bluuebunny/crossref_metadata_2025_split huggingface dataset:
Functional in vivo studies of the Neurospora crassa cys‐14 gene upstream region: importance of CYS3‐binding sites for regulated expression
Authors: Li Qunhui , Zhou Liwei , Marzluf George A. 
Published: October 2006

Summary Sulphate transport in Neurospora crassa is achieved by two distinct sulphate permeases, I and II, encoded by the cys‐13 and cys‐14 genes, respectively. The synthesis of both sulphate permeases is subject to sulphur repression and requires the global positive‐acting regulatory protein CYS3. CYS3, a bZIP DNA‐binding protein, regulates cys‐14 expression at the transcriptional level and binds in vitro specifically to three DNA‐recognition sites, A, B, and C, in the cys‐14 upstream region. In vivo functional analysis of the cys‐14 promoter was carried out with 5’deletions and by deletions or mutations of CYS3 DNA‐binding sites. The most distal CYS3‐binding site, C, located 1.4kb upstream of the transcriptional start site, is necessary and sufficient to mediate strong transcriptional activation by CYS3; moreover, site C was able to function equally well when it was located at variable distances upstream of the cys‐14 gene. Site B, located 1 kb upstream, alone is able to support a moderate degree of cys‐14 expression. Site A is not required and does not appear to play any functional role in cys‐14 expression, even though it is in close proximity to the transcriptional start site. The presence of multiple copies of CYS3‐binding elements A or B in the cys‐14 promoter results in a parallel increase of regulated gene expression. When a transforming cys‐14 gene becomes integrated at ectopic locations in the host genome, it can be expressed in an unregulated fashion, presumably by coming under the control of other promoter elements. Our results also suggested that at least one enzyme in the sulphate catabolic pathway requires a functional CYS3 protein for expression.
Involvement of the sensor kinase EnvZ in the in vivo activation of the response‐regulator PhoB by acetyl phosphate
Authors: Kim Soo‐Ki , Wilmes‐Riesenberg Mary R. , Wanner Barry L. 
Published: October 2006

SummaryThree signalling pathways lead to activation of the phosphate (Pho) regulon by phosphorylation of the response‐regulator PhoB in Escherichia coil One pathway responds to the extracellular inorganic phosphate (Pi) level and leads to activation by the Pi sensor kinase, PhoR. The other two pathways are Pi independent and are apparent in the absence of PhoR. One Pi‐independent pathway responds to the level of an unknown catabolite and leads to activation by the catabolite regulatory sensor kinase, CreC (originally called PhoM); the other Pi‐independent pathway responds to acetyl phosphate and leads to activation by a process requiring acetyl phosphate. Here we show that activation of PhoB by acetyl phosphate can require the sensor kinase EnvZ. Accordingly, we propose that the in vivo activation of PhoB by acetyl phosphate (and perhaps other two‐component response‐regulators as well) probably always requires a certain kinase that can vary depending upon the growth conditions.
Virulence and vaccine potential ofSalmonella typhlmuriummutants deficient in the expression of the RpoS (σs) regulon
Authors: Coynault Colette , Robbe‐Saule Véronique , Norel Françoise 
Published: October 2006

SummaryThe alternative sigma factor RpoS (σs) is required forSalmonellavirulence in mice. We report the immunizing capacity ofSalmonella typhimurlum rpoSandrpoS aroAmutants to protect susceptible BALB/c mice against subsequent oral challenge with virulent S.typhimurium.When administered orally or intraperitoneally,rpoSderivatives of the mouse‐virulent S.typhimuriumstrains, C52 and SL1344, were highly attenuated and were efficient single‐dose live vaccines.rpoS aroAmutants were more attenuated than corresponding singlearoAorrpoSmutants, as assessed after oral or intraperitoneal administration, but retained significant ability to protect mice against salmonellosis.Salmonella rpoSandrpoS aroAmutants therefore deserve serious consideration for rational vaccine design. Consistent with this,Salmonella typhiTy2, a ‘wild‐type’ strain used widely for the development of human live‐vaccine candidates against typhoid fever, was shown to be defective forrpoS.In addition, our results demonstrate thatrpoSnot only controls the growth and persistence of S.typhimuriumin deep lymphoid organs, but also plays a role during the initial stages of oral infection.
Measurement and Significance of 3‐Nitrotyrosine in Systemic Lupus Erythematosus
Authors: Khan F. , Siddiqui A. A. , Ali R. 
Published: October 2006

AbstractNitration of free and protein associated tyrosine represents, in vivo, a mechanism that can severely compromise the cell function. The detection of 3‐nitrotyrosine (3‐NT) in pathological tissues is suggestive of the occurrence of nitrating pathways and has been identified as a marker of inflammation and a stable end product of increased reactive nitrogen intermediate production. Protein nitration occurs in many disease conditions including systemic lupus erythematosus (SLE). In this study we show that the level of both free and protein bound 3‐NT, which is produced by reactive nitrogen species (RNS)‐dependent oxidative damage, is elevated in patients with SLE and that there is a possible role of RNS‐modified epitopes in the aetiology of the disease. Commercially available poly L‐tyrosine was exposed to nitrating species, inducing nitration in tyrosine residues. Immunoglobulin‐G (IgG) purified on Protein‐A‐Sepharose matrix from 24 SLE patients was studied for their recognition of native and nitrated poly L‐tyrosine by direct binding and competition enzyme‐linked immunosorbent assay (ELISA). The formation of immune complex between SLE IgG and nitrated poly L‐tyrosine was visualized by gel retardation assay. Free 3‐NT in patients’ sera was detected and quantitated by high performance liquid chromatography whereas protein‐bound 3‐NT was analysed by Western blotting and the concentration was calculated by sandwich ELISA. The concentration of free 3‐NT was found to be 1.4 ± 0.09 μm whereas the concentration of protein bound 3‐NT was 96.52 ± 21.12 μm nitrated bovine serum albumin equivalents/mg protein, which was significantly higher when compared with healthy controls. Elevated level of 3‐NT was observed in SLE patients using two different techniques, when compared with healthy subjects confirms the overproduction of RNS in the pathogenesis of human SLE.
Impact of Influenza Vaccine Formulation with a Detailed Analysis of the Cytokine Response
Authors: Szyszko E. , Brokstad K. , Cox R. J. , Hovden A.‐O. , Madhun A. , Haaheim L. R. 
Published: October 2006

AbstractVaccination provides the most effective method of limiting the impact of influenza. Inactivated influenza vaccines are available in three formulations and more information needs to be generated on how antigen presented in different vaccine formulations influences the subsequent immune response. In the present study, we have investigated the effect of two different influenza vaccine formulations on the resulting antibody and cytokine responses and compared these responses with influenza infection. Mice were vaccinated intramuscularly with one or two doses of whole or split virus vaccine or alternatively intranasally infected with influenza virus. Lymphocytes were isolated from spleen cells and stimulatedin vitrofor 24 or 72 h for analysis of cytokine profile at the gene expression and at the protein level. Additionally, whole blood was collected and the serum antibody response investigated by haemagglutination inhibition (HI) and enzyme‐linked immunosorbent assay (ELISA). We found that one dose of whole virus vaccine induced higher antibody and cytokine responses and thus was more immunogenic in unprimed mice than split virus vaccine. Whole virus vaccine induced a strong IFN‐γ(type 1) immune response after one dose of vaccine and a more mixed cytokine response after the second dose. Split virus vaccine induced a type 2 response, particularly after two vaccine doses. Our results show that two doses of vaccine (both vaccine formulation) were more effective in induction of Th2 type of cytokines and thus indicate that both the formulation and also the number of vaccine doses substantially influences the magnitude and quality of the immune response.
Modulation of Chemokine Gene Expression in CD133+ Cord Blood‐Derived Human Mast Cells by Cyclosporin A and Dexamethasone
Authors: Holm M. , Kvistgaard H. , Dahl C. , Andersen H. B. , Hansen T. K. , Schiøtz P. O. , Junker S. 
Published: October 2006

AbstractWe have recently developed a protocol for generating huge numbers of mature and functional mast cells from in vitro differentiated umbilical cord blood cells. Using CD133 as a positive selection marker to isolate haematopoietic progenitors we routinely expand the number of recovered cells at least 150‐fold, which vastly exceeds the yields of conventional protocols using CD34+ cells as a source of progenitors. Taking advantage of the large quantities of in vitro differentiated mast cells, here we assess at the levels of transcription and translation the kinetics of chemokine gene induction following receptor mediated mast cell activation or following pharmacological activation of specific signal transduction cascades that become activated upon classical FcɛRI receptor crosslinking. We demonstrate that chemokine genes encoding IL‐8, MCP‐1, MIP‐1α, and MIP‐1β are induced with different kinetics and with different amplitudes in a receptor activation dependent manner, and that these events can be mimicked using pharmacological agents which activate distinct signal transduction pathways. These findings were corroborated by adding immunomodulators such as cyclosporin A and dexamethasone prior to mast cell activation. Finally, we demonstrate that the same modulators added after mast cell activation can differentially quench ongoing chemokine gene induction. Thus, considering the vast yields of mast cells, our protocol is valuable not only for studying regulation of gene expression in mast cells in general, but also as an experimental tool to develop better and more balanced treatments of mast cell related disorders.
Immune Regulatory T Cells in Siblings of Children Suffering from Type 1 Diabetes Mellitus
Authors: Michalek J. , Vrabelova Z. , Hrotekova Z. , Kyr M. , Pejchlova M. , Kolouskova S. , Faresjö M. , Stechova K. 
Published: October 2006

AbstractPatients with type 1 diabetes are suffering from defects in immune regulatory cells. Their siblings may be at increased risk of type 1 diabetes especially if they are carriers of certain human leucocyte antigen (HLA) alleles. In a prospective non‐randomized study, we intended to evaluate 31 healthy siblings of paediatric patients with type 1 diabetes and explore immune regulatory populations of CD4+CD25+ T cells and natural killer (NK) T cells. Tested siblings of type 1 diabetes patients were stratified according to the HLA‐associated risk of possible diabetes development. Immune regulatory function of CD4+CD25+ T cells was tested in vitro. Significant differences in CD4+CD25+ but not in NK T cells have been identified. Siblings of type 1 diabetes patients carrying high risk HLA alleles (DQA1*05, DQB1*0201, DQB1*0302) had significantly lower number of immune regulatory CD4+CD25+ T cells than the age‐matched healthy controls or siblings carrying low‐risk HLA alleles (DQB1*0301, DQB1*0603, DQB1*0602). Regulatory function of CD4+CD25+ T cells demonstrated a dose‐escalation effect. In siblings of type 1 diabetes patients, the defect in immune regulatory CD4+CD25+ T cells exists in association with genetic HLA‐linked risk for type 1 diabetes.
Benznidazole Treatment during Early‐indeterminate Chagas’ Disease Shifted the Cytokine Expression by Innate and Adaptive Immunity Cells toward a Type 1‐modulated Immune Profile
Authors: Sathler‐Avelar R. , Vitelli‐Avelar D.M. , Massara R.L. , Borges J.D. , Lana M. , Teixeira‐Carvalho A. , Dias J.C.P. , Elói‐Santos S.M. , Martins‐Filho O.A. 
Published: October 2006

Abstract Trypanosoma cruzi‐infected children was treated with benznidazole (Bz) during the early‐indeterminate disease (E‐IND) and the cytokine pattern of innate and adaptive immune compartments were evaluated prior to the treatment and 1 year after it. At first, we observed that the ex vivo cytokine profile of circulating leukocytes from E‐IND (n = 6) resembled the one observed for healthy schoolchildren (n = 7). Additionally, in vitro stimulation with T. cruzi antigens drove the E‐IND cytokine pattern toward a mixed immune profile with higher levels of IFN‐γ+, TNF‐α+ and IL‐4+ NK cells, increased numbers of IFN‐γ+, TNF‐α+ and IL‐10+ CD4+ T cells in addition to enhanced frequency of TNF‐α+/IL‐4+ CD19+ lymphocytes. Interestingly, upon T. cruzi antigen in vitro stimulation, E‐IND CD8+ lymphocytes displayed a selective enhancement of IFN‐γ expression, accounting for a global type 1‐modulated cytokine microenvironment. A shift toward a type 1‐modulated profile was also the hallmark of Bz‐treated children (E‐INDT). In this context, despite the mixed overall ex vivo cytokine profile observed for NK and CD8+ T cells, increased ability of these leukocytes to produce IFN‐γ in response to T. cruzi antigens was reported. Most noteworthy was the IL‐10 production evidenced at T lymphocytes, mainly CD4+ cells, as well as B lymphocytes, both ex vivo and upon antigen stimulation. Together, these findings gave evidence that NK cells and CD8+ T lymphocytes are the major sources of IFN‐γ, a pivotal cytokine for successful therapeutic response in human Chagas’ disease. Moreover, our data have also brought additional information, pointing out IL‐10 production by CD4+ cells and B lymphocytes, as the putative key element for parasite clearance in the absence of deleterious tissue damage.
Pre‐attack stress‐load, appraisals, and coping in children's responses to the 9/11 terrorist attacks
Authors: Lengua Liliana J. , Long Anna C. , Meltzoff Andrew N. 
Published: October 2006

Background:  Appraisal and coping following a disaster are important factors in children's post‐traumatic stress (PTS) symptoms. However, little is known about predictors of disaster coping responses. This study examined stress‐load, appraisals and coping styles measured prior to the September 11 terrorist attacks as predictors of 9/11‐specific appraisals, coping and PTS.Methods:  A community sample of children and parents (N = 143) participating in an ongoing study were interviewed by phone approximately 1 month following 9/11.Results:  Pre‐attack stress‐load, appraisal and coping styles predicted children's 9/11‐specific appraisals, coping, and PTS. 9/11‐specific threat appraisals and avoidant coping predicted higher PTS and mediated the effects of pre‐attack stress‐load and threat appraisal.Conclusions:  Pre‐disaster stress‐load, appraisal and coping styles predict disaster‐specific appraisal and coping, which in turn, contribute to PTS. Coping interventions might mitigate PTS symptoms following a disaster.
Efficacy of Interpersonal Psychotherapy‐Adolescent Skills Training: an indicated preventive intervention for depression
Authors: Young Jami F. , Mufson Laura , Davies Mark 
Published: October 2006

Background:  Indicated interventions for adolescents with elevated depressive symptoms may help decrease rates of depression. The current study reports on the efficacy of Interpersonal Psychotherapy‐Adolescent Skills Training (IPT‐AST), a group indicated preventive intervention.Methods:  Forty‐one adolescents with elevated depression symptoms were randomized to receive either IPT‐AST or school counseling (SC) as delivered by guidance counselors and social workers. Adolescents in the two intervention conditions were compared on depression symptoms, overall functioning, and depression diagnoses post‐intervention and at 3‐month and 6‐month follow‐up.Results:  Adolescents who received IPT‐AST had significantly fewer depression symptoms and better overall functioning post‐intervention and at follow‐up. Adolescents in IPT‐AST also reported fewer depression diagnoses than adolescents in usual care.Conclusions:  These results provide preliminary evidence of the efficacy of IPT‐AST as an intervention for adolescents with subthreshold depression. Future research is needed to confirm the efficacy of IPT‐AST in a larger and more diverse sample and to determine its long‐term impact on depression symptoms and depression diagnoses.
Selective Loss of N‐Methyl‐d‐Aspartate‐Sensitive l‐[3H]Glutamate Binding Sites in Rat Brain Following Portacaval Anastomosis
Authors: Peterson Christine , Giguere Jean‐Francois , Cotman Carl W. , Butterworth Roger F. 
Published: October 2006

AbstractExcitatory amino acids have been implicated in the pathogenesis of hepatic encephalopathy. In the present study, kainate, quisqualate and N‐methyl‐d‐aspartate (NMDA) subclasses of l‐glutamate receptors were measured in adult rat brain by quantitative receptor autoradiography following surgical construction of an end‐to‐side portacaval anastomosis (PCA). PCA resulted in sustained hyperammonemia and decreased binding of l‐glutamate to the NMDA receptor when compared to sham‐operated controls. Decreases in binding ranged from 17 to 39% in several regions of cerebral cortex, hippocampus, striatum, and thalamus. Binding to quisqualate and kainate receptor subtypes was not altered. PCA leads to astrocytic changes in brain but does not result in any measurable loss of neuronal integrity. It is therefore proposed that decreased glutamate binding to the NMDA receptor following PCA results from increased extracellular glutamate caused by decreased reuptake into perineuronal astrocytes and a compensatory down‐regulation of these receptors. Such changes could be of pathophysiological significance in hepatic encephalopathy.
Phorbol Esters Induce Neurotransmitter Release in Cholinergic Synaptosomes from Torpedo Electric Organ
Authors: Guitart Xavier , Marsal Jordi , Solsona Carles 
Published: October 2006

AbstractThe effect of phorbol esters and so the involvement of Ca2+/phospholipid‐dependent protein kinase (protein kinase C; PKC) in the release of acetylcholine (ACh) was studied using Torpedo electric organ synaptosomes. 12‐O‐Tetradecanoylphorbol 13‐acetate (TPA), a known activator of PKC, induced neurotransmitter release in a concentration‐dependent manner and increased the potassium‐evoked release of ACh. The effect of TPA was shown to be independent of the extrasynaptosomal calcium concentration. TPA‐induced ACh release was reversed by H‐7, an inhibitor of PKC activity. This drug showed no effect on potassium‐evoked ACh release. Botulinum toxin, a strong blocker of potassium‐induced ACh release in that synaptosomal preparation, showed no inhibitory effect on the TPA‐induced ACh release. Our results suggest that activation of PKC potentiates the release of an ACh pool that is not releasable by potassium depolarization, independently of the extracellular calcium concentration.
Characterization of Functional Neuropeptide Y Receptors in a Human Neuroblastoma Cell Line
Authors: Gordon Eric A , Kohout Trudy A. , Fishman Peter H. 
Published: October 2006

AbstractWe identified receptors for neuropeptide Y (NPY) on an established human neuroblastoma cell line, SK‐N‐MC, which are functionally coupled to adenylate cyclase through the inhibitory guanine nucleotide‐binding protein of adenylate cyclase, Gi Intact SK‐N‐MC cells bound radiolabeled NPY with a KD of 2 nM and contained ˜83,000 receptors/cell. Unlabeled porcine and human NPY and structurally related porcine peptide YY (PYY) competed with labeled NPY for binding to the receptors. NPY inhibited cyclic AMP accumulation in SK‐N‐MC cells stimulated by isoproterenol, dopamine, vasoactive intestinal peptide, cholera toxin, and forskolin. NPY inhibited isoproterenol‐stimulated cyclic AMP production in a dose‐dependent manner, with half‐maximal inhibition at 0.5 nM NPY. Porcine and human NPY and porcine PYY gave similar dose‐response curves. NPY also inhibited basal and isoproterenol‐stimulated adenylate cyclase activity in disrupted cells. Pertussis toxin treatment of the cells completely blocked the ability of NPY to inhibit cyclic AMP production and adenylate cyclase activity. The toxin catalyzed the ADP‐ribosylation of a 41‐kDa protein in SK‐N‐MC cells that corresponds to Gi. The receptors on SK‐N‐MC cells appeared to be specific for NPY, as other neurotransmitter drugs, such as α‐adrenergic, dopaminergic. muscarinic, and serotonergic antagonists, did not compete for either NPY binding or NPY inhibition of adenylate cyclase. Thus, SK‐N‐MC cells may be a useful model for investigating NPY receptors and NPY‐mediated signal trans‐duction.
Aluminum Uptake by Neuroblastoma Cells
Authors: Shi Biao , Haug Alfred 
Published: October 2006

AbstractAluminum uptake studies in viable neuroblastoma cells were performed. Aluminum uptake was largely dependent on the pH of the suspension medium. At physiological pH values, cells were apparently unable to incorporate detectable amounts of aluminum in the absence of proper mediators. Aluminum uptake was enhanced as the pH decreased, attaining a plateau at about pH 6.0. In experiments with 2 × 106 cells/ml, pH 6.0, and 25 μM aluminum in the medium, aluminum incorporation reached saturation at 5 nmol of aluminum/mg of cellular protein, accounting for 60–70% of aluminum added. At pH 6.0, cells showed a large capacity for accumulating aluminum; about 70% of intracellular aluminum was associated with the post‐mitochondrial fraction. At neutral pH, application of apotransferrin seemed to facilitate aluminum translocation into cells via membrane receptors. Fatty acids were also capable of mediating aluminum uptake at neutral pH, probably by forming aluminum‐fatty acid complexes. Low molecular weight aluminum chelators, e.g., citrate, inhibited aluminum uptake. Treatment of cells with energy metabolism blockers had virtually no influence on aluminum uptake, indicative of passive mechanisms. The results suggest that aluminum uptake occurs via different modes dependent on growth conditions, such as medium pH.
1‐Methyl‐4‐Phenyl‐1,2,3,6‐Tetrahydropyridine‐Resistant, Flat‐Cell PC12 Variants Having a Partial Loss of Transformed Phenotype
Authors: Andersen Julie K. , Zhang Mao‐Bin , Zhong Xiao‐Hui , Rozenberg Yanina Y. , Howard Bruce D. 
Published: October 2006

AbstractWe have cloned and characterized two variants of PC12 cells. MPT1 cells were selected by their resistance to 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP), and variant 2068 was isolated nonselectively as a large, flat‐cell variant commonly occurring in PC12 cultures. Variant 2068 cells also exhibit resistance to MPTP. Karyotype analysis revealed that these variants are true derivatives of wild‐type PC12 cells; however, each variant is tetraploid, whereas the wild‐type parent is diploid. The two variants contain an altered level and composition of lactate dehydrogenase isoenzymes, which could account for a previously described difference in lactate metabolism. Both variants exhibit a partial loss of transformed phenotype in culture in that they are nonrefractile, grow in monolayers, and fail to multiply in soft agar. We suggest that this alteration in transformed phenotype may result in altered mitochondria and lactate dehydrogenase and thus account for their resistance to MPTP. Compared with wild‐type PC12 cells, MPT1 cells have a decreased level of fos mRNA and an increased level of myc mRNA; the latter results from an increased level of transcription of exon 1 of the myc gene. Studies with hybrid cells obtained by fusing MPT1 cells with wild‐type‐like cells show that most, but not all, of the parameters of the MPT1 phenotype predominate.