis C within 7 days after injection. Furthermore, HCV protein levels in liver tissue also decreased in a CD4 and CD8 T-cell-dependent manner. Consistent with these results, we showed that rVV-N25 immunization induced a robust CD8 T-cell immune response that was specific to the HCV nonstructural protein 2. We also demonstrated that the onset of chronic hepatitis in CN2-29((+/-/MxCre((+/- mice was mainly attributable to inflammatory cytokines, (tumor necrosis factor TNF-α and (interleukin IL-6. Thus, our generated mice model should be useful for further investigation of the immunological processes associated with persistent expression of HCV proteins because these mice had not developed immune tolerance to the HCV antigen. In addition, we propose that rVV-N25 could be developed as an effective therapeutic vaccine.

  3. Elongation Factor Tu and Heat Shock Protein 70 Are Membrane-Associated Proteins from Mycoplasma ovipneumoniae Capable of Inducing Strong Immune Response in Mice.

    Science.gov (United States)

    Jiang, Fei; He, Jinyan; Navarro-Alvarez, Nalu; Xu, Jian; Li, Xia; Li, Peng; Wu, Wenxue

    2016-01-01

    Chronic non-progressive pneumonia, a disease that has become a worldwide epidemic has caused considerable loss to sheep industry. Mycoplasma ovipneumoniae (M. ovipneumoniae) is the causative agent of interstitial pneumonia in sheep, goat and bighorn. We here have identified by immunogold and immunoblotting that elongation factor Tu (EF-Tu) and heat shock protein 70 (HSP 70) are membrane-associated proteins on M. ovipneumonaiea. We have evaluated the humoral and cellular immune responses in vivo by immunizing BALB/c mice with both purified recombinant proteins rEF-Tu and rHSP70. The sera of both rEF-Tu and rHSP70 treated BALB/c mice demonstrated increased levels of IgG, IFN-γ, TNF-α, IL-12(p70), IL-4, IL-5 and IL-6. In addition, ELISPOT assay showed significant increase in IFN-γ+ secreting lymphocytes in the rHSP70 group when compared to other groups. Collectively our study reveals that rHSP70 induces a significantly better cellular immune response in mice, and may act as a Th1 cytokine-like adjuvant in immune response induction. Finally, growth inhibition test (GIT) of M. ovipneumoniae strain Y98 showed that sera from rHSP70 or rEF-Tu-immunized mice inhibited in vitro growth of M. ovipneumoniae. Our data strongly suggest that EF-Tu and HSP70 of M. ovipneumoniae are membrane-associated proteins capable of inducing antibody production, and cytokine secretion. Therefore, these two proteins may be potential candidates for vaccine development against M. ovipneumoniae infection in sheep.

  4. Elongation Factor Tu and Heat Shock Protein 70 Are Membrane-Associated Proteins from Mycoplasma ovipneumoniae Capable of Inducing Strong Immune Response in Mice.

    Directory of Open Access Journals (Sweden)

    Fei Jiang

    Full Text Available Chronic non-progressive pneumonia, a disease that has become a worldwide epidemic has caused considerable loss to sheep industry. Mycoplasma ovipneumoniae (M. ovipneumoniae is the causative agent of interstitial pneumonia in sheep, goat and bighorn. We here have identified by immunogold and immunoblotting that elongation factor Tu (EF-Tu and heat shock protein 70 (HSP 70 are membrane-associated proteins on M. ovipneumonaiea. We have evaluated the humoral and cellular immune responses in vivo by immunizing BALB/c mice with both purified recombinant proteins rEF-Tu and rHSP70. The sera of both rEF-Tu and rHSP70 treated BALB/c mice demonstrated increased levels of IgG, IFN-γ, TNF-α, IL-12(p70, IL-4, IL-5 and IL-6. In addition, ELISPOT assay showed significant increase in IFN-γ+ secreting lymphocytes in the rHSP70 group when compared to other groups. Collectively our study reveals that rHSP70 induces a significantly better cellular immune response in mice, and may act as a Th1 cytokine-like adjuvant in immune response induction. Finally, growth inhibition test (GIT of M. ovipneumoniae strain Y98 showed that sera from rHSP70 or rEF-Tu-immunized mice inhibited in vitro growth of M. ovipneumoniae. Our data strongly suggest that EF-Tu and HSP70 of M. ovipneumoniae are membrane-associated proteins capable of inducing antibody production, and cytokine secretion. Therefore, these two proteins may be potential candidates for vaccine development against M. ovipneumoniae infection in sheep.

  5. Recombinant protein rMBP-NAP restricts tumor progression by triggering antitumor immunity in mouse metastatic lung cancer.

    Science.gov (United States)

    Wang, Ting; Du, Mingxuan; Ji, Zhenyu; Ding, Cong; Wang, Chengbo; Men, Yingli; Liu, Shimeng; Liang, Taotao; Liu, Xin; Kang, Qiaozhen

    2018-02-01

    Recombinant Helicobacter pylori neutrophil-activating protein fused with maltose-binding protein (rMBP-NAP), a potential TLR2 ligand, was reported to possess immunomodulatory effects on in situ tumors in our previous study. In the present work, we attempt to elucidate the effect of rMBP-NAP at the local immune modulation in B16-F10-induced metastatic lung cancer. Our results demonstrated that growth of B16-F10 melanoma metastases in the lung was significantly arrested after rMBP-NAP treatment, along with marked reduction in metastatic lung nodules and significant increase in survival. The treatment induced both local and systemic immune responses, which were associated with higher influx of CD4 + /CD8 + T cells and drove toward Th1-like and cytotoxic immune environment. Moreover, rMBP-NAP also showed significant anti-angiogenic activity by reducing vascularization in lung tumor sections. rMBP-NAP could induce antitumor immunity through activating Th1 cells and producing pro-inflammatory cytokines, which are responsible for the effective cytotoxic immune response against cancer progression. Our findings indicate that rMBP-NAP might be a novel antitumor therapeutic strategy.

  6. Serological characterization of guinea pigs infected with H3N2 human influenza or immunized with hemagglutinin protein

    Directory of Open Access Journals (Sweden)

    Bushnell Ruth V

    2010-08-01

    Full Text Available Abstract Background Recent and previous studies have shown that guinea pigs can be infected with, and transmit, human influenza viruses. Therefore guinea pig may be a useful animal model for better understanding influenza infection and assessing vaccine strategies. To more fully characterize the model, antibody responses following either infection/re-infection with human influenza A/Wyoming/03/2003 H3N2 or immunization with its homologous recombinant hemagglutinin (HA protein were studied. Results Serological samples were collected and tested for anti-HA immunoglobulin by ELISA, antiviral antibodies by hemagglutination inhibition (HI, and recognition of linear epitopes by peptide scanning (PepScan. Animals inoculated with infectious virus demonstrated pronounced viral replication and subsequent serological conversion. Animals either immunized with the homologous HA antigen or infected, showed a relatively rapid rise in antibody titers to the HA glycoprotein in ELISA assays. Antiviral antibodies, measured by HI assay, were detectable after the second inoculation. PepScan data identified both previously recognized and newly defined linear epitopes. Conclusions Infection and/or recombinant HA immunization of guinea pigs with H3N2 Wyoming influenza virus resulted in a relatively rapid production of viral-specific antibody thus demonstrating the strong immunogenicity of the major viral structural proteins in this animal model for influenza infection. The sensitivity of the immune response supports the utility of the guinea pig as a useful animal model of influenza infection and immunization.

  7. Human cytomegalovirus tegument protein pUL83 inhibits IFI16-mediated DNA sensing for immune evasion.

    Science.gov (United States)

    Li, Tuo; Chen, Jin; Cristea, Ileana M

    2013-11-13

    Nuclear sensing of viral DNA has emerged as an essential step in innate immune responses against herpesviruses. Here, we provide mechanistic insight into host recognition of human cytomegalovirus (HCMV) and subsequent immune evasion by this prominent DNA virus. We establish that the interferon-inducible protein IFI16 acts as a nuclear DNA sensor following HCMV infection, binding viral DNA and triggering expression of antiviral cytokines via the STING-TBK1-IRF3 signaling pathway. The HCMV tegument protein pUL83 inhibits this response by interacting with the IFI16 pyrin domain, blocking its oligomerization upon DNA sensing and subsequent immune signals. pUL83 disrupts IFI16 by concerted action of its N- and C-terminal domains, in which an evolutionarily conserved N-terminal pyrin association domain (PAD) binds IFI16. Additionally, phosphorylation of the N-terminal domain modulates pUL83-mediated inhibition of pyrin aggregation. Collectively, our data elucidate the interplay between host DNA sensing and HCMV immune evasion, providing targets for restoring antiviral immunity. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. Immune responses of a chimaeric protein vaccine containing Mycoplasma hyopneumoniae antigens and LTB against experimental M. hyopneumoniae infection in pigs.

    Science.gov (United States)

    Marchioro, Silvana B; Sácristan, Rubén Del Pozo; Michiels, Annelies; Haesebrouck, Freddy; Conceição, Fabricio R; Dellagostin, Odir A; Maes, Dominiek

    2014-08-06

    A recombinant chimaeric protein containing three Mycoplasma hyopneumoniae antigens (C-terminal portion of P97, heat shock protein P42, and NrdF) fused to an adjuvant, the B subunit of heat-labile enterotoxin of Escherichia coli (LTB), was used to immunize pigs against enzootic pneumonia. The systemic and local immune responses, as well as the efficacy of the chimaeric protein in inducing protection against experimental M. hyopneumoniae infection were evaluated. In total, 60 male piglets, purchased from a M. hyopneumoniae-free herd, at 4 weeks of age were randomly allocated to six different experimental groups of 10 animals each: recombinant chimaeric protein by intramuscular (IM) (1) or intranasal (IN) (2) administration, commercial bacterin by IM administration (3), and the adjuvant LTB by IM (4, control group A) or IN (5, control group B) administration. All groups were immunized at 24 and 38 days of age and challenged at 52 days of age. The sixth group that was not challenged was used as the negative control (IN [n=5] or IM [n=5] administration of the LTB adjuvant). Compared with the non-challenged group, administration of the chimaeric protein induced significant (Phyopneumoniae infection in pigs. This lack of effectiveness points towards the need for further studies to improve the efficacy of this subunit-based vaccine approach. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. Glycosylation of Candida albicans cell wall proteins is critical for induction of innate immune responses and apoptosis of epithelial cells.

    Directory of Open Access Journals (Sweden)

    Jeanette Wagener

    Full Text Available C. albicans is one of the most common fungal pathogen of humans, causing local and superficial mucosal infections in immunocompromised individuals. Given that the key structure mediating host-C. albicans interactions is the fungal cell wall, we aimed to identify features of the cell wall inducing epithelial responses and be associated with fungal pathogenesis. We demonstrate here the importance of cell wall protein glycosylation in epithelial immune activation with a predominant role for the highly branched N-glycosylation residues. Moreover, these glycan moieties induce growth arrest and apoptosis of epithelial cells. Using an in vitro model of oral candidosis we demonstrate, that apoptosis induction by C. albicans wild-type occurs in early stage of infection and strongly depends on intact cell wall protein glycosylation. These novel findings demonstrate that glycosylation of the C. albicans cell wall proteins appears essential for modulation of epithelial immunity and apoptosis induction, both of which may promote fungal pathogenesis in vivo.

  10. DMBT1 encodes a protein involved in the immune defense and in epithelial differentiation and is highly unstable in cancer

    DEFF Research Database (Denmark)

    Mollenhauer, J; Herbertz, S; Holmskov, U

    2000-01-01

    into the functions of DMBT1 by expression analyses and studies with a monoclonal antibody against the protein. The DMBT1 mRNA is expressed throughout the immune system, and Western blot studies demonstrated that isoforms of DMBT1 are identical to the collectin-binding protein gp-340, a glycoprotein that is involved...... in the respiratory immune defense. Immunohistochemical analyses revealed that DMBT1 is produced by both tumor-associated macrophages and tumor cells and that it is deregulated in glioblastoma multiforme in comparison to normal brain tissue. Our data further suggest that the proteins CRP-ductin and hensin, both...... of which have been implicated in epithelial differentiation, are the DMBT1 orthologs in mice and rabbits, respectively. These findings and the spatial and temporal distribution of DMBT1 in fetal and adult epithelia suggest that DMBT1 further plays a role in epithelial development. Rearrangements of DMBT1...

  11. Immune evasion activities of accessory proteins Vpu, Nef and Vif are conserved in acute and chronic HIV-1 infection.

    Science.gov (United States)

    Mlcochova, Petra; Apolonia, Luis; Kluge, Silvia F; Sridharan, Aishwarya; Kirchhoff, Frank; Malim, Michael H; Sauter, Daniel; Gupta, Ravindra K

    2015-08-01

    Heterosexual HIV-1 transmission has been identified as a genetic bottleneck and a single transmitted/founder (T/F) variant with reduced sensitivity to type I interferon initiates productive infection in most cases. We hypothesized that particularly active accessory protein(s) may confer T/F viruses with a selective advantage in establishing HIV infection. Thus, we tested vpu, vif and nef alleles from six T/F and six chronic (CC) viruses in assays for 9 immune evasion activities involving the counteraction of interferon-stimulated genes and modulation of ligands known to activate innate immune cells. All functions were highly conserved with no significant differences between T/F and CC viruses, suggesting that these accessory protein functions are important throughout the course of infection. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  12. Immune responses elicited against rotavirus middle layer protein VP6 inhibit viral replication in vitro and in vivo

    Science.gov (United States)

    Lappalainen, Suvi; Pastor, Ana Ruth; Tamminen, Kirsi; López-Guerrero, Vanessa; Esquivel-Guadarrama, Fernando; Palomares, Laura A; Vesikari, Timo; Blazevic, Vesna

    2014-01-01

    Rotavirus (RV) is a common cause of severe gastroenteritis (GE) in children worldwide. Live oral RV vaccines protect against severe RVGE, but the immune correlates of protection are not yet clearly defined. Inner capsid VP6 protein is a highly conserved, abundant, and immunogenic RV protein, and VP6-specific mucosal antibodies, especially IgA, have been implicated to protect against viral challenge in mice. In the present study systemic and mucosal IgG and IgA responses were induced by immunizing BALB/c mice intranasally with a combination of recombinant RV VP6 protein (subgroup II [SGII]) and norovirus (NoV) virus-like particles (VLPs) used in a candidate vaccine. Following immunization mice were challenged orally with murine RV strain EDIMwt (SG non-I-non-II, G3P10[16]). In order to determine neutralizing activity of fecal samples, sera, and vaginal washes (VW) against human Wa RV (SGII, G1P1A[8]) and rhesus RV (SGI, G3P5B[3]), the RV antigen production was measured with an ELISA-based antigen reduction neutralization assay. Only VWs of immunized mice inhibited replication of both RVs, indicating heterotypic protection of induced antibodies. IgA antibody depletion and blocking experiments using recombinant VP6 confirmed that neutralization was mediated by anti-VP6 IgA antibodies. Most importantly, after the RV challenge significant reduction in viral shedding was observed in feces of immunized mice. These results suggest a significant role for mucosal RV VP6-specific IgA for the inhibition of RV replication in vitro and in vivo. In addition, these results underline the importance of non-serotype-specific immunity induced by the conserved subgroup-specific RV antigen VP6 in clearance of RV infection. PMID:25424814

  13. Functions of innate and acquired immune system are reduced in domestic pigeons (Columba livia domestica) given a low protein diet

    Science.gov (United States)

    Mabuchi, Yuko; Frankel, Theresa L.

    2016-01-01

    Racing pigeons are exposed to and act as carriers of diseases. Dietary protein requirement for their maintenance has not been determined experimentally despite their being domesticated for over 7000 years. A maintenance nitrogen (protein) requirement (MNR) for pigeons was determined in a balance study using diets containing 6, 10 and 14% crude protein (CP). Then, the effects of feeding the diets were investigated to determine whether they were adequate to sustain innate and acquired immune functions. Nitrogen intake from the 6% CP diet was sufficient to maintain nitrogen balance and body weight in pigeons. However, the immune functions of phagocytosis, oxidative burst and lymphocyte proliferation in pigeons fed this diet were reduced compared with those fed 10 and 14% CP diets. Pigeons given the 6 and 10% CP diets had lower antibody titres following inoculation against Newcastle disease (ND) than those on the 14% CP diet. A confounding factor found on autopsy was the presence of intestinal parasites in some of the pigeons given the 6 and 10% CP diets; however, none of the pigeons used to measure MNR or acquired immunity to ND were infested with parasites. In conclusion, neither the 6 nor 10% CP diets adequately sustained acquired immune function of pigeons. PMID:27069640

  14. Passive immunization to outer membrane proteins MLP and PAL does not protect mice from sepsis.

    Science.gov (United States)

    Valentine, Catherine H; Hellman, Judith; Beasley-Topliffe, Laura K; Bagchi, Aranya; Warren, H Shaw

    2006-01-01

    Multiple older studies report that immunoglobulin directed to rough mutant bacteria, such as E. coli J5, provides broad protection against challenge with heterologous strains of Gram-negative bacteria. This protection was initially believed to occur through binding of immunoglobulin to bacterial lipopolysaccharide (LPS). However, hundreds of millions of dollars have been invested in attempting to develop clinically-effective anti-LPS monoclonal antibodies without success, and no study has shown that IgG from this antiserum binds LPS. Identification of the protective mechanism would facilitate development of broadly protective human monoclonal antibodies for treating sepsis. IgG from this antiserum binds 2 bacterial outer membrane proteins: murein lipoprotein (MLP) and peptidoglycan-associated lipoprotein (PAL). Both of these outer membrane proteins are highly conserved, have lipid domains that are anchored in the bacterial membrane, are shed from bacteria in blebs together with LPS, and activate cells through Toll-like receptor 2. Our goal in the current work was to determine if passive immunization directed to MLP and PAL protects mice from Gram-negative sepsis. Neither monoclonal nor polyclonal IgG directed to MLP or PAL conferred survival protection in 3 different models of sepsis: cecal ligation and puncture, an infected burn model, and an infected fibrin clot model mimicking peritonitis. Our results are not supportive of the hypothesis that either anti-MLP or anti-PAL IgG are the protective antibodies in the previously described anti-rough mutant bacterial antisera. These studies suggest that a different mechanism of protection is involved.

  15. Immunization of mice by Hollow Mesoporous Silica Nanoparticles as carriers of Porcine Circovirus Type 2 ORF2 Protein

    Directory of Open Access Journals (Sweden)

    Guo Hui-Chen

    2012-06-01

    Full Text Available Abstract Backgroud Porcine circovirus type 2 (PCV2 is a primary etiological agent of post-weaning multi-systemic wasting syndrome (PMWS, which is a disease of increasing importance to the pig industry worldwide. Hollow mesoporous silica nanoparticles (HMSNs have gained increasing interest for use in vaccines. Methods To study the potential of HMSNs for use as a protein delivery system or vaccine carriers. HMSNs were synthesized by a sol–gel/emulsion(oil-in-water/ethanol method, purified PCV2 GST-ORF2-E protein was loaded into HMSNs, and the resulting HMSN/protein mixture was injected into mice. The uptake and release profiles of protein by HMSNs in vitro were investigated. PCV2 GST-ORF2-E specific antibodies and secretion of IFN-γ were detected by enzyme-linked immunosorbent assays, spleen lymphocyte proliferation was measured by the MTS method, and the percentage of CD4+ and CD8+ were determined by flow cytometry. Results HMSNs were found to yield better binding capacities and delivery profiles of proteins; the specific immune response induced by PCV2 GST-ORF2-E was maintained for a relatively long period of time after immunization with the HMSN/protein complex. Conclusion The findings suggest that HMSNs are good protein carriers and have high potential for use in future applications in therapeutic drug delivery.

  16. Cancer associated aberrant protein o-glycosylation can modify antigen processing and immune response

    DEFF Research Database (Denmark)

    Madsen, Caroline B; Petersen, Cecilie; Lavrsen, Kirstine

    2012-01-01

    Aberrant glycosylation of mucins and other extracellular proteins is an important event in carcinogenesis and the resulting cancer associated glycans have been suggested as targets in cancer immunotherapy. We assessed the role of O-linked GalNAc glycosylation on antigen uptake, processing......, and presentation on MHC class I and II molecules. The effect of GalNAc O-glycosylation was monitored with a model system based on ovalbumin (OVA)-MUC1 fusion peptides (+/- glycosylation) loaded onto dendritic cells co-cultured with IL-2 secreting OVA peptide-specific T cell hybridomas. To evaluate the in vivo...... response to a cancer related tumor antigen, Balb/c or B6.Cg(CB)-Tg(HLA-A/H2-D)2Enge/J (HLA-A2 transgenic) mice were immunized with a non-glycosylated or GalNAc-glycosylated MUC1 derived peptide followed by comparison of T cell proliferation, IFN-¿ release, and antibody induction. Gal...

  17. The C-type lectin-like domain containing proteins Clec-39 and Clec-49 are crucial for Caenorhabditis elegans immunity against Serratia marcescens infection.

    Science.gov (United States)

    Miltsch, S M; Seeberger, P H; Lepenies, B

    2014-07-01

    Caenorhabditis elegans exhibits protective immunity against a variety of fungal and bacterial pathogens. Since C. elegans lacks an adaptive immune system, pathogen recognition is mediated entirely by innate immunity. To date, little is known about the involvement of pattern recognition receptors (PRRs) in pathogen sensing as part of the C. elegans immunity. C-type lectin-like domain (CTLD) containing proteins represent a superfamily of PRRs. A large number of genes encoding for CTLD proteins are present in the C. elegans genome, however the role of CTLD proteins in bacterial recognition and antibacterial immunity has not yet been determined. In this study, we investigated the function of selected C. elegans CTLD proteins during infection with the Gram-negative bacterium Serratia marcescens. Wild-type and CTLD gene-deficient C. elegans strains were compared in their susceptibility to S. marcescens infection. Interestingly, survival and egg laying were significantly reduced in strains deficient for clec-39 and clec-49 indicating a role for both CTLD proteins in C. elegans immune defense against bacteria as evidenced by using S. marcescens infection. Binding studies with recombinantly expressed Clec-39-Fc and Clec-49-Fc fusion proteins revealed that both CTLD proteins recognized live bacteria in a Ca(2+)-independent manner. This study provides insight into the role of CTLD proteins in C. elegans immunity and demonstrates their function during bacterial infection. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Protein malnutrition impairs the immune response and influences the severity of infection in a hamster model of chronic visceral leishmaniasis.

    Directory of Open Access Journals (Sweden)

    Eugenia Carrillo

    Full Text Available Leishmaniasis remains one of the world's most devastating neglected tropical diseases. It mainly affects developing countries, where it often co-exists with chronic malnutrition, one of the main risk factors for developing the disease. Few studies have been published, however, on the relationship between leishmaniasis progression and malnutrition. The present paper reports the influence of protein malnutrition on the immune response and visceral disease development in adult hamsters infected with Leishmania infantum fed either standard or low protein diets. The low protein diet induced severe malnutrition in these animals, and upon infection with L. infantum 33% had severe visceral leishmaniasis compared to only 8% of animals fed the standard diet. The infected, malnourished animals showed notable leukocyte depletion, mild specific antibody responses, impairment of lymphoproliferation, presence of parasites in blood (16.67% of the hamsters and significant increase of the splenic parasite burden. Animals fed standard diet suffered agranulocytosis and monocytopenia, but showed stronger specific immune responses and had lower parasite loads than their malnourished counterparts. The present results show that protein malnutrition promotes visceral leishmaniasis and provide clues regarding the mechanisms underlying the impairment of the immune system.

  19. Mycoplasma hyopneumoniae and Mycoplasma flocculare differential domains from orthologous surface proteins induce distinct cellular immune responses in mice.

    Science.gov (United States)

    Leal, Fernanda Munhoz Dos Anjos; Virginio, Veridiana Gomes; Martello, Carolina Lumertz; Paes, Jéssica Andrade; Borges, Thiago J; Jaeger, Natália; Bonorino, Cristina; Ferreira, Henrique Bunselmeyer

    2016-07-15

    Mycoplasma hyopneumoniae and Mycoplasma flocculare are two genetically close species found in the swine respiratory tract. Despite their similarities, while M. hyopneumoniae is the causative agent of porcine enzootic pneumonia, M. flocculare is a commensal bacterium. Genomic and transcriptional comparative analyses so far failed to explain the difference in pathogenicity between these two species. We then hypothesized that such difference might be, at least in part, explained by amino acid sequence and immunological or functional differences between ortholog surface proteins. In line with that, it was verified that approximately 85% of the ortholog surface proteins from M. hyopneumoniae 7448 and M. flocculare present one or more differential domains. To experimentally assess possible immunological implications of this kind of difference, the extracellular differential domains from one pair of orthologous surface proteins (MHP7448_0612, from M. hyopneumoniae, and MF_00357, from M. flocculare) were expressed in E. coli and used to immunize mice. The recombinant polypeptides (rMHP61267-169 and rMF35767-196, respectively) induced distinct cellular immune responses. While, rMHP61267-169 induced both Th1 and Th2 responses, rMF35767-196 induced just an early pro-inflammatory response. These results indicate that immunological properties determined by differential domains in orthologous surface protein might play a role in pathogenicity, contributing to elicit specific and differential immune responses against each species. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Protective immunity provided by HLA-A2 epitopes for fusion and hemagglutinin proteins of measles virus

    International Nuclear Information System (INIS)

    Oh, Sang Kon; Stegman, Brian; Pendleton, C. David; Ota, Martin O.; Pan, C.-H.; Griffin, Diane E.; Burke, Donald S.; Berzofsky, Jay A.

    2006-01-01

    Natural infection and vaccination with a live-attenuated measles virus (MV) induce CD8 + T-cell-mediated immune responses that may play a central role in controlling MV infection. In this study, we show that newly identified human HLA-A2 epitopes from MV hemagglutinin (H) and fusion (F) proteins induced protective immunity in HLA-A2 transgenic mice challenged with recombinant vaccinia viruses expressing F or H protein. HLA-A2 epitopes were predicted and synthesized. Five and four peptides from H and F, respectively, bound to HLA-A2 molecules in a T2-binding assay, and four from H and two from F could induce peptide-specific CD8 + T cell responses in HLA-A2 transgenic mice. Further experiments proved that three peptides from H (H9-567, H10-250, and H10-516) and one from F protein (F9-57) were endogenously processed and presented on HLA-A2 molecules. All peptides tested in this study are common to 5 different strains of MV including Edmonston. In both A2K b and HHD-2 mice, the identified peptide epitopes induced protective immunity against recombinant vaccinia viruses expressing H or F. Because F and H proteins induce neutralizing antibodies, they are major components of new vaccine strategies, and therefore data from this study will contribute to the development of new vaccines against MV infection

  1. Induction of neutralizing antibodies specific for the envelope proteins of the koala retrovirus by immunization with recombinant proteins or with DNA.

    Science.gov (United States)

    Fiebig, Uwe; Dieckhoff, Britta; Wurzbacher, Christian; Möller, Annekathrin; Kurth, Reinhard; Denner, Joachim

    2015-04-30

    The koala retrovirus (KoRV) is the result of a transspecies transmission of a gammaretrovirus with fatal consequences for the new host. Like many retroviruses, KoRV induces lymphoma, leukemia and an immunodeficiency that is associated with opportunistic infections in the virus-infected animals. We recently reported the induction of neutralizing antibodies by immunization with the recombinant ectodomain of the transmembrane envelope protein p15E of KoRV. Since the neutralization titers of the p15E-specific sera were only moderate, we investigated the use of the surface envelope protein gp70 to induce neutralizing antibodies. We immunized rats and goats with the recombinant gp70 protein of the KoRV, an unglycosylated protein of 52kD (rgp70/p52) or with the corresponding DNA. In parallel we immunized with recombinant rp15E or with a combination of rp15E and rgp70/p52. In all cases binding and neutralizing antibodies were induced. The gp70-specific sera had titers of neutralizing antibodies that were 15-fold higher than the p15E-specific sera. Combining rp15E and rgp70/p52 did not significantly increase neutralizing titers compared to rgp70/p52 alone. High titers of neutralizing antibodies specific for gp70 were also induced by immunization with DNA. Since KoRV and PERV are closely related, we investigated cross-neutralization of the antisera. The antisera against p15E and gp70 of PERV and KoRV inhibited infection by both viruses. The envelope proteins of the KoRV may therefore form the basis of an effective preventive vaccine to protect uninfected koalas from infection and possibly an immunotherapeutic treatment for those already infected.

  2. The Verticillium-specific protein VdSCP7 localizes to the plant nucleus and modulates immunity to fungal infections.

    Science.gov (United States)

    Zhang, Lisha; Ni, Hao; Du, Xuan; Wang, Sheng; Ma, Xiao-Wei; Nürnberger, Thorsten; Guo, Hui-Shan; Hua, Chenlei

    2017-07-01

    Fungal pathogens secrete effector proteins to suppress plant basal defense for successful colonization. Resistant plants, however, can recognize effectors by cognate R proteins to induce effector-triggered immunity (ETI). By analyzing secretomes of the vascular fungal pathogen Verticillium dahliae, we identified a novel secreted protein VdSCP7 that targets the plant nucleus. The green fluorescent protein (GFP)-tagged VdSCP7 gene with either a mutated nuclear localization signal motif or with additional nuclear export signal was transiently expressed in Nicotiana benthamiana, and investigated for induction of plant immunity. The role of VdSCP7 in V. dahliae pathogenicity was characterized by gene knockout and complementation, and GFP labeling. Expression of the VdSCP7 gene in N. benthamiana activated both salicylic acid and jasmonate signaling, and altered the plant's susceptibility to the pathogens Botrytis cinerea and Phytophthora capsici. The immune response activated by VdSCP7 was highly dependent on its initial extracellular secretion and subsequent nuclear localization in plants. Knockout of the VdSCP7 gene significantly enhanced V. dahliae aggressiveness on cotton. GFP-labeled VdSCP7 is secreted by V. dahliae and accumulates in the plant nucleus. We conclude that VdSCP7 is a novel effector protein that targets the host nucleus to modulate plant immunity, and suggest that plants can recognize VdSCP7 to activate ETI during fungal infection. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  3. IgG immune responses to different proteins of Helicobacter Pylori as defined by immunoblot assay

    Directory of Open Access Journals (Sweden)

    Raeiszadeh M

    2000-09-01

    Full Text Available Helicobacter pylori (H.pylori is an etiologic factor for chronic gastritis and peptic ulcers. Serological testing of H.pylori infection is common in Iran, as other parts of the world. There are geographical variations in the humoral immune response to various H. pylori strains in different parts of the worl. We studied the immunogenic proteins of H.pylori by means of an Immunoblot assay with antigens of H.pylori strains isolated in Iran. Sera of 64 patients suffering from dyspepsia were analyzed to determine antibodlies which were good marker of infection and the antibody patterns associated with peptic ulcer.54 out of 64 dyspeptic patients were infected by H. pylori based on positive culture or positive results of both rapid urease test and direct examination. 14 out of fity-four had peptic ulcers and the rest were catagoriied as patients with non-ulcer dyspepsia. Some of them had multiple erosions in the gut or deodenum. Tweny –two major bands were identified by immunoblot. Of these, IgG antibodies against 10 protients, and they produced immunoreative bands at 14, 16, 22, 26, 32 , 32, 44, 87, 92, 120 Kda. Antibody patterns were not identical in the patients. The presence of at least one band at 14, 16, 22, 26, 32, 35Kda was the best marker of infection(sensitivity, 90% and specificity, 80% Major serological cross reactions were found at moderate molecular weight bands (50, 52, 54, 60, 66 KDa. The presence of at least one band at 14, 16, 22, 26, 32, 35Kda was the best marker of infection (sensitivity, 90% and specificity, 80%. Major serological crossreactions were found at moderate molerate molecular weight bands (50, 52, 54, 60, 66 KDa. The presence of antibodies to 120 Kda protein (Cag A and 87 Kda Protein (Vac A were not associated with the presence of peptic ulcers. These were in contradiction to results obtained across Europe and U.S but in agreement with Asian studies. However the presence of at least one band at either 32 or 35 Kda was

  4. G-protein coupled receptor signaling architecture of mammalian immune cells.

    Directory of Open Access Journals (Sweden)

    Natalia Polouliakh

    Full Text Available A series of recent studies on large-scale networks of signaling and metabolic systems revealed that a certain network structure often called "bow-tie network" are observed. In signaling systems, bow-tie network takes a form with diverse and redundant inputs and outputs connected via a small numbers of core molecules. While arguments have been made that such network architecture enhances robustness and evolvability of biological systems, its functional role at a cellular level remains obscure. A hypothesis was proposed that such a network function as a stimuli-reaction classifier where dynamics of core molecules dictate downstream transcriptional activities, hence physiological responses against stimuli. In this study, we examined whether such hypothesis can be verified using experimental data from Alliance for Cellular Signaling (AfCS that comprehensively measured GPCR related ligands response for B-cell and macrophage. In a GPCR signaling system, cAMP and Ca2+ act as core molecules. Stimuli-response for 32 ligands to B-Cells and 23 ligands to macrophages has been measured. We found that ligands with correlated changes of cAMP and Ca2+ tend to cluster closely together within the hyperspaces of both cell types and they induced genes involved in the same cellular processes. It was found that ligands inducing cAMP synthesis activate genes involved in cell growth and proliferation; cAMP and Ca2+ molecules that increased together form a feedback loop and induce immune cells to migrate and adhere together. In contrast, ligands without a core molecules response are scattered throughout the hyperspace and do not share clusters. G-protein coupling receptors together with immune response specific receptors were found in cAMP and Ca2+ activated clusters. Analyses have been done on the original software applicable for discovering 'bow-tie' network architectures within the complex network of intracellular signaling where ab initio clustering has been

  5. G-protein coupled receptor signaling architecture of mammalian immune cells.

    Science.gov (United States)

    Polouliakh, Natalia; Nock, Richard; Nielsen, Frank; Kitano, Hiroaki

    2009-01-01

    A series of recent studies on large-scale networks of signaling and metabolic systems revealed that a certain network structure often called "bow-tie network" are observed. In signaling systems, bow-tie network takes a form with diverse and redundant inputs and outputs connected via a small numbers of core molecules. While arguments have been made that such network architecture enhances robustness and evolvability of biological systems, its functional role at a cellular level remains obscure. A hypothesis was proposed that such a network function as a stimuli-reaction classifier where dynamics of core molecules dictate downstream transcriptional activities, hence physiological responses against stimuli. In this study, we examined whether such hypothesis can be verified using experimental data from Alliance for Cellular Signaling (AfCS) that comprehensively measured GPCR related ligands response for B-cell and macrophage. In a GPCR signaling system, cAMP and Ca2+ act as core molecules. Stimuli-response for 32 ligands to B-Cells and 23 ligands to macrophages has been measured. We found that ligands with correlated changes of cAMP and Ca2+ tend to cluster closely together within the hyperspaces of both cell types and they induced genes involved in the same cellular processes. It was found that ligands inducing cAMP synthesis activate genes involved in cell growth and proliferation; cAMP and Ca2+ molecules that increased together form a feedback loop and induce immune cells to migrate and adhere together. In contrast, ligands without a core molecules response are scattered throughout the hyperspace and do not share clusters. G-protein coupling receptors together with immune response specific receptors were found in cAMP and Ca2+ activated clusters. Analyses have been done on the original software applicable for discovering 'bow-tie' network architectures within the complex network of intracellular signaling where ab initio clustering has been implemented as well

  6. Spodoptera frugiperda X-tox protein, an immune related defensin rosary, has lost the function of ancestral defensins.

    Directory of Open Access Journals (Sweden)

    Delphine Destoumieux-Garzón

    Full Text Available BACKGROUND: X-tox proteins are a family of immune-related proteins only found in Lepidoptera and characterized by imperfectly conserved tandem repeats of several defensin-like motifs. Previous phylogenetic analysis of X-tox genes supported the hypothesis that X-tox have evolved from defensins in a lineage-specific gene evolution restricted to Lepidoptera. In this paper, we performed a protein study in which we asked whether X-tox proteins have conserved the antimicrobial functions of their ancestral defensins and have evolved as defensin reservoirs. METHODOLOGY/PRINCIPAL FINDINGS: We followed the outcome of Spod-11-tox, an X-tox protein characterized in Spodoptera frugiperda, in bacteria-challenged larvae using both immunochemistry and antimicrobial assays. Three hours post infection, the Spod-11-tox protein was expressed in 80% of the two main classes of circulating hemocytes (granulocytes and plasmatocytes. Located in secretory granules of hemocytes, Spod-11-tox was never observed in contact with microorganisms entrapped within phagolyzosomes showing that Spod-11-tox is not involved in intracellular pathogen killing. In fact, the Spod-11-tox protein was found to be secreted into the hemolymph of experimentally challenged larvae. In order to determine antimicrobial properties of the Spod-11-tox protein, it was consequently fractionated according to a protocol frequently used for antimicrobial peptide purification. Over the course of purification, the anti-Spod-11-tox immunoreactivity was found to be dissociated from the antimicrobial activity. This indicates that Spod-11-tox is not processed into bioactive defensins in response to a microbial challenge. CONCLUSIONS/SIGNIFICANCE: Altogether, our results show that X-tox proteins have not evolved as defensin reservoirs and have lost the antimicrobial properties of the ancestral insect defensins. The lepidopteran X-tox protein family will provide a valuable and tractable model to improve our

  7. Spodoptera frugiperda X-tox protein, an immune related defensin rosary, has lost the function of ancestral defensins.

    Science.gov (United States)

    Destoumieux-Garzón, Delphine; Brehelin, Michel; Bulet, Philippe; Boublik, Yvan; Girard, Pierre-Alain; Baghdiguian, Stephen; Zumbihl, Robert; Escoubas, Jean-Michel

    2009-08-27

    X-tox proteins are a family of immune-related proteins only found in Lepidoptera and characterized by imperfectly conserved tandem repeats of several defensin-like motifs. Previous phylogenetic analysis of X-tox genes supported the hypothesis that X-tox have evolved from defensins in a lineage-specific gene evolution restricted to Lepidoptera. In this paper, we performed a protein study in which we asked whether X-tox proteins have conserved the antimicrobial functions of their ancestral defensins and have evolved as defensin reservoirs. We followed the outcome of Spod-11-tox, an X-tox protein characterized in Spodoptera frugiperda, in bacteria-challenged larvae using both immunochemistry and antimicrobial assays. Three hours post infection, the Spod-11-tox protein was expressed in 80% of the two main classes of circulating hemocytes (granulocytes and plasmatocytes). Located in secretory granules of hemocytes, Spod-11-tox was never observed in contact with microorganisms entrapped within phagolyzosomes showing that Spod-11-tox is not involved in intracellular pathogen killing. In fact, the Spod-11-tox protein was found to be secreted into the hemolymph of experimentally challenged larvae. In order to determine antimicrobial properties of the Spod-11-tox protein, it was consequently fractionated according to a protocol frequently used for antimicrobial peptide purification. Over the course of purification, the anti-Spod-11-tox immunoreactivity was found to be dissociated from the antimicrobial activity. This indicates that Spod-11-tox is not processed into bioactive defensins in response to a microbial challenge. Altogether, our results show that X-tox proteins have not evolved as defensin reservoirs and have lost the antimicrobial properties of the ancestral insect defensins. The lepidopteran X-tox protein family will provide a valuable and tractable model to improve our knowledge on the molecular evolution of defensins, a class of innate immune effectors largely

  8. Neural Network Enhanced Structure Determination of Osteoporosis, Immune System, and Radiation Repair Proteins, Phase I

    Data.gov (United States)

    National Aeronautics and Space Administration — The proposed innovation will utilize self learning neural network technology to determine the structure of osteoporosis, immune system disease, and excess radiation...

  9. Interference of outer membrane protein PalA with protective immunity against Actinobacillus pleuropneumoniae infections in vaccinated pigs.

    Science.gov (United States)

    van den Bosch, Han; Frey, Joachim

    2003-09-08

    The role of antibodies to the outer membrane protein PalA of Actinobacillus pleuropneumoniae in protective immunity was studied in pigs vaccinated with purified PalA alone and PalA in combination with toxoids of the RTX toxins ApxI and ApxII using an established challenge model with the virulent serotype 1 of A. pleuropneumoniae. Pigs that developed antibody titers against PalA after immunization were more significantly affected by challenge with A. pleuropneumoniae serotype 1. Following challenge, pigs that were immunized with PalA showed more severe respiratory symptoms, had a higher mortality rate and died faster. They also displayed much more severe lung lesions after necropsy than animals not immunized with PalA. Pigs that were immunized with toxoids of the two cytotoxins ApxI and ApxII were protected against challenge with A. pleuropneumoniae. In contrast, the protective efficacy of the ApxI and ApxII vaccine was completely lost when it was supplemented with PalA. Hence, antibodies induced against the outer membrane protein PalA of A. pleuropneumoniae aggravated the consequences of infection and counteracted the protective effect of anti-ApxI and anti-ApxII antibodies. Due to the high similarity between protein analogues of PalA from various bacteria of the Pasteurellaceae family such as P6 of Haemophilus influenzae or 16kDa Omp of Pasteurella multocida, this deleterious effect of PalA in vaccination should be taken into consideration in the development of vaccines against infections with other Pasteurellaceae.

  10. Effects of the RNA-binding protein, KSRP, on innate immune response against Helicobacter pylori infection in mice.

    Science.gov (United States)

    Li, Ningzhe; Cao, Mei; Yi, Sijun; Cheng, Juan; Wang, Lei; Tao, Yuwei; Wu, Daoyan; Peng, Jingshan; Zhang, Mao; Qi, Panpan; Zhao, Jian

    2018-01-08

    Helicobacter pylori (H. pylori) contributes to various gastric diseases such as chronic gastritis, gastric ulcer, and gastric carcinoma. Host innate immune response against the pathogen plays a significant role in elimination of pathogen infection. Importantly, pathogen elimination is closely related to numerous inflammatory-related genes that participate in complex biological response of cells to harmful stimuli. Here we studied effects of the KH-type splicing regulatory protein (KSRP), a RNA-binding protein, on innate immune response against H. pylori infection. We found that H. pylori infection downregulated KSRP expression directly, and that KSRP overexpression repressed upregulation of CXCL-2 expression induced by H. pylori and facilitated H. pylori proliferation in vitro. Similarly, KSRP overexpression in H. pylori mice also facilitated H. pylori proliferation and colonization, and induced more severe gastric mucosal damage. Intriguingly, CXCL-2 and HMOX-1 were upregulated in H. pylori infected mice after KSRP overexpression. This difference in expression of these genes implicated that KSRP was closely associated with and directly participated in the innate immune response against H. pylori. These results were beneficial for understanding the in vivo function of KSRP on innate immune response against pathogen infection. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. The Mycoplasma hyopneumoniae recombinant heat shock protein P42 induces an immune response in pigs under field conditions.

    Science.gov (United States)

    Jorge, Sérgio; de Oliveira, Natasha Rodrigues; Marchioro, Silvana Beutinger; Fisch, Andressa; Gomes, Charles Klazer; Hartleben, Cláudia Pinho; Conceição, Fabricio Rochedo; Dellagostin, Odir Antonio

    2014-09-01

    Enzootic pneumonia (EP), resulting from Mycoplasma hyopneumoniae infection is one of the most prevalent diseases in pigs and is a major cause of economic losses to the swine industry worldwide. EP is often controlled by vaccination with inactivated, adjuvanted whole-cell bacterin. However, these bacterins provide only partial protection and do not prevent M. hyopneumoniae colonization. Attempts to develop vaccines that are more efficient have made use of the recombinant DNA technology. The objective of this study was to assess the potential of recombinant M. hyopneumoniae heat shock protein P42 in vaccine preparations against EP, using piglets housed under field conditions in a M. hyopneumoniae-positive farm. The cellular and humoral immune responses were elicited after a single intramuscular inoculation of rP42 in an oil-based adjuvant, or in conjunction with whole-cell vaccine preparation. The production of INF-γ and IL-10 cytokines was quantified in the supernatant of the cultured mononuclear cells. The rP42 emulsified in oil-based adjuvant was able to trigger a strong humoral immune response. Further, it induced a cellular immune response, accompanied by the production of antibodies that reacted with the native M. hyopneumoniae protein. The rP42 mediated induction of cellular and humoral immune response in the host suggests that rP42 emulsified in an oil-based adjuvant holds promise as an effective recombinant subunit vaccine against EP. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Herpes simplex virus 1 infection dampens the immediate early antiviral innate immunity signaling from peroxisomes by tegument protein VP16.

    Science.gov (United States)

    Zheng, Chunfu; Su, Chenhe

    2017-02-21

    Herpes simplex virus 1 (HSV-1) is an archetypal member of the alphaherpesvirus subfamily with a large genome encoding over 80 proteins, many of which play a critical role in virus-host interactions and immune modulation. Upon viral infections, the host cells activate innate immune responses to restrict their replications. Peroxisomes, which have long been defined to regulate metabolic activities, are reported to be important signaling platforms for antiviral innate immunity. It has been verified that signaling from peroxisomal MAVS (MAVS-Pex) triggers a rapid interferon (IFN) independent IFN-stimulated genes (ISGs) production against invading pathogens. However, little is known about the interaction between DNA viruses such as HSV-1 and the MAVS-Pex mediated signaling. HSV-1 could activate the MAVS-Pex signaling pathway at a low multiplicity of infection (MOI), while infection at a high MOI dampens MAVS-Pex induced immediately early ISGs production. A high-throughput screen assay reveals that HSV-1 tegument protein VP16 inhibits the immediate early ISGs expression downstream of MAVS-Pex signaling. Moreover, the expression of ISGs was recovered when VP16 was knockdown with its specific short hairpin RNA. HSV-1 blocks MAVS-Pex mediated early ISGs production through VP16 to dampen the immediate early antiviral innate immunity signaling from peroxisomes.

  13. A Small Cysteine-Rich Protein from the Asian Soybean Rust Fungus, Phakopsora pachyrhizi, Suppresses Plant Immunity.

    Directory of Open Access Journals (Sweden)

    Mingsheng Qi

    2016-09-01

    Full Text Available The Asian soybean rust fungus, Phakopsora pachyrhizi, is an obligate biotrophic pathogen causing severe soybean disease epidemics. Molecular mechanisms by which P. pachyrhizi and other rust fungi interact with their host plants are poorly understood. The genomes of all rust fungi encode many small, secreted cysteine-rich proteins (SSCRP. While these proteins are thought to function within the host, their roles are completely unknown. Here, we present the characterization of P. pachyrhizi effector candidate 23 (PpEC23, a SSCRP that we show to suppress plant immunity. Furthermore, we show that PpEC23 interacts with soybean transcription factor GmSPL12l and that soybean plants in which GmSPL12l is silenced have constitutively active immunity, thereby identifying GmSPL12l as a negative regulator of soybean defenses. Collectively, our data present evidence for a virulence function of a rust SSCRP and suggest that PpEC23 is able to suppress soybean immune responses and physically interact with soybean transcription factor GmSPL12l, a negative immune regulator.

  14. A Small Cysteine-Rich Protein from the Asian Soybean Rust Fungus, Phakopsora pachyrhizi, Suppresses Plant Immunity.

    Science.gov (United States)

    Qi, Mingsheng; Link, Tobias I; Müller, Manuel; Hirschburger, Daniela; Pudake, Ramesh N; Pedley, Kerry F; Braun, Edward; Voegele, Ralf T; Baum, Thomas J; Whitham, Steven A

    2016-09-01

    The Asian soybean rust fungus, Phakopsora pachyrhizi, is an obligate biotrophic pathogen causing severe soybean disease epidemics. Molecular mechanisms by which P. pachyrhizi and other rust fungi interact with their host plants are poorly understood. The genomes of all rust fungi encode many small, secreted cysteine-rich proteins (SSCRP). While these proteins are thought to function within the host, their roles are completely unknown. Here, we present the characterization of P. pachyrhizi effector candidate 23 (PpEC23), a SSCRP that we show to suppress plant immunity. Furthermore, we show that PpEC23 interacts with soybean transcription factor GmSPL12l and that soybean plants in which GmSPL12l is silenced have constitutively active immunity, thereby identifying GmSPL12l as a negative regulator of soybean defenses. Collectively, our data present evidence for a virulence function of a rust SSCRP and suggest that PpEC23 is able to suppress soybean immune responses and physically interact with soybean transcription factor GmSPL12l, a negative immune regulator.

  15. PGRP-LB is a maternally transmitted immune milk protein that influences symbiosis and parasitism in tsetse’s offspring

    Science.gov (United States)

    Wang, Jingwen; Aksoy, Serap

    2012-01-01

    Beneficial microbe functions range from host dietary supplementation to development and maintenance of host immune system. In mammals, newborn progeny are quickly colonized with a symbiotic fauna that is provisioned in mother’s milk and that closely resembles that of the parent. Tsetse fly (Diptera: Glossinidae) also depends on the obligate symbiont Wigglesworthia for nutritional supplementation, optimal fecundity, and immune system development. Tsetse progeny develop one at a time in an intrauterine environment and receive nourishment and symbionts in mother’s milk. We show that the host Peptidoglycan Recognition Protein (PGRP-LB) is expressed only in adults and is a major component of the milk that nourishes the developing progeny. The amidase activity associated with PGRP-LB may scavenge the symbiotic peptidoglycan and prevent the induction of tsetse's Immune Deficiency pathway that otherwise can damage the symbionts. Reduction of PGRP-LB experimentally diminishes female fecundity and damages Wigglesworthia in the milk through induction of antimicrobial peptides, including Attacin. Larvae that receive less maternal PGRP-LB give rise to adults with fewer Wigglesworthia and hyperimmune responses. Such adults also suffer dysregulated immunity, as indicated by the presence of higher trypanosome densities in parasitized adults. We show that recPGRP-LB has antimicrobial and antitrypanosomal activities that may regulate symbiosis and impact immunity. Thus, PGRP-LB plays a pivotal role in tsetse’s fitness by protecting symbiosis against host-inflicted damage during development and by controlling parasite infections in adults that can otherwise reduce host fecundity. PMID:22689989

  16. PGRP-LB is a maternally transmitted immune milk protein that influences symbiosis and parasitism in tsetse's offspring.

    Science.gov (United States)

    Wang, Jingwen; Aksoy, Serap

    2012-06-26

    Beneficial microbe functions range from host dietary supplementation to development and maintenance of host immune system. In mammals, newborn progeny are quickly colonized with a symbiotic fauna that is provisioned in mother's milk and that closely resembles that of the parent. Tsetse fly (Diptera: Glossinidae) also depends on the obligate symbiont Wigglesworthia for nutritional supplementation, optimal fecundity, and immune system development. Tsetse progeny develop one at a time in an intrauterine environment and receive nourishment and symbionts in mother's milk. We show that the host Peptidoglycan Recognition Protein (PGRP-LB) is expressed only in adults and is a major component of the milk that nourishes the developing progeny. The amidase activity associated with PGRP-LB may scavenge the symbiotic peptidoglycan and prevent the induction of tsetse's Immune Deficiency pathway that otherwise can damage the symbionts. Reduction of PGRP-LB experimentally diminishes female fecundity and damages Wigglesworthia in the milk through induction of antimicrobial peptides, including Attacin. Larvae that receive less maternal PGRP-LB give rise to adults with fewer Wigglesworthia and hyperimmune responses. Such adults also suffer dysregulated immunity, as indicated by the presence of higher trypanosome densities in parasitized adults. We show that recPGRP-LB has antimicrobial and antitrypanosomal activities that may regulate symbiosis and impact immunity. Thus, PGRP-LB plays a pivotal role in tsetse's fitness by protecting symbiosis against host-inflicted damage during development and by controlling parasite infections in adults that can otherwise reduce host fecundity.

  17. Thirty Minutes of Hypobaric Hypoxia Provokes Alterations of Immune Response, Haemostasis, and Metabolism Proteins in Human Serum

    Directory of Open Access Journals (Sweden)

    Jochen Hinkelbein

    2017-08-01

    Full Text Available Hypobaric hypoxia (HH during airline travel induces several (patho- physiological reactions in the human body. Whereas severe hypoxia is investigated thoroughly, very little is known about effects of moderate or short-term hypoxia, e.g. during airline flights. The aim of the present study was to analyse changes in serum protein expression and activation of signalling cascades in human volunteers staying for 30 min in a simulated altitude equivalent to airline travel. After approval of the local ethics committee, 10 participants were exposed to moderate hypoxia (simulation of 2400 m or 8000 ft for 30 min in a hypobaric pressure chamber. Before and after hypobaric hypoxia, serum was drawn, centrifuged, and analysed by two-dimensional gel electrophoresis (2-DIGE and matrix-assisted laser desorption/ionization followed by time-of-flight mass spectrometry (MALDI-TOF. Biological functions of regulated proteins were identified using functional network analysis (GeneMania®, STRING®, and Perseus® software. In participants, oxygen saturation decreased from 98.1 ± 1.3% to 89.2 ± 1.8% during HH. Expression of 14 spots (i.e., 10 proteins: ALB, PGK1, APOE, GAPDH, C1QA, C1QB, CAT, CA1, F2, and CLU was significantly altered. Bioinformatic analysis revealed an association of the altered proteins with the signalling cascades “regulation of haemostasis” (four proteins, “metabolism” (five proteins, and “leukocyte mediated immune response” (five proteins. Even though hypobaric hypoxia was short and moderate (comparable to an airliner flight, analysis of protein expression in human subjects revealed an association to immune response, protein metabolism, and haemostasis

  18. Neuroendocrine-immune interaction: regulation of inflammation via G-protein coupled receptors

    NARCIS (Netherlands)

    Verburg-van Kemenade, B.M.L.; Aa, van der L.M.; Chadzinska, M.K.

    2013-01-01

    Neuroendocrine- and immune systems interact in a bi-directional fashion to communicate the status of pathogen recognition to the brain and the immune response is influenced by physiological changes. The network of ligands and their receptors involved includes cytokines and chemokines,

  19. Cancer associated aberrant protein O-glycosylation can modify antigen processing and immune response.

    Directory of Open Access Journals (Sweden)

    Caroline B Madsen

    Full Text Available Aberrant glycosylation of mucins and other extracellular proteins is an important event in carcinogenesis and the resulting cancer associated glycans have been suggested as targets in cancer immunotherapy. We assessed the role of O-linked GalNAc glycosylation on antigen uptake, processing, and presentation on MHC class I and II molecules. The effect of GalNAc O-glycosylation was monitored with a model system based on ovalbumin (OVA-MUC1 fusion peptides (+/- glycosylation loaded onto dendritic cells co-cultured with IL-2 secreting OVA peptide-specific T cell hybridomas. To evaluate the in vivo response to a cancer related tumor antigen, Balb/c or B6.Cg(CB-Tg(HLA-A/H2-D2Enge/J (HLA-A2 transgenic mice were immunized with a non-glycosylated or GalNAc-glycosylated MUC1 derived peptide followed by comparison of T cell proliferation, IFN-γ release, and antibody induction. GalNAc-glycosylation promoted presentation of OVA-MUC1 fusion peptides by MHC class II molecules and the MUC1 antigen elicited specific Ab production and T cell proliferation in both Balb/c and HLA-A2 transgenic mice. In contrast, GalNAc-glycosylation inhibited the presentation of OVA-MUC1 fusion peptides by MHC class I and abolished MUC1 specific CD8+ T cell responses in HLA-A2 transgenic mice. GalNAc glycosylation of MUC1 antigen therefore facilitates uptake, MHC class II presentation, and antibody response but might block the antigen presentation to CD8+ T cells.

  20. Characterization of Ixodes ricinus Fibrinogen-Related Proteins (Ixoderins Discloses Their Function in the Tick Innate Immunity

    Directory of Open Access Journals (Sweden)

    Helena Honig Mondekova

    2017-12-01

    Full Text Available Ticks are important vectors of serious human and animal disease-causing organisms, but their innate immunity can fight invading pathogens and may have the ability to reduce or block transmission to mammalian hosts. Lectins, sugar-binding proteins, can distinguish between self and non-self-oligosaccharide motifs on pathogen surfaces. Although tick hemolymph possesses strong lectin activity, and several lectins have already been isolated and characterized, little is known about the implementation of these molecules in tick immunity. Here, we have described and functionally characterized fibrinogen-related protein (FReP lectins in Ixodes ticks. We have shown that the FReP family contains at least 27 genes (ixoderins, ixo that could, based on phylogenetic and expression analyses, be divided into three groups with differing degrees of expansion. By using RNA interference-mediated gene silencing (RNAi we demonstrated that IXO-A was the main lectin in tick hemolymph. Further, we found that ixoderins were important for phagocytosis of Gram-negative bacteria and yeasts by tick hemocytes and that their expression was upregulated upon injection of microbes, wounding, or after blood feeding. However, although the tick hemocytes could swiftly phagocytose Borrelia afzelii spirochetes, their transmission and burst of infection in mice was not altered. Our results demonstrate that tick ixoderins are crucial immune proteins that work as opsonins in the tick hemolymph, targeting microbes for phagocytosis or lysis.

  1. Isolation and Purification of an Antibacterial Protein from Immune Induced Haemolymph of American Cockroach,Periplaneta americana.

    Science.gov (United States)

    Basseri, Hamid Reza; Dadi-Khoeni, Amir; Bakhtiari, Ronak; Abolhassani, Mandan; Hajihosseini-Baghdadabadi, Reza

    2016-12-01

    Antimicrobial peptides play a role as effectors substances in the immunity of vertebrate and invertebrate hosts. In the current study, antimicrobial peptide was isolated from the haemolymph of the American cockroach, Periplaneta americana . Micrococcus luteus as Gram-positive bacteria and Escherichia coli as Gram-negative bacteria were candidate for injection. Induction was done by injecting both bacteria into the abdominal cavity of two groups of cockroaches separately. The haemolymphs were collected 24 hours after post injection and initially tested against both bacteria. Subsequently, the immune induced haemolymph was purified by high performance liquid chromatography (HPLC) to separate the proteins responsible for the antibacterial activity. The non-induced haemolymph did not show any activity against both bacteria whereas induced haemolymph exhibited high activity against M. luteus but did less against E. coli . Two fractions showed antibacterial activity against M. luteus. Finally the molecular weight of the isolated antibacterial proteins were determined as 72 kDa and 62 kDa using SDS-PAGE. Induced haemolymph of American cockroaches has the ability to produce peptides to combat against Gram-positive bacteria when an immune challenge is mounted. Further work has to be done to sequence of the protein, which it would be advantageous.

  2. The human cytomegalovirus tegument protein pp65 (pUL83): a key player in innate immune evasion.

    Science.gov (United States)

    Biolatti, Matteo; Dell'Oste, Valentina; De Andrea, Marco; Landolfo, Santo

    2018-01-31

    The germline encoded proteins serving as "pattern recognition receptors" (PRRs) constitute the earliest step in the innate immune response by recognizing the "pathogen-associated molecular patterns" (PAMPs) that comprise microbe nucleic acids and proteins usually absent from healthy hosts. Upon detection of exogenous nucleic acid two different innate immunity signaling cascades are activated. The first culminates in the production of chemokines, cytokines, and type I interferons (IFN-I), while the second leads to inflammasome complex formation. Human cytomegalovirus (HCMV), a member of the -herpesvirus subfamily, is a wide spread pathogen that infects a vast majority of the world's population. The virion has an icosahedral capsid that contains a linear dsDNA genome of approximately 240 kb, surrounded by an outer lipid envelope and a proteinaceous tegument containing several viral proteins. Despite the numerous and multifaceted antiviral effects of IFNs and cytokines, HCMV is able to invade, multiply, and establish persistent infection in healthy human hosts. To achieve this goal the virus has developed different strategies to block the IFN-I response and to alter the physiological outcomes of the IFN-inducible genes. This article focuses on HCMV tegument pp65 by reviewing its mechanisms of action in favoring virus evasion from the host innate immune response.

  3. Two mechanistically distinct immune evasion proteins of cowpox virus combine to avoid antiviral CD8 T cells

    OpenAIRE

    Byun, Minji; Verweij, Marieke C.; Pickup, David J.; Wiertz, Emmanuel J. H. J.; Hansen, Ted H.; Yokoyama, Wayne M.

    2009-01-01

    Downregulation of MHC class I on the cell surface is an immune evasion mechanism shared by many DNA viruses including cowpox virus. Previously, a cowpox virus protein, CPXV203, was shown to downregulate MHC class I. Here, we report that CPXV12 is the only other MHC class I regulating protein of cowpox virus and it uses a mechanism distinct from that of CPXV203. Whereas CPXV203 retains fully assembled MHC class I by exploiting the KDEL-mediated endoplasmic reticulum retention pathway, CPXV12 b...

  4. Loss of Arabidopsis thaliana Dynamin-Related Protein 2B reveals separation of innate immune signaling pathways.

    Directory of Open Access Journals (Sweden)

    John M Smith

    2014-12-01

    Full Text Available Vesicular trafficking has emerged as an important means by which eukaryotes modulate responses to microbial pathogens, likely by contributing to the correct localization and levels of host components necessary for effective immunity. However, considering the complexity of membrane trafficking in plants, relatively few vesicular trafficking components with functions in plant immunity are known. Here we demonstrate that Arabidopsis thaliana Dynamin-Related Protein 2B (DRP2B, which has been previously implicated in constitutive clathrin-mediated endocytosis (CME, functions in responses to flg22 (the active peptide derivative of bacterial flagellin and immunity against flagellated bacteria Pseudomonas syringae pv. tomato (Pto DC3000. Consistent with a role of DRP2B in Pattern-Triggered Immunity (PTI, drp2b null mutant plants also showed increased susceptibility to Pto DC3000 hrcC-, which lacks a functional Type 3 Secretion System, thus is unable to deliver effectors into host cells to suppress PTI. Importantly, analysis of drp2b mutant plants revealed three distinct branches of the flg22-signaling network that differed in their requirement for RESPIRATORY BURST OXIDASE HOMOLOGUE D (RBOHD, the NADPH oxidase responsible for flg22-induced apoplastic reactive oxygen species production. Furthermore, in drp2b, normal MAPK signaling and increased immune responses via the RbohD/Ca2+-branch were not sufficient for promoting robust PR1 mRNA expression nor immunity against Pto DC3000 and Pto DC3000 hrcC-. Based on live-cell imaging studies, flg22-elicited internalization of the plant flagellin-receptor, FLAGELLIN SENSING 2 (FLS2, was found to be partially dependent on DRP2B, but not the closely related protein DRP2A, thus providing genetic evidence for a component, implicated in CME, in ligand-induced endocytosis of FLS2. Reduced trafficking of FLS2 in response to flg22 may contribute in part to the non-canonical combination of immune signaling defects

  5. Consequences of protein supplementation for anorexia, expression of immunity and plasma leptin concentrations in parasitized ewes of two breeds.

    Science.gov (United States)

    Zaralis, Konstantinos; Tolkamp, Bert J; Houdijk, Jos G M; Wylie, Alastair R G; Kyriazakis, Ilias

    2009-02-01

    The periparturient relaxation of immunity (PPRI) against parasites in ewes has a nutritional basis. We investigated whether ewes experience a reduction in food intake (anorexia) during PPRI and if the magnitude of anorexia is affected by host production potential and dietary protein supplementation. We also investigated whether nematode infection is linked to plasma leptin concentrations in periparturient ewes. The experiment was a 2 x 2 x 2 factorial design. Two breeds of twin-bearing/lactating ewes (Greyface cross, G (n 32) and Scottish Blackface, B (n 32)) were used. Half of the ewes were trickle infected with 30,000 larvae of the abomasal parasite Teladorsagia circumcincta per week and the other half were not. During the experiment, all ewes had ad libitum access to a low-protein diet that provided less protein than the recommended allowance. In addition, half of the ewes received a protein supplement that resulted in protein intakes that exceeded recommendations. Nematode infection resulted in a breakdown of immunity to parasites and a reduction in food intake in both breeds. The breeds differed in the extent of PPRI (G ewes having higher faecal egg counts than B ewes), but not in the magnitude of anorexia. Protein supplementation resulted in a reduction in faecal egg counts, but had no effect on the magnitude of anorexia. Plasma leptin concentrations changed significantly over time, but were not affected by protein supplementation or infection. It is concluded that infection with T. circumcincta in periparturient ewes results in anorexia that is not alleviated by protein supplementation and seems unrelated to plasma leptin concentrations.

  6. Sequence variations and protein expression levels of the two immune evasion proteins Gpm1 and Pra1 influence virulence of clinical Candida albicans isolates.

    Directory of Open Access Journals (Sweden)

    Shanshan Luo

    Full Text Available Candida albicans, the important human fungal pathogen uses multiple evasion strategies to control, modulate and inhibit host complement and innate immune attack. Clinical C. albicans strains vary in pathogenicity and in serum resistance, in this work we analyzed sequence polymorphisms and variations in the expression levels of two central fungal complement evasion proteins, Gpm1 (phosphoglycerate mutase 1 and Pra1 (pH-regulated antigen 1 in thirteen clinical C. albicans isolates. Four nucleotide (nt exchanges, all representing synonymous exchanges, were identified within the 747-nt long GPM1 gene. For the 900-nt long PRA1 gene, sixteen nucleotide exchanges were identified, which represented synonymous, as well as non-synonymous exchanges. All thirteen clinical isolates had a homozygous exchange (A to G at position 73 of the PRA1 gene. Surface levels of Gpm1 varied by 8.2, and Pra1 levels by 3.3 fold in thirteen tested isolates and these differences influenced fungal immune fitness. The high Gpm1/Pra1 expressing candida strains bound the three human immune regulators more efficiently, than the low expression strains. The difference was 44% for Factor H binding, 51% for C4BP binding and 23% for plasminogen binding. This higher Gpm1/Pra1 expressing strains result in enhanced survival upon challenge with complement active, Factor H depleted human serum (difference 40%. In addition adhesion to and infection of human endothelial cells was increased (difference 60%, and C3b surface deposition was less effective (difference 27%. Thus, variable expression levels of central immune evasion protein influences immune fitness of the human fungal pathogen C. albicans and thus contribute to fungal virulence.

  7. Sequence variations and protein expression levels of the two immune evasion proteins Gpm1 and Pra1 influence virulence of clinical Candida albicans isolates.

    Science.gov (United States)

    Luo, Shanshan; Hipler, Uta-Christina; Münzberg, Christin; Skerka, Christine; Zipfel, Peter F

    2015-01-01

    Candida albicans, the important human fungal pathogen uses multiple evasion strategies to control, modulate and inhibit host complement and innate immune attack. Clinical C. albicans strains vary in pathogenicity and in serum resistance, in this work we analyzed sequence polymorphisms and variations in the expression levels of two central fungal complement evasion proteins, Gpm1 (phosphoglycerate mutase 1) and Pra1 (pH-regulated antigen 1) in thirteen clinical C. albicans isolates. Four nucleotide (nt) exchanges, all representing synonymous exchanges, were identified within the 747-nt long GPM1 gene. For the 900-nt long PRA1 gene, sixteen nucleotide exchanges were identified, which represented synonymous, as well as non-synonymous exchanges. All thirteen clinical isolates had a homozygous exchange (A to G) at position 73 of the PRA1 gene. Surface levels of Gpm1 varied by 8.2, and Pra1 levels by 3.3 fold in thirteen tested isolates and these differences influenced fungal immune fitness. The high Gpm1/Pra1 expressing candida strains bound the three human immune regulators more efficiently, than the low expression strains. The difference was 44% for Factor H binding, 51% for C4BP binding and 23% for plasminogen binding. This higher Gpm1/Pra1 expressing strains result in enhanced survival upon challenge with complement active, Factor H depleted human serum (difference 40%). In addition adhesion to and infection of human endothelial cells was increased (difference 60%), and C3b surface deposition was less effective (difference 27%). Thus, variable expression levels of central immune evasion protein influences immune fitness of the human fungal pathogen C. albicans and thus contribute to fungal virulence.

  8. Molecular characterization of the insect immune protein hemolin and its high induction during embryonic diapause in the gypsy moth, Lymantria dispar

    Science.gov (United States)

    K.-Y. Lee; F. M. Horodyski; A. P. Valaitis; D. L. Denlinger

    2002-01-01

    During the embryonic (pharate first instar) diapause of the gypsy moth, Lymantria dispar, a 55 kDa protein is highly up-regulated in the gut. We now identify that protein as hemolin, an immune protein in the immunoglobulin superfamily. We isolated a gypsy moth hemolin cDNA and demonstrated a high degree of similarity with hemolins from three other...

  9. Analysis of Globodera rostochiensis effectors reveals conserved functions of SPRYSEC proteins in suppressing and eliciting plant immune responses

    KAUST Repository

    Ali, Shawkat

    2015-08-11

    Potato cyst nematodes (PCNs), including Globodera rostochiensis (Woll.), are important pests of potato. Plant parasitic nematodes produce multiple effector proteins, secreted from their stylets, to successfully infect their hosts. These include proteins delivered to the apoplast and to the host cytoplasm. A number of effectors from G. rostochiensis predicted to be delivered to the host cytoplasm have been identified, including several belonging to the secreted SPRY domain (SPRYSEC) family. SPRYSEC proteins are unique to members of the genus Globodera and have been implicated in both the induction and the repression of host defense responses. We have tested the properties of six different G. rostochiensis SPRYSEC proteins by expressing them in Nicotiana benthamiana and N. tabacum. We have found that all SPRYSEC proteins tested are able to suppress defense responses induced by NB-LRR proteins as well as cell death induced by elicitors, suggesting that defense repression is a common characteristic of members of this effector protein family. At the same time, GrSPRYSEC-15 elicited a defense responses in N. tabacum, which was found to be resistant to a virus expressing GrSPRYSEC-15. These results suggest that SPRYSEC proteins may possess characteristics that allow them to be recognized by the plant immune system.

  10. Suppression of host immune response by the core protein of hepatitis C virus: possible implications for hepatitis C virus persistence.

    Science.gov (United States)

    Large, M K; Kittlesen, D J; Hahn, Y S

    1999-01-15

    Hepatitis C virus (HCV) is a major human pathogen causing mild to severe liver disease worldwide. This positive strand RNA virus is remarkably efficient at establishing chronic infections. Although a high rate of genetic variability may facilitate viral escape and persistence in the face of Ag-specific immune responses, HCV may also encode proteins that facilitate evasion of immunological surveillance. To address the latter possibility, we examined the influence of specific HCV gene products on the host immune response to vaccinia virus in a murine model. Various vaccinia/HCV recombinants expressing different regions of the HCV polyprotein were used for i.p. inoculation of BALB/c mice. Surprisingly, a recombinant expressing the N-terminal half of the polyprotein (including the structural proteins, p7, NS2, and a portion of NS3; vHCV-S) led to a dose-dependent increase in mortality. Increased mortality was not observed for a recombinant expressing the majority of the nonstructural region or for a negative control virus expressing the beta-galactosidase protein. Examination of T cell responses in these mice revealed a marked suppression of vaccinia-specific CTL responses and a depressed production of IFN-gamma and IL-2. By using a series of vaccinia/HCV recombinants, we found that the HCV core protein was sufficient for immunosuppression, prolonged viremia, and increased mortality. These results suggest that the HCV core protein plays an important role in the establishment and maintenance of HCV infection by suppressing host immune responses, in particular the generation of virus-specific CTLs.

  11. Protein Malnutrition Modifies Innate Immunity and Gene Expression by Intestinal Epithelial Cells and Human Rotavirus Infection in Neonatal Gnotobiotic Pigs.

    Science.gov (United States)

    Vlasova, Anastasia N; Paim, Francine C; Kandasamy, Sukumar; Alhamo, Moyasar A; Fischer, David D; Langel, Stephanie N; Deblais, Loic; Kumar, Anand; Chepngeno, Juliet; Shao, Lulu; Huang, Huang-Chi; Candelero-Rueda, Rosario A; Rajashekara, Gireesh; Saif, Linda J

    2017-01-01

    Malnutrition affects millions of children in developing countries, compromising immunity and contributing to increased rates of death from infectious diseases. Rotavirus is a major etiological agent of childhood diarrhea in developing countries, where malnutrition is prevalent. However, the interactions between the two and their combined effects on immune and intestinal functions are poorly understood. In this study, we used neonatal gnotobiotic (Gn) pigs transplanted with the fecal microbiota of a healthy 2-month-old infant (HIFM) and fed protein-deficient or -sufficient bovine milk diets. Protein deficiency induced hypoproteinemia, hypoalbuminemia, hypoglycemia, stunting, and generalized edema in Gn pigs, as observed in protein-malnourished children. Irrespective of the diet, human rotavirus (HRV) infection early, at HIFM posttransplantation day 3 (PTD3), resulted in adverse health effects and higher mortality rates (45 to 75%) than later HRV infection (PTD10). Protein malnutrition exacerbated HRV infection and affected the morphology and function of the small intestinal epithelial barrier. In pigs infected with HRV at PTD10, there was a uniform decrease in the function and/or frequencies of natural killer cells, plasmacytoid dendritic cells, and CD103 + and apoptotic mononuclear cells and altered gene expression profiles of intestinal epithelial cells (chromogranin A, mucin 2, proliferating cell nuclear antigen, SRY-Box 9, and villin). Thus, we have established the first HIFM-transplanted neonatal pig model that recapitulates major aspects of protein malnutrition in children and can be used to evaluate physiologically relevant interventions. Our findings provide an explanation of why nutrient-rich diets alone may lack efficacy in malnourished children. IMPORTANCE Malnutrition and rotavirus infection, prevalent in developing countries, individually and in combination, affect the health of millions of children, compromising their immunity and increasing the rates

  12. Echinoderm immunity.

    Science.gov (United States)

    Smith, L Courtney; Ghosh, Julie; Buckley, Katherine M; Clow, Lori A; Dheilly, Nolwenn M; Haug, Tor; Henson, John H; Li, Chun; Lun, Cheng Man; Majeske, Audrey J; Matranga, Valeria; Nair, Sham V; Rast, Jonathan P; Raftos, David A; Roth, Mattias; Sacchi, Sandro; Schrankel, Catherine S; Stensvåg, Klara

    2010-01-01

    A survey for immune genes in the genome for the purple sea urchin has shown that the immune system is complex and sophisticated. By inference, immune responses of all echinoderms maybe similar. The immune system is mediated by several types of coelomocytes that are also useful as sensors of environmental stresses. There are a number of large gene families in the purple sea urchin genome that function in immunity and of which at least one appears to employ novel approaches for sequence diversification. Echinoderms have a simpler complement system, a large set of lectin genes and a number of antimicrobial peptides. Profiling the immune genes expressed by coelomocytes and the proteins in the coelomic fluid provide detailed information about immune functions in the sea urchin. The importance of echinoderms in maintaining marine ecosystem stability and the disastrous effects of their removal due to disease will require future collaborations between ecologists and immunologists working towards understanding and preserving marine habitats.

  13. Immunization with Brugia malayi Myosin as Heterologous DNA Prime Protein Boost Induces Protective Immunity against B. malayi Infection in Mastomys coucha.

    Directory of Open Access Journals (Sweden)

    Jyoti Gupta

    Full Text Available The current control strategies employing chemotherapy with diethylcarbamazine, ivermectin and albendazole have reduced transmission in some filaria-endemic areas, there is growing interest for complementary approaches, such as vaccines especially in light of threat of parasite developing resistance to mainstay drugs. We earlier demonstrated recombinant heavy chain myosin of B. malayi (Bm-Myo as a potent vaccine candidate whose efficacy was enhanced by heterologous DNA prime/protein boost (Myo-pcD+Bm-Myo vaccination in BALB/c mice. BALB/c mouse though does not support the full developmental cycle of B. malayi, however, the degree of protection may be studied in terms of transformation of challenged infective larvae (L3 to next stage (L4 with an ease of delineating the generated immunological response of host. In the current investigation, DNA vaccination with Bm-Myo was therefore undertaken in susceptible rodent host, Mastomys coucha (M. coucha which sustains the challenged L3 and facilitates their further development to sexually mature adult parasites with patent microfilaraemia. Immunization schedule consisted of Myo-pcD and Myo-pcD+Bm-Myo followed by B. malayi L3 challenge and the degree of protection was evaluated by observing microfilaraemia as well as adult worm establishment. Myo-pcD+Bm-Myo immunized animals not only developed 78.5% reduced blood microfilarial density but also decreased adult worm establishment by 75.3%. In addition, 75.4% of the recovered live females revealed sterilization over those of respective control animals. Myo-pcD+Bm-Myo triggered higher production of specific IgG and its isotypes which induced marked cellular adhesion and cytotoxicity (ADCC to microfilariae (mf and L3 in vitro. Both Th1 and Th2 cytokines were significantly up-regulated displaying a mixed immune response conferring considerable protection against B. malayi establishment by engendering a long-lasting effective immune response and therefore emerges

  14. Structural and functional annotation of human FAM26F: A multifaceted protein having a critical role in the immune system.

    Science.gov (United States)

    Malik, Uzma; Javed, Aneela; Ali, Amjad; Asghar, Kashif

    2017-01-15

    Human immune system is a complex amalgam of a greatly diverse ensemble comprising of various cellular and non-cellular components, including proteins. FAM26F (family with sequence similarity 26, member F) is a relatively recently identified gene reported to play important role in diverse immune responses. Numerous studies have reported FAM26F to be differentially expressed in several viral, bacterial and parasitic infections, in certain pathophysiological conditions such as heart and liver transplantation, and in several cancers. FAM26F has also been found to be upregulated by various stimulants such as polyI:C, LPS, INF gamma and TNF alpha, and via various anticipated pathways including TLR3, TLR4 IFN-β and Dectin-1. Moreover, the synergistic expression of FAM26F on both NK-cells and myeloid dendritic cells is required to activate NK-cells against tumors via its cytoplasmic tail, thus emphasizing the therapeutic potential of FAM26F for NK sensitive tumors. Although a considerable amount of evidence is present regarding the potential role of FAM26F in immune modulation, the exact function and modulatory pathways of this gene are yet to be elucidated. We aimed to completely characterize FAM26F in order to apprehend its function and role in the immune responses. The results revealed human FAM26F to be located at chromosomal position 6q22.1. FAM26F mRNA contains 1141bp coding region encoding a 315 amino acid long, stable protein that has been well-conserved throughout evolution. It is a signal peptide deprived transmembrane protein that is secreted through non-classical pathway. The presence of a single well-conserved Ca_hom_mod domain indicated FAM26F to be a cation channel involved in the transport of molecules. A potential N-glycosylation and 14 phosphorylation sites were also predicted, along with four interacting partners of FAM26F. The secondary and tertiary structures of FAM26F were determined. Moreover, the presence of an immunoglobulin-like fold in FAM26F

  15. Composition and Variation of Macronutrients, Immune Proteins, and Human Milk Oligosaccharides in Human Milk From Nonprofit and Commercial Milk Banks.

    Science.gov (United States)

    Meredith-Dennis, Laura; Xu, Gege; Goonatilleke, Elisha; Lebrilla, Carlito B; Underwood, Mark A; Smilowitz, Jennifer T

    2018-02-01

    When human milk is unavailable, banked milk is recommended for feeding premature infants. Milk banks use processes to eliminate pathogens; however, variability among methods exists. Research aim: The aim of this study was to compare the macronutrient (protein, carbohydrate, fat, energy), immune-protective protein, and human milk oligosaccharide (HMO) content of human milk from three independent milk banks that use pasteurization (Holder vs. vat techniques) or retort sterilization. Randomly acquired human milk samples from three different milk banks ( n = 3 from each bank) were analyzed for macronutrient concentrations using a Fourier transform mid-infrared spectroscopy human milk analyzer. The concentrations of IgA, IgM, IgG, lactoferrin, lysozyme, α-lactalbumin, α antitrypsin, casein, and HMO were analyzed by mass spectrometry. The concentrations of protein and fat were significantly ( p milk samples that had undergone retort sterilization had significantly less immune-protective proteins and total and specific HMOs compared with samples that had undergone Holder and vat pasteurization. These data suggest that further analysis of the effect of retort sterilization on human milk components is needed prior to widespread adoption of this process.

  16. Lloviu virus VP24 and VP35 proteins function as innate immune antagonists in human and bat cells

    International Nuclear Information System (INIS)

    Feagins, Alicia R.; Basler, Christopher F.

    2015-01-01

    Lloviu virus (LLOV) is a new member of the filovirus family that also includes Ebola virus (EBOV) and Marburg virus (MARV). LLOV has not been cultured; however, its genomic RNA sequence indicates the coding capacity to produce homologs of the EBOV and MARV VP24, VP35, and VP40 proteins. EBOV and MARV VP35 proteins inhibit interferon (IFN)-alpha/beta production and EBOV VP35 blocks activation of the antiviral kinase PKR. The EBOV VP24 and MARV VP40 proteins inhibit IFN signaling, albeit by different mechanisms. Here we demonstrate that LLOV VP35 suppresses Sendai virus induced IFN regulatory factor 3 (IRF3) phosphorylation, IFN-α/β production, and PKR phosphorylation. Additionally, LLOV VP24 blocks tyrosine phosphorylated STAT1 binding to karyopherin alpha 5 (KPNA5), STAT1 nuclear accumulation, and IFN-induced gene expression. LLOV VP40 lacks detectable IFN antagonist function. These activities parallel EBOV IFN inhibitory functions. EBOV and LLOV VP35 and VP24 proteins also inhibit IFN responses in bat cells. These data suggest that LLOV infection will block innate immune responses in a manner similar to EBOV. - Highlights: • Lloviu virus (LLOV) is a new member of the filovirus family. • LLOV VP35 blocks IRF3 phosphorylation, IFN-α/β production and PKR phosphorylation. • LLOV VP24 inhibits IFN responses by targeting phospho-STAT1 KPNA interaction. • Infection by LLOV may block innate immune responses in a manner similar to EBOV.

  17. Lloviu virus VP24 and VP35 proteins function as innate immune antagonists in human and bat cells

    Energy Technology Data Exchange (ETDEWEB)

    Feagins, Alicia R.; Basler, Christopher F., E-mail: chris.basler@mssm.edu

    2015-11-15

    Lloviu virus (LLOV) is a new member of the filovirus family that also includes Ebola virus (EBOV) and Marburg virus (MARV). LLOV has not been cultured; however, its genomic RNA sequence indicates the coding capacity to produce homologs of the EBOV and MARV VP24, VP35, and VP40 proteins. EBOV and MARV VP35 proteins inhibit interferon (IFN)-alpha/beta production and EBOV VP35 blocks activation of the antiviral kinase PKR. The EBOV VP24 and MARV VP40 proteins inhibit IFN signaling, albeit by different mechanisms. Here we demonstrate that LLOV VP35 suppresses Sendai virus induced IFN regulatory factor 3 (IRF3) phosphorylation, IFN-α/β production, and PKR phosphorylation. Additionally, LLOV VP24 blocks tyrosine phosphorylated STAT1 binding to karyopherin alpha 5 (KPNA5), STAT1 nuclear accumulation, and IFN-induced gene expression. LLOV VP40 lacks detectable IFN antagonist function. These activities parallel EBOV IFN inhibitory functions. EBOV and LLOV VP35 and VP24 proteins also inhibit IFN responses in bat cells. These data suggest that LLOV infection will block innate immune responses in a manner similar to EBOV. - Highlights: • Lloviu virus (LLOV) is a new member of the filovirus family. • LLOV VP35 blocks IRF3 phosphorylation, IFN-α/β production and PKR phosphorylation. • LLOV VP24 inhibits IFN responses by targeting phospho-STAT1 KPNA interaction. • Infection by LLOV may block innate immune responses in a manner similar to EBOV.

  18. Relationship between SPRY and B30.2 protein domains. Evolution of a component of immune defence?

    Science.gov (United States)

    Rhodes, David A; de Bono, Bernard; Trowsdale, John

    2005-01-01

    SPRY and B30.2 are homologous domains which can be identified in 11 protein families encoded in the human genome. These include cell surface receptors of the immunoglobulin super-family (BTNs), negative regulators of the JAK/STAT pathway (SOCS-box SSB1–4) and proteins encoded by the numerous TRIM genes. Collectively, proteins containing SPRY and B30.2 domains cover a wide range of functions, including regulation of cytokine signalling (SOCS), RNA metabolism (DDX1, hnRNPs), intracellular calcium release (RyR receptors), immunity to retroviruses (TRIM5alpha) as well as regulatory and developmental processes (HERC1, Ash2L). In order to clarify the evolutionary relationship between the two domains, we compiled a curated database of SPRY and B30.2-domain sequences. We show that while SPRY domains are evolutionarily ancient, B30.2 domains, found in BTN and TRIM proteins, are a more recent evolutionary adaptation, comprising the combination of SPRY with an additional domain, PRY. The combination of SPRY and PRY to produce B30.2 domains may have been selected and maintained as a component of immune defence. PMID:16313355

  19. Dengue-1 envelope protein domain III along with PELC and CpG oligodeoxynucleotides synergistically enhances immune responses.

    Directory of Open Access Journals (Sweden)

    Chen-Yi Chiang

    Full Text Available The major weaknesses of subunit vaccines are their low immunogenicity and poor efficacy. Adjuvants can help to overcome some of these inherent defects with subunit vaccines. Here, we evaluated the efficacy of the newly developed water-in-oil-in-water multiphase emulsion system, termed PELC, in potentiating the protective capacity of dengue-1 envelope protein domain III. Unlike aluminum phosphate, dengue-1 envelope protein domain III formulated with PELC plus CpG oligodeoxynucleotides induced neutralizing antibodies against dengue-1 virus and increased the splenocyte secretion of IFN-γ after in vitro re-stimulation. The induced antibodies contained both the IgG1 and IgG2a subclasses. A rapid anamnestic neutralizing antibody response against a live dengue virus challenge was elicited at week 26 after the first immunization. These results demonstrate that PELC plus CpG oligodeoxynucleotides broaden the dengue-1 envelope protein domain III-specific immune responses. PELC plus CpG oligodeoxynucleotides is a promising adjuvant for recombinant protein based vaccination against dengue virus.

  20. Dietary Animal Plasma Proteins Improve the Intestinal Immune Response in Senescent Mice.

    Science.gov (United States)

    Miró, Lluïsa; Garcia-Just, Alba; Amat, Concepció; Polo, Javier; Moretó, Miquel; Pérez-Bosque, Anna

    2017-12-11

    Increased life expectancy has promoted research on healthy aging. Aging is accompanied by increased non-specific immune activation (inflammaging) which favors the appearance of several disorders. Here, we study whether dietary supplementation with spray-dried animal plasma (SDP), which has been shown to reduce the activation of gut-associated lymphoid tissue (GALT) in rodents challenged by S. aureus enterotoxin B (SEB), and can also prevent the effects of aging on immune system homeostasis. We first characterized GALT in a mouse model of accelerated senescence (SAMP8) at different ages (compared to mice resistant to accelerated senescence; SAMR1). Second, we analyzed the SDP effects on GALT response to an SEB challenge in SAMP8 mice. In GALT characterization, aging increased the cell number and the percentage of activated Th lymphocytes in mesenteric lymph nodes and Peyer's patches (all, p response to the SEB challenge, young mice showed increased expression of intestinal IL-6 and TNF-α, as well as lymphocyte recruitment and activation (all, p immune response of senescent mice to the SEB challenge was weak, since SEB did not change cell recruitment or the percentage of activated Th lymphocytes. Mice supplemented with SDP showed improved capacity to respond to the SEB challenge, similar to the response of the young mice. These results indicate that senescent mice have an impaired mucosal immune response characterized by unspecific GALT activation and a weak specific immune response. SDP supplementation reduces non-specific basal immune activation, allowing for the generation of specific responses.

  1. The G protein-coupled receptor FSHR-1 is required for the Caenorhabditis elegans innate immune response.

    Science.gov (United States)

    Powell, Jennifer R; Kim, Dennis H; Ausubel, Frederick M

    2009-02-24

    Innate immunity is an ancient defense system used by both vertebrates and invertebrates. Previously characterized innate immune responses in plants and animals are triggered by detection of pathogens using specific receptors, which typically use a leucine-rich repeat (LRR) domain to bind molecular patterns associated with infection. The nematode Caenorhabditis elegans uses defense pathways conserved with vertebrates; however, the mechanism by which C. elegans detects pathogens is unknown. We screened all LRR-containing transmembrane receptors in C. elegans and identified the G protein-coupled receptor FSHR-1 as an important component of the C. elegans immune response to Gram-negative and Gram-positive bacterial pathogens. FSHR-1 acts in the C. elegans intestine, the primary site of exposure to ingested pathogens. FSHR-1 signals in parallel to the known p38 MAPK pathway but converges to regulate the transcriptional induction of an overlapping but nonidentical set of antimicrobial effectors. FSHR-1 may act generally to boost the nematode immune response, or it may function as a pathogen receptor.

  2. The Salmonella Effector Protein SopA Modulates Innate Immune Responses by Targeting TRIM E3 Ligase Family Members.

    Directory of Open Access Journals (Sweden)

    Jana Kamanova

    2016-04-01

    Full Text Available Salmonella Typhimurium stimulates inflammatory responses in the intestinal epithelium, which are essential for its ability to replicate within the intestinal tract. Stimulation of these responses is strictly dependent on the activity of a type III secretion system encoded within its pathogenicity island 1, which through the delivery of effector proteins, triggers signaling pathways leading to inflammation. One of these effectors is SopA, a HECT-type E3 ligase, which is required for the efficient stimulation of inflammation in an animal model of Salmonella Typhimurium infection. We show here that SopA contributes to the stimulation of innate immune responses by targeting two host E3 ubiquitin ligases, TRIM56 and TRIM65. We also found that TRIM65 interacts with the innate immune receptor MDA5 enhancing its ability to stimulate interferon-β signaling. Therefore, by targeting TRIM56 and TRIM65, SopA can stimulate signaling through two innate immune receptors, RIG-I and MDA5. These findings describe a Salmonella mechanism to modulate inflammatory responses by directly targeting innate immune signaling mechanisms.

  3. Depression of leukocyte protein synthesis, immune function and growth performance induced by high environmental temperature in broiler chickens

    Science.gov (United States)

    Kamel, Nancy N.; Ahmed, Ayman M. H.; Mehaisen, Gamal M. K.; Mashaly, Magdi M.; Abass, Ahmed O.

    2017-09-01

    In tropical and semitropical regions, raising broiler chickens out of their thermal comfort zone can cause an added economic loss in the poultry industry. The cause for the deleterious effects on immunity and growth performance of broilers under high environmental temperatures is still poorly understood. Therefore, the aim of the current investigation was to evaluate the effect of heat stress on leukocytes protein synthesis and immune function as a possible direct cause of low performance in broiler chickens under such condition. In this study, 300 one-day-old male broiler chicks (Cobb500™) were randomly assigned into 2 groups with 5 replicates of 30 chicks each. From 21 to 42 days of age, one group was exposed to non-stressed condition at 24 °C and 50% relative humidity (control group), while the other group was exposed to heat stress at 35 °C and 50% relative humidity (HS group). At 42 days of age, blood samples were collected from each group to evaluate stress indicators, immune function, and leukocytes protein synthesis. Production performance was also recorded. Noteworthy, protein synthesis in leukocytes was significantly ( P group by 38% compared to control group. In contrast, the phosphorylation level on threonine 56 site (Thr56) of eukaryotic elongation factor (eEF2), which indicates the suppression of protein translation process through altering the protein elongation phase, was significantly threefold higher in HS group than in control ( P group, respectively). Moreover, results on the broiler performance indicate that HS birds had a significant ( P < 0.05) lower body weight gain by 58%, lower feed consumption by 39%, higher conversion ratio by 27%, and higher mortality by more than three times, compared to control birds. In conclusion, our results demonstrate that the inhibition of leukocyte protein synthesis through increasing the level of eEF2 Thr56 phosphorylation may play a key role in the observed decrease in immune function and growth

  4. Peanut protein structure, polyphenol content and immune response to peanut proteins in vivo are modulated by laccase.

    Science.gov (United States)

    Mihajlovic, L; Radosavljevic, J; Nordlund, E; Krstic, M; Bohn, T; Smit, J; Buchert, J; Cirkovic Velickovic, T

    2016-05-18

    Food texture can be improved by enzyme-mediated covalent cross-linking of different food components, such as proteins and carbohydrates. Cross-linking changes the biological and immunological properties of proteins and may change the sensitizing potential of food allergens. In this study we applied a microbial polyphenol oxidase, laccase, to cross-link peanut proteins. The size and morphology of the obtained cross-linked proteins were analyzed by electrophoresis and electron microscopy. Structural changes in proteins were analyzed by CD spectroscopy and by using specific antibodies to major peanut allergens. The bioavailability of peanut proteins was analyzed using a Caco-2 epithelial cell model. The in vivo sensitizing potential of laccase-treated peanut proteins was analyzed using a mouse model of food allergy. Finally, peanut polyphenols were analyzed by UHPLC-MS/MS, before and after the enzymatic reaction with laccase. Laccase treatment of peanut proteins yielded a covalently cross-linked material, with the modified tertiary structure of peanut proteins, improved bioavailability of Ara h 2 (by 70 fold, p polyphenol content and profile by HPLC-MS/MS revealed that laccase treatment depleted the peanut extract of polyphenol compounds leaving mostly isorhamnetin derivatives and procyanidin dimer B-type in detectable amounts. Treatment of complex food extracts rich in polyphenols with laccase results in both protein cross-linking and modification of polyphenol compounds. These extensively cross-linked proteins have unchanged potency to induce allergic sensitization in vivo, but certain immunomodulatory changes were observed.

  5. Chapter 11 Unexpected Turns and Twists in Structure/Function of PR-Proteins that Connect Energy Metabolism and Immunity

    KAUST Repository

    Narasimhan, Meena L.

    2009-01-01

    Innate immunity in plants is manifested by a complex array of antimicrobial processes that includes induction of sets of pathogenesis-related (PR) proteins. The availability of genomic data has made clear that each PR-protein family in a species is represented by several genes. Microarray data in public databases show that in most families, including the PR-5 family surveyed here, the expression of only few family members is defense associated. Genetic studies show that depending on their nutrient acquisition strategy, pathogens induce distinct but overlapping sets of PR genes, suggesting a connection to energy or resource allocation. PR-5 proteins have a clearly recognizable structure that is referred to as the thaumatin (THN) domain, which can be overlapped with mammalian Complement 1q-tumor necrosis factor (C1q-TNF) domains such as that of the mammalian hormone adiponectin. The occurrence of THN domain proteins is widespread. Similarities between THN domain proteins and mammalian C1q-TNF family proteins include their ligands and their subcellular locations. Osmotin (tobacco PR-5c) regulates energy balance signaling in mammalian cells by interaction with adiponectin receptors by a pathway that shares components with plant energy and stress signaling pathways. These data suggest additional roles for PR-5 proteins, as scaffolds and/or in signaling, particularly in regulating energy balance. © 2009 Elsevier Ltd. All rights reserved.

  6. Boltzmann energy-based image analysis demonstrates that extracellular domain size differences explain protein segregation at immune synapses.

    Directory of Open Access Journals (Sweden)

    Nigel J Burroughs

    2011-08-01

    Full Text Available Immune synapses formed by T and NK cells both show segregation of the integrin ICAM1 from other proteins such as CD2 (T cell or KIR (NK cell. However, the mechanism by which these proteins segregate remains unclear; one key hypothesis is a redistribution based on protein size. Simulations of this mechanism qualitatively reproduce observed segregation patterns, but only in certain parameter regimes. Verifying that these parameter constraints in fact hold has not been possible to date, this requiring a quantitative coupling of theory to experimental data. Here, we address this challenge, developing a new methodology for analysing and quantifying image data and its integration with biophysical models. Specifically we fit a binding kinetics model to 2 colour fluorescence data for cytoskeleton independent synapses (2 and 3D and test whether the observed inverse correlation between fluorophores conforms to size dependent exclusion, and further, whether patterned states are predicted when model parameters are estimated on individual synapses. All synapses analysed satisfy these conditions demonstrating that the mechanisms of protein redistribution have identifiable signatures in their spatial patterns. We conclude that energy processes implicit in protein size based segregation can drive the patternation observed in individual synapses, at least for the specific examples tested, such that no additional processes need to be invoked. This implies that biophysical processes within the membrane interface have a crucial impact on cell:cell communication and cell signalling, governing protein interactions and protein aggregation.

  7. IRE1/bZIP60-mediated unfolded protein response plays distinct roles in plant immunity and abiotic stress responses.

    Directory of Open Access Journals (Sweden)

    Adrian A Moreno

    Full Text Available Endoplasmic reticulum (ER-mediated protein secretion and quality control have been shown to play an important role in immune responses in both animals and plants. In mammals, the ER membrane-located IRE1 kinase/endoribonuclease, a key regulator of unfolded protein response (UPR, is required for plasma cell development to accommodate massive secretion of immunoglobulins. Plant cells can secrete the so-called pathogenesis-related (PR proteins with antimicrobial activities upon pathogen challenge. However, whether IRE1 plays any role in plant immunity is not known. Arabidopsis thaliana has two copies of IRE1, IRE1a and IRE1b. Here, we show that both IRE1a and IRE1b are transcriptionally induced during chemically-induced ER stress, bacterial pathogen infection and treatment with the immune signal salicylic acid (SA. However, we found that IRE1a plays a predominant role in the secretion of PR proteins upon SA treatment. Consequently, the ire1a mutant plants show enhanced susceptibility to a bacterial pathogen and are deficient in establishing systemic acquired resistance (SAR, whereas ire1b is unaffected in these responses. We further demonstrate that the immune deficiency in ire1a is due to a defect in SA- and pathogen-triggered, IRE1-mediated cytoplasmic splicing of the bZIP60 mRNA, which encodes a transcription factor involved in the expression of UPR-responsive genes. Consistently, IRE1a is preferentially required for bZIP60 splicing upon pathogen infection, while IRE1b plays a major role in bZIP60 processing upon Tunicamycin (Tm-induced stress. We also show that SA-dependent induction of UPR-responsive genes is altered in the bzip60 mutant resulting in a moderate susceptibility to a bacterial pathogen. These results indicate that the IRE1/bZIP60 branch of UPR is a part of the plant response to pathogens for which the two Arabidopsis IRE1 isoforms play only partially overlapping roles and that IRE1 has both bZIP60-dependent and bZIP60-independent

  8. Ectopic expression of X-linked lymphocyte-regulated protein pM1 renders tumor cells resistant to antitumor immunity.

    Science.gov (United States)

    Kang, Tae Heung; Noh, Kyung Hee; Kim, Jin Hee; Bae, Hyun Cheol; Lin, Ken Y; Monie, Archana; Pai, Sara I; Hung, Chien-Fu; Wu, T-C; Kim, Tae Woo

    2010-04-15

    Tumor immune escape is a major obstacle in cancer immunotherapy, but the mechanisms involved remain poorly understood. We have previously developed an immune evasion tumor model using an in vivo immune selection strategy and revealed Akt-mediated immune resistance to antitumor immunity induced by various cancer immunotherapeutic agents. In the current study, we used microarray gene analysis to identify an Akt-activating candidate molecule overexpressed in immune-resistant tumors compared with parental tumors. X-linked lymphocyte-regulated protein pM1 (XLR) gene was the most upregulated in immune-resistant tumors compared with parental tumor cells. Furthermore, the retroviral transduction of XLR in parental tumor cells led to activation of Akt, resulting in upregulation of antiapoptotic proteins and the induction of immune resistance phenotype in parental tumor cells. In addition, we found that transduction of parental tumor cells with other homologous genes from the mouse XLR family, such as synaptonemal complex protein 3 (SCP3) and XLR-related, meiosis-regulated protein (XMR) and its human counterpart of SCP3 (hSCP3), also led to activation of Akt, resulting in the upregulation of antiapoptotic proteins and induction of immune resistance phenotype. Importantly, characterization of a panel of human cervical cancers revealed relatively higher expression levels of hSCP3 in human cervical cancer tissue compared with normal cervical tissue. Thus, our data indicate that ectopic expression of XLR and its homologues in tumor cells represents a potentially important mechanism for tumor immune evasion and serves as a promising molecular target for cancer immunotherapy. (c) 2010 AACR.

  9. Systematic identification of anti-interferon function on hepatitis C virus genome reveals p7 as an immune evasion protein.

    Science.gov (United States)

    Qi, Hangfei; Chu, Virginia; Wu, Nicholas C; Chen, Zugen; Truong, Shawna; Brar, Gurpreet; Su, Sheng-Yao; Du, Yushen; Arumugaswami, Vaithilingaraja; Olson, C Anders; Chen, Shu-Hua; Lin, Chung-Yen; Wu, Ting-Ting; Sun, Ren

    2017-02-21

    Hepatitis C virus (HCV) encodes mechanisms to evade the multilayered antiviral actions of the host immune system. Great progress has been made in elucidating the strategies HCV employs to down-regulate interferon (IFN) production, impede IFN signaling transduction, and impair IFN-stimulated gene (ISG) expression. However, there is a limited understanding of the mechanisms governing how viral proteins counteract the antiviral functions of downstream IFN effectors due to the lack of an efficient approach to identify such interactions systematically. To study the mechanisms by which HCV antagonizes the IFN responses, we have developed a high-throughput profiling platform that enables mapping of HCV sequences critical for anti-IFN function at high resolution. Genome-wide profiling performed with a 15-nt insertion mutant library of HCV showed that mutations in the p7 region conferred high levels of IFN sensitivity, which could be alleviated by the expression of WT p7 protein. This finding suggests that p7 protein of HCV has an immune evasion function. By screening a liver-specific ISG library, we identified that IFI6-16 significantly inhibits the replication of p7 mutant viruses without affecting WT virus replication. In contrast, knockout of IFI6-16 reversed the IFN hypersensitivity of p7 mutant virus. In addition, p7 was found to be coimmunoprecipitated with IFI6-16 and to counteract the function of IFI6-16 by depolarizing the mitochondria potential. Our data suggest that p7 is a critical immune evasion protein that suppresses the antiviral IFN function by counteracting the function of IFI6-16.

  10. Adenovirus E1B 19-kilodalton protein modulates innate immunity through apoptotic mimicry.

    Science.gov (United States)

    Radke, Jay R; Grigera, Fernando; Ucker, David S; Cook, James L

    2014-03-01

    Cells that undergo apoptosis in response to chemical or physical stimuli repress inflammatory reactions, but cells that undergo nonapoptotic death in response to such stimuli lack this activity. Whether cells dying from viral infection exhibit a cell death-type modulatory effect on inflammatory reactions is unknown. We compared the effects on macrophage inflammatory responses of cells dying an apoptotic or a nonapoptotic death as a result of adenoviral infection. The results were exactly opposite to the predictions from the conventional paradigm. Cells dying by apoptosis induced by infection with an adenovirus type 5 (Ad5) E1B 19-kilodalton (E1B 19K) gene deletion mutant did not repress macrophage NF-κB activation or cytokine responses to proinflammatory stimuli, whereas cells dying a nonapoptotic death from infection with E1B 19K-competent, wild-type Ad5 repressed these macrophage inflammatory responses as well as cells undergoing classical apoptosis in response to chemical injury. The immunorepressive, E1B 19K-related cell death activity depended upon direct contact of the virally infected corpses with responder macrophages. Replacement of the viral E1B 19K gene with the mammalian Bcl-2 gene in cis restored the nonapoptotic, immunorepressive cell death activity of virally infected cells. These results define a novel function of the antiapoptotic, adenoviral E1B 19K protein that may limit local host innate immune inflammation during accumulation of virally infected cells at sites of infection and suggest that E1B 19K-deleted, replicating adenoviral vectors might induce greater inflammatory responses to virally infected cells than E1B 19K-positive vectors, because of the net effect of their loss-of-function mutation. We observed that cells dying a nonapoptotic cell death induced by adenovirus infection repressed macrophage proinflammatory responses while cells dying by apoptosis induced by infection with an E1B 19K deletion mutant virus did not repress macrophage

  11. The use of 125I-labelled protein A for the detection of humoral immunity of gross murine leukaemia virus

    International Nuclear Information System (INIS)

    Holmes, H.C.; Tuffrey, M.; Barnes, R.D.; Steuden, J.; Hilgers, J.

    1980-01-01

    A solid-phase radioimmunoassay utilising binding of 125 I-labelled protein A to antibodies bound to virus adsorbed onto microtitre plates was shown to be suitable for detection of humoral immunity to Gross murine leukaemia virus (MuLV). The specificity of the reaction was shown by the fact that only homologous or closely related viruses effectively inhibited binding of antibodies to adsorbed virus. With this method a low level of spontaneous humoral immunity was demonstrated in sera from AKR/Crc mice, a strain with high concentrations of endogenous virus, whereas little or no anti-viral activity was found in CBA/H-T6Crc, a subline that does not appear to express MuLV. (Auth.)

  12. Improvement of In Vivo Expression of Genes Delivered by Self-Amplifying RNA Using Vaccinia Virus Immune Evasion Proteins

    Science.gov (United States)

    Beissert, Tim; Koste, Lars; Perkovic, Mario; Walzer, Kerstin C.; Erbar, Stephanie; Selmi, Abderraouf; Diken, Mustafa; Kreiter, Sebastian; Türeci, Özlem; Sahin, Ugur

    2017-01-01

    Among nucleic acid–based delivery platforms, self-amplifying RNA (saRNA) vectors are of increasing interest for applications such as transient expression of recombinant proteins and vaccination. saRNA is safe and, due to its capability to amplify intracellularly, high protein levels can be produced from even minute amounts of transfected templates. However, it is an obstacle to full exploitation of this platform that saRNA induces a strong innate host immune response. In transfected cells, pattern recognition receptors sense double-stranded RNA intermediates and via activation of protein kinase R (PKR) and interferon signaling initiate host defense measures including a translational shutdown. To reduce pattern recognition receptor stimulation and unleash suppressed saRNA translation, this study co-delivered non-replicating mRNA encoding vaccinia virus immune evasion proteins E3, K3, and B18. It was shown that E3 is far superior to K3 or B18 as a highly potent blocker of PKR activation and of interferon (IFN)-β upregulation. B18, in contrast, is superior in controlling OAS1, a key IFN-inducible gene involved in viral RNA degradation. By combining all three vaccinia proteins, the study achieved significant suppression of PKR and IFN pathway activation in vitro and enhanced expression of saRNA-encoded genes of interest both in vitro and in vivo. This approach promises to overcome key hurdles of saRNA gene delivery. Its application may improve the bioavailability of the encoded protein, and reduce the effective dose and correspondingly the cost of goods of manufacture in the various fields where saRNA utilization is envisioned. PMID:28877647

  13. Improvement of In Vivo Expression of Genes Delivered by Self-Amplifying RNA Using Vaccinia Virus Immune Evasion Proteins.

    Science.gov (United States)

    Beissert, Tim; Koste, Lars; Perkovic, Mario; Walzer, Kerstin C; Erbar, Stephanie; Selmi, Abderraouf; Diken, Mustafa; Kreiter, Sebastian; Türeci, Özlem; Sahin, Ugur

    2017-12-01

    Among nucleic acid-based delivery platforms, self-amplifying RNA (saRNA) vectors are of increasing interest for applications such as transient expression of recombinant proteins and vaccination. saRNA is safe and, due to its capability to amplify intracellularly, high protein levels can be produced from even minute amounts of transfected templates. However, it is an obstacle to full exploitation of this platform that saRNA induces a strong innate host immune response. In transfected cells, pattern recognition receptors sense double-stranded RNA intermediates and via activation of protein kinase R (PKR) and interferon signaling initiate host defense measures including a translational shutdown. To reduce pattern recognition receptor stimulation and unleash suppressed saRNA translation, this study co-delivered non-replicating mRNA encoding vaccinia virus immune evasion proteins E3, K3, and B18. It was shown that E3 is far superior to K3 or B18 as a highly potent blocker of PKR activation and of interferon (IFN)-β upregulation. B18, in contrast, is superior in controlling OAS1, a key IFN-inducible gene involved in viral RNA degradation. By combining all three vaccinia proteins, the study achieved significant suppression of PKR and IFN pathway activation in vitro and enhanced expression of saRNA-encoded genes of interest both in vitro and in vivo. This approach promises to overcome key hurdles of saRNA gene delivery. Its application may improve the bioavailability of the encoded protein, and reduce the effective dose and correspondingly the cost of goods of manufacture in the various fields where saRNA utilization is envisioned.

  14. Heat shock proteins and experimental autoimmune encephalomyelitis (EAE): I. Immunization with a peptide of the myelin protein 2',3' cyclic nucleotide 3' phosphodiesterase that is cross-reactive with a heat shock protein alters the course of EAE.

    Science.gov (United States)

    Birnbaum, G; Kotilinek, L; Schlievert, P; Clark, H B; Trotter, J; Horvath, E; Gao, E; Cox, M; Braun, P E

    1996-05-15

    We describe sequence similarity and immunologic cross-reactivity between a peptide of the mycobacterial hsp, HSP65, and the myelin protein 2',3' cyclic nucleotide 3' phosphodiesterase (CNP). We demonstrate that immunization with the homologous cross-reactive CNP peptide (hsp-CNP peptide) has significant biological consequences. Rats immunized with hsp-CNP peptide in either complete Freund's adjuvant (CFA) or incomplete Freund's adjuvant (IFA) produce large amounts of peptide-specific antibody. Isotypes of antibodies in animals immunized with peptide in CFA are IgG1 and IgG2a. Isotypes of antibodies in rats immunized with peptide in IFA are predominantly IgG1, with low titers of IgG2a. T cell proliferative responses to HSP65 are present in rats immunized with peptide in CFA. T cell responses to HSP65 initially are absent in rats immunized with peptide in IFA but develop over time. T cell proliferative responses to hsp-CNP peptide were not detected. None of the groups of rats developed clinical or histologic evidence of experimental autoimmune encephalomyelitis (EAE). To induce EAE, rats preimmunized with hsp-CNP peptide were challenged with guinea pig spinal cord (GPSC) emulsified in CFA. Rats preimmunized with peptide in CFA developed severe EAE. Rats preimmunized with hsp-CNP peptide in IFA were protected from EAE, with both a lower incidence and severity of disease. Injecting the murine monoclonal antibody recognizing the shared HSP65 and CNP epitope did not protect against EAE. Our data suggest that a Th2 pattern of immune response to a CNP peptide that itself is non-encephalitogenic protects against EAE. Immune responses to either hsp or myelin proteins cross-reactive with hsp may play an important role in the development of EAE.

  15. Sublingual immunization with the phosphate-binding-protein (PstS) reduces oral colonization by Streptococcus mutans.

    Science.gov (United States)

    Ferreira, E L; Batista, M T; Cavalcante, R C M; Pegos, V R; Passos, H M; Silva, D A; Balan, A; Ferreira, L C S; Ferreira, R C C

    2016-10-01

    Bacterial ATP-binding cassette (ABC) transporters play a crucial role in the physiology and pathogenicity of different bacterial species. Components of ABC transporters have also been tested as target antigens for the development of vaccines against different bacterial species, such as those belonging to the Streptococcus genus. Streptococcus mutans is the etiological agent of dental caries, and previous studies have demonstrated that deletion of the gene encoding PstS, the substrate-binding component of the phosphate uptake system (Pst), reduced the adherence of the bacteria to abiotic surfaces. In the current study, we generated a recombinant form of the S. mutans PstS protein (rPstS) with preserved structural features, and we evaluated the induction of antibody responses in mice after sublingual mucosal immunization with a formulation containing the recombinant protein and an adjuvant derived from the heat-labile toxin from enterotoxigenic Escherichia coli strains. Mice immunized with rPstS exhibited systemic and secreted antibody responses, measured by the number of immunoglobulin A-secreting cells in draining lymph nodes. Serum antibodies raised in mice immunized with rPstS interfered with the adhesion of bacteria to the oral cavity of naive mice challenged with S. mutans. Similarly, mice actively immunized with rPstS were partially protected from oral colonization after challenge with the S. mutans NG8 strain. Therefore, our results indicate that S. mutans PstS is a potential target antigen capable of inducing specific and protective antibody responses after sublingual administration. Overall, these observations raise interesting perspectives for the development of vaccines to prevent dental caries. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  16. The p36 Isoform of Murine Cytomegalovirus m152 Protein Suffices for Mediating Innate and Adaptive Immune Evasion

    Science.gov (United States)

    Fink, Annette; Renzaho, Angeliqué; Reddehase, Matthias J.; Lemmermann, Niels A. W.

    2013-01-01

    The MHC-class I (MHC-I)-like viral (MHC-Iv) m152 gene product of murine cytomegalovirus (mCMV) was the first immune evasion molecule described for a member of the β-subfamily of herpesviruses as a paradigm for analogous functions of human cytomegalovirus proteins. Notably, by interacting with classical MHC-I molecules and with MHC-I-like RAE1 family ligands of the activatory natural killer (NK) cell receptor NKG2D, it inhibits presentation of antigenic peptides to CD8 T cells and the NKG2D-dependent activation of NK cells, respectively, thus simultaneously interfering with adaptive and innate immune recognition of infected cells. Although the m152 gene product exists in differentially glycosylated isoforms whose individual contributions to immune evasion are unknown, it has entered the scientific literature as m152/gp40, based on the quantitatively most prominent isoform but with no functional justification. By construction of a recombinant mCMV in which all three N-glycosylation sites are mutated (N61Q, N208Q, and N241Q), we show here that N-linked glycosylation is not essential for functional interaction of the m152 immune evasion protein with either MHC-I or RAE1. These data add an important functional detail to recent structural analysis of the m152/RAE1γ complex that has revealed N-glycosylations at positions Asn61 and Asn208 of m152 distant from the m152/RAE1γ interface. PMID:24351798

  17. Immunization with FSHβ fusion protein antigen prevents bone loss in a rat ovariectomy-induced osteoporosis model

    Energy Technology Data Exchange (ETDEWEB)

    Geng, Wenxin; Yan, Xingrong; Du, Huicong; Cui, Jihong; Li, Liwen, E-mail: liven@nwu.edu.cn; Chen, Fulin, E-mail: chenfl@nwu.edu.cn

    2013-05-03

    Highlights: •A GST-FSH fusion protein was successfully expressed in E. coli. •Immunization with GST-FSH antigen can raise high-titer anti-FSH polyclonal sera. •Anti-FSH polyclonal sera can neutralize osteoclastogenic effect of FSH in vitro. •FSH immunization can prevent bone loss in a rat osteoporosis model. -- Abstract: Osteoporosis, a metabolic bone disease, threatens postmenopausal women globally. Hormone replacement therapy (HTR), especially estrogen replacement therapy (ERT), is used widely in the clinic because it has been generally accepted that postmenopausal osteoporosis is caused by estrogen deficiency. However, hypogonadal α and β estrogen receptor null mice were only mildly osteopenic, and mice with either receptor deleted had normal bone mass, indicating that estrogen may not be the only mediator that induces osteoporosis. Recently, follicle-stimulating hormone (FSH), the serum concentration of which increases from the very beginning of menopause, has been found to play a key role in postmenopausal osteoporosis by promoting osteoclastogenesis. In this article, we confirmed that exogenous FSH can enhance osteoclast differentiation in vitro and that this effect can be neutralized by either an anti-FSH monoclonal antibody or anti-FSH polyclonal sera raised by immunizing animals with a recombinant GST-FSHβ fusion protein antigen. Moreover, immunizing ovariectomized rats with the GST-FSHβ antigen does significantly prevent trabecular bone loss and thereby enhance the bone strength, indicating that a FSH-based vaccine may be a promising therapeutic strategy to slow down bone loss in postmenopausal women.

  18. Partial protective effect of intranasal immunization with recombinant Toxoplasma gondii rhoptry protein 17 against toxoplasmosis in mice.

    Directory of Open Access Journals (Sweden)

    Hai-Long Wang

    Full Text Available Toxoplasma gondii (T. gondii is an obligate intracellular protozoan parasite that infects a variety of mammals, including humans. An effective vaccine for this parasite is therefore needed. In this study, RH strain T. gondii rhoptry protein 17 was expressed in bacteria as a fusion with glutathione S-transferase (GST and the recombinant proteins (rTgROP17 were purified via GST-affinity chromatography. BALB/c mice were nasally immunised with rTgROP17, and induction of immune responses and protection against chronic and lethal T. gondii infections were investigated. The results revealed that mice immunised with rTgROP17 produced high levels of specific anti-rTgROP17 IgGs and a mixed IgG1/IgG2a response of IgG2a predominance. The systemic immune response was associated with increased production of Th1 (IFN-γand IL-2 and Th2 (IL-4 cytokines, and enhanced lymphoproliferation (stimulation index, SI in the mice immunised with rTgROP17. Strong mucosal immune responses with increased secretion of TgROP17-specific secretory IgA (SIgA in nasal, vaginal and intestinal washes were also observed in these mice. The vaccinated mice displayed apparent protection against chronic RH strain infection as evidenced by their lower liver and brain parasite burdens (59.17% and 49.08%, respectively than those of the controls. The vaccinated mice also exhibited significant protection against lethal infection of the virulent RH strain (survival increased by 50% compared to the controls. Our data demonstrate that rTgROP17 can trigger strong systemic and mucosal immune responses against T. gondii and that ROP17 is a promising candidate vaccine for toxoplasmosis.

  19. Pertussis toxin targets the innate immunity through DAP12, FcRγ, and MyD88 adaptor proteins.

    Science.gov (United States)

    Phongsisay, Vongsavanh; Iizasa, Ei'ichi; Hara, Hiromitsu; Yoshida, Hiroki

    2017-04-01

    Activation of the innate immunity by adjuvants, such as pertussis toxin (PTX), in the presence of autoreactive lymphocytes and antigen mimicry is sufficient to trigger autoimmunity. Toll-like, C-type lectin, and immunglobulin-like receptors play an important role in the innate immunity by sensing a variety of microbial products through several adaptor proteins, including MyD88, DAP12, and FcRγ. This study investigated the interaction between PTX and innate immune components. The direct interactions between coated PTX and receptor-fusion proteins were examined using ELISA-based binding assays. Functionally, PTX-binding receptors could be classified into two groups: inhibition (DAP12-coupled TREM2, ITIM-bearing SIRPα, SIGNR1/SIGNR3/DCSIGN) and activation (MyD88-associated TLR4, DAP12-coupled LMIR5/CD300b, FcRγ-coupled LMIR8/CD300c, CLEC9A, MGL-1). DAP12, MyD88, and FcRγ were selected for further investigation. A comprehensive analysis of gene transcription showed that PTX up-regulated the expression of various inflammatory mediators. DAP12 deficiency resulted in reduction or enhancement of inflammatory responses in a cytokine-specific manner. PTX was able to activate the TREM2-DAP12 signalling pathway. PTX induced lower expression of inflammatory mediators in the absence of FcRγ alone and substantially lost its inflammatory capacity in the absence of both FcRγ and MyD88. PTX was able to activate the MyD88-NF-κB signalling pathway in the presence of TLR2 or TLR4. The inflammatory activity of PTX was completely lost by heating. These results demonstrate that PTX targets the innate immunity through DAP12, FcRγ, and MyD88 providing new insights into the immunobiology of PTX. Copyright © 2016 Elsevier GmbH. All rights reserved.

  20. The p36 isoform of murine cytomegalovirus m152 protein suffices for mediating innate and adaptive immune evasion.

    Science.gov (United States)

    Fink, Annette; Renzaho, Angeliqué; Reddehase, Matthias J; Lemmermann, Niels A W

    2013-12-16

    The MHC-class I (MHC-I)-like viral (MHC-Iv) m152 gene product of murine cytomegalovirus (mCMV) was the first immune evasion molecule described for a member of the β-subfamily of herpesviruses as a paradigm for analogous functions of human cytomegalovirus proteins. Notably, by interacting with classical MHC-I molecules and with MHC-I-like RAE1 family ligands of the activatory natural killer (NK) cell receptor NKG2D, it inhibits presentation of antigenic peptides to CD8 T cells and the NKG2D-dependent activation of NK cells, respectively, thus simultaneously interfering with adaptive and innate immune recognition of infected cells. Although the m152 gene product exists in differentially glycosylated isoforms whose individual contributions to immune evasion are unknown, it has entered the scientific literature as m152/gp40, based on the quantitatively most prominent isoform but with no functional justification. By construction of a recombinant mCMV in which all three N-glycosylation sites are mutated (N61Q, N208Q, and N241Q), we show here that N-linked glycosylation is not essential for functional interaction of the m152 immune evasion protein with either MHC-I or RAE1. These data add an important functional detail to recent structural analysis of the m152/RAE1g complex that has revealed N-glycosylations at positions Asn61 and Asn208 of m152 distant from the m152/RAE1g interface.

  1. Recombinant flagellin-PAL fusion protein of Legionella pneumophila induced cell-mediated and protective immunity against bacteremia in BALB/c mice.

    Science.gov (United States)

    Mohabati Mobarez, Ashraf; Ahmadrajabi, Roya; Khoramabadi, Nima; Salmanian, Ali Hatef

    2017-09-08

    We report a new recombinant fusion protein composed of full-length Legionella pneumophila flagellin A and peptidoglycan-associated lipoprotein (PAL), rFLA-PAL, capable of inducing protective immunity against L. pneumophila. The recombinant protein was over expressed in Escherichia coli strain BL21 (DE3) using pET-28a (+) expression vector (pET28a-flaA-pal) and purified by Ni 2+ exchange chromatography. Immunological properties of rFLA-PAL were assessed in a mouse model. Female BALB/c mice, immunized with rFLA-PAL, exhibited a rapid increase in serum antibody concentration against each of its protein portions. Furthermore, a strong activation of both innate and adaptive cell-mediated immunity was observed as indicated by antigen-specific splenocyte proliferation, IFN-γ and IL-12 production, and early production of TNF-α in the serum and in splenocyte cultures which were separately assessed against PAL and FLA. BALB/c mice were challenged with a lethal dose of L. pneumophila intravenously. In a 10-days follow-up after intravenous lethal challenge with L. pneumophila, a 100% survival rate was observed for mice immunized with rFLA-PAL, same as for those immunized with a sublethal dose of L. pneumophila. Based on the potent immune responses observed in mice immunized with rFLA-PAL, this recombinant fusion protein could be a potential vaccine candidate against the intracellular pathogen L. pneumophila.

  2. Polymorphism in liver-stage malaria vaccine candidate proteins: immune evasion and implications for vaccine design.

    Science.gov (United States)

    Flanagan, Katie L; Wilson, Kirsty L; Plebanski, Magdalena

    2016-01-01

    The pre-erythrocytic stage of infection by malaria parasites represents a key target for vaccines that aim to eradicate malaria. Two important broad immune evasion strategies that can interfere with vaccine efficacy include the induction of dendritic cell (DC) dysfunction and regulatory T cells (Tregs) by blood-stage malaria parasites, leading to inefficient priming of T cells targeting liver-stage infections. The parasite also uses 'surgical strike' strategies, whereby polymorphism in pre-erythrocytic antigens can interfere with host immunity. Specifically, we review how even single amino acid changes in T cell epitopes can lead to loss of binding to major histocompatibility complex (MHC), lack of cross-reactivity, or antagonism and immune interference, where simultaneous or sequential stimulation with related variants of the same T cell epitope can cause T cell anergy or the conversion of effector to immunosuppressive T cell phenotypes.

  3. Elicitation of strong immune responses by a DNA vaccine expressing a secreted form of hepatitis C virus envelope protein E2 in murine and porcine animal models

    DEFF Research Database (Denmark)

    Li, Yiping; Kang, H.N.; Babiuk, L.A.

    2006-01-01

    boosting with a recombinant E2 protein vaccine formulated with CpG ODN and 10% Emulsigen. The immunogenicity of HCV E2 vaccines was analyzed by ELISA for antibody responses, MTT assay for lymphocyte proliferation, ELISPOT for the number of interferon-gamma secreting cells, and cytotoxic T lymphocyte assays...... and shifted the immune response towards Th2-like ones in piglets. CONCLUSION: A DNA vaccine expressing a secreted form of HCV E2 protein elicited E2-specific immune responses in mice and piglets. Recombinant E2 protein vaccination following DNA immunization significantly increased the antibody response......AIM: To characterize the immunogenicity of a hepatitis C virus (HCV) E2 DNA vaccine alone or with a protein vaccine boost in murine and porcine animal models. METHODS: A DNA vaccine expressing a secreted form of HCV E2 protein was constructed and used to vaccinate mice and piglets with or without...

  4. Accelerated evolution of innate immunity proteins in social insects: adaptive evolution or relaxed constraint?

    Science.gov (United States)

    Harpur, Brock A; Zayed, Amro

    2013-07-01

    The genomes of eusocial insects have a reduced complement of immune genes-an unusual finding considering that sociality provides ideal conditions for disease transmission. The following three hypotheses have been invoked to explain this finding: 1) social insects are attacked by fewer pathogens, 2) social insects have effective behavioral or 3) novel molecular mechanisms for combating pathogens. At the molecular level, these hypotheses predict that canonical innate immune pathways experience a relaxation of selective constraint. A recent study of several innate immune genes in ants and bees showed a pattern of accelerated amino acid evolution, which is consistent with either positive selection or a relaxation of constraint. We studied the population genetics of innate immune genes in the honey bee Apis mellifera by partially sequencing 13 genes from the bee's Toll pathway (∼10.5 kb) and 20 randomly chosen genes (∼16.5 kb) sequenced in 43 diploid workers. Relative to the random gene set, Toll pathway genes had significantly higher levels of amino acid replacement mutations segregating within A. mellifera and fixed between A. mellifera and A. cerana. However, levels of diversity and divergence at synonymous sites did not differ between the two gene sets. Although we detect strong signs of balancing selection on the pathogen recognition gene pgrp-sa, many of the genes in the Toll pathway show signatures of relaxed selective constraint. These results are consistent with the reduced complement of innate immune genes found in social insects and support the hypothesis that some aspect of eusociality renders canonical innate immunity superfluous.

  5. Dietary Animal Plasma Proteins Improve the Intestinal Immune Response in Senescent Mice

    Directory of Open Access Journals (Sweden)

    Lluïsa Miró

    2017-12-01

    Full Text Available Increased life expectancy has promoted research on healthy aging. Aging is accompanied by increased non-specific immune activation (inflammaging which favors the appearance of several disorders. Here, we study whether dietary supplementation with spray-dried animal plasma (SDP, which has been shown to reduce the activation of gut-associated lymphoid tissue (GALT in rodents challenged by S. aureus enterotoxin B (SEB, and can also prevent the effects of aging on immune system homeostasis. We first characterized GALT in a mouse model of accelerated senescence (SAMP8 at different ages (compared to mice resistant to accelerated senescence; SAMR1. Second, we analyzed the SDP effects on GALT response to an SEB challenge in SAMP8 mice. In GALT characterization, aging increased the cell number and the percentage of activated Th lymphocytes in mesenteric lymph nodes and Peyer’s patches (all, p < 0.05, as well as the expression of IL-6 and TNF-α in intestinal mucosa (both, p < 0.05. With respect to GALT response to the SEB challenge, young mice showed increased expression of intestinal IL-6 and TNF-α, as well as lymphocyte recruitment and activation (all, p < 0.05. However, the immune response of senescent mice to the SEB challenge was weak, since SEB did not change cell recruitment or the percentage of activated Th lymphocytes. Mice supplemented with SDP showed improved capacity to respond to the SEB challenge, similar to the response of the young mice. These results indicate that senescent mice have an impaired mucosal immune response characterized by unspecific GALT activation and a weak specific immune response. SDP supplementation reduces non-specific basal immune activation, allowing for the generation of specific responses.

  6. Peanut protein structure, polyphenol content and immune response to peanut proteins in vivo are modulated by laccase

    NARCIS (Netherlands)

    Mihajlovic, L; Radosavljevic, J; Nordlund, E; Krstic, M; Bohn, T; Smit, Joost; Buchert, J; Cirkovic Velickovic, T

    2016-01-01

    Food texture can be improved by enzyme-mediated covalent cross-linking of different food components, such as proteins and carbohydrates. Cross-linking changes the biological and immunological properties of proteins and may change the sensitizing potential of food allergens. In this study we applied

  7. Plasmodium vivax Reticulocyte Binding Proteins Are Key Targets of Naturally Acquired Immunity in Young Papua New Guinean Children.

    Directory of Open Access Journals (Sweden)

    Camila T França

    2016-09-01

    Full Text Available Major gaps in our understanding of Plasmodium vivax biology and the acquisition of immunity to this parasite hinder vaccine development. P. vivax merozoites exclusively invade reticulocytes, making parasite proteins that mediate reticulocyte binding and/or invasion potential key vaccine or drug targets. While protein interactions that mediate invasion are still poorly understood, the P. vivax Reticulocyte-Binding Protein family (PvRBP is thought to be involved in P. vivax restricted host-cell selectivity.We assessed the binding specificity of five members of the PvRBP family (PvRBP1a, PvRBP1b, PvRBP2a, PvRBP2b, PvRBP2-P2 and a non-binding fragment of PvRBP2c to normocytes or reticulocytes. PvRBP2b was identified as the only reticulocyte-specific binder (P<0.001, whereas the others preferentially bound to normocytes (PvRBP1a/b P≤0.034, or showed comparable binding to both (PvRBP2a/2-P2, P = 0.38. Furthermore, we measured levels of total and IgG subclasses 1, 2, 3 and 4 to the six PvRBPs in a cohort of young Papua New Guinean children, and assessed their relationship with prospective risk of P. vivax malaria. Children had substantial, highly correlated (rho = 0.49-0.82, P<0.001 antibody levels to all six PvRBPs, with dominant IgG1 and IgG3 subclasses. Both total IgG (Incidence Rate Ratio [IRR] 0.63-0.73, P = 0.008-0.041 and IgG1 (IRR 0.56-0.69, P = 0.001-0.035 to PvRBP2b and PvRBP1a were strongly associated with reduced risk of vivax-malaria, independently of age and exposure.These results demonstrate a diversity of erythrocyte-binding phenotypes of PvRBPs, indicating binding to both reticulocyte-specific and normocyte-specific ligands. Our findings provide further insights into the naturally acquired immunity to P. vivax and highlight the importance of PvRBP proteins as targets of naturally acquired humoral immunity. In-depth studies of the role of PvRBPs in P. vivax invasion and functional validation of the role of anti-PvRBP antibodies in

  8. Novel Bat Influenza Virus NS1 Proteins Bind Double-Stranded RNA and Antagonize Host Innate Immunity.

    Science.gov (United States)

    Turkington, Hannah L; Juozapaitis, Mindaugas; Kerry, Philip S; Aydillo, Teresa; Ayllon, Juan; García-Sastre, Adolfo; Schwemmle, Martin; Hale, Benjamin G

    2015-10-01

    We demonstrate that novel bat HL17NL10 and HL18NL11 influenza virus NS1 proteins are effective interferon antagonists but do not block general host gene expression. Solving the RNA-binding domain structures revealed the canonical NS1 symmetrical homodimer, and RNA binding required conserved basic residues in this domain. Interferon antagonism was strictly dependent on RNA binding, and chimeric bat influenza viruses expressing NS1s defective in this activity were highly attenuated in interferon-competent cells but not in cells unable to establish antiviral immunity. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  9. The p36 Isoform of Murine Cytomegalovirus m152 Protein Suffices for Mediating Innate and Adaptive Immune Evasion

    OpenAIRE

    Fink, Annette; Renzaho, Angeliqué; Reddehase, Matthias J.; Lemmermann, Niels A. W.

    2013-01-01

    The MHC-class I (MHC-I)-like viral (MHC-Iv) m152 gene product of murine cytomegalovirus (mCMV) was the first immune evasion molecule described for a member of the β-subfamily of herpesviruses as a paradigm for analogous functions of human cytomegalovirus proteins. Notably, by interacting with classical MHC-I molecules and with MHC-I-like RAE1 family ligands of the activatory natural killer (NK) cell receptor NKG2D, it inhibits presentation of antigenic peptides to CD8 T cells and the NKG2D-d...

  10. Immune evasion by varicelloviruses : the identification of a new family of TAP-inhibiting proteins

    NARCIS (Netherlands)

    Koppers-Lalić, Danijela

    2007-01-01

    From the earliest times of their evolution, multi-cellular organisms have been defending themselves against infectious agents like nucleic acids, viruses, bacteria, fungi and parasites. Continuous selection pressure resulted in the development of sophisticated immune systems, which in their adaptive

  11. Inactivation of colicin Y by intramembrane helix–helix interaction with its immunity protein

    Czech Academy of Sciences Publication Activity Database

    Šmajs, D.; Doležalová, M.; Macek, Pavel; Žídek, L.

    2008-01-01

    Roč. 275, č. 21 (2008), s. 5325-5331 ISSN 1742-464X Institutional research plan: CEZ:AV0Z50200510 Keywords : colicin immunity * colicin y * helix-helix interaction Subject RIV: CE - Biochemistry Impact factor: 3.139, year: 2008

  12. Interleukin-17-induced protein lipocalin 2 is dispensable for immunity to oral candidiasis.

    Science.gov (United States)

    Ferreira, Maria Carolina; Whibley, Natasha; Mamo, Anna J; Siebenlist, Ulrich; Chan, Yvonne R; Gaffen, Sarah L

    2014-03-01

    Oropharyngeal candidiasis (OPC; thrush) is an opportunistic fungal infection caused by the commensal microbe Candida albicans. Immunity to OPC is strongly dependent on CD4+ T cells, particularly those of the Th17 subset. Interleukin-17 (IL-17) deficiency in mice or humans leads to chronic mucocutaneous candidiasis, but the specific downstream mechanisms of IL-17-mediated host defense remain unclear. Lipocalin 2 (Lcn2; 24p3; neutrophil gelatinase-associated lipocalin [NGAL]) is an antimicrobial host defense factor produced in response to inflammatory cytokines, particularly IL-17. Lcn2 plays a key role in preventing iron acquisition by bacteria that use catecholate-type siderophores, and lipocalin 2(-/-) mice are highly susceptible to infection by Escherichia coli and Klebsiella pneumoniae. The role of Lcn2 in mediating immunity to fungi is poorly defined. Accordingly, in this study, we evaluated the role of Lcn2 in immunity to oral infection with C. albicans. Lcn2 is strongly upregulated following oral infection with C. albicans, and its expression is almost entirely abrogated in mice with defective IL-17 signaling (IL-17RA(-/-) or Act1(-/-) mice). However, Lcn2(-/-) mice were completely resistant to OPC, comparably to wild-type (WT) mice. Moreover, Lcn2 deficiency mediated protection from OPC induced by steroid immunosuppression. Therefore, despite its potent regulation during C. albicans infection, Lcn2 is not required for immunity to mucosal candidiasis.

  13. Human Secretory Immune Response to Fatty Acid-Binding Protein Fraction from Giardia lamblia

    Science.gov (United States)

    Hasan, S. M. T.; Maachee, M.; Córdova, O. M.; Diaz de la Guardia, R.; Martins, M.; Osuna, A.

    2002-01-01

    The secretory immune response in humans infected with Giardia lamblia was studied by using saliva samples and an 8-kDa antigen capable of binding fatty acids. This antigen was not recognized by saliva samples from healthy individuals. The antigen may be useful in diagnostic studies of G. lamblia infection. PMID:11895992

  14. Human Secretory Immune Response to Fatty Acid-Binding Protein Fraction from Giardia lamblia

    OpenAIRE

    Hasan, S. M. T.; Maachee, M.; Córdova, O. M.; Diaz de la Guardia, R.; Martins, M.; Osuna, A.

    2002-01-01

    The secretory immune response in humans infected with Giardia lamblia was studied by using saliva samples and an 8-kDa antigen capable of binding fatty acids. This antigen was not recognized by saliva samples from healthy individuals. The antigen may be useful in diagnostic studies of G. lamblia infection.

  15. Intramammary Immunization of Pregnant Mice with Staphylococcal Protein A Reduces the Post-Challenge Mammary Gland Bacterial Load but Not Pathology.

    Directory of Open Access Journals (Sweden)

    Jully Gogoi-Tiwari

    Full Text Available Protein A, encoded by the spa gene, is one of the major immune evading MSCRAMM of S. aureus, demonstrated to be prevalent in a significant percentage of clinical bovine mastitis isolates in Australia. Given its' reported significance in biofilm formation and the superior performance of S. aureus biofilm versus planktonic vaccine in the mouse mastitis model, it was of interest to determine the immunogenicity and protective potential of Protein A as a potential vaccine candidate against bovine mastitis using the mouse mastitis model. Pregnant Balb/c mice were immunised with Protein A emulsified in an alum-based adjuvant by subcutaneous (s/c or intramammary (i/mam routes. While humoral immune response of mice post-immunization were determined using indirect ELISA, cell-mediated immune response was assessed by estimation of interferon-gamma (IFN-γ produced by protein A-stimulated splenocyte supernatants. Protective potential of Protein A against experimental mastitis was determined by challenge of immunized versus sham-vaccinated mice by i/mam route, based upon manifestation of clinical symptoms, total bacterial load and histopathological damage to mammary glands. Significantly (p<0.05 higher levels of IgG1 isotype were produced in mice immunized by the s/c route. In contrast, significantly higher levels of the antibody isotype IgG2a were produced in mice immunized by the i/mam route (p<0.05. There was significant reduction (p<0.05 in bacterial loads of the mammary glands of mice immunized by Protein A regardless of the route of immunization, with medium level of clinical symptoms observed up to day 3 post-challenge. However, Protein A vaccine failed to protect immunized mice post-challenge with biofilm producing encapsulated S. aureus via i/mam route, regardless of the route of immunization, as measured by the level of mammary tissue damage. It was concluded that, Protein A in its' native state was apparently not a suitable candidate for inclusion

  16. Comparison of potential protection conferred by three immunization strategies (protein/protein, DNA/DNA, and DNA/protein) against Brucella infection using Omp2b in BALB/c Mice.

    Science.gov (United States)

    Golshani, Maryam; Rafati, Sima; Nejati-Moheimani, Mehdi; Ghasemian, Melina; Bouzari, Saeid

    2016-12-25

    In the present study, immunogenicity and protective efficacy of the Brucella outer membrane protein 2b (Omp2b) was evaluated in BALB/c mice using Protein/Protein, DNA/DNA and DNA/Protein vaccine strategies. Immunization of mice with three vaccine regimens elicited a strong specific IgG response (higher IgG2a titers over IgG1 titers) and provided Th1-oriented immune response. Vaccination of BALB/c mice with the DNA/Pro regimen induced higher levels of IFN-γ/IL-2 and conferred more protection levels against B. melitenisis and B. abortus challenge than did the protein or DNA alone. In conclusion, Omp2b is able to stimulate specific immune responses and to confer cross protection against B. melitensis and B. abortus infection. Therefore, it could be introduced as a new potential candidate for the development of a subunit vaccine against Brucella infection. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. The construction of recombinant Lactobacillus casei expressing BVDV E2 protein and its immune response in mice.

    Science.gov (United States)

    Bhuyan, Anjuman Ara; Memon, Atta Muhammad; Bhuiyan, Ali Akbar; Zhonghua, Li; Zhang, Bingzhou; Ye, Shiyi; Mengying, Li; He, Qi-Gai

    2018-03-20

    Bovine viral diarrhea virus (BVDV) is the etiological agent of BVD causes substantial economic losses and endemic in world-wide cattle population. Mucosal immunity plays an important role in protection against BVDV infection and Lactobacillus casei is believed as an excellent live vaccine vector for expressing foreign genes. In this study, we have constructed a novel recombinant L. casei/pELX1-E2 strain expressing the most immunogenic E2 antigen of BVDV; using growth phage dependent surface expression system pELX1. The expression of E2 protein was verified by SDS-PAGE, Western blotting, and Immunofluorescence microscopic analysis. The immune responses triggered by the E2 producing recombinant L. casei were evaluated in BALB/c mice revealed that oral and intranasal (IN) administration of the recombinant strain was able to induce a significantly higher level of specific anti-E2 mucosal IgA and serum IgG as well as the greater level of cellular response by IFN-γ and IL-12 than those of intramuscular (IM) and control groups of mice. However, IN inoculation was found the most potent route of immunization. The ability of the recombinant strain to induce serum neutralizing antibody against BVDV and reduced viral load after viral challenge indicated better protection of BVDV infection. Therefore, this recombinant L. casei expressing E2 could be a safe and promising mucosal vaccine candidate against BVD. Copyright © 2018 Elsevier B.V. All rights reserved.

  18. Coated protein nanoclusters from influenza H7N9 HA are highly immunogenic and induce robust protective immunity.

    Science.gov (United States)

    Wang, Li; Chang, Timothy Z; He, Yuan; Kim, Jong R; Wang, Shelly; Mohan, Teena; Berman, Zachary; Tompkins, S Mark; Tripp, Ralph A; Compans, Richard W; Champion, Julie A; Wang, Bao-Zhong

    2017-01-01

    Recurring influenza viruses pose an annual threat to public health. A time-saving, cost-effective and egg-independent influenza vaccine approach is important particularly when responding to an emerging pandemic. We fabricated coated, two-layer protein nanoclusters from recombinant trimeric hemagglutinin from an avian-origin H7N9 influenza A virus as an approach for vaccine development in response to an emerging pandemic. Assessment of the virus-specific immune responses and protective efficacy in mice immunized with the nanoclusters demonstrated that the vaccine candidates were highly immunogenic, able to induce protective immunity and long-lasting humoral antibody responses to this virus without the use of adjuvants. Because the advantages of the highly immunogenic coated nanoclusters also include rapid productions in an egg-independent system, this approach has great potential for influenza vaccine production not only in response to an emerging pandemic, but also as a replacement for conventional seasonal influenza vaccines. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. Circulating immune complexes, immunoglobulin G, salivary proteins and salivary immunoglobulin A in patients with Sjögren's syndrome

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    Hadži-Mihailović Miloš

    2009-01-01

    Full Text Available Introduction. Sjögren's syndrome (SS is a chronic autoimmune disorder, with its major clinical manifestations resulting from changes in exocrine glands. Objective The aim of this study was to evaluate serum concentrations of circulating immune complexes (CIC and immunoglobulin G (IgG, and salivary proteins (SP and salivary immunoglobulin A (sIgA in 40 patients with SS, and to correlate these values among themselves, as well as with the unstimulated salivary flow rate (USFR and the duration of disease. Methods. The total of 40 patients were included in this research. CIC was determined using the solution of polyethylene glycol and IgG with the standard procedure of radial immunodiffusion. SP was investigated by the method of Lowry and sIgA was separated from the whole saliva using the method of immune chromatography. Results. The values of most of the studied parameters exceeded the normal range in a high degree: CIC 72.5%, IgG 70%, SP 80%. The concentrations of CIC were significantly higher in the patients with the duration of disease less than 10 years. With the decrease of USFR, the concentration of sIgA and IgG were increased with statistical significance. Conclusion The increased prevalence of abnormal values of CIC, IgG and SP indicate that the patients with SS have developed a higher level of immune reactivity. These results could be useful in diagnosis and disease activity monitoring.

  20. Protein Kinase C-theta (PKC-theta in Natural Killer (NK cell function and anti-tumor immunity

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    Alberto eAnel

    2012-07-01

    Full Text Available The protein kinase C-theta (PKCtheta, which is essential for T cell function and survival, is also required for efficient anti-tumor immune surveillance. Natural killer (NK cells, which express PKCtheta, play a prominent role in this process, mainly by elimination of tumor cells with reduced or absent major histocompatibility complex class-I (MHC-I expression. This justifies the increased interest of the use of activated NK cells in anti-tumor immunotherapy in the clinic. The in vivo development of MHC-I-deficient tumors is much favored in PKCtheta-/- mice compared with wild-type mice. Recent data offer some clues on the mechanism that could explain the important role of PKCtheta in NK cell-mediated anti-tumor immune surveillance: some studies show that PKCtheta is implicated in signal transduction and anti-tumoral activity of NK cells elicited by interleukin (IL-12 or IL-15, while others show that it is implicated in NK cell functional activation mediated by certain killer activating receptors (KAR. Alternatively, the possibility that PKCtheta is involved in NK cell degranulation is discussed, since recent data indicate that it is implicated in microtubule-organizing center (MTOC polarization to the immune synapse in CD4+ T cells. The implication of PKC isoforms in degranulation has been more extensively studied in CTL, and these studies will be also summarized.

  1. A novel lumazine synthase molecule from Brucella significantly promotes the immune-stimulation effects of antigenic protein.

    Science.gov (United States)

    Du, Z Q; Wang, J Y

    2015-10-27

    Brucella, an intracellular parasite that infects some livestock and humans, can damage or destroy the reproductive system of livestock. The syndrome is referred to as brucellosis and often occurs in pastoral areas; it is contagious from livestock to humans. In this study, the intact Brucella suis outer membrane protein 31 (omp31) gene was cloned, recombinantly expressed, and examined as a subunit vaccine candidate. The intact Brucella lumazine synthase (bls) gene was cloned and recombinantly expressed to study polymerization function in vitro. Non-reducing gel electrophoresis showed that rBs-BLS existed in different forms in vitro, including as a dimer and a pentamer. An enzyme-linked immunosorbent assay result showed that rOmp31 protein could induce production of an antibody in rabbits. However, the rOmp31-BLS fusion protein could elicit a much higher antibody titer in rabbits; this construct involved fusion of the Omp31 molecule with the BLS molecule. Our results indicate that Omp31 is involved in immune stimulation, while BLS has a polymerizing function based on rOmp31-BLS fusion protein immunogenicity. These data suggest that Omp31 is an ideal subunit vaccine candidate and that the BLS molecule is a favorable transport vector for antigenic proteins.

  2. Staphylococcus aureus surface protein SdrE binds complement regulator factor H as an immune evasion tactic.

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    Julia A Sharp

    Full Text Available Similar to other highly successful invasive bacterial pathogens, Staphylococcus aureus recruits the complement regulatory protein factor H (fH to its surface to inhibit the alternative pathway of complement. Here, we report the identification of the surface-associated protein SdrE as a fH-binding protein using purified fH overlay of S. aureus fractionated cell wall proteins and fH cross-linking to S. aureus followed by mass spectrometry. Studies using recombinant SdrE revealed that rSdrE bound significant fH whether from serum or as a purified form, in both a time- and dose-dependent manner. Furthermore, rSdrE-bound fH exhibited cofactor functionality for factor I (fI-mediated cleavage of C3b to iC3b which correlated positively with increasing amounts of fH. Expression of SdrE on the surface of the surrogate bacterium Lactococcus lactis enhanced recruitment of fH which resulted in increased iC3b generation. Moreover, surface expression of SdrE led to a reduction in C3-fragment deposition, less C5a generation, and reduced killing by polymorphonuclear cells. Thus, we report the first identification of a S. aureus protein associated with the staphylococcal surface that binds factor H as an immune evasion mechanism.

  3. The Recombinant Sea Urchin Immune Effector Protein, rSpTransformer-E1, Binds to Phosphatidic Acid and Deforms Membranes

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    Cheng Man Lun

    2017-05-01

    Full Text Available The purple sea urchin, Strongylocentrotus purpuratus, possesses a sophisticated innate immune system that functions without adaptive capabilities and responds to pathogens effectively by expressing the highly diverse SpTransformer gene family (formerly the Sp185/333 gene family. The swift gene expression response and the sequence diversity of SpTransformer cDNAs suggest that the encoded proteins have immune functions. Individual sea urchins can express up to 260 distinct SpTransformer proteins, and their diversity suggests that different versions may have different functions. Although the deduced proteins are diverse, they share an overall structure of a hydrophobic leader, a glycine-rich N-terminal region, a histidine-rich region, and a C-terminal region. Circular dichroism analysis of a recombinant SpTransformer protein, rSpTransformer-E1 (rSpTrf-E1 demonstrates that it is intrinsically disordered and transforms to α helical in the presence of buffer additives and binding targets. Although native SpTrf proteins are associated with the membranes of perinuclear vesicles in the phagocyte class of coelomocytes and are present on the surface of small phagocytes, they have no predicted transmembrane region or conserved site for glycophosphatidylinositol linkage. To determine whether native SpTrf proteins associate with phagocyte membranes through interactions with lipids, when rSpTrf-E1 is incubated with lipid-embedded nylon strips, it binds to phosphatidic acid (PA through both the glycine-rich region and the histidine-rich region. Synthetic liposomes composed of PA and phosphatidylcholine show binding between rSpTrf-E1 and PA by fluorescence resonance energy transfer, which is associated with leakage of luminal contents suggesting changes in lipid organization and perhaps liposome lysis. Interactions with liposomes also change membrane curvature leading to liposome budding, fusion, and invagination, which is associated with PA clustering induced

  4. The Recombinant Sea Urchin Immune Effector Protein, rSpTransformer-E1, Binds to Phosphatidic Acid and Deforms Membranes.

    Science.gov (United States)

    Lun, Cheng Man; Samuel, Robin L; Gillmor, Susan D; Boyd, Anthony; Smith, L Courtney

    2017-01-01

    The purple sea urchin, Strongylocentrotus purpuratus , possesses a sophisticated innate immune system that functions without adaptive capabilities and responds to pathogens effectively by expressing the highly diverse SpTransformer gene family (formerly the Sp185/333 gene family). The swift gene expression response and the sequence diversity of SpTransformer cDNAs suggest that the encoded proteins have immune functions. Individual sea urchins can express up to 260 distinct SpTransformer proteins, and their diversity suggests that different versions may have different functions. Although the deduced proteins are diverse, they share an overall structure of a hydrophobic leader, a glycine-rich N-terminal region, a histidine-rich region, and a C-terminal region. Circular dichroism analysis of a recombinant SpTransformer protein, rSpTransformer-E1 (rSpTrf-E1) demonstrates that it is intrinsically disordered and transforms to α helical in the presence of buffer additives and binding targets. Although native SpTrf proteins are associated with the membranes of perinuclear vesicles in the phagocyte class of coelomocytes and are present on the surface of small phagocytes, they have no predicted transmembrane region or conserved site for glycophosphatidylinositol linkage. To determine whether native SpTrf proteins associate with phagocyte membranes through interactions with lipids, when rSpTrf-E1 is incubated with lipid-embedded nylon strips, it binds to phosphatidic acid (PA) through both the glycine-rich region and the histidine-rich region. Synthetic liposomes composed of PA and phosphatidylcholine show binding between rSpTrf-E1 and PA by fluorescence resonance energy transfer, which is associated with leakage of luminal contents suggesting changes in lipid organization and perhaps liposome lysis. Interactions with liposomes also change membrane curvature leading to liposome budding, fusion, and invagination, which is associated with PA clustering induced by rSpTrf-E1

  5. Enhanced protection in mice induced by immunization with inactivated whole viruses compare to spike protein of middle east respiratory syndrome coronavirus.

    Science.gov (United States)

    Deng, Yao; Lan, Jiaming; Bao, Linlin; Huang, Baoying; Ye, Fei; Chen, Yingzhu; Yao, Yanfeng; Wang, Wenling; Qin, Chuan; Tan, Wenjie

    2018-04-04

    The persistent public health threat of infection with Middle East respiratory syndrome coronavirus (MERS-CoV) highlights the need for an effective and safe MERS-CoV vaccine. In this study, we prepared and vaccinated mice with either a Spike (S) protein or inactivated whole MERS-CoV (IV) with a combined adjuvant (alum+CpG) as a vaccine formulation. Similar levels of the anti-S protein IgG response and neutralizing activity were induced by both the S protein and IV vaccines. In addition, immune responses against three other structural proteins, the envelope (E), membrane (M), and nucleocapsid (N) proteins, were also detected in sera of mice that received IV. No antigen-specific T-cell immunity was detected after vaccination based on the interferon-γ ELISpot assay. Mice were transduced with Ad5-hDPP4 after the final immunization and were then challenged with MERS-CoV (1 × 10 5 plaque-forming units). Compared with the control group (adjuvant alone), mice immunized with the S protein or IV showed slightly lower pathological damage in the lung, as well as reduced antigen expression and lung virus titers. Mice that received IV formulations also showed increased protective immunity (almost no live virus was isolated from the lung). In conclusion, our data indicate that immunization with our IV formulation induced enhanced protection in mice compared to immunization with the S protein against MERS-CoV, which should be further tested in camels and clinical trials.

  6. The Interplay between Radioresistant Caco-2 Cells and the Immune System Increases Epithelial Layer Permeability and Alters Signaling Protein Spectrum

    Science.gov (United States)

    Morini, Jacopo; Babini, Gabriele; Barbieri, Sofia; Baiocco, Giorgio; Ottolenghi, Andrea

    2017-01-01

    Colorectal cancer is one of the most frequent type of cancer, with a higher incidence in the developed countries. Colorectal cancer is usually managed with both surgeries, chemotherapy and radiotherapy. Radiotherapy has the well-known advantage of targeting the tumor, minimizing normal tissue exposure. Nevertheless, during radiation treatment, exposure of healthy tissues is of great concern, in particular because of the effects on the intestinal barrier functions and on cells belonging to the immune system. The functional role of intestinal barrier in avoiding paracellular trafficking and controlling bacterial spread from gut it is well known and it is due to the presence of tight junction complexes. However, intestinal barrier is fundamental in participating to the interplay with immune system, especially considering the gut-associated lymphoid tissue. Until few years ago, radiotherapy was considered to bear only a depressive action on the immune system. However, it is now recognized that the release of pro-inflammatory signals and phenotypic changes in tumoral cells due to ionizing radiation could trigger the immune system against the tumor. In this work, we address how intestinal barrier functions are perturbed by X-ray doses in the range 0–10 Gy, focusing on the interplay between tumoral cells and the immune system. To this aim, we adopted a coculture model in which Caco-2 cells can be grown in presence/absence of peripheral blood mononuclear cells (PBMC). We focused our attention on changes in the proliferation, trans-epithelial electrical resistance (TEER), cytokine release, and proteins of the junctional complexes. Our results indicate a high radioresistance of Caco-2 in the investigated dose range, and an increased permeability of the tumoral cell layer due to the presence of PBMC. This is found to be correlated with activation of PBMC, inhibiting the apoptotic pathway, with the enhancement of cytokine release and with variation of tight junction

  7. Genetically modified anthrax lethal toxin safely delivers whole HIV protein antigens into the cytosol to induce T cell immunity

    Science.gov (United States)

    Lu, Yichen; Friedman, Rachel; Kushner, Nicholas; Doling, Amy; Thomas, Lawrence; Touzjian, Neal; Starnbach, Michael; Lieberman, Judy

    2000-07-01

    Bacillus anthrax lethal toxin can be engineered to deliver foreign proteins to the cytosol for antigen presentation to CD8 T cells. Vaccination with modified toxins carrying 8-9 amino acid peptide epitopes induces protective immunity in mice. To evaluate whether large protein antigens can be used with this system, recombinant constructs encoding several HIV antigens up to 500 amino acids were produced. These candidate HIV vaccines are safe in animals and induce CD8 T cells in mice. Constructs encoding gag p24 and nef stimulate gag-specific CD4 proliferation and a secondary cytotoxic T lymphocyte response in HIV-infected donor peripheral blood mononuclear cells in vitro. These results lay the foundation for future clinical vaccine studies.

  8. Regulation of Toll-like receptor 4-mediated immune responses through Pasteurella multocida toxin-induced G protein signalling

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    Hildebrand Dagmar

    2012-08-01

    Full Text Available Abstract Background Lipopolysaccharide (LPS-triggered Toll-like receptor (TLR 4-signalling belongs to the key innate defence mechanisms upon infection with Gram-negative bacteria and triggers the subsequent activation of adaptive immunity. There is an active crosstalk between TLR4-mediated and other signalling cascades to secure an effective immune response, but also to prevent excessive inflammation. Many pathogens induce signalling cascades via secreted factors that interfere with TLR signalling to modify and presumably escape the host response. In this context heterotrimeric G proteins and their coupled receptors have been recognized as major cellular targets. Toxigenic strains of Gram-negative Pasteurella multocida produce a toxin (PMT that constitutively activates the heterotrimeric G proteins Gαq, Gα13 and Gαi independently of G protein-coupled receptors through deamidation. PMT is known to induce signalling events involved in cell proliferation, cell survival and cytoskeleton rearrangement. Results Here we show that the activation of heterotrimeric G proteins through PMT suppresses LPS-stimulated IL-12p40 production and eventually impairs the T cell-activating ability of LPS-treated monocytes. This inhibition of TLR4-induced IL-12p40 expression is mediated by Gαi-triggered signalling as well as by Gβγ-dependent activation of PI3kinase and JNK. Taken together we propose the following model: LPS stimulates TLR4-mediated activation of the NFĸB-pathway and thereby the production of TNF-α, IL-6 and IL-12p40. PMT inhibits the production of IL-12p40 by Gαi-mediated inhibition of adenylate cyclase and cAMP accumulation and by Gβγ-mediated activation of PI3kinase and JNK activation. Conclusions On the basis of the experiments with PMT this study gives an example of a pathogen-induced interaction between G protein-mediated and TLR4-triggered signalling and illustrates how a bacterial toxin is able to interfere with the host’s immune

  9. The impact of RGS and other G-protein regulatory proteins on Gαi-mediated signaling in immunity.

    Science.gov (United States)

    Kehrl, John H

    2016-08-15

    Leukocyte chemoattractant receptors are members of the G-protein coupled receptor (GPCR) family. Signaling downstream of these receptors directs the localization, positioning and homeostatic trafficking of leukocytes; as well as their recruitment to, and their retention at, inflammatory sites. Ligand induced changes in the molecular conformation of chemoattractant receptors results in the engagement of heterotrimeric G-proteins, which promotes α subunits to undergo GTP/GDP exchange. This results in the functional release of βγ subunits from the heterotrimers, thereby activating downstream effector molecules, which initiate leukocyte polarization, gradient sensing, and directional migration. Pertussis toxin ADP ribosylates Gαi subunits and prevents chemoattractant receptors from triggering Gαi nucleotide exchange. The use of pertussis toxin revealed the essential importance of Gαi subunit nucleotide exchange for chemoattractant receptor signaling. More recent studies have identified a range of regulatory mechanisms that target these receptors and their associated heterotrimeric G-proteins, thereby helping to control the magnitude, kinetics, and duration of signaling. A failure in these regulatory pathways can lead to impaired receptor signaling and immunopathology. The analysis of mice with targeted deletions of Gαi isoforms as well as some of these G-protein regulatory proteins is providing insights into their roles in chemoattractant receptor signaling. Published by Elsevier Inc.

  10. Evolution of the Immune Response against Recombinant Proteins (TcpA, TcpB, and FlaA as a Candidate Subunit Cholera Vaccine

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    Neda Molaee

    2017-01-01

    Full Text Available Vibrio cholerae is the causative agent of cholera and annually leads to death of thousands of people around the globe. Two factors in the pathogenesis of this bacterium are its pili and flagella. The main subunits of pili TcpA, TcpB, and FlaA are the constituent subunit of flagella. In this study, we studied the ability of pili and flagella subunits to stimulate immune responses in mice. After amplification of TcpA, TcpB, and FlaA genes using PCR, they were cloned in expression plasmids. After production of the above-mentioned proteins by using IPTG, the proteins were purified and then approved using immunoblot method. After injection of the purified proteins to a mice model, immune response stimulation was evaluated by measuring the levels of IgG1 and IgG2a antibody titers, IL5 and IFN-γ. Immune response stimulation against pili and flagella antigens was adequate. Given the high levels of IL5 titer and IgG1 antibody, the stimulated immune response was toward Th1. Humoral immune response stimulation is of key importance in prevention of cholera. Our immunological analysis shows the appropriate immune response in mice model after vaccination with recombinant proteins. The high level of IL5 and low level of IFN-γ show the activation of Th2 cell response.

  11. Signal transduction of Helicobacter pylori during interaction with host cell protein receptors of epithelial and immune cells

    Science.gov (United States)

    Pachathundikandi, Suneesh Kumar; Tegtmeyer, Nicole; Backert, Steffen

    2013-01-01

    Helicobacter pylori infections can induce pathologies ranging from chronic gastritis, peptic ulceration to gastric cancer. Bacterial isolates harbor numerous well-known adhesins, vacuolating cytotoxin VacA, protease HtrA, urease, peptidoglycan, and type IV secretion systems (T4SS). It appears that H. pylori targets more than 40 known host protein receptors on epithelial or immune cells. A series of T4SS components such as CagL, CagI, CagY, and CagA can bind to the integrin α5β1 receptor. Other targeted membrane-based receptors include the integrins αvβ3, αvβ5, and β2 (CD18), RPTP-α/β, GP130, E-cadherin, fibronectin, laminin, CD46, CD74, ICAM1/LFA1, T-cell receptor, Toll-like receptors, and receptor tyrosine kinases EGFR, ErbB2, ErbB3, and c-Met. In addition, H. pylori is able to activate the intracellular receptors NOD1, NOD2, and NLRP3 with important roles in innate immunity. Here we review the interplay of various bacterial factors with host protein receptors. The contribution of these interactions to signal transduction and pathogenesis is discussed. PMID:24280762

  12. Binding of the Fap2 protein of Fusobacterium nucleatum to human inhibitory receptor TIGIT protects tumors from immune cell attack.

    Science.gov (United States)

    Gur, Chamutal; Ibrahim, Yara; Isaacson, Batya; Yamin, Rachel; Abed, Jawad; Gamliel, Moriya; Enk, Jonatan; Bar-On, Yotam; Stanietsky-Kaynan, Noah; Coppenhagen-Glazer, Shunit; Shussman, Noam; Almogy, Gideon; Cuapio, Angelica; Hofer, Erhard; Mevorach, Dror; Tabib, Adi; Ortenberg, Rona; Markel, Gal; Miklić, Karmela; Jonjic, Stipan; Brennan, Caitlin A; Garrett, Wendy S; Bachrach, Gilad; Mandelboim, Ofer

    2015-02-17

    Bacteria, such as Fusobacterium nucleatum, are present in the tumor microenvironment. However, the immunological consequences of intra-tumoral bacteria remain unclear. Here, we have shown that natural killer (NK) cell killing of various tumors is inhibited in the presence of various F. nucleatum strains. Our data support that this F. nucleatum-mediated inhibition is mediated by human, but not by mouse TIGIT, an inhibitory receptor present on all human NK cells and on various T cells. Using a library of F. nucleatum mutants, we found that the Fap2 protein of F. nucleatum directly interacted with TIGIT, leading to the inhibition of NK cell cytotoxicity. We have further demonstrated that tumor-infiltrating lymphocytes expressed TIGIT and that T cell activities were also inhibited by F. nucleatum via Fap2. Our results identify a bacterium-dependent, tumor-immune evasion mechanism in which tumors exploit the Fap2 protein of F. nucleatum to inhibit immune cell activity via TIGIT. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Immunization with the Recombinant Cholera Toxin B Fused to Fimbria 2 Protein Protects against Bordetella pertussis Infection

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    Noelia Olivera

    2014-01-01

    Full Text Available This study examined the immunogenic properties of the fusion protein fimbria 2 of Bordetella pertussis (Fim2—cholera toxin B subunit (CTB in the intranasal murine model of infection. To this end B. pertussis Fim2 coding sequence was cloned downstream of the cholera toxin B subunit coding sequence. The expression and assembly of the fusion protein into pentameric structures (CTB-Fim2 were evaluated by SDS-PAGE and monosialotetrahexosylgaglioside (GM1-ganglioside enzyme-linked immunosorbent assay (ELISA. To evaluate the protective capacity of CTB-Fim2, an intraperitoneal or intranasal mouse immunization schedule was performed with 50 μg of CTB-Fim2. Recombinant (rFim2 or purified (BpFim2 Fim2, CTB, and phosphate-buffered saline (PBS were used as controls. The results showed that mice immunized with BpFim2 or CTB-Fim2 intraperitoneally or intranasally presented a significant reduction in bacterial lung counts compared to control groups (P<0.01 or P<0.001, resp.. Moreover, intranasal immunization with CTB-Fim2 induced significant levels of Fim2-specific IgG in serum and bronchoalveolar lavage (BAL and Fim2-specific IgA in BAL. Analysis of IgG isotypes and cytokines mRNA levels showed that CTB-Fim2 results in a mixed Th1/Th2 (T-helper response. The data presented here provide support for CTB-Fim2 as a promising recombinant antigen against Bordetella pertussis infection.

  14. Disruption of Protein Mannosylation Affects Candida guilliermondii Cell Wall, Immune Sensing, and Virulence

    OpenAIRE

    Navarro-Arias, María J.; Defosse, Tatiana A.; Dementhon, Karine; Csonka, Katalin; Mellado-Mojica, Erika; Dias Valério, Aline; González-Hernández, Roberto J.; Courdavault, Vincent; Clastre, Marc; Hernández, Nahúm V.; Pérez-García, Luis A.; Singh, Dhirendra K.; Vizler, Csaba; Gácser, Attila; Almeida, Ricardo S.

    2016-01-01

    The fungal cell wall contains glycoproteins that interact with the host immune system. In the prominent pathogenic yeast Candida albicans, Pmr1 acts as a Golgi-resident ion pump that provides cofactors to mannosyltransferases, regulating the synthesis of mannans attached to glycoproteins. To gain insight into a putative conservation of such a crucial process within opportunistic yeasts, we were particularly interested in studying the role of the PMR1 homolog in a low-virulent species that rar...

  15. Genome-Wide Profiling of Humoral Immune Response to Coxiella burnetii Infection by Protein Microarray

    Science.gov (United States)

    2010-01-01

    study of brucellosis in Lima, Peru and were approved by the Comite de Etica of Universidad Peruana Cayetano Heredia, Lima, Peru and the Comite de...be targeted by the early humoral immune response in vaccinated cattle [33], and in acutely infected guinea pigs of the Nine Mile strain in phase I...34]. ELISA-based assays using CBU1910 were able to distinguish vaccinated cattle from those naturally exposed [33]. Moreover. CBU1910 vaccinated

  16. CpG oligodeoxynucleotide-enhanced humoral immune response and production of antibodies to prion protein PrPSc in mice immunized with 139A scrapie-associated fibrils.

    Science.gov (United States)

    Spinner, Daryl S; Kascsak, Regina B; Lafauci, Giuseppe; Meeker, Harry C; Ye, Xuemin; Flory, Michael J; Kim, Jae Il; Schuller-Levis, Georgia B; Levis, William R; Wisniewski, Thomas; Carp, Richard I; Kascsak, Richard J

    2007-06-01

    Prion diseases are characterized by conversion of the cellular prion protein (PrP(C)) to a protease-resistant conformer, the srapie form of PrP (PrP(Sc)). Humoral immune responses to nondenatured forms of PrP(Sc) have never been fully characterized. We investigated whether production of antibodies to PrP(Sc) could occur in PrP null (Prnp(-/-)) mice and further, whether innate immune stimulation with the TLR9 agonist CpG oligodeoxynucleotide (ODN) 1826 could enhance this process. Whether such stimulation could raise anti-PrP(Sc) antibody levels in wild-type (Prnp(+/+)) mice was also investigated. Prnp(-/-) and Prnp(+/+) mice were immunized with nondenatured 139A scrapie-associated fibrils (SAF), with or without ODN 1826, and were tested for titers of PrP-specific antibodies. In Prnp(-/-) mice, inclusion of ODN 1826 in the immunization regime increased anti-PrP titers more than 13-fold after two immunizations and induced, among others, antibodies to an N-terminal epitope, which were only present in the immune repertoire of mice receiving ODN 1826. mAb 6D11, derived from such a mouse, reacts with the N-terminal epitope QWNK in native and denatured forms of PrP(Sc) and recombinant PrP and exhibits a K(d) in the 10(-)(11) M range. In Prnp(+/+) mice, ODN 1826 increased anti-PrP levels as much as 84% after a single immunization. Thus, ODN 1826 potentiates adaptive immune responses to PrP(Sc) in 139A SAF-immunized mice. These results represent the first characterization of humoral immune responses to nondenatured, infectious PrP(Sc) and suggest methods for optimizing the generation of mAbs to PrP(Sc), many of which could be used for diagnosis and treatment of prion diseases.

  17. Immunization of a wild koala population with a recombinant Chlamydia pecorum Major Outer Membrane Protein (MOMP) or Polymorphic Membrane Protein (PMP) based vaccine: New insights into immune response, protection and clearance.

    Science.gov (United States)

    Desclozeaux, Marion; Robbins, Amy; Jelocnik, Martina; Khan, Shahneaz Ali; Hanger, Jon; Gerdts, Volker; Potter, Andrew; Polkinghorne, Adam; Timms, Peter

    2017-01-01

    We assessed the effects of two different single-dose anti-Chlamydia pecorum (C. pecorum) vaccines (containing either Major Outer Membrane Protein (3MOMP) or Polymorphic Membrane Protein (Pmp) as antigens) on the immune response of a group of wild koalas. Both vaccines elicited a systemic humoral response as seen by the production of anti-chlamydial IgG antibodies in more than 90% of vaccinated koalas. A mucosal immune response was also observed, with an increase in Chlamydia-specific mucosal IgG and/or IgA antibodies in some koalas post-vaccination. Both vaccines elicited a cell-mediated immune response as measured by the production of the cytokines IFN-γ and IL-17 post-vaccination. To determine the level of protection provided by the vaccines under natural conditions we assessed C. pecorum infection loads and chlamydial disease status of all vaccinated koalas pre- and post-vaccination, compared to a non-vaccinated cohort from the same habitat. The MOMP vaccinated koalas that were infected on the day of vaccination showed significant clearance of their infection at 6 months post-vaccination. In contrast, the number of new infections in the PMP vaccine was similar to the control group, with some koalas progressing to disease. Genotyping of the ompA gene from the C. pecorum strains infecting the vaccinated animals, identified genetic variants of ompA-F genotype and a new genotype ompA-O. We found that those animals that were the least well protected became infected with strains of C. pecorum not covered by the vaccine. In conclusion, a single dose vaccine formulated with either recombinant PmpG or MOMP can elicit both cell-mediated and humoral (systemic and mucosal) immune responses, with the MOMP vaccine showing clearance of infection in all infected koalas. Although the capability of our vaccines to stimulate an adaptive response and be protective needs to be fully evaluated, this work illustrates the necessity to combine epitopes most relevant to a large panel of

  18. From immune response to cancer : a spot on the low molecular weight protein tyrosine phosphatase

    NARCIS (Netherlands)

    Souza, A. C. S.; Azoubel, S.; Queiroz, K. C. S.; Peppelenbosch, M. P.; Ferreira, C. V.

    Reversible tyrosine phosphorylation is a key posttranslational regulatory modification of proteins in all eukaryotic cells in normal and pathological processes. Recently a pivotal janus-faced biological role of the low molecular weight protein tyrosine phosphatase (LMWPTP) has become clear. On the

  19. Peptide fragments induce a more rapid immune response than intact proteins in earthworms.

    Science.gov (United States)

    Hanusová, R; Tucková, L; Halada, P; Bezouska, K; Bilej, M

    1999-01-01

    The effect of in vivo proteolytic processing of protein antigen was studied in Eisenia foetida earthworms. Parenteral administration of the protein antigen induces elevated levels of an antigen-binding protein (ABP) which recognizes the protein used for stimulation. When the protein antigen is administered simultaneously with nontoxic serine proteinase inhibitor, ABP levels remain close to background. On the other hand, the in vivo adaptive response of earthworms to peptide fragments obtained by coelomic fluid digestion of the foreign antigen occurs even in the presence of proteinase inhibitor and, moreover, is significantly faster as compared to the response to intact antigen. These findings confirm the role of proteolytic processing in earthworms. MALDI mass spectrometric analysis of the fragments after coelomic fluid digestion has revealed the presence of the peptide fragments with molecular weights in the mass range 700-1100 Da.

  20. Identifying protein biomarkers in predicting disease severity of dengue virus infection using immune-related protein microarray.

    Science.gov (United States)

    Soe, Hui Jen; Yong, Yean K; Al-Obaidi, Mazen M Jamil; Raju, Chandramathi Samudi; Gudimella, Ranganath; Manikam, Rishya; Sekaran, Shamala Devi

    2018-02-01

    Dengue virus is one of the most widespread flaviviruses that re-emerged throughout recent decades. The progression from mild dengue to severe dengue (SD) with the complications such as vascular leakage and hemorrhage increases the fatality rate of dengue. The pathophysiology of SD is not entirely clear. To investigate potential biomarkers that are suggestive of pathogenesis of SD, a small panel of serum samples selected from 1 healthy individual, 2 dengue patients without warning signs (DWS-), 2 dengue patients with warning signs (DWS+), and 5 patients with SD were subjected to a pilot analysis using Sengenics Immunome protein array. The overall fold changes of protein expressions and clustering heat map revealed that PFKFB4, TPM1, PDCL3, and PTPN20A were elevated among patients with SD. Differential expression analysis identified that 29 proteins were differentially elevated greater than 2-fold in SD groups than DWS- and DWS+. From the 29 candidate proteins, pathways enrichment analysis also identified insulin signaling and cytoskeleton pathways were involved in SD, suggesting that the insulin pathway may play a pivotal role in the pathogenesis of SD.

  1. The structure of a contact-dependent growth-inhibition (CDI) immunity protein from Neisseria meningitidis MC58

    Energy Technology Data Exchange (ETDEWEB)

    Tan, Kemin; Johnson, Parker M.; Stols, Lucy; Boubion, Bryan; Eschenfeldt, William; Babnigg, Gyorgy; Hayes, Christopher S.; Joachimiak, Andrezj; Goulding, Celia W.

    2015-05-20

    Contact-dependent growth inhibition (CDI) is an important mechanism of intercellular competition between neighboring Gram-negative bacteria. CDI systems encode large surface-exposed CdiA effector proteins that carry a variety of C-terminal toxin domains (CdiA-CTs). All CDI+bacteria also produce CdiI immunity proteins that specifically bind to the cognate CdiA-CT and neutralize its toxin activity to prevent auto-inhibition. Here, the X-ray crystal structure of a CdiI immunity protein fromNeisseria meningitidisMC58 is presented at 1.45 Å resolution. The CdiI protein has structural homology to the Whirly family of RNA-binding proteins, but appears to lack the characteristic nucleic acid-binding motif of this family. Sequence homology suggests that the cognate CdiA-CT is related to the eukaryotic EndoU family of RNA-processing enzymes. A homology model is presented of the CdiA-CT based on the structure of the XendoU nuclease fromXenopus laevis. Molecular-docking simulations predict that the CdiA-CT toxin active site is occluded upon binding to the CdiI immunity protein. Together, these observations suggest that the immunity protein neutralizes toxin activity by preventing access to RNA substrates.

  2. Increasing the immune activity of exosomes: the effect of miRNA-depleted exosome proteins on activating dendritic cell/cytokine-induced killer cells against pancreatic cancer* #

    Science.gov (United States)

    Que, Ri-sheng; Lin, Cheng; Ding, Guo-ping; Wu, Zheng-rong; Cao, Li-ping

    2016-01-01

    Background: Tumor-derived exosomes were considered to be potential candidates for tumor vaccines because they are abundant in immune-regulating proteins, whereas tumor exosomal miRNAs may induce immune tolerance, thereby having an opposite immune function. Objective: This study was designed to separate exosomal protein and depleted exosomal microRNAs (miRNAs), increasing the immune activity of exosomes for activating dendritic cell/cytokine-induced killer cells (DC/CIKs) against pancreatic cancer (PC). Methods: PC-derived exosomes (PEs) were extracted from cultured PANC-1 cell supernatants and then ruptured; this was followed by ultrafiltered exosome lysates (UELs). DCs were stimulated with lipopolysaccharide (LPS), PE, and UEL, followed by co-culture with CIKs. The anti-tumor effects of DC/CIKs against PC were evaluated by proliferation and killing rates, tumor necrosis factor-α (TNF-α) and perforin secretion. Exosomal miRNAs were depleted after lysis and ultrafiltration, while 128 proteins were retained, including several immune-activating proteins. Results: UEL-stimulated DC/CIKs showed a higher killing rate than LPS- and PE-stimulated DC/CIKs. Conclusions: miRNA-depleted exosome proteins may be promising agonists for specifically activating DC/CIKs against PC. PMID:27143262

  3. Increasing the immune activity of exosomes: the effect of miRNA-depleted exosome proteins on activating dendritic cell/cytokine-induced killer cells against pancreatic cancer.

    Science.gov (United States)

    Que, Ri-Sheng; Lin, Cheng; Ding, Guo-Ping; Wu, Zheng-Rong; Cao, Li-Ping

    2016-05-01

    Tumor-derived exosomes were considered to be potential candidates for tumor vaccines because they are abundant in immune-regulating proteins, whereas tumor exosomal miRNAs may induce immune tolerance, thereby having an opposite immune function. This study was designed to separate exosomal protein and depleted exosomal microRNAs (miRNAs), increasing the immune activity of exosomes for activating dendritic cell/cytokine-induced killer cells (DC/CIKs) against pancreatic cancer (PC). PC-derived exosomes (PEs) were extracted from cultured PANC-1 cell supernatants and then ruptured; this was followed by ultrafiltered exosome lysates (UELs). DCs were stimulated with lipopolysaccharide (LPS), PE, and UEL, followed by co-culture with CIKs. The anti-tumor effects of DC/CIKs against PC were evaluated by proliferation and killing rates, tumor necrosis factor-α (TNF-α) and perforin secretion. Exosomal miRNAs were depleted after lysis and ultrafiltration, while 128 proteins were retained, including several immune-activating proteins. UEL-stimulated DC/CIKs showed a higher killing rate than LPS- and PE-stimulated DC/CIKs. miRNA-depleted exosome proteins may be promising agonists for specifically activating DC/CIKs against PC.

  4. Recombinant Protein Containing B-Cell Epitopes of Different Loxosceles Spider Toxins Generates Neutralizing Antibodies in Immunized Rabbits

    Directory of Open Access Journals (Sweden)

    Sabrina de Almeida Lima

    2018-04-01

    Full Text Available Loxoscelism is the most important form of araneism in South America. The treatment of these accidents uses heterologous antivenoms obtained from immunization of production animals with crude loxoscelic venom. Due to the scarcity of this immunogen, new alternatives for its substitution in antivenom production are of medical interest. In the present work, three linear epitopes for Loxosceles astacin-like protease 1 (LALP-1 (SLGRGCTDFGTILHE, ENNTRTIGPFDYDSIMLYGAY, and KLYKCPPVNPYPGGIRPYVNV and two for hyaluronidase (LiHYAL (NGGIPQLGDLKAHLEKSAVDI and ILDKSATGLRIIDWEAWR from Loxosceles intermedia spider venom were identified by SPOT-synthesis technique. One formerly characterized linear epitope (DFSGPYLPSLPTLDA of sphingomyelinase D (SMase D SMase-I from Loxosceles laeta was also chosen to constitute a new recombinant multiepitopic protein. These epitopes were combined with a previously produced chimeric multiepitopic protein (rCpLi composed by linear and conformational B-cell epitopes from SMase D from L. intermedia venom, generating a new recombinant multiepitopic protein derived from loxoscelic toxins (rMEPLox. We demonstrated that rMEPLox is non-toxic and antibodies elicited in rabbits against this antigen present reactivity in ELISA and immunoblot assays with Brazilian L. intermedia, L. laeta, L. gaucho, and L. similis spider venoms. In vivo and in vitro neutralization assays showed that anti-rMEPLox antibodies can efficiently neutralize the sphingomyelinase, hyaluronidase, and metalloproteinase activity of L. intermedia venom. This study suggests that this multiepitopic protein can be a suitable candidate for experimental vaccination approaches or for antivenom production against Loxosceles spp. venoms.

  5. Influenza A Virus Protein PA-X Contributes to Viral Growth and Suppression of the Host Antiviral and Immune Responses.

    Science.gov (United States)

    Hayashi, Tsuyoshi; MacDonald, Leslie A; Takimoto, Toru

    2015-06-01

    Influenza virus infection causes global inhibition of host protein synthesis in infected cells. This host shutoff is thought to allow viruses to escape from the host antiviral response, which restricts virus replication and spread. Although the mechanism of host shutoff is unclear, a novel viral protein expressed by ribosomal frameshifting, PA-X, was found to play a major role in influenza virus-induced host shutoff. However, little is known about the impact of PA-X expression on currently circulating influenza A virus pathogenicity and the host antiviral response. In this study, we rescued a recombinant influenza A virus, A/California/04/09 (H1N1, Cal), containing mutations at the frameshift motif in the polymerase PA gene (Cal PA-XFS). Cal PA-XFS expressed significantly less PA-X than Cal wild type (WT). Cal WT, but not Cal PA-XFS, induced degradation of host β-actin mRNA and suppressed host protein synthesis, supporting the idea that PA-X induces host shutoff via mRNA decay. Moreover, Cal WT inhibited beta interferon (IFN-β) expression and replicated more rapidly than Cal PA-XFS in human respiratory cells. Mice infected with Cal PA-XFS had significantly lower levels of viral growth and greater expression of IFN-β mRNA in their lungs than mice infected with Cal WT. Importantly, more antihemagglutinin and neutralizing antibodies were produced in Cal PA-XFS-infected mice than in Cal WT-infected mice, despite the lower level of virus replication in the lungs. Our data indicate that PA-X of the pandemic H1N1 virus has a strong impact on viral growth and the host innate and acquired immune responses to influenza virus. Virus-induced host protein shutoff is considered to be a major factor allowing viruses to evade innate and acquired immune recognition. We provide evidence that the 2009 H1N1 influenza A virus protein PA-X plays a role in virus replication and inhibition of host antiviral response by means of its host protein synthesis shutoff activity both in vitro

  6. Expression of a hantavirus N protein and its efficacy as antigen in immune assays

    Directory of Open Access Journals (Sweden)

    L.T.M. Figueiredo

    2008-07-01

    Full Text Available Hantavirus cardiopulmonary syndrome (HCPS has been recognized as an important public heath problem. Five hantaviruses associated with HCPS are currently known in Brazil: Juquitiba, Araraquara, Laguna Negra-like, Castelo dos Sonhos, and Anajatuba viruses. The laboratory diagnosis of HCPS is routinely carried out by the detection of anti-hantavirus IgM and/or IgG antibodies. The present study describes the expression of the N protein of a hantavirus detected in the blood sample of an HCPS patient. The entire S segment of the virus was amplified and found to be 1858 nucleotides long, with an open reading frame of 1287 nucleotides that encodes a protein of 429 amino acids. The nucleotide sequence described here showed a high identity with the N protein gene of Araraquara virus. The entire N protein was expressed using the vector pET200D and the Escherichia coli BL21 strain. The expression of the recombinant protein was confirmed by the detection of a 52-kDa protein by Western blot using a pool of human sera obtained from HCPS patients, and by specific IgG detection in five serum samples of HCPS patients tested by ELISA. These results suggest that the recombinant N protein could be used as an antigen for the serological screening of hantavirus infection.

  7. Recombinant paracoccin reproduces the biological properties of the native protein and induces protective Th1 immunity against Paracoccidioides brasiliensis infection.

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    Ana Claudia Paiva Alegre

    2014-04-01

    Full Text Available BACKGROUND: Paracoccin is a dual-function protein of the yeast Paracoccidioides brasiliensis that has lectin properties and N-acetylglucosaminidase activities. Proteomic analysis of a paracoccin preparation from P. brasiliensis revealed that the sequence matched that of the hypothetical protein encoded by PADG-3347 of isolate Pb-18, with a polypeptide sequence similar to the family 18 endochitinases. These endochitinases are multi-functional proteins, with distinct lectin and enzymatic domains. METHODOLOGY/PRINCIPAL FINDINGS: The multi-exon assembly and the largest exon of the predicted ORF (PADG-3347, was cloned and expressed in Escherichia coli cells, and the features of the recombinant proteins were compared to those of the native paracoccin. The multi-exon protein was also used for protection assays in a mouse model of paracoccidioidomycosis. CONCLUSIONS/SIGNIFICANCE: Our results showed that the recombinant protein reproduced the biological properties described for the native protein-including binding to laminin in a manner that is dependent on carbohydrate recognition-showed N-acetylglucosaminidase activity, and stimulated murine peritoneal macrophages to produce high levels of TNF-α and nitric oxide. Considering the immunomodulatory potential of glycan-binding proteins, we also investigated whether prophylactic administration of recombinant paracoccin affected the course of experimental paracoccidioidomycosis in mice. In comparison to animals injected with vehicle (controls, mice treated with recombinant paracoccin displayed lower pulmonary fungal burdens and reduced pulmonary granulomas. These protective effects were associated with augmented pulmonary levels of IL-12 and IFN-γ. We also observed that injection of paracoccin three days before challenge was the most efficient administration protocol, as the induced Th1 immunity was balanced by high levels of pulmonary IL-10, which may prevent the tissue damage caused by exacerbated

  8. Acyclovir Therapy Reduces the CD4+ T Cell Response against the Immunodominant pp65 Protein from Cytomegalovirus in Immune Competent Individuals.

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    Annette Pachnio

    Full Text Available Cytomegalovirus (CMV infects the majority of the global population and leads to the development of a strong virus-specific immune response. The CMV-specific CD4+ and CD8+ T cell immune response can comprise between 10 and 50% of the T cell pool within peripheral blood and there is concern that this may impair immunity to other pathogens. Elderly individuals with the highest magnitude of CMV-specific immune response have been demonstrated to be at increased risk of mortality and there is increasing interest in interventions that may serve to moderate this. Acyclovir is an anti-viral drug with activity against a range of herpes viruses and is used as long term treatment to suppress reactivation of herpes simplex virus. We studied the immune response to CMV in patients who were taking acyclovir to assess if therapy could be used to suppress the CMV-specific immune response. The T cell reactivity against the immunodominant late viral protein pp65 was reduced by 53% in people who were taking acyclovir. This effect was seen within one year of therapy and was observed primarily within the CD4+ response. Acyclovir treatment only modestly influenced the immune response to the IE-1 target protein. These data show that low dose acyclovir treatment has the potential to modulate components of the T cell response to CMV antigen proteins and indicate that anti-viral drugs should be further investigated as a means to reduce the magnitude of CMV-specific immune response and potentially improve overall immune function.

  9. Construction of Lactococcus lactis expressing secreted and anchored Eimeria tenella 3-1E protein and comparison of protective immunity against homologous challenge.

    Science.gov (United States)

    Ma, Chunli; Zhang, Lili; Gao, Mingyang; Ma, Dexing

    2017-07-01

    Two novel plasmids pTX8048-SP-Δ3-1E and pTX8048-SP-NAΔ3-1E-CWA were constructed. The plasmids were respectively electrotransformed into L. lactis NZ9000 to generate strain of L. lactis/pTX8048-SP-Δ3-1E in which 3-1E protein was expressed in secretion, and L. lactis/pTX8048-SP-NAΔ3-1E-CWA on which 3-1E protein was covalently anchored to the surface of bacteria cells. The expression of target proteins were examined by Western blot. The live lactococci expressing secreted 3-1E protein, anchored 3-1E protein, and cytoplasmic 3-1E protein was administered orally to chickens respectively, and the protective immunity and efficacy were compared by animal experiment. The results showed oral immunization to chickens with recombinant lactococci expressing anchored 3-1E protein elicited high 3-1E-specific serum IgG, increased high proportion of CD4 + and CD8α + cells in spleen, alleviated average lesion score in cecum, decreased the oocyst output per chicken compared to lactococci expressing cytoplasmic or secreted 3-1E protein. Taken together, these findings indicated the surface anchored Eimeria protein displayed by L. lacits can induce protective immunity and partial protection against homologous infection. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Molecular characterization of the recombinant protein RmLTI-BmCG-LTB: Protective immunity against Rhipicephalus (Boophilus microplus.

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    Bárbara Guimarães Csordas

    Full Text Available The bovine tick Rhipicephalus (Boophilus microplus is found in several tropical and subtropical regions of the world. This parasite transmits pathogens that cause disease, such as babesiosis (Babesia bovis and B. bigemina and anaplasmosis (Anaplasma marginale. Tick infestations cause enormous livestock losses, and controlling tick infestations and the transmission of tick-borne diseases remains a challenge for the livestock industry. Because the currently available commercial vaccines offer only partial protection against R. (B. microplus, there is a need for more efficient vaccines. Several recombinant antigens have been evaluated using different immunization strategies, and they show great promise. This work describes the construction and immunological characterization of a multi-antigen chimera composed of two R. (B. microplus antigens (RmLTI and BmCG and one Escherichia coli antigen (B subunit, LTB. The immunogenic regions of each antigen were selected and combined to encode a single polypeptide. The gene was cloned and expressed in E. coli. For all of the experiments, two groups (treated and control of four Angus heifers (3-6 months old were used. The inoculation was performed via intramuscular injection with 200 μg of purified recombinant chimeric protein and adjuvated. The chimeric protein was recognized by specific antibodies against each subunit and by sera from cattle inoculated with the chimera. Immunization of RmLTI-BmCG-LTB cattle reduced the number of adult female ticks by 6.29% and vaccination of cattle with the chimeric antigen provided 55.6% efficacy against R. (B. microplus infestation. The results of this study indicate that the novel chimeric protein is a potential candidate for the future development of a more effective vaccine against R. (B. microplus.

  11. Molecular characterization of the recombinant protein RmLTI-BmCG-LTB: Protective immunity against Rhipicephalus (Boophilus) microplus.

    Science.gov (United States)

    Csordas, Bárbara Guimarães; Cunha, Rodrigo Casquero; Garcia, Marcos Valério; da Silva, Sérgio Silva; Leite, Fábio Leivas; Andreotti, Renato

    2018-01-01

    The bovine tick Rhipicephalus (Boophilus) microplus is found in several tropical and subtropical regions of the world. This parasite transmits pathogens that cause disease, such as babesiosis (Babesia bovis and B. bigemina) and anaplasmosis (Anaplasma marginale). Tick infestations cause enormous livestock losses, and controlling tick infestations and the transmission of tick-borne diseases remains a challenge for the livestock industry. Because the currently available commercial vaccines offer only partial protection against R. (B.) microplus, there is a need for more efficient vaccines. Several recombinant antigens have been evaluated using different immunization strategies, and they show great promise. This work describes the construction and immunological characterization of a multi-antigen chimera composed of two R. (B.) microplus antigens (RmLTI and BmCG) and one Escherichia coli antigen (B subunit, LTB). The immunogenic regions of each antigen were selected and combined to encode a single polypeptide. The gene was cloned and expressed in E. coli. For all of the experiments, two groups (treated and control) of four Angus heifers (3-6 months old) were used. The inoculation was performed via intramuscular injection with 200 μg of purified recombinant chimeric protein and adjuvated. The chimeric protein was recognized by specific antibodies against each subunit and by sera from cattle inoculated with the chimera. Immunization of RmLTI-BmCG-LTB cattle reduced the number of adult female ticks by 6.29% and vaccination of cattle with the chimeric antigen provided 55.6% efficacy against R. (B.) microplus infestation. The results of this study indicate that the novel chimeric protein is a potential candidate for the future development of a more effective vaccine against R. (B.) microplus.

  12. Effects of montelukast sodium combined with pidotimod on acute phase protein and immune function in children with acute bronchitis

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    Jing Wang

    2017-08-01

    Full Text Available Objective: To observe the effects of montelukast sodium combined with pidotimod on acute phase protein (APP and indexes of immunologic function in pediatric acute bronchitis treatment. Methods: A total of 180 cases children with acute bronchitis acted as research objects were randomly divided into control group (n=65 and observation group (n=63. On the basis of conventional therapy, control group was treated by plus pidotimod. On this base, observation group was treated with montelukast sodium. The changes of acute phase proteins (CRP, HP, a1-AAG and CER and immune function (CD3+ , CD4+ , CD8+ and CD4+ /CD8+ levels before and after treatment were observed after 2 months. Results: Before treatment, CRP, HP, a1-AAG, CER, CD3+ , CD4+ , CD8+ and CD4+ /CD8+ levels of two groups had no statistically significant difference; CRP, HP, a1-AAG, CER, and CD8+ levels of control and observation groups decreased significantly after treatment, the decreases of observation group were more obvious than that of control group, and the levels after treatment were significantly lower than that of control groups. The levels of CD3+ , CD4+ and CD4+ /CD8+ in two groups after treatment were significantly higher than those before treatment. For observation group, the levels of CD3+ , CD4+ and CD4+ /CD8+ increased more significantly after treatment, which were significantly higher than that of the control group. Conclusion: Using Montelukast sodium combined with pidotimod can effectively reduce the children's acute phase protein levels, improve immune function, which has clinical value for the treatment of children with acute bronchitis.

  13. Phosphorylation of mouse immunity-related GTPase (IRG resistance proteins is an evasion strategy for virulent Toxoplasma gondii.

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    Tobias Steinfeldt

    Full Text Available Virulence of complex pathogens in mammals is generally determined by multiple components of the pathogen interacting with the functional complexity and multiple layering of the mammalian immune system. It is most unusual for the resistance of a mammalian host to be overcome by the defeat of a single defence mechanism. In this study we uncover and analyse just such a case at the molecular level, involving the widespread intracellular protozoan pathogen Toxoplasma gondii and one of its most important natural hosts, the house mouse (Mus musculus. Natural polymorphism in virulence of Eurasian T. gondii strains for mice has been correlated in genetic screens with the expression of polymorphic rhoptry kinases (ROP kinases secreted into the host cell during infection. We show that the molecular targets of the virulent allelic form of ROP18 kinase are members of a family of cellular GTPases, the interferon-inducible IRG (immunity-related GTPase proteins, known from earlier work to be essential resistance factors in mice against avirulent strains of T. gondii. Virulent T. gondii strain ROP18 kinase phosphorylates several mouse IRG proteins. We show that the parasite kinase phosphorylates host Irga6 at two threonines in the nucleotide-binding domain, biochemically inactivating the GTPase and inhibiting its accumulation and action at the T. gondii parasitophorous vacuole membrane. Our analysis identifies the conformationally active switch I region of the GTP-binding site as an Achilles' heel of the IRG protein pathogen-resistance mechanism. The polymorphism of ROP18 in natural T. gondii populations indicates the existence of a dynamic, rapidly evolving ecological relationship between parasite virulence factors and host resistance factors. This system should be unusually fruitful for analysis at both ecological and molecular levels since both T. gondii and the mouse are widespread and abundant in the wild and are well-established model species with

  14. Novel Role for Protein Inhibitor of Activated STAT 4 (PIAS4) in the Restriction of Herpes Simplex Virus 1 by the Cellular Intrinsic Antiviral Immune Response.

    Science.gov (United States)

    Conn, Kristen L; Wasson, Peter; McFarlane, Steven; Tong, Lily; Brown, James R; Grant, Kyle G; Domingues, Patricia; Boutell, Chris

    2016-05-01

    Small ubiquitin-like modifier (SUMO) is used by the intrinsic antiviral immune response to restrict viral pathogens, such as herpes simplex virus 1 (HSV-1). Despite characterization of the host factors that rely on SUMOylation to exert their antiviral effects, the enzymes that mediate these SUMOylation events remain to be defined. We show that unconjugated SUMO levels are largely maintained throughout infection regardless of the presence of ICP0, the HSV-1 SUMO-targeted ubiquitin ligase. Moreover, in the absence of ICP0, high-molecular-weight SUMO-conjugated proteins do not accumulate if HSV-1 DNA does not replicate. These data highlight the continued importance for SUMO signaling throughout infection. We show that the SUMO ligase protein inhibitor of activated STAT 4 (PIAS4) is upregulated during HSV-1 infection and localizes to nuclear domains that contain viral DNA. PIAS4 is recruited to sites associated with HSV-1 genome entry through SUMO interaction motif (SIM)-dependent mechanisms that are destabilized by ICP0. In contrast, PIAS4 accumulates in replication compartments through SIM-independent mechanisms irrespective of ICP0 expression. Depletion of PIAS4 enhances the replication of ICP0-null mutant HSV-1, which is susceptible to restriction by the intrinsic antiviral immune response. The mechanisms of PIAS4-mediated restriction are synergistic with the restriction mechanisms of a characterized intrinsic antiviral factor, promyelocytic leukemia protein, and are antagonized by ICP0. We provide the first evidence that PIAS4 is an intrinsic antiviral factor. This novel role for PIAS4 in intrinsic antiviral immunity contrasts with the known roles of PIAS proteins as suppressors of innate immunity. Posttranslational modifications with small ubiquitin-like modifier (SUMO) proteins regulate multiple aspects of host immunity and viral replication. The protein inhibitor of activated STAT (PIAS) family of SUMO ligases is predominantly associated with the suppression of

  15. Isolation and Purification of an Antibacterial Protein from Immune Induced Haemolymph of American Cockroach, Periplaneta americana

    OpenAIRE

    Hamid Reza Basseri; Amir Dadi-Khoeni; Ronak Bakhtiari; Mandan Abolhassani; Reza Hajihosseini-Baghdadabadi

    2016-01-01

    Background: Antimicrobial peptides play a role as effectors substances in the immunity of vertebrate and inverte­brate hosts. In the current study, antimicrobial peptide was isolated from the haemolymph of the American cock­roach, Periplaneta americana.Methods: Micrococcus luteus as Gram-positive bacteria and Escherichia coli as Gram-negative bacteria were candi­date for injection. Induction was done by injecting both bacteria into the abdominal cavity of two groups of cock­roaches separately...

  16. Novel Sperm and Gonadotropin-releasing Hormone-based Recombinant Fusion Protein: Achievement of 100% Contraceptive Efficacy by Co-immunization of Male and Female Mice.

    Science.gov (United States)

    Minhas, Vidisha; Shrestha, Abhinav; Wadhwa, Neerja; Singh, Rita; Gupta, Satish Kumar

    2016-12-01

    Improvements in long-term female contraception can be achieved by vaccinating with sperm-derived proteins. Here, recombinant proteins comprising either (i) N- (amino acid residues 1-80) or C- (amino acid residues 76-126) terminal fragments of mouse sperm protein 17 (Sp17) fused to the promiscuous T non-B cell epitope of tetanus toxoid (TT), amino acid residues 830-844 followed by di-lysine linker (KK) (TT-KK-Sp17 N or TT-KK-Sp17 C , respectively) or (ii) mouse equatorin (amino acid residues 21-185) fused to the T non-B cell epitope of bovine RNase (amino acid residues 94-104) were expressed in Escherichia coli. Immunization of female FVB/J mice, using alum as an adjuvant, led to the generation of high antibody titers against the above proteins. Antibodies against both N- and C-terminal fragments of Sp17 reacted with the entire capacitated mouse spermatozoa, whereas those against equatorin reacted exclusively with the equatorial region. Despite the reactivity of all immune sera, only sera from mice immunized with TT-KK-Sp17 N and TT-KK-Sp17 C significantly reduced mouse in vitro fertilization. Mating studies of the immunized females with un-immunized male mice revealed the highest infertility in the TT-KK-Sp17 C -immunized group. In an attempt to further boost the immune response, the C-terminal fragment of Sp17 was expressed as fusion protein with a tandem repeat of gonadotropin-releasing hormone (GnRH) (Sp17 C -GnRH 2 ). Immunization of both male and female mice with Sp17 C -GnRH 2 led to higher contraceptive efficacy compared to mice immunized with TT-KK-Sp17 C . Interestingly, mating studies wherein partners were both immunized with Sp17 C -GnRH 2 showed a complete failure of female mice to conceive. Thus, immunization of both males and females with Sp17 C -GnRH 2 has the potential to increase contraceptive efficacy. Mol. Reprod. Dev. 83: 1048-1059, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  17. Mass spectrometry-based proteomic exploration of the human immune system: focus on the inflammasome, global protein secretion, and T cells.

    Science.gov (United States)

    Nyman, Tuula A; Lorey, Martina B; Cypryk, Wojciech; Matikainen, Sampsa

    2017-05-01

    The immune system is our defense system against microbial infections and tissue injury, and understanding how it works in detail is essential for developing drugs for different diseases. Mass spectrometry-based proteomics can provide in-depth information on the molecular mechanisms involved in immune responses. Areas covered: Summarized are the key immunology findings obtained with MS-based proteomics in the past five years, with a focus on inflammasome activation, global protein secretion, mucosal immunology, immunopeptidome and T cells. Special focus is on extracellular vesicle-mediated protein secretion and its role in immune responses. Expert commentary: Proteomics is an essential part of modern omics-scale immunology research. To date, MS-based proteomics has been used in immunology to study protein expression levels, their subcellular localization, secretion, post-translational modifications, and interactions in immune cells upon activation by different stimuli. These studies have made major contributions to understanding the molecular mechanisms involved in innate and adaptive immune responses. New developments in proteomics offer constantly novel possibilities for exploring the immune system. Examples of these techniques include mass cytometry and different MS-based imaging approaches which can be widely used in immunology.

  18. Robust immunity and heterologous protection against influenza in mice elicited by a novel recombinant NP-M2e fusion protein expressed in E. coli.

    Science.gov (United States)

    Wang, Wenling; Huang, Baoying; Jiang, Tao; Wang, Xiuping; Qi, Xiangrong; Gao, Yingying; Tan, Wenjie; Ruan, Li

    2012-01-01

    The 23-amino acid extracellular domain of matrix 2 protein (M2e) and the internal nucleoprotein (NP) of influenza are highly conserved among viruses and thus are promising candidate antigens for the development of a universal influenza vaccine. Various M2e- or NP-based DNA or viral vector vaccines have been shown to have high immunogenicity; however, high cost, complicated immunization procedures, and vector-specific antibody responses have restricted their applications. Immunization with an NP-M2e fusion protein expressed in Escherichia coli may represent an alternative strategy for the development of a universal influenza vaccine. cDNA encoding M2e was fused to the 3' end of NP cDNA from influenza virus A/Beijing/30/95 (H3N2). The fusion protein (NM2e) was expressed in E. coli and isolated with 90% purity. Mice were immunized with recombinant NM2e protein along with aluminum hydroxide gel and/or CpG as adjuvant. NM2e plus aluminum hydroxide gel almost completely protected the mice against a lethal (20 LD(50)) challenge of heterologous influenza virus A/PR/8/34. The NM2e fusion protein expressed in E. coli was highly immunogenic in mice. Immunization with NM2e formulated with aluminum hydroxide gel protected mice against a lethal dose of a heterologous influenza virus. Vaccination with recombinant NM2e fusion protein is a promising strategy for the development of a universal influenza vaccine.

  19. Genome-wide comparison of the protein-coding repertoire reveals fast evolution of immune-related genes in cephalochordates and Osteichthyes superclass.

    Science.gov (United States)

    Zhang, Qi-Lin; Xu, Bin; Wang, Xiu-Qiang; Yuan, Ming-Long; Chen, Jun-Yuan

    2018-01-02

    Amphioxus is used to investigate the origin and evolution of vertebrates. To better understand the characteristics of genome evolution from cephalochordates to Osteichthyes, we conducted a genome-wide pairwise comparison of protein-coding genes within amphioxus (a comparable group) and parallel analyses within Osteichthyes (two comparable groups). A batch of fast-evolving genes in each comparable group was identified. Of these genes, the most fast-evolving genes (top 20) were scrutinized, most of which were involved in immune system. An analysis of the fast-evolving genes showed that they were enriched into gene ontology (GO) terms and pathways primarily involved in immune-related functions. Similarly, this phenomenon was detected within Osteichthyes, and more well-known and abundant GO terms and pathways involving innate immunity were found in Osteichthyes than in cephalochordates. Next, we measured the expression responses of four genes belonging to metabolism or energy production-related pathways to lipopolysaccharide challenge in the muscle, intestine or skin of B. belcheri ; three of these genes ( HMGCL , CYBS and MDH2 ) showed innate immune responses. Additionally, some genes involved in adaptive immunity showed fast evolution in Osteichthyes, such as those involving "intestinal immune network for IgA production" or "T-cell receptor signaling pathway". In this study, the fast evolution of immune-related genes in amphioxus and Osteichthyes was determined, providing insights into the evolution of immune-related genes in chordates.

  20. Active immunization with recombinant GnRH fusion protein in boars reduces both testicular development and mRNA expression levels of GnRH receptor in pituitary.

    Science.gov (United States)

    Fang, Fugui; Li, Haidong; Liu, Ya; Zhang, Yunhai; Tao, Yong; Li, Yunsheng; Cao, Hongguo; Wang, Suolu; Wang, Lin; Zhang, Xiaorong

    2010-06-01

    Immunization using recombinant maltose binding protein-gonadotropin releasing hormone (MBP-GnRH6) altered both testicular development and transcription of the pituitary GnRH receptor (GnRHR) gene in boars. Scrotal measurement and blood samples were taken at 4-week interval after immunization at 9 weeks of age. The concentrations of testosterone and anti-GnRH antibodies in serum were determined by radioimmunoassay and enzyme-linked immunosorbent assay, respectively. The results showed that active immunization with MBP-GnRH6 increased the serum concentration of anti-GnRH antibodies (Pimmunized animals as compared with MBP immunized boars. MBP-GnRH6 immunized pigs exhibited mounting behavior 4 weeks later than MBP immunized boars. No mature spermatozoa were observed from MBP-GnRH6 immunized animals. By real-time quantitative PCR analysis, the amount of GnRHR mRNA in the pituitary tissue was found to be significantly lower in MBP-GnRH6 immunized animals than in controls (P<0.05). These data demonstrate that recombinant MBP-GnRH6 was effective in immunological castration in boars. Copyright (c) 2010 Elsevier B.V. All rights reserved.

  1. Innate immune response, intestinal morphology and microbiota changes in Senegalese sole fed plant protein diets with probiotics or autolysed yeast.

    Science.gov (United States)

    Batista, S; Medina, A; Pires, M A; Moriñigo, M A; Sansuwan, K; Fernandes, J M O; Valente, L M P; Ozório, R O A

    2016-08-01

    The effects of using plant ingredients in Senegalese sole (Solea senegalensis) diet on immune competence and intestine morphology and microbial ecology are still controversial. Probiotics or immunostimulants can potentially alter the intestinal microbiota in a way that protects fish against pathogens. The current study aimed to examine the intestine histology and microbiota and humoral innate immune response in juvenile sole fed diets with low (35 %) or high (72 %) content of plant protein (PP) ingredients supplemented with a multispecies probiotic bacteria or autolysed yeast. Fish fed the probiotic diet had lower growth performance. Lysozyme and complement activities were significantly higher in fish fed PP72 diets than in their counterparts fed PP35 diets after 17 and 38 days of feeding. At 2 days of feeding, fish fed unsupplemented PP72 showed larger intestine section area and longer villus than fish fed unsupplemented PP35. At 17 days of feeding, fish fed unsupplemented PP72 showed more goblet cells than the other dietary groups, except the group fed yeast supplemented PP35 diet. High dietary PP level, acutely stimulate fish innate immune defence of the fish after 2 and 17 days of feeding. However, this effect does not occur after 73 days of feeding, suggesting a habituation to dietary treatments and/or immunosuppression, with a reduction in the number of the goblet cells. Fish fed for 38 days with diets supplemented with autolysed yeast showed longer intestinal villus. The predominant bacteria found in sole intestine were Vibrio sp. and dietary probiotic supplementation caused a reduction in Vibrio content, regardless of the PP level.

  2. Immune responses to HBsAg conjugated to protein D of non-typeable Haemophilus influenzae in mice.

    Directory of Open Access Journals (Sweden)

    Qiudong Su

    Full Text Available Hepatitis B vaccine that contains an aluminum hydroxide adjuvant induces apoptotic death of Hepa 1-6 cells. Difficult-to-degrade chemical additives in vaccines effectively enhance vaccine immunogenicity, but also affect the host tissue. Identification of bio-molecules that are readily degraded and compatible in vivo as an adjuvant is important for vaccine research. The hapten-carrier effect suggests that stimulation of helper T (Th cells by carrier adjuvants is feasible. Protein D (PD of non-typeable Haemophilus influenzae covalently conjugated to some polysaccharide vaccines has been confirmed to convert T-cell independent (TI antigens into T-cell dependent (TD antigens, and elicit strong T-cell responses ultimately. Herein, we would substitube PD for aluminum hydroxide adjuvant in Hepatitis B vaccine.Truncated PD (amino acids 20-364 was expressed in Escherichia coli and purified by (NH42SO4 precipitation and DEAE chromatography. After evaluation of antigenicity by western blotting, PD was covalently conjugated to yeast-derived recombinant HBsAg by cross-linking with glutaraldehyde. Intramuscular immunization with the conjugate induced higher level of HBsAg-specific antibody than did HBsAg alone (p < 0.05, and was comparable to commercial Hepatitis B vaccine. During the surveillance period (days 35-105, anti-HBs titers were hold high. Moreover, the conjugated vaccine enhanced Th1 immune responses, while Th2 responses were also activated and induced an antibody response, as determined by IFN-γ ELISPOT and IgG1/IgG2a ratio assays.Recombinant truncated PD covalently conjugated to HBsAg antigen enhanced the immunogenicity of the antigen in mice simultaneously by humoral and cellular immune response, which would facilitate therapeutic hepatitis B vaccines.

  3. Humoral and cellular immunity to Plasmodium falciparum merozoite surface protein 1 and protection from infection with blood-stage parasites.

    Science.gov (United States)

    Moormann, Ann M; Sumba, Peter Odada; Chelimo, Kiprotich; Fang, Hua; Tisch, Daniel J; Dent, Arlene E; John, Chandy C; Long, Carole A; Vulule, John; Kazura, James W

    2013-07-01

     Acquired immunity to malaria develops with increasing age and repeated infections. Understanding immune correlates of protection from malaria would facilitate vaccine development and identification of biomarkers that reflect changes in susceptibility resulting from ongoing malaria control efforts.  The relationship between immunoglobulin G (IgG) antibody and both interferon γ (IFN-γ) and interleukin 10 (IL-10) responses to the 42-kD C-terminal fragment of Plasmodium falciparum merozoite surface protein 1 (MSP142) and the risk of (re)infection were examined following drug-mediated clearance of parasitemia in 94 adults and 95 children in an area of holoendemicity of western Kenya.  Positive IFN-γ enzyme-linked immunosorbent assay (ELISA) and enzyme-linked immunosorbent spot assay (ELISPOT) responses to MSP142 3D7 were associated with delayed time to (re)infection, whereas high-titer IgG antibodies to MSP142 3D7 or FVO alleles were not independently predictive of the risk of (re)infection. When IFN-γ and IL-10 responses were both present, the protective effect of IFN-γ was abrogated. A Cox proportional hazard model including IFN-γ, IL-10, MSP142 3D7 IgG antibody responses, hemoglobin S genotype, age, and infection status at baseline showed that the time to blood-stage infection correlated positively with IFN-γ responses and negatively with IL-10 responses, younger age, and asymptomatic parasitemia.  Evaluating combined allele-specific cellular and humoral immunity elicited by malaria provides a more informative measure of protection relative to evaluation of either measure alone.

  4. Effects of Protein-Iron Complex Concentrate Supplementation on Iron Metabolism, Oxidative and Immune Status in Preweaning Calves

    Directory of Open Access Journals (Sweden)

    Robert Kupczyński

    2017-07-01

    Full Text Available The objective of this study was to determine the effects of feeding protein-iron complex (PIC on productive performance and indicators of iron metabolism, hematology parameters, antioxidant and immune status during first 35 days of a calf’s life. Preparation of the complex involved enzymatic hydrolysis of milk casein (serine protease from Yarrowia lipolytica yeast. Iron chloride was then added to the hydrolyzate and lyophilizate. Calves were divided into treated groups: LFe (low iron dose 10 g/day calf of protein-iron complex, HFe (height iron dose 20 g/day calf, and control group. Dietary supplements containing the lower dose of concentrate had a significant positive effect on iron metabolism, while the higher dose of concentrate resulted in increase of total iron binding capacity (TIBC, saturation of transferrin and decrease of and unsaturated iron binding capacity (UIBC, which suggest iron overload. Additionally, treatment with the lower dose of iron remarkably increased the antioxidant parameters, mainly total antioxidant (TAS and glutathione peroxidase activity (GPx. Higher doses of PIC were related to lower total antioxidant status. IgG, IgM, insulin, glucose, TNFα and IGF-1 concentration did not change significantly in either group after supplementation. In practice, the use of protein-iron complex concentrate requires taking into account the iron content in milk replacers and other feedstuffs.

  5. Search for Partner Proteins of A. thaliana Immunophilins Involved in the Control of Plant Immunity

    Directory of Open Access Journals (Sweden)

    Inna A. Abdeeva

    2018-04-01

    Full Text Available The involvement of plant immunophilins in multiple essential processes such as development, various ways of adapting to biotic and abiotic stresses, and photosynthesis has already been established. Previously, research has demonstrated the involvement of three immunophilin genes (AtCYP19-1/ROC3, AtFKBP65/ROF2, and AtCYP57 in the control of plant response to invasion by various pathogens. Current research attempts to identify host target proteins for each of the selected immunophilins. As a result, candidate interactors have been determined and confirmed using a yeast 2-hybrid (Y2H system for protein–protein interaction assays. The generation of mutant isoforms of ROC3 and AtCYP57 harboring substituted amino acids in the in silico-predicted active sites became essential to achieving significant binding to its target partners. This data shows that ROF2 targets calcium-dependent lipid-binding domain-containing protein (At1g70790; AT1 and putative protein phosphatase (At2g30020; АТ2, whereas ROC3 interacts with GTP-binding protein (At1g30580; ENGD-1 and RmlC-like cupin (At5g39120. The immunophilin AtCYP57 binds to putative pyruvate decarboxylase-1 (Pdc1 and clathrin adaptor complex-related protein (At5g05010. Identified interactors confirm our previous findings that immunophilins ROC3, ROF2, and AtCYP57 are directly involved with stress response control. Further, these findings extend our understanding of the molecular functional pathways of these immunophilins.

  6. Immunization with FSHβ fusion protein antigen prevents bone loss in a rat ovariectomy-induced osteoporosis model.

    Science.gov (United States)

    Geng, Wenxin; Yan, Xingrong; Du, Huicong; Cui, Jihong; Li, Liwen; Chen, Fulin

    2013-05-03

    Osteoporosis, a metabolic bone disease, threatens postmenopausal women globally. Hormone replacement therapy (HTR), especially estrogen replacement therapy (ERT), is used widely in the clinic because it has been generally accepted that postmenopausal osteoporosis is caused by estrogen deficiency. However, hypogonadal α and β estrogen receptor null mice were only mildly osteopenic, and mice with either receptor deleted had normal bone mass, indicating that estrogen may not be the only mediator that induces osteoporosis. Recently, follicle-stimulating hormone (FSH), the serum concentration of which increases from the very beginning of menopause, has been found to play a key role in postmenopausal osteoporosis by promoting osteoclastogenesis. In this article, we confirmed that exogenous FSH can enhance osteoclast differentiation in vitro and that this effect can be neutralized by either an anti-FSH monoclonal antibody or anti-FSH polyclonal sera raised by immunizing animals with a recombinant GST-FSHβ fusion protein antigen. Moreover, immunizing ovariectomized rats with the GST-FSHβ antigen does significantly prevent trabecular bone loss and thereby enhance the bone strength, indicating that a FSH-based vaccine may be a promising therapeutic strategy to slow down bone loss in postmenopausal women. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. Fungal melanin stimulates surfactant protein D-mediated opsonization of and host immune response to Aspergillus fumigatus spores.

    Science.gov (United States)

    Sze Wah Wong, Sarah; Rani, Manjusha; Dodagatta-Marri, Eswari; Ibrahim-Granet, Oumaima; Kishore, Uday; Bayry, Jagadeesh; Latgé, Jean-Paul; Sahu, Arvind; Madan, Taruna; Aimanianda, Vishukumar

    2018-02-05

    Surfactant protein D (SP-D), a C-type lectin and pattern-recognition soluble factor, plays an important role in immune surveillance to detect and eliminate human pulmonary pathogens. SPD has been shown to protect against infections with the most ubiquitous airborne fungal pathogen, Aspergillus fumigatus, but the fungal surface component(s) interacting with SP-D is unknown. Here, we show that SP-D binds to melanin pigment on the surface of A. fumigatus dormant spores (conidia). SP-D also exhibited an affinity to two cell-wall polysaccharides of A. fumigatus, galactomannan (GM) and galactosaminogalactan (GAG). The immunolabeling pattern of SP-D was punctate on the conidial surface and was uniform on germinating conidia, in accordance with the localization of melanin, GM, and GAG. We also found that the collagen-like domain of SP-D is involved in its interaction with melanin, whereas its carbohydrate-recognition domain recognized GM and GAG. Unlike unopsonized conidia, SPD-opsonized conidia were phagocytosed more efficiently and stimulated the secretion of proinflammatory cytokines by human monocytederived macrophages. Further, SP-D -/- mice challenged intranasally with wild-type conidia or melanin ghosts (i.e. hollow melanin spheres) displayed significantly reduced proinflammatory cytokines in the lung compared with wild-type mice. In summary, SP-D binds to melanin present on the dormant A. fumigatus conidial surface, facilitates conidial phagocytosis, and stimulates the host immune response. Copyright © 2018, The American Society for Biochemistry and Molecular Biology.

  8. Sublingual vaccination with sonicated Salmonella proteins and mucosal adjuvant induces mucosal and systemic immunity and protects mice from lethal enteritis.

    Science.gov (United States)

    Huang, Ching-Feng; Wu, Tzee-Chung; Wu, Chia-Chao; Lee, Chin-Cheng; Lo, Wen-Tsung; Hwang, Kwei-Shuai; Hsu, Mu-Ling; Peng, Ho-Jen

    2011-07-01

    Salmonella enteritidis is one of the most common pathogens of enteritis. Most experimental vaccines against Salmonella infection have been applied through injections. This is a new trial to explore the effect of sublingual administration of Salmonella vaccines on systemic and mucosal immunity. Adult BALB/c mice were sublingually vaccinated with sonicated Salmonella proteins (SSP) alone, or plus adjuvant CpG DNA (CpG) or cholera toxin (CT). They were boosted 2 weeks later. Saliva specific secretory IgA (SIgA) antibody responses were significantly stimulated in the mice vaccinated with SSP only or together with CpG or CT. Whereas the mice sublingually vaccinated with SSP and CpG had higher spleen cell IFN-γ production and serum specific IgG2a antibody responses, those receiving SSP and CT showed enhanced spleen cell IL-4, IL-5 and IL-6 production, and serum specific IgG1 antibody responses. After oral challenge with live S. enteritidis, the same strain of the source of SSP, immune protection in those sublingually vaccinated with SSP and CpG or CT was found to prevent intestinal necrosis and to render a higher survival rate. In conclusion, sublingual vaccination together with mucosal adjuvant CpG or CT is a simple but effective way against enteric bacterial pathogens. © 2011 The Authors. APMIS © 2011 APMIS.

  9. Dual RNAseq shows the human mucosal immunity protein, MUC13, is a hallmark of Plasmodium exoerythrocytic infection

    KAUST Repository

    LaMonte, Gregory

    2017-10-03

    The exoerythrocytic stage of Plasmodium malaria infection is a critical window for prophylactic intervention. Using a genome-wide dual RNA sequencing of flow-sorted infected and uninfected hepatoma cells we identify the human mucosal immunity gene, Mucin13 (MUC13), as strongly upregulated during Plasmodium exoerythrocytic hepatic-stage infection. We confirm that MUC13 expression is upregulated in hepatoma cell lines and primary hepatocytes. In immunofluorescence assays, host MUC13 protein expression distinguishes infected cells from adjacent uninfected cells and shows similar colocalization with parasite biomarkers such as UIS4 and HSP70. We further show that localization patterns are species independent, distinguishing both P. berghei and P. vivax infected cells, and that MUC13 can be used to identify compounds that inhibit parasite replication in hepatocytes across all Human-infecting Plasmodium species. This data presents a novel interface of host-parasite interactions in Plasmodium, in that a component of host mucosal immunity is reprogrammed to assist the progression of infection.

  10. Structural Insights into Immune Recognition of the Severe Acute Respiratory Syndrome Coronavirus S Protein Receptor Binding Domain

    Energy Technology Data Exchange (ETDEWEB)

    Pak, J.; Sharon, C; Satkunarajah, M; Thierry, C; Cameron, C; Kelvin, D; Seetharaman, J; Cochrane, A; Plummer, F; et. al.

    2009-01-01

    The spike (S) protein of the severe acute respiratory syndrome coronavirus (SARS-CoV) is responsible for host cell attachment and fusion of the viral and host cell membranes. Within S the receptor binding domain (RBD) mediates the interaction with angiotensin-converting enzyme 2 (ACE2), the SARS-CoV host cell receptor. Both S and the RBD are highly immunogenic and both have been found to elicit neutralizing antibodies. Reported here is the X-ray crystal structure of the RBD in complex with the Fab of a neutralizing mouse monoclonal antibody, F26G19, elicited by immunization with chemically inactivated SARS-CoV. The RBD-F26G19 Fab complex represents the first example of the structural characterization of an antibody elicited by an immune response to SARS-CoV or any fragment of it. The structure reveals that the RBD surface recognized by F26G19 overlaps significantly with the surface recognized by ACE2 and, as such, suggests that F26G19 likely neutralizes SARS-CoV by blocking the virus-host cell interaction.

  11. High-protein enteral nutrition enriched with immune-modulating nutrients vs standard high-protein enteral nutrition and nosocomial infections in the ICU: a randomized clinical trial.

    Science.gov (United States)

    van Zanten, Arthur R H; Sztark, François; Kaisers, Udo X; Zielmann, Siegfried; Felbinger, Thomas W; Sablotzki, Armin R; De Waele, Jan J; Timsit, Jean-François; Honing, Marina L H; Keh, Didier; Vincent, Jean-Louis; Zazzo, Jean-Fabien; Fijn, Harvey B M; Petit, Laurent; Preiser, Jean-Charles; van Horssen, Peter J; Hofman, Zandrie

    2014-08-06

    Enteral administration of immune-modulating nutrients (eg, glutamine, omega-3 fatty acids, selenium, and antioxidants) has been suggested to reduce infections and improve recovery from critical illness. However, controversy exists on the use of immune-modulating enteral nutrition, reflected by lack of consensus in guidelines. To determine whether high-protein enteral nutrition enriched with immune-modulating nutrients (IMHP) reduces the incidence of infections compared with standard high-protein enteral nutrition (HP) in mechanically ventilated critically ill patients. The MetaPlus study, a randomized, double-blind, multicenter trial, was conducted from February 2010 through April 2012 including a 6-month follow-up period in 14 intensive care units (ICUs) in the Netherlands, Germany, France, and Belgium. A total of 301 adult patients who were expected to be ventilated for more than 72 hours and to require enteral nutrition for more than 72 hours were randomized to the IMHP (n = 152) or HP (n = 149) group and included in an intention-to-treat analysis, performed for the total population as well as predefined medical, surgical, and trauma subpopulations. High-protein enteral nutrition enriched with immune-modulating nutrients vs standard high-protein enteral nutrition, initiated within 48 hours of ICU admission and continued during the ICU stay for a maximum of 28 days. The primary outcome measure was incidence of new infections according to the Centers for Disease Control and Prevention (CDC) definitions. Secondary end points included mortality, Sequential Organ Failure Assessment (SOFA) scores, mechanical ventilation duration, ICU and hospital lengths of stay, and subtypes of infections according CDC definitions. There were no statistically significant differences in incidence of new infections between the groups: 53% (95% CI, 44%-61%) in the IMHP group vs 52% (95% CI, 44%-61%) in the HP group (P = .96). No statistically significant differences were

  12. Disruption of Protein Mannosylation Affects Candida guilliermondii Cell Wall, Immune Sensing, and Virulence

    Directory of Open Access Journals (Sweden)

    María J. Navarro-Arias

    2016-12-01

    Full Text Available The fungal cell wall contains glycoproteins that interact with the host immune system. In the prominent pathogenic yeast Candida albicans, Pmr1 acts as a Golgi-resident ion pump that provides cofactors to mannosyltransferases, regulating the synthesis of mannans attached to glycoproteins. To gain insight into a putative conservation of such a crucial process within opportunistic yeasts, we were particularly interested in studying the role of the PMR1 homolog in a low-virulent species that rarely causes candidiasis, Candida guilliermondii. We disrupted C. guilliermondii PMR1 and found that loss of Pmr1 affected cell growth and morphology, biofilm formation, susceptibility to cell wall perturbing agents, mannan levels, and the wall composition and organization. Despite there was a significant increment in the amount of β1,3-glucan exposed at the wall surface, this positively influenced only the ability of the mutant to stimulate IL-10 production by human monocytes, suggesting that recognition of both mannan and β1,3-glucan, is required to stimulate strong levels of pro-inflammatory cytokines. Accordingly, our results indicate C. guilliermondii sensing by monocytes was critically dependent on the recognition of N-linked mannans and β1,3-glucan, as reported in other Candida species. In addition, chemical remotion of cell wall O-linked mannans was found to positively influence the recognition of C. guilliermondii by human monocytes, suggesting that O-linked mannans mask other cell wall components from immune cells. This observation contrasts with that reported in C. albicans. Finally, mice infected with C. guilliermondii pmr1 null mutant cells had significantly lower fungal burdens compared to animals challenged with the parental strain. Accordingly, the null mutant showed inability to kill larvae in the Galleria mellonella infection model. This study thus demonstrates that mannans are relevant for the C. guilliermondii-host interaction, with

  13. Disruption of Protein Mannosylation AffectsCandida guilliermondiiCell Wall, Immune Sensing, and Virulence.

    Science.gov (United States)

    Navarro-Arias, María J; Defosse, Tatiana A; Dementhon, Karine; Csonka, Katalin; Mellado-Mojica, Erika; Dias Valério, Aline; González-Hernández, Roberto J; Courdavault, Vincent; Clastre, Marc; Hernández, Nahúm V; Pérez-García, Luis A; Singh, Dhirendra K; Vizler, Csaba; Gácser, Attila; Almeida, Ricardo S; Noël, Thierry; López, Mercedes G; Papon, Nicolas; Mora-Montes, Héctor M

    2016-01-01

    The fungal cell wall contains glycoproteins that interact with the host immune system. In the prominent pathogenic yeast Candida albicans , Pmr1 acts as a Golgi-resident ion pump that provides cofactors to mannosyltransferases, regulating the synthesis of mannans attached to glycoproteins. To gain insight into a putative conservation of such a crucial process within opportunistic yeasts, we were particularly interested in studying the role of the PMR1 homolog in a low-virulent species that rarely causes candidiasis, Candida guilliermondii . We disrupted C. guilliermondii PMR1 and found that loss of Pmr1 affected cell growth and morphology, biofilm formation, susceptibility to cell wall perturbing agents, mannan levels, and the wall composition and organization. Despite the significant increment in the amount of β1,3-glucan exposed at the wall surface, this positively influenced only the ability of the mutant to stimulate IL-10 production by human monocytes, suggesting that recognition of both mannan and β1,3-glucan, is required to stimulate strong levels of pro-inflammatory cytokines. Accordingly, our results indicate C. guilliermondii sensing by monocytes was critically dependent on the recognition of N -linked mannans and β1,3-glucan, as reported in other Candida species. In addition, chemical remotion of cell wall O -linked mannans was found to positively influence the recognition of C. guilliermondii by human monocytes, suggesting that O -linked mannans mask other cell wall components from immune cells. This observation contrasts with that reported in C. albicans . Finally, mice infected with C. guilliermondii pmr1 Δ null mutant cells had significantly lower fungal burdens compared to animals challenged with the parental strain. Accordingly, the null mutant showed inability to kill larvae in the Galleria mellonella infection model. This study thus demonstrates that mannans are relevant for the C. guilliermondii -host interaction, with an atypical role for O

  14. Modification of the plasma complement protein profile by exogenous estrogens is indicative of a compromised immune competence in marine medaka (Oryzias melastigma).

    Science.gov (United States)

    Dong, Miao; Seemann, Frauke; Humble, Joseph L; Liang, Yimin; Peterson, Drew R; Ye, Rui; Ren, Honglin; Kim, Hui-Su; Lee, Jae-Seong; Au, Doris W T; Lam, Yun Wah

    2017-11-01

    Growing evidence suggests that the immune system of teleost is vulnerable to xenoestrogens, which are ubiquitous in the marine environment. This study detected and identified the major circulatory immune proteins deregulated by 17α-ethinylestradiol (EE2), which may be linked to fish susceptibility to pathogens in the marine medaka, Oryzias melastigma. Fish immune competence was determined using a host resistance assay to pathogenic bacteria Edwardsiella tarda. Females were consistently more susceptible to infection-induced mortality than males. Exposure to EE2 could narrow the sex gap of mortality by increasing infection-induced death in male fish. Proteomic analysis revealed that the major plasma immune proteins of adult fish were highly sexually dimorphic. EE2 induced pronounced sex-specific changes in the plasma proteome, with the male plasma composition clearly becoming "feminised". Male plasma was found to contain a higher level of fibrinogens, WAP63 and ependymin-2-like protein, which are involved in coagulation, inflammation and regeneration. For the first time, we demonstrated that expression of C1q subunit B (C1Q), an initiating factor of the classical complement pathway, was higher in males and was suppressed in both sexes in response to EE2 and bacterial challenge. Moreover, cleavage and post-translational modification of C3, the central component of the complement system, could be altered by EE2 treatment in males (C3dg down; C3g up). Multiple regression analysis indicated that C1Q is possibly an indicator of fish survival, which warrants further confirmation. The findings support the potential application of plasma immune proteins for prognosis/diagnosis of fish immune competence. Moreover, this study provides the first biochemical basis of the sex-differences in fish immunity and how these differences might be modified by xenoestrogens. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. The non-structural protein 5 and matrix protein are antigenic targets of T cell immunity to genotype 1 porcine reproductive and respiratory syndrome viruses

    Directory of Open Access Journals (Sweden)

    Helen eMokhtar

    2016-02-01

    Full Text Available The porcine reproductive and respiratory syndrome virus (PRRSV is the cause of one of the most economically important diseases affecting swine worldwide. Efforts to develop a next-generation vaccine have largely focussed on envelope glycoproteins to target virus-neutralising antibody responses. However, these approaches have failed to demonstrate the necessary efficacy to progress towards market. T cells are crucial to the control of many viruses through cytolysis and cytokine secretion. Since control of PRRSV infection is not dependent on the development of neutralising antibodies, it has been proposed that T cell mediated immunity plays a key role. We therefore hypothesised that conserved T cell antigens represent prime candidates for the development a novel PRRS vaccine. Antigens were identified by screening a proteome-wide synthetic peptide library with T cells from cohorts of pigs rendered immune by experimental infections with a closely-related (subtype 1 or divergent (subtype 3 PRRSV-1 strain. Dominant T cell IFN-γ responses were directed against the non-structural protein 5 (NSP5, and to a lesser extent, the matrix (M protein. The majority of NSP5-specific CD8 T cells and M-specific CD4 T cells expressed a putative effector memory phenotype and were polyfunctional as assessed by co-expression of TNF-α and mobilisation of the cytotoxic degranulation marker CD107a. Both antigens were generally well conserved amongst strains of both PRRSV genotypes. Thus M and NSP5 represent attractive vaccine candidate T cell antigens which should be evaluated further in the context of PRRSV vaccine development.

  16. Calcium/Calmodulin-Dependent Protein Kinase IV Mediates IFN-γ-Induced Immune Behaviors in Skeletal Muscle Cells

    Directory of Open Access Journals (Sweden)

    RuiCai Gu

    2018-03-01

    Full Text Available Background/Aims: Whether calcium/calmodulin-dependent protein kinase IV (CaMKIV plays a role in regulating immunologic features of muscle cells in inflammatory environment, as it does for immune cells, remains mostly unknown. In this study, we investigated the influence of endogenous CaMKIV on the immunological characteristics of myoblasts and myotubes received IFN-γ stimulation. Methods: C2C12 and murine myogenic precursor cells (MPCs were cultured and differentiated in vitro, in the presence of pro-inflammatory IFN-γ. CaMKIV shRNA lentivirus transfection was performed to knockdown CaMKIV gene in C2C12 cells. pEGFP-N1-CaMKIV plasmid was delivered into knockout cells for recovering intracellular CaMKIV gene level. CREB1 antagonist KG-501 was used to block CREB signal. qPCR, immunoblot analysis, or immunofluorescence was used to detect mRNA and protein levels of CaMKIV, immuno-molecules, or pro-inflammatory cytokines and chemokines. Co-stimulatory molecules expression was assessed by FACS analysis. Results: IFN-γ induces the expression or up-regulation of MHC-I/II and TLR3, and the up-regulation of CaMKIV level in muscle cells. In contrast, CaMKIV knockdown in myoblasts and myotubes leads to expression inhibition of the above immuno-molecules. As well, CaMKIV knockdown selectively inhibits pro-inflammatory cytokines/chemokines, and co-stimulatory molecules expression in IFN-γ treated myoblasts and myotubes. Finally, CaMKIV knockdown abolishes IFN-γ induced CREB pathway molecules accumulation in differentiated myotubes. Conclusions: CaMKIV can be induced to up-regulate in muscle cells under inflammatory condition, and positively mediates intrinsic immune behaviors of muscle cells triggered by IFN-γ.

  17. Viral double-strand RNA-binding proteins can enhance innate immune signaling by toll-like Receptor 3.

    Directory of Open Access Journals (Sweden)

    Yvonne Lai

    Full Text Available Toll-like Receptor 3 (TLR3 detects double-stranded (ds RNAs to activate innate immune responses. While poly(I:C is an excellent agonist for TLR3 in several cell lines and in human peripheral blood mononuclear cells, viral dsRNAs tend to be poor agonists, leading to the hypothesis that additional factor(s are likely required to allow TLR3 to respond to viral dsRNAs. TLR3 signaling was examined in a lung epithelial cell line by quantifying cytokine production and in human embryonic kidney cells by quantifying luciferase reporter levels. Recombinant 1b hepatitis C virus polymerase was found to enhance TLR3 signaling in the lung epithelial BEAS-2B cells when added to the media along with either poly(I:C or viral dsRNAs. The polymerase from the genotype 2a JFH-1 HCV was a poor enhancer of TLR3 signaling until it was mutated to favor a conformation that could bind better to a partially duplexed RNA. The 1b polymerase also co-localizes with TLR3 in endosomes. RNA-binding capsid proteins (CPs from two positive-strand RNA viruses and the hepadenavirus hepatitis B virus (HBV were also potent enhancers of TLR3 signaling by poly(I:C or viral dsRNAs. A truncated version of the HBV CP that lacked an arginine-rich RNA-binding domain was unable to enhance TLR3 signaling. These results demonstrate that several viral RNA-binding proteins can enhance the dsRNA-dependent innate immune response initiated by TLR3.

  18. Dysfunction of Arabidopsis MACPF domain protein activates programmed cell death via tryptophan metabolism in MAMP-triggered immunity.

    Science.gov (United States)

    Fukunaga, Satoshi; Sogame, Miho; Hata, Masaki; Singkaravanit-Ogawa, Suthitar; Piślewska-Bednarek, Mariola; Onozawa-Komori, Mariko; Nishiuchi, Takumi; Hiruma, Kei; Saitoh, Hiromasa; Terauchi, Ryohei; Kitakura, Saeko; Inoue, Yoshihiro; Bednarek, Paweł; Schulze-Lefert, Paul; Takano, Yoshitaka

    2017-01-01

    Plant immune responses triggered upon recognition of microbe-associated molecular patterns (MAMPs) typically restrict pathogen growth without a host cell death response. We isolated two Arabidopsis mutants, derived from accession Col-0, that activated cell death upon inoculation with nonadapted fungal pathogens. Notably, the mutants triggered cell death also when treated with bacterial MAMPs such as flg22. Positional cloning identified NSL1 (Necrotic Spotted Lesion 1) as a responsible gene for the phenotype of the two mutants, whereas nsl1 mutations of the accession No-0 resulted in necrotic lesion formation without pathogen inoculation. NSL1 encodes a protein of unknown function containing a putative membrane-attack complex/perforin (MACPF) domain. The application of flg22 increased salicylic acid (SA) accumulation in the nsl1 plants derived from Col-0, while depletion of isochorismate synthase 1 repressed flg22-inducible lesion formation, indicating that elevated SA is needed for the cell death response. nsl1 plants of Col-0 responded to flg22 treatment with an RBOHD-dependent oxidative burst, but this response was dispensable for the nsl1-dependent cell death. Surprisingly, loss-of-function mutations in PEN2, involved in the metabolism of tryptophan (Trp)-derived indole glucosinolates, suppressed the flg22-induced and nsl1-dependent cell death. Moreover, the increased accumulation of SA in the nsl1 plants was abrogated by blocking Trp-derived secondary metabolite biosynthesis, whereas the nsl1-dependent hyperaccumulation of PEN2-dependent compounds was unaffected when the SA biosynthesis pathway was blocked. Collectively, these findings suggest that MAMP-triggered immunity activates a genetically programmed cell death in the absence of the functional MACPF domain protein NSL1 via Trp-derived secondary metabolite-mediated activation of the SA pathway. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

  19. Erwinia amylovora effector protein Eop1 suppresses PAMP-triggered immunity in Malus

    Science.gov (United States)

    Erwinia amylovora (Ea) utilizes a type three secretion system (T3SS) to deliver effector proteins into plant host cells. Several Ea effectors have been identified based on their sequence similarity to plant and animal bacterial pathogen effectors; however, the function of the majority of Ea effecto...

  20. C-reactive protein collaborates with plasma lectins to boost immune response against bacteria

    DEFF Research Database (Denmark)

    Ng, PM; Le Saux, A; Lee, CM

    2007-01-01

    Although human C-reactive protein (CRP) becomes upregulated during septicemia, its role remains unclear, since purified CRP showed no binding to many common pathogens. Contrary to previous findings, we show that purified human CRP (hCRP) binds to Salmonella enterica, and that binding is enhanced ...

  1. An Eimeria vaccine candidate based on Eimeria tenella immune mapped protein 1 and the TLR-5 agonist Salmonella typhimurium FliC flagellin

    Energy Technology Data Exchange (ETDEWEB)

    Yin, Guangwen; Qin, Mei [National Animal Protozoa Laboratory and College of Veterinary Medicine, China Agricultural University, Beijing 100193 (China); Liu, Xianyong [National Animal Protozoa Laboratory and College of Veterinary Medicine, China Agricultural University, Beijing 100193 (China); Key Laboratory of Zoonosis, China Ministry of Agriculture and College of Veterinary Medicine, China Agricultural University, Beijing 100193 (China); Suo, Jingxia; Tang, Xinming; Tao, Geru [National Animal Protozoa Laboratory and College of Veterinary Medicine, China Agricultural University, Beijing 100193 (China); Han, Qian [Department of Biochemistry, Virginia Tech, Blacksburg, VA 24061 (United States); Suo, Xun [National Animal Protozoa Laboratory and College of Veterinary Medicine, China Agricultural University, Beijing 100193 (China); Key Laboratory of Zoonosis, China Ministry of Agriculture and College of Veterinary Medicine, China Agricultural University, Beijing 100193 (China); Wu, Wenxue, E-mail: labboard@126.com [National Animal Protozoa Laboratory and College of Veterinary Medicine, China Agricultural University, Beijing 100193 (China); Key Laboratory of Zoonosis, China Ministry of Agriculture and College of Veterinary Medicine, China Agricultural University, Beijing 100193 (China)

    2013-10-25

    Highlights: •We found a new protective protein – (IMPI) in Eimeria tenella. •EtIMP1-flagellin fusion protein is an effective immunogen against Eimeria infection. •Flagellin can be as an apicomplexan parasite vaccine adjuvant in chickens. -- Abstract: Immune mapped protein-1 (IMP1) is a new protective protein in apicomplexan parasites, and exits in Eimeria tenella. But its structure and immunogenicity in E. tenella are still unknown. In this study, IMPI in E. tenella was predicted to be a membrane protein. To evaluate immunogenicity of IMPI in E. tenella, a chimeric subunit vaccine consisting of E. tenella IMP1 (EtIMP1) and a molecular adjuvant (a truncated flagellin, FliC) was constructed and over-expressed in Escherichia coli and its efficacy against E. tenella infection was evaluated. Three-week-old AA broiler chickens were vaccinated with the recombinant EtIMP1-truncated FliC without adjuvant or EtIMP1 with Freund’s Complete Adjuvant. Immunization of chickens with the recombinant EtIMP1-truncated FliC fusion protein resulted in stronger cellular immune responses than immunization with only recombinant EtIMP1 with adjuvant. The clinical effect of the EtIMP1-truncated FliC without adjuvant was also greater than that of the EtIMP1 with adjuvant, which was evidenced by the differences between the two groups in body weight gain, oocyst output and caecal lesions of E. tenella-challenged chickens. The results suggested that the EtIMP1-flagellin fusion protein can be used as an effective immunogen in the development of subunit vaccines against Eimeria infection. This is the first demonstration of antigen-specific protective immunity against avian coccidiosis using a recombinant flagellin as an apicomplexan parasite vaccine adjuvant in chickens.

  2. An Eimeria vaccine candidate based on Eimeria tenella immune mapped protein 1 and the TLR-5 agonist Salmonella typhimurium FliC flagellin

    International Nuclear Information System (INIS)

    Yin, Guangwen; Qin, Mei; Liu, Xianyong; Suo, Jingxia; Tang, Xinming; Tao, Geru; Han, Qian; Suo, Xun; Wu, Wenxue

    2013-01-01

    Highlights: •We found a new protective protein – (IMPI) in Eimeria tenella. •EtIMP1-flagellin fusion protein is an effective immunogen against Eimeria infection. •Flagellin can be as an apicomplexan parasite vaccine adjuvant in chickens. -- Abstract: Immune mapped protein-1 (IMP1) is a new protective protein in apicomplexan parasites, and exits in Eimeria tenella. But its structure and immunogenicity in E. tenella are still unknown. In this study, IMPI in E. tenella was predicted to be a membrane protein. To evaluate immunogenicity of IMPI in E. tenella, a chimeric subunit vaccine consisting of E. tenella IMP1 (EtIMP1) and a molecular adjuvant (a truncated flagellin, FliC) was constructed and over-expressed in Escherichia coli and its efficacy against E. tenella infection was evaluated. Three-week-old AA broiler chickens were vaccinated with the recombinant EtIMP1-truncated FliC without adjuvant or EtIMP1 with Freund’s Complete Adjuvant. Immunization of chickens with the recombinant EtIMP1-truncated FliC fusion protein resulted in stronger cellular immune responses than immunization with only recombinant EtIMP1 with adjuvant. The clinical effect of the EtIMP1-truncated FliC without adjuvant was also greater than that of the EtIMP1 with adjuvant, which was evidenced by the differences between the two groups in body weight gain, oocyst output and caecal lesions of E. tenella-challenged chickens. The results suggested that the EtIMP1-flagellin fusion protein can be used as an effective immunogen in the development of subunit vaccines against Eimeria infection. This is the first demonstration of antigen-specific protective immunity against avian coccidiosis using a recombinant flagellin as an apicomplexan parasite vaccine adjuvant in chickens

  3. The Potato Nucleotide-binding Leucine-rich Repeat (NLR) Immune Receptor Rx1 Is a Pathogen-dependent DNA-deforming Protein.

    Science.gov (United States)

    Fenyk, Stepan; Townsend, Philip D; Dixon, Christopher H; Spies, Gerhard B; de San Eustaquio Campillo, Alba; Slootweg, Erik J; Westerhof, Lotte B; Gawehns, Fleur K K; Knight, Marc R; Sharples, Gary J; Goverse, Aska; Pålsson, Lars-Olof; Takken, Frank L W; Cann, Martin J

    2015-10-09

    Plant nucleotide-binding leucine-rich repeat (NLR) proteins enable cells to respond to pathogen attack. Several NLRs act in the nucleus; however, conserved nuclear targets that support their role in immunity are unknown. Previously, we noted a structural homology between the nucleotide-binding domain of NLRs and DNA replication origin-binding Cdc6/Orc1 proteins. Here we show that the NB-ARC (nucleotide-binding, Apaf-1, R-proteins, and CED-4) domain of the Rx1 NLR of potato binds nucleic acids. Rx1 induces ATP-dependent bending and melting of DNA in vitro, dependent upon a functional P-loop. In situ full-length Rx1 binds nuclear DNA following activation by its cognate pathogen-derived effector protein, the coat protein of potato virus X. In line with its obligatory nucleocytoplasmic distribution, DNA binding was only observed when Rx1 was allowed to freely translocate between both compartments and was activated in the cytoplasm. Immune activation induced by an unrelated NLR-effector pair did not trigger an Rx1-DNA interaction. DNA binding is therefore not merely a consequence of immune activation. These data establish a role for DNA distortion in Rx1 immune signaling and define DNA as a molecular target of an activated NLR. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. The Potato Nucleotide-binding Leucine-rich Repeat (NLR) Immune Receptor Rx1 Is a Pathogen-dependent DNA-deforming Protein*

    Science.gov (United States)

    Fenyk, Stepan; Townsend, Philip D.; Dixon, Christopher H.; Spies, Gerhard B.; de San Eustaquio Campillo, Alba; Slootweg, Erik J.; Westerhof, Lotte B.; Gawehns, Fleur K. K.; Knight, Marc R.; Sharples, Gary J.; Goverse, Aska; Pålsson, Lars-Olof; Takken, Frank L. W.; Cann, Martin J.

    2015-01-01

    Plant nucleotide-binding leucine-rich repeat (NLR) proteins enable cells to respond to pathogen attack. Several NLRs act in the nucleus; however, conserved nuclear targets that support their role in immunity are unknown. Previously, we noted a structural homology between the nucleotide-binding domain of NLRs and DNA replication origin-binding Cdc6/Orc1 proteins. Here we show that the NB-ARC (nucleotide-binding, Apaf-1, R-proteins, and CED-4) domain of the Rx1 NLR of potato binds nucleic acids. Rx1 induces ATP-dependent bending and melting of DNA in vitro, dependent upon a functional P-loop. In situ full-length Rx1 binds nuclear DNA following activation by its cognate pathogen-derived effector protein, the coat protein of potato virus X. In line with its obligatory nucleocytoplasmic distribution, DNA binding was only observed when Rx1 was allowed to freely translocate between both compartments and was activated in the cytoplasm. Immune activation induced by an unrelated NLR-effector pair did not trigger an Rx1-DNA interaction. DNA binding is therefore not merely a consequence of immune activation. These data establish a role for DNA distortion in Rx1 immune signaling and define DNA as a molecular target of an activated NLR. PMID:26306038

  5. Analysis of skin and secretions of Dybowski's frogs (Rana dybowskii) exposed to Staphylococcus aureus or Escherichia coli identifies immune response proteins.

    Science.gov (United States)

    Xiao, Xiang-Hong; Miao, Hui-Min; Xu, Yi-Gang; Zhang, Jing-Yu; Chai, Long-Hui; Xu, Jia-Jia

    2014-04-01

    The aim of the present study was to investigate responses in Dybowski's frogs (Rana dybowskii) exposed to bacteria, using proteomic and transcriptomic approaches. Staphylococcus aureus and Escherichia coli were used as representative Gram-positive and Gram-negative bacteria, respectively, in an infectious challenge model. Frog skin and skin secretions were collected and protein expression in infected frogs compared to control frogs by two-dimensional gel electrophoresis, silver staining, and image analysis. Proteins that demonstrated differential expression were analysed by mass spectrometry and identified by searching protein databases. More than 180 protein spots demonstrated differential expression in E. coli- or S. aureus-challenged groups and, of these, more than 55 spots were up- or down-regulated at least sixfold, post-infection. Proteins with a potential function in the immune response were identified, such as stathmin 1a, annexin A1, superoxide dismutase A, C-type lectin, lysozyme, antimicrobial peptides, cofilin-1-B, mannose receptor, histone H4, prohormone convertase 1, carbonyl reductase 1 and some components of the Toll-like receptor (TLR) signalling pathway. These molecules are potential candidates for further investigation of immune mechanisms in R. dybowskii; in particular, TLR-mediated responses, which might be activated in frogs exposed to pathogenic bacteria as part of innate immune defence, but which might also impact on adaptive immunity to infection. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Both IIC and IID Components of Mannose Phosphotransferase System Are Involved in the Specific Recognition between Immunity Protein PedB and Bacteriocin-Receptor Complex.

    Directory of Open Access Journals (Sweden)

    Wanli Zhou

    Full Text Available Upon exposure to exogenous pediocin-like bacteriocins, immunity proteins specifically bind to the target receptor of the mannose phosphotransferase system components (man-PTS IIC and IID, therefore preventing bacterial cell death. However, the specific recognition of immunity proteins and its associated target receptors remains poorly understood. In this study, we constructed hybrid receptors to identify the domains of IIC and/or IID recognized by the immunity protein PedB, which confers immunity to pediocin PA-1. Using Lactobacillus plantarum man-PTS EII mutant W903, the IICD components of four pediocin PA-1-sensitive strains (L. plantarum WQ0815, Leuconostoc mesenteroides 05-43, Lactobacillus salivarius REN and Lactobacillus acidophilus 05-172 were respectively co-expressed with the immunity protein PedB. Well-diffusions assays showed that only the complex formed by LpIICD from L. plantarum WQ0815 with pediocin PA-1 could be recognized by PedB. In addition, a two-step PCR approach was used to construct hybrid receptors by combining LpIIC or LpIID recognized by PedB with the other three heterologous IID or IIC compounds unrecognized by PedB, respectively. The results showed that all six hybrid receptors were recognized by pediocin PA-1. However, when IIC or IID of L. plantarum WQ0815 was replaced with any corresponding IIC or IID component from L. mesenteroides 05-43, L. salivarius REN and L. acidophilus 05-172, all the hybrid receptors could not be recognized by PedB. Taken altogether, we concluded that both IIC and IID components of the mannose phosphotransferase system play an important role in the specific recognition between the bacteriocin-receptor complex and the immunity protein PedB.

  7. Effects of protein-carbohydrate supplementation on immunity and resistance training outcomes: a double-blind, randomized, controlled clinical trial.

    Science.gov (United States)

    Naclerio, Fernando; Larumbe-Zabala, Eneko; Ashrafi, Nadia; Seijo, Marco; Nielsen, Birthe; Allgrove, Judith; Earnest, Conrad P

    2017-02-01

    To examine the impact of ingesting hydrolyzed beef protein, whey protein, and carbohydrate on resistance training outcomes, body composition, muscle thickness, blood indices of health and salivary human neutrophil peptides (HNP1-3), as reference of humoral immunity followed an 8-week resistance training program in college athletes. Twenty-seven recreationally physically active males and females (n = 9 per treatment) were randomly assigned to one of the three groups: hydrolyzed beef protein, whey protein, or non-protein isoenergetic carbohydrate. Treatment consisted of ingesting 20 g of supplement, mixed with orange juice, once a day immediately post-workout or before breakfast on non-training days. Measurements were performed pre- and post-intervention on total load (kg) lifted at the first and last workout, body composition (via plethysmography) vastus medialis thickness (mm) (via ultrasonography), and blood indices of health. Salivary HNP1-3 were determined before and after performing the first and last workout. Salivary concentration and secretion rates of the HNP1-3 decreased in the beef condition only from pre-first-workout (1.90 ± 0.83 μg/mL; 2.95 ± 2.83 μg/min, respectively) to pre-last-workout (0.92 ± 0.63 μg/mL, p = 0.025, d = 1.03; 0.76 ± 0.74 μg/min, p = 0.049, d = 0.95), and post-last-workout (0.95 ± 0.60 μg/mL, p = 0.032, d = 1.00; 0.59 ± 0.52 μg/min, p = 0.027, d = 1.02). No other significant differences between groups were observed. Supplementation with a carbohydrate-protein beverage may support resistance training outcomes in a comparable way as the ingestion of only carbohydrate. Furthermore, the ingestion of 20 g of hydrolyzed beef protein resulted in a decreased level and secretion rates of the HNP1-3 from baseline with no negative effect on blood indices of health.

  8. Immunoediting and persistence of antigen-specific immunity in patients who have previously been vaccinated with NY-ESO-1 protein formulated in ISCOMATRIX™.

    Science.gov (United States)

    Nicholaou, Theo; Chen, Weisan; Davis, Ian D; Jackson, Heather M; Dimopoulos, Nektaria; Barrow, Catherine; Browning, Judy; Macgregor, Duncan; Williams, David; Hopkins, Wendie; Maraskovsky, Eugene; Venhaus, Ralph; Pan, Linda; Hoffman, Eric W; Old, Lloyd J; Cebon, Jonathan

    2011-11-01

    NY-ESO-1 protein formulated in ISCOMATRIX™ results in CD4+, CD8+ T cell and antibody-mediated immunity. We evaluated persistence of immunity, relapse-free survival and tumour antigen expression upon relapse in patients vaccinated in an earlier trial. Immunity was measured in 28 patients with resected NY-ESO-1-expressing tumours (melanoma 25, breast 3) 252-1,155 days (median = 681) after vaccination. In the earlier vaccination, trial patients received NY-ESO-1 with ISCOMATRIX™ adjuvant at three protein doses 10 μg, 30 μg or 100 μg (n = 14); 100 μg NY-ESO-1 protein (n = 8) or placebo (n = 6), together with 1 μg of intradermal (ID) NY-ESO-1 protein twice for DTH skin testing. Immune responses assessed in the current study included antibody titres, circulating NY-ESO-1-specific T cells and DTH reactivity 2 days after DTH skin testing with NY-ESO-1 protein (1 μg) or peptides (10 μg). Relapse-free survival was determined for 42 melanoma patients. On relapse NY-ESO-1 and HLA, class I was assessed by immunohistochemistry in 17. Persisting anti-NY-ESO-1 immunity was detected in 10/14 recipients who had previously received vaccine with ISCOMATRIX™ adjuvant. In contrast, immunity only persisted in 3/14 who received 100 μg un-adjuvanted NY-ESO-1 protein (3/8) or 2 μg DTH protein (0/6) P = 0.02. Hence, persisting NY-ESO-1 immunity was associated with prior adjuvant. Tumour NY-ESO-1 or HLA class I was downregulated in participants who relapsed suggesting immunoediting had occurred. Immunoediting suggests that a signal of anti-tumour activity was observed in high-risk resected melanoma patients vaccinated with NY-ESO-1/ISCOMATRIX™. This was associated with measurable persisting immunity in the majority of vaccinated subjects tested. A prospective randomised trial has been undertaken to confirm these results.

  9. Membrane-localized extra-large G proteins and Gbg of the heterotrimeric G proteins form functional complexes engaged in plant immunity in Arabidopsis.

    Science.gov (United States)

    Maruta, Natsumi; Trusov, Yuri; Brenya, Eric; Parekh, Urvi; Botella, José Ramón

    2015-03-01

    In animals, heterotrimeric G proteins, comprising Ga, Gb, and Gg subunits, are molecular switches whose function tightly depends on Ga and Gbg interaction. Intriguingly, in Arabidopsis (Arabidopsis thaliana), multiple defense responses involve Gbg, but not Ga. We report here that the Gbg dimer directly partners with extra-large G proteins (XLGs) to mediate plant immunity. Arabidopsis mutants deficient in XLGs, Gb, and Gg are similarly compromised in several pathogen defense responses, including disease development and production of reactive oxygen species. Genetic analysis of double, triple, and quadruple mutants confirmed that XLGs and Gbg functionally interact in the same defense signaling pathways. In addition, mutations in XLG2 suppressed the seedling lethal and cell death phenotypes of BRASSINOSTEROID INSENSITIVE1-associated receptor kinase1-interacting receptor-like kinase1 mutants in an identical way as reported for Arabidopsis Gb-deficient mutants. Yeast (Saccharomyces cerevisiae) three-hybrid and bimolecular fluorescent complementation assays revealed that XLG2 physically interacts with all three possible Gbg dimers at the plasma membrane. Phylogenetic analysis indicated a close relationship between XLGs and plant Ga subunits, placing the divergence point at the dawn of land plant evolution. Based on these findings, we conclude that XLGs form functional complexes with Gbg dimers, although the mechanism of action of these complexes, including activation/deactivation, must be radically different form the one used by the canonical Ga subunit and are not likely to share the same receptors. Accordingly, XLGs expand the repertoire of heterotrimeric G proteins in plants and reveal a higher level of diversity in heterotrimeric G protein signaling.

  10. Effect of β-1, 3 glucan binding protein based zinc oxide nanoparticles supplemented diet on immune response and disease resistance in Oreochromis mossambicus against Aeromonas hydrophila.

    Science.gov (United States)

    Anjugam, Mahalingam; Vaseeharan, Baskaralingam; Iswarya, Arokiadhas; Gobi, Narayanan; Divya, Mani; Thangaraj, Merlin P; Elumalai, Preetham

    2018-03-06

    Recently, several immunostimulants such as β-glucan, microbial and plant products have been used as dietary supplements to combat disease outbreaks in aquaculture. The present study investigates the potential of Portunus pelagicus β-1, 3 glucan binding protein based zinc oxide nanoparticles (Ppβ-GBP-ZnO NPs) supplemented diet on growth, immune response and disease resistance in Mozambique tilapia, Oreochromis mossambicus. The immune-related protein β-GBP was purified from the haemolymph of P. pelagicus using Sephadex G-100 affinity column chromatography. Ppβ-GBP-ZnO NPs was physico- chemically characterized and experimental feed was formulated. Fish were separately fed with commercial diet (control-group I) and Ppβ-GBP (group II, III, IV), Ppβ-GBP-ZnO NPs (group V, VI, VII), chem-ZnO NPs (VIII, IX, X) mixed diet at the concentration of 0.001%, 0.002% and 0.004% respectively. Triplicate groups of O. mossambicus were fed with experimental diets twice a day for 30 days. Fish receiving Ppβ-GBP-ZnO NPs supplemented diet showed a significant increase (P immune responses (myeloperoxidase activity, lysozyme activity and reactive oxygen species activity) and humoral immune responses (complement activity, antiprotease activity and alkaline phosphatase activity) were evaluated at an interval of 15 days during the feeding trial. Results demonstrate that both cellular and humoral immune responses were substantially increased (P immune system and survival of O. mossambicus. Copyright © 2018 Elsevier Ltd. All rights reserved.

  11. DNA vaccine encoding myristoylated membrane protein (MMP) of rock bream iridovirus (RBIV) induces protective immunity in rock bream (Oplegnathus fasciatus).

    Science.gov (United States)

    Jung, Myung-Hwa; Nikapitiya, Chamilani; Jung, Sung-Ju

    2018-02-01

    Rock bream iridovirus (RBIV) causes severe mass mortalities in rock bream (Oplegnathus fasciatus) in Korea. In this study, we investigated the potential of viral membrane protein to induce antiviral status protecting rock bream against RBIV infection. We found that fish administered with ORF008L (myristoylated membrane protein, MMP) vaccine exhibited significantly higher levels of survival compared to ORF007L (major capsid protein, MCP). Moreover, ORF008L-based DNA vaccinated fish showed significant protection at 4 and 8 weeks post vaccination (wpv) than non-vaccinated fish after infected with RBIV (6.7 × 10 5 ) at 23 °C, with relative percent survival (RPS) of 73.36% and 46.72%, respectively. All of the survivors from the first RBIV infection were strongly protected (100% RPS) from re-infected with RBIV (1.1 × 10 7 ) at 100 dpi. In addition, the MMP (ORF008L)-based DNA vaccine significantly induced the gene expression of TLR3 (14.2-fold), MyD88 (11.6-fold), Mx (84.7-fold), ISG15 (8.7-fold), PKR (25.6-fold), MHC class I (13.3-fold), Fas (6.7-fold), Fas ligand (6.7-fold), caspase9 (17.0-fold) and caspase3 (15.3-fold) at 7 days post vaccination in the muscle (vaccine injection site). Our results showed the induction of immune responses and suggest the possibility of developing preventive measures against RBIV using myristoylated membrane protein-based DNA vaccine. Copyright © 2018 Elsevier Ltd. All rights reserved.

  12. Haemato-biochemical, non-specific immunity, antioxidant capacity and histopathological changes in Labeo rohita fingerlings fed rubber protein isolate.

    Science.gov (United States)

    Fawole, Femi John; Sahu, N P; Jain, K K; Gupta, S; Rajendran, K V; Shamna, N; Poojary, Nalini

    2017-06-01

    A 60-day feeding trial was conducted to evaluate the haemato-biochemical, innate immune response, antioxidant capacity and histopathological changes in Labeo rohita fingerlings fed rubber protein isolates (RPI). One hundred and eighty fingerlings (average weight 4.45 ± 0.01 g) were distributed into five experimental groups in triplicate and fed with isonitrogenous and isocaloric diets. Soybean protein isolate (SPI) served as the reference diet (Control), and the treatment diets were formulated as RPI 25 , RPI 50 , RPI 75 and RPI 100 replacing 25, 50, 75 and 100% of SPI protein, respectively. The growth performance indices like final body weight (9.54-10.27 g), net weight gain (5.09-5.84 g), metabolic growth rate (4.54-5.02) and feed efficiency ratio (0.60-0.65) among the various groups were not significantly different (P > 0.05). All the haematological parameters, except red blood cells, showed no significant differences compared with the control group (P > 0.05). The immuno-biochemical parameters like albumin, globulin, total immunoglobulin, respiratory burst and lysozyme activities among the various groups did not differ significantly (P > 0.05). The stress enzyme such as superoxide dismutase (SOD), catalase (CAT) and oxidative stress marker malondialdehyde (MDA) showed no significant difference (P > 0.05). Histopathological examination of the liver revealed no marked changes. In summary, the results showed that RPI was well utilised by the fish and its inclusion did not generate any oxidative-induced stress, thus, RPI may be suggested as a potential replacement for SPI in fish diets without any detrimental effects. Hence, protein isolation offers a unique opportunity for the utilisation of rubber seed meal.

  13. Conserved charged amino acids within Sendai virus C protein play multiple roles in the evasion of innate immune responses.

    Directory of Open Access Journals (Sweden)

    Takashi Irie

    Full Text Available One of the accessory proteins of Sendai virus (SeV, C, translated from an alternate reading frame of P/V mRNA has been shown to function at multiple stages of infection in cell cultures as well as in mice. C protein has been reported to counteract signal transduction by interferon (IFN, inhibit apoptosis induced by the infection, enhance the efficiency of budding of viral particles, and regulate the polarity of viral genome-length RNA synthesis to maximize production of infectious particles. In this study, we have generated a series of SeV recombinants containing substitutions of highly conserved, charged residues within the C protein, and characterized them together with previously-reported C'/C(-, 4C(-, and F170S recombinant viruses in infected cell cultures in terms of viral replication, cytopathogenicity, and antagonizing effects on host innate immunity. Unexpectedly, the amino acid substitutions had no or minimal effect on viral growth and viral RNA synthesis. However, all the substitutions of charged amino acids resulted in the loss of a counteracting effect against the establishment of an IFN-alpha-mediated anti-viral state. Infection by the virus (Cm2' containing mutations at K77 and D80 induced significant IFN-beta production, severe cytopathic effects, and detectable amounts of viral dsRNA production. In addition to the Cm2' virus, the virus containing mutations at E114 and E115 did not inhibit the poly(I:C-triggered translocation of cellular IRF-3 to the nucleus. These results suggest that the C protein play important roles in viral escape from induction of IFN-beta and cell death triggered by infection by means of counteracting the pathway leading to activation of IRF-3 as well as of minimizing viral dsRNA production.

  14. Conserved charged amino acids within Sendai virus C protein play multiple roles in the evasion of innate immune responses.

    Science.gov (United States)

    Irie, Takashi; Nagata, Natsuko; Igarashi, Tomoki; Okamoto, Isao; Sakaguchi, Takemasa

    2010-05-19

    One of the accessory proteins of Sendai virus (SeV), C, translated from an alternate reading frame of P/V mRNA has been shown to function at multiple stages of infection in cell cultures as well as in mice. C protein has been reported to counteract signal transduction by interferon (IFN), inhibit apoptosis induced by the infection, enhance the efficiency of budding of viral particles, and regulate the polarity of viral genome-length RNA synthesis to maximize production of infectious particles. In this study, we have generated a series of SeV recombinants containing substitutions of highly conserved, charged residues within the C protein, and characterized them together with previously-reported C'/C(-), 4C(-), and F170S recombinant viruses in infected cell cultures in terms of viral replication, cytopathogenicity, and antagonizing effects on host innate immunity. Unexpectedly, the amino acid substitutions had no or minimal effect on viral growth and viral RNA synthesis. However, all the substitutions of charged amino acids resulted in the loss of a counteracting effect against the establishment of an IFN-alpha-mediated anti-viral state. Infection by the virus (Cm2') containing mutations at K77 and D80 induced significant IFN-beta production, severe cytopathic effects, and detectable amounts of viral dsRNA production. In addition to the Cm2' virus, the virus containing mutations at E114 and E115 did not inhibit the poly(I:C)-triggered translocation of cellular IRF-3 to the nucleus. These results suggest that the C protein play important roles in viral escape from induction of IFN-beta and cell death triggered by infection by means of counteracting the pathway leading to activation of IRF-3 as well as of minimizing viral dsRNA production.

  15. Outer Membrane Proteins of Brucella abortus Vaccinal and Field Strains and their Immune Response in Buffaloes

    Directory of Open Access Journals (Sweden)

    Rukhshanda Munir*, M. Afzal1, M. Hussain2, S. M. S. Naqvi3 and A. Khanum3

    2010-04-01

    Full Text Available Outer membrane proteins (OMPs of three strains of B. abortus i.e. S19, RB51 and a local field isolate of biotype 1 were isolated through disrupting cells to generate membranes by centrifugation and sodium lauryl sarcosinate solubilisation of inner membrane proteins. Distinct OMP profiles of each strain were seen on SDS-PAGE. SDS-PAGE analysis of S19 and field isolate revealed eight protein bands in each strain. The OMPs of S19 had molecular masses 89.0, 73.0, 53.7, 49.0, 38.0, 27.0, 22.3, and 17.7 kDa, while field isolate had OMPs of 151.3, 89.0, 75.8, 67.6, 37.0, 27.0, 24.0 and 19.0 kDa. B. abortus RB51 yielded 11 OMP bands ranging from 12.5 to 107.1 kDa, with 34.2, 15.8 and 12.5 kDa as additional OMPs. Western immunoblot analysis using antisera raised against all three strains in buffaloes indicated an almost similar pattern of immuno-reactive OMPs in S19 and field strain. Two OMPs of molecular weight 37-38 and 19 kDa were immuno-reactive in all strains in buffaloes. There is possibility of use of these OMPs in a recombinant vaccine for B. abortus. A distinct protein of molecular weight of 151.3 kDa was identified in field strain but not in both vaccine strains of B. abortus. Use of this OMP in a diagnostic assay may differentiate between vaccinated and infected animals.

  16. Serum Immune-Related Proteins are Differentially Expressed during Hibernation in the American Black Bear

    OpenAIRE

    Chow, Brian A.; Donahue, Seth W.; Vaughan, Michael R.; McConkey, Brendan; Vijayan, Mathilakath M.

    2013-01-01

    Hibernation is an adaptation to conserve energy in the face of extreme environmental conditions and low food availability that has risen in several animal phyla. This phenomenon is characterized by reduced metabolic rate (∼25% of the active basal metabolic rate in hibernating bears) and energy demand, while other physiological adjustments are far from clear. The profiling of the serum proteome of the American black bear (Ursus americanus) may reveal specific proteins that are differentially m...

  17. West Nile virus differentially modulates the unfolded protein response to facilitate replication and immune evasion.

    Science.gov (United States)

    Ambrose, Rebecca L; Mackenzie, Jason M

    2011-03-01

    For intracellular survival it is imperative that viruses have the capacity to manipulate various cellular responses, including metabolic and biosynthetic pathways. The unfolded protein response (UPR) is induced by various external and internal stimuli, including the accumulation of misfolded proteins in the endoplasmic reticulum (ER). Our previous studies have indicated that the replication and assembly of the flavivirus West Nile virus strain Kunjin virus (WNV(KUN)) is intimately associated with the ER. Thus, we sought to determine whether the UPR was induced during WNV(KUN) infection. WNV(KUN) induces UPR signaling during replication, which is coordinated with peak replication. Interestingly, signaling is biased toward the ATF6/IRE-1 arm of the response, with high levels of Xbp-1 activation but negligible eukaryotic translation initiation factor 2α phosphorylation and downstream transcription. We show that the PERK-mediated response may partially regulate replication, since external UPR stimulation had a limiting effect on early replication events and cells deficient for PERK demonstrated increased replication and virus release. Significantly, we show that the WNV(KUN) hydrophobic nonstructural proteins NS4A and NS4B are potent inducers of the UPR, which displayed a high correlation in inhibiting Jak-STAT signaling in response to alpha interferon (IFN-α). Sequential removal of the transmembrane domains of NS4A showed that reducing hydrophobicity decreased UPR signaling and restored IFN-α-mediated activation. Overall, these results suggest that WNV(KUN) can stimulate the UPR to facilitate replication and that the induction of a general ER stress response, regulated by hydrophobic WNV(KUN) proteins, can potentiate the inhibition of the antiviral signaling pathway.

  18. Macrophage Immune Response Suppression by Recombinant Mycobacterium tuberculosis Antigens, the ESAT-6, CFP-10, and ESAT-6/CFP-10 Fusion Proteins

    Science.gov (United States)

    Seghatoleslam, Atefeh; Hemmati, Mina; Ebadat, Saeedeh; Movahedi, Bahram; Mostafavi-Pour, Zohreh

    2016-01-01

    Background: Macrophage immune responses are affected by the secretory proteins of Mycobacterium tuberculosis (Mtb). This study aimed to examine the immune responses of macrophages to Mtb secretory antigens, namely ESAT-6, CFP-10, and ESAT-6/CFP-10. Methods: THP-1 cells (a human monocytic cell line) were cultured and differentiated to macrophages by phorbol 12-myristate 13-acetate. The cytotoxicity of the recombinant Mtb proteins was assessed using the MTT assay. Two important immune responses of macrophages, namely NO and ROS production, were measured in response to the ESAT-6, CFP-10, and ESAT-6/CFP-10 antigens. The data were analyzed using one-way ANOVA with SPSS, version 16, and considered significant at Pproteins markedly reduced macrophage immune response. The treatment of the THP-1-differentiated cells with ESAT-6, CFP-10, and ESAT-6/CFP-10 reduced NO and ROS production. The treated THP-1-differentiated cells exhibited less inducible NO synthase activity than did the untreated cells. No toxic effect on macrophage viability was observed for the applied proteins at the different concentrations. Conclusion: It seems that the decline in macrophage immune response is due to the suppression of NO and ROS production pathways without any effect on cell viability. PMID:27365551

  19. A novel tetravalent formulation combining the four aggregated domain III-capsid proteins from dengue viruses induces a functional immune response in mice and monkeys.

    Science.gov (United States)

    Suzarte, Edith; Gil, Lázaro; Valdés, Iris; Marcos, Ernesto; Lazo, Laura; Izquierdo, Alienys; García, Angélica; López, Lázaro; Álvarez, Maylin; Pérez, Yusleydis; Castro, Jorge; Romero, Yaremis; Guzmán, María G; Guillén, Gerardo; Hermida, Lisset

    2015-08-01

    Our group developed a subunit vaccine candidate against dengue virus based on two different viral regions: the domain III of the envelope protein and the capsid protein. The novel chimeric protein from dengue-2 virus [domain III-capsid (DIIIC-2)], when presented as aggregated incorporating oligodeoxynucleotides, induced anti-viral and neutralizing antibodies, a cellular immune response and conferred significant protection to mice and monkeys. The remaining constructs were already obtained and properly characterized. Based on this evidence, this work was aimed at assessing the immune response in mice of the chimeric proteins DIIIC of each serotype, as monovalent and tetravalent formulations. Here, we demonstrated the immunogenicity of each protein in terms of humoral and cell-mediated immunity, without antigen competition on the mixture forming the formulation tetra DIIIC. Accordingly, significant protection was afforded as measured by the limited viral load in the mouse encephalitis model. The assessment of the tetravalent formulation in non-human primates was also conducted. In this animal model, it was demonstrated that the formulation induced neutralizing antibodies and memory cell-mediated immune response with IFN-γ-secreting and cytotoxic capacity, regardless the route of immunization used. Taken together, we can assert that the tetravalent formulation of DIIIC proteins constitutes a promising vaccine candidate against dengue virus, and propose it for further efficacy experiments in monkeys or in the dengue human infection model, as it has been recently proposed. © The Japanese Society for Immunology. 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. Detection of the Host Immune Response to Burkholderia mallei Heat-Shock Proteins GroEL and DnaK in a Glanders Patient and Infected Mice

    Science.gov (United States)

    2007-01-01

    host cell. A GroEL-like protein in Actinobacillus actinomycetemcomitans, a patho- gen associated with periodontal disease , was found in extracellular...shock proteins in protection from and pathogenesis of infectious diseases . Clin Microbiol Rev 12:19–39. Zugel U, Schoel B, Yamamoto S, Hengel H...us Disease 59 (2007) 137–147 www.elsevier.com/locate/diagmicrobioDiagnostic Microbiology and InfectioDetection of the host immune response to

  1. Epicutaneous immunization with protein antigen TNP-Ig and NOD2 ligand muramyl dipeptide (MDP) reverses skin-induced suppression of contact hypersensitivity.

    Science.gov (United States)

    Majewska-Szczepanik, Monika; Dorożyńska, Iwona; Strzępa, Anna; Szczepanik, Marian

    2014-02-01

    Epicutaneous (EC) immunization offers a new method of a needle-free and self-administrable immunization by using a topically applied patch to deliver vaccine. We have previously shown that EC immunization with hapten-conjugated protein antigen TNP-Ig prior to hapten sensitization inhibits Th1-mediated contact hypersensitivity (CHS) in mice. Our further work showed that EC immunization with TNP-Ig and Toll-like receptor (TLR) ligands prior to hapten sensitization reverses skin-induced suppression. Animal model of contact hypersensitivity was used to study reversal of skin-induced suppression. Current work showed that EC immunization with protein antigen TNP-Ig and MDP NOD2 agonist - muramyldipeptide (L isoform) reverses skin-induced suppression of CHS. On the other hand L18-MDP NOD2 agonist - muramyldipeptide with a C18 fatty acid chain and MDP control - negative control for MDP - muramyldipeptide (D isoform, inactive) did not reverse skin-induced suppression. "Transfer in" experiment showed that reversal of skin-induced suppression can be adoptively transferred with lymphoid cells isolated from donors EC treated with TNP-Ig and MDP NOD2 agonist. Moreover, experiment employing two non-cross-reacting antigens TNP-Ig and OX-Ig proved that reversal of skin-induced suppression is antigen specific. Additionally, lymph node cells isolated from mice EC immunized with TNP-Ig and MDP NOD2 agonist produced increased level of IFN-γ suggesting that this cytokine might be involved in reversal of skin-induced suppression. This work shows that EC immunization with protein antigen plus NOD2 ligand MDP may be a potential tool to increase the immunogenicity of weekly immunogenic antigens. Copyright © 2014 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  2. Induction of Immune Tolerance to Foreign Protein via Adeno-Associated Viral Vector Gene Transfer in Mid-Gestation Fetal Sheep

    Science.gov (United States)

    Davey, Marcus G.; Riley, John S.; Andrews, Abigail; Tyminski, Alec; Limberis, Maria; Pogoriler, Jennifer E.; Partridge, Emily; Olive, Aliza; Hedrick, Holly L.; Flake, Alan W.; Peranteau, William H.

    2017-01-01

    A major limitation to adeno-associated virus (AAV) gene therapy is the generation of host immune responses to viral vector antigens and the transgene product. The ability to induce immune tolerance to foreign protein has the potential to overcome this host immunity. Acquisition and maintenance of tolerance to viral vector antigens and transgene products may also permit repeat administration thereby enhancing therapeutic efficacy. In utero gene transfer (IUGT) takes advantage of the immunologic immaturity of the fetus to induce immune tolerance to foreign antigens. In this large animal study, in utero administration of AAV6.2, AAV8 and AAV9 expressing green fluorescent protein (GFP) to ~60 day fetal sheep (term: ~150 days) was performed. Transgene expression and postnatal immune tolerance to GFP and viral antigens were assessed. We demonstrate 1) hepatic expression of GFP 1 month following in utero administration of AAV6.2.GFP and AAV8.GFP, 2) in utero recipients of either AAV6.2.GFP or AAV8.GFP fail to mount an anti-GFP antibody response following postnatal GFP challenge and lack inflammatory cellular infiltrates at the intramuscular site of immunization, 3) a serotype specific anti-AAV neutralizing antibody response is elicited following postnatal challenge of in utero recipients of AAV6.2 or AAV8 with the corresponding AAV serotype, and 4) durable hepatic GFP expression was observed up to 6 months after birth in recipients of AAV8.GFP but expression was lost between 1 and 6 months of age in recipients of AAV6.2.GFP. The current study demonstrates, in a preclinical large animal model, the potential of IUGT to achieve host immune tolerance to the viral vector transgene product but also suggests that a single exposure to the vector capsid proteins at the time of IUGT is inadequate to induce tolerance to viral vector antigens. PMID:28141818

  3. Reversal of epigenetic silencing of MHC class I chain-related protein A and B improves immune recognition of Merkel cell carcinoma.

    Science.gov (United States)

    Ritter, Cathrin; Fan, Kaiji; Paulson, Kelly G; Nghiem, Paul; Schrama, David; Becker, Jürgen C

    2016-02-23

    Merkel cell carcinoma (MCC) is a virally associated cancer characterized by its aggressive behavior and strong immunogenicity. Both viral infection and malignant transformation induce expression of MHC class I chain-related protein (MIC) A and B, which signal stress to cells of the immune system via Natural Killer group 2D (NKG2D) resulting in elimination of target cells. However, despite transformation and the continued presence of virally-encoded proteins, MICs are only expressed in a minority of MCC tumors in situ and are completely absent on MCC cell lines in vitro. This lack of MIC expression was due to epigenetic silencing via MIC promoter hypo-acetylation; indeed, MIC expression was re-induced by pharmacological inhibition of histone deacetylases (HDACs) both in vitro and in vivo. This re-induction of MICs rendered MCC cells more sensitive to immune-mediated lysis. Thus, epigenetic silencing of MICs is an important immune escape mechanism of MCCs.

  4. Endoplasmic reticulum chaperone glucose regulated protein 170-Pokemon complexes elicit a robust antitumor immune response in vivo.

    Science.gov (United States)

    Yuan, Bangqing; Xian, Ronghua; Wu, Xianqu; Jing, Junjie; Chen, Kangning; Liu, Guojun; Zhou, Zhenhua

    2012-07-01

    Previous evidence suggested that the stress protein grp170 can function as a highly efficient molecular chaperone, binding to large protein substrates and acting as a potent vaccine against specific tumors when purified from the same tumor. In addition, Pokemon can be found in almost all malignant tumor cells and is regarded to be a promising candidate for the treatment of tumors. However, the potential of the grp170-Pokemon chaperone complex has not been well described. In the present study, the natural chaperone complex between grp170 and the Pokemon was formed by heat shock, and its immunogenicity was detected by ELISPOT and (51)Cr-release assays in vitro and by tumor bearing models in vivo. Our results demonstrated that the grp170-Pokemon chaperone complex could elicit T cell responses as determined by ELISPOT and (51)Cr-release assays. In addition, immunized C57BL/6 mice were challenged with subcutaneous (s.c.) injection of Lewis cancer cells to induce primary tumors. Treatment of mice with the grp170-Pokemon chaperone complex also significantly inhibited tumor growth and prolonged the life span of tumor-bearing mice. Our results indicated that the grp170-Pokemon chaperone complex might represent a powerful approach to tumor immunotherapy and have significant potential for clinical application. Copyright © 2012 Elsevier GmbH. All rights reserved.

  5. Sibiriline, a new small chemical inhibitor of receptor-interacting protein kinase 1, prevents immune-dependent hepatitis.

    Science.gov (United States)

    Le Cann, Fabienne; Delehouzé, Claire; Leverrier-Penna, Sabrina; Filliol, Aveline; Comte, Arnaud; Delalande, Olivier; Desban, Nathalie; Baratte, Blandine; Gallais, Isabelle; Piquet-Pellorce, Claire; Faurez, Florence; Bonnet, Marion; Mettey, Yvette; Goekjian, Peter; Samson, Michel; Vandenabeele, Peter; Bach, Stéphane; Dimanche-Boitrel, Marie-Thérèse

    2017-09-01

    Necroptosis is a regulated form of cell death involved in several disease models including in particular liver diseases. Receptor-interacting protein kinases, RIPK1 and RIPK3, are the main serine/threonine kinases driving this cell death pathway. We screened a noncommercial, kinase-focused chemical library which allowed us to identify Sibiriline as a new inhibitor of necroptosis induced by tumor necrosis factor (TNF) in Fas-associated protein with death domain (FADD)-deficient Jurkat cells. Moreover, Sib inhibits necroptotic cell death induced by various death ligands in human or mouse cells while not protecting from caspase-dependent apoptosis. By using competition binding assay and recombinant kinase assays, we demonstrated that Sib is a rather specific competitive RIPK1 inhibitor. Molecular docking analysis shows that Sib is trapped closed to human RIPK1 adenosine triphosphate-binding site in a relatively hydrophobic pocket locking RIPK1 in an inactive conformation. In agreement with its RIPK1 inhibitory property, Sib inhibits both TNF-induced RIPK1-dependent necroptosis and RIPK1-dependent apoptosis. Finally, Sib protects mice from concanavalin A-induced hepatitis. These results reveal the small-molecule Sib as a new RIPK1 inhibitor potentially of interest for the treatment of immune-dependent hepatitis. © 2017 Federation of European Biochemical Societies.

  6. Potent Immune Modulation by MEDI6383, an Engineered Human OX40 Ligand IgG4P Fc Fusion Protein.

    Science.gov (United States)

    Oberst, Michael D; Auge, Catherine; Morris, Chad; Kentner, Stacy; Mulgrew, Kathy; McGlinchey, Kelly; Hair, James; Hanabuchi, Shino; Du, Qun; Damschroder, Melissa; Feng, Hui; Eck, Steven; Buss, Nicholas; de Haan, Lolke; Pierce, Andrew J; Park, Haesun; Sylwester, Andrew; Axthelm, Michael K; Picker, Louis; Morris, Nicholas P; Weinberg, Andrew; Hammond, Scott A

    2018-03-15

    Ligation of OX40 (CD134, TNFRSF4) on activated T cells by its natural ligand (OX40L, CD252, TNFSF4) enhances cellular survival, proliferation, and effector functions such as cytokine release and cellular cytotoxicity. We engineered a recombinant human OX40L IgG4P Fc fusion protein termed MEDI6383 that assembles into a hexameric structure and exerts potent agonist activity following engagement of OX40. MEDI6383 displayed solution phase agonist activity that was enhanced when the fusion protein was clustered by Fc gamma receptors (FcγRs) on the surface of adjacent cells. The resulting co-stimulation of OX40 on T cells induced NF-κB promoter activity in OX40-expressing T cells and induced Th1-type cytokine production, proliferation, and resistance to regulatory T cell (Treg)-mediated suppression. MEDI6383 enhanced the cytolytic activity of tumor-reactive T cells and reduced tumor growth in the context of an alloreactive human T cell:tumor cell admix model in immunocompromised mice. Consistent with the role of OX40 costimulation in the expansion of memory T cells, MEDI6383 administered to healthy non-human primates elicited peripheral blood CD4 and CD8 central and effector memory T cell proliferation as well as B cell proliferation. Together, these results suggest that OX40 agonism has the potential to enhance anti-tumor immunity in human malignancies. Copyright ©2018, American Association for Cancer Research.

  7. Serotype 3 pneumococci sequester platelet-derived human thrombospondin-1 via the adhesin and immune evasion protein Hic.

    Science.gov (United States)

    Binsker, Ulrike; Kohler, Thomas P; Krauel, Krystin; Kohler, Sylvia; Habermeyer, Johanna; Schwertz, Hansjörg; Hammerschmidt, Sven

    2017-04-07

    Streptococcus pneumoniae serotype 3 strains emerge frequently within clinical isolates of invasive diseases. Bacterial invasion into deeper tissues is associated with colonization and immune evasion mechanisms. Thus, pneumococci express a versatile repertoire of surface proteins sequestering and interacting specifically with components of the human extracellular matrix and serum. Hic, a PspC-like pneumococcal surface protein, possesses vitronectin and factor H binding activity. Here, we show that heterologously expressed Hic domains interact, similar to the classical PspC molecule, with human matricellular thrombospondin-1 (hTSP-1). Binding studies with isolated human thrombospondin-1 and various Hic domains suggest that the interaction between hTSP-1 and Hic differs from binding to vitronectin and factor H. Binding of Hic to hTSP-1 is inhibited by heparin and chondroitin sulfate A, indicating binding to the N-terminal globular domain or type I repeats of hTSP-1. Competitive inhibition experiments with other pneumococcal hTSP-1 adhesins demonstrated that PspC and PspC-like Hic recognize similar domains, whereas PavB and Hic can bind simultaneously to hTSP-1. In conclusion, Hic binds specifically hTSP-1; however, truncation in the N-terminal part of Hic decreases the binding activity, suggesting that the full length of the α-helical regions of Hic is required for an optimal interaction. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  8. Membrane protein nanoclustering as a functional unit of immune cells : from nanoscopy to single molecule dynamics

    OpenAIRE

    Torreño Piña, Juan Andrés

    2015-01-01

    Premi Extraordinari de Doctorat, promoció 2014-2015. Àmbit de Ciències State-of-the-art biophysical techniques featuring high temporal and spatial resolution have allowed for the first time the direct visualization of individual transmembrane proteins on the cell membrane. These techniques have revealed that a large amount of molecular components of the cell membrane do not organize in a random manner but they rather grouped together forming so-called clusters at the nanoscale. Moreover, t...

  9. Molecular mechanisms of viral immune evasion proteins to inhibit MHC class I antigen processing and presentation.

    Science.gov (United States)

    Zhou, Fang

    2009-01-01

    Viral products inhibit MHC class I antigen processing and presentation via three major pathways: inhibition of major histocompatibility complex (MHC) class I expression on cells, blockade of peptide trafficking and loading on MHC class I molecules, and inhibition of peptide generation in host cells. Viral products also interfere with IFN-gamma -mediated JAK/STAT signal transduction in cells. These results imply that viral proteins probably inhibit the function of IFN-gamma in MHC class I antigen presentation via inactivation of JAK/STAT signal transduction in host cells. Mechanisms of viral products to inhibit IFN-gamma -mediated MHC class I antigen presentation were summarized in this literature review.

  10. The Potato Nucleotide-Binding Leucine-Rich Repeat (NLR) Immune Receptor Rx1 is a Pathogen Dependent DNA-Deforming Protein

    NARCIS (Netherlands)

    Fenyk, S.; Townsend, P.D.; Dixon, C.H.; Spies, G.B.; Campillo, A.S.E.; Slootweg, E.J.; Westerhof, L.B.; Gawehns, F.K.K.; Knight, M.R.; Sharples, G.J.; Goverse, A.; Palsson, L.O.; Takken, F.L.W.; Cann, M.J.

    2015-01-01

    Plant NLR proteins enable cells to respond to pathogen attack. Several NLRs act in the nucleus, however, conserved nuclear targets that support their role in immunity are unknown. Previously we noted a structural homology between the NB domain of NLRs and DNA replication origin-binding Cdc6/Orc1

  11. CD27 instructs CD4+ T cells to provide help for the memory CD8+ T cell response after protein immunization

    NARCIS (Netherlands)

    Xiao, Yanling; Peperzak, Victor; Keller, Anna M.; Borst, Jannie

    2008-01-01

    For optimal quality, memory CD8(+) T cells require CD4(+) T cell help. We have examined whether CD4(+) T cells require CD27 to deliver this help, in a model of intranasal OVA protein immunization. CD27 deficiency reduced the capacity of CD4(+) T cells to support Ag-specific CD8(+) T cell

  12. Synergistic effect of embryo vaccination with Eimeria profilin and Clostridium perfringens NetB proteins on inducing protective immunity against necrotic enteritis in broiler chickens

    Science.gov (United States)

    The effects of embryo vaccination with Eimeria profilin plus Clostridium perfringens NetB toxin proteins in combination with the Montanide IMS-OVO adjuvant on the chicken immune response to necrotic enteritis were investigated using an E. maxima/C. perfringens co-infection model. Eighteen-day-old br...

  13. LC-MS/MS analysis of visceral and subcutaneous adipose tissue proteomes in young goats with focus on innate immunity and inflammation related proteins

    DEFF Research Database (Denmark)

    Restelli, Laura; Codrea, Marius Cosmin; Savoini, Giovanni

    2014-01-01

    and visceral adipose tissues of goat, focusing on proteins involved in immune and inflammatory response. A 2-D LC-MS/MS approach followed by cluster analysis shows a clear distinction between subcutaneous and visceral fat tissue proteomes, and qualitative RT-PCR based analysis of 30 potential adipokines...

  14. Apoplastic Venom Allergen-like Proteins of Cyst Nematodes Modulate the Activation of Basal Plant Innate Immunity by Cell Surface Receptors

    NARCIS (Netherlands)

    Lozano Torres, J.L.; Wilbers, R.H.P.; Warmerdam, S.; Finkers-Tomczak, A.M.; Diaz Granados Muñoz, A.; Schaik, van C.C.; Helder, J.; Bakker, J.; Goverse, A.; Schots, A.; Smant, G.

    2014-01-01

    Despite causing considerable damage to host tissue during the onset of parasitism, nematodes establish remarkably persistent infections in both animals and plants. It is thought that an elaborate repertoire of effector proteins in nematode secretions suppresses damage-triggered immune responses of

  15. Humoral immune response against proteins E6 and E7 in cervical carcinoma patients positive for human papilloma virus type 16 during treatment and follow-up

    NARCIS (Netherlands)

    Baay, M. F.; Duk, J. M.; Burger, M. P.; de Bruijn, H. W.; Stolz, E.; Herbrink, P.

    1999-01-01

    To investigate the humoral immune response to transforming proteins E6 and E7 of human papillomavirus type 16 before and after treatment and during follow-up, consecutive serum samples from 36 cervical cancer patients whose tumours were found to contain human papillomavirus type 16 DNA by use of the

  16. Humoral immune response against proteins E6 and E7 in cervical carcinoma patients positive for human papilloma virus type 16 during treatment and follow-up

    NARCIS (Netherlands)

    Baay, MFD; Duk, JM; Burger, MPM; de Bruijn, HWA; Stolz, E; Herbrink, P

    To investigate the humoral immune response to transforming proteins E6 and E7 of human papillomavirus type 16 before and after treatment and during follow-up, consecutive serum samples from 36 cervical cancer patients whose tumours were found to contain human papillomavirus type 16 DNA by use of the

  17. Killing machines: three pore-forming proteins of the immune system

    Science.gov (United States)

    McCormack, Ryan; de Armas, Lesley; Shiratsuchi, Motoaki

    2014-01-01

    The evolution of early multicellular eukaryotes 400–500 million years ago required a defensive strategy against microbial invasion. Pore-forming proteins containing the membrane-attack-complex-perforin (MACPF) domain were selected as the most efficient means to destroy bacteria or virally infected cells. The mechanism of pore formation by the MACPF domain is distinctive in that pore formation is purely physical and unspecific. The MACPF domain polymerizes, refolds, and inserts itself into bilayer membranes or bacterial outer cell walls. The displacement of surface lipid/carbohydrate molecules by the polymerizing MACPF domain creates clusters of large, water-filled holes that destabilize the barrier function and provide access for additional anti-bacterial or anti-viral effectors to sensitive sites that complete the destruction of the invader via enzymatic or chemical attack. The highly efficient mechanism of anti-microbial defense by a combined physical and chemical strategy using pore-forming MACPF-proteins has been retargeted during evolution of vertebrates and mammals for three purposes: (1) to kill extracellular bacteria C9/polyC9 evolved in conjunction with complement, (2) to kill virus infected and cancer cells perforin-1/polyperforin-1 CTL evolved targeted by NK and CTL, and (3) to kill intracellular bacteria transmembrane perforin-2/putative polyperforin-2 evolved targeted by phagocytic and nonphagocytic cells. Our laboratory has been involved in the discovery and description of each of the three pore-formers that will be reviewed here. PMID:24293008

  18. Mucosal immunization with PspA (Pneumococcal surface protein A-adsorbed nanoparticles targeting the lungs for protection against pneumococcal infection.

    Directory of Open Access Journals (Sweden)

    Tasson C Rodrigues

    Full Text Available Burden of pneumonia caused by Streptococcus pneumoniae remains high despite the availability of conjugate vaccines. Mucosal immunization targeting the lungs is an attractive alternative for the induction of local immune responses to improve protection against pneumonia. Our group had previously described the development of poly(glycerol adipate-co-ω-pentadecalactone (PGA-co-PDL polymeric nanoparticles (NPs adsorbed with Pneumococcal surface protein A from clade 4 (PspA4Pro within L-leucine microcarriers (nanocomposite microparticles-NCMPs for mucosal delivery targeting the lungs (NP/NCMP PspA4Pro. NP/NCMP PspA4Pro was now used for immunization of mice. Inoculation of this formulation induced anti-PspA4Pro IgG antibodies in serum and lungs. Analysis of binding of serum IgG to intact bacteria showed efficient binding to bacteria expressing PspA from clades 3, 4 and 5 (family 2, but no binding to bacteria expressing PspA from clades 1 and 2 (family 1 was observed. Both mucosal immunization with NP/NCMP PspA4Pro and subcutaneous injection of the protein elicited partial protection against intranasal lethal pneumococcal challenge with a serotype 3 strain expressing PspA from clade 5 (PspA5. Although similar survival levels were observed for mucosal immunization with NP/NCMP PspA4Pro and subcutaneous immunization with purified protein, NP/NCMP PspA4Pro induced earlier control of the infection. Conversely, neither immunization with NP/NCMP PspA4Pro nor subcutaneous immunization with purified protein reduced bacterial burden in the lungs after challenge with a serotype 19F strain expressing PspA from clade 1 (PspA1. Mucosal immunization with NP/NCMP PspA4Pro targeting the lungs is thus able to induce local and systemic antibodies, conferring protection only against a strain expressing PspA from the homologous family 2.

  19. Inhibition of antiviral innate immunity by birnavirus VP3 protein via blockage of viral double-stranded RNA binding to the host cytoplasmic RNA detector MDA5.

    Science.gov (United States)

    Ye, Chengjin; Jia, Lu; Sun, Yanting; Hu, Boli; Wang, Lun; Lu, Xingmeng; Zhou, Jiyong

    2014-10-01

    Chicken MDA5 (chMDA5), the sole known pattern recognition receptor for cytoplasmic viral RNA in chickens, initiates type I interferon (IFN) production. Infectious bursal disease virus (IBDV) evades host innate immunity, but the mechanism is unclear. We report here that IBDV inhibited antiviral innate immunity via the chMDA5-dependent signaling pathway. IBDV infection did not induce efficient type I interferon (IFN) production but antagonized the antiviral activity of beta interferon (IFN-β) in DF-1 cells pretreated with IFN-α/β. Dual-luciferase assays and inducible expression systems demonstrated that IBDV protein VP3 significantly inhibited IFN-β expression stimulated by naked IBDV genomic double-stranded RNA (dsRNA). The VP3 protein competed strongly with chMDA5 to bind IBDV genomic dsRNA in vitro and in vivo, and VP3 from other birnaviruses also bound dsRNA. Site-directed mutagenesis confirmed that deletion of the VP3 dsRNA binding domain restored IFN-β expression. Our data demonstrate that VP3 inhibits antiviral innate immunity by blocking binding of viral genomic dsRNA to MDA5. MDA5, a known pattern recognition receptor and cytoplasmic viral RNA sensor, plays a critical role in host antiviral innate immunity. Many pathogens escape or inhibit the host antiviral immune response, but the mechanisms involved are unclear for most pathogens. We report here that birnaviruses inhibit host antiviral innate immunity via the MDA5-dependent signaling pathway. The antiviral innate immune system involving IFN-β did not function effectively during birnavirus infection, and the viral protein VP3 significantly inhibited IFN-β expression stimulated by naked viral genomic dsRNA. We also show that VP3 blocks MDA5 binding to viral genomic dsRNA in vitro and in vivo. Our data reveal that birnavirus-encoded viral protein VP3 is an inhibitor of the antiviral innate immune response and inhibits the antiviral innate immune response via the MDA5-dependent signaling pathway

  20. Mucosal Immunization with Newcastle Disease Virus Vector Coexpressing HIV-1 Env and Gag Proteins Elicits Potent Serum, Mucosal, and Cellular Immune Responses That Protect against Vaccinia Virus Env and Gag Challenges.

    Science.gov (United States)

    Khattar, Sunil K; Manoharan, Vinoth; Bhattarai, Bikash; LaBranche, Celia C; Montefiori, David C; Samal, Siba K

    2015-07-21

    Newcastle disease virus (NDV) avirulent strain LaSota was used to coexpress gp160 Env and p55 Gag from a single vector to enhance both Env-specific and Gag-specific immune responses. The optimal transcription position for both Env and Gag genes in the NDV genome was determined by generating recombinant NDV (rNDV)-Env-Gag (gp160 located between the P and M genes and Gag between the HN and L genes), rNDV-Gag-Env (Gag located between the P and M genes and gp160 between the HN and L genes), rNDV-Env/Gag (gp160 followed by Gag located between the P and M genes), and rNDV-Gag/Env (Gag followed by gp160 located between the P and M genes). All the recombinant viruses replicated at levels similar to those seen with parental NDV in embryonated chicken eggs and in chicken fibroblast cells. Both gp160 and Gag proteins were expressed at high levels in cell culture, with gp160 found to be incorporated into the envelope of NDV. The Gag and Env proteins expressed by all the recombinants except rNDV-Env-Gag self-assembled into human immunodeficiency virus type 1 (HIV-1) virus-like particles (VLPs). Immunization of guinea pigs by the intranasal route with these rNDVs produced long-lasting Env- and Gag-specific humoral immune responses. The Env-specific humoral and mucosal immune responses and Gag-specific humoral immune responses were higher in rNDV-Gag/Env and rNDV-Env/Gag than in the other recombinants. rNDV-Gag/Env and rNDV-Env/Gag were also more efficient in inducing cellular as well as protective immune responses to challenge with vaccinia viruses expressing HIV-1 Env and Gag in mice. These results suggest that vaccination with a single rNDV coexpressing Env and Gag represents a promising strategy to enhance immunogenicity and protective efficacy against HIV. A safe and effective vaccine that can induce both systemic and mucosal immune responses is needed to control HIV-1. In this study, we showed that coexpression of Env and Gag proteins of HIV-1 performed using a single

  1. Cathepsin L is an immune-related protein in Pacific abalone (Haliotis discus hannai)--Purification and characterization.

    Science.gov (United States)

    Shen, Jian-Dong; Cai, Qiu-Feng; Yan, Long-Jie; Du, Cui-Hong; Liu, Guang-Ming; Su, Wen-Jin; Ke, Caihuan; Cao, Min-Jie

    2015-12-01

    Cathepsin L, an immune-related protein, was purified from the hepatopancreas of Pacific abalone (Haliotis discus hannai) by ammonium sulfate precipitation and column chromatographies of SP-Sepharose and Sephacryl S-200 HR. Purified cathepsin L appeared as two bands with molecular masses of 28.0 and 28.5 kDa (namely cathepsin La and Lb) on SDS-PAGE under reducing conditions, suggesting that it is a glycoprotein. Peptide mass fingerprinting (PMF) analysis revealed that peptide fragments of 95 amino acid residues was high similarity to cathepsin L of pearl oyster (Pinctada fucata). The optimal temperature and pH of cathepsin L were 35 °C and pH 5.5. Cathepsin L was particularly inhibited by cysteine proteinase inhibitors of E-64 and leupeptin, while it was activated by metalloproteinase inhibitors EDTA and EGTA. The full-length cathepsin L cDNA was further cloned from the hepatopancreas by rapid PCR amplification of cDNA ends (RACE). The open reading frame of the enzyme was 981 bp, encoding 327 amino acid residues, with a conserved catalytic triad (Cys134, His273 and Asn293), a potential N-glycosylation site and conserved ERFNIN, GNYD, and GCGG motifs, which are characteristics of cathepsin L. Western blot and proteinase activity analysis revealed that the expression and enzyme activity of cathepsin L were significantly up-regulated in hepatopancreas at 8 h following Vibrio parahaemolyticus infection, demonstrating that cathepsin L is involved in the innate immune system of abalone. Our present study for the first time reported the purification, characterization, molecular cloning, and tissue expression of cathepsin L in abalone. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. Respiratory innate immune proteins differentially modulate the neutrophil respiratory burst response to influenza A virus

    DEFF Research Database (Denmark)

    White, Mitchell R; Crouch, Erika; Vesona, Jenny

    2005-01-01

    Oxidants and neutrophils contribute to lung injury during influenza A virus (IAV) infection. Surfactant protein (SP)-D plays a pivotal role in restricting IAV replication and inflammation in the first several days after infection. Despite its potent anti-inflammatory effects in vivo, preincubation...... of IAV with SP-D in vitro strongly increases neutrophil respiratory burst responses to the virus. Several factors are shown to modify this apparent proinflammatory effect of SP-D. Although multimeric forms of SP-D show dose-dependent augmentation of respiratory burst responses, trimeric, single-arm forms...... either show no effect or inhibit these responses. Furthermore, if neutrophils are preincubated with multimeric SP-D before IAV is added, oxidant responses to the virus are significantly reduced. The ability of SP-D to increase neutrophil uptake of IAV can be dissociated from enhancement of oxidant...

  3. Schistosoma mansoni tegument protein Sm29 is able to induce a Th1-type of immune response and protection against parasite infection.

    Directory of Open Access Journals (Sweden)

    Fernanda C Cardoso

    Full Text Available BACKGROUND: Schistosomiasis continues to be a significant public health problem. This disease affects 200 million people worldwide and almost 800 million people are at risk of acquiring the infection. Although vaccine development against this disease has experienced more failures than successes, encouraging results have recently been obtained using membrane-spanning protein antigens from the tegument of Schistosoma mansoni. Our group recently identified Sm29, another antigen that is present at the adult worm tegument surface. In this study, we investigated murine cellular immune responses to recombinant (r Sm29 and tested this protein as a vaccine candidate. METHODS AND FINDINGS: We first show that Sm29 is located on the surface of adult worms and lung-stage schistosomula through confocal microscopy. Next, immunization of mice with rSm29 engendered 51%, 60% and 50% reduction in adult worm burdens, in intestinal eggs and in liver granuloma counts, respectively (p<0.05. Protective immunity in mice was associated with high titers of specific anti-Sm29 IgG1 and IgG2a and elevated production of IFN-gamma, TNF-alpha and IL-12, a typical Th1 response. Gene expression analysis of worms recovered from rSm29 vaccinated mice relative to worms from control mice revealed a significant (q<0.01 down-regulation of 495 genes and up-regulation of only 22 genes. Among down-regulated genes, many of them encode surface antigens and proteins associated with immune signals, suggesting that under immune attack schistosomes reduce the expression of critical surface proteins. CONCLUSION: This study demonstrates that Sm29 surface protein is a new vaccine candidate against schistosomiasis and suggests that Sm29 vaccination associated with other protective critical surface antigens is the next logical strategy for improving protection.

  4. Schistosoma mansoni tegument protein Sm29 is able to induce a Th1-type of immune response and protection against parasite infection.

    Science.gov (United States)

    Cardoso, Fernanda C; Macedo, Gilson C; Gava, Elisandra; Kitten, Gregory T; Mati, Vitor L; de Melo, Alan L; Caliari, Marcelo V; Almeida, Giulliana T; Venancio, Thiago M; Verjovski-Almeida, Sergio; Oliveira, Sergio C

    2008-10-01

    Schistosomiasis continues to be a significant public health problem. This disease affects 200 million people worldwide and almost 800 million people are at risk of acquiring the infection. Although vaccine development against this disease has experienced more failures than successes, encouraging results have recently been obtained using membrane-spanning protein antigens from the tegument of Schistosoma mansoni. Our group recently identified Sm29, another antigen that is present at the adult worm tegument surface. In this study, we investigated murine cellular immune responses to recombinant (r) Sm29 and tested this protein as a vaccine candidate. We first show that Sm29 is located on the surface of adult worms and lung-stage schistosomula through confocal microscopy. Next, immunization of mice with rSm29 engendered 51%, 60% and 50% reduction in adult worm burdens, in intestinal eggs and in liver granuloma counts, respectively (p<0.05). Protective immunity in mice was associated with high titers of specific anti-Sm29 IgG1 and IgG2a and elevated production of IFN-gamma, TNF-alpha and IL-12, a typical Th1 response. Gene expression analysis of worms recovered from rSm29 vaccinated mice relative to worms from control mice revealed a significant (q<0.01) down-regulation of 495 genes and up-regulation of only 22 genes. Among down-regulated genes, many of them encode surface antigens and proteins associated with immune signals, suggesting that under immune attack schistosomes reduce the expression of critical surface proteins. This study demonstrates that Sm29 surface protein is a new vaccine candidate against schistosomiasis and suggests that Sm29 vaccination associated with other protective critical surface antigens is the next logical strategy for improving protection.

  5. No evidence for impaired humoral immunity to pneumococcal proteins in Australian Aboriginal children with otitis media.

    Science.gov (United States)

    Thornton, Ruth B; Kirkham, Lea-Ann S; Corscadden, Karli J; Coates, Harvey L; Vijayasekaran, Shyan; Hillwood, Jessica; Toster, Sophie; Edminston, Phillipa; Zhang, Guicheng; Keil, Anthony; Richmond, Peter C

    2017-01-01

    The Australian Aboriginal population experiences disproportionately high rates of otitis media (OM). Streptococcus pneumoniae is one of the main pathogens responsible for OM and currently no vaccine offering cross strain protection exists. Vaccines consisting of conserved antigens to S. pneumoniae may reduce the burden of OM in high-risk populations; however no data exists examining naturally acquired antibody in Aboriginal children with OM. Serum and salivary IgA and IgG were measured against the S. pneumoniae antigens PspA1 and 2, CbpA and Ply in a cross sectional study of 183 children, including 36 non-Aboriginal healthy control children and 70 Aboriginal children and 77 non-Aboriginal children undergoing surgery for OM using a multiplex bead assay. Significant differences were observed between the 3 groups for serum anti-PspA1 IgA, anti-CbpA and anti-Ply IgG and for all salivary antibodies assessed. Aboriginal children with a history of OM had significantly higher antibody titres than non-Aboriginal healthy children with no history of OM and non-Aboriginal children with a history of OM for several proteins in serum and saliva. Non-Aboriginal children with a history of OM had significantly higher salivary anti-PspA1 IgG than healthy children, while all other titres were comparable between the groups. Conserved vaccine candidate proteins from S. pneumoniae induce serum and salivary antibody responses in Aboriginal and non-Aboriginal children with a history of OM. Aboriginal children do not have an impaired antibody response to the antigens measured from S. pneumoniae and they may represent vaccine candidates in Indigenous populations. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  6. Heat Shock Protein 70 Enhances Mucosal Immunity against Human Norovirus When Coexpressed from a Vesicular Stomatitis Virus Vector

    Science.gov (United States)

    Ma, Yuanmei; Duan, Yue; Wei, Yongwei; Liang, Xueya; Niewiesk, Stefan; Oglesbee, Michael

    2014-01-01

    ABSTRACT Human norovirus (NoV) accounts for 95% of nonbacterial gastroenteritis worldwide. Currently, there is no vaccine available to combat human NoV as it is not cultivable and lacks a small-animal model. Recently, we demonstrated that recombinant vesicular stomatitis virus (rVSV) expressing human NoV capsid protein (rVSV-VP1) induced strong immunities in mice (Y. Ma and J. Li, J. Virol. 85:2942–2952, 2011). To further improve the safety and efficacy of the vaccine candidate, heat shock protein 70 (HSP70) was inserted into the rVSV-VP1 backbone vector. A second construct was generated in which the firefly luciferase (Luc) gene was inserted in place of HSP70 as a control for the double insertion. The resultant recombinant viruses (rVSV-HSP70-VP1 and rVSV-Luc-VP1) were significantly more attenuated in cell culture and viral spread in mice than rVSV-VP1. At the inoculation dose of 1.0 × 106 PFU, rVSV-HSP70-VP1 triggered significantly higher vaginal IgA than rVSV-VP1 and significantly higher fecal and vaginal IgA responses than rVSV-Luc-VP1, although serum IgG and T cell responses were similar. At the inoculation dose of 5.0 × 106 PFU, rVSV-HSP70-VP1 stimulated significantly higher T cell, fecal, and vaginal IgA responses than rVSV-VP1. Fecal and vaginal IgA responses were also significantly increased when combined vaccination of rVSV-VP1 and rVSV-HSP70 was used. Collectively, these data indicate that (i) insertion of an additional gene (HSP70 or Luc) into the rVSV-VP1 backbone further attenuates the VSV-based vaccine in vitro and in vivo, thus improving the safety of the vaccine candidate, and (ii) HSP70 enhances the human NoV-specific mucosal and T cell immunities triggered by a VSV-based human NoV vaccine. IMPORTANCE Human norovirus (NoV) is responsible for more than 95% of acute nonbacterial gastroenteritis worldwide. Currently, there is no vaccine for this virus. Development of a live attenuated vaccine for human NoV has not been possible because it is

  7. Bovine milk proteome in the first 9 days: protein interactions in maturation of the immune and digestive system of the newborn.

    Science.gov (United States)

    Zhang, Lina; Boeren, Sjef; Hageman, Jos A; van Hooijdonk, Toon; Vervoort, Jacques; Hettinga, Kasper

    2015-01-01

    In order to better understand the milk proteome and its changes from colostrum to mature milk, samples taken at seven time points in the first 9 days from 4 individual cows were analyzed using proteomic techniques. Both the similarity in changes from day 0 to day 9 in the quantitative milk proteome, and the differences in specific protein abundance, were observed among four cows. One third of the quantified proteins showed a significant decrease in concentration over the first 9 days after calving, especially in the immune proteins (as much as 40 fold). Three relative high abundant enzymes (XDH, LPL, and RNASE1) and cell division and proliferation protein (CREG1) may be involved in the maturation of the gastro-intestinal tract. In addition, high correlations between proteins involved in complement and blood coagulation cascades illustrates the complex nature of biological interrelationships between milk proteins. The linear decrease of protease inhibitors and proteins involved in innate and adaptive immune system implies a protective role for protease inhibitor against degradation. In conclusion, the results found in this study not only improve our understanding of the role of colostrum in both host defense and development of the newborn calf but also provides guidance for the improvement of infant formula through better understanding of the complex interactions between milk proteins.

  8. Bovine milk proteome in the first 9 days: protein interactions in maturation of the immune and digestive system of the newborn.

    Directory of Open Access Journals (Sweden)

    Lina Zhang

    Full Text Available In order to better understand the milk proteome and its changes from colostrum to mature milk, samples taken at seven time points in the first 9 days from 4 individual cows were analyzed using proteomic techniques. Both the similarity in changes from day 0 to day 9 in the quantitative milk proteome, and the differences in specific protein abundance, were observed among four cows. One third of the quantified proteins showed a significant decrease in concentration over the first 9 days after calving, especially in the immune proteins (as much as 40 fold. Three relative high abundant enzymes (XDH, LPL, and RNASE1 and cell division and proliferation protein (CREG1 may be involved in the maturation of the gastro-intestinal tract. In addition, high correlations between proteins involved in complement and blood coagulation cascades illustrates the complex nature of biological interrelationships between milk proteins. The linear decrease of protease inhibitors and proteins involved in innate and adaptive immune system implies a protective role for protease inhibitor against degradation. In conclusion, the results found in this study not only improve our understanding of the role of colostrum in both host defense and development of the newborn calf but also provides guidance for the improvement of infant formula through better understanding of the complex interactions between milk proteins.

  9. Mucosal priming of the murine immune system against enterohemorrhagic Escherichia coli O157:H7 using Lactococcus lactis expressing the type III secretion system protein EspB.

    Science.gov (United States)

    Ahmed, B; Loos, M; Vanrompay, D; Cox, E

    2013-03-15

    Enterohemorrhagic Escherichia coli (EHEC), particularly E. coli serotype O157:H7, has been responsible for multiple human outbreaks of hemorrhagic colitis and hemolytic uremic syndrome worldwide. Humans become infected by direct or indirect contact with faeces of asymptomatic EHEC shedding ruminants. Currently there is no human or animal vaccine available against EHEC infection. EHEC use a type III secretion system (T3SS) to colonize the intestine and therefore eliciting mucosal immunity against T3SS proteins could be a potential vaccination strategy. To develop such a mucosal vaccine, EspB - a significant member of the T3SS - was intracellularly expressed in Lactococcus lactis (LL-EspB) and this strain was used to immunize BALB/c mice orally. Ten days post-immunization, no specific antibody response was detected in serum or faeces of immunized mice. However, statistically significant (PT3SS protein, EspB. Nevertheless, an optimized EspB expression in L. lactis may be required to improve the mucosal immune response. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. Inactivation of the alpha C protein antigen gene, bca, by a novel shuttle/suicide vector results in attenuation of virulence and immunity in group B Streptococcus.

    Science.gov (United States)

    Li, J; Kasper, D L; Ausubel, F M; Rosner, B; Michel, J L

    1997-11-25

    The alpha C protein of group B Streptococcus (GBS) is a major surface-associated antigen. Although its role in the biology and virulence of GBS has not been defined, it is opsonic and capable of eliciting protective immunity. The alpha C protein is widely distributed among clinical isolates and is a potential protein carrier and antigen in conjugate vaccines to prevent GBS infections. The structural gene for the alpha C protein, bca, has been cloned and sequenced. The protein encoded by bca is related to a class of surface-associated proteins of gram-positive cocci involved in virulence and immunity. To investigate the potential roles of the alpha C protein, bca null mutants were generated in which the bca gene was replaced with a kanamycin resistance cassette via homologous recombination using a novel shuttle/suicide vector. Studies of lethality in neonatal mice showed that the virulence of the bca null mutants was attenuated 5- to 7-fold when compared with the isogenic wild-type strain A909. Significant differences in mortality occurred in the first 24 h, suggesting that the role of the alpha antigen is important in the initial stages of the infection. In contrast to A909, bca mutants were no longer killed by polymorphonuclear leukocytes in the presence of alpha-specific antibodies in an in vitro opsonophagocytic assay. In contrast to previous studies, alpha antigen expression does not appear to play a role in resistance to opsonophagocytosis in the absence of alpha-specific antibodies. In addition, antibodies to the alpha C protein did not passively protect neonatal mice from lethal challenge with bca mutants, suggesting that these epitopes are uniquely present within the alpha antigen as expressed from the bca gene. Therefore, the alpha C protein is important in the pathogenesis of GBS infection and is a target for protective immunity in the development of GBS vaccines.

  11. Btp Proteins from Brucella abortus Modulate the Lung Innate Immune Response to Infection by the Respiratory Route

    Science.gov (United States)

    Hielpos, Maria Soledad; Ferrero, Mariana C.; Fernández, Andrea G.; Falivene, Juliana; Vanzulli, Silvia; Comerci, Diego J.; Baldi, Pablo C.

    2017-01-01

    Although inhalation of infected aerosols is a frequent route for Brucella infection in humans, it rarely causes pulmonary clinical manifestations, suggesting a mild or nearly absent local inflammatory response. The goal of this study was to characterize the early innate immune response to intratracheal infection with Brucella abortus in mice and to evaluate whether it is modulated by this pathogen. After infection with 106 CFU of B. abortus, the pulmonary bacterial burden at 7 days post-infection (p.i.) was comparable to the initial inoculum, despite an initial transient decline. Brucella was detected in spleen and liver as early as 1 day p.i. IL-1β and MCP-1 increased at 3 days p.i., whereas IL-12, KC, TNF-α, and IFN-γ only increased at 7 days p.i. Histological examination did not reveal peribronchial or perivascular infiltrates in infected mice. Experiments were conducted to evaluate if the limited inflammatory lung response to B. abortusis caused by a bacterial mechanism of TLR signaling inhibition. Whereas inoculation of E. coli LPS to control mice [phosphate-buffered saline (PBS)/LPS] caused lung inflammation, almost no histological changes were observed in mice preinfected intratracheally with B. abortus (WT/LPS). We speculated that the Brucella TIR-containing proteins (Btps) A and B, which impair TLR signaling in vitro, may be involved in this modulation. After LPS challenge, mice preinfected with the B. abortus btpAbtpB double mutant exhibited a stronger pulmonary polymorphonuclear infiltrate than WT/LPS mice, although milder than that of the PBS/LPS group. In addition, lungs from B. abortus btpAbtpB-infected mice presented a stronger inflammatory infiltrate than those infected with the WT strain, and at day 7 p.i., the pulmonary levels of KC, MCP-1, and IL-12 were higher in mice infected with the mutant. This study shows that B. abortus infection produces a mild proinflammatory response in murine lungs, partially due to immune modulation by

  12. Btp Proteins from Brucella abortus Modulate the Lung Innate Immune Response to Infection by the Respiratory Route.

    Science.gov (United States)

    Hielpos, Maria Soledad; Ferrero, Mariana C; Fernández, Andrea G; Falivene, Juliana; Vanzulli, Silvia; Comerci, Diego J; Baldi, Pablo C

    2017-01-01

    Although inhalation of infected aerosols is a frequent route for Brucella infection in humans, it rarely causes pulmonary clinical manifestations, suggesting a mild or nearly absent local inflammatory response. The goal of this study was to characterize the early innate immune response to intratracheal infection with Brucella abortus in mice and to evaluate whether it is modulated by this pathogen. After infection with 10 6  CFU of B. abortus , the pulmonary bacterial burden at 7 days post-infection (p.i.) was comparable to the initial inoculum, despite an initial transient decline. Brucella was detected in spleen and liver as early as 1 day p.i. IL-1β and MCP-1 increased at 3 days p.i., whereas IL-12, KC, TNF-α, and IFN-γ only increased at 7 days p.i. Histological examination did not reveal peribronchial or perivascular infiltrates in infected mice. Experiments were conducted to evaluate if the limited inflammatory lung response to B. abortus is caused by a bacterial mechanism of TLR signaling inhibition. Whereas inoculation of E. coli LPS to control mice [phosphate-buffered saline (PBS)/LPS] caused lung inflammation, almost no histological changes were observed in mice preinfected intratracheally with B. abortus (WT/LPS). We speculated that the Brucella TIR-containing proteins (Btps) A and B, which impair TLR signaling in vitro , may be involved in this modulation. After LPS challenge, mice preinfected with the B. abortus btpAbtpB double mutant exhibited a stronger pulmonary polymorphonuclear infiltrate than WT/LPS mice, although milder than that of the PBS/LPS group. In addition, lungs from B. abortus btpAbtpB -infected mice presented a stronger inflammatory infiltrate than those infected with the WT strain, and at day 7 p.i., the pulmonary levels of KC, MCP-1, and IL-12 were higher in mice infected with the mutant. This study shows that B. abortus infection produces a mild proinflammatory response in murine lungs, partially due to immune modulation

  13. Btp Proteins from Brucella abortus Modulate the Lung Innate Immune Response to Infection by the Respiratory Route

    Directory of Open Access Journals (Sweden)

    Maria Soledad Hielpos

    2017-08-01

    Full Text Available Although inhalation of infected aerosols is a frequent route for Brucella infection in humans, it rarely causes pulmonary clinical manifestations, suggesting a mild or nearly absent local inflammatory response. The goal of this study was to characterize the early innate immune response to intratracheal infection with Brucella abortus in mice and to evaluate whether it is modulated by this pathogen. After infection with 106 CFU of B. abortus, the pulmonary bacterial burden at 7 days post-infection (p.i. was comparable to the initial inoculum, despite an initial transient decline. Brucella was detected in spleen and liver as early as 1 day p.i. IL-1β and MCP-1 increased at 3 days p.i., whereas IL-12, KC, TNF-α, and IFN-γ only increased at 7 days p.i. Histological examination did not reveal peribronchial or perivascular infiltrates in infected mice. Experiments were conducted to evaluate if the limited inflammatory lung response to B. abortusis caused by a bacterial mechanism of TLR signaling inhibition. Whereas inoculation of E. coli LPS to control mice [phosphate-buffered saline (PBS/LPS] caused lung inflammation, almost no histological changes were observed in mice preinfected intratracheally with B. abortus (WT/LPS. We speculated that the Brucella TIR-containing proteins (Btps A and B, which impair TLR signaling in vitro, may be involved in this modulation. After LPS challenge, mice preinfected with the B. abortus btpAbtpB double mutant exhibited a stronger pulmonary polymorphonuclear infiltrate than WT/LPS mice, although milder than that of the PBS/LPS group. In addition, lungs from B. abortus btpAbtpB-infected mice presented a stronger inflammatory infiltrate than those infected with the WT strain, and at day 7 p.i., the pulmonary levels of KC, MCP-1, and IL-12 were higher in mice infected with the mutant. This study shows that B. abortus infection produces a mild proinflammatory response in murine lungs, partially due to immune

  14. Cauliflower mosaic virus protein P6 inhibits signaling responses to salicylic acid and regulates innate immunity.

    Directory of Open Access Journals (Sweden)

    Andrew J Love

    Full Text Available Cauliflower mosaic virus (CaMV encodes a multifunctional protein P6 that is required for translation of the 35S RNA and also acts as a suppressor of RNA silencing. Here we demonstrate that P6 additionally acts as a pathogenicity effector of an unique and novel type, modifying NPR1 (a key regulator of salicylic acid (SA- and jasmonic acid (JA-dependent signaling and inhibiting SA-dependent defence responses We find that that transgene-mediated expression of P6 in Arabidopsis and transient expression in Nicotiana benthamiana has profound effects on defence signaling, suppressing expression of representative SA-responsive genes and increasing expression of representative JA-responsive genes. Relative to wild-type Arabidopsis P6-expressing transgenics had greatly reduced expression of PR-1 following SA-treatment, infection by CaMV or inoculation with an avirulent bacterial pathogen Pseudomonas syringae pv tomato (Pst. Similarly transient expression in Nicotiana benthamiana of P6 (including a mutant form defective in translational transactivation activity suppressed PR-1a transcript accumulation in response to Agrobacterium infiltration and following SA-treatment. As well as suppressing the expression of representative SA-regulated genes, P6-transgenic Arabidopsis showed greatly enhanced susceptibility to both virulent and avirulent Pst (titres elevated 10 to 30-fold compared to non-transgenic controls but reduced susceptibility to the necrotrophic fungus Botrytis cinerea. Necrosis following SA-treatment or inoculation with avirulent Pst was reduced and delayed in P6-transgenics. NPR1 an important regulator of SA/JA crosstalk, was more highly expressed in the presence of P6 and introduction of the P6 transgene into a transgenic line expressing an NPR1:GFP fusion resulted in greatly increased fluorescence in nuclei even in the absence of SA. Thus in the presence of P6 an inactive form of NPR1 is mislocalized in the nucleus even in uninduced plants

  15. A novel fusion protein domain III-capsid from dengue-2, in a highly aggregated form, induces a functional immune response and protection in mice

    International Nuclear Information System (INIS)

    Valdes, Iris; Bernardo, Lidice; Gil, Lazaro; Pavon, Alekis; Lazo, Laura; Lopez, Carlos; Romero, Yaremis; Menendez, Ivon; Falcon, Viviana; Betancourt, Lazaro; Martin, Jorge; Chinea, Glay; Silva, Ricardo; Guzman, Maria G.; Guillen, Gerardo; Hermida, Lisset

    2009-01-01

    Based on the immunogenicity of domain III from the Envelope protein of dengue virus as well as the proven protective capacity of the capsid antigen, we have designed a novel domain III-capsid chimeric protein with the goal of obtaining a molecule potentially able to induce both humoral and cell-mediated immunity (CMI). After expression of the recombinant gene in Escherichia coli, the domain III moiety retained its antigenicity as evaluated with anti-dengue sera. In order to explore alternatives for modulating the immunogenicity of the protein, it was mixed with oligodeoxynucleotides in order to obtain particulated aggregates and then immunologically evaluated in mice in comparison with non-aggregated controls. Although the humoral immune response induced by both forms of the protein was equivalent, the aggregated variant resulted in a much stronger CMI as measured by in vitro IFN-γ secretion and protection experiments, mediated by CD4 + and CD8 + cells. The present work provides additional evidence in support for a crucial role of CMI in protection against dengue virus and describes a novel vaccine candidate against the disease based on a recombinant protein that can stimulate both arms of the acquired immune system.

  16. A Functional Toll-Interacting Protein Variant Is Associated with Bacillus Calmette-Guérin-Specific Immune Responses and Tuberculosis.

    Science.gov (United States)

    Shah, Javeed A; Musvosvi, Munyaradzi; Shey, Muki; Horne, David J; Wells, Richard D; Peterson, Glenna J; Cox, Jeffery S; Daya, Michelle; Hoal, Eileen G; Lin, Lin; Gottardo, Raphael; Hanekom, Willem A; Scriba, Thomas J; Hatherill, Mark; Hawn, Thomas R

    2017-08-15

    The molecular mechanisms that regulate tuberculosis susceptibility and bacillus Calmette-Guérin (BCG)-induced immunity are mostly unknown. However, induction of the adaptive immune response is a critical step in host control of Mycobacterium tuberculosis. Toll-interacting protein (TOLLIP) is a ubiquitin-binding protein that regulates innate immune responses, including Toll-like receptor signaling, which initiate adaptive immunity. TOLLIP variation is associated with susceptibility to tuberculosis, but the mechanism by which it regulates tuberculosis immunity is poorly understood. To identify functional TOLLIP variants and evaluate the role of TOLLIP variation on innate and adaptive immune responses to mycobacteria and susceptibility to tuberculosis. We used human cellular immunology approaches to characterize the role of a functional TOLLIP variant on monocyte mRNA expression and M. tuberculosis-induced monocyte immune functions. We also examined the association of TOLLIP variation with BCG-induced T-cell responses and susceptibility to latent tuberculosis infection. We identified a functional TOLLIP promoter region single-nucleotide polymorphism, rs5743854, which was associated with decreased TOLLIP mRNA expression in infant monocytes. After M. tuberculosis infection, TOLLIP-deficient monocytes demonstrated increased IL-6, increased nitrite, and decreased bacterial replication. The TOLLIP-deficiency G/G genotype was associated with decreased BCG-specific IL-2 + CD4 + T-cell frequency and proliferation. This genotype was also associated with increased susceptibility to latent tuberculosis infection. TOLLIP deficiency is associated with decreased BCG-specific T-cell responses and increased susceptibility to tuberculosis. We hypothesize that the heightened antibacterial monocyte responses after vaccination of TOLLIP-deficient infants are responsible for decreased BCG-specific T-cell responses. Activating TOLLIP may provide a novel adjuvant strategy for BCG

  17. Regulation of Exacerbated Immune Responses in Human Peripheral Blood Cells by Hydrolysed Egg White Proteins

    Science.gov (United States)

    Lozano-Ojalvo, Daniel; Molina, Elena; López-Fandiño, Rosina

    2016-01-01

    The anti-allergic potential of egg white protein hydrolysates (from ovalbumin, lysozyme and ovomucoid) was evaluated as their ability to hinder cytokine and IgE production by Th2-skewed human peripheral blood mononuclear cells (PBMCs), as well as the release of pro-inflammatory factors and generation of reactive oxygen species from Th1-stimulated peripheral blood leukocytes (PBLs). The binding to IgE of egg allergic patients was determined and the peptides present in the hydrolysates were identified. The hydrolysates with alcalase down-regulated the production of Th2-biased cytokines and the secretion of IgE to the culture media of Th2-skewed PBMCs, and they significantly neutralized oxidative stress in PBLs. The hydrolysates of ovalbumin and ovomucoid with pepsin helped to re-establish the Th1/Th2 balance in Th2-biased PBMCs, while they also inhibited the release of pro-inflammatory mediators and reduced oxidative stress in PBLs treated with inflammatory stimuli. The hydrolysates with alcalase, in addition to equilibrating Th2 differentiation, exhibited a low IgE-binding. Therefore, they would elicit mild allergic reactions while retaining T cell-stimulating abilities, which might correlate with an anti-allergic benefit. PMID:27007699

  18. Regulation of Exacerbated Immune Responses in Human Peripheral Blood Cells by Hydrolysed Egg White Proteins.

    Directory of Open Access Journals (Sweden)

    Daniel Lozano-Ojalvo

    Full Text Available The anti-allergic potential of egg white protein hydrolysates (from ovalbumin, lysozyme and ovomucoid was evaluated as their ability to hinder cytokine and IgE production by Th2-skewed human peripheral blood mononuclear cells (PBMCs, as well as the release of pro-inflammatory factors and generation of reactive oxygen species from Th1-stimulated peripheral blood leukocytes (PBLs. The binding to IgE of egg allergic patients was determined and the peptides present in the hydrolysates were identified. The hydrolysates with alcalase down-regulated the production of Th2-biased cytokines and the secretion of IgE to the culture media of Th2-skewed PBMCs, and they significantly neutralized oxidative stress in PBLs. The hydrolysates of ovalbumin and ovomucoid with pepsin helped to re-establish the Th1/Th2 balance in Th2-biased PBMCs, while they also inhibited the release of pro-inflammatory mediators and reduced oxidative stress in PBLs treated with inflammatory stimuli. The hydrolysates with alcalase, in addition to equilibrating Th2 differentiation, exhibited a low IgE-binding. Therefore, they would elicit mild allergic reactions while retaining T cell-stimulating abilities, which might correlate with an anti-allergic benefit.

  19. A single immunization with a recombinant canine adenovirus expressing the rabies virus G protein confers protective immunity against rabies in mice

    International Nuclear Information System (INIS)

    Li Jianwei; Faber, Milosz; Papaneri, Amy; Faber, Marie-Luise; McGettigan, James P.; Schnell, Matthias J.; Dietzschold, Bernhard

    2006-01-01

    Rabies vaccines based on live attenuated rabies viruses or recombinant pox viruses expressing the rabies virus (RV) glycoprotein (G) hold the greatest promise of safety and efficacy, particularly for oral immunization of wildlife. However, while these vaccines induce protective immunity in foxes, they are less effective in other animals, and safety concerns have been raised for some of these vaccines. Because canine adenovirus 2 (CAV2) is licensed for use as a live vaccine for dogs and has an excellent efficacy and safety record, we used this virus as an expression vector for the RVG. The recombinant CAV2-RV G produces virus titers similar to those produced by wild-type CAV2, indicating that the RVG gene does not affect virus replication. Comparison of RVG expressed by CAV2-RV G with that of vaccinia-RV G recombinant virus (V-RG) revealed similar amounts of RV G on the cell surface. A single intramuscular or intranasal immunization of mice with CAV2-RVG induced protective immunity in a dose-dependent manner, with no clinical signs or discomfort from the virus infection regardless of the route of administration or the amount of virus

  20. Immunization of Mice by BCG Formulated HCV Core Protein Elicited Higher Th1-Oriented Responses Compared to Pluronic-F127 Copolymer

    Science.gov (United States)

    Yazdanian, Maryam; Memarnejadian, Arash; Mahdavi, Mehdi; Sadat, Seyed Mehdi; Motevali, Fatemeh; Vahabpour, Rouhollah; Khanahmad, Hossein; Siadat, Seyed Davar; Aghasadeghi, Mohammad Reza; Roohvand, Farzin

    2013-01-01

    Background A supreme vaccine for Hepatitis C virus (HCV) infection should elicit strong Th1-oriented cellular responses. In the absence of a Th1-specific adjuvant, immunizations by protein antigens generally induce Th2-type and weak cellular responses. Objectives To evaluate the adjuvant effect of BCG in comparison with nonionic copolymer-Pluronic F127 (F127) as a classic adjuvant in the formulation of HCV core protein (HCVcp) as a candidate vaccine for induction of Th1 immune responses. Materials and Methods Expression of N-terminally His-Tagged HCVcp (1-122) by pIVEX2.4a-core vector harboring the corresponding gene under the control of arabinose-inducible (araBAD) promoter was achieved in BL21-AI strain of E.coli and purified through application of nitrilotriacetic acid (Ni-NTA) chromatography. Mice were immunized subcutaneously (s.c.) in base of the tail with 100 μl of immunogen (F127+HCVcp or BCG+HCVcp; 5 μgHCVcp/mouse/dose) or control formulations (PBS, BCG, F127) at weeks 0, 3, 6. Total and subtypes of IgG, as well as cellular immune responses (Proliferation, In vivo CTL and IFN-γ/IL-4 ELISpot assays against a strong and dominant H2-d restricted, CD8+-epitopic peptide, core 39-48; RRGPRLGVRA of HCVcp) were compared in each group of immunized animals. Results Expression and purification of core protein around the expected size (21 kDa) was confirmed by Western blotting. The HCVcp + BCG vaccinated mice showed significantly higher lymphocyte proliferation and IFN-γ production but lower levels of cell lysis (45% versus 62% in CTL assay) than the HCVcp+F127 immunized animals. “Besides, total anti-core IgG and IgG1 levels were significantly higher in HCVcp + F127 immunized mice as compared to HCVcp + BCG vaccinated animals, indicating relatively higher efficacy of F127 for the stimulation of humoral and Th2-oriented immune responses”. Conclusions Results showed that HCVcp + BCG induced a moderate CTL and mixed Th1/Th2 immune responses with higher levels of

  1. Immunization against Clostridium perfringens cells elicits protection against Clostridium tetani in mouse model: identification of cross-reactive proteins using proteomic methodologies

    Directory of Open Access Journals (Sweden)

    Singh Lokendra

    2008-11-01

    Full Text Available Abstract Background Clostridium tetani and Clostridium perfringens are among the medically important clostridial pathogens causing dis