bluuebunny/crossref_metadata_2025 huggingface dataset:
The important role of actinin-like protein (AcnA) in cytokinesis and apical dominance of hyphal cells in Aspergillus nidulans
Authors: Wang Jinjun , Hu Hongqin , Wang Sha , Shi Jie , Chen Shaochun , Wei Hua , Xu Xushi , Lu Ling
Published: July 2009

The actin cytoskeleton is involved in many processes in eukaryotic cells, including interaction with a wide variety of actin-binding proteins such as the actin-capping proteins, the actin filament nucleators and the actin cross-linking proteins. Here, we report the identification and characterization of an actinin-like protein (AcnA) from the filamentous fungusAspergillus nidulans. Not only did the depletion of AcnA byalcA(p) promoter repression or the deletion of AcnA result in explicit abnormalities in septation and conidiation, but also theacnAmutants induced a loss of apical dominance in cells with dichotomous branching, in which a new branch was formed by splitting the existing tip in two. Consequently, the colony showed flabellate edges. Moreover, we found that the localization of the GFP–AcnA fusion was quite dynamic. In the isotropic expansion phase of the germinated spore, GFP–AcnA was organized as cortical patches with cables lining the cell wall. Subsequently, GFP–AcnA was localized to the actively growing hyphal tips and to the sites of septation in the form of combined double contractile rings. Our data suggest that AcnA plays an important role in cytokinesis and apical dominance of hyphal cells, possibly via actin-dependent polarization maintenance and medial ring establishment inA. nidulans. This is the first report, to our knowledge, of the function of an actinin-like protein in filamentous fungi.
Identification of Rv3852 as a nucleoid-associated protein in Mycobacterium tuberculosis
Authors: Werlang Isabel C. R. , Schneider Cristopher Z. , Mendonça Jordana D. , Palma Mario S. , Basso Luiz A. , Santos Diógenes S.
Published: July 2009

Tuberculosis remains the major cause of mortality due to a bacterial pathogen,Mycobacterium tuberculosis. The molecular mechanisms of infection and persistence have not been completely elucidated for this pathogen. Studies involving nucleoid-associated proteins (NAPs), which have been related to the control and influence of virulence genes in pathogenic bacteria, can help unveil the virulence process ofM. tuberculosis. Here, we describe the initial characterization of an ORF for anM. tuberculosisputative NAP. TheRv3852gene was cloned and expressed, and its product purified to homogeneity. A qualitative protein–DNA binding assay was carried out by gel-retardation and the protein affinity for specific DNA sequences was assessed quantitatively by surface plasmon resonance (SPR). A stoichiometry of 10 molecules of monomeric protein per molecule of DNA was determined. The monophasic apparent dissociation rate constant values increased to a saturable level as a function of protein concentration, yielding two limiting values for the molecular recognition of proU2 DNA. A protein–DNA binding mechanism is proposed. In addition, functional complementation studies with anEscherichia coli hnsmutant reinforce the likelihood that the Rv3852 protein represents a novel NAP inM. tuberculosis.
Channelrhodopsins ofVolvox carteriAre Photochromic Proteins That Are Specifically Expressed in Somatic Cells under Control of Light, Temperature, and the Sex Inducer
Authors: Kianianmomeni Arash , Stehfest Katja , Nematollahi Ghazaleh , Hegemann Peter , Hallmann Armin
Published: July 2009

Channelrhodopsins are light-gated ion channels involved in the photoresponses of microalgae. Here, we describe the characterization of two channelrhodopsins, Volvox channelrhodopsin-1 (VChR1) and VChR2, from the multicellular green alga Volvox carteri. Both are encoded by nuclear single copy genes and are highly expressed in the small biflagellated somatic cells but not in the asexual reproductive cells (gonidia). Expression of both VChRs increases after cell cleavage and peaks after completion of embryogenesis, when the biosynthesis of the extracellular matrix begins. Likewise, expression of both transcripts increases after addition of the sex-inducer protein, but VChR2 is induced much more than VChR1. The expression of VChR1 is specifically promoted by extended dark periods, and heat stress reduces predominantly VChR1 expression. Expression of both VChRs increased under low light conditions, whereas cold stress and wounding reduced expression. Both VChRs were spectroscopically studied in their purified recombinant forms. VChR2 is similar to the ChR2 counterpart from Chlamydomonas reinhardtii with respect to its absorption maximum (460 nm) and photocycle dynamics. In contrast, VChR1 absorbs maximally at 540 nm at low pH (D540), shifting to 500 nm at high pH (D500). Flash photolysis experiments showed that after light excitation, the D540 dark state bleaches and at least two photoproducts, P600 and P500, are sequentially populated during the photocycle. We hypothesize that VChR2 is a general photoreceptor that is responsible for the avoidance of blue light and might play a key role in sexual development, whereas VChR1 is the main phototaxis photoreceptor under vegetative conditions, as it is more specifically adapted to environmental conditions and the developmental stages of Volvox.
A Structural Basis for the pH-Dependent Xanthophyll Cycle in Arabidopsis thaliana
Authors: Arnoux Pascal , Morosinotto Tomas , Saga Giorgia , Bassi Roberto , Pignol David
Published: July 2009

Plants adjust their photosynthetic activity to changing light conditions. A central regulation of photosynthesis depends on the xanthophyll cycle, in which the carotenoid violaxanthin is converted into zeaxanthin in strong light, thus activating the dissipation of the excess absorbed energy as heat and the scavenging of reactive oxygen species. Violaxanthin deepoxidase (VDE), the enzyme responsible for zeaxanthin synthesis, is activated by the acidification of the thylakoid lumen when photosynthetic electron transport exceeds the capacity of assimilatory reactions: at neutral pH, VDE is a soluble and inactive enzyme, whereas at acidic pH, it attaches to the thylakoid membrane where it binds its violaxanthin substrate. VDE also uses ascorbate as a cosubstrate with a pH-dependent K  m that may reflect a preference for ascorbic acid. We determined the structures of the central lipocalin domain of VDE (VDEcd) at acidic and neutral pH. At neutral pH, VDEcd is monomeric with its active site occluded within a lipocalin barrel. Upon acidification, the barrel opens up and the enzyme appears as a dimer. A channel linking the two active sites of the dimer can harbor the entire carotenoid substrate and thus may permit the parallel deepoxidation of the two violaxanthin β-ionone rings, making VDE an elegant example of the adaptation of an asymmetric enzyme to its symmetric substrate.
Successful major and minor surgery using factor VIII inhibitor bypassing activity in patients with haemophilia A and inhibitors
Authors: LAUROUA P. , FERRER A.‐M. , GUÉRIN V.
Published: July 2009

Summary.  Surgeries are being increasingly performed in patients with haemophilia A and high‐titre inhibitors. Optimal bypassing agent regimens need further delineation. Data pertaining to surgeries from 1989 to 2004 at a single centre were retrospectively analysed. Patients received a standardized factor eight inhibitor bypassing activity (FEIBA) dose for both major and minor elective or emergency surgeries. The standard FEIBA dose was 70 U kg−1 per infusion. FEIBA was infused at 9 and 1 h before and 8 h after operation. Infusions were routinely repeated every 8 h afterward. Haemostatic efficacy was assessed on the basis of blood loss, occurrence of haematoma and transfusion requirements. Seven adult patients underwent a total of 12 operations: 10 major and two minor. Ten procedures were elective. The median cumulative numbers of infusions and days of therapy were 46 and 17, respectively. Cumulative total FEIBA consumption was a median of 3185 U kg−1. Observed blood losses, haematoma incidence and transfusion requirements were comparable to those expected for noncoagulopathic patients undergoing similar procedures. The only large haematoma occurred after a hip prosthesis operation and resolved under continuing FEIBA treatment. There were no cases of disseminated intravascular coagulation or other thromboembolic complications. FEIBA provides an effective and safe first‐line peri‐ and postoperative haemostatic therapy for patients with haemophilia A and inhibitors, allowing both major and minor operations to be successfully performed.
Thrombin generation and platelet activation induced by rFVIIa (NovoSeven®) and NN1731 in a reconstituted cell‐based model mimicking haemophilia conditions
Authors: ALJAMALI M. N. , KJALKE M. , HEDNER U. , EZBAN M. , TRANHOLM M.
Published: July 2009

Summary.  Replacement therapy with factor VIII (FVIII) and factor IX (FIX) is routinely used in haemophilia patients with haemophilia A and B, respectively, while recombinant activated FVII (rFVIIa) has proven to induce haemostasis in haemophilia patients with inhibitors. To evaluate the effect of therapeutic intervention in patients with residual factor activities, the effects of increasing concentrations of rFVIIa or NN1731 on thrombin generation and platelet activation were measured in a cell‐based model system mimicking severe, moderate and mild haemophilia A or B. Purified monocytes stimulated to express tissue factor and non‐activated platelets from peripheral blood of healthy donors were incubated with a mixture of purified human coagulation factors in the absence or presence of increasing concentrations of FVIII or FIX. Sub‐samples were analysed for thrombin activity and platelet activation measured as exposure of P‐selectin by flow cytometry. Dose‐dependent increases in thrombin generation and platelet activation were observed following increasing concentrations of rFVIIa or NN1731 in both haemophilia A‐ and B‐like conditions. At 25 nm rFVIIa, which nears the peak levels in patient plasma after 90 μg kg−1 intravenous dosing, the effects on maximum thrombin generation rate (maxTG) at 1–10% FVIII were comparable to those at 100% and 200% FVIII in the absence of rFVIIa. Normalization of maxTG required 500 nm rFVIIa and 25 nm NN1731 or 25–100 nm rFVIIa and 5 nm NN1731 in severe or moderate/mild haemophilia A and haemophilia B, respectively. This suggests that NN1731 holds its promise as a future bypassing agent for haemophilia patients with and without inhibitors.
Ethanol lock therapy for the treatment of catheter‐related infections in haemophilia patients
Authors: RAJPURKAR M. , BOLDT‐MACDONALD K. , MCLENON R. , CALLAGHAN M. U. , CHITLUR M. , LUSHER J. M. , BECKER C.
Published: July 2009

Summary.  Central venous access devices (CVAD) are increasingly being used for optimal delivery of clotting factor concentrates in patients with haemophilia with poor peripheral venous access. The utility of CVAD is particularly well recognized in young patients starting factor prophylaxis and in patients with inhibitors undergoing immune tolerance induction (ITI). A catheter‐related infection (CRI) remains the most common complication of CVAD in haemophilia patients and is the most frequent indication for its removal. Additionally, in some patients the infection results in significant morbidity and mortality and also contributes to failure of the ITI regimen. Ethanol‐lock therapy (ELT) is a treatment modality that has been used to treat CRI in patients with indwelling catheters for home parenteral nutrition and chemotherapy. The aim of this study was to report the success in treating CRI in haemophilia patients using ELT. Three severe haemophilia A patients undergoing ITI regimen who developed CVAD infections resistant to conventional management with antibiotics were treated by ELT according to the institutional technique. All three patients responded well to ELT with clearance of the CVAD infection. There were no adverse side effects. To our knowledge, this is the first report of ELT in patients with haemophilia. The role of ELT needs to be investigated in larger studies for treatment of CRI in patients with bleeding disorders.
Influence of phylogeny and ploidy on species ranges of North American angiosperms
Authors: Martin Sara L , Husband Brian C.
Published: July 2009

1. Species differ widely in the size and ecological characteristics of their geographic ranges. This variation reflects, in part, differences in ecological tolerance among taxa, but the influence of phylogeny and evolutionary divergence through genome duplication remains controversial.

2. Here, we examine the evolutionary sources of variation in species range area, latitudinal position and overlap, and six range climatic variables (maximum, minimum and breadth of temperature and precipitation) for North American angiosperms. We sampled two diploid and one polyploid species from each of 144 genera and examined sources of variation in a nested taxonomic analysis and congeneric species pairs comparison.

3. For all range characteristics studied, a significant portion of the variance among species (mean 53%, range 37–60%) was attributable to taxonomic membership above species (i.e. genus, family and order). Four of the seven range attributes were significantly correlated between diploid congeners (e.g. range arears = 0.65) and between heteroploid (diploid and polyploid) congeners (e.g. range arears = 0.62). The strength of correlations between heteroploid relatives was not consistently different than correlations between diploids.

4. No differences in mean range breadth (range area, temperature breadth, precipitation breadth) or climatological position (minimum and maximum temperature and precipitation) were observed between diploid and polyploid congeners. Polyploids were not more likely to occupy northern latitudinal positions and were no more displaced from diploids than other congeneric diploids.

5.Synthesis. Our results suggest that, at the scale of North America, the geographical and ecological aspects of species ranges are influenced by phylogenetic history. Furthermore, genome duplication does not cause or allow greater divergence in species range characteristics than occurs between related diploid species.
Improving geriatric transitional care through inter‐professional care teams
Authors: Blewett Lynn A. , Johnson Kelli , McCarthy Teresa , Lackner Thomas , Brandt Barbara
Published: July 2009

Objectives  The aim of this study was to examine the impact of the use of an inter‐professional care team on patient length of stay and payer charges in a geriatric transitional care unit.

Methods  An analysis of de‐identified administrative records for transitional care patients for the 12‐month period (2003–2004) cared for by the inter‐professional team (n = 163) and cared for by traditional single provider care model (n = 176) was carried out. We conducted logistic regression on length of stay and charges controlling for patient demographics and acuity levels.

Results  The inter‐professional care team patients had significantly shorter lengths of stay, fewer patient days and lower total charges. Patient diagnosis and acuity were similar across groups.

Conclusion  This study provides empirical evidence of the impact of an inter‐professional care model in providing cost‐effective transitional care in a nursing home setting. Evidence of shorter lengths of stay, shorter patient days and lower charges suggests benefit in the development and financing of inter‐professional care teams for transitional care services.
Linking impulsivity to dysfunctional thought control and insomnia: a structural equation model
Authors: SCHMIDT RALPH E. , GAY PHILIPPE , GHISLETTA PAOLO , VAN DER LINDEN MARTIAL
Published: July 2009

According to cognitive models of insomnia, excessive mental activity at bedtime may be viewed as an important impediment to the process of falling asleep. A further assumption of these models is that ‘cognitive arousal’ may be perpetuated and exacerbated by counterproductive strategies of thought management. As yet, little is known about factors that may predispose people to rely on these strategies when confronted with thoughts that keep them awake at night. This study examined the relations between impulsivity, use of different thought‐control strategies and insomnia severity. A sample of 391 university students completed the UPPS Impulsive Behavior Scale, the Thought Control Questionnaire Insomnia‐Revised and the Insomnia Severity Index. Correlation analyses revealed that two facets of impulsivity (urgency and lack of perseverance), two strategies of thought control (aggressive suppression and worry) and insomnia severity were positively associated. Follow‐up structural equation modeling analyses showed that the two mentioned thought‐control strategies mediated the effects of the two facets of impulsivity on sleep problems. These findings extend existing cognitive accounts of insomnia by suggesting how predisposing and perpetuating factors may be related: specific personality traits may incline individuals to respond with dysfunctional thought‐control strategies to unwanted mental activity at night.
An RNA trap helps bacteria get the most out of chitosugars
Authors: Vogel Jörg
Published: July 2009

Small regulatory RNAs (sRNAs) are well known to command bacterial protein synthesis by modulating the translation and decay of target mRNAs. Most sRNAs are specifically regulated by a cognate transcription factor under certain growth or stress conditions. Investigations of the conserved Hfq‐dependent MicM sRNA in Escherichia coli (article by Poul Valentin‐Hansen and colleagues in this issue of Molecular Microbiology) and in Salmonella have unravelled a novel type of gene regulation in which the chitobiose operon mRNA acts as an RNA trap to degrade the constitutively expressed MicM sRNA, thereby alleviating MicM‐mediated repression of the synthesis of the YbfM porin that is required for chitosugar uptake. The results suggest that ‘target’ mRNAs might be both prey and also predators of sRNAs.
Genetic testing in familial AD and FTD: Mutation and phenotype spectrum in a Danish cohort
Authors: Lindquist SG , Schwartz M , Batbayli M , Waldemar G , Nielsen JE
Published: July 2009

Autosomal dominantly transmitted Alzheimer's disease (AD) and frontotemporal dementia (FTD) are genetically heterogeneous disorders. To date, three genes have been identified in which mutations cause early‐onset autosomal dominant inherited AD: APP, PSEN1, and PSEN2. Mutations in two genes on chromosome 17, the MAPT and the PGRN genes, are associated with autosomal dominant inherited FTD. The aim of this study was to characterize the mutation spectrum and describe genotype‐phenotype correlations in families with inherited dementia. The identification of novel mutations and/or atypical genotype‐phenotype correlations contributes to further characterizing the disorders. DNA‐samples from the 90 index cases from a Danish referral‐based cohort representing families with presumed autosomal dominant inherited AD or FTD were screened for mutations in the known genes with sequencing, denaturing high‐performance liquid chromatography (DHPLC) and multiplex ligation‐dependent probe amplification (MLPA) techniques. Seven presumed pathogenic mutations (two PSEN1, one PSEN2, one APP, one MAPT, and two PGRN) were identified, including a novel PSEN2 mutation (V393M). No dosage aberrations were identified.
Leprosy in a renal transplant recipient: review of the literature
Authors: Guditi S. , Ram R. , Ismal K.M. , Sahay M. , Dakshinamurthy K.V. , Girish N. , Prasad N.
Published: July 2009

Abstract: A 52‐year‐old male underwent living‐related renal transplantation. He received prednisolone, azathioprine, and cyclosporine as immunosuppression protocol. Eleven years after transplantation, he developed pyrexia with multiple nodular lesions on his limbs, trunk, and face. Skin biopsy and smears showed the presence of numerous acid‐fast bacilli with 5% sulfuric acid indicative of Mycobacterium leprae. He was initiated on multidrug therapy (MDT) including dapsone, clofazimine, and rifampicin. After 2 years of MDT, he developed new multiple erythematous, tender subcutaneous nodules in crops over his face and upper limbs. Skin biopsies and histopathological examination confirmed the diagnosis of type 2 lepra reaction or erythema nodosa leprosum. He was managed with an increase in the dose of prednisolone and thalidomide. He was continued on MDT.
Context‐Related Variation in the Vocal Growling Behaviour of the Domestic Dog (Canis familiaris)
Authors: Taylor Anna M. , Reby David , McComb Karen
Published: July 2009

We investigated whether the growls of domestic dogs, Canis familiaris, exhibit predictable variations in a range of key acoustic parameters when they are given in two contrasting experimental contexts, and whether humans are sensitive to such variation. In a standardized experimental paradigm, an experimenter visited 32 domestic dogs and generated an aggression context and a play context. In these contexts, 204 isolated growls were recorded and subsequently acoustically analysed. Contrary to previous findings on barks, fundamental and formant frequencies of growls did not vary between the two contexts. However, growls from the aggression context were significantly longer than growls from the play context. Additionally, the temporal structure of the vocal sequences containing growls differed significantly between the contexts. In a series of psychoacoustic experiments, human listeners were not able to discriminate between growls recorded in the two contexts, however both aggressiveness and playfulness ratings were strongly influenced by dog weight, with larger dogs being perceived as more aggressive than smaller dogs. However, participants were able to attribute aggressiveness and playfulness to synthesized sequences in which growls occurred at a two rates, typical of either the aggression or the play context. We conclude that context‐related variation in growling behaviour resides in the temporal structure, rather than in the acoustic composition of isolated growls, and furthermore that human listeners are able to attribute context on the basis of growling rate. In addition, human appear to have an intuitive acoustic bias toward perceiving larger dogs as more aggressive.
Adverse reactions to azathioprine cannot be predicted by thiopurine S‐methyltransferase genotype in Japanese patients with inflammatory bowel disease
Authors: Takatsu Noritaka , Matsui Toshiyuki , Murakami Yuji , Ishihara Hiroshi , Hisabe Takashi , Nagahama Takashi , Maki Shinichirou , Beppu Takahiro , Takaki Yasuhiro , Hirai Fumihito , Yao Kenshi
Published: July 2009

Background and Aims:  Azathioprine (AZA) is associated with a high frequency of adverse reactions. We examined polymorphism of the thiopurine S‐methyltransferase (TPMT) gene to determine whether the TPMT genotype would be a predictive marker for the development of adverse reactions to AZA.

Methods:  The frequency of TPMT mutations was investigated in 147 Japanese inflammatory bowel disease (IBD) patients retrospectively. In these subjects, the presence of four mutant alleles (TPMT*2, *3B, *3C and *8) was determined by direct sequencing. The incidence of adverse reactions among patients carrying wild‐type TPMT was investigated. The blood level of 6‐thioguanine nucleotide (6‐TGN) was measured and analyzed in 47 patients with wild‐type TPMT. The results were analyzed in relation to the concomitant use of aminosalicylates (ASA).

Results:  Of the 147 patients, 144 (98.0%) were wild‐type for TPMT (TPMT*1/*1) and three (2.0%) carried a mutant TPMT allele (TPMT*1/*3C). The incidence of adverse reactions was 33.3% (38/114) in the wild‐type group. Leukopenia (WBC ≤ 3000/µL) was seen in 15.8% of the patients with wild‐type TPMT. 6‐TGN levels varied among 47 patients with wild‐type TPMT. The blood levels of 6‐TGN were significantly higher in the patients receiving concomitant ASA treatment compared with those not receiving concomitant ASA treatment (P = 0.0033).

Conclusion:  The frequency of TPMT gene mutations is low among Japanese IBD patients. The incidence of adverse reactions to AZA was high, even in patients carrying wild‐type TPMT. It is concluded that determination of TPMT genotype may not be useful in Japanese IBD patients to predict adverse reactions to AZA.
Efficacy of the bell and pad alarm therapy for nocturnal enuresis
Authors: Gim Cindy SY , Lillystone David , Caldwell Patrina HY
Published: July 2009

Aim:  (i) To determine the efficacy of bell and pad alarm therapy as an initial and relapse treatment for nocturnal enuresis; (ii) to explore risk factors for treatment failure; and (iii) to explore risk factors for relapse within 12 months of successful bell and pad alarm therapy.

Methods:  A 22‐item questionnaire was sent to 240 children who received bell and pad alarm therapy in a 6‐year period via a community centre. The questionnaire recorded demographic characteristics of the child, length of the first bell and pad alarm therapy, outcome of initial treatment and relapse information.

Results:  The initial response and relapse rates of bell and pad alarm therapy were 84 and 30%, respectively. Female gender, absence of diurnal symptoms and willingness to use alarm therapy were associated with better treatment outcomes. Treatment success was associated with shorter treatment length. The success rate of repeating alarm therapy after relapse was 78%, with an average length of treatment of 10 weeks.

Conclusion:  The bell and pad alarm therapy is an effective treatment for nocturnal enuresis both as initial therapy and after relapse. The association between patient characteristics and treatment response found in our study may help inform clinicians of likely treatment outcomes, and identify those who may need a different approach.
Epithelial‐mesenchymal transition induced by transforming growth factor‐β1 in mouse tracheal epithelial cells
Authors: KUROISHI Shigeki , SUDA Takafumi , FUJISAWA Tomoyuki , IDE Kyotaro , INUI Naoki , NAKAMURA Yutaro , NAKAMURA Hirotoshi , CHIDA Kingo
Published: July 2009

Background and objective:  Epithelial‐mesenchymal transition (EMT) is the process by which differentiated epithelial cells undergo a phenotypic transition to mesenchymal cells. This process may occur in certain fibrotic diseases that involve airway remodelling. However, few studies have directly proved the occurrence of EMT in primary cultures of airway epithelial cells. The aim of this study was to clarify whether airway epithelial cells can differentiate into mesenchymal cells through EMT.

Methods:  Mouse tracheal epithelial cells (mTEC) were cultured in an air–liquid interface (ALI) culture system, in the presence or absence of transforming growth factor‐β1 (TGF‐β1). The expression of mesenchymal and epithelial cell markers was examined by immunofluorescence staining and western blotting. Secretion of matrix metalloproteinase (MMP)‐9 into the culture medium was measured by ELISA. The phenotype of epithelial cells involved in EMT was also examined by immunofluorescence staining.

Results:  Immunofluorescence staining and western blotting revealed that TGF‐β1 treatment for 14 days induced the expression of the mesenchymal markers, α‐smooth muscle actin (α‐SMA) and vimentin, and reduced the expression of the epithelial markers, zonula occludens‐1 (Zo‐1) and occludin. In addition, α‐SMA and Zo‐1 were colocalized within individual cells treated with TGF‐β1. Concentrations of MMP‐9 in the culture medium were significantly higher in TGF‐β1‐treated mTEC than in untreated cells. The basal cell markers, cytokeratin‐5 and cytokeratin‐17, were colocalized within the cells expressing α‐SMA.

Conclusions:  EMT was induced by TGF‐β1 in primary cultures of mTEC, suggesting that airway epithelial cells, possibly the basal cells, may be involved in airway remodelling through EMT.
Early recurrence of Pneumocystis jiroveci pneumonia in two HIV‐infected patients: Linking infection relapse and immune reconstitution syndrome
Authors: KOLDITZ Martin , HALANK Michael , BANDT Dirk , SPORNRAFT‐RAGALLER Petra , HÖFFKEN Gert
Published: July 2009

This case report describes two HIV‐positive patients with Pneumocystis jiroveci pneumonia who relapsed within 1 week of starting secondary prophylaxis and antiretroviral treatment. Diagnostic and therapeutic approaches as well as the need to establish risk factors for infection recurrence or early immune reconstitution syndrome are highlighted.
Novel method for sputum induction using the Lung Flute in patients with suspected pulmonary tuberculosis
Authors: FUJITA Akira , MURATA Kengo , TAKAMORI Mikio
Published: July 2009

Background and objective:  The Lung Flute is a small self‐powered audio device that generates sound waves, which vibrate in tracheobronchial secretions. This was a preliminary trial to evaluate the usefulness of the Lung Flute for sputum sampling in patients suspected of pulmonary tuberculosis (TB).

Methods:  Thirty‐four patients who were not expectorating sputum, but for whom sputum examination was required for the differential diagnosis of TB or other diseases, were enrolled in the study. Patients were instructed to blow out fast and hard through the Lung Flute and to repeat this for a total 20 sets of two blows each.

Results:  Using the Lung Flute, sputum samples were collected within 10 or 20 min from 30 of 34 patients (88%). The device permitted a rapid diagnosis of TB in seven of 15 confirmed TB cases. In three patients acid‐fast bacillus smears were positive. In four patients acid‐fast bacillus smears were negative, but PCR tests for TB were positive. Hyperventilation‐related symptoms occurred in three patients.

Conclusions:  The application of the Lung Flute may represent a promising technique for the rapid diagnosis of pulmonary TB.
Interaction of the C‐terminal tail region of the metabotropic glutamate receptor 7 with the protein kinase C substrate PICK1
Authors: El Far Oussama , Airas José , Wischmeyer Erhardt , Nehring Ralf B. , Karschin Andreas , Betz Heinrich
Published: July 2009

Group III metabotropic glutamate receptors (mGluRs) are highly enriched in the presynaptic terminals of glutamatergic synapses where they mediate feedback inhibition of neurotransmitter release. Here, we used the yeast two‐hybrid system to identify a direct interaction of the C‐terminal tail region of mGluR7 with the rat homologue of the protein kinase C substrate PICK1. This interaction is specifically mediated by the very C‐terminal amino acids of the receptor and can be reconstituted in human embryonic kidney 293 cells by transfection of full‐length mGluR7 and PICK1 cDNAs. Quantitative β‐galactosidase assays revealed that among the different group III mGluRs, mGluR7 is the major PICK1 binding partner although other subfamily members can also interact with PICK1. These data indicate that PDZ domain‐containing proteins might contribute to the presynaptic localization of group III mGluRs.
Differential roles of corticotropin‐releasing factor receptor subtypes 1 and 2 in opiate withdrawal and in relapse to opiate dependence
Authors: Lu Lin , Liu Dehong , Ceng Xiabo , Ma Lan
Published: July 2009

The possible effects on the morphine withdrawal signs of the nonspecific corticotropin‐releasing factor (CRF) receptor antagonist α‐helical CRF, the selective CRF receptor subtype 1 antagonist CP‐154,526 and the selective CRF receptor subtype 2 antagonist antisauvagine‐30 (AS‐30) were investigated in rats. The most withdrawal signs, including jumping, teeth chatter, writhing, shakes, lacrimation, piloerection, irritability and diarrhoea, were attenuated by pretreatment with α‐helical CRF (10 µg i.c.v.) and CP‐154,526 (30 mg/kg i.p.). However, no morphine withdrawal signs except for diarrhea were significantly affected by pretreatment with AS‐30 (10 µg, i.c.v.). To investigate the possible role of different CRFR antagonists (α‐helical CRF, CP‐154,526 and AS‐30) in relapse to opiate dependence, the 28‐day extinction of morphine‐conditioned place preference (CPP) was used. The morphine‐CPP disappeared following a 28‐day extinction and then was reactivated by a single injection of 10 mg/kg morphine. Pretreatment with α‐helical CRF (10 µg, i.c.v.) and CP‐154,526 (30 mg/kg, i.p.) could significantly block this reactivation of morphine‐CPP. In contrast, pretreatment with AS‐30 (1 or 10 µg i.c.v.) did not affect this reactivation of morphine‐CPP. The present study demonstrated that activation of the CRF receptor is involved in morphine withdrawal signs and relapse to morphine dependence, and that the role of CRF receptor subtypes 1 and 2 in withdrawal and reactivation of morphine dependence is not identical. CRF receptor subtype 1, but not subtype 2, is largely responsible for the action of the CRF system on opiate dependence. These results suggest that the CRF receptor antagonists, particularly the CRF receptor subtype 1 antagonist, might be of some value in the treatment and prevention of drug dependence.
Interactions between imidazoline binding sites and dopamine levels in the rat nucleus accumbens
Authors: Barrot Michel , Rettori Marie‐Claire , Guardiola‐Lemaître Béatrice , Jarry Christian , Le Moal Michel , Piazza Pier Vincenzo
Published: July 2009

Imidazoline binding sites are present in the striatal complex and in the extended amygdala and have been implicated in mood disorders. In this report we analysed the influence of these sites on the functional activity of the mesolimbic dopaminergic transmission, one of the major brain systems involved in the regulation of motivation and reward. We studied the effects of two imidazoline ligands, S23229 and S23230 (respectively S(+) and R(–) enantiomers of the S22687 or (5‐[2‐methyl phenoxy methyl] 1,3‐oxazolin‐2‐yl) amine), on extracellular dopamine in the nucleus accumbens using microdialysis in freely moving rats. We compared these imidazoline ligands to cocaine, a dopamine uptake blocker known to increase extracellular dopamine concentrations. S23229 dose‐dependently increased extracellular dopamine and locomotor activity. S23230 dose‐dependently increased extracellular dopamine and produced a near‐significant dose–effect on locomotor activity. S23229 had a stronger efficacy than S23230 and increased dopamine levels in the nucleus accumbens at an extent similar to the one of cocaine. These results suggest that central imidazoline binding sites could contribute to the functional regulation of the mesolimbic dopaminergic system.
A brain slice model for in vitro analyses of astrocytic gap junction and connexin43 regulation: actions of ischemia, glutamate and elevated potassium
Authors: Nagy J. I. , Li W. E. I.
Published: July 2009

Brain slices prepared from adult rats and maintained for up to 3 h in vitro were used to investigate the effects of pharmacological treatments on the phosphorylation state, immunolabelling characteristics and ultrastructural localization of astrocytic gap junctions and connexin43 (Cx43). Slices deprived of glucose/oxygen to mimic ischemia or those exposed to 1 mm glutamate for 1 h exhibited Cx43 dephosphorylation, epitope masking and gap junction internalization as revealed by Western blotting and Cx43 immunolocalization with various antibodies. Treatment with 15 mm K+ caused Cx43 dephosphorylation without junction internalization. The effects of glutamate and K+ were completely blocked by the N‐methyl‐d‐aspartate (NMDA) glutamate receptor antagonist 2‐amino‐5‐phosphonovalerate (APV), which acts largely on neuronal NMDA receptors, suggesting neuronal mediation of glial gap junction responses to these treatments. Astrocytes contained a dephosphorylated form of Cx43 with a typical migration profile at 41 kDa as well as novel, apparently dephosphorylated or partially phosphorylated, forms migrating at 43 kDa. These results indicate that slices prepared from adult brain can serve as a convenient model to investigate the molecular basis and receptor‐mediated mechanisms underlying astrocytic Cx43 responses that have been observed in vivo following cerebral ischemia or neural activation. These processes can be related in part to neuronal regulation of astrocytic gap junctional coupling state, which is also amenable to analysis in brain slices.
Subcellular compartmentalization of a potassium channel (Kv1.4): preferential distribution in dendrites and dendritic spines of neurons in the dorsal cochlear nucleus
Authors: Juiz José M. , Luján Rafael , Domínguez del Toro Eduardo , Fuentes V. , Ballesta Juan J. , Criado Manuel
Published: July 2009

Voltage‐dependent ion channels have specific patterns of distribution along the neuronal plasma membrane of dendrites, cell bodies and axons, which need to be unravelled in order to understand their contribution to neuronal excitability and firing patterns. We have investigated the subcellular compartmentalization of Kv1.4, a transient, fast‐inactivating potassium channel, in fusiform cells and related interneurons of the rat dorsal cochlear nucleus. A polyclonal antibody which binds to a region near the N‐terminus domain of a Kv1.4 channel was raised in rabbits. Using a high‐resolution combination of immunocytochemical methods, Kv1.4 was localized mainly in the apical dendritic trunks and cell bodies of fusiform cells, as well as in dendrites and cell bodies of interneurons of the dorsal cochlear nucleus, likely cartwheel cells. Quantitative immunogold immunocytochemistry revealed a pronounced distal to proximal gradient in the dendrosomatic distribution of Kv1.4. In plasma membrane localizations, Kv1.4 was preferentially present in dendritic spines, either in the spine neck or in perisynaptic locations, always away from the postsynaptic density. These findings indicate that Kv1.4 is largely distributed in dendritic compartments of fusiform and cartwheel cells of the dorsal cochlear nucleus. Its preferential localization in dendritic spines, where granule cell axons make powerful excitatory synapses, suggests a role for this voltage‐dependent ion channel in the regulation of dendritic excitability and excitatory inputs.
Topographical projection from the superior colliculus to the nucleus of the brachium of the inferior colliculus in the ferret: convergence of visual and auditory information
Authors: Doubell Timothy P. , Baron Jérôme , Skaliora Irini , King Andrew J.
Published: July 2009

The normal maturation of the auditory space map in the deeper layers of the ferret superior colliculus (SC) depends on signals provided by the superficial visual layers, but it is unknown where or how these signals influence the developing auditory responses. Here we report that tracer injections in the superficial layers label axons with en passant and terminal boutons, both in the deeper layers of the SC and in their primary source of auditory input, the nucleus of the brachium of the inferior colliculus (nBIC). Electron microscopy confirmed that biocytin‐labelled SC axons form axodendritic synapses on nBIC neurons. Injections of biotinylated dextran amine in the nBIC resulted in anterograde labelling in the deeper layers of the SC, as well as retrogradely labelled superficial and deep SC neurons, whose distribution varied systematically with the rostrocaudal placement of the injection sites in the nBIC. Topographical order in the projection from the SC to the ipsilateral nBIC was confirmed using fluorescent microspheres. We demonstrated the existence of functional SC‐nBIC connections by making whole‐cell current‐clamp recordings from young ferret slices. Both monosynaptic and polysynaptic EPSPs were generated by electrical stimulation of either the superficial or deep SC layers. In addition to unimodal auditory units, both visual and bimodal visual–auditory units were recorded in the nBIC in vivo and their incidence was higher in juvenile ferrets than in adults. The SC‐nBIC circuit provides a potential means by which visual and other sensory or premotor signals may be delivered to the nBIC to calibrate the representation of auditory space.
Effect of feeding on Fos protein expression in sheep hypothalamus with special reference to the supraoptic and paraventricular nuclei: an immunohistochemical study
Authors: Chaillou Elodie , Baumont René , Tramu Gérard , Tillet Yves
Published: July 2009

The hypothalamus plays an important role in the control of food intake in different species, but there is little relevant information for ruminants like sheep. In order to study the putative role of several hypothalamic nuclei in food intake in sheep, Fos expression, a marker of cellular activity, was compared by immunohistochemistry between fed and unfed ewes. The expression of Fos protein was stimulated in the supraoptic nucleus of fed ewes, whereas it was increased in the paraventricular nucleus of unfed animals. In the latter nucleus, Fos immunoreactivity was mainly localized close to the third ventricle, an area corresponding to the parvocellular system of the nucleus, but never in the magnocellular system. In the paraventricular nucleus, the number of corticotrophin releasing factor‐immunoreactive neurons and the number of Fos/corticotrophin releasing factor double‐labelled neurons were not affected by feeding or lack of feeding. The number of Fos‐immunoreactive neurons was higher in the lateral septum, the infundibular, the ventromedial and in the dorsomedial nuclei of unfed ewes than in those of fed ewes. Our results show for the first time that the dorsomedial and ventromedial nuclei are involved in the control of feeding in sheep as in rodents. The supraoptic nucleus of sheep is activated by the same conditions as in rodents but, conversely, the paraventricular nucleus is activated in unfed sheep, whereas in rodents and primates, this nucleus is activated by satiety as well as by fasting. In sheep, unlike in rodents, corticotrophin releasing factor did not appear to be involved in short‐term regulation of food intake.
Assignment of ecto‐nucleoside triphosphate diphosphohydrolase‐1/cd39 expression to microglia and vasculature of the brain
Authors: Braun Norbert , Sévigny Jean , Robson Simon C. , Enjyoji Keiichi , Guckelberger Olaf , Hammer Klaus , Di Virgilio Francesco , Zimmermann Herbert
Published: July 2009

Extracellular nucleotides are ubiquitous extracellular mediators that interact with and activate nucleotide type 2 (P2) receptors. These receptors initiate a wide variety of signalling pathways that appear important for functional associations between neurons and glial cells and for the regulation of blood flow, haemostatic and inflammatory reactions in the brain. Ectonucleotidases are extracellular nucleotide‐metabolizing enzymes that modulate P2 receptor‐mediated signalling by the regulated hydrolysis of these agonists. A considerable number of ectoenzyme species with partially overlapping substrate and tissue distributions have been described. Major candidates for expression in the brain are members of the ecto‐nucleoside triphosphate diphosphohydrolase (E‐NTPDase or CD39) family. The production of cd39–/–mice and specific reagents have enabled us to analyse the specific cellular distribution of NTPDase1 (CD39), the prototype member of the enzyme family, in the mouse brain. Using monospecific antibodies and enzyme histochemical staining, we have identified NTPDase1 as a major ectonucleotidase associated with both microglia and the endothelial and smooth muscle cells of the vasculature. NTPDase1 is not expressed by neurons and astrocytes. Additional unidentified ectonucleotidase functional activity is observed at lower levels throughout the brain parenchyma. NTPDase1 may regulate P2 receptor‐mediated functions of microglia as well as influence nucleotide signalling between neurons or astrocytes that are associated with multiple microglial ramifications. The expression of NTPDase1 by cerebrovascular endothelial and smooth muscle cells also suggests involvement in the regulation of blood flow and thrombogenesis.
cDNA cloning and expression of a γ‐aminobutyric acidAreceptor ε‐subunit in rat brain
Authors: Moragues Nathalie , Ciofi Philippe , Lafon Pierrette , Odessa Marie‐Françoise , Tramu Gérard , Garret Maurice
Published: July 2009

A cDNA encoding a GABAAreceptor subunit was isolated from rat brain. The predicted protein is 70% identical to the human ε‐subunit. It was recently reported [Sinkkonenet al. (2000),J. Neurosci.,20, 3588–3595] that the rodent ε‐subunit mRNA encoded an additional sequence (≈400 residues). We provide evidence that human and rat ε‐subunit are similar in size. The distribution of cells expressing the GABAAε‐subunit was examined in the rat brain.In situhybridization histochemistry revealed that ε‐subunit mRNA is expressed by neurons located in septal and preoptic areas, as well as in various hypothalamic nuclei, including paraventricular, arcuate, dorsomedial and medial tuberal nuclei. The mRNA was also detected in major neuronal groups with broad‐range influence, such as the cholinergic (basal nucleus), dopaminergic (substantia nigra compacta), serotonergic (raphe nuclei), and noradrenergic (locus coeruleus) systems. Immunohistochemistry using an affinity‐purified antiserum directed towards the N‐terminal sequence unique to the rat ε‐subunit revealed the presence of ε‐subunit immunoreactivity over the somatodendritic domain of neurons with a distribution closely matching that of mRNA‐expressing cells. Moreover, usingin situhybridization, α3, θ and ε GABAAsubunit mRNAs were all detected with an overlapping distribution in neurons of the dorsal raphe and the locus coeruleus. Our results suggest that novel GABAAreceptors may regulate, neuroendocrine and modulatory systems in the brain.
Dopaminergic innervation of the pallidum in the normal state, in MPTP‐treated monkeys and in parkinsonian patients
Authors: Jan Caroline , François Chantal , Tandé Dominique , Yelnik Jérôme , Tremblay Léon , Agid Yves , Hirsch Etienne
Published: July 2009

The aim of the present study was to characterize the dopaminergic innervation of the pallidum in primates (humans and Cercopithecus aethiops). Firstly, in monkeys, biotin dextran amine was injected into dopaminergic areas, and the anterogradely labelled axons were reconstructed from serial sections and analysed in the pallidum. Secondly, in parkinsonian patients and MPTP‐treated monkeys, the dopaminergic innervation of the pallidum was studied using tyrosine hydroxylase‐positive fibre quantification. Our study revealed that dopaminergic areas A8 and A9 innervated the two pallidal segments. Individual axonal arborizations displayed a great heterogeneity. Some dopaminergic axons crossed the pallidum without branching, other axons made small terminal arborizations in a restricted region of one pallidal segment, whereas others developed dense arborizations covering extended areas in the two pallidal segments. This heterogeneous organization suggests that dopamine could directly modulate the pallidum using either a point‐to‐point or a diffuse projection pattern. A statistically significant loss of dopaminergic fibres in the internal (−43%) and external pallidum (−39.6%) of humans, and in the internal (−54.3%) and external pallidum (−59%) of monkeys was revealed in parkinsonian states. The consequences of this alteration are still unknown but it might participate in the triggering of motor symptoms observed in Parkinson's disease.