bluuebunny/crossref_metadata_2025 huggingface dataset:
Allergen‐induced interleukin‐9 production in vitro: correlation with atopy in human adults and comparison with interleukin‐5 and interleukin‐13
Authors: Devos S. , Cormont F. , Vrtala S. , Hooghe‐Peters E. , Pirson F. , Van Snick J.
Published: January 2006

Background
The contribution of IL‐9 to human atopy is supported by genetic studies. However, IL‐9 production in response to allergen in vitro has been reported only in children.

Objective
Study IL‐9 induction by allergen in adults, compare it with IL‐5 and IL‐13 and evaluate its association with atopy.

Methods
Peripheral blood mononuclear cell (PBMC) from control adults and from atopic patients were cultured with various allergens or phytohaemagglutinin (PHA) and secreted IL‐5, IL‐9 and IL‐13 were measured by ELISA.

Results
IL‐9 was produced in response to Dermatophagoides pteronyssinus (Der p) by PBMC from Der p‐hypersensitive adults at levels equivalent to those induced by PHA but with slower kinetics. The induction of IL‐9 was allergen specific, reflecting donor RAST profile. In Der p‐triggered reactions of non‐atopic and atopic subjects, IL‐9 showed the highest selectivity for atopics, IL‐5 and IL‐13 being produced more frequently in non‐atopic donors. Significant correlations with specific IgE titres were found for IL‐9 with all allergens tested (Der p and two peptides of Bet v 1 birch allergen). For IL‐5 and IL‐13, they were in the same range for Der p but more variable for birch allergens. Patterns of cytokine production by individual patients in response to allergen reflected these differences: for Der p, IL‐5, IL‐9 and IL‐13 productions were strongly correlated but for birch IL‐5 differed from the latter two. The in vitro production of IL‐9 reflected clinical hypersensitivity profiles and was higher in individuals with asthma than in those with disease limited to rhinitis and/or conjunctivitis.

Conclusions
Allergen‐triggered IL‐9 production in vitro is an excellent marker for atopy in adults given its virtual absence in allergen‐stimulated PBMC from non‐atopic individuals and its correlation with allergen‐specific IgE and asthma.
The diagnosis of Brazil nut allergy using history, skin prick tests, serum‐specific immunoglobulin E and food challenges
Authors: Ridout S. , Matthews S. , Gant C. , Twiselton R. , Dean T. , Arshad S. H.
Published: January 2006

Background
Allergy to Brazil nut is a relatively common nut allergy and can be fatal. However, the evidence is lacking regarding the best approach to its diagnosis.

Objective
We sought to determine the relative merits of history, skin prick testing, measurement of serum‐specific IgE and challenge in the diagnosis of Brazil nut allergy.

Methods
Fifty‐six children and adults with a history of an allergic reaction to Brazil nut or evidence of sensitization were investigated by questionnaire (n=56), skin prick tests (SPTs) (n=53), measurement of serum‐specific IgE to Brazil nut (n=54) and double‐blind, placebo‐controlled labial, and if necessary oral, challenges (n=19).

Results
Brazil nut allergy occurred in highly atopic individuals of any age with a strong family history of atopy. In 24 of 56 (43%), the history of an immediate reaction was sufficient to make a diagnosis with confidence and an oral challenge was considered unsafe. Of the 19 subjects undertaking the ‘gold standard’ test of a double‐blind, placebo‐controlled, food challenge, all six subjects with a SPT of at least 6 mm had a positive challenge and all three subjects with a SPT of 0 mm had a negative challenge. In the remaining 10 (53%) subjects, where SPT was between 1 and 5 mm and serum‐specific IgE was less than 3.5 kU/L, an oral challenge was performed resulting in three positive and seven negative challenges.

Conclusion
A combination of history, SPT and serum‐specific IgE was adequate in achieving a diagnosis in the majority (77%) patients with suspected Brazil nut allergy. However, a doubtful history with SPT between 1 and 5 mm, or a serum‐specific IgE less than 3.5 kU/L may require an oral challenge to help determine the risk of a Brazil nut allergic reaction.
Heart specific up‐regulation of genes for B‐type and C‐type natriuretic peptide receptors in diabetic mice
Authors: Christoffersen C. , Bartels E. D. , Nielsen L. B.
Published: January 2006

Background  Diabetes may cause cardiomyopathy characterized by cardiac fibrosis. Recent studies of genetically modified mice have elucidated a role of the natriuretic peptides (NP), type‐A and type‐B (ANP and BNP), and their common receptor [natriuretic peptide receptor (NPR), type‐A] in development of cardiac fibrosis. The role of NP type‐C (CNP) and NPR type‐B (NPR‐B) in the heart is less well established. In this study we examined if diabetes alters heart expression of the genes encoding the NP and its receptors.

Materials and methods  Cardiac mRNA was quantified by real‐time PCR in diabetic streptozotocin (STZ)‐treated and ob/ob‐mice and nondiabetic control mice.

Results  The ob/ob‐mice with type‐II diabetes displayed highly significant increases of the cardiac mRNA expression of NPR‐B and NPR‐C while the expression levels of NPR‐A, ANP, BNP, and CNP mRNA were similar in ob/ob‐mice and controls. Mice with STZ‐induced type‐I diabetes also showed an increase of heart NPR‐B mRNA expression at 12 weeks, but not at 3, 6 or 9 weeks after STZ‐treatment. The ANP and NPR‐C mRNA expressions were only altered after 3 weeks, whereas BNP, CNP and NPR‐A mRNA expressions were not altered in STZ‐treated‐mouse hearts at any of the time points.

Conclusions  The results show that diabetes in mice confers increased NPR‐B gene expression in the heart, suggesting that increased NPR‐B signalling may affect development of diabetic cardiomyopathy.
Green tea polyphenol (–)‐epigallocatechin‐3‐gallate inhibits ethanol‐induced activation of pancreatic stellate cells
Authors: Asaumi H. , Watanabe S. , Taguchi M. , Tashiro M. , Nagashio Y. , Nomiyama Y. , Nakamura H. , Otsuki M.
Published: January 2006

Background  Activated pancreatic stellate cells (PSCs) play a central role in the pathogenesis of pancreatic fibrogenesis and inflammation. Ethanol, a major cause of chronic pancreatitis, directly induces PSC activation and oxidative stress. Inhibition of PSC activation or stimulation to PSC might be an effective therapeutic strategy for the prevention of pancreatic fibrosis, and (–)‐epigallocatechin‐3‐gallate (EGCG), a major component of green tea extracts, is a potent antioxidant of polyphenols. Therefore, we examined the mechanisms through which ethanol induces oxidative stress on PSCs and evaluated the effect of EGCG on activation and cell functions of ethanol‐stimulated PSCs.

Materials and methods  The PSCs were isolated from the pancreas of male Wister rats with Nycodenz gradient methods and cells between passages one and four were used. Isolated PSCs were cultured with ethanol (50 mM) in the absence or presence of EGCG (5 µM or 25 µM).

Results  The EGCG pre‐treatment abolished ethanol‐induced lipid peroxidation of the cell membrane, loss of total superoxide dismutase (SOD) activity and suppressed ethanol‐induced gene expressions of Mn‐ and Cu/Zn‐SOD. EGCG also suppressed ethanol‐induced p38 mitogen‐activated protein (MAP) kinase phosphorylation, α‐smooth muscle actin production in PSCs and activated transforming growth factor‐β1 secretion into the medium. Furthermore, EGCG inhibited ethanol‐induced type‐I procollagen production and collagen secretion. In addition, EGCG inhibited transformation of freshly isolated cells to activated myofibroblast‐like phenotype.

Conclusions  Our results suggest that green tea and polyphenols could prevent pancreatic fibrosis by inhibiting PSC activation through the antioxidative effect.
Hypoxic inhibition of human myometrial contractions in vitro: implications for the regulation of parturition
Authors: Bugg G. J. , Riley M. J. , Johnston T. A. , Baker P. N. , Taggart M. J.
Published: January 2006

Background  Insufficient tissue oxygenation is a likely contribution to weak, inco‐ordinate human uterine contractile activity characteristic of prolonged, dysfunctional labour. However, the direct effects of hypoxia on human myometrial contractility has, surprisingly, not yet been detailed. Therefore, we report the influence of hypoxia on spontaneous and agonist‐induced carbachol, prostaglandin (PGF2α), and oxytocin contractions of myometria from nonpregnant and pregnant women.

Materials and methods  Uterine biopsies were obtained from pregnant women at term undergoing elective Caesarean section and nonpregnant women undergoing hysterectomy. Myometrial strips were equilibrated at 37 °C in normoxic physiological salt solution (95% air/5% CO2) and the influence of hypoxia (95% N2/5% CO2) on contractility was investigated.

Results  Hypoxia resulted in a significant reduction in spontaneous contractile function; nonpregnant tissue was less resistant to the deleterious effects of hypoxia. Agonist‐induced contractions, while being more resistant to hypoxia than spontaneous contractions, were also significantly inhibited. In myometria of pregnant women the PGF2α‐ or oxytocin‐induced contractility was more resistant to hypoxia than carbachol. Finally, the inhibitory actions of hypoxia were exacerbated with repeated oxytocin administration with a more severe effect on contractile integral than on initial phasic contraction amplitude.

Conclusions  We detail, for the first time, the effects of hypoxia on contractility of human myometria from nonpregnant and pregnant women. Physiologically important uterotonic agents are more resistant to the effects of hypoxia than spontaneous contractions although repeated stimulation with oxytocin during hypoxia results in progressively less force. The results indicate that if significant hypoxia occurs in vivo then it is a likely contributory factor to the pathways underlying prolonged dysfunctional labour.
Influence of a subsequent meal on the oro‐cecal transit time of a solid test meal
Authors: Priebe M. G. , Wachters‐Hagedoorn R. E. , Landman K. , Heimweg J. , Elzinga H. , Vonk R. J.
Published: January 2006

Background  Oro‐cecal transit time (OCTT) is determined for clinical diagnostics of intestinal complaints and research purposes. Ingestion of a subsequent meal during the test period shortens the OCTT of a liquid test meal (glucose solution), as previously reported. This study was conducted to determine whether the same phenomenon occurs after ingestion of a solid test meal.

Materials and methods  The OCTT of a pancake was measured with the lactose‐[13C]‐ureide breath test on two occasions in 28 volunteers. All the volunteers took the same subsequent meal once at 4 h and at 6 h after ingestion of the pancake.

Results  In 16 of the 56 tests no increase in breath‐13CO2 was observed. No statistically significant difference was found between the OCTTs of the test meal after ingestion of the subsequent meal at 4 h or 6 h (367; 311–405 min and 290; 370–405 min, median quartiles, respectively) (P = 0·14, n = 18). Only a subgroup (n = 4) with a short OCTT in the test with the 4‐h subsequent meal (278; 259–296 min) tended to have a longer OCTT in the test with the 6‐h subsequent meal (390; 379–401 min; P = 0·059).

Conclusion  The effect of the ingestion of a subsequent meal on the transit time of a test meal is shown to be dependent on the physical form and/or caloric content of the test meal.
Effects of antibodies obtained from patients with dilated cardiomyopathy on the function of isolated rat hearts
Authors: Staudt Y. , Trimpert C. , Birkenmeier K. , Krieg T. , Bemmann T. , Beug D. , Felix S. B. , Staudt A.
Published: January 2006

Background  Cardiac autoantibodies may play a pathophysiological role in cardiac dysfunction of patients suffering from dilated cardiomyopathy (DCM). Immunoadsorption (IA), which removes antibodies from patients’ plasma, may consequently improve cardiac function in DCM. The functional effects of DCM antibodies are only partly understood.

Materials and methods  DCM patients (n = 10) were treated with IA by application of antibody columns directed against human immunoglobulin (Ig). IA was also performed with plasma taken from 10 healthy donors (controls). The antibodies eliminated and purified by IA were collected and dialysed. Rat hearts were isolated and perfused retrogradely via the aorta in Langendorff mode. During constant‐pressure and constant‐volume perfusion of the hearts, the influence of diluted antibodies on contractility, relaxation, and on coronary perfusion was analysed.

Results  Antibodies obtained from controls had no effect on contractility and relaxation of isolated perfused hearts during constant‐pressure and constant‐volume perfusion. In contrast, during constant‐pressure perfusion, collected DCM antibodies caused immediate and dose‐related reduction of contractility (dLVP/dtmax: dilution −1 : 32 = −7·1 ± 1·1%; dilution −1 : 2 = −20·1 ± 2·1%; P < 0·001) and diastolic relaxation (dLVP/dtmin: dilution −1 : 32 = −11·1 ± 1·5%; dilution −1 : 2 = −23·9 ± 2·2%; P < 0·001). The heart rate did not change significantly in either group. The effects of DCM antibodies on contractility and relaxation remained detectable during constant‐volume perfusion. The observed reduction of contractility and diastolic relaxation was accompanied by impairment of coronary perfusion.

Conclusion  In the rat heart, antibodies obtained from DCM patients may impair contractility and relaxation, and thereby probably also coronary perfusion.
Lipoprotein(a), type 2 diabetes and vascular risk in coronary patients
Authors: Saely C. H. , Koch L. , Schmid F. , Marte T. , Aczel S. , Langer P. , Hoefle G. , Drexel H.
Published: January 2006

Background  Lipoprotein(a) [Lp(a)] is an important cardiovascular risk factor in the general population. However, prospective data on the vascular risk conferred by Lp(a) in patients with diabetes mellitus are scarce and controversial. It is not known whether the diabetic state affects the association of Lp(a) with vascular events among coronary patients.

Design  We measured Lp(a) in 587 consecutive patients undergoing coronary angiography for the evaluation of coronary artery disease. The incidence of vascular events was recorded over 4 years.

Results  At baseline, Lp(a) was significantly lower in patients with type 2 diabetes (T2DM) (n = 136) than in nondiabetic individuals (11 (0·8–30) mg dL−1 vs. 16 (0·8–51) mg dL−1; P = 0·025). Prospectively, Lp(a) was a strong and independent predictor of vascular events in nondiabetic patients (standardized adjusted hazard ratio (HR) = 1·461 (1·121–1·904); P = 0·005), but not in patients with T2DM [HR = 0·812 (0·539–1·223); P = 0·320]. An interaction term diabetes × Lp(a) was significant (P = 0·008), indicating that Lp(a) was a significantly stronger predictor of vascular events in nondiabetic patients than in patients with T2DM.

Conclusions  Lp(a) in diabetic coronary patients is low and not associated with the incidence of vascular events. Although measurement of Lp(a) provides useful information in nondiabetic coronary patients, it is of little value in coronary patients with T2DM.
Protein kinase A‐dependent increase in WAVE2 expression induced by the focal adhesion protein vinexin
Authors: Mitsushima Masaru , Sezaki Takuhito , Akahane Rie , Ueda Kazumitsu , Suetsugu Shiro , Takenawa Tadaomi , Kioka Noriyuki
Published: January 2006

The focal adhesion protein vinexin is a member of a family of adaptor proteins that are thought to participate in the regulation of cell adhesion, cytoskeletal reorganization, and growth factor signaling. Here, we show that vinexin β increases the amount of and reduces the mobility on SDS‐PAGE of Wiskott‐Aldrich syndrome protein family verprolin‐homologous protein (WAVE) 2 protein, which is a key factor modulating actin polymerization in migrating cells. This mobility retardation disappeared after in vitro phosphatase treatment. Co‐immunoprecipitation assays revealed the interaction of vinexin β with WAVE2 as well as WAVE1 and N‐WASP. Vinexin β interacts with the proline‐rich region of WAVE2 through the first and second SH3 domains of vinexin β. Mutations disrupting the interaction impaired the ability of vinexin β to increase the amount of WAVE2 protein. Treatments with proteasome inhibitors increased the amount of WAVE2, but did not have an additive effect with vinexin β. Inhibition of protein kinase A (PKA) activity suppressed the vinexin‐induced increase in WAVE2 protein, while activation of PKA increased WAVE2 expression without vinexin β. These results suggest that vinexin β regulates the proteasome‐dependent degradation of WAVE2 in a PKA‐dependent manner.
Structural relationship of curcumin derivatives binding to the BRCT domain of human DNA polymerase λ
Authors: Takeuchi Toshifumi , Ishidoh Tomomi , Iijima Hiroshi , Kuriyama Isoko , Shimazaki Noriko , Koiwai Osamu , Kuramochi Kouji , Kobayashi Susumu , Sugawara Fumio , Sakaguchi Kengo , Yoshida Hiromi , Mizushina Yoshiyuki
Published: January 2006

We previously reported that phenolic compounds, petasiphenol and curcumin (diferuloylmethane), were a selective inhibitor of DNA polymerase λ (pol λ) in vitro. The purpose of this study was to investigate the molecular structural relationship of curcumin and 13 chemically synthesized derivatives of curcumin. The inhibitory effect on pol λ (full‐length, i.e. intact pol λ including the BRCA1 C‐ terminal [BRCT] domain) by some derivatives was stronger than that by curcumin, and monoacetylcurcumin (compound 13) was the strongest pol λ inhibitor of all the compounds tested, achieving 50% inhibition at a concentration of 3.9 µm. The compound did not influence the activities of replicative pols such as α, δ, and ɛ. It had no effect on pol β activity either, although the three‐dimensional structure of pol β is thought to be highly similar to that of pol λ. Compound 13 did not inhibit the activity of the C‐terminal catalytic domain of pol λ including the pol β‐like core, in which the BRCT motif was deleted from its N‐terminal region. MALDI‐TOF MS analysis demonstrated that compound 13 bound selectively to the N‐terminal domain of pol λ, but did not bind to the C‐terminal region. Based on these results, the pol λ‐inhibitory mechanism of compound 13 is discussed.
Epstein–Barr virus infection negatively impacts the CXCR4‐dependent migration of tonsillar B cells
Authors: Ehlin‐Henriksson Barbro , Mowafi Frida , Klein George , Nilsson Anna
Published: January 2006

The primary Epstein–Barr virus (EBV) infection occurs in the oropharynx, where the virus infects B cells and subsequently establishes latency in the memory B‐cell compartment. EBV has previously been shown to induce changes in the cell surface expression of several chemokine receptors in cell lines and the transfection of EBNA2 or LMP1 into a B‐cell‐lymphoma‐derived cell line decreased the expression of CXCR4. We show that in vitro EBV infection reduces the expression of CXCR4 on primary tonsil B cells already 43 hr after infection. Furthermore, EBV infection affects the chemotactic response to stromal cell‐derived factor (SDF‐1)α/CXCL12, the ligand for CXCR4, with a reduction of SDF‐1α‐induced migration. To clarify whether this reduced migration is EBV‐specific or a consequence of cell activation, tonsillar B cells were either infected with EBV, activated with anti‐CD40 and interleukin‐4 (IL‐4) or kept in medium. Activation by anti‐CD40 and IL‐4 decreased the CXCR4 expression but the CD40 + IL‐4‐stimulated cells showed no reduction of chemotactic efficacy. Our finding suggests that changing the SDF‐1α response of the EBV‐infected B cells may serve the viral strategy by directing the infected cells into the extrafollicular areas, rather than retaining them in the lymphoepithelium.
Blockade of OX40‐ligand after initial triggering of the T helper 2 response inhibits mercuric chloride‐induced autoimmunity
Authors: MacPhee Iain A. M. , Yagita Hideo , Oliveira David B. G.
Published: January 2006

Mercuric chloride (HgCl2)‐induced autoimmunity in Brown Norway rats is a spontaneously resolving autoimmune response driven by the activation of T helper type 2 lymphocytes (Th2 cells). Treatment with antibody to OX40‐ligand (OX40‐L) from the time of the first HgCl2 injection for 12 days had little effect. Delayed treatment commenced 8 days after the first HgCl2 injection significantly suppressed immunoglobulin E production, splenomegaly, weight loss and mortality. This makes OX40/OX40‐L signalling an attractive therapeutic target for Th2‐driven autoimmune diseases. Intravenous administration of the murine antibody to OX‐40‐L elicited a vigorous anti‐mouse immunoglobulin antibody response that was significantly enhanced compared to the response to control immunoglobulin. It is likely that this response significantly reduced the plasma half‐life of the anti‐OX40‐L antibody and this observation has clear implications for the interpretation of data from experiments where anti‐OX40‐L is used in vivo.
CelTOS, a novel malarial protein that mediates transmission to mosquito and vertebrate hosts
Authors: Kariu Tohru , Ishino Tomoko , Yano Kazuhiko , Chinzei Yasuo , Yuda Masao
Published: January 2006

The malarial parasite has two hosts in its life cycle, a vertebrate and a mosquito. We report here that malarial invasion into these hosts is mediated by a protein, designated cell‐traversal protein for ookinetes and sporozoites (CelTOS), which is localized to micronemes that are organelles for parasite invasive motility. Targeted disruption of the CelTOS gene in Plasmodium berghei reduced parasite infectivity in the mosquito host approximately 200‐fold. The disruption also reduced the sporozoite infectivity in the liver and almost abolished its cell‐passage ability. Liver infectivity was restored in Kupffer cell‐depleted rats, indicating that CelTOS is necessary for sporozoite passage from the circulatory system to hepatocytes through the liver sinusoidal cell layer. Electron microscopic analysis revealed that celtos‐disrupted ookinetes invade the midgut epithelial cell by rupturing the cell membrane, but then fail to cross the cell, indicating that CelTOS is necessary for migration through the cytoplasm. These results suggest that conserved cell‐passage mechanisms are used by both sporozoites and ookinetes to breach host cellular barriers. Elucidation of these mechanisms might lead to novel antimalarial strategies to block parasite's transmission.
The ISWI and CHD1 chromatin remodelling activities influence ADH2 expression and chromatin organization
Authors: Xella Barbara , Goding Colin , Agricola Eleonora , Di Mauro Ernesto , Caserta Micaela
Published: January 2006

Nucleosome remodelling complexes play a key role in gene activation in response to environmental changes by driving promoter chromatin to reach an accessible configuration. They also mediate genome‐wide chromatin organization, although their role in processes other than activation‐related chromatin remodelling are poorly understood. The Saccharomyces cerevisiae ADH2 gene represents an excellent model for understanding the role of chromatin structure and remodelling in gene regulation. Following glucose depletion, highly positioned promoter nucleosomes are destabilized leading to strictly regulated kinetics of transcriptional activation. Nevertheless, no chromatin remodelling activities responsible for establishing or remodelling ADH2 chromatin structure have been identified to date. Here we show that the absence of the Isw1 and Chd1 ATP‐dependent chromatin remodelling activities delays the maximal expression of ADH2 without impairing the chromatin remodelling that occurs upon activation. Instead, a destabilized chromatin structure on the ADH2 coding and termination region is observed in the absence of Isw1 or Chd1 in repressing conditions. The specific Isw1 complex involved in this nucleosome repositioning is Isw1b because the deletion of Ioc2 and Ioc4, but not of Ioc3, causes the same phenotype as the deletion of Isw1. Moreover, the lack of Chd1 combined with the absence of Isw1 and Isw2 impairs nucleosome spacing along the ADH2 gene, and genome‐wide in S. cerevisiae. Thus, the ISWI and Chd1 remodelling factors are not only involved in transcription‐related chromatin remodelling, but also are required to maintain a specific chromatin configuration across the yeast genome.
Mycobacterium tuberculosis prcBA genes encode a gated proteasome with broad oligopeptide specificity
Authors: Lin Gang , Hu Guiqing , Tsu Christopher , Kunes Yune Z. , Li Huilin , Dick Lawrence , Parsons Thomas , Li Ping , Chen Zhiqiang , Zwickl Peter , Weich Nadine , Nathan Carl
Published: January 2006

Genes predicted to be associated with the putative proteasome of Mycobacterium tuberculosis (Mtb) play a critical role in defence of the bacillus against nitrosative stress. However, proteasomes are uncommon in eubacteria and it remains to be established whether Mtb's prcBA genes in fact encode a proteasome. We found that coexpression of recombinant PrcB and PrcA in Escherichia coli over a prolonged period at 37°C allowed formation of an α7β7β7α7, 750 kDa cylindrical stack of four rings in which all 14 β‐subunits were proteolytically processed to expose the active site threonine. In contrast to another Actinomycete, Rhodococcus erythropolis, Mtb's β‐chain propeptide was not required for particle assembly. Peptidolytic activity of the 750 kDa particle towards a hydrophobic oligopeptide was nearly two orders of magnitude less than that of the Rhodococcus 20S proteasome, and unlike eukaryotic and archaeal proteasomes, activity of the Mtb 750 kDa particle could not be stimulated by SDS, Mg2+ or Ca2+. Electron microscopy revealed what appeared to be obstructed α‐rings in the Mtb 750 kDa particle. Deletion of the N‐terminal octapeptide from Mtb's α‐chain led to disappearance of the apparent obstruction and a marked increase of peptidolytic activity. Unlike proteasomes isolated from other Actinomycetes, the open‐gate Mtb mutant 750 kDa particle cleaved oligopeptides not only after hydrophobic residues but also after basic, acidic and small, neutral amino acids. Thus, Mtb encodes a broadly active, gated proteasome that may work in concert with an endogenous activator.
CRZ1, a target of the calcineurin pathway in Candida albicans
Authors: Karababa Mahir , Valentino Emilio , Pardini Giacomo , Coste Alix T. , Bille Jacques , Sanglard Dominique
Published: January 2006

Calcineurin is a major player in calcium‐dependent signal transduction pathways of eukaryotes. Calcineurin acts on transcription factors (e.g. CRZ1 in Saccharomyces cerevisiae) and governs the expression of genes in a species‐dependent fashion. In Candida albicans, the calcineurin pathway is involved in tolerance to antifungal agents, cation homeostasis and virulence. However, the components of the calcineurin pathway are still poorly investigated in this yeast species. Taking S. cerevisiae as a model to reconstitute this pathway, two CRZ1‐like genes, CRZ1 and CRZ2 (for calcineurin‐responsive zinc finger 1 and 2 genes), were found with C2H2 zinc finger domains. Only CRZ1 was able to restore the calcium hypersusceptibility of a S. cerevisiae crz1Δ mutant and to mediate calcium‐dependent gene expression in this yeast species. Several experiments showed that CRZ1 was dependent on calcineurin in C. albicans: (i) phenotypic analysis of a crz1Δ/Δ mutant showed impaired growth as compared with the wild type in the presence of cations (Ca2+, Mn2+) as does a mutant lacking calcineurin subunit A (cnaΔ/Δ) and (ii) a green fluorescent protein (GFP)–Crz1p fusion protein showed a calcium‐ and calcineurin‐dependent nuclear localization. To further analyse the relationship between calcineurin and CRZ1, a comprehensive analysis of calcineurin/Crz1p‐dependent gene expression following addition of Ca2+ (200 mM) was performed. Among the expression of 264 genes altered by at least twofold, the upregulation of 60 genes was dependent on both calcineurin and CRZ1. Interestingly, a motif [5′‐G(C/T)GGT‐3′] with similarity to the target sequence of Crz1p (GNGGCG/TCA) from S. cerevisiae was identified as a putative regulatory sequence in the upstream regions of these calcineurin/Crz1p‐dependent genes. However, additional experiments showed that calcineurin may have other targets in addition to CRZ1. First, CRZ1 was not involved in tolerance to antifungal agents (fluconazole, terbinafine) on the opposite to calcineurin. Second, CRZ1 was only moderately influencing virulence in a mice model of infection which is in sharp contrast to the strong avirulence of cnaΔ/Δ mutant in the same animal model. Even though this work establishes CRZ1 as a calcineurin target, further studies are needed to identify other calcineurin‐dependent elements in C. albicans.
Inactivation of the fibronectin‐binding adhesin gene bbk32 significantly attenuates the infectivity potential of Borrelia burgdorferi
Authors: Seshu J. , Esteve‐Gassent M. Dolores , Labandeira‐Rey Maria , Kim Jung Hwa , Trzeciakowski Jerome P. , Höök Magnus , Skare Jonathan T.
Published: January 2006

Borrelia burgdorferi, the aetiological agent of Lyme disease, utilizes multiple adhesins to interact with both the arthropod vector and mammalian hosts it colonizes. One such adhesive molecule is a surface‐exposed fibronectin‐binding lipoprotein, designated BBK32. Previous characterization of BBK32‐mediated fibronectin binding has been limited to biochemical analyses due to the difficulty in mutagenizing infectious isolates of B. burgdorferi. Here we report an alternative method to inactivate bbk32 via allelic exchange through use of a low‐passage variant of B. burgdorferi strain B31 that is more readily transformed. The resulting mutant does not synthesize BBK32, exhibits reduced fibronectin binding in solid phase assays and manifests decreased interactions with mouse fibroblast cells relative to both the infectious parent and genetic complement. Furthermore, the bbk32 knockout was significantly attenuated in the murine model of Lyme disease, whereas a genetically complemented control was not, indicating that BBK32 is necessary for maximal B. burgdorferi infection in the mouse. To our knowledge this is the first mutational analysis of a surface exposed, functional borrelial lipoprotein adhesin whose activity is associated with the mammalian host environment. By analogy with other pathogens that utilize fibronectin binding as an important virulence determinant, the borrelial fibronectin–BBK32 interaction is likely to be important in B. burgdorferi‐specific pathogenic mechanisms, particularly in the context of dissemination, secondary colonization and/or persistence.
Ammonium permease‐based sensing mechanism for rapid ammonium activation of the protein kinase A pathway in yeast
Authors: Van Nuland An , Vandormael Patrick , Donaton Monica , Alenquer Marta , Lourenço Artur , Quintino Edgar , Versele Matthias , Thevelein Johan M.
Published: January 2006

In the yeast Saccharomyces cerevisiae starvation for nitrogen on a glucose‐containing medium causes entrance into G0 and downregulation of all targets of the PKA pathway. Re‐addition of a nitrogen source in the presence of glucose causes rapid activation of trehalase and other PKA targets. Trehalase activation upon ammonium re‐supplementation is dependent on PKA activity, but not on its regulatory subunit nor is it associated with an increase in cAMP. In nitrogen‐starved cells, ammonium transport and activation of trehalase are most active in strains expressing either the Mep2 or Mep1 ammonium permease, as opposed to Mep3. The non‐metabolizable ammonium analogue, methylamine, also triggers activation of trehalase when transported by Mep2 but not when taken up by diffusion. Inhibition of ammonium incorporation into metabolism did not prevent signalling. Extensive site‐directed mutagenesis of Mep2 showed that transport and signalling were generally affected in a similar way, although they could be separated partially by specific mutations. Our results suggest an ammonium permease‐based sensing mechanism for rapid activation of the PKA pathway. Mutagenesis of Asn246 to Ala in Mep2 abolished transport and signalling with methylamine but had no effect with ammonium. The plant AtAmt1;1, AtAmt1;2, AtAmt1;3 and AtAmt2 ammonium transporters sustained transport and trehalase activation to different extents. Specific mutations in Mep2 affected the activation of trehalase differently from induction of pseudohyphal differentiation. We also show that Mep permease involvement in PKA control is different from their role in haploid invasive growth, in which Mep1 sustains and Mep2 inhibits, in a way independent of the ammonium level in the medium.
GTP drives myosin light chain 1 interaction with the class V myosin Myo2 IQ motifs via a Sec2 RabGEF‐mediated pathway
Authors: Bielli Pamela , Casavola Elena Caroli , Biroccio Antonino , Urbani Andrea , Ragnini‐Wilson Antonella
Published: January 2006

The yeast myosin light chain 1 (Mlc1p) belongs to a branch of the calmodulin superfamily and is essential for vesicle delivery at the mother‐bud neck during cytokinesis due to is ability to bind to the IQ motifs of the class V myosin Myo2p. While calcium binding to calmodulin promotes binding/release from the MyoV IQ motifs, Mlc1p is unable to bind calcium and the mechanism of its interaction with target motifs has not been clarified. The presence of Mlc1p in a complex with the Rab/Ypt Sec4p and with Myo2p suggests a role for Mlc1p in regulating Myo2p cargo binding/release by responding to the activation of Rab/Ypt proteins. Here we show that GTP or GTPγS potently stimulate Mlc1p interaction with Myo2p IQ motifs. The C‐terminus of the Rab/Ypt GEF Sec2p, but not Sec4p activation, is essential for this interaction. Interestingly, overexpression of constitutively activated Ypt32p, a Rab/Ypt protein that acts upstream of Sec4p, stimulates Mlc1p/Myo2p interaction similarly to GTP although a block of Ypt32 GTP binding does not completely abolish the GTP‐mediated Mlc1p/Myo2p interaction. We propose that Mlc1p/Myo2p interaction is stimulated by a signal that requires Sec2p and activation of Ypt32p.
A new protein superfamily includes two novel 3‐methyladenine DNA glycosylases from Bacillus cereus, AlkC and AlkD
Authors: Alseth Ingrun , Rognes Torbjørn , Lindbäck Toril , Solberg Inger , Robertsen Kristin , Kristiansen Knut Ivan , Mainieri Davide , Lillehagen Lucy , Kolstø Anne‐Brit , Bjørås Magnar
Published: January 2006

Soil bacteria are heavily exposed to environmental methylating agents such as methylchloride and may have special requirements for repair of alkylation damage on DNA. We have used functional complementation of an Escherichia coli tag alkA mutant to screen for 3‐methyladenine DNA glycosylase genes in genomic libraries of the soil bacterium Bacillus cereus. Three genes were recovered: alkC, alkD and alkE. The amino acid sequence of AlkE is homologous to the E. coli AlkA sequence. AlkC and AlkD represent novel proteins without sequence similarity to any protein of known function. However, iterative and indirect sequence similarity searches revealed that AlkC and AlkD are distant homologues of each other within a new protein superfamily that is ubiquitous in the prokaryotic kingdom. Homologues of AlkC and AlkD were also identified in the amoebas Entamoeba histolytica and Dictyostelium discoideum, but no other eukaryotic counterparts of the superfamily were found. The alkC and alkD genes were expressed in E. coli and the proteins were purified to homogeneity. Both proteins were found to be specific for removal of N‐alkylated bases, and showed no activity on oxidized or deaminated base lesions in DNA. B. cereus AlkC and AlkD thus define novel families of alkylbase DNA glycosylases within a new protein superfamily.
Replication‐associated gene dosage effects shape the genomes of fast‐growing bacteria but only for transcription and translation genes
Authors: Couturier Etienne , Rocha Eduardo P. C.
Published: January 2006

The bidirectional replication of bacterial genomes leads to transient gene dosage effects. Here, we show that such effects shape the chromosome organisation of fast‐growing bacteria and that they correlate strongly with maximal growth rate. Surprisingly the predicted maximal number of replication rounds shows little if any phylogenetic inertia, suggesting that it is a very labile trait. Yet, a combination of theoretical and statistical analyses predicts that dozens of replication forks may be simultaneously present in the cells of certain species. This suggests a strikingly efficient management of the replication apparatus, of replication fork arrests and of chromosome segregation in such cells. Gene dosage effects strongly constrain the position of genes involved in translation and transcription, but not other highly expressed genes. The relative proximity of the former genes to the origin of replication follows the regulatory dependencies observed under exponential growth, as the bias is stronger for RNA polymerase, then rDNA, then ribosomal proteins and tDNA. Within tDNAs we find that only the positions of the previously proposed ‘ubiquitous’ tRNA, which translate the most frequent codons in highly expressed genes, show strong signs of selection for gene dosage effects. Finally, we provide evidence for selection acting upon genome organisation to take advantage of gene dosage effects by identifying a positive correlation between genome stability and the number of simultaneous replication rounds. We also show that gene dosage effects can explain the over‐representation of highly expressed genes in the largest replichore of genomes containing more than one chromosome. Together, these results demonstrate that replication‐associated gene dosage is an important determinant of chromosome organisation and dynamics, especially among fast‐growing bacteria.
A C‐terminal translocation signal is necessary, but not sufficient for type IV secretion of the Helicobacter pylori CagA protein
Authors: Hohlfeld Sabine , Pattis Isabelle , Püls Jürgen , Plano Gregory V. , Haas Rainer , Fischer Wolfgang
Published: January 2006

Type IV secretion systems are increasingly recognized as important virulence determinants of Gram‐negative bacterial pathogens. While the examination of several type IV‐secreted proteins suggested that their secretion depends on C‐terminal signals, the nature of these signals and their conservation among different systems remain unclear. Here, we have characterized the secretion signal of the Helicobacter pylori CagA protein, which is translocated by the Cag type IV secretion apparatus into eucaryotic cells. The production of fusion proteins of CagA and green fluorescent protein (GFP) did not result in translocation of GFP to epithelial cells, but a fusion of GFP with the CagA C‐terminus exerted a dominant‐negative effect upon wild‐type CagA translocation. We show that CagA translocation depends on the presence of its 20 C‐terminal amino acids, containing an array of positively charged residues. Interestingly, these positive charges are neither necessary nor sufficient for CagA translocation, but replacing the C‐terminal region of CagA with that of other type IV‐secreted proteins reconstitutes CagA translocation competence. Using a novel type IV translocation assay with a phosphorylatable peptide tag, we show that removal of the N‐terminal part of the CagA protein renders the protein translocation‐incompetent as well. Thus, the Cag type IV secretion system seems to diverge from other systems not only with respect to its composition and architecture, but also in terms of substrate recognition and transport.
Oropharyngeal dysphagia after the acute phase of stroke: predictors of aspiration
Authors: Terré R. , Mearin F.
Published: January 2006

Abstract  Oropharyngeal dysphagia is frequent during the acute phase of stroke, but most patients recover. Dysphagia is related to higher incidence of aspiration, pneumonia and death. Frequently neither clinical history nor neurological evaluation predicts the presence of aspiration. In 64 patients not recovered from severe stroke after the acute phase with clinically suspected oropharyngeal dysphagia we investigated: (i) the correlation between clinical manifestations and videofluoroscopic findings; (ii) predictive factors of aspiration and silent aspiration. Clinical examination showed that 44% had impaired gag reflex, 47% cough during oral feeding, and 13% changes in voice after swallowing. Videofluoroscopy revealed some abnormality in 87%: 53% in the oral phase and 84% in the pharyngeal phase (aspiration in 66%; half being silent). Impaired pharyngeal safety was more frequent in posterior territory lesions and patients with a history of pneumonia (P < 0.01). No correlation was found between clinical evaluation findings and presence of aspiration. Silent aspirations were more frequent in patients with previous orotracheal intubation (P < 0.05) and abnormalities in velopharyngeal reflexes (P < 0.05). We concluded that in patients not recovered from severe stroke after the acute phase and with suspected oropharyngeal dysphagia, clinical evaluation is of scant use in predicting aspiration and silent aspiration. Videofluoroscopic examination is mandatory in these patients.
Long‐lasting changes in small intestinal transport following the recovery fromTrichinella spiralisinfection
Authors: Venkova K. , Greenwood‐van Meerveld B.
Published: January 2006

Abstract Changes in intestinal motility and visceral sensitivity are found after resolution of acute enteric inflammation. The study investigates whether a transient nematode‐induced intestinal inflammation may result in long‐lasting remodelling of epithelial transport. Ferrets infected with Trichinella spiralis or sham‐infected animals were euthanized on day 10, 30 or 60 postinfection (PI) and the jejunum was isolated. The net transport of electrolytes was measured electrophysiologically as transmucosal short‐circuit current (Isc) and responses to electrical field stimulation (EFS: 1–32 Hz) or secretagogues were investigated. Myeloperoxidase (MPO) activity, a marker of mucosal inflammation, was maximal during the enteric stage of T. spiralis infection (day 10 PI) and returned to normal on days 30 and 60 PI. Mucosal inflammation caused a reduction in basal Isc, increased electrical conductance (G) and decreased the maximal responses to EFS, carbachol or histamine. On days 30 and 60 PI the inflammation resolved and basal electrogenic transport appeared normal; however, the secretion induced by EFS, carbachol or histamine remained suppressed. Moreover, EFS‐induced responses were shifted from predominantly cholinergic in controls to non‐cholinergic in the infected animals. The results suggest that a transient small intestinal inflammation causes a long‐term remodelling of epithelial function.
Tinea capitis in eastern Nepal
Authors: Jha Bibeka Nand , Garg Vijay Kumar , Agrawal Sudha , Khanal Basudha , Agarwalla Arun
Published: January 2006

Background  Tinea capitis is an increasing public health concern throughout the world. The clinical types and etiological agents vary from time to time and place to place. This study was undertaken to identify the etiological agents and to determine the clinico‐etiological correlation of tinea capitis in eastern Nepal.

Methods  Sixty‐nine clinically diagnosed cases of tinea capitis were enrolled in this study. Hair roots and skin scrapings were collected from each patient and subjected to microscopy and culture for identification of fungal hyphae and spores.

Results  Tinea capitis accounted for 4.6% of all dermatophyte infections: 68.1% occurred in patients below the age of 11 years with a male to female ratio of 1 : 1.9. “Gray patch” was the most common clinical type (52.2%), followed by “black dot” (17.4%), seborrhoeic dermatitis (13%), alopecia areata (11.6%) and pustular (4.3%). Direct microscopy of hair was positive in 62.3% of patients. Culture positivity was found in 56.7% of patients. Common isolated organisms were Trichophyton violaceum (48.71%), T. mentagrophytes (15.38%), T. tonsurans (12.82%), Microsporum canis (7.69%), T. rubrum and M. gypseum (5.12% each), and M. audouinii and M. nanum (2.56% each).

Conclusions  Trichophyton violaceum was the most common pathogen of tinea capitis. The clinical manifestations were variable and “gray patch” was the most common clinical presentation in this part of the world.
A new gastrointestinal finding in Proteus syndrome: report of a case of multiple colonic hemangiomas
Authors: Farajzadeh Saideh , Zahedi Mohammad Javad , Moghaddam Sodaif Darvish
Published: January 2006

A patient with Proteus syndrome presented with lower gastrointestinal bleeding due to multiple colonic hemangiomas, a finding which has not been described previously in this syndrome. The patient was a 20‐year‐old man with features fulfilling the diagnostic criteria for Proteus syndrome. He fulfilled both general criteria (mosaic distribution of the lesions, progressive course and sporadic occurrence) and specific criteria (including epidermal nevus, disproportionate overgrowth of limbs and vascular malformations). Fiberoptic colonoscopy revealed multiple hemangiomas, 0.5–1 cm in diameter, on the left side of the colon. Some gastrointestinal complications have been reported in patients with Proteus syndrome, including rectal polyps, colonic lipomatosis, atrophy of the intestinal villi and intestinal affection with fatty wall thickening but, as far as we are aware, colonic hemangiomas have not previously been reported in this syndrome.
Epidemiology of pemphigus in south‐western Iran: A 10‐year retrospective study (1991–2000)
Authors: Salmanpour R. , Shahkar H. , Namazi M. R. , Rahman‐Shenas M. R.
Published: January 2006

Background  Pemphigus is a disease demonstrating an uneven geographic distribution. This is the first epidemiologic study of pemphigus in Iran.

Methods  Data were collected from the files of all newly registered patients with pemphigus at the sole dermatologic referral center for south‐western Iran located in the city of Shiraz, during the period 1991–2000. The data were analyzed with regard to age, sex, subtypes of pemphigus, clinical presentation, and season of onset.

Results  The results showed that the mean incidence of the disease for the overall population in Shiraz was 0.67/105/year. The male to female ratio was 1 : 1.33. The mean age at onset of the disease was 38 years (range, 12–93 years). The disease was most common in the third to fifth decades (62.1% of the overall patients). Pemphigus vulgaris was the most commonly encountered type, followed by pemphigus foliaceus (87.7% and 9%, respectively). The first manifestations of the illness most often occurred during winter (30.8% of the patients).

Conclusions  The results reveal a high incidence of pemphigus vulgaris in south‐western Iran compared with that encountered in other countries. Our patients also had a relatively young age at onset and a female predominance.