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Ultrasound assisted regioselective sulfonation of aromatic compounds with sulfuric acid.
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A simple and convenient methodology for selective sulfonation of aromatic compounds using sulfuric acid under sonication is described. The present methodology shows a considerable enhancement in the reaction rate along with improved selectivity compared with the reactions performed under silent conditions. The effect of various parameters such as agitation speed, sulfuric acid concentration, and temperature on reaction system have been investigated and are explained on the basis of ultrasonically generated cavitational effects.
| 19,014,895
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The formation of argpyrimidine, a methylglyoxal-arginine adduct, in the nucleus of neural cells.
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Methylglyoxal (MG) is an endogenous metabolite in glycolysis and forms stable adducts primarily with arginine residues of intracellular proteins. The biological role of this modification in cell function is not known. In the present study, we found that a MG-detoxification enzyme glyoxalase I (GLO1) is mainly expressed in the ventricular zone (VZ) at embryonic day 16 which neural stem and progenitor cells localize. Moreover, immunohistochemical analysis revealed that argpyrimidine, a major MG-arginine adduct, is predominantly produced in cortical plate neurons not VZ during cerebral cortex development and is exclusively located in the nucleus. Immunoblotting experiment showed that the formation of argpyrimidine occurs on some nuclear proteins of cortical neurons. To our knowledge, this is first report of the argpyrimidine formation in the nucleus of neuron. These findings suggest that GLO1, which is dominantly expressed in the embryonic VZ, reduces the intracellular level of MG and suppresses the formation of argpyrimidine in neural stem and progenitor cells. Argpyrimidine may contribute to the neural differentiation and/or the maintenance of the differentiated state via the modification of nuclear proteins.
| 19,014,907
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Aldo-keto reductases from the AKR1B subfamily: retinoid specificity and control of cellular retinoic acid levels.
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NADP(H)-dependent cytosolic aldo-keto reductases (AKRs) have been added to the group of enzymes which contribute to oxidoreductive conversions of retinoids. Recently, we found that two members from the AKR1B subfamily (AKR1B1 and AKRB10) were active in the reduction of all-trans- and 9-cis-retinaldehyde, with K(m) values in the micromolar range, but with very different k(cat) values. With all-trans-retinaldehyde, AKR1B10 shows a much higher k(cat) value than AKR1B1 (18 min(-1)vs. 0.37 min(-1)) and a catalytic efficiency comparable to that of the best retinaldehyde reductases. Structural, molecular dynamics and site-directed mutagenesis studies on AKR1B1 and AKR1B10 point that subtle differences at the entrance of their retinoid-binding site, especially at position 125, are determinant for the all-trans-retinaldehyde specificity of AKR1B10. Substitutions in the retinoid cyclohexene ring, analyzed here further, also influence such specificity. Overall it is suggested that the rate-limiting step in the reaction mechanism with retinaldehyde differs between AKR1B1 and AKR1B10. In addition, we demonstrate here that enzymatic activity of AKR1B1 and AKR1B10 lowers all-trans- and 9-cis-retinoic acid-dependent trans-activation in living cells, indicating that both enzymes may contribute to pre-receptor regulation of retinoic acid and retinoid X nuclear receptors. This result supports that overexpression of AKR1B10 in cancer (an updated review on this topic is included) may contribute to dedifferentiation and tumor development.
| 19,014,918
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Epidemiological association between fasting plasma glucose and shortened APTT.
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To investigate the relationship between fasting plasma glucose (FPG) and shortened activated partial thromboplastin time (APTT), an independent risk factor for thrombosis. We analyzed outpatients' results of coagulation tests and FPG. When compared with euglycemic subjects, those with impaired fasting glucose and diabetes displayed significantly shortened APTTs. This previously unreported association deserves scrutiny, because APTT is relatively inexpensive and it might identify diabetic patients at major risk of thrombosis.
| 19,014,926
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Negative elongation factor NELF controls transcription of immediate early genes in a stimulus-specific manner.
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The transcription rate of immediate early genes (IEGs) is controlled directly by transcription elongation factors at the transcription elongation step. Negative elongation factor (NELF) and 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) sensitivity-inducing factor (DSIF) stall RNA polymerase II (pol II) soon after transcription initiation. Upon induction of IEG transcription, DSIF is converted into an accelerator for pol II elongation. To address whether and how NELF as well as DSIF controls overall IEG transcription, its expression was reduced using stable RNA interference in GH4C1 cells. NELF knock-down reduced thyrotropin-releasing hormone (TRH)-induced transcription of the IEGs c-fos, MKP-1, and junB. In contrast, epidermal growth factor (EGF)-induced transcription of these IEGs was unaltered or even slightly increased by NELF knock-down. Thus, stable knock-down of NELF affects IEG transcription stimulation-specifically. Conversely, DSIF knock-down reduced both TRH- and EGF-induced transcription of the three IEGs. Interestingly, TRH-induced activation of the MAP kinase pathway, a pathway essential for transcription of the three IEGs, was down-regulated by NELF knock-down. Thus, stable knock-down of NELF, by modulating intracellular signaling pathways, caused stimulation-specific loss of IEG transcription. These observations indicate that NELF controls overall IEG transcription via multiple mechanisms both directly and indirectly.
| 19,014,935
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Orientobilharzia turkestanicum is a member of Schistosoma genus based on phylogenetic analysis using ribosomal DNA sequences.
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In the present study, samples representing Orientobilharzia turkestanicum from cattle, sheep, cashmere goat and goat in Heilongjiang Province, China, were characterized and grouped genetically by sequences of internal transcribed spacer (ITS, including ITS-1 and ITS2) and 28S ribosomal DNA (28S rDNA). The ITS and 28S rDNA were amplified by polymerase chain reaction (PCR) and then sequenced and compared with that of other members of the Schistosomatidae available in GenBank, and phylogenetic relationships between them were re-constructed using the neighbor-joining and maximum parsimony methods. The lengths of ITS-1, ITS-2 and 28S rDNA sequences for all O. turkestanicum samples from different hosts were 384bp, 331bp and 1304bp, respectively. While the ITS-1 sequences of O. turkestanicum from each of the four different hosts, and ITS-2 of O. turkestanicum from cattle, sheep and cashmere goat were identical, respectively, the ITS-2 of O. turkestanicum from goat differed from that of O. turkestanicum from cattle, sheep and cashmere goat by one nucleotide. The 28S rDNA sequences of O. turkestanicum from sheep and cashmere goat were identical, but differed from that of O. turkestanicum from cattle and goat by two nucleotides, with the latter two also having identical 28S rDNA sequence. Phylogenetic analyses based on the combined sequences of the ITS-1 and ITS-2, or the 28S rDNA sequences placed O. turkestanicum within the genus Schistosoma, and it was phylogenetically closer to the African schistosome group than to the Asian schistosome group. These results should have implications for studying the origin and evolution of O. turkestanicum and other members of the Schistosomatidae.
| 19,014,940
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Presentation and long-term follow-up of refractory celiac disease: comparison of type I with type II.
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Refractory celiac disease (RCD) was recently subdivided into 2 subtypes (RCD I and II) based on a normal or abnormal phenotype of intraepithelial lymphocytes (IELs), respectively. It is not clear, however, if these 2 entities differ in their presentation at diagnosis or long-term outcome. We compared the clinical and biological characteristics of RCD I and RCD II at diagnosis, the risk of developing an overt lymphoma, and the predictive factors of survival. Medical files of 14 patients with RCD I and 43 with RCD II were analyzed retrospectively. Predictive factors of overt lymphoma and survival were studied in univariate and multivariate analyses. At diagnosis, malnutrition, ulcerative jejunitis, and lymphocytic gastritis were more common in patients with RCD II than RCD I (P< .05). Overt lymphomas occurred in 2 patients with RCD I and 16 with RCD II. In the univariate analysis, abnormal IEL phenotype and increased age at diagnosis of RCD were predictive factors for overt lymphoma. Abnormal IEL phenotype (P< .01), clonality (P= .01), and overt lymphoma (P= .001) predicted short survival time. Only abnormal IEL phenotype (P= .03) and overt lymphoma (P= .04) were predictive in the multivariate analysis. The 5-year survival rate was 93% in patients with RCD I and 44% with RCD II. RCD II has a much more severe presentation and prognosis than patients with RCD I; <44% of patients with RCD II survive 5 years after diagnosis. Abnormal IEL phenotype is a predictive factor but not a necessary condition for the development of overt lymphoma.
| 19,014,942
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SIFamide illustrates the rapid evolution in Arthropod neuropeptide research.
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This review is focussed on SIFamide. This neuropeptide was discovered as a result of an extensive purification process, typical for 20th century physiology, of an extract of 350,000 flesh flies. Our knowledge of SIFamide greatly expanded since the first publication in 1996. Describing the minor and major findings on this peptide is our lead to summarise a number of innovations that recently became common in research on Arthropods. Mass spectrometry, nanoLC, whole mount immunocytochemistry, genome sequencing, deorphanizing receptors and functional gene knock downs are aspects that dramatically improved and changed peptide research. Some of the techniques mentioned in this review were of course applied before 1996, but they were not widespread. Although the focus of the review is on insects we incorporated the data of SIFamide in Crustaceans as well. SIFamide illustrates that crustaceans and insects might have more in common than was previously anticipated. Today, six isoforms of SIFamide are discovered in many crustaceans, several insects and a tick. The sequence of SIFamide is extremely conserved among these species. Deorphanizing its receptor in Drosophila, learned that both the ligand and receptor are impressively conserved, pointing at a crucial function. Immunohistochemistry and mass spectrometry data reveal that SIFamide is present in the crustacean brain and gut, but restricted to four neurons in the insect pars intercerebralis. The immunoreactive patterns in the brain refer to a neuromodulatory role in combining visual, tactile and olfactory input. Eventually, targeted cell ablation and RNAi revealed that SIFamide modulates sexual behaviour in fruit flies.
| 19,014,945
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Active school transport, physical activity levels and body weight of children and youth: a systematic review.
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Active school transport (AST) may be an important source of children's physical activity (PA). Innovative solutions that increase PA time for children, without putting added pressure on the school curriculum, merit consideration. Before implementing such solutions, it is important to demonstrate that active school transport is associated with health-related outcomes. Following a standardized protocol, we conducted a systematic review of published research to address this question and explore whether children who actively commute to school also have a healthier body weight. Online searches of 5 electronic databases were conducted. Potential studies were screened on the basis of objective measures of physical activity. Thirteen studies were included in this review. Nine studies demonstrated that children who actively commute to school accumulate significantly more PA and two studies reported that they expended significantly more kilocalories per day. Where studies examined body weight (n=10), only one reported active commuters having a lower body weight. These studies demonstrate that active school commuters tend to be more physically active overall than passive commuters. However, evidence for the impact of AST in promoting healthy body weights for children and youth is not compelling.
| 19,014,963
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In-tube transfection improves the efficiency of gene transfer in primary neuronal cultures.
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To facilitate genetic studies in primary neurons, we analyzed the efficiency of cationic lipid-mediated plasmid DNA transfection using adherent and acutely dissociated neuronal suspensions derived from embryonic mouse cortical tissue. Compared to transfections using adherent cultures, the in-tube procedure enhanced the delivery of a GFP reporter plasmid between four- to eightfold depending on the age of the harvested embryo. The procedure required relatively brief complex incubation times, and supported the transfection of cells expressing the neuronal markers NeuN and TuJ1 with improved uniformity in transfection events across the well surface. To demonstrate the utility of this approach in studying the genetic mechanisms controlling neuron development, we provide data regarding the role of the bZIP transcription factor c/EBP-beta in regulating neurite outgrowth. It is anticipated that this in vitro protocol will facilitate the identification of novel genes involved in both developmental and disease-relevant signaling pathways.
| 19,014,969
|
Chronic alcohol treatment in rats alters sleep by fragmenting periods of vigilance cycling in the light period with extended wakenings.
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Studies have shown that disturbed sleep produced by chronic alcohol abuse in humans can predict relapse drinking after periods of abstinence. How alcohol produces disturbed sleep remains unknown. In this study we used a novel analysis of sleep to examine the effects of alcohol on sleep patterns in rats. This analysis separates waking into multiple components and defines a period labeled vigilance cycling (VC) in which the rat rapidly cycles through various vigilance states. These VC episodes are separated by long duration wake (LDW) periods. We find that 6 weeks of alcohol (6% in a liquid diet) caused fragmentation of extended VC episodes that normally occur in the light period. However, total daily amounts of slow-wave sleep (SWS) and rapid-eye movement sleep (REMS) remained constant. The daily amount of wake, SWS, and REMS remained constant because the alcohol treated rats increased the amount of VC in the dark period, and the sleep nature of VC in the dark period became more intense. In addition, we observed more wake and less REMS early in the light period in alcohol treated rats. All effects completely reversed by day 16 of alcohol withdrawal. Comparison of the effects of chronic alcohol to acute alcohol exposure demonstrated the effects of chronic alcohol are due to adaptation and not the acute presence of alcohol. The effects of chronic alcohol treatment in rats mimic the effects reported in humans (REMS suppression, difficulty falling asleep, and difficulty remaining asleep).
| 19,014,977
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Reactive oxygen species regulate properties of transformation in UROtsa cells exposed to monomethylarsonous acid by modulating MAPK signaling.
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UROtsa cells exposed to 50 nM monomethylarsonous acid [MMA(III)] for 52 wk (MSC52) achieved hyperproliferation, anchorage independent growth, and enhanced tumorgenicity. MMA(III) has been shown to induce reactive oxygen species (ROS), which can lead to activation of signaling cascades causing stress-related proliferation of cells and even cellular transformation. Previous research established the acute activation of MAPK signaling cascade by ROS produced by MMA(III) as well as chronic up regulation of COX-2 and EGFR in MSC52 cells. To determine if ROS played a role in the chronic pathway perturbations by acting as secondary messengers, activation of Ras was determined in UROtsa cells [exposed to MMA(III) for 0-52 wk] and found to be increased through 52 wk most dramatically after 20 wk of exposure. Ras has been shown to cause an increase in O2(-) and be activated by increases in O2(-), making ROS important to study in the transformation process. COX-2 upregulation in MSC52 cells was confirmed by real time RT-PCR. By utilizing both antioxidants or specific COX inhibitors, it was shown that COX-2 upregulation was dependent on ROS, specifically, O2(-). In addition, because previous research established the importance of MAPK activation in phenotypic changes associated with transformation in MSC52 cells, it was hypothesized that ROS play a role in maintaining phenotypic characteristics of the malignant transformation of MSC52 cells. Several studies have demonstrated that cancer cells have lowered superoxide dismutase (MnSOD) activity and protein levels. Increasing levels of MnSOD have been shown to suppress the malignant phenotype of cells. SOD was added to MSC52 cells resulting in slower proliferation rates (doubling time=42h vs. 31h). ROS scavengers of OH also slowed proliferation rates of MSC52 cells. To further substantiate the importance of ROS in these properties of transformation in MSC52 cells, anchorage independent growth was assessed after the addition of antioxidants, both enzymatic and non-enzymatic. Scavengers of OH, and O2(-) blocked the colony formation of MSC52 cells. These data support the role for the involvement of ROS in properties of transformation of UROtsa cells exposed to MMA(III).
| 19,014,992
|
Molecular cloning, characterization and expression of two tryptophan hydroxylase (TPH-1 and TPH-2) genes in the hypothalamus of Atlantic croaker: down-regulation after chronic exposure to hypoxia.
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Recently we discovered that hypoxia causes marked impairment of reproductive neuroendocrine function in Atlantic croaker, a marine teleost, which is due to a decline in hypothalamic serotonergic activity. As a first step in understanding the molecular responses of the hypothalamic serotonergic system to hypoxia, we cloned and characterized the genes for the enzymes regulating the rate-limiting step in serotonin biosynthesis, tryptophan hydroxylase (TPH-1 and TPH-2) in the croaker brain. The full-length croaker TPH-1 and TPH-2 cDNAs contain open reading frames encoding proteins with 479 and 487 amino acids, respectively, which are highly homologous to the TPH-1 (76-93%) and TPH-2 (64-92%) proteins of other vertebrates. Croaker TPH-1 and TPH-2 mRNA expression was detected throughout the brain but was greatest in the hypothalamic region. Both Northern blot analysis and real-time PCR showed that TPH-1 (transcript size approximately 2.1 kb) and TPH-2 ( approximately 1.9 kb) mRNA levels were significantly decreased in the hypothalami of croaker exposed for 2 weeks to hypoxic conditions compared with those in fish exposed to normoxic conditions. Immunohistochemistry of hypothalamic neurons with TPH antibodies showed reduced expression of TPHs in hypoxia-exposed fish compared with normoxic fish. Western blot analysis confirmed that hypoxia caused a marked decline in hypothalamic TPH protein levels, which was associated with decreases in hypothalamic TPH enzyme activity and 5-hydroxytryptophan levels. These results suggest that TPH is a major site of hypoxia-induced down-regulation of serotonergic function in croaker brains. Moreover, they provide the first evidence that hypoxia decreases the expression of TPH transcripts in vertebrate brains.
| 19,015,006
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No elevated genomic damage in children and adolescents with attention deficit/hyperactivity disorder after methylphenidate therapy.
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Attention-deficit/hyperactivity disorder (ADHD) is the most frequent psychiatric disorder in children and adolescents and is often treated with methylphenidate (MPH), resulting in MPH exposure in more than 1% of all children in many countries. A 2005 report on cytogenetic effects in peripheral lymphocytes from 12 ADHD children treated for 3 months with MPH raised questions about its genetic toxicity and potential carcinogenicity. In 2007, we described no elevated micronucleus frequency in 21 children after 3 months of MPH-treatment; since the difference between the two studies could not be explained we now enlarged the overall sample size, and added a healthy control group, a new chronically treated group and positive control slides. Furthermore, micronuclei were analyzed in a second tissue, buccal mucosa. A healthy control group (23 individuals), a chronically MPH-treated (>12 months) group (21 patients), and a drug naïve group of ADHD-affected children (26 patients), which was analyzed again after 3 months (17 patients) and 6 months (11 patients), provided samples for analysis of micronucleated lymphocytes. With inclusion of 14 previously obtained blood samples, an overall group size of 31 patients was reached for the comparison of the 3 months observation time with before for micronucleated lymphocytes. For buccal mucosa cells, an additional inclusion of 10 more chronically treated patients (no lymphocytes donated) yielded sample numbers of 22 (healthy), 17 (chronically treated), 23 (ADHD drug naïve), 14 (3 months) and 11 (6 months). No significant alteration in genomic damage as seen as micronucleus frequency in peripheral lypmphocytes or buccal mucosa cells was detected after MPH treatment. No indication for genomic damage induced by MPH was obtained in this study, as in our previous study. Together with our previous study, our overall number of MPH-treated patients is now 68 (30 chronically treated, 38 prospectively followed), plus 23 healthy controls. Ongoing studies in the USA, as well as continuation of recently published epidemiological cancer incidence analysis should provide additional reassurance for MPH-treated ADHD patients.
| 19,015,014
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Morinda citrifolia Linn (Noni): in vivo and in vitro reproductive toxicology.
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Morinda citrifolia Linn (syn. Noni) is a plant widely used as food and medicine worldwide but there are no toxicological tests about this plant focused on reproduction. To investigate possible endocrine activity and toxic effect on the reproductive system of Wistar rats by exposure of aqueous extract of the Morinda citrifolia. Two experimental protocols in vivo were developed, (a) uterotrophic assay and (b) in utero and lactational assay, and one test in vitro to investigate the effect on the contractility of pregnant uteri isolated from rats (doses of the extract: 7.5, 75 and 750 mg/kg). The uterotrophic assay indicates presence of in vivo antiestrogenic activity of extract at doses of 7.5 and 750 mg/kg. The in utero and lactation exposure showed that the treatment with extract at the dose of 7.5mg/kg induced a reduction of 50% in parturition index and an increase of 74% in postimplantation losses index. The in vitro test showed that uteri from rats treated with 7.5mg/kg of the extract presented a 50% reduction on contraction induced by arachidonic acid. The exposure of aqueous extract of Morinda citrifolia in Wistar rats induced reproductive toxicity in nonlinear dose-response.
| 19,015,020
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Protection of cerulein-induced pancreatic fibrosis by pancreas-specific expression of Smad7.
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Pancreatic fibrosis is the hallmark of chronic pancreatitis, currently an incurable disease. Pancreatitis fibrosis is caused by deposition of extracellular matrix (ECM) and the underlying pathological mechanism remains unclear. In addition to its broad biological activities, TGF-beta is a potent pro-fibrotic factor and many in vitro studies using cell systems have implicated a functional role of TGF-beta in the pathogenesis of pancreatic fibrosis. We analyzed the in vivo role of TGF-beta pathway in pancreatic fibrosis in this study. Smad7, an intracellular inhibitory protein that antagonizes TGF-beta signaling, was specifically expressed in the pancreas using a transgenic mouse model. Chronic pancreatitis was induced in the mouse with repeated administration of cerulein. Smad7 expression in the pancreas was able to significantly inhibit cerulein-induced pancreatic fibrosis. Consistently, the protein levels of collagen I and fibronectin were decreased in the Smad7 transgenic mice. In addition, alpha-smooth muscle actin, a marker of activated pancreas stellate cells, was reduced in the transgenic mice. Taken together, these data indicate that inhibition of TGF-beta signaling by Smad7 is able to protect cerulein-induced pancreatic fibrosis in vivo.
| 19,015,026
|
The roles of the FGF signal in zebrafish embryos analyzed using constitutive activation and dominant-negative suppression of different FGF receptors.
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The roles of the FGF family growth factors and their receptors (FGFRs) in zebrafish embryos were examined using variously modified versions of the four FGFR genes (fgfr1-4). Constitutively active forms of all of the examined FGFRs (ca-FGFRs) caused dorsalization, brain caudalization, and secondary axis formation, indicating that the main FGF signal transduction downstream of the receptor is highly similar among FGFRs. All of the membrane-bound type of dominant-negative FGFRs (mdn-FGFRs) derived from the four fgfr genes, which interfere with endogenous FGFRs, produced posterior truncation, as previously reported in both Xenopus and zebrafish. mdn-FGFR3c had the strongest effects on embryos, progressively disrupting the posterior structure as the dose increased. At the highest dose, only the forebrain was formed. At lower doses, mdn-FGFR3c mainly suppressed the paraxial mesoderm. The co-injection of mRNA for different mdn-FGFRs and FGFs resulted in diverse suppression spectra of the respective FGFRs against FGFs. Only mdn-FGFR3c severely suppressed all of the FGFs examined. We also examined the effects of the secretory type of dominant-negative FGFRs (sdn-FGFRs), which are released from cells and trap FGF ligands. Only sdn-FGFR3c resulted in the characteristic effect of selectively disrupting the isthmic development, as well as the tailbud. The co-injection of the mRNA for sdn-FGFRs and FGFs suggested that sdn-FGFR3c inhibits FGFs of the FGF8 subfamily, which is consistent with its specific effects on development. We discuss the implications of our findings obtained in the present study.
| 19,015,027
|
Mass spectrometry-based footprinting of protein-protein interactions.
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We present a high-resolution mass spectrometric (MS) footprinting method enabling identification of contact amino acids in protein-protein complexes. The method is based on comparing surface topologies of a free protein versus its complex with the binding partner using differential accessibility of small chemical group selective modifying reagents. Subsequent MS analysis reveals the individual amino acids selectively shielded from modification in the protein-protein complex. The current report focuses on probing interactions between full-length HIV-1 integrase and its principal cellular partner lens epithelium-derived growth factor. This method has a generic application and is particularly attractive for studying large protein-protein interactions that are less amenable for crystallographic or NMR analysis.
| 19,015,031
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Rejection of pulse related artefact (PRA) from continuous electroencephalographic (EEG) time series recorded during functional magnetic resonance imaging (fMRI) using constraint independent component analysis (cICA).
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Rejection of the pulse related artefact (PRA) from electroencephalographic (EEG) time series recorded simultaneously with fMRI data is difficult, particularly during NREM sleep because of the similarities between sleep slow waves and PRA, in both temporal and frequency domains and the need to work with non-averaged data. Here we introduce an algorithm based on constrained independent component analysis (cICA) for PRA removal. This method has several advantages: (1) automatic detection of the components corresponding to the PRA; (2) stability of the solution and (3) computational treatability. Using multichannel EEG recordings obtained in a 3 T MR scanner, with and without concomitant fMRI acquisition, we provide evidence for the sensitivity and specificity of the method in rejecting PRA in various sleep and waking conditions.
| 19,015,033
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The left amygdala knows fear: laterality in the amygdala response to fearful eyes.
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The detection of threat is a role that the amygdala plays well, evidenced by its increased response to fearful faces in human neuroimaging studies. A critical element of the fearful face is an increase in eye white area (EWA), hypothesized to be a significant cue in activating the amygdala. However, another important social signal that can increase EWA is a lateral shift in gaze direction, which also serves to orient attention to potential threats. It is unknown how the amygdala differentiates between these increases in EWA and those that are specifically associated with fear. Using functional magnetic resonance imaging, we show that the left amygdala distinguished between fearful eyes and gaze shifts despite similar EWA increases whereas the right amygdala was less discriminatory. Additional analyses also revealed selective hemispheric response patterns in the left fusiform gyrus. Our data show clear hemispheric differences in EWA-based fear activation, suggesting the existence of parallel mechanisms that code for emotional face information.
| 19,015,094
|
Ventral frontal cortex in children: morphology, social cognition and femininity/masculinity.
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The ventral frontal cortex (VFC) has been shown to differ morphologically between sexes. Social cognition, which many studies demonstrate involves the VFC, also differs between sexes, with females being more adept than males. In a previous study of subregions of the VFC in our lab, in an adult population, size of the straight gyrus (SG) but not the orbitofrontal cortex (OFC), differed between sexes and correlated with better performance on a test of social cognition and with greater identification with feminine characteristics. To investigate the relationship between VFC structure and social cognition in children, VFC gray matter volumes were measured on MRIs from 37 boys and 37 girls aged 7 to 17. The VFC was subdivided into the OFC and SG. Subjects were also administered a test of social perceptiveness and a rating scale of femininity/masculinity. In contrast to our findings in adults, the SG was slightly smaller in girls than boys. In girls, but not boys, smaller SG volumes significantly correlated with better social perception and higher identification with feminine traits. No volume differences by sex or significant correlations were found with the OFC. These data suggest a complex relationship between femininity, social cognition and SG morphology.
| 19,015,107
|
Self-reflection across time: cortical midline structures differentiate between present and past selves.
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The processing of personal changes across time and the ability to differentiate between representations of present and past selves are crucial for developing a mature sense of identity. In this study, we explored the neural correlates of self-reflection across time using functional magnetic resonance imaging (fMRI). College undergraduates were asked to reflect on their own psychological characteristics and those of an intimate other, for both the present time period (i.e. at college) and a past time period (i.e. high school years) that involved significant personal changes. Cortical midline structures (CMS) were commonly recruited by the four reflective tasks (reflecting on the present self, past self, present other and past other), relative to a control condition (making valence judgments). More importantly, however, the degree of activity in CMS also varied significantly according to the target of reflection, with the ventral and dorsal medial prefrontal cortex and the posterior cingulate cortex being more recruited when reflecting on the present self than when reflecting on the past self or when reflecting on the other person. These findings suggest that CMS may contribute to differentiate between representations of present and past selves.
| 19,015,116
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AutDB: a gene reference resource for autism research.
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Recent advances in studies of Autism Spectrum Disorders (ASD) has uncovered many new candidate genes and continues to do so at an accelerated pace. To address the genetic complexity of ASD, we have developed AutDB (http://www.mindspec.org/autdb.html), a publicly available web-portal for on-going collection, manual annotation and visualization of genes linked to the disorder. We present a disease-driven database model in AutDB where all genes connected to ASD are collected and classified according to their genetic variation: candidates identified from genetic association studies, rare single gene mutations and genes linked to syndromic autism. Gene entries are richly annotated for their relevance to autism, along with an in-depth view of their molecular functions. The content of AutDB originates entirely from the published scientific literature and is organized to optimize its use by the research community. The main focus of this resource is to provide an up-to-date, annotated list of ASD candidate genes in the form of reference dataset for interrogating molecular mechanisms underlying the disorder. Our model for consolidated knowledge representation in genetically complex disorders could be replicated to study other such disorders.
| 19,015,121
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A cytoplasmic variant of the KH-type splicing regulatory protein serves as a decay-promoting factor for phosphoglycerate kinase 2 mRNA in murine male germ cells.
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Phosphoglycerate kinase 2 (PGK2) is a germ cell-specific protein whose mRNA is translationally regulated in the mammalian testis. Using RNA affinity chromatography with the 3'-untranslated region (UTR) of Pgk2 mRNA and adult testis extracts, several associated proteins including a novel isoform of the AU-rich element RNA-binding protein and KH-type splicing regulatory protein (KSRP) were identified. KSRP, a protein of approximately 75 kDa, is widely expressed in somatic and germ cells where it is primarily nuclear. In addition to the approximately 75-kDa KSRP, a approximately 52-kD KSRP, t-KSRP, is present in the cytoplasm of a subpopulation of germ cells. t-KSRP binds directly to a 93-nt sequence (designated the F1 region) of the 3'-UTR of the Pgk2 mRNA and destabilizes Pgk2 mRNA constructs in testis extracts and in transfected cells. We conclude that this testicular variant of the multifunctional nucleic acid-binding protein, KSRP, serves as a decay-promoting factor for Pgk2 mRNA in male germ cells.
| 19,015,122
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Selection of oligonucleotides for whole-genome microarrays with semi-automatic update.
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Oligonucleotide microarray probes are designed to match specific transcripts present in databases that are regularly updated. As a consequence probes should be checked every new database release. We thus developed an informatics tool allowing the semi-automatic update of probe collections of long oligonucleotides and applied it to the mouse RefSeq database. http://www.bio.espci.fr/sol/
| 19,015,129
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Prediction of translation initiation site for microbial genomes with TriTISA.
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We report a new and simple method, TriTISA, for accurate prediction of translation initiation site (TIS) of microbial genomes. TriTISA classifies all candidate TISs into three categories based on evolutionary properties, and characterizes them in terms of Markov models. Then, it employs a Bayesian methodology for the selection of true TIS with a non-supervised, iterative procedure. Assessment on experimentally verified TIS data shows that TriTISA is overall better than all other methods of the state-of-the-art for microbial genome TIS prediction. In particular, TriTISA is shown to have a robust accuracy independent of the quality of initial annotation. The C++ source code is freely available under the GNU GPL license via http://mech.ctb.pku.edu.cn/protisa/TriTISA.
| 19,015,130
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Fish oil, but not flaxseed oil, decreases inflammation and prevents pressure overload-induced cardiac dysfunction.
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Clinical studies suggest that intake of omega-3 polyunsaturated fatty acids (omega-3 PUFA) may lower the incidence of heart failure. Dietary supplementation with omega-3 PUFA exerts metabolic and anti-inflammatory effects that could prevent left ventricle (LV) pathology; however, it is unclear whether these effects occur at clinically relevant doses and whether there are differences between omega-3 PUFA from fish [eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] and vegetable sources [alpha-linolenic acid (ALA)]. We assessed the development of LV remodelling and pathology in rats subjected to aortic banding treated with omega-3 PUFA over a dose range that spanned the intake of humans taking omega-3 PUFA supplements. Rats were fed a standard food or diets supplemented with EPA+DHA or ALA at 0.7, 2.3, or 7% of energy intake. Without supplementation, aortic banding increased LV mass and end-systolic and -diastolic volumes. ALA supplementation had little effect on LV remodelling and dysfunction. In contrast, EPA+DHA dose-dependently increased EPA and DHA, decreased arachidonic acid in cardiac membrane phospholipids, and prevented the increase in LV end-diastolic and -systolic volumes. EPA+DHA resulted in a dose-dependent increase in the anti-inflammatory adipokine adiponectin, and there was a strong correlation between the prevention of LV chamber enlargement and plasma levels of adiponectin (r = -0.78). Supplementation with EPA+DHA had anti-aggregatory and anti-inflammatory effects as evidenced by decreases in urinary thromboxane B(2) and serum tumour necrosis factor-alpha. Dietary supplementation with omega-3 PUFA derived from fish, but not from vegetable sources, increased plasma adiponectin, suppressed inflammation, and prevented cardiac remodelling and dysfunction under pressure overload conditions.
| 19,015,135
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Regulation of human dUTPase gene expression and p53-mediated transcriptional repression in response to oxaliplatin-induced DNA damage.
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Deoxyuridine triphosphate nucleotidohydrolase (dUTPase) catalyzes the hydrolysis of dUTP to dUMP and PPi. Although dUTP is a normal intermediate in DNA synthesis, its accumulation and misincorporation into DNA is lethal. Importantly, uracil misincorporation is a mechanism of cytotoxicity induced by fluoropyrimidine chemotherapeutic agents including 5-fluorouracil (5-FU) and elevated expression of dUTPase is negatively correlated with clinical response to 5-FU-therapy. In this study we performed the first functional characterization of the dUTPase promoter and demonstrate a role for E2F-1 and Sp1 in driving dUTPase expression. We establish a direct role for both mutant and wild-type forms of p53 in modulating dUTPase promoter activity. Treatment of HCT116 p53(+/+) cells with the DNA-damaging agent oxaliplatin induced a p53-dependent transcriptional downregulation of dUTPase not observed in the isogenic null cell line. Oxaliplatin treatment induced enrichment of p53 at the dUTPase promoter with a concomitant reduction in Sp1. The suppression of dUTPase by oxaliplatin promoted increased levels of dUTP that was enhanced by subsequent addition of fluoropyrimidines. The novel observation that oxaliplatin downregulates dUTPase expression may provide a mechanistic basis contributing to the synergy observed between 5-FU and oxaliplatin in the clinic. Furthermore, these studies provide the first evidence of a direct transcriptional link between the essential enzyme dUTPase and the tumor suppressor p53.
| 19,015,155
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Opioid-mediated muscle afferents inhibit central motor drive and limit peripheral muscle fatigue development in humans.
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We investigated the role of somatosensory feedback from locomotor muscles on central motor drive (CMD) and the development of peripheral fatigue during high-intensity endurance exercise. In a double-blind, placebo-controlled design, eight cyclists randomly performed three 5 km time trials: control, interspinous ligament injection of saline (5K(Plac), L3-L4) or intrathecal fentanyl (5K(Fent), L3-L4) to impair cortical projection of opioid-mediated muscle afferents. Peripheral quadriceps fatigue was assessed via changes in force output pre- versus postexercise in response to supramaximal magnetic femoral nerve stimulation (DeltaQ(tw)). The CMD during the time trials was estimated via quadriceps electromyogram (iEMG). Fentanyl had no effect on quadriceps strength. Impairment of neural feedback from the locomotor muscles increased iEMG during the first 2.5 km of 5K(Fent) versus 5K(Plac) by 12 +/- 3% (P < 0.05); during the second 2.5 km, iEMG was similar between trials. Power output was also 6 +/- 2% higher during the first and 11 +/- 2% lower during the second 2.5 km of 5K(Fent) versus 5K(Plac) (both P < 0.05). Capillary blood lactate was higher (16.3 +/- 0.5 versus 12.6 +/- 1.0%) and arterial haemoglobin O(2) saturation was lower (89 +/- 1 versus 94 +/- 1%) during 5K(Fent) versus 5K(Plac). Exercise-induced DeltaQ(tw) was greater following 5K(Fent) versus 5K(Plac) (-46 +/- 2 versus -33 +/- 2%, P < 0.001). Our results emphasize the critical role of somatosensory feedback from working muscles on the centrally mediated determination of CMD. Attenuated afferent feedback from exercising locomotor muscles results in an overshoot in CMD and power output normally chosen by the athlete, thereby causing a greater rate of accumulation of muscle metabolites and excessive development of peripheral muscle fatigue.
| 19,015,193
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Agonist- and antagonist-induced conformational changes of loop F and their contributions to the rho1 GABA receptor function.
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Binding of gamma-aminobutyric acid (GABA) to its receptor initiates a conformational change to open the channel, but the mechanism of the channel activation is not well understood. To this end, we scanned loop F (K210-F227) in the N-terminal domain of the rho1 GABA receptor expressed in Xenopus oocytes using a site-specific fluorescence technique. We detected GABA-induced fluorescence changes at six positions (K210, K211, L216, K217, T218 and I222). At these positions the fluorescence changes were dose dependent and highly correlated to the current dose-response, but with lower Hill coefficients. The competitive antagonist 3-aminopropyl(methyl)phosphinic acid (3-APMPA) induced fluorescence changes in the same direction at the four middle or lower positions. The non-competitive antagonist picrotoxin blocked nearly 50% of GABA-induced fluorescence changes at T218 and I222, but only <20% at K210 and K217 and 0% at K211 and L216 positions. Interestingly, the picrotoxin-blocked fraction of the GABA-induced fluorescence changes was highly correlated to the Hill coefficient of the GABA-induced dose-dependent fluorescence change. The PTX-insensitive mutant L216C exhibited the lowest Hill coefficient, similar to that in binding. Thus, the PTX-sensitive fraction reflects the conformational change related to channel gating, whereas the PTX-insensitive fraction represents a binding effect. The binding effect is further supported by the picrotoxin resistance of a competitive antagonist-induced fluorescence change. A cysteine accessibility test further confirmed that L216C and K217C partially line the binding pocket, and I222C became more exposed by GABA. Our results are consistent with a mechanism that an outward movement of the lower part of loop F is coupled to the channel activation.
| 19,015,197
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Adaptation of reactive and voluntary saccades: different patterns of adaptation revealed in the antisaccade task.
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Sensorimotor adaptation restores and maintains the accuracy of goal-directed movements. It remains unclear whether these adaptive mechanisms modify actions by controlling peripheral premotor stages that send commands to the effectors and/or earlier processing stages involved in registration of target location. Here, we studied the effect of adaptation of saccadic eye movements, a well-established model of sensorimotor adaptation, in an antisaccade task. This task introduces a clear spatial dissociation between the actual target direction and the requested saccade direction because the correct movement direction is in the opposite direction from the target location. We used this requirement of a vector inversion to assess the level(s) of saccadic adaptation for two different types of adapted saccades. In two different experiments, we tested the transfer to antisaccades of the adaptation in one direction of reactive saccades to jumping targets and of scanning voluntary saccades within a target array. In the first experiment, we found that adaptation of reactive saccades transferred only to antisaccades in the adapted direction. In contrast, in the second experiment, adaptation of scanning voluntary saccades transferred to antisaccades in both the adapted and non-adapted directions. We conclude that adaptation of reactive saccades acts only downstream of the vector inversion required in the antisaccade task, whereas adaptation of voluntary saccades has a distributed influence, acting both upstream and downstream of vector inversion.
| 19,015,199
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High-density lipoprotein: why all the fuss?
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The current search for new treatments to combat coronary heart disease (CHD) is centred on increasing HDL-cholesterol. The failure of the CETP inhibitor torcetrapib may force a rethink. This perspective briefly reviews the antiatherosclerotic properties of HDL and ways HDL-cholesterol concentration can be raised, but argues - in light of the fact that HDL-cholesterol concentration does not reflect the protective properties of HDL particles - that this approach is flawed and a different approach, targeting know antiatherosclerotic components of HDL, is required.
| 19,015,220
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Extracting respiratory data from pulse oximeter plethysmogram traces in newborn infants.
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To investigate whether valid respiratory data can be extracted from the pulse oximeter plethysmographic (pleth) trace in healthy newborn infants, pleth data were collected from the foot, and respiratory airflow was simultaneously measured using a facemask. The pleth waveform was analysed using fast Fourier transform (FFT), low-pass filtering (LPF), and by plotting the peak-to-peak amplitude variation (PtP). Using FFT in 14 term infants, the median (range) respiratory rate from the pleth signal was 43 (30-65) breaths/min, and from the flow signal it was 44 (30-67) breaths/min (median difference 0.01 breaths/min, p>0.05). Both LPF and PtP analysis yielded waveforms with a frequency similar to the respiratory rate. Respiratory information, including respiratory rate and a respiratory-like waveform, can reliably be extracted from the pleth trace of a standard pulse oximeter in newborn infants. Such analysis may be clinically useful for non-invasive assessment of respiratory problems in infants and young children.
| 19,015,221
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Detection of cryptic pathogenic copy number variations and constitutional loss of heterozygosity using high resolution SNP microarray analysis in 117 patients referred for cytogenetic analysis and impact on clinical practice.
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Microarray genome analysis is realising its promise for improving detection of genetic abnormalities in individuals with mental retardation and congenital abnormality. Copy number variations (CNVs) are now readily detectable using a variety of platforms and a major challenge is the distinction of pathogenic from ubiquitous, benign polymorphic CNVs. The aim of this study was to investigate replacement of time consuming, locus specific testing for specific microdeletion and microduplication syndromes with microarray analysis, which theoretically should detect all known syndromes with CNV aetiologies as well as new ones. Genome wide copy number analysis was performed on 117 patients using Affymetrix 250K microarrays. 434 CNVs (195 losses and 239 gains) were found, including 18 pathogenic CNVs and 9 identified as "potentially pathogenic". Almost all pathogenic CNVs were larger than 500 kb, significantly larger than the median size of all CNVs detected. Segmental regions of loss of heterozygosity larger than 5 Mb were found in 5 patients. Genome microarray analysis has improved diagnostic success in this group of patients. Several examples of recently discovered "new syndromes" were found suggesting they are more common than previously suspected and collectively are likely to be a major cause of mental retardation. The findings have several implications for clinical practice. The study revealed the potential to make genetic diagnoses that were not evident in the clinical presentation, with implications for pretest counselling and the consent process. The importance of contributing novel CNVs to high quality databases for genotype-phenotype analysis and review of guidelines for selection of individuals for microarray analysis is emphasised.
| 19,015,223
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USP19 deubiquitinating enzyme supports cell proliferation by stabilizing KPC1, a ubiquitin ligase for p27Kip1.
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p27(Kip1) is a cyclin-dependent kinase inhibitor that regulates the G(1)/S transition. Increased degradation of p27(Kip1) is associated with cellular transformation. Previous work demonstrated that the ubiquitin ligases KPC1/KPC2 and SCF(Skp2) ubiquitinate p27(Kip1) in G(1) and early S, respectively. The regulation of these ligases remains unclear. We report here that the USP19 deubiquitinating enzyme interacts with and stabilizes KPC1, thereby modulating p27(Kip1) levels and cell proliferation. Cells depleted of USP19 by RNA interference exhibited an inhibition of cell proliferation, progressing more slowly from G(0)/G1 to S phase, and accumulated p27(Kip1). This increase in p27(Kip1) was associated with normal levels of Skp2 but reduced levels of KPC1. The overexpression of KPC1 or the use of p27(-/-) cells inhibited significantly the growth defect observed upon USP19 depletion. KPC1 was ubiquitinated in vivo and stabilized by proteasome inhibitors and by overexpression of USP19, and it also coimmunoprecipitated with USP19. Our results identify USP19 as the first deubiquitinating enzyme that regulates the stability of a cyclin-dependent kinase inhibitor and demonstrate that progression through G(1) to S phase is, like the metaphase-anaphase transition, controlled in a hierarchical, multilayered fashion.
| 19,015,242
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Characterization of the binding specificity of K88ac and K88ad fimbriae of enterotoxigenic Escherichia coli by constructing K88ac/K88ad chimeric FaeG major subunits.
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Enterotoxigenic Escherichia coli (ETEC) strains expressing K88 (F4) fimbriae are the major cause of diarrhea in young pigs. Three antigenic variants of K88 fimbriae (K88ab, K88ac, and K88ad) have been identified among porcine ETEC strains. Each K88 fimbrial variant shows a unique pattern in binding to different receptors on porcine enterocytes. Such variant specificity in fimbrial binding is believed to be controlled by the major subunit (FaeG) of the K88 fimbriae, because the genes coding for the only other fimbrial subunit are identical among the three variants. Uniqueness in binding to host receptors may be responsible for differences in the virulence levels of porcine diarrhea disease caused by K88 ETEC strains. To better understand the relationships between the structure of FaeG proteins and fimbrial binding function, and perhaps virulence in disease, we constructed and expressed various K88ac/K88ad faeG gene chimeras and characterized the binding activity of each K88 chimeric fimbria. After verifying biosynthesis of the chimeric fimbriae, we examined their binding specificities in bacterial adherence assays by using porcine brush border vesicles that are specific to either the K88ac or K88ad fimbria. Results showed that each fimbria switched binding specificity to that of the reciprocal type when a peptide comprising amino acids 125 to 163 was exchanged with that of its counterpart. Substitutions of a single amino acid within this region negatively affected the binding capacity of each fimbria. These data indicate that the peptide including amino acids 125 to 163 of the FaeG subunit is essential for K88 variant-specific binding.
| 19,015,246
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Humoral immunity against capsule polysaccharide protects the host from magA+ Klebsiella pneumoniae-induced lethal disease by evading Toll-like receptor 4 signaling.
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Klebsiella pneumoniae magA (for mucoviscosity-associated gene A) is linked to the pathogenesis of primary pyogenic liver abscess, but the underlying mechanism by which magA increases pathogenicity is not well elucidated. In this study, we investigated the role of the capsular polysaccharides (CPS) in the pathogenesis of magA(+) K. pneumoniae by comparing host immunity to magA(+) K. pneumoniae and a DeltamagA mutant. We found that Toll-like receptor 4 recognition by magA(+) K. pneumoniae was hampered by the mucoviscosity of the magA(+) K. pneumoniae CPS. Interestingly, monoclonal antibodies (MAbs) against magA(+) K. pneumoniae CPS recognized all of the K1 strains tested but not the DeltamagA and non-K1 strains. Moreover, the anti-CPS MAbs protected mice from magA(+) K. pneumoniae-induced liver abscess formation and lethality. This indicates that the K1 epitope is a promising target for vaccine development, and anti-CPS MAbs has great potential to protect host from K1 strain-induced mortality and morbidity in diabetic and other immunocompromised patients in the future.
| 19,015,249
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Rapamycin-induced Gln3 dephosphorylation is insufficient for nuclear localization: Sit4 and PP2A phosphatases are regulated and function differently.
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Gln3, the major activator of nitrogen catabolite repression (NCR)-sensitive transcription, is often used as an assay of Tor pathway regulation in Saccharomyces cerevisiae. Gln3 is cytoplasmic in cells cultured with repressive nitrogen sources (Gln) and nuclear with derepressive ones (Pro) or after treating Gln-grown cells with the Tor inhibitor, rapamycin (Rap). In Raptreated or Pro-grown cells, Sit4 is posited to dephosphorylate Gln3, which then dissociates from a Gln3-Ure2 complex and enters the nucleus. However, in contrast with this view, Sit4-dependent Gln3 dephosphorylation is greater in Gln than Pro. Investigating this paradox, we show that PP2A (another Tor pathway phosphatase)-dependent Gln3 dephosphorylation is regulated oppositely to that of Sit4, being greatest in Pro- and least in Gln-grown cells. It thus parallels nuclear Gln3 localization and NCR-sensitive transcription. However, because PP2A is not required for nuclear Gln3 localization in Pro, PP2A-dependent Gln3 dephosphorylation and nuclear localization are likely parallel responses to derepressive nitrogen sources. In contrast, Rap-induced nuclear Gln3 localization absolutely requires all four PP2A components (Pph21/22, Tpd3, Cdc55, and Rts1). In pph21Delta22Delta, tpd3Delta, or cdc55Delta cells, however, Gln3 is dephosphorylated to the same level as in Rap-treated wild-type cells, indicating Rap-induced Gln3 dephosphorylation is insufficient to achieve nuclear localization. Finally, PP2A-dependent Gln3 dephosphorylation parallels conditions where Gln3 is mostly nuclear, while Sit4-dependent and Rap-induced dephosphorylation parallels those where Gln3 is mostly cytoplasmic, suggesting the effects of these phosphatases on Gln3 may occur in different cellular compartments.
| 19,015,262
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The genetic basis of hereditary medullary thyroid cancer: clinical implications for the surgeon, with a particular emphasis on the role of prophylactic thyroidectomy.
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Medullary thyroid cancer (MTC) may occur either sporadically or on a hereditary basis. Hereditary MTC may be observed with either multiple endocrine neoplasia syndromes (MEN 2A and MEN 2B) or as familial MTC (FMTC). Despite the rarity of these syndromes, early diagnosis is especially important, since MTC is a lethal disease if not promptly and appropriately treated. Recently, the development of genetic testing and direct DNA analysis allows the identification of asymptomatic patients. Surgical prophylaxis should be considered in these cases, ideally to prevent the development of MTC. During the recent decade, the concept of 'codon-directed' timing of prophylactic surgery emerged as a reasonable strategy in the management of these patients. Currently, genetic analysis offers the possibility to define genotype-phenotype correlations and to adjust the time of prophylactic surgery. Hereditary MTC is a model of genetically determined cancer in which both diagnostic and therapeutic strategies rely on the identification of specific mutations.
| 19,015,274
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Aire controls the differentiation program of thymic epithelial cells in the medulla for the establishment of self-tolerance.
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The roles of autoimmune regulator (Aire) in the expression of the diverse arrays of tissue-restricted antigen (TRA) genes from thymic epithelial cells in the medulla (medullary thymic epithelial cells [mTECs]) and in organization of the thymic microenvironment are enigmatic. We approached this issue by creating a mouse strain in which the coding sequence of green fluorescent protein (GFP) was inserted into the Aire locus in a manner allowing concomitant disruption of functional Aire protein expression. We found that Aire(+) (i.e., GFP(+)) mTECs were the major cell types responsible for the expression of Aire-dependent TRA genes such as insulin 2 and salivary protein 1, whereas Aire-independent TRA genes such as C-reactive protein and glutamate decarboxylase 67 were expressed from both Aire(+) and Aire(-) mTECs. Remarkably, absence of Aire from mTECs caused morphological changes together with altered distribution of mTECs committed to Aire expression. Furthermore, we found that the numbers of mTECs that express involucrin, a marker for terminal epidermal differentiation, were reduced in Aire-deficient mouse thymus, which was associated with nearly an absence of Hassall's corpuscle-like structures in the medulla. Our results suggest that Aire controls the differentiation program of mTECs, thereby organizing the global mTEC integrity that enables TRA expression from terminally differentiated mTECs in the thymic microenvironment.
| 19,015,306
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TIMs: central regulators of immune responses.
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Exhaustion of T cell responses during chronic viral infections has been observed in both mouse and man and has been attributed to up-regulation of PD-1 on the surface of exhausted T cells. In patients with chronic human HIV infection, T cell exhaustion leads to opportunistic infections associated with AIDS. However, not all the exhausted T cells express PD-1, suggesting that other molecules may be involved in the phenotype. A new study now demonstrates a central role for T cell immunoglobulin and mucin domain-containing protein-3 (TIM-3) in T cell exhaustion during chronic HIV infection and suggests that TIM-3 may be a novel therapeutic target in chronic viral diseases.
| 19,015,312
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RhoA-ROCK and p38MAPK-MSK1 mediate vitamin D effects on gene expression, phenotype, and Wnt pathway in colon cancer cells.
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The active vitamin D metabolite 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) inhibits proliferation and promotes differentiation of colon cancer cells through the activation of vitamin D receptor (VDR), a transcription factor of the nuclear receptor superfamily. Additionally, 1,25(OH)(2)D(3) has several nongenomic effects of uncertain relevance. We show that 1,25(OH)(2)D(3) induces a transcription-independent Ca(2+) influx and activation of RhoA-Rho-associated coiled kinase (ROCK). This requires VDR and is followed by activation of the p38 mitogen-activated protein kinase (p38MAPK) and mitogen- and stress-activated kinase 1 (MSK1). As shown by the use of chemical inhibitors, dominant-negative mutants and small interfering RNA, RhoA-ROCK, and p38MAPK-MSK1 activation is necessary for the induction of CDH1/E-cadherin, CYP24, and other genes and of an adhesive phenotype by 1,25(OH)(2)D(3). RhoA-ROCK and MSK1 are also required for the inhibition of Wnt-beta-catenin pathway and cell proliferation. Thus, the action of 1,25(OH)(2)D(3) on colon carcinoma cells depends on the dual action of VDR as a transcription factor and a nongenomic activator of RhoA-ROCK and p38MAPK-MSK1.
| 19,015,318
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Comparison of the performances of two in-house rapid methods for antitubercular drug susceptibility testing.
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Resistance to rifampin (rifampicin), isoniazid, and streptomycin of 69 Mycobacterium tuberculosis isolates was analyzed by an in-house method based on mycobacteriophage D29 and a colorimetric micromethod. Both methods showed sensitivity and specificity values ranging from 93% to 100%. These simple methods offer an option for drug resistance assessment of M. tuberculosis.
| 19,015,333
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Antileishmanial activity of 1,3,4-thiadiazolium-2-aminide in mice infected with Leishmania amazonensis.
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The efficacy of two mesoionic derivatives (MI-H-H and MI-4-OCH(3)) was evaluated in CBA/J mice infected with Leishmania amazonensis. Treatment with these compounds demonstrated that the MI-4-OCH(3) derivative and the reference drug meglumine antimoniate (Glucantime) presented significant activity relative to an untreated control. No apparent hepatic or renal toxicity due to these mesoionic compounds was found.
| 19,015,338
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Statistical model to evaluate in vivo activities of antimalarial drugs in a Plasmodium cynomolgi-macaque model for Plasmodium vivax malaria.
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Preclinical animal models informing antimalarial drug development are scarce. We have used asexual erythrocytic Plasmodium cynomolgi infections of rhesus macaques to model Plasmodium vivax during preclinical development of compounds targeting parasite phospholipid synthesis. Using this malaria model, we accumulated data confirming highly reproducible infection patterns, with self-curing parasite peaks reproducibly preceding recrudescence peaks. We applied nonlinear mixed-effect (NLME) models, estimating treatment effects in three drug studies: G25 (injected) and the bisthiazolium prodrugs TE4gt and TE3 (oral). All compounds fully cured P. cynomolgi-infected macaques, with significant effects on parasitemia height and time of peak. Although all three TE3 doses tested were fully curative, NLME models discriminated dose-dependent differential pharmacological antimalarial activity. By applying NLME modeling treatment effects are readily quantified. Such drug development studies are more informative and contribute to reduction and refinement in animal experimentation.
| 19,015,340
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Safety, pharmacokinetics, and antiretroviral activity of multiple doses of ibalizumab (formerly TNX-355), an anti-CD4 monoclonal antibody, in human immunodeficiency virus type 1-infected adults.
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Ibalizumab (formerly TNX-355) is a humanized monoclonal antibody that binds CD4, the primary receptor for human immunodeficiency virus type 1 (HIV-1), and inhibits the viral entry process. A phase lb multidose study of the safety, pharmacokinetics, and antiviral activity of ibalizumab was conducted with 22 HIV-1-infected patients. Nineteen patients were randomized to receive either 10 mg/kg of body weight weekly (arm A) or a 10-mg/kg loading dose followed by 6 mg/kg every 2 weeks (arm B) intravenously for 9 weeks. Three patients were assigned to receive 25 mg/kg every 2 weeks for five doses (arm C). During the study, the patients remained off other antiretrovirals or continued a stable failing regimen. Treatment with ibalizumab resulted in substantial reductions in HIV-1 RNA levels (0.5 to 1.7 log(10)) in 20 of 22 subjects. In most patients, HIV-1 RNA fell to nadir levels after 1 to 2 weeks of treatment and then returned to baseline despite continued treatment. Baseline viral isolates were susceptible to ibalizumab in vitro, regardless of coreceptor tropism. Emerging resistance to ibalizumab was manifested by reduced maximal percent inhibition in a single-cycle HIV infectivity assay. Resistant isolates remained CD4 dependent and were susceptible to enfuvirtide in vitro. Complete coating of CD4(+) T-cell receptors was correlated with serum ibalizumab concentrations. There was no evidence of CD4(+) T-cell depletion in ibalizumab-treated patients. Ibalizumab was not immunogenic, and no serious drug-related adverse effects occurred. In conclusion, ibalizumab administered either weekly or biweekly was safe and well tolerated and demonstrated antiviral activity. Further studies with ibalizumab in combination with standard antiretroviral treatments are warranted.
| 19,015,347
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Activity of levofloxacin alone and in combination with a DnaK inhibitor against gram-negative rods, including levofloxacin-resistant strains.
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Synergy time-kill testing of levofloxacin alone and in combination with CHP-105, a representative DnaK inhibitor, against 50 gram-negative rods demonstrated that 34 of the 50 strains tested showed significant synergy between levofloxacin and CHP-105 after 12 h and 24 h. Fourteen of these 34 organisms were quinolone resistant (levofloxacin MICs of > or =4 microg/ml).
| 19,015,359
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Multistrain virus dynamics with mutations: a global analysis.
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We consider within-host virus models with n >or= 2 strains and allow mutation between the strains. If there is no mutation, a Lyapunov function establishes global stability of the steady state corresponding to the fittest strain. For small perturbations, this steady state persists, perhaps with small concentrations of some or all other strains, depending on the connectivity of the graph describing all possible mutations. Moreover, using a perturbation result due to Smith & Waltman (1999), we show that this steady state also preserves global stability.
| 19,015,367
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Object representations for multiple visual categories overlap in lateral occipital and medial fusiform cortex.
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How representations of visual objects are maintained across changes in viewpoint is a central issue in visual perception. Whether neural processes underlying view-invariant recognition involve distinct subregions within extrastriate visual cortex for distinct categories of visual objects remains unresolved. We used event-related functional magnetic resonance imaging in 16 healthy volunteers to map visual cortical areas responding to a large set (156) of exemplars from 3 object categories (faces, houses, and chairs), each repeated once after a variable time lag (3-7 intervening stimuli). Exemplars were repeated with the same viewpoint (but different retinal size) or with different viewpoint and size. The task was kept constant across object categories (judging items as "young" vs. "old"). We identified object-selective adaptation effects by comparing neural responses to the first presentation versus repetition of each individual exemplar. We found that exemplar-specific adaptation effects partly overlapped with regions showing category-selective responses (as identified using a separate localizer scan). These included the lateral fusiform gyrus (FG) for faces, parahippocampal gyrus for houses, and lateral occipital complex (LOC) for chairs. In face-selective fusiform gyrus (FG), adaptation effects occurred only for faces repeated with the same viewpoint, but not with a different viewpoint, confirming previous studies using faces only. By contrast, a region in right medial FG, adjacent to but nonoverlapping with the more lateral and face-selective FG, showed repetition effects for faces and to a lesser extent for other objects, regardless of changes in viewpoint or in retinal image-size. Category- and viewpoint-independent repetition effects were also found in bilateral LOC. Our results reveal a common neural substrate in bilateral LOC and right medial FG underlying view-invariant and category-independent recognition for multiple object identities, with only a relative preference for faces in medial FG but no selectivity in LOC.
| 19,015,371
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B-cell count and survival: differentiating chronic lymphocytic leukemia from monoclonal B-cell lymphocytosis based on clinical outcome.
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The diagnosis of chronic lymphocytic leukemia (CLL) in asymptomatic patients has historically been based on documenting a characteristic lymphocyte clone and the presence of lymphocytosis. There are minimal data regarding which lymphocyte parameter (absolute lymphocyte count [ALC] or B-cell count) and what threshold should be used for diagnosis. We analyzed the relationship of ALC and B-cell count with clinical outcome in 459 patients with a clonal population of CLL phenotype to determine (1) whether the CLL diagnosis should be based on ALC or B-cell count, (2) what lymphocyte threshold should be used for diagnosis, and (3) whether any lymphocyte count has independent prognostic value after accounting for biologic/molecular prognostic markers. B-cell count and ALC had similar value for predicting treatment-free survival (TFS) and overall survival as continuous variables, but as binary factors, a B-cell threshold of 11 x 10(9)/L best predicted survival. B-cell count remained an independent predictor of TFS after controlling for ZAP-70, IGHV, CD38, or fluorescence in situ hybridization (FISH) results (all P < .001). These analyses support basing the diagnosis of CLL on B-cell count and retaining the size of the B-cell count in the diagnostic criteria. Using clinically relevant criteria to distinguish between monoclonal B-cell lymphocytosis (MBL) and CLL could minimize patient distress caused by labeling asymptomatic people at low risk for adverse clinical consequences as having CLL.
| 19,015,397
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The angular branch: maximizing the scapular pedicle in head and neck reconstruction.
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To evaluate the scapular free flap based on the angular artery in complex head and neck reconstruction. Case series. A tertiary referral center. A series of 25 osteocutaneous scapular flaps was performed from August 2000 through January 2005. Of these 25 flaps, 7 procedures of scapular bone solely vascularized by the angular artery and vein were performed to reconstruct head and neck defects. The angular vessels were used to reach the neck for anastomosis in midfacial reconstruction (n = 2), to carry a separate second bone flap in complex oromandibular defects (n = 2), and to reach the contralateral neck for anastomosis in through-and-through oromandibular defects encompassing overlying facial skin (n = 3). Pedicle length and flap viability. Postoperative bone scans revealed all bone segments to be vascularized. The pedicle length originating from the circumflex scapular vessels varied from 6.7 to 9.0 cm (mean length, 7.5 cm). The pedicle length of the angular vessels varied from 13.0 to 15.0 cm (mean length, 14.1 cm), a mean length of 6.6 cm longer than the circumflex scapular flap. Vein grafts were not necessary to perform remote anastomoses with the additional pedicle length. The angular vessels can reliably supply the scapula. Use of the angular vessels over the circumflex scapular vessels increases the bone pedicle length by a mean length of 6.6 cm (88%) and is a useful technique to avoid vein grafting for remote anastomosis.
| 19,015,454
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Modulation of tumor cell growth in vivo by extracellular matrix metalloprotease inducer.
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To investigate if loss of extracellular matrix metalloprotease inducer (EMMPRIN) will inhibit the growth of head and neck squamous cell carcinoma (HNSCC) tumor cell lines in vivo. Tumor cell-derived EMMPRIN is highly overexpressed in HNSCC and is thought to be induced by surrounding fibroblasts to stimulate matrix metalloproteases, which modulate tumor cell invasion, growth, and angiogenesis. In vivo study using FaDu tumor xenografts. Academic research facility. Severe combined immunodeficiency (SCID) mice. The HNSCC cell line FaDu was transfected with EMMPRIN (FaDu/E), control vector (FaDu), or plasmid-expressing small-interfering RNA against EMMPRIN (FaDu/siE). Tumor cells combined with fibroblast cells were xenografted onto the flank of SCID mice. Tumors were measured biweekly over 4 weeks, at which time the mice were killed, and tumor samples were analyzed for proliferation (Ki-67 immunohistochemical analysis), vascularization (factor VIII staining), and apoptosis (TUNEL [terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling] assay). Growth of head and neck cancer cell lines genetically engineered to express variable levels of EMMPRIN. Tumor growth positively correlated and animal survival negatively correlated with increasing EMMPRIN expression. FaDu/E tumor growth was significantly larger at 4 weeks compared with FaDu tumors (P = .006). Similarly, the control vector-transfected FaDu tumors were significantly larger than FaDu/siE (P < .001). Immunohistochemical analysis demonstrated increased Ki-67 in EMMPRIN-transfected cells, without a significant change in the rate of apoptosis between groups. Vascular density and tumor formation rate also increased significantly with EMMPRIN expression. This study suggests that anti-EMMPRIN-targeted therapy may prove to be a novel treatment option in HNSCC.
| 19,015,455
|
Gastrointestinal tract recovery in patients undergoing bowel resection: results of a randomized trial of alvimopan and placebo with a standardized accelerated postoperative care pathway.
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To investigate the efficacy and safety of alvimopan, 12 mg, administered orally 30 to 90 minutes preoperatively and twice daily postoperatively in conjunction with a standardized accelerated postoperative care pathway for managing postoperative ileus after bowel resection. This multicenter, randomized, placebo-controlled, double-blind, phase 3 trial enrolled adult patients undergoing partial bowel resection with primary anastomosis by laparotomy and scheduled to receive intravenous, opioid-based, patient-controlled analgesia. A standardized accelerated postoperative care pathway including early ambulation, oral feeding, and postoperative nasogastric tube removal was used to facilitate gastrointestinal (GI) tract recovery in all of the patients. The primary end point was time to GI-2 recovery (toleration of solid food and first bowel movement). Secondary end points included time to GI-3 recovery (toleration of solid food and first flatus or bowel movement), hospital discharge order written, and actual hospital discharge. Postoperative length of hospital stay based on calendar day of hospital discharge order written, opioid consumption, and overall postoperative ileus-related morbidity were recorded. Alvimopan, 12 mg, was well tolerated and significantly accelerated GI-2 recovery, GI-3 recovery, and actual hospital discharge compared with a standardized accelerated postoperative care pathway alone (hazard ratio = 1.5, 1.5, and 1.4, respectively; P < .001 for all). Time to hospital discharge order written as measured by hazard ratio (1.4) and by postoperative calendar days (mean for alvimopan, 5.2 days; mean for placebo, 6.2 days) was also accelerated. Opioid consumption was comparable between groups, and alvimopan was associated with reduced postoperative ileus-related morbidity compared with placebo. Alvimopan, 12 mg, administered 30 to 90 minutes before and twice daily after bowel resection is well tolerated, accelerates GI tract recovery, and reduces postoperative ileus-related morbidity without compromising opioid analgesia.
| 19,015,469
|
Nipple-sparing mastectomy update: one hundred forty-nine procedures and clinical outcomes.
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To describe our experience with patients who underwent the nipple-sparing mastectomy procedure developed and standardized at our institution and to report clinical outcomes for those patients with a breast cancer diagnosis. Prospective study for consecutive nipple-sparing mastectomy procedures. Multidisciplinary breast center at a large tertiary care facility. One hundred ten consecutive patients underwent nipple-sparing mastectomy between July 2001 and June 2007. Nipple-sparing mastectomy was offered to carefully screened patients; the nipple-areola tissue was cored and sent for histologic frozen-section analysis intraoperatively. Assessment of nipple-areola cored tissue for neoplastic involvement; postoperative stability of retained nipple-areola complex; and clinical outcomes. Data were available for 149 nipple-sparing mastectomies performed on 110 patients. No procedure performed for prevention had neoplastic involvement of the cored nipple-areola tissue, while 9 procedures performed for breast cancer treatment were found to have neoplastic involvement. Postoperatively, 2 patients had partial loss of the nipple-areola complex due to sloughing and a third patient developed an infection that required surgical removal of the nipple-areola complex. Among patients with breast cancer, none with ductal carcinoma in situ has developed a recurrence, while 4 patients with infiltrating breast cancer have, including 2 patients with distant metastases only, a third with a chest wall recurrence, and a fourth with an axillary recurrence. A low incidence of neoplastic involvement of the nipple-areola cored tissue leads to successful completion of nipple-sparing mastectomy for most patients.
| 19,015,470
|
GM1/GalNAc-GD1a complex: a target for pure motor Guillain-Barre syndrome.
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GM1 and GalNAc-GD1a are located on the axolemma of the motor nerves and are believed to be the antigens associated with pure motor Guillain-Barré syndrome (GBS). Furthermore, GM1 and GalNAc-GD1a may exist nearby and colocalize on the axolemma. Ganglioside complex (GSC) antigens associated with GM1 or GalNAc-GD1a can be target antigens in pure motor GBS. We investigated GBS sera for antibodies to a GSC consisting of GM1 and GalNAc-GD1a (GM1/GalNAc-GD1a) and analyzed the clinical and electrophysiologic findings of patients with antibodies to GM1/GalNAc-GD1a. Sera from 224 patients with GBS were surveyed for antibodies to GSCs consisting of two of nine gangliosides (GM1, GM2, GM3, GD1a, GD3, GT1a, GT1b, GQ1b, and GalNAc-GD1a). We analyzed the clinical and electrophysiologic features of patients with IgG antibodies to the GM1/GalNAc-GD1a complex. Ten patients with GBS had IgG antibodies to the GM1/GalNAc-GD1a complex. The clinical findings of the 10 patients with GBS were characterized by preserved sensory system and infrequent cranial nerve deficits. According to the criteria established by Hadden et al., electrodiagnostic studies showed a demyelinating pattern in four patients and axonal neuropathy pattern in two. Early motor conduction block at intermediate nerve segments was found in five patients. GM1 and GalNAc-GD1a may form a complex in the axolemma at nodes of Ranvier or paranodes of the motor nerves, and may be a target antigen in pure motor Guillain-Barré syndrome, especially in the form of acute motor conduction block neuropathy.
| 19,015,484
|
Early-onset dementia with prolonged occipital seizures: an atypical case of Kufs disease.
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Kufs disease is the adult-onset form of neuronal ceroid lipofuscinosis (NCL). Its two clinical phenotypes are type A (progressive myoclonus epilepsy with dementia) and type B (behavioral abnormalities and dementia, associated with pyramidal and extrapyramidal signs). We describe the clinical evolution of an atypical case characterized by progressive dementia and focal occipital seizures. A healthy 37-year-old woman began showing memory deficits and behavioral disturbances (apathy, lack of inhibitions, untidiness). After 4 years, she developed rare clusters of tonic-clonic seizures, as well as focal seizures originating from the temporo-occipital regions, clinically associated with visual hallucinations, wandering, and agitation. When she was 44 years old, neuropsychological assessment revealed severe frontotemporal dementia. MRI showed cortical atrophy and, on T2-weighted images, hypointensity of the basal ganglia, and hyperintensity and reduction of the deep white matter. On the basis of these findings, a diagnosis of Kufs disease was hypothesized. A skin biopsy was negative, but electron microscopy examination of a right frontal lobe brain biopsy revealed the presence of typical storage material (fingerprint inclusions). The patient never developed myoclonus or extrapyramidal signs. Kufs disease is difficult to diagnose on account of its heterogeneous clinical pattern and pathologic features, and the lack of a specific genetic locus alteration. The neuropsychological pattern and MRI findings observed in patients with early-onset frontotemporal dementia and seizure disorder suggest that Kufs disease should be considered in their differential diagnosis. Extracerebral biopsy can be nondiagnostic, and when alternative diagnoses have been ruled out, cerebral biopsy should be considered.
| 19,015,486
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Perceptual representations in false recognition and priming of pictures.
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Using a new procedure, we investigate whether imagination can induce false memory by creating a perceptual representation. Participants studied pictures and words with and without an imagery task and at test performed both a direct recognition test and an indirect perceptual identification test on pictorial stimuli. Corrected false recognition rates were 7% for pictures studied in word form (Experiment 1), 26% for pictures imagined once (Experiment 2), and 48% for pictures imagined multiple times (Experiment 3), although on the indirect test, no priming was found for these items. Furthermore, a perceptual/conceptual imagery manipulation did not affect the tendency to claim that imagined items had been studied as pictures (Experiment 4). These results suggest that the false memories reported on direct tests are not driven by perceptual representations.
| 19,015,501
|
Exploring knotting mechanisms in protein folding.
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One of the most striking topological features to be found in a protein is that of a distinct knot formed by the path of the polypeptide backbone. Such knotted structures represent some of the smallest "self-tying" knots observed in Nature. Proteins containing a knot deep within their structure add an extra complication to the already challenging protein-folding problem; it is not obvious how, during the process of folding, a substantial length of polypeptide chain manages to spontaneously thread itself through a loop. Here, we probe the folding mechanism of YibK, a homodimeric alpha/beta-knot protein containing a deep trefoil knot at its carboxy terminus. By analyzing the effect of mutations made in the knotted region of the protein we show that the native structure in this area remains undeveloped until very late in the folding reaction. Single-site destabilizing mutations made in the knot structure significantly affect only the folding kinetics of a late-forming intermediate and the slow dimerization step. Furthermore, we find evidence to suggest that the heterogeneity observed in the denatured state is not caused by isomerization of the single cis proline bond as previously thought, but instead could be a result of the knotting mechanism. These results allow us to propose a folding model for YibK where the threading of the polypeptide chain and the formation of native structure in the knotted region of the protein occur independently as successive events.
| 19,015,517
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The structure of corepressor Dax-1 bound to its target nuclear receptor LRH-1.
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The Dax-1 protein is an enigmatic nuclear receptor that lacks an expected DNA binding domain, yet functions as a potent corepressor of nuclear receptors. Here we report the structure of Dax-1 bound to one of its targets, liver receptor homolog 1 (LRH-1). Unexpectedly, Dax-1 binds to LRH-1 using a new module, a repressor helix built from a family conserved sequence motif, PCFXXLP. Mutations in this repressor helix that are linked with human endocrine disorders dissociate the complex and attenuate Dax-1 function. The structure of the Dax-1:LRH-1 complex provides the molecular mechanism for the function of Dax-1 as a potent transcriptional repressor.
| 19,015,525
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Orthology, function and evolution of accessory gland proteins in the Drosophila repleta group.
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The accessory gland proteins (Acps) of Drosophila have become a model for the study of reproductive protein evolution. A major step in the study of Acps is to identify biological causes and consequences of the observed patterns of molecular evolution by comparing species groups with different biology. Here we characterize the Acp complement of Drosophila mayaguana, a repleta group representative. Species of this group show important differences in ecology and reproduction as compared to other Drosophila. Our results show that the extremely high rates of Acp evolution previously found are likely to be ubiquitous among species of the repleta group. These evolutionary rates are considerably higher than the ones observed in other Drosophila groups' Acps. This disparity, however, is not accompanied by major differences in the estimated number of Acps or in the functional categories represented as previously suggested. Among the genes expressed in accessory glands of D. mayaguana almost half are likely products of recent duplications. This allowed us to test predictions of the neofunctionalization model for gene duplication and paralog evolution in a more or less constrained timescale. We found that positive selection is a strong force in the early divergence of these gene pairs.
| 19,015,541
|
A high-density single nucleotide polymorphism map for Neurospora crassa.
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We report the discovery and validation of a set of single nucleotide polymorphisms (SNPs) between the reference Neurospora crassa strain Oak Ridge and the Mauriceville strain (FGSC 2555), of sufficient density to allow fine mapping of most loci. Sequencing of Mauriceville cDNAs and alignment to the completed genomic sequence of the Oak Ridge strain identified 19,087 putative SNPs. Of these, a subset was validated by cleaved amplified polymorphic sequence (CAPS), a simple and robust PCR-based assay that reliably distinguishes between SNP alleles. Experimental confirmation resulted in the development of 250 CAPS markers distributed evenly over the genome. To demonstrate the applicability of this map, we used bulked segregant analysis followed by interval mapping to locate the csp-1 mutation to a narrow region on LGI. Subsequently, we refined mapping resolution to 74 kbp by developing additional markers, resequenced the candidate gene, NCU02713.3, in the mutant background, and phenocopied the mutation by gene replacement in the WT strain. Together, these techniques demonstrate a generally applicable and straightforward approach for the isolation of novel genes from existing mutants. Data on both putative and validated SNPs are deposited in a customized public database at the Broad Institute, which encourages augmentation by community users.
| 19,015,548
|
Metabolic Syndrome and the Risk of Ischemic Heart Disease and Stroke among Middle-Aged Japanese.
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Limited information is available regarding risk of cardiovascular disease and trends for the metabolic syndrome in Asia. We examined the impact of the metabolic syndrome and its components on risk of cardiovascular disease among middle-aged Japanese according to four criteria. We followed 2,613 subjects from a rural Japanese community who participated in cardiovascular health examinations between 1990 and 1993. After 27,477 person-years of follow-up through 2003, there were 42 incidents of ischemic heart disease, 73 total strokes (54 ischemic and 18 hemorrhagic), and 115 total cases of cardiovascular disease. The metabolic syndrome was defined according to the National Cholesterol Education Program Adult Treatment Panel III (NCEP-ATPIII), American Heart Association/National Heart, Lung, and Blood Institute (AHA/NHLBI), International Diabetes Federation (IDF), and Japanese criteria. The multivariable hazard ratios (95%CI) associated with the metabolic syndrome based on NCEP-ATPIII criteria were 2.1 (1.1-4.0) for ischemic heart disease, 1.7 (1.0-2.7) for total stroke, 2.0 (1.2-3.5) for ischemic stroke, 1.1 (0.4-2.8) for hemorrhagic stroke, 2.0 (1.3-3.1) for ischemic cardiovascular disease, and 1.7 (1.2-2.5) for total cardiovascular disease. The population-attributable fractions of the metabolic syndrome based on NCEP-ATPIII criteria were 26-27% for ischemic heart disease and ischemic stroke and 20% for total cardiovascular disease. The metabolic syndrome based on AHA/NHLBI, IDF and Japanese criteria had weaker associations with risk of cardiovascular disease, and the association with risk of ischemic heart disease was not statistically significant. The metabolic syndrome based on NCEP-ATP III criteria predicted risks of ischemic heart disease, ischemic stroke and total cardiovascular disease, whereas that based on three other criteria predicted them less effectively.
| 19,015,596
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Association of serum lipocalin-type prostaglandin D synthase levels with subclinical atherosclerosis in untreated asymptomatic subjects.
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Recent studies suggest that lipocalin-type prostaglandin (PG) D synthase (L-PGDS), which converts PGH2 to PGD2, is implicated in the pathogenesis of atherosclerosis. However, clinical evidence for the association between serum L-PGDS levels and atherosclerosis has not been reported. In this study, we measured the serum L-PGDS concentration using sandwich enzyme-linked immunosorbent assay (ELISA) and investigated the association with traditional cardiovascular risk factors and surrogate atherosclerotic indices, such as the maximum score of the intima-media complex thickness of the carotid artery (C-IMT(max)) and the brachial-ankle pulse wave velocity (ba-PWV), in 500 non-treated asymptomatic subjects. The serum concentration of L-PGDS was 0.56+/-0.01 (mean+/-SEM, range 0.25-1.27, median 0.54) mg/L. Serum L-PGDS levels increased with age and were higher in men than in women. Serum L-PGDS was higher in subjects with hypertension and increased with increasing numbers of the traditional atherosclerotic risk factors. When the subjects were divided into four groups according to the levels of serum L-PGDS, the age-adjusted values of C-IMT(max) and ba-PWV were significantly increased in subjects with higher serum L-PGDS levels (quartile 3 and quartile 4) compared to those in the lowest quartile (quartile 1), for both genders. Multiple regression analysis including risk factors revealed that serum L-PGDS was an independent determinant for ba-PWV (beta=0.130, p<0.001). Serum L-PGDS tended to associate with C-IMT(max) but was not statistically significant (beta=0.084, p=0.075). In conclusion, our results suggest that an increase in serum L-PGDS concentration is associated with the progression of atherosclerosis.
| 19,015,601
|
Superior vena cava syndrome caused by an intravascular thrombosis due to underlying prostate carcinoma.
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Superior vena cava (SVC) syndrome is usually caused by malignant tumors or their lymph node metastases oppressing a SVC. However, we encountered a case of SVC syndrome that was caused by a thrombus in the SVC, which we considered as a manifestation of Trousseau's syndrome triggered by underlying prostate cancer. A 60-year-old man patient complained of facial swelling. Physical examinations suggested SVC syndrome; enhanced CT and MRI demonstrated the presence of thrombus in the SVC accompanied by multiple mediastinal and axillary lymph node swelling. Histological examination of both percutaneous transluminally aspirated thrombus via a catheter through jugular vein and the axillary lymph nodes included metastatic prostate cancer. Although the ultrasonic and MR images were not compatible with the prostate cancer, needle biopsies from the prostate established the diagnosis. The SVC syndrome as an initial manifestation of underlying unknown malignancy and also due to intravascular thrombosis caused by cancer metastasis to the vascular wall is extremely uncommon.
| 19,015,618
|
Synthesis and structure of new 3,3,9,9-tetrasubstituted-2,4,8,10-tetraoxaspiro[5.5]undecane derivatives.
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The configurational and conformational behavior of some new 3,3,9,9-tetrasubstituted-2,4,8,10-tetraoxaspiro[5.5]undecane derivatives with axial chirality was investigated by conformational analysis and variable temperature NMR experiments.
| 19,015,624
|
PML tumor suppressor is regulated by HIPK2-mediated phosphorylation in response to DNA damage.
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The promyelocytic leukemia (PML) tumor suppressor protein, a central regulator of cell proliferation and apoptosis, is frequently fused to the retinoic acid receptor-alpha (RARalpha) in acute PML. Here we show the interaction of PML with another tumor suppressor protein, the serine/threonine kinase homeodomain-interacting protein kinase (HIPK2). In response to DNA damage, HIPK2 phosphorylates PML at serines 8 and 38. Although HIPK2-mediated phosphorylation of PML occurs early during the DNA damage response, the oncogenic PML-RARalpha fusion protein is phosphorylated with significantly delayed kinetics. DNA damage or HIPK2 expression leads to the stabilization of PML and PML-RARalpha proteins. The N-terminal phosphorylation sites contribute to the DNA damage-induced PML SUMOylation and are required for the ability of PML to cooperate with HIPK2 for the induction of cell death.
| 19,015,637
|
Predicting the ideal serum creatinine level following kidney transplantation.
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This Practice Point commentary reviews a study by Sberro et al. that evaluated formulae to predict the lowest measured serum creatinine concentration in kidney transplant recipients following surgery. The objective of the study was to ascertain a simple means of identifying patients with inappropriately high serum creatinine concentrations, who are in need of further investigation. A prediction formula based on the recipient's age and weight and the donor's preoperative estimated creatinine clearance, as calculated from the Cockcroft-Gault equation, showed the strongest correlation, the greatest precision, the lowest positive bias, and the second highest 30% accuracy with the lowest observed serum creatinine concentration in the recipient. This study provides a simple means of predicting the lowest serum creatinine concentration following kidney transplantation. However, the sensitivity, specificity and other diagnostic characteristics of the equation need to be determined in a prospective study before this approach can be recommended in routine clinical practice.
| 19,015,653
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Remission of metabolic syndrome following a 15-week low-calorie lifestyle change program for weight loss.
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To evaluate the rate of remission of metabolic syndrome (Met Syn) among patients undergoing a brief, low-calorie lifestyle change weight loss intervention and to compare the baseline characteristics of patients who were remitted and not remitted from Met Syn at post-treatment. Obese adults (N=36) meeting criteria for Met Syn enrolled in an outpatient fee-for-service behavioral weight loss intervention. Participants were assessed on key Met Syn variables (waist circumference, blood pressure, triglycerides, high-density lipoprotein (HDL) cholesterol and fasting blood glucose) at pre- and post-treatment. The majority of patients (61%) responded to treatment after a 9.9% mean weight loss. Although Met Syn responders did not differ significantly from Met Syn non-responders on any baseline Met Syn criterion variable, responders had significantly lower baseline body mass indices (BMI; kg/m(2)) and met criteria for fewer baseline Met Syn variables. As expected, Met Syn responders, compared with Met Syn non-responders, had significantly lower post-treatment waist circumference, systolic and diastolic blood pressures, triglycerides and fasting blood glucose. Patient groups did not differ significantly on weight lost (kg or %), or on the proportion of patients losing > or =10% of initial body weight. In a community population, Met Syn responds to weight loss through a low-calorie lifestyle intervention; for some patients, however, the recommended 10% weight loss may not be enough for Met Syn remission.
| 19,015,662
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Fundamental-mode cutoff in liquid-filled Y-shaped microstructured fibers with Ge-doped core.
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We report on the cutoff characteristics of a Ge-doped Y-shaped microstructured fiber in which the holes are filled with a liquid of refractive index higher than silica but lower than the Ge-doped core. It is found that the cutoff wavelength was very sensitive to temperature variations as a result of the refractive index changes of the liquid. The basic properties of such a fiber permit the fabrication of wideband tunable short-pass filters, as well as temperature sensors with high sensitivity. A temperature sensitivity of 25 nm/degrees C is reported.
| 19,015,673
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Numerical multiplexing and demultiplexing of digital holographic information for remote reconstruction in amplitude and phase.
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We investigate the possibility to multiplexing (Mux) and demultiplexing (de-Mux) numerically digital holograms (DHs) with the aim of optimizing their storage and/or transmission process. The DHs are multiplexed and demultiplexed thanks to the unique property of the digital holography to numerically manage the complex wavefields. We show that it is possible to retrieve correctly quantitative information about the amplitude and phase of one hundred DHs. This result can be useful to transmit efficiently, in terms of reduced amount of data, the DHs from the recording head to a remote display unit.
| 19,015,690
|
Route to the minimum pulse duration in normal-dispersion fiber lasers.
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The factors that control the pulse duration in all-normal-dispersion lasers are identified. To minimize the pulse duration the cavity dispersion should be as small as possible. For fixed dispersion increasing pulse energy leads to shorter pulses with more structured spectra. Experiments performed with ordinary single-mode fiber at 1 microm wavelength agree reasonably with numerical simulations and produce clean approximately 80 fs pulses. The simulations indicate that 30 fs pulses can be reached at higher energies.
| 19,015,693
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Visible photon-avalanche upconversion in Ho3+ singly doped beta-Na(Y1.5Na0.5)F6 under 980 nm excitation.
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Visible IR-to-green photon-avalanche upconversion is reported in an Ho3+ singly doped beta-Na(Y1.5Na0.5)F6 crystal under 980 nm excitation. Upconverted green, red, and IR emissions are observed at 540, 645, and 751 nm, respectively. Temporal evolution and excitation power dependent upconversion intensity are measured, suggesting that a photon-avalanche mechanism is responsible for the upconversion process. It is believed that an efficient cross relaxation (5S2,5I8)-->(5I6,5I6) mainly performs the population of 5I6 excited state, resulting in the intense photon-avalanche upconversion emission in the synthesized samples.
| 19,015,698
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Mid-infrared optical combs from a compact amplified Er-doped fiber oscillator.
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By means of a difference-frequency-generation (DFG) process driven by a two-branch Er-doped fiber laser at a stabilized 100 MHz repetition rate, broadly tunable pulses from 5 to 12 microm are generated with an unprecedented power level of around 100 microW. The mid-IR pulse train is expected to exhibit an harmonic comb structure as a result of the cancellation of the carrier-envelope offset frequency resulting from the DFG process.
| 19,015,704
|
Warmer weather linked to tick attack and emergence of severe rickettsioses.
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The impact of climate on the vector behaviour of the worldwide dog tick Rhipicephalus sanguineus is a cause of concern. This tick is a vector for life-threatening organisms including Rickettsia rickettsii, the agent of Rocky Mountain spotted fever, R. conorii, the agent of Mediterranean spotted fever, and the ubiquitous emerging pathogen R. massiliae. A focus of spotted fever was investigated in France in May 2007. Blood and tissue samples from two patients were tested. An entomological survey was organised with the study of climatic conditions. An experimental model was designed to test the affinity of Rh. sanguineus for biting humans in variable temperature conditions. Serological and/or molecular tools confirmed that one patient was infected by R. conorii, whereas the other was infected by R. massiliae. Dense populations of Rh. sanguineus were found. They were infected with new genotypes of clonal populations of either R. conorii (24/133; 18%) or R. massiliae (13/133; 10%). April 2007 was the warmest since 1950, with summer-like temperatures. We show herein that the human affinity of Rh. sanguineus was increased in warmer temperatures. In addition to the originality of theses cases (ophthalmic involvements, the second reported case of R. massiliae infection), we provide evidence that this cluster of cases was related to a warming-mediated increase in the aggressiveness of Rh. sanguineus, leading to increased human attacks. From a global perspective, we predict that as a result of globalisation and warming, more pathogens transmitted by the brown dog tick may emerge in the future.
| 19,015,724
|
Self-assembly in monoelaidin aqueous dispersions: direct vesicles to cubosomes transition.
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In the present study, synchrotron small-angle X-ray scattering (SAXS) and Cryo-TEM were used to characterize the temperature-induced structural transitions of monoelaidin (ME) aqueous dispersion in the presence of the polymeric stabilizer F127. We prove that the direct transition from vesicles to cubosomes by heating this dispersion is possible. The obtained results were compared with the fully hydrated bulk ME phase. Our results indicate the formation of ME dispersion, which is less stable than that based on the congener monoolein (MO). In addition, the temperature-dependence behavior significantly differs from the fully hydrated bulk phase. SAXS findings indicate a direct L(alpha)-V(2) internal transition in the dispersion. While the transition temperature is conserved in the dispersion, the formed cubosomes with internal Im3m symmetry clearly contain more water and this ordered interior is retained over a wider temperature range as compared to its fully hydrated bulk system. At 25 degrees C, Cryo-TEM observations reveal the formation of most likely closely packed onion-like vesicles. Above the lamellar to non-lamellar phase transition at 65 degrees C, flattened cubosomes with an internal nanostructure are observed. However, they have only arbitrary shapes and thus, their morphology is significantly different from that of the well-shaped analogous MO cubosome and hexosome particles. Our study reveals a direct liposomes-cubosomes transition in ME dispersion. The obtained results suggest that the polymeric stabilizer F127 especially plays a significant role in the membrane fusion processes. F127 incorporates in considerable amount into the internal nanostructure and leads to the formation of a highly swollen Im3m phase.
| 19,015,726
|
Allele dependent silencing of COL1A2 using small interfering RNAs.
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Osteogenesis imperfecta (OI) is generally caused by a dominant mutation in Collagen I, encoded by the genes COL1A1 and COL1A2. To date there is no satisfactory therapy for OI, but inactivation of the mutant allele through small interfering RNAs (siRNA) is a promising approach, as siRNAs targeting each allele of a polymorphism could be used for allele-specific silencing irrespective of the location of the actual mutations. In this study we examined the allele dependent effects of several tiled siRNAs targeting a region surrounding an exonic COL1A2 T/C polymorphism (rs1800222) in heterozygous primary human bone cells. Relative abundances of COL1A2 alleles were determined by cDNA sequencing and overall COL1A2 abundance was analyzed by quantitative PCR. One of the siRNAs decreased overall COL1A2 abundance by 71% of which 75% was due to silencing of the targeted T-allele. In conclusion, allele-preferential silencing of Collagen type I genes may be a future therapeutic approach for OI.
| 19,015,742
|
An innovative method to evaluate the suture compliance in sealing the surgical wound lips.
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The increasing number of surgical procedures performed with local anesthesia, followed by immediate patient discharge from the hospital, emphasizes the need for a tight waterproof suture that is capable of maintaining its tensile strength in the postoperative phase when the wound tumescence, edema due to the anesthetic drug, and surgical trauma disappear. Moreover, the issue of having an accurate surgical wound closure is very relevant in vivo in order to prevent hemorrhage and exogenous microbial infections. This study aimed at designing a new a lab technique that could be used for evaluating the best surgical material. Using such a technique, we compared the wound-lip-sealing properties of three commonly-used suture threads, namely polyurethane, polypropylene, and polyamide. The mechanical properties of same-size suture threads made from polyurethane, polypropylene, and polyamide, were compared in order to define the one that possess the best elastic properties by being able to counteract the tension-relaxation process in the first 12 hours following surgery. The tension holding capacity of the suture materials was measured in both in vivo and in vitro experiments. The surface area of the scar associated with the three different suture threads was measured and compared, and the permeability of the three different suture threads was assessed at 0 minute, 2 minute, 4 minute, 6 minute, and 8 minute- interval. Results showed that polyurethane suture threads had significantly (P < 0.05) better tensile strength, elongation endurance before breakage, and better elasticity coefficient as compared to polypropylene and polyamide suture threads. Moreover, polyurethane suture threads were significantly (P < 0.05) more impermeable as compared to the other two suture thread types (polypropylene and polyamide). This impermeability was also associated with a tighter wound-lip-sealing ability, and with significantly (P < 0.05) less scar formation. Among the main concerns that surgeons, physicians, and patients often have is the development infection, oozing, and scar at the incision site following suturing. This always raises the question about which suture to use to avoid the above problems. This study provides evidence that the new technique developed in our lab could be used to compare the wound-lip sealing properties of different surgical suture threads. Using such a technique, the results show that polyurethane is significantly better than other commonly-used suture threads, like polypropylene and polyamide, in relation to wound sealing and scar formation.
| 19,015,745
|
Protective effects of urinary trypsin inhibitor on systemic inflammatory response induced by lipopolysaccharide.
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Urinary trypsin inhibitor (UTI), a serine protease inhibitor, has been widely used in Japan as a drug for patients with acute inflammatory disorders such as disseminated intravascular coagulation (DIC), shock, and pancreatitis. Recent in vitro studies have demonstrated that serine protease inhibitors may have anti-inflammatory properties beyond their inhibition of neutrophil elastase at the site of inflammation. However, the therapeutic effects of UTI in vivo remain unclear. In this review, we introduce the roles of UTI in the experimental systemic inflammatory response induced by both intraperitoneal and intratracheal administration of lipopolysaccharide using UTI deficient and wild-type mice. Our experiments suggest that UTI can protect against systemic inflammatory response and subsequent organ injury induced by bacterial endotoxin, at least partly, through the inhibition of proinflammatory cytokine and chemokine expression. UTI may therefore present an attractive "rescue" therapeutic option for systemic inflammatory response syndromes such as DIC, acute lung injury, and multiple organ dysfunction.
| 19,015,747
|
Hysteroscopic sterilization: history and current methods.
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For many practicing obstetrician-gynecologists, tubal ligation was the gold standard by which female sterilization techniques were measured. Yet gynecologic surgeons have simultaneously sought to occlude the fallopian tubes transcervically to avoid discomfort and complications associated with transabdominal approaches. In this review, the history of transcervical sterilization is discussed. Past, current, and upcoming techniques are reviewed. This article focuses on interval sterilization techniques, thus removing post-vaginal and post-cesarean delivery tubal ligations from the discussion.
| 19,015,762
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The effects of self-aggregation on the vibrational circular dichroism and optical rotation measurements of glycidol.
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The noncovalent interactions between glycidol molecules in the CDCl3 solvent have been studied by means of vibrational absorption (VA), vibrational circular dichroism (VCD), optical rotation (OR) spectroscopy and density functional theory (DFT) calculations. The concentration dependence of the VA and VCD spectra and OR measurements at five excitation wavelengths, i.e. 589, 578, 546, 436 and 365 nm, has been reported. To model the effects of self-aggregation of glycidol on the measurements, the energetic and conformational properties of the glycidol monomer, dimer and trimer were evaluated and the corresponding VA, VCD and OR spectra were simulated. The results show that at 0.2 M or lower concentrations the self-aggregation of glycidol is negligible since the simulated VA and VCD spectra, with the contribution from only the monomeric glycidol conformers, reproduce well the features in the experimental spectra. At 3.5 M, the binary conformers dominate, while at the intermediate concentration of 1.1 M, both the monomeric and binary conformers are important. The comparison of the experimental and theoretical OR values supports the above conclusions. This work shows the potential of using multiple chiroptical spectroscopic methods in combination with theoretical calculations to probe the self-aggregation process of chiral molecules in solution. Such studies can in turn help to achieve reliably absolute configuration determinations in the cases when chiral molecules self-aggregate profusely.
| 19,015,782
|
Anatomical study of the greater palatine artery and related structures of the palatal vault: considerations for palate as the subepithelial connective tissue graft donor site.
|
Palate is considered as a tissue graft donor site for dental surgical procedures. Therefore, the aim of this study was to investigate the anatomy of palatal structures, such as greater palatine artery, greater palatine foramen, and incisive fossa, in order to consider their topography at planning the graft dimensions and reduce the potential risk of injury of greater palatine artery. Direct inspection of 41 Thai cadavers was performed. The results showed the statistically significant differences as for the length of female and male palates (p = 0.017); however, vertical measurements were equally distributed in examined population. Main location of greater palatine foramen was palatal to the second molar (35.7%), as well as, interproximal to the second and third molars (35.7%) in women, and palatal to the second molar in men (65%). GPA was branching most frequently at the level of first premolar (38%) and at first and second molars together (43%) in women. In men, the branching on the alveolar process side was commonly observed at the level of first and second premolars together (56%), and at the level of second and third molars together (32%). In the area between maxillary first premolar and second molar, it appeared possible to harvest a connective tissue graft measuring at least 5 mm in height. The results of this research will provide the useful data for other comparative studies and for assisting periodontologists in planning the dimensions and harvesting the subepithelial connective tissue grafts from palate.
| 19,015,806
|
Culturable bacteria in glacial meltwater at 6,350 m on the East Rongbuk Glacier, Mount Everest.
|
Culturable bacteria in the glacial meltwater in the ablation zones of glacier at high altitude (6,350 m) on Mt Everest were isolated and identified by 16S rRNA amplification and sequencing. The obtained sequences revealed the presence of members of alpha, beta, and gamma-Proteobacteria, Actinobacteria, and Firmicutes, with the Actinobacteria dominant in the studied habitat. All 16S rRNA sequences were similar to previously determined sequences, ranging from 97 to 99% identical values. The strains isolated from meltwater were distinctly different from those recovered from a cryoconite hole and under glacier habitat. The majority of the isolates' nearest neighbors were from the permafrost, dust, soil, plant, and aqueous environments. The Biolog bioassay and growth test under different temperatures suggested that the culturable bacteria in glacial meltwater could be divided into three categories in terms of their survival strategies: Group I sensitive to temperature change but versatile in utilization of carbon substrates (capable of utilization of about 70% of the Biolog carbon substrates); Group II tolerant to variable temperature and less capable of carbon utilization (less than half of the Biolog carbon species can be used); Group III slow in growth and weak in carbon utilization (only a few Biolog carbon substrates can be used).
| 19,015,814
|
Cloning, sequence analysis, and expression of a gene encoding Chromobacterium sp. DS-1 cholesterol oxidase.
|
Chromobacterium sp. strain DS-1 produces an extracellular cholesterol oxidase that is very stable at high temperatures and in the presence of organic solvents and detergents. In this study, we cloned and sequenced the structural gene encoding the cholesterol oxidase. The primary translation product was predicted to be 584 amino acid residues. The mature product is composed of 540 amino acid residues. The amino acid sequence of the product showed significant similarity (53-62%) to the cholesterol oxidases from Burkholderia spp. and Pseudomonas aeruginosa. The DNA fragment corresponding to the mature enzyme was subcloned in the pET-21d(+) expression vector and expressed as an active product in Escherichia coli. The cholesterol oxidase produced from the recombinant E. coli was purified to homogeneity. The physicochemical properties were similar to those of native enzyme purified from strain DS-1. K(m) and V(max) values of the cholesterol oxidase were estimated from Lineweaver-Burk plots. The V(max)/K(m) ratio of the enzyme was higher than those of commercially available cholesterol oxidases. The circular dichroism spectral analysis of the recombinant DS-1 enzyme and Burkholderia cepacia ST-200 cholesterol oxidase showed that the conformational stability of the DS-1 enzyme was higher than that of B. cepacia ST-200 enzyme at higher temperatures.
| 19,015,844
|
Elementary mode analysis: a useful metabolic pathway analysis tool for characterizing cellular metabolism.
|
Elementary mode analysis is a useful metabolic pathway analysis tool to identify the structure of a metabolic network that links the cellular phenotype to the corresponding genotype. The analysis can decompose the intricate metabolic network comprised of highly interconnected reactions into uniquely organized pathways. These pathways consisting of a minimal set of enzymes that can support steady state operation of cellular metabolism represent independent cellular physiological states. Such pathway definition provides a rigorous basis to systematically characterize cellular phenotypes, metabolic network regulation, robustness, and fragility that facilitate understanding of cell physiology and implementation of metabolic engineering strategies. This mini-review aims to overview the development and application of elementary mode analysis as a metabolic pathway analysis tool in studying cell physiology and as a basis of metabolic engineering.
| 19,015,845
|
Disposition of desipramine, a sensitive cytochrome P450 2D6 substrate, when coadministered with intravenous temsirolimus.
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Intravenous (i.v.) temsirolimus, a novel inhibitor of mammalian target of rapamycin (mTOR), is approved for treatment of renal cell carcinoma. In vitro studies with pooled human liver microsomes showed that temsirolimus and its principal metabolite, sirolimus, inhibit the CYP2D6 isozyme (K(i) = 1.5 and 5 microM, respectively), indicating potential for pharmacokinetic interaction with agents that are substrates of CYP2D6. This 2-period study in healthy subjects investigated the pharmacokinetics of a single oral 50-mg dose of the CYP2D6 substrate desipramine, first without and subsequently with a single coadministered i.v. 25-mg dose of temsirolimus. The study population consisted of 25 males and 1 female; 10 were black, 12 were white, and 4 were of other races. Plasma and whole blood samples were available from all 26 subjects in period 1 following oral desipramine and from 23 subjects in period 2 following oral desipramine and i.v. temsirolimus coadministration. The 90% confidence intervals for least squares geometric mean ratios of desipramine and 2-hydroxy-desipramine C(max), AUC(T), and AUC were within 80-125%, indicating that parameter differences did not manifest into clinically relevant exposure changes. A single i.v. 25-mg dose of temsirolimus, alone or with desipramine, was well tolerated in healthy subjects. A single i.v. 25-mg dose of temsirolimus did not alter disposition of desipramine. Temsirolimus i.v. 25 mg may be safely administered with agents metabolized through the CYP2D6 pathway, but vigilance for drug interaction is warranted in patients with advanced malignancies.
| 19,015,855
|
Protamine-mediated DNA coating remarkably improves bombardment transformation efficiency in plant cells.
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We have developed a method by which remarkably higher efficiencies of transient and stable transformation were achieved in bombardment transformation of plants. Over fivefold increase in transient gus gene expression was achieved when rice or maize suspension cells were bombarded with gold particles coated with plasmid DNA in the presence of protamine instead of the conventional spermidine. A 3.3-fold improvement in stable transformation efficiency was also observed using rice suspension cells with the new coating approach. The coated protamine-plasmid DNA complex resisted degradation by a DNase or by rice cell extract much longer than the spermidine-plasmid DNA complex. The results from this study suggest that protamine protects plasmid DNA longer than spermidine when being delivered inside the cells, probably by forming a nano-scale complex, and thus helps improve the efficiency of particle bombardment-mediated plant transformation.
| 19,015,859
|
Effects of low ambient temperature on heart rate variability during sleep in humans.
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The effects of cold exposure on heart rate variability (HRV) during sleep were examined. Eight male subjects slept under three different conditions: 3 degrees C, 50-80% relative humidity (RH) [3]; 10 degrees C, 50% RH [10]; and 17 degrees C 50% RH [17]. No significant differences were observed in HRV during rapid eye movement sleep (REM) and wakefulness. The ratio of the low frequency (LF) to high frequency component (HF) of HRV (LF/HF) significantly differed among the conditions during stage 2 and slow wave sleep (SWS) that decreased as the ambient temperature decreased. The normalized LF [LF/(LF + HF)] significantly decreased in 3 and 10 than in 17 during SWS. In low ambient temperature, predominant cardiac parasympathetic activity during stage 2 with no significant difference during REM and wakefulness may cause variations in HRV at transition from stage 2 to REM and wakefulness. These results may partly explain the peak in adverse cardiac events during winter.
| 19,015,871
|
Frequent inactivation of RUNX3 by promoter hypermethylation and protein mislocalization in oral squamous cell carcinomas.
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RUNX3 is a functionally important component in transforming growth factor-beta (TGF-beta) mediated signaling pathway. Epigenetic silencing expression of RUNX3, as well as aberrant cytoplasmic retention of RUNX3 protein are causally involved in gastric carcinogenesis. Here, we examined the expression of RUNX3 gene and protein in oral squamous cell carcinomas (OSCCs) and analyzed the methylation status of RUNX3 promoter region. About 10 normal oral mucosa and 30 OSCCs were collected to examine RUNX3 expression by RT-PCR analysis and immunohistochemistry assay using anti-RUNX3 monoclonal antibody R3-6E9. Methylation-specific PCR was carried out on the same specimens to analyze the methylation status of RUNX3 promoter. In addition, the stored paraffin-embedded specimens, including 40 oral leucoplakia (OLK) and 120 OSCCs, were examined by immunohistochemistry assay. RUNX3 gene and protein were underexpressed in OSCCs due to promoter hypermethylation. Protein mislocalization occurred frequently. Both downregulation of RUNX3 protein expression (P = 0.001) and protein mislocalization (P = 0.001) were correlated with the differentiation grades in OSCCs. RUNX3 plays an important role in oral carcinogenesis. It may be a useful diagnostic marker and a potential therapeutic target for OSCC.
| 19,015,875
|
Community use of the amplatzer atrial septal defect occluder: results of the multicenter MAGIC atrial septal defect study.
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The objective of this study was to determine the initial safety and results of unrestricted multi-institution routine community use of the Amplatzer Septal Occluder (ASO) for atrial septal defect (ASD) closure. A multicenter, nonrandomized prospective study was performed in 13 pediatric cardiology centers from November 2004 to September 2007. Data were collected at the time of cardiac catheterization and 1 day postimplant. Four hundred seventy-eight patients underwent cardiac catheterization for ASO device closure of an ASD. The median age was 6 years (range, <1-83 years), and the mean weight was 37.2 kg (range, 2.6-148 kg). Procedural success was 96% (458/478 patients), with deficient rims being the major single reason for failed implantation (9/20). Major and minor complication rates were 1.1% and 4.8%, respectively, and were not different between simple and complex ASD groups. Success at 24 h was 99.4% (333/335) in the simple ASD group and 100% (120/120) in the complex ASD group. The presence of large defects, the presence of multiple defects, the use of multiple devices, and a weight <8 kg were significantly associated with a residual shunt (small to moderate) at 24 h. In conclusion, the ASO device in routine clinical practice for simple and complex ASD closure has an immediate safety and effectiveness profile equal to that reported in the initial pivotal FDA trial for simple ASDs. Based on the evolution in care posed by the ASO and the lack of consensus on patient selection in complex ASDs, this study points out the need to redefine the optimal patient and possibly broaden the indications for device closure of ASDs.
| 19,015,911
|
Accelerated ageing: from mechanism to therapy through animal models.
|
Ageing research benefits from the study of accelerated ageing syndromes such as Hutchinson-Gilford progeria syndrome (HGPS), characterized by the early appearance of symptoms normally associated with advanced age. Most HGPS cases are caused by a mutation in the gene LMNA, which leads to the synthesis of a truncated precursor of lamin A known as progerin that lacks the target sequence for the metallopotease FACE-1/ZMPSTE24 and remains constitutively farnesylated. The use of Face-1/Zmpste24-deficient mice allowed us to demonstrate that accumulation of farnesylated prelamin A causes severe abnormalities of the nuclear envelope, hyper-activation of p53 signalling, cellular senescence, stem cell dysfunction and the development of a progeroid phenotype. The reduction of prenylated prelamin A levels in genetically modified mice leads to a complete reversal of the progeroid phenotype, suggesting that inhibition of protein farnesylation could represent a therapeutic option for the treatment of progeria. However, we found that both prelamin A and its truncated form progerin can undergo either farnesylation or geranylgeranylation, revealing the need of targeting both activities for an efficient treatment of HGPS. Using Face-1/Zmpste24-deficient mice as model, we found that a combination of statins and aminobisphosphonates inhibits both types of modifications of prelamin A and progerin, improves the ageing-like symptoms of these mice and extends substantially their longevity, opening a new therapeutic possibility for human progeroid syndromes associated with nuclear-envelope defects. We discuss here the use of this and other animal models to investigate the molecular mechanisms underlying accelerated ageing and to test strategies for its treatment.
| 19,015,945
|
Analysis of vibroarthrographic signals with features related to signal variability and radial-basis functions.
|
Knee-joint sounds or vibroarthrographic (VAG) signals contain diagnostic information related to the roughness, softening, breakdown, or the state of lubrication of the articular cartilage surfaces. Objective analysis of VAG signals provides features for pattern analysis, classification, and noninvasive diagnosis of knee-joint pathology of various types. We propose parameters related to signal variability for the analysis of VAG signals, including an adaptive turns count and the variance of the mean-squared value computed during extension, flexion, and a full swing cycle of the leg, for the purpose of classification as normal or abnormal, that is, screening. With a database of 89 VAG signals, screening efficiency of up to 0.8570 was achieved, in terms of the area under the receiver operating characteristics curve, using a neural network classifier based on radial-basis functions, with all of the six proposed features. Using techniques for feature selection, the turns counts for the flexion and extension parts of the VAG signals were chosen as the top two features, leading to an improved screening efficiency of 0.9174. The proposed methods could lead to objective criteria for improved selection of patients for clinical procedures and reduce healthcare costs.
| 19,015,987
|
Clinical and epidemiological characteristics of thyroid hemiagenesis: ultrasound screening in patients with thyroid disease and normal population.
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Thyroid hemiagenesis is a rare form of thyroid dysgenesis, in which one thyroid lobe fails to develop. The true prevalence of this rare abnormality is about 0.05-0.2% in normal population. We aimed to determine prevalence of thyroid hemiagenesis in patients with various thyroid disorders and a normal population in a mild to moderate iodine-deficient area. The clinical and thyroid ultrasonography records of 4,833 patients who presented with various thyroid disorders were reviewed. In addition, ultrasonographic data of two large surveys carried out for the community screening of iodine status of children (n = 4,772) and thyroid disorders of adult subjects (n = 2,935) were analyzed. In patients with thyroid disorders, we found 12 cases with thyroid hemiagenesis (0.25%). Thyroid hemiagenesis was due to the agenesis of the left lobe in all cases. The underlying thyroid diseases were Hashimoto's thyroiditis (n = 4), euthyroid multinodular goiter (n = 4), and toxic adenoma (n = 1). Three subjects have no underlying thyroid disease. In ultrasonography screening of normal population, altogether, the absence of the left lobe was detected in only two cases, indicating a true prevalence of thyroid hemiagenesis of 0.025%. None of the reviewed patients had thyroid dysfunction. Our community-based data is in accordance with previous studies in terms of prevalence and male-to-female ratio.
| 19,016,002
|
What is (still not) known of the mechanism by which electroporation mediates gene transfer and expression in cells and tissues.
|
Cell membranes can be transiently permeabilized under application of electric pulses. This treatment allows hydrophilic therapeutic molecules, such as anticancer drugs and DNA, to enter into cells and tissues. This process, called electropermeabilization or electroporation, has been rapidly developed over the last decade to deliver genes to tissues and organs, but there is a general agreement that very little is known about what is really occurring during membrane electropermeabilization. It is well accepted that the entry of small molecules, such as anticancer drugs, occurs mostly through simple diffusion after the pulse while the entry of macromolecules, such as DNA, occurs through a multistep mechanism involving the electrophoretically driven interaction of the DNA molecule with the destabilized membrane during the pulse and then its passage across the membrane. Therefore, successful DNA electrotransfer into cells depends not only on cell permeabilization but also on the way plasmid DNA interacts with the plasma membrane and, once into the cytoplasm, migrates towards the nucleus. The focus of this review is to describe the different aspects of what is known of the mechanism of membrane permeabilization and associated gene transfer and, by doing so, what are the actual limits of the DNA delivery into cells.
| 19,016,008
|
Epidermal growth factor receptor as a predictor of tumor response to preoperative chemoradiation in locally advanced gastric carcinoma.
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The purpose of our study was a retrospective evaluation whether the intensity of epidermal growth factor receptor (EGFR) expression predicts tumor response to preoperative chemoradiotherapy in patients with locally advanced gastric carcinoma. Thirty-six patients with gastric adenocarcinoma (cT2-4 or N+) were studied. Preoperative treatment consisted of 30-45 Gy of gastric irradiation with continuous 5-fluorouracil and weekly cisplatin. Surgical resection was performed 4-6 weeks later. EGFR expression in pretreatment tumor biopsies was assessed by immunohistochemistry. Level of EGFR expression was determined from the intensity and extent of staining. Tumor response was defined as a reduction of at least one T-stage level and/or finding of intense tumor regression in histopathologic examination. Seventeen patients responded to preoperative chemoradiation -- 8 patients (22%) had pathologic complete response, 9 patients (25%) were downstaged. Positive EGFR expression was found in 8 tumors (22%), and represented a significant predictive marker of poor tumor response in multivariate logistic regression analysis (p = 0.015). Response to chemoradiotherapy was found in 60% (16/28) of EGFR negative patients and in 13% (1/8) of EGFR positive patients (p = 0.044). None of the eight EGFR positive patients achieved pathologic complete response in comparison with 8/28 (29%) of patients with EGFR negative staining (p = 0.16). EGFR may represent a molecular marker predictive for poor response to preoperative chemoradiotherapy in locally advanced gastric carcinoma.
| 19,016,018
|
[Local recurrence following hepatic radiofrequency ablation: diagnosis and treatment].
|
Radiofrequency ablation (RFA) is an established treatment in irresectable malignant liver disease. The most severe constraint is re-occurrence at site of ablation. Whereas factors influencing local recurrence rates have been determined, little is known about the timespan within local recurrence (LR) is to be expected, and further treatment options. In the presented trial, RFA was performed using two different types of monopolar devices. All procedures were conducted under general anesthesia. Follow-up examinations took part after 3, 6, 12 months and annually. 149 RFAs in 125 patients were enrolled. Percutaneous access was chosen in 74 cases (50%), laparoscopic in 15 (10%) and open surgical in 60 cases (40%). Indications were primary liver tumors in 99 (67%) and metastases in 50 cases (33%). Overall LR rate was 29.5% on a per-patient- and 19.7% on a per-tumor-basis. The majority of LRs (71%) occurred within 9 months after the RFA despite observations beyond 2 years following the treatment (Figure 1). 75% of LR could be treated by targeted interventions (RFA, n = 18, 53%, laser-induced thermo therapy (LITT), n = 2.6%, brachytherapy, n = 2.6% or transarterial chemoembolisation (TACE), n = 2.6%) or resection (n = 6.18%); 4 patients underwent liver transplantation (12%) (Figure 2). Local recurrence can be considered rather common after RFA. It is observed during the first 3 years of the follow-up period, and schedules have to be designed according to this finding. Follow-on treatment is feasible in approximately 75% of LR. Factors influencing the secondary success of repeated procedures have yet to be determined.
| 19,016,019
|
Treatment options for high-risk T1 bladder cancer: status quo and future perspectives of radiochemotherapy.
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To review the standards and new developments in diagnosis and management of high-risk T1 bladder cancer with emphasis on the role of radiotherapy (RT) and radiochemotherapy (RCT). A systematic review of the literature on developments in diagnosis and management of high-risk T1 bladder cancer was performed. First transurethral resection (TUR), as radical as safely possible, supported by fluorescence cystoscopy, shows higher detection and decreased recurrence rates. An immediate single postoperative instillation with a chemotherapeutic drug reduces the relative risk of recurrence by 40%. A second TUR is recommended to assess residual tumor. For adjuvant intravesical therapy, bacille Calmette-Guérin (BCG) demonstrated the highest efficacy. Early cystectomy should be reserved for selected patients. A recent phase III trial comparing RT versus conservative treatment in T1 G3 tumors could not show any advantage for RT. Data from Erlangen, Germany, using combined RCT in 80% of the patients, compare favorably with most of the contemporary BCG series. Results of intravesical therapy are still unsatisfying and early cystectomy is associated with morbidity and mortality. RT alone proved not superior to other conservative treatment strategies. However, data on RCT are promising and demonstrate an alternative to intravesical therapy and radical cystectomy.
| 19,016,022
|
Seed displacements after permanent brachytherapy for prostate cancer in dependence on the prostate level.
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To evaluate seed displacements after permanent prostate brachytherapy considering different prostate levels. In 61 patients, postimplant CT scans were performed 1 day and 1 month after an implant with stranded seeds. Seed and prostate surface displacements were determined relative to pelvic bones. Four groups of seed locations were selected: seeds at the base (n = 305; B), at the apex (n = 305; A), close to the urethra (n = 306; U), and close to the rectal wall (n = 204; R). The length of two strands (always containing four seeds) per patient was measured in all CT scans and compared. The largest inferior seed displacements were found at the base: mean 5.3 mm (B), 2.2 mm (A), 2.7 mm (U), 3.3 mm (R; p < 0.001). Posterior displacements predominated both at the base and the central region: mean 2.2 mm (B), 2.0 mm (U), 0.8 mm (A), -0.6 mm (R; p < 0.001). With a decreasing edema between day 1 and 30 (mean prostate volume of 51 cm(3) vs. 41 cm(3); p < 0.001), a mean caudal prostate base displacement of 3.9 mm was found, whereas the mean inward displacement ranged from 1.2 to 1.6 mm at the remaining borders (lateral, anterior, posterior, apical). The analysis of the strand lengths revealed an implant compression between day 1 and 30 (mean 1.7 mm; p < 0.001). The largest prostate tissue and seed displacements were observed at the prostate base, associated with an implant compression. Predominantly inferior and posterior displacements implicate consequential smaller preplanning margins at the apex and the posterior prostate.
| 19,016,041
|
Adipose tissue and skeletal muscle plasticity modulates metabolic health.
|
Obesity, accumulation of adipose tissue, develops when energy intake exceeds energy expenditure. Adipose tissue is essential for buffering the differences between energy intake and expenditure by accumulating lipids while skeletal muscle is the energy burning machine. Here we adopted the concept that (i) adipose tissue ability to regulate the storage capacity for lipids as well as (ii) dynamic regulation of muscle and adipose tissue secretory and metabolic activity is important for maintaining the metabolic health. This might be at least in part related to tissue plasticity, a phenomenon enabling dynamic modulation of the tissue phenotype in different physiological and pathophysiological situations. Recent advances in our understanding of the complex endocrine function of adipose tissue in regulating lipid metabolism, adipogenesis, angiogenesis, extracellular matrix remodelling, inflammation and oxidative stress prompted us to review the role of tissue plasticity--dynamic changes in adipose tissue and skeletal muscle metabolic and endocrine phenotype--in determining the difference between metabolic health and disease.
| 19,016,045
|
Maternal coherence in the Adult Attachment Interview is linked to maternal reminiscing and to children's self concept.
|
The role of maternal attachment representations in mother-child reminiscing and children's self concept was assessed in a sample of 31 New Zealand mothers and their 5.5-year-old children. Mothers participated in the Adult Attachment Interview (AAI; Main, Goldwyn, & Hesse, 2002) and reminisced about everyday past events with their children. Children participated in the Children's Self View Questionnaire (Eder, 1990) to measure interpersonal and intrapersonal aspects of their self concept. Maternal coherence on the AAI was positively correlated with mothers' elaborative reminiscing and with interpersonal aspects of children's self concept. Mothers' states of mind with respect to attachment may enable open and elaborative reminiscing with their children, and may also indirectly lead to children's self-concept development.
| 19,016,052
|
Ethylene glycol poisoning presenting with a falsely elevated lactate level.
|
Early diagnosis of ethylene glycol poisoning is crucial in order to prevent morbidity and mortality. However, diagnosis can sometimes be delayed because of the false elevation of lactate in some chemistry analyzers as a result of the interference of glycolate, a metabolite of ethylene glycol. We present a case of ethylene glycol poisoning presenting with a falsely elevated lactate level on a blood gas analyzer in the emergency department. Given the fact that nowadays there is a marked increase in use of point-of-care analyzers, one should be aware of possible false readings since they use different methods of measuring compared with clinical chemistry analyzers. On the other hand, measuring a "lactate gap" using two different technologies, only one of which is sensitive to glycolate, could be a clinically efficient way to make the diagnosis of advanced ethylene glycol poisoning in the emergency department or other critical care setting.
| 19,016,054
|
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