bluuebunny/crossref_metadata_2025 huggingface dataset:
Mutation of α1GT-type Calcium Channels in Mice Does Not Change Anesthetic Requirements for Loss of the Righting Reflex and Minimum Alveolar Concentration but Delays the Onset of Anesthetic Induction
Authors: Petrenko Andrey B. , Tsujita Mika , Kohno Tatsuro , Sakimura Kenji , Baba Hiroshi
Published: May 2007

Background

T-type calcium channels regulate neuronal membrane excitability and participate in a number of physiologic and pathologic processes in the central nervous system, including sleep and epileptic activity. Volatile anesthetics inhibit native and recombinant T-type calcium channels at concentrations comparable to those required to produce anesthesia. To determine whether T-type calcium channels are involved in the mechanisms of anesthetic action, the authors examined the effects of general anesthetics in mutant mice lacking alpha1G T-type calcium channels.

Methods

The hypnotic effects of volatile and intravenous anesthetics administered to mutant and C57BL/6 control mice were evaluated using the behavioral endpoint of loss of righting reflex. To investigate the immobilizing effects of volatile anesthetics in mice, the minimum alveolar concentration (MAC) values were determined using the tail-clamp method.

Results

The 50% effective concentration for loss of righting reflex and MAC values for volatile anesthetics were not altered after alpha1G channel knockout. However, mutant mice required significantly more time to develop anesthesia/hypnosis after exposure to isoflurane, halothane, and sevoflurane and after intraperitoneal administration of pentobarbital.

Conclusions

The 50% effective concentration for loss of righting reflex and MAC values for the volatile anesthetics were not altered after alpha1G calcium channel knockout, indicating that normal functioning of alpha1G calcium channels is not required for the maintenance of anesthetic hypnosis and immobility. However, the timely induction of anesthesia/hypnosis by volatile anesthetic agents and some intravenous anesthetic agents may require the normal functioning of these channel subunits.
Mechanical Pain Hypersensitivity after Incisional Surgery Is Enhanced in Rats Subjected to Neonatal Peripheral Inflammation
Authors: Chu Ya-Chun , Chan Kwok-Hon , Tsou Mei-Yung , Lin Su-Man , Hsieh Ying-Chou , Tao Yuan-Xiang
Published: May 2007

Background

Neonatal pain and inflammation may lead to a long-term effect on nociceptive processing in adults. The current study examined the characteristics of postoperative incisional pain behaviors in adult rats that were subjected to neonatal peripheral inflammation.

Methods

Rat pups received a subcutaneous injection of saline or carrageenan into the plantar surface of the left hind paw at postnatal day 1. Naive pups were used as the control. Paw withdrawal thresholds to punctuate mechanical stimuli were examined at postnatal days 35, 42, and 49. After rats received a plantar incision on the left or right hind paw at postnatal day 50, paw withdrawal thresholds were measured at 4 h, 1 day, 2 days, 3 days, 5 days, and 7 days after incision. In addition, spinal cord Fos expression was detected at 2 h after incision. Finally, the effects of intrathecal N-methyl-D-aspartate receptor antagonists DL-2-amino-5-phosphonovaleric acid and dizocilpine and the nitric oxide synthase inhibitor L-N-nitro-arginine methylester on incisional pain were examined at 4 h after incision.

Results

Although the rats subjected to neonatal peripheral carrageenan injection developed mechanical hypoalgesia in bilateral hind paws at baseline, they displayed increased spinal cord Fos expression at 2 h and exaggerated mechanical pain hypersensitivity at 4 h (but not at other time points) after plantar incision. Intrathecal DL-2-amino-5-phosphonovaleric acid, dizocilpine, and L-N-nitro-arginine methylester significantly attenuated incision-induced mechanical pain hypersensitivity at 4 h after incision in the neonatally carrageenan-treated rats, but not in the naive or neonatally saline-treated rats.

Conclusions

The authors' results suggest that early inflammatory insults during the neonatal period could produce excessive incision-associated mechanical pain hypersensitivity in adult rats. Spinal cord N-methyl-D-aspartate receptors and downstream nitric oxide signaling might contribute to this abnormal pain hypersensitivity, although the mechanisms underlying the long-term effect of neonatal inflammation are still unclear.
Oral Gabapentin Activates Spinal Cholinergic Circuits to Reduce Hypersensitivity after Peripheral Nerve Injury and Interacts Synergistically with Oral Donepezil
Authors: Hayashida Ken-ichiro , Parker Renée , Eisenach James C.
Published: May 2007

Background

Gabapentin administration into the brain of mice reduces nerve injury-induced hypersensitivity and is blocked by intrathecal atropine and enhanced by intrathecal neostigmine. The authors tested the relevance of these findings to oral therapy by examining the efficacy of oral gabapentin to reduce hypersensitivity after nerve injury in rats and its interaction with the clinically used cholinesterase inhibitor, donepezil.

Methods

Male rats with hypersensitivity after spinal nerve ligation received gabapentin orally, intrathecally, and intracerebroventricularly with or without intrathecal atropine, and withdrawal threshold to paw pressure was determined. The effects of oral gabapentin and donepezil alone and in combination on withdrawal threshold were determined in an isobolographic design.

Results

Gabapentin reduced hypersensitivity to paw pressure by all routes of administration, and was more potent and with a quicker onset after intracerebroventricular than intrathecal injection. Intrathecal atropine reversed the effect of intracerebroventricular and oral gabapentin. Oral gabapentin and donepezil interacted in a strongly synergistic manner, with an observed efficacy at one tenth the predicted dose of an additive interaction. The gabapentin-donepezil combination was reversed by intrathecal atropine.

Conclusions

Although gabapentin may relieve neuropathic pain by actions at many sites, these results suggest that its actions in the brain to cause spinal cholinergic activation predominate after oral administration. Side effects, particularly nausea, cannot be accurately determined on rats. Nevertheless, oral donepezil is well tolerated by patients in the treatment of Alzheimer dementia, and the current study provides the rationale for clinical study of combination of gabapentin and donepezil to treat neuropathic pain.
Molecular analysis of the NSP4 and VP6 genes of rotavirus strains recovered from hospitalized children in Rio de Janeiro, Brazil
Authors: Araújo Irene Trigueiros , Heinemann Marcos Bryan , Mascarenhas Joana D'Arc P. , Assis Rosane M. Santos , Fialho Alexandre Madi , Leite José Paulo G.
Published: May 2007

Group A rotaviruses are the main cause of acute gastroenteritis in children throughout the world. The two outer capsid proteins, VP4 and VP7, define the P and G genotypes, respectively. Rotaviruses with P[8]G1, P[4]G2, P[8]G3 and P[8]G4 genotypes are predominant in infecting humans and the G9 genotype is emerging in most continents as the fifth most common G type worldwide. The inner capsid protein VP6 is responsible for subgroup (SG) specificities, allowing classification of rotaviruses into SG I, SG II, SG I+II and SG non-I-non-II. The non-structural protein 4 (NSP4) encoded by segment 10 has a role in viral morphogenesis and five genetic groups have been described, NSP4 genotypes A–E. The aim of this investigation was to characterize the NSP4 and VP6 genes of rotavirus strains recovered from hospitalized children. Thirty rotavirus strains were submitted to RT-PCR followed by sequencing and phylogenetic analysis. Among the different G and P genotype combinations, two distinct genetic groups could be recognized for the NSP4 gene. Twenty-eight clustered with NSP4 genotype B. The two P[4]G2 strains fell into NSP4 genotype A and clustered distinctly, with a 100 % bootstrap value. The strains distinguished within a group were closely related to each other at the nucleotide and amino acid levels. A phylogenetic tree was constructed for the VP6 gene including the human strains RMC100, E210, Wa, US1205 and 1076, and the animal strains Gott, NCDV, SA-11, FI-14 and EW. This is the first report on Brazilian rotavirus strains describing NSP4 genotype A strains associated with VP6 SG I, and NSP4 genotype B strains associated with VP6 SG II.
Identification of a group of shigella-like isolates as Shigella boydii 20
Authors: Grimont Francine , Lejay-Collin Monique , Talukder Kaisar A. , Carle Isabelle , Issenhuth Sylvie , Le Roux Karine , Grimont Patrick A. D.
Published: May 2007

Infections by Shigella species are an important cause of diarrhoeal disease worldwide. Of 4198 Shigella isolates received by the French National Reference Centre for Escherichia coli and Shigella, 180 from patients with diarrhoea and dysentery in 2000–2004 did not react with any available polyclonal rabbit antisera used to identify the established Shigella serogroups. This study describes the molecular and phenotypic characteristics of these isolates in seroagglutination tests, molecular serotyping (rfb-RFLP and fliC-RFLP), ribotyping, detection of invasivity and enterotoxins genes, and antibiotic sensitivity. All isolates gave biochemical reactions typical of Shigella boydii, were mannitol-positive and indole-negative. They all carried invasion-associated genes, enterotoxin 2 [ShET-2] and an IS630 sequence. They had a unique ribotype that was distinct from all other Shigella and E. coli patterns. Further characterization by rfb-RFLP clearly distinguished this serogroup from all other Shigella or E. coli O-groups. The fliC-RFLP pattern corresponded to P4, an F-pattern which is associated with 10 different serogroups of S. boydii. A new antiserum prepared against strain 00-977 agglutinated all 180 isolates and cross-agglutination and absorption studies with anti-00-977 serum and anti-CDC 99-4528 (reference for the newly described S. boydii serogroup 20) serum showed identical antigenic structure. Furthermore, strains 00-977 and CDC 99-4528 had the same molecular serotype, ribotype and virulence genes.
Two diallyl sulphides derived from garlic inhibit meticillin-resistant Staphylococcus aureus infection in diabetic mice
Authors: Tsao Shih-Ming , Liu Wen-Hu , Yin Mei-Chin
Published: May 2007

The inhibitory effect of diallyl sulphide (DAS) and diallyl disulphide (DADS) against meticillin-resistant Staphylococcus aureus (MRSA) infection in diabetic mice was studied. The influence of these agents on the plasma levels of fibronectin, C-reactive protein (CRP), fibrinogen, interleukin (IL)-6 and tumour necrosis factor-α (TNF-α), and on the activity of plasminogen activator inhibitor-1 (PAI-1), antithrombin III (AT-III) and protein C, in MRSA-infected diabetic mice was examined. To induce diabetes, mice were treated intraperitoneally with streptozotocin for 5 consecutive days. Ten clinical MRSA isolates obtained from infected patients were used in this study. Diabetic mice were infected by injecting 200 μl MRSA/PBS suspension containing 107 c.f.u. via the tail vein. At day 4 post-infection, 200 μl DAS or DADS was administrated twice orally with an interval of 12 h. Eight hours after each administration, the blood and organs of mice were collected. Results showed that DAS and DADS significantly decreased MRSA viability in the kidney (P <0.05), with administration of each agent twice showing a greater inhibitory effect than when given once (P <0.05). MRSA infection in diabetic mice significantly elevated the plasma levels of IL-6 and TNF-α (P <0.05). DAS or DADS given once did not affect the plasma levels of IL-6 and TNF-α (P >0.05); however, DAS or DADS given twice significantly decreased the plasma levels of both IL-6 and TNF-α (P <0.05). DAS and DADS treatments also significantly reduced the plasma levels of CRP, fibronectin and fibrinogen (P <0.05). DAS or DADS treatment did not affect PAI-1 activity (P >0.05), but DAS or DADS given twice significantly increased AT-III activity (P <0.05). DADS given twice elevated protein C activity (P <0.05). MRSA infection significantly increased malondialdehyde levels in the kidney and spleen (P <0.05), and these levels were significantly decreased by treatment with DAS or DADS (P <0.05). These data suggest that DAS and DADS could provide multiple protective functions against MRSA infection in diabetic mice.
Role of AmpD, OprF and penicillin-binding proteins in β-lactam resistance in clinical isolates of Pseudomonas aeruginosa
Authors: Bratu Simona , Landman David , Gupta Jyoti , Quale John
Published: May 2007

In this study, the mechanisms leading to increased chromosomal AmpCβ-lactamase expression and the contributory roles of the outer-membrane protein OprF and penicillin-binding proteins were analysed in 33 characterized clinical isolates ofPseudomonas aeruginosa. The genesampDandampEwere analysed by PCR and DNA sequencing. Expression of the geneoprFwas assessed using real-time RT-PCR, and penicillin-binding proteins were analysed using a chemiluminescence assay. Several of the isolates with increasedampCexpression had major deletions affectingampD, although in some isolates the mechanism of increasedampCexpression could not be ascertained. Occasional isolates had increased expression of bothampCandoprFbut remained susceptible to cephalosporins, suggesting that increasedβ-lactamase activity could not offset increased outer-membrane permeability. There were no discernible changes in penicillin-binding proteins. Genomic deletions inampDwere observed in selected clinical isolates ofP. aeruginosawith increased expression of the AmpCβ-lactamase. For some isolates, cephalosporin resistance was dependent upon the interplay ofampCandoprFexpression.
Diversity of biofilms produced by quorum-sensing-deficient clinical isolates of Pseudomonas aeruginosa
Authors: Schaber J. Andy , Hammond Adrienne , Carty Nancy L. , Williams Simon C. , Colmer-Hamood Jane A. , Burrowes Ben H. , Dhevan Vijian , Griswold John A. , Hamood Abdul N.
Published: May 2007

The quorum-sensing (QS) systems control several virulence attributes of Pseudomonas aeruginosa. Five QS-deficient P. aeruginosa clinical isolates (CI) that were obtained from wound (CI-1), tracheal (CI-2, CI-3, CI-4) and urinary tract (CI-5) infections had previously been characterized. In this study, a flow-through continuous-culture system was utilized to examine in detail the biofilms formed by these isolates in comparison with the P. aeruginosa prototrophic strain PAO1. Analysis of the biofilms by confocal laser scanning microscopy and COMSTAT image analysis at 1 and 7 days post-inoculation showed that the isolates produced diverse biofilms. In comparison with PAO1, the CI produced biofilms that scarcely or partially covered the surface at day 1, although CI-1 produced larger microcolonies. At day 7, CI-2 and CI-4 produced mature biofilms denser than that produced by PAO1, while the biofilm formed by CI-1 changed very little from day 1. CI-1 was defective in both swarming and twitching motilities, and immunoblotting analysis confirmed that it produced a reduced level of PilA protein. The twitching-motility defect of CI-1 was not complemented by a plasmid carrying intact pilA. In the 48 h colony biofilm assay, the CI varied in susceptibility to imipenem, gentamicin and piperacillin/tazobactam. These results suggest that: (1) the isolates produced biofilms with different structures and densities from that of PAO1; (2) biofilm formation by the isolates was not influenced by either the isolation site or the QS deficiencies of the isolates; (3) the behaviour of CI-1 in the different biofilm systems may be due to its lack of swarming motility and type IV pilus-related twitching motility.
Recurrent infection with genetically identical pneumococcal isolates in a patient with interleukin-1 receptor-associated kinase-4 deficiency
Authors: Szabó Judit , Dobay Orsolya , Erdős Melinda , Borbély Ágnes , Rozgonyi Ferenc , Maródi László
Published: May 2007

Interleukin-1 receptor-associated kinase (IRAK)-4 deficiency is a rare primary immunodeficiency disorder characterized by severe, invasive infections with Streptococcus pneumoniae. Using the PFGE technique a genetic linkage was found between two S. pneumoniae serotype 14 isolates causing arthritis and meningitis at 3 and 5½ years of age, respectively, in a boy with IRAK-4 deficiency. This finding suggested that patients with IRAK-4 deficiency may harbour persistent strains of pneumococci. Alternatively, reinfection with strains from close contacts of the patient might cause recurrent invasive disease. It is proposed that eradication of pneumococci from the nasopharynx, and immunization of household contacts may prevent recurrent infection in IRAK-4-deficient patients.
Effects of vaccination by a recombinant antigen ureB138 (a segment of the β-subunit of urease) against Helicobacter pylori infection
Authors: Morihara Fumiko , Fujii Ryoji , Hifumi Emi , Nishizono Akira , Uda Taizo
Published: May 2007

Helicobacter pylorihas to counteract acidity during colonization in the stomach. The most important region for the enzymic activity ofH. pyloriurease, consisting of 138 aa (ureB138), was determined by a comparison of the homology of amino acid sequences, and a structural analysis, between urease ofH. pyloriand various other species. This region was expressed inEscherichia colias a fusion protein with glutathione S-transferase (GST), which was cleaved by PreScission protease between the GST moiety and ureB138. The ureB138 protein was then purified by gel filtration. The polyclonal antibody (pAb) induced by immunization with the purified ureB138 could suppress urease activity by about 50 %, while the pAb against theH. pyloriurease did not show any inhibitory effect at all. Immunohistochemical analysis indicated that the ureB138-specific pAb specifically recognized theH. pyloriinfecting human gastric tissues. The effects of vaccination of recombinant ureB138 against infection by this organism were also examined. Specific IgG and IgA antibodies againstH. pyloriurease were induced in the serum of mice immunized with ureB138. A reduction in the number of colonizingH. pyloriwas observed in mice treated with ureB138 compared to ones treated with BSA and infection control mice. In the protected mice, severe gastritis characterized by marked infiltration of mononuclear cells was noted compared with the gastritis observed in unprotected mice. Immunohistochemical staining for IgA in gastric mucosa showed that the number of mice positively stained with IgA was significantly higher in ureB138-vaccinated mice than in non-vaccinated mice. This indicates that local IgA antibody and severe post-immunization gastritis correlate well with the protection of mice againstH. pyloriinfection.
Role of the plasmid-encoded tet(O) gene in tetracycline-resistant clinical isolates of Campylobacter jejuni and Campylobacter coli
Authors: Dasti Javid Iqbal , Groß Uwe , Pohl Sven , Lugert Raimond , Weig Michael , Schmidt-Ott Ruprecht
Published: May 2007

The prevalence of tetracycline resistance, tetracycline MICs andtet(O) gene localization were investigated in 83Campylobacterisolates from patients suffering from acute gastroenteritis in Germany. Combined biochemical and molecular markers identified 74 isolates (89 %) asCampylobacterjejuni, including seven atypical isolates that failed to hydrolyse hippurate, and nine isolates (11 %) asCampylobactercoli. Tetracycline resistance was detected in six out of nineCampylobacter coliisolates (67 %) and 13 out of 74C. jejuniisolates (18 %). Low-level tetracycline resistance was observed forC. coli(MIC 16 μg ml−1for all strains), whereasC. jejunishowed high-level resistance (MIC >256 μg ml−1for all strains). Both low- and high-level tetracycline resistance was associated with the presence of thetet(O) gene. InC. jejuni,tet(O) was plasmid-encoded in 54 % of tetracycline-resistant isolates, whereas inC. coli,tet(O) appeared to be located on the chromosome.
Post-antifungal effects of the antifungal compound 2-(3,4-dimethyl-2,5-dihydro-1H-pyrrol-2-yl)-1-methylethyl pentanoate on Aspergillus fumigatus
Authors: Dabur Rajesh , Mandal T. K. , Sharma G. L.
Published: May 2007

The post-antifungal effect (PAFE) of the antifungal compound 2-(3,4-dimethyl-2,5-dihydro-1H-pyrrol-2-yl)-1-methylethyl pentanoate (DHP) upon Aspergillus fumigatus was investigated. The conidia of A. fumigatus were exposed to DHP at concentrations of 1× and 4× MIC90 for variable times at 37 °C. Amphotericin B (AmB)-treated or drug-free controls were included in the study. DHP as well as AmB exposure resulted in prolonged lag phases of the turbidimetric growth curves. Both the treatments gave rise to delayed growth, with lag phases of 11 h upon treatment with a concentration of 4× MIC90 for 4 h. Furthermore, it was observed that DHP inhibited the expression of three A. fumigatus secretory proteins of 18, 42 and 55 kDa. One protein of 42 kDa was found to be a metalloprotease, which is an important virulence factor. Analysis of time-dependent antigenic profiles showed the early expression of high-molecular-mass antigens. Expression of low-molecular-mass antigens started after 24 h culture. The antigens of A. fumigatus that are expressed during the early phase of growth were observed to be adversely affected after treatment with DHP. Although the mechanism of action of DHP to inhibit these proteins/antigens is unknown, the observations may be valuable to understand their role in the virulence of the pathogen, as well as the antigen-mediated responses caused by A. fumigatus.
Diversity of antibiotic resistance integrative and conjugative elements among haemophili
Authors: Dimopoulou Ioanna D. , Kartali Sofia I. , Harding Rosalind M. , Peto Tim E. A. , Crook Derrick W.
Published: May 2007

The objective of this study was to investigate the sequence diversity in a single country of a family of integrative and conjugative elements (ICEs) that are vectors of antibiotic resistance in Haemophilus influenzae and Haemophilus parainfluenzae, and test the hypothesis that they emerged from a single lineage. Sixty subjects aged 9 months – 13 years were recruited and oropharyngeal samples cultured. Up to 10 morphologically distinct Pasteurellaceae spp. were purified, and then the species were determined and differentiated by partial sequence analysis of 16S rDNA and mdh, respectively. ICEs were detected by PCR directed at five genes distributed evenly across the ICE. These amplicons were sequenced and aligned by the neighbour-joining algorithm. A total of 339 distinguishable isolates were cultured. ICEs with all 5 genes present were found in 9 of 110 (8 %) H. influenzae and 21 of 211 (10 %) H. parainfluenzae, respectively. ICEs were not detected among the other Pasteurellaceae. A total of 20 of 60 (33 %) children carried at least 1 oropharyngeal isolate with an ICE possessing all 5 genes. One of the five genes, integrase, however, consisted of two lineages, one of which was highly associated with H. influenzae. The topology of neighbour-joining trees of the remaining four ICE genes was compared and showed a lack of congruence; though, the genes form a common pool among H. influenzae and H. parainfluenzae. This family of antibiotic resistance ICEs was prevalent among the children studied, was genetically diverse, formed a large gene pool, transferred between H. influenzae and H. parainfluenzae, lacked population structure and possessed features suggestive of panmixia, all indicating it has not recently emerged from a single source.
Prosthetic valve endocarditis due to Neisseria elongata subsp. elongata in a patient with Klinefelter's syndrome
Authors: Evans Morgan , Yazdani Farah , Malnick Henry , Shah Jayesh J. , Turner David P. J.
Published: May 2007

A case is reported of prosthetic valve endocarditis due to Neisseria elongata subsp. elongata in a patient with Klinefelter's syndrome. This is believed to be only the third case of endocarditis reported due to this subspecies. N. elongata is difficult to identify, and is morphologically and biochemically similar to Kingella spp. Sequencing of the 16S rRNA gene is useful for identification. The patient was successfully treated with amoxicillin and gentamicin, followed by ceftriaxone.
Real-time PCRs for detection of Trichomonas vaginalis
β-tubulin and 18S rRNA genes in female genital specimens
Authors: Simpson Paul , Higgins Geoff , Qiao Ming , Waddell Russell , Kok Tuckweng
Published: May 2007

Trichomonas vaginalis is the cause of one of the most common types of vaginitis, trichomoniasis. The incidence of trichomoniasis in developed countries has decreased substantially during the past decade, but high prevalence of this disease can still be found in rural and remote areas of Australia. Clinical manifestations of symptomatic women are generally non-specific, but include vaginal discharge, vaginitis and irritation. T. vaginalis infection has also been linked to the increased risk of human immunodeficiency virus transmission. Current diagnosis of T. vaginalis relies on the visualization of motile organisms in a wet-mount preparation. Culture is used mainly in reference laboratories. The latter two methods require viable organisms and would not be suitable for use where transportation of specimens can be delayed. Two real-time fluorescence resonance energy transfer (FRET) hybridization probe PCR assays were used in this study to test for T. vaginalis DNA, targeting the β-tubulin and 18S rRNA genes. We tested 500 randomly selected female patients, in an STD setting, for T. vaginalis DNA. The FRET PCRs targeting the β-tubulin gene and the 18S rRNA gene detected 96 % (85/89) and 100 % (89/89) , respectively, of the positive specimens (first-void urine sample or genital swabs). Wet-mount microscopy was performed on 76 of these PCR-positive specimens and showed a sensitivity of 38 % (29/76). The prevalence, by PCR, of trichomoniasis was 18 % in this study. The two real-time PCRs developed in this study, targeting different genetic regions of the organism, provide a rapid, sensitive and specific diagnosis of T. vaginalis infection.
PAR‐2 activation regulates IL‐8 and GRO‐α synthesis by NF‐κB, but not RANTES, IL‐6, eotaxin or TARC expression in nasal epithelium
Authors: Rudack C. , Steinhoff M. , Mooren F. , Buddenkotte J. , Becker K. , Von Eiff C. , Sachse F.
Published: May 2007

BackgroundThe effects of protease‐activated receptor‐2 (PAR‐2) stimulation on inflammation mechanisms of chronic rhinosinusitis (CRS) are still unknown.

MethodsPAR‐2 receptor expression was investigated by immunohistochemistry and Taqman mRNA analysis in the mucosa of different rhinosinusitis entities. In primary nasal epithelial cell cultures, the function of PAR‐2 and its ability to produce CXC, CC chemokines, and IL‐6 were measured by calcium mobilization and stimulation tests. Inhibition tests were performed using cortisone, serine protease inhibitors, cysteine protease inhibitors, Pertussis toxin (PTX) and nuclear transcription factor (NF‐κB) inhibition (BAY 11‐7085). Signal transduction pathways were analysed by electromobility shift assays (EMSA) and NF‐κB binding studies.

ResultsThe expression of PAR‐2 was found to be increased in CRS specimens. The activation of PAR by trypsin or PAR‐2‐specific activating peptide (AP) caused an increase in cytosolic calcium, as well as the release of the CXC chemokines IL‐8 and growth‐related oncogene (GRO)‐α, but not the release of CC chemokines or IL‐6. AP‐induced CXC chemokine was sensitive to PTX and activation of NF‐κB was inhibited by BAY11‐7085. Furthermore, a serine protease inhibitor significantly inhibited chemokine synthesis stimulated by trypsin and culture supernatants of staphylococci, whereas steroids and cysteine protease inhibitors had little effect.

ConclusionPAR‐2 plays a role in serine protease‐mediated regulation – staphylococcal and non‐staphylococcal origin – of IL‐8 and GRO‐α in nasal epithelial cells, but not in the regulation of CC chemokines. PAR‐2 may therefore be involved in the pathophysiology of CRS and NP at different sites of activation, namely (i) proteases, (ii) the PAR‐2 receptor itself or (iii) the application of novel agents that block NF‐κB/IκB‐α signalling.
Mutational analysis of immunoglobulin E‐binding epitopes of β‐casein and β‐lactoglobulin showed a heterogeneous pattern of critical amino acids between individual patients and pooled sera
Authors: Cocco R. R. , Järvinen K.‐M. , Han N. , Beyer K. , Sampson H. A.
Published: May 2007

Background
For immunotherapeutic approaches, ‘critical’ amino acids (AAs) within allergenic epitopes are replaced with alternate AAs to eliminate IgE antibody binding.

Objective
To determine the critical AAs for IgE binding in β‐casein and β‐lactoglobulin (BLG).

Methods
Peptides of 10–14 AAs in length were synthesized on a derivatized cellulose membrane with single AA substitutions (alanine or glycine) at each position. Membranes were incubated with a pool of sera from 15 cow's milk‐allergic patients and individual sera from six of the 15 patients. In cases where no decrease in binding occurred with a single AA substitution, peptides with two AA substitutions were generated and labelled.

Results
Using pooled patient sera, single AA substitutions led to complete elimination of binding to six of 11 peptides for β‐casein and to all six peptides for BLG. Substituting two AAs led to an elimination of binding to four of the remaining five β‐casein epitopes. However, in three of the 11 modified β‐casein peptides and five of the six BLG peptides, no decrease in IgE binding occurred in at least one individual patient. For these patients, critical AAs other than those defined by the patient serum pool were identified, indicating a heterogeneous pattern of IgE recognition.

Conclusion
These results indicate that AAs critical for IgE binding are more heterogeneous than initially defined by pooled milk‐allergic patient sera. For future immunotherapeutic interventions with mutated peptides, critical AAs should also be identified with individual patient sera to account for heterogeneity of IgE binding between patients.
Association of the single‐nucleotide polymorphism and haplotype of the interleukin 18 gene with atopic dermatitis in Koreans
Authors: Kim E. , Lee J.‐E. , Namkung J.‐H. , Park J.‐H. , Kim S. , Shin E.‐S. , Cho E.‐Y. , Yang J.‐M.
Published: May 2007

Background
IL‐18 is a proinflammatory cytokine that plays an important role for T‐helper type 1 (Th1) and Th2 cytokine in the presence/absence of IL‐12. It has been recently shown that human IL‐18 plays a role in atopic dermatitis (AD) by enhancing IL‐4 and IL‐13 production and by stimulating the synthesis of IgE.

Objective
We wanted to evaluate the associations of single‐nucleotide polymorphism (SNP) and the haplotype in the IL‐18 gene, hence we performed genotyping for the SNPs in the IL‐18 gene in AD patients and normal controls.

Method
We genotyped three SNPs from the IL‐18 gene for the 1120 case–control samples (646 AD patients and 474 normal controls). We measured the serum IL‐18, IL‐4 and IL‐13 concentrations in 74 individuals (25 ADe, 25 ADi and 24 controls) by performing ELISA.

Result
The rs795467 SNP and haplotype T–T–C were significantly associated with AD, and especially between the ADe and normal control groups (P=0.03 and 0.01). The serum IL‐18 concentration was higher in the AD group than in the normal controls without any correlation with the rs795467 polymorphism. We did not find any correlations between the serum IL‐18 levels and the SCORing atopic dermatitis index, the blood eosinophil counts and the ECP, and there was no correlation between the serum IL‐18 levels and the serum IL‐4 and IL‐13 levels.

Conclusion
We found that the one SNP and the haplotype T–T–C were strongly associated with the allergic type of AD, but not with the non‐allergic intrinsic type. These data support the hypothesis that IL‐18 up‐regulates IgE production, yet more experiments will be needed to prove the in vivo involvement of Th2 cytokine.
Induction of tolerance after establishment of peanut allergy by the food allergy herbal formula‐2 is associated with up‐regulation of interferon‐γ
Authors: Qu C. , Srivastava K. , Ko J. , Zhang T. F. , Sampson H. A. , Li X.‐M.
Published: May 2007

Background
Peanut (PN)‐anaphylaxis is potentially life threatening. We previously reported that a Chinese herbal medicine preparation, food allergy herbal formula‐2 (FAHF‐2), prevented peanut allergy (PNA) in mice when administered during sensitization.

Objective
To investigate whether FAHF‐2 also can prevent anaphylactic reactions when administered to mice with established PNA and, if so, whether protection would persist after cessation of therapy.

Methods
C3H/HeJ mice sensitized and boosted over 8 weeks with a standard protocol known to establish PN hypersensitivity received seven weeks of FAHF‐2 treatment or water as a sham treatment. Mice were subsequently challenged with PN at week 14 (1‐day post‐therapy) and week 18 (4‐week post‐therapy) to evaluate the efficacy and persistence of FAHF‐2 treatment by assessing anaphylactic scores, core body temperatures and plasma histamine levels. Serum PN‐specific antibody levels and cytokine profiles from splenocytes and mesenteric lymph node (MLN) cells were also determined.

Results
All sham‐treated mice challenged at weeks 14 and 18 showed anaphylactic symptoms. In contrast, FAHF‐2‐treated mice showed no sign of anaphylactic reactions. PN‐specific IgE levels in FAHF‐2‐treated mice also were reduced whereas IgG2a levels were increased. Furthermore, MLN cells from FAHF‐2‐treated mice produced markedly less IL‐4 and IL‐5, but more IFN‐γ, and contained increased numbers of IFN‐γ‐producing CD8+ cells as compared with sham‐treated mice.

Conclusion
FAHF‐2 treatment established PN tolerance in this model, which persisted for at least 4‐week post‐treatment. This result was associated with modulation of intestinal T helper type 1 cell (Th1) and Th2 cytokine production, and with increased numbers of mesenteric IFN‐γ‐producing CD8+ cells.
Genetically glycosylated ovomucoid third domain can modulate Immunoglobulin E antibody production and cytokine response in BALB/c mice
Authors: Rupa P. , Nakamura S. , Mine Y.
Published: May 2007

Background
Food allergies are on the rise and it is estimated that in North America, 8% of the children and 4% of the adults have food allergies. Food allergies tend to occur more often in children than in adults due to their immature digestive and immune systems. Hen's egg is among the most common cause of food‐induced allergic reactions in North America.

Objective
The present study was undertaken to investigate the role of N‐glycans of the third domain of ovomucoid in IgE binding and modulation of allergen‐specific immune response in BALB/c mice.

Methods
The cDNA encoding the third domain of ovomucoid was inserted into the yeast genome and expressed in Pichia pastoris X‐33 cells, under the control of the glyceraldehyde‐3‐phosphate (GAP) dehydrogenase promoter for constitutive expression to obtain a post‐translationally modified and functionally active ovomucoid third domain. Upon expression, the protein was secreted into the extracellular medium and was purified by size exclusion chromatography. The recombinant protein was produced at 10 mg/L of the culture supernatant. BALB/c mice were sensitized with the recombinant and native forms of glycosylated ovomucoid third domain antigen. The allergic response of the native and the recombinant glycosylated forms of ovomucoid third domain antigens were compared using antibody and cytokine measurements.

Results
ELISA tests indicated a significant decrease in specific IgE antibodies to the recombinant N‐linked glycosylated form (P‐Gly), when compared with the native glycosylated form (DIII+) using mice sera. Immunization with P‐Gly induced the production of IFN‐γ [T‐helper type 1 (Th1) response] and lowered the production of IL‐4 (Th2 response), and a skewed balance towards the Th1 cytokine demonstrated that P‐Gly has a modulating ability on Th1/Th2 balance to down‐regulate Th2 response. Furthermore, N‐linked glycan (N28) in the third domain of ovomucoid was shown to be associated with suppression of the allergic response.

Conclusion
Therefore, we can conclude that P‐Gly facilitates and contributes to the discovery of new molecular target for the development of a safe and specific therapeutic vaccine for the treatment of egg allergy, and oligosaccharides do seem to play a major role in the suppression of IgE‐binding activity.
CD4+ T cells from mice with intestinal immediate‐type hypersensitivity induce airway hyperreactivity
Authors: Ozdemir C. , Sel S. , Schöll I. , Yildirim A. Ö. , Bluemer N. , Garn H. , Ackermann U. , Wegmann M. , Barlan I. B. , Renz H. , Sel S.
Published: May 2007

Background
A subset of food‐allergic patients does not only respond clinically with symptoms in the gastro‐intestinal tract but also with asthmatic reactions.

Objective
The aim of this study was to analyse whether CD4+ T cells from mice with intestinal immediate‐hypersensitivity reactions to food allergen are involved in the development of experimental asthma.

Methods
BALB/c mice were intraperitoneally sensitized to ovalbumin (OVA), followed by repeated intra‐gastric (i.g.) OVA challenges. Control animals were either sham‐sensitized or sham‐challenged with phosphate‐buffered saline (PBS). Duodenum, jejunum, ileum and colon were histologically examined. CD4+ T cells from mesenteric lymph nodes were transferred from various donor groups into recipient mice that received either OVA or PBS aerosol challenges. Recipients were analysed by measurements of lung function using head‐out body‐plethysmography and examination of broncho‐alveolar lavage and lung histology.

Results
The highest levels of OVA‐specific IgE antibody levels were detected in OVA‐sensitized and OVA‐challenged mice. Throughout the lower intestinal tract, a marked infiltration with eosinophils was observed, and goblet cell numbers as well as goblet cell area were significantly increased. The villus/crypt ratio was decreased compared with controls. The transfer of CD4+ T cells from mesenteric lymph nodes of OVA‐sensitized and OVA‐challenged mice triggered airway hyperreactivity and eosinophilic airway inflammation in recipients aerosol challenged with OVA, but not with PBS.

Conclusion
We conclude that CD4+ T cells from mesenteric lymph nodes of mice with allergen‐induced immediate‐type hypersensitivity reactions in the gut are able to transfer the phenotype of experimental asthma.
Ses i 6, the sesame 11S globulin, can activate basophils and shows cross‐reactivity with walnut in vitro
Authors: Wallowitz M. L. , Chen R. J. Y. , Tzen J. T. C. , Teuber S. S.
Published: May 2007

Background
Sesame allergy is increasingly being reported, and multi‐sensitization to peanut and tree nuts has been described. The clinical relevance and cross‐reactivity of many sesame proteins, such as Ses i 6, are unknown.

Objective
The aims of this study were to perform a preliminary examination of the cross‐reactivity of Ses i 6 in vitro, examine the ability of Ses i 6 to activate basophils in a modified basophil activation test (mBAT), and assess whether such an assay may help to distinguish between potentially relevant and irrelevant IgE reactivity towards 11S globulin proteins.

Methods
Inhibition immunoblotting and chicken anti‐rJug r 4 antibodies were used to determine the cross‐reactivity of rSes i 6. Basophils from atopic donors were stripped of resident IgE before passive sensitization with food‐allergic sera and challenged with protein extracts or recombinant protein. Basophil activation was measured using two activation markers, CD203c and CD63, via flow cytometry.

Results
IgE immunoblotting showed cross‐reactivity between rJug r 4 and rSes i 6 using sera from two human donors and chicken IgY. Additionally, rSes i 6 activated basophils passively sensitized with sesame‐allergic sera. Cross‐reactive serum from a sesame‐allergic but walnut‐tolerant donor was not able to activate basophils when challenged by walnut extract despite IgE reactivity to walnut determined by immunoblotting.

Conclusions
The sesame 11S globulin shows partial immunological cross‐reactivity with walnut, and although it is classified as a minor allergen, activated basophils sensitized with serum from seven out of eleven sesame‐allergic donors. Additionally, the mBAT may help distinguish between clinically relevant and irrelevant in vitro IgE cross‐reactivity of seed storage proteins in nuts and seeds and thus warrants use in further studies.
Frequency and significance of immediate contact reactions to peanut in peanut‐sensitive children
Authors: Wainstein B. K. , Kashef S. , Ziegler M. , Jelley D. , Ziegler J. B.
Published: May 2007

Background
Parents of atopic children frequently report, and are alarmed by, contact reactions to foods. Some schools restrict foods due to concerns regarding possible systemic reactions following contact in allergic children.

Objective
We aimed to determine the frequency with which peanut‐sensitive children exhibited contact sensitivity to peanut butter and to assess the significance of such reactions.

Methods
One gram of peanut butter was applied directly to the skin of 281 children who were skin prick test (SPT) positive to peanut (immediate skin application food test; I‐SAFT). The test was considered positive if one or more weals were present when the patch was removed after 15 min. A subset of children then underwent an open‐label oral challenge with graded amounts of peanut protein.

Results
During 3515 clinic visits, 330 I‐SAFT tests for peanut contact sensitivity were performed; 136 (41%) were positive. The mean SPT diameter was 10 mm in the I‐SAFT‐positive children and 8.5 mm in the I‐SAFT‐negative children (t‐test, P<0.0001). No child had a systemic reaction following topical application of peanut butter. Eighty‐four children had 85 oral challenges after blinded, placebo‐controlled I‐SAFT testing. Challenge was positive in 26/32 of those with a positive I‐SAFT and negative in only 6/32. Challenge was also positive in 26/53 but negative in 27/53 of those with a negative I‐SAFT (sensitivity 50%, specificity 82%, χ2, P=0.003).

Conclusion
A minority of children sensitized to peanut (positive SPT) develop localized urticaria from prolonged skin contact with peanut butter. No tested subjects, including ones with systemic reactions upon oral challenge, developed a systemic reaction to prolonged skin exposure to peanut. Therefore, systemic reactions resulting from this mode of contact with peanut butter appear highly unlikely.
Alterations of food antigen‐specific serum immunoglobulins G and E antibodies in patients with irritable bowel syndrome and functional dyspepsia
Authors: Zuo X. L. , Li Y. Q. , Li W. J. , Guo Y. T. , Lu X. F. , Li J. M. , Desmond P. V.
Published: May 2007

Background
Post‐prandial worsening of symptoms as well as adverse reactions to one or more foods are common in the patients with functional gastrointestinal diseases, such as irritable bowel syndrome (IBS) and functional dyspepsia (FD). However, the role played by true food allergy in the pathogenesis of these diseases is still controversial and there are no well‐established tests to identify food allergy in this condition.

Objective
To investigate serum food antigen‐specific IgG, IgE antibody and total IgE antibody titres in controls and patients with IBS and FD, and to correlate symptoms with the food antigen‐specific IgG titres in IBS and FD patients.

Methods
Thirty‐seven IBS patients, 28 FD patients and 20 healthy controls participated in this study. Serum IgG and IgE antibody titres to 14 common foods including beef, chicken, codfish, corn, crab, eggs, mushroom, milk, pork, rice, shrimp, soybean, tomatoes and wheat were analysed by ELISA. Serum total IgE titres were also measured. Last, symptomatology was assessed in the study.

Results
IBS patients had significantly higher titres of IgG antibody to crab (P=0.000), egg (P=0.000), shrimp (P=0.000), soybean (P=0.017) and wheat (P=0.004) than controls. FD patients had significantly higher titres of IgG antibody to egg (P=0.000) and soybean (P=0.017) than controls. The percentage of individuals with detectable positive food antigen‐specific IgE antibodies of the three groups did not show any significant differences (P=0.971). There were no significant differences between IBS patients, FD patients and controls in the serum total IgE antibody titres (P=0.978). Lastly, no significant correlation was seen between symptom severity and serum food antigen‐specific IgG antibody titres both in IBS and FD patients.

Conclusion
Serum IgG antibody titres to some common foods increased in IBS and FD patients compared to controls. But there is no significant correlation between symptom severity and elevated serum food antigen‐specific IgG antibodies in these patients.
Maternal allergy influences the proliferation of neonatal T cells expressing CCR4, CXCR5 or CD103
Authors: Haddeland U. , Brandtzaeg P. , Nakstad B.
Published: May 2007

Background
Elevated proliferative response to allergen in cord blood mononuclear cells (CBMCs) is related to subsequent allergy development of the neonate and has been suggested as a screening marker for high allergy risk.

Objective
To characterize the proliferating cells in CBMCs from a neonatal group influenced by maternal allergy compared with a control group without known allergic heredity.

Methods
CBMCs were stimulated with bovine β‐lactoglobulin (β‐LG) and proliferation was analysed by radioactive thymidine incorporation and expressed both as the traditional stimulation index (SI) and SI corrected by eliminating non‐specific proliferation. After β‐LG combined with endotoxin stimulation, cellular expression of IL‐4 and IFN‐γ mRNA was determined by quantitative RT‐PCR and adhesion as well as chemokine receptors were analysed by three‐colour flow cytometry in proliferating T cells (CD3+Ki‐67+).

Results
The percentage of CCR4+ cells correlated weakly with concurrent IL‐4 expression (rS=0.5, P<0.05), while CXCR3 correlated strongly with IFN‐γ expression (rS=0.83, P<0.001). In the allergy risk group, the percentage of proliferating T cells expressing CCR4 or integrin αE (CD103) was significantly reduced compared with the control group, while CXCR5 and the corrected SI were relatively increased (CCR4: P=0.01; integrin αE: P=0.03; CXCR5: P=0.04; SI: P=0.04).

Conclusion
Our results implied delayed maturation of immune functions involved in cellular migration, cell–cell interaction and immunoregulatory functions in neonates with hereditary allergy risk. The alterations observed in this subject group suggested that the corrected SI as well as proliferation of CCR4+, CXCR5+ or CD103+ T cells in allergen‐stimulated CBMCs might serve as early screening markers for allergy risk.