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Find video protocols related to scientific articles indexed in Pubmed.
The complete mitochondrial genome of the mudsnail Cipangopaludina cathayensis (Gastropoda: Viviparidae).
Mitochondrial DNA
PUBLISHED: 10-17-2014
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Abstract We present the complete mitochondrial genome of Cipangopaludina cathayensis in this study. The mitochondrial genome is 17,157?bp in length, containing 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes. All of them are encoded on the heavy strand except 7 tRNA genes on the light strand. Overall nucleotide compositions of the light strand are 44.51% of A, 26.74% of T, 20.48% of C and 8.28% of G. All the protein-coding genes start with ATG initiation codon except ATP6 with ATA and ND4 with TTG, and 2 types of termination codons are TAA (ATP6, ND2, COX1, COX2, ATP8, ND1, ND6, Cytb, COX3, ND4) and TAG (ND4L, ND5, ND3). There are 29 intergenic spacers and 5 gene overlaps. The tandem repeat sequences are observed in COX2, tRNA(Asp), ATP6, tRNA(Cys), S-rRNA, ND1, Cytb, ND4 and COX3 genes. Gene arrangement and distribution are different from the typical vertebrates. The absence of D-loop is consistent with the Gastropoda, but at least one lengthy non-coding region is essential regulatory element for the initiation of transcription and replication.
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The complete mitochondrial genome of the giant African snail Achatina fulica (Mollusca: Achatinidae).
Mitochondrial DNA
PUBLISHED: 09-19-2014
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Abstract We present the complete mitochondrial genome of the Achatina fulica in this study. The results show that the mitochondrial genome is 15,057?bp in length, which is comprised of 13 protein-coding genes, 2 rRNA genes, 21 tRNA genes. The nucleotide compositions of the light strand are 35.47% of A, 27.97% of T 19.46% of C, and 17.10% of G. Except the ND3, 7 tRNA, ATP6, ATP8, COX3 and 12S-rRNA on the light strand, the rest are encoded on the heavy strand. Five types of inferred initiation codons are ATA (ND1, ND5), GTG (ND6), ATG (COX3, COX2), ATT (ND4) and TTG (COX1, ND2, ND3, ND4L, ATP6, ATP8, Cytb), and 3 types of inferred termination codons are T (COX3, ND2), TAA (ND1, ND4L, ND5, ND6, ATP6), and TAG (ND3, ND4, COX1, COX2, Cytb, ATP8). There are 24 intergenic spacers and 6 gene overlaps. The tandem repeat sequence (total 52?bp) of (AATAATT)n is observed in 16S-rRNA. Gene arrangement and distribution are inconsistent with the typical vertebrates.
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[Gender differences of Pomacea canaliculata in cold tolerance].
Ying Yong Sheng Tai Xue Bao
PUBLISHED: 09-17-2014
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In order to understand whether Pomacea canaliculata exhibits gender differences in cold resistance, the survival rate and supercooling point of both P. canaliculata females and males were investigated at low temperature. The changes in physiological and biochemical indices in the body of P. canaliculata before and after the winter were also examined. The results showed that the mean supercooling point of females was -6.83 degrees C, which was significantly lower than that of the males (-6.26 degrees C). With the arrival of winter, the amounts of bound water, glycerol, lipid and glycogen of P. canaliculata increased, while the amount of free water decreased. These indices except glycerol showed significant differences between males and females. More than 90% non-acclimated snails could survive at the temperature of 10 degrees C for 7 days. The survival rate of snails exhibited significant differences in sex and shell height after being transferred at 5 degrees C for 7 days. In addition, more females were recorded during field investigation. Based on these results, we concluded that the females had higher cold tolerance than the males. This finding would provide some references for further investigation of ecological adaptation, natural sex ratio and cold tolerance mechanisms of P. canaliculata.
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[Screening bioactive compounds inhibiting influenza virus from isatidis radix by ultrafiltration mass spectrometry].
Zhongguo Zhong Yao Za Zhi
PUBLISHED: 09-11-2014
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In vitro neuraminidase inhibition assays and ultrafiltration liquid chromatography with diodearray detector coupled to time of flight mass spectrometer (UPLC-DAD-TOF-MS) were combined to screen bioactive compounds inhibiting neuraminidase from Isatidis Radix. By comparing the compounds from Isatidis Radix before and after ultrafiltration, we found that arginine, goitrin and adenosinea can bind with neuraminidase, and the binding degree of the three compounds were (36.23 +/- 1.12)%, (32.54 +/- 1.02)% and (9.38 +/- 0.47)%, respectively. The IC50 of arginine and goitrin were (1.16 +/- 0.02), (1.20 +/- 0.02) g x L(-1), respectively. While the IC50 of adenosinea was higher than 500 g x L(-1). The results showed that arginine and goitrin might be the main compounds with antiviral activity of Isatidis Radix. This study may provide a useful method for the screening of bioactive compounds and quality control of Isatidis Radix.
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Reinnervated nerves contribute to the secretion function and regeneration of denervated submandibular glands in rabbits.
Eur. J. Oral Sci.
PUBLISHED: 09-10-2014
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This study aimed to investigate the contribution of redistributed nerves in the secretory function and regeneration of a denervated submandibular gland (SMG). The postganglionic parasympathetic and sympathetic denervated SMGs of rabbits were wrapped in polyester or acellular dermal matrices to block nerve regeneration either partially or completely. Submandibular glands were removed 4, 8, 16, and 24 wk after the operation and examined histologically. Furthermore, the aquaporin-5 (AQP5), muscarinic-3 (M3), and ?1-adrenergic receptors were evaluated by immunofluorescence and western blot analysis. After denervation, salivary flow was decreased and acinar cells were atrophic, and the expression levels of the M3, ?1-adrenergic, and AQP5 receptors were decreased. However, both impaired secretion function and atrophic parenchyma were gradually ameliorated with the growing redistribution of parasympathetic and sympathetic nerves. Apoptosis was markedly inhibited and expression of the M3, ?1-adrenergic, and AQP5 receptors was increased after reinnervation. In contrast, SMGs without reinnervated nerves maintained hyposecretion and atrophic parenchyma. In conclusion, reinnervated nerves in a rabbit's denervated SMG played an important role in the secretion function and regeneration of SMGs via up-regulation of the expression of neurotransmitter receptors and AQP5.
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Acetylome analysis reveals diverse functions of lysine acetylation in Mycobacterium tuberculosis.
Mol. Cell Proteomics
PUBLISHED: 09-01-2014
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The lysine acetylation of proteins is a reversible posttranslational modification that plays a critical regulatory role in both eukaryotes and prokaryotes. Mycobacterium tuberculosis is a facultative intracellular pathogen and the causative agent of tuberculosis. Increasing evidence shows that lysine acetylation may play an important role in the pathogenesis of M. tuberculosis. However, only a few acetylated proteins of M. tuberculosis are known, presenting a major obstacle to understanding the functional roles of reversible lysine acetylation in this pathogen. We performed a global acetylome analysis of M. tuberculosis H37Ra by combining protein/peptide prefractionation, antibody enrichment, and LC-MS/MS. In total, we identified 226 acetylation sites in 137 proteins of M. tuberculosis H37Ra. The identified acetylated proteins were functionally categorized into an interaction map and shown to be involved in various biological processes. Consistent with previous reports, a large proportion of the acetylation sites were present on proteins involved in glycolysis/gluconeogenesis, the citrate cycle, and fatty acid metabolism. A NAD+-dependent deacetylase (MRA_1161) deletion mutant of M. tuberculosis H37Ra was constructed and its characterization showed a different colony morphology, reduced biofilm formation, and increased tolerance of heat stress. Interestingly, lysine acetylation was found, for the first time, to block the immunogenicity of a peptide derived from a known immunogen, HspX, suggesting that lysine acetylation plays a regulatory role in immunogenicity. Our data provide the first global survey of lysine acetylation in M. tuberculosis. The dataset should be an important resource for the functional analysis of lysine acetylation in M. tuberculosis and facilitate the clarification of the entire metabolic networks of this life-threatening pathogen.
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[A method of screening the antitumor lead compounds based on the dynamic bio-response profile of cells].
Yao Xue Xue Bao
PUBLISHED: 08-26-2014
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The study is to report the establishment of a method of screening the antitumor compounds based on the dynamic bio-response profile of cells to make up for the shortages of conventional end-point tests such as tedious operation and low sensitivity. Based on the principle of electric impedance of cells, the real-time cell electronic sensing (RT-CES) system was used to monitor the effect of epirubicin (EPI), cisplatinum (DDP) and carboplatin (CBP) on the growth of HepG2 cells, with the cell index (CI), half maximal inhibitory concentration (IC50) and detachment curve as evaluation indexes. Meanwhile, cell counting kit-8 (CCK-8) and microscopy were applied for verification. The results showed that CI curve could sensitively real-time profile the inhibitory effect of model drugs on HepG2 cells. The IC50 of EPI, DDP and CBP were 0.53 +/- 0.04, 9.79 +/- 0.26 and 597.00 +/- 3.79 microg x mL(-1), respectively. What's more, the significant differences of detachment curves of the three drugs indicated that their functional mechanisms might be different, this is consistent with the literature. The RT-CES system with non-invasive, label-free and real-time characteristics could be used to monitor the bio-response profile of the three drugs to HepG2 cells, allowing to qualitatively and quantitatively distinguish the antitumor activities of the three drugs, and could be a complementary method for the present screening of antitumor compounds.
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Design and synthesis of novel 1,2,3-triazole-pyrimidine hybrids as potential anticancer agents.
Eur J Med Chem
PUBLISHED: 08-05-2014
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A series of novel 1,2,3-triazole-pyrimidine hybrids were designed, synthesized and evaluated for their anticancer activity against four selected cancer cell lines (MGC-803, EC-109, MCF-7 and B16-F10). Most of the synthesized compounds exhibited moderate to good activity against all the cancer cell lines selected. Compound 17 showed the most excellent anticancer activity with single-digit micromolar IC50 values ranging from 1.42 to 6.52 ?M. Further mechanism studies revealed that compound 17 could obviously inhibit the proliferation of EC-109 cancer cells by inducing apoptosis and arresting the cell cycle at G2/M phase.
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The S28H mutation on mNeptune generates a brighter near-infrared monomeric fluorescent protein with improved quantum yield and pH-stability.
Acta Biochim. Biophys. Sin. (Shanghai)
PUBLISHED: 07-25-2014
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For living deep-tissue imaging, the optical window favorable for light penetration is in near-infrared wavelengths, which requires fluorescent proteins with emission spectra in the near-infrared region. Here, we report that a single mutant Ser28His of mNeptune with a near-infrared (?650 nm) emission maxima of 652 nm is found to improve the brightness, photostability, and pH stability when compared with its parental protein mNeptune, while it remains as a monomer, demonstrating that there is still plenty of room to improve the performance of the existing near infrared fluorescence proteins by directed evolution.
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Detection of Bacillus anthracis spores by super-paramagnetic lateral-flow immunoassays based on "Road Closure"
Biosens Bioelectron
PUBLISHED: 07-01-2014
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Detection of Bacillus anthracis in the field, whether as a natural infection or as a biothreat remains challenging. Here we have developed a new lateral-flow immunochromatographic assay (LFIA) for B. anthracis spore detection based on the fact that conjugates of B. anthracis spores and super-paramagnetic particles labeled with antibodies will block the pores of chromatographic strips and form retention lines on the strips, instead of the conventionally reported test lines and control lines in classic LFIA. As a result, this new LFIA can simultaneously realize optical, magnetic and naked-eye detection by analyzing signals from the retention lines. As few as 500-700 pure B. anthracis spores can be recognized with CV values less than 8.31% within 5min of chromatography and a total time of 20min. For powdery sample tests, this LFIA can endure interference from 25% (w/v) milk, 10% (w/v) baking soda and 10% (w/v) starch without any sample pre-treatment, and has a corresponding detection limit of 6×10(4) spores/g milk powder, 2×10(5) spores/g starch and 5×10(5) spores/g baking soda. Compared with existing methods, this new approach is very competitive in terms of sensitivity, specificity, cost and ease of operation. This proof-of-concept study can also be extended for detection of many other large-sized analytes.
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Standardized treatment of Chinese medicine decoction for cancer pain patients with opioid-induced constipation: a multi-center prospective randomized controlled study.
Chin J Integr Med
PUBLISHED: 06-28-2014
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To observe the efficacy and the influence on quality of life (QOL) of syndrome differentiation treatment with Chinese medicine (CM) for opioid-induced constipation as well as the safety and influence on analgesic effect of opioids.
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Dynamic monitoring of the cytotoxic effects of protoberberine alkaloids from Rhizoma Coptidis on HepG2 cells using the xCELLigence system.
Chin J Nat Med
PUBLISHED: 06-28-2014
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To investigate the cytotoxic effects of the six protoberberine alkaloids (PAs) from Rhizoma Coptidis on HepG2 cells.
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[A novel method for testing sterility of injections based on biothermodynamics].
Yao Xue Xue Bao
PUBLISHED: 06-26-2014
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This study aims at trying to establish a novel method of sterility test for injections based on biothermodynamics, in order to overcome the deficiencies of routine sterility tests such as long detecting cycle, low sensitivity and prone to misjudgments. A biothermodynamics method was adopted to rapidly detect the microorganism contamination of injections by monitoring the heat metabolism during the growth of microbe. The growth rate equal to or greater than zero and the heat power difference of P(i) and P(0) with three folds higher than the noise of baseline were chosen as indexes to study the heat change rule of microbe. In this way, the effectiveness of the new method to detect strains required by conventional sterility test or in injection samples was also investigated. Results showed that the Gram-positive bacteria, Gram-negative bacteria and fungi demanded by sterility testing methodology could be detected by biothermodynamics method within 10 hours, with the sensitivity lower than 100 CFU x mL(-1). Meanwhile, this method was successfully applied to the sterility test of Compound Yinchen injection (FFYC), Shuanghuanglian powder injection (SHL) and Compound Triamcinolone injection (TAND) which were sterilized with different degrees. Therefore, the biothermodynamics method, with advantages of fast detection and high sensitivity, could be a complementary solution for conventional sterility tests.
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Activity of Earthworm in Latosol Under Simulated Acid Rain Stress.
Bull Environ Contam Toxicol
PUBLISHED: 06-01-2014
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Acid rain is still an issue of environmental concerns. This study investigated the impacts of simulated acid rain (SAR) upon earthworm activity from the Latosol (acidic red soil). Laboratory experiment was performed by leaching the soil columns grown with earthworms (Eisenia fetida) at the SAR pH levels ranged from 2.0 to 6.5 over a 34-day period. Results showed that earthworms tended to escape from the soil and eventually died for the SAR at pH = 2.0 as a result of acid toxicity. The catalase activity in the earthworms decreased with the SAR pH levels, whereas the superoxide dismutases activity in the earthworms showed a fluctuate pattern: decreasing from pH 6.5 to 5.0 and increasing from pH 5.0 to 4.0. Results implied that the growth of earthworms was retarded at the SAR pH ? 3.0.
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Sustained running in rats administered corticosterone prevents the development of depressive behaviors and enhances hippocampal neurogenesis and synaptic plasticity without increasing neurotrophic factor levels.
Cell Transplant
PUBLISHED: 05-13-2014
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We have previously shown that voluntary running acts as an anxiolytic and ameliorates deficits in hippocampal neurogenesis and spatial learning. It also reduces depression-like behaviors that are normally observed in rats that were administered either low (30 mg/kg) or moderate (40 mg/kg) doses of corticosterone (CORT). However, the protective effects of running were absent in rats treated with a high (50 mg/kg) dose of CORT. We examined whether allowing animals to exercise for 2 weeks prior and/or concurrently with the administration of 50 mg/kg CORT treatment could have similar protective effects. We examined hippocampal neurogenesis using immunohistochemical staining of proliferative and survival cells with the thymidine analogs (BrdU, CIdU, and IdU). In addition, we monitored synaptic protein expression and quantified the levels of neurotrophic factors in these animals as well as performing behavioral analyses (forced swim test and sucrose preference test). Our results indicate that the depressive phenotype and reductions in neurogenesis that normally accompany high CORT administration could only be prevented by allowing animals to exercise both prior to and concurrently with the CORT administration period. These animals also showed increases in both synaptophysin and PSD-95 protein levels, but surprisingly, neither brain-derived neurotrophic factor (BDNF) nor insulin-like growth factor 1 (IGF-1) levels were increased in these animals. The results suggest that persistent exercise can strengthen resilience to stress by promoting hippocampal neurogenesis and increasing synaptic protein levels, thereby reducing the deleterious effects of stress.
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Water Extract of Fructus Hordei Germinatus Shows Antihyperprolactinemia Activity via Dopamine D2 Receptor.
Evid Based Complement Alternat Med
PUBLISHED: 05-12-2014
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Objective. Fructus Hordei Germinatus is widely used in treating hyperprolactinemia (hyperPRL) as a kind of Chinese traditional herb in China. In this study, we investigated the anti-hyperPRL activity of water extract of Fructus Hordei Germinatus (WEFHG) and mechanism of action. Methods. Effect of WEFHG on serum prolactin (PRL), estradiol (E2), progesterone (P), follicle-stimulating hormone (FSH), luteinizing hormone (LH), and hypothalamus protein kinase A (PKA) and cyclic adenosine monophosphate (cAMP) levels of hyperPRL rats were investigated. And effect of WEFHG on PRL secretion, D2 receptors, and dopamine transporters (DAT) was studied in MMQ, GH3, and PC12 cells, respectively. Results. WEFHG reduced the secretion of PRL in hyperPRL rats effectively. In MMQ cell, treatment with WEFHG at 1-5?mg/mL significantly suppressed PRL secretion and synthesis. Consistent with a D2-action, WEFHG did not affect PRL in rat pituitary lactotropic tumor-derived GH3 cells that lack the D2 receptor expression but significantly increased the expression of D2 receptors and DAT in PC12 cells. In addition, WEFHG reduced the cAMP and PKA levels of hypothalamus in hyperPRL rats significantly. Conclusions. WEFHG showed anti-hyperPRL activity via dopamine D2 receptor, which was related to the second messenger cAMP and PKA.
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In vivo imaging of protein-protein and RNA-protein interactions using novel far-red fluorescence complementation systems.
Nucleic Acids Res.
PUBLISHED: 05-09-2014
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Imaging of protein-protein and RNA-protein interactions in vivo, especially in live animals, is still challenging. Here we developed far-red mNeptune-based bimolecular fluorescence complementation (BiFC) and trimolecular fluorescence complementation (TriFC) systems with excitation and emission above 600 nm in the 'tissue optical window' for imaging of protein-protein and RNA-protein interactions in live cells and mice. The far-red mNeptune BiFC was first built by selecting appropriate split mNeptune fragments, and then the mNeptune-TriFC system was built based on the mNeptune-BiFC system. The newly constructed mNeptune BiFC and TriFC systems were verified as useful tools for imaging protein-protein and mRNA-protein interactions, respectively, in live cells and mice. We then used the new mNeptune-TriFC system to investigate the interactions between human polypyrimidine-tract-binding protein (PTB) and HIV-1 mRNA elements as PTB may participate in HIV mRNA processing in HIV activation from latency. An interaction between PTB and the 3'long terminal repeat region of HIV-1 mRNAs was found and imaged in live cells and mice, implying a role for PTB in regulating HIV-1 mRNA processing. The study provides new tools for in vivo imaging of RNA-protein and protein-protein interactions, and adds new insight into the mechanism of HIV-1 mRNA processing.
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Construction of a chimeric lysin Ply187N-V12C with extended lytic activity against staphylococci and streptococci.
Microb Biotechnol
PUBLISHED: 05-05-2014
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Developing chimeric lysins with a wide lytic spectrum would be important for treating some infections caused by multiple pathogenic bacteria. In the present work, a novel chimeric lysin (Ply187N-V12C) was constructed by fusing the catalytic domain (Ply187N) of the bacteriophage lysin Ply187 with the cell binding domain (146-314aa, V12C) of the lysin PlyV12. The results showed that the chimeric lysin Ply187N-V12C had not only lytic activity similar to Ply187N against staphylococcal strains but also extended its lytic activity to streptococci and enterococci, such as Streptococcus?dysgalactiae, Streptococcus agalactiae, Streptococcus pyogenes, Enterococcus faecium and Enterococcus faecalis, which Ply187N could not lyse. Our work demonstrated that generating novel chimeric lysins with an extended lytic spectrum was feasible through fusing a catalytic domain with a cell-binding domain from lysins with lytic spectra across multiple genera.
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Acs is essential for propionate utilization in Escherichia coli.
Biochem. Biophys. Res. Commun.
PUBLISHED: 04-29-2014
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Bacteria like Escherichia coli can use propionate as sole carbon and energy source. All pathways for degradation of propionate start with propionyl-CoA. However, pathways of propionyl-CoA synthesis from propionate and their regulation mechanisms have not been carefully examined in E. coli. In this study, roles of the acetyl-CoA synthetase encoding gene acs and the NAD(+)-dependent protein deacetylase encoding gene cobB on propionate utilization in E. coli were investigated. Results from biochemical analysis showed that, reversible acetylation also modulates the propionyl-CoA synthetase activity of Acs. Subsequent genetic analysis revealed that, deletion of acs in E. coli results in blockage of propionate utilization, suggesting that acs is essential for propionate utilization in E. coli. Besides, deletion of cobB in E. coli also results in growth defect, but only under lower concentrations of propionate (5mM and 10mM propionate), suggesting the existence of other propionyl-CoA synthesis pathways. In combination with previous observations, our data implies that, for propionate utilization in E. coli, a primary amount of propionyl-CoA seems to be required, which is synthesized by Acs.
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Estimating impacts of land use on groundwater quality using trilinear analysis.
Environ Monit Assess
PUBLISHED: 04-27-2014
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Groundwater is connected to the landscape above and is thus affected by the overlaying land uses. This study evaluated the impacts of land uses upon groundwater quality using trilinear analysis. Trilinear analysis is a display of experimental data in a triangular graph. Groundwater quality data collected from agricultural, septic tank, forest, and wastewater land uses for a 6-year period were used for the analysis. Results showed that among the three nitrogen species (i.e., nitrate and nitrite (NO(x)), dissolved organic nitrogen (DON), and total organic nitrogen (TON)), NO(x) had a high percentage and was a dominant species in the groundwater beneath the septic tank lands, whereas TON was a major species in groundwater beneath the forest lands. Among the three phosphorus species, namely the particulate phosphorus (PP), dissolved ortho phosphorus (PO4(3-)) and dissolved organic phosphorus (DOP), there was a high percentage of PP in the groundwater beneath the septic tank, forest, and agricultural lands. In general, Ca was a dominant cation in the groundwater beneath the septic tank lands, whereas Na was a dominant cation in the groundwater beneath the forest lands. For the three major anions (i.e., F(-), Cl(-), and SO4(2-)), F(-) accounted for <1% of the total anions in the groundwater beneath the forest, wastewater, and agricultural lands. Impacts of land uses on groundwater Cd and Cr distributions were not profound. This study suggests that trilinear analysis is a useful technique to characterize the relationship between land use and groundwater quality.
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Phosphoproteomic analysis provides novel insights into stress responses in Phaeodactylum tricornutum, a model diatom.
J. Proteome Res.
PUBLISHED: 04-18-2014
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Protein phosphorylation on serine, threonine, and tyrosine (Ser/Thr/Tyr) is well established as a key regulatory posttranslational modification used in signal transduction to control cell growth, proliferation, and stress responses. However, little is known about its extent and function in diatoms. Phaeodactylum tricornutum is a unicellular marine diatom that has been used as a model organism for research on diatom molecular biology. Although more than 1000 protein kinases and phosphatases with specificity for Ser/Thr/Tyr residues have been predicted in P. tricornutum, no phosphorylation event has so far been revealed by classical biochemical approaches. Here, we performed a global phosphoproteomic analysis combining protein/peptide fractionation, TiO(2) enrichment, and LC-MS/MS analyses. In total, we identified 264 unique phosphopeptides, including 434 in vivo phosphorylated sites on 245 phosphoproteins. The phosphorylated proteins were implicated in the regulation of diverse biological processes, including signaling, metabolic pathways, and stress responses. Six identified phosphoproteins were further validated by Western blotting using phospho-specific antibodies. The functions of these proteins are discussed in the context of signal transduction networks in P. tricornutum. Our results advance the current understanding of diatom biology and will be useful for elucidating the phosphor-relay signaling networks in this model diatom.
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A systemic analysis of S-1 regimens for treatment of patients with colon cancer.
Asian Pac. J. Cancer Prev.
PUBLISHED: 04-11-2014
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Fluorouracil-based regimens have been widely accepted and recommended in the guidelines for treating patients with early or advanced staged colon cancer, although results are controversial. Here we performed a systemic analysis to evaluate the impact of S-1 based regimens on response and survival of patients with colon cancer.
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Rapid detection of New Delhi metallo-?-lactamase gene and variants coding for carbapenemases with different activities by use of a PCR-based in vitro protein expression method.
J. Clin. Microbiol.
PUBLISHED: 03-26-2014
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New Delhi metallo-?-lactamase (NDM)-producing bacteria are considered potential global health threats. It is necessary to monitor NDM-1 and its variants in clinical isolates in order to understand the NDM-1 epidemic and the impact of its variants on ?-lactam resistance. To reduce the lengthy time needed for cloning and expression of NDM-1 variants, a novel PCR-based in vitro protein expression (PCR-P) method was used to detect blaNDM-1 and its variants coding for carbapenemases with different activities (functional variants). The PCR-P method combined a long-fragment real-time quantitative PCR (LF-qPCR) with in vitro cell-free expression to convert the blaNDM-1 amplicons into NDM for carbapenemase assay. The method could screen for blaNDM-1 within 3 h with a detection limit of 5 copies and identify functional variants within 1 day. Using the PCR-P to analyze 5 recent blaNDM-1 variants, 2 functional variants, blaNDM-4 and blaNDM-5, were revealed. In the initial testing of 23 clinical isolates, the PCR-P assay correctly found 8 isolates containing blaNDM-1. This novel method provides the first integrated approach for rapidly detecting the full-length blaNDM-1 and revealing its functional variants in clinical isolates.
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Dorsal raphe neurons signal reward through 5-HT and glutamate.
Neuron
PUBLISHED: 03-25-2014
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The dorsal raphe nucleus (DRN) in the midbrain is a key center for serotonin (5-hydroxytryptamine; 5-HT)-expressing neurons. Serotonergic neurons in the DRN have been theorized to encode punishment by opposing the reward signaling of dopamine neurons. Here, we show that DRN neurons encode reward, but not punishment, through 5-HT and glutamate. Optogenetic stimulation of DRN Pet-1 neurons reinforces mice to explore the stimulation-coupled spatial region, shifts sucrose preference, drives optical self-stimulation, and directs sensory discrimination learning. DRN Pet-1 neurons increase their firing activity during reward tasks, and this activation can be used to rapidly change neuronal activity patterns in the cortex. Although DRN Pet-1 neurons are often associated with 5-HT, they also release glutamate, and both neurotransmitters contribute to reward signaling. These experiments demonstrate the ability of DRN neurons to organize reward behaviors and might provide insights into the underlying mechanisms of learning facilitation and anhedonia treatment.
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Intrathecal administration of mesenchymal stem cells reduces the reactive oxygen species and pain behavior in neuropathic rats.
Korean J Pain
PUBLISHED: 03-12-2014
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Neuropathic pain induced by spinal or peripheral nerve injury is very resistant to common pain killers, nerve block, and other pain management approaches. Recently, several studies using stem cells suggested a new way to control the neuropatic pain. In this study, we used the spinal nerve L5 ligation (SNL) model to investigate whether intrathecal rat mesenchymal stem cells (rMSCs) were able to decrease pain behavior, as well as the relationship between rMSCs and reactive oxygen species (ROS).
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Synthesis and preliminary biological evaluation of 1,2,3-triazole-Jaspine B hybrids as potential cytotoxic agents.
Eur J Med Chem
PUBLISHED: 03-08-2014
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Two series of more available novel 1,2,3-triazole-Jaspine B hybrids were efficiently synthesized employing click chemistry approach and evaluated for their cytotoxic activities against three human cancer cell lines (EC-9706, MGC-803 and MCF-7). Among them, compound 14h showed excellent inhibition against MCF-7 (IC50 = 1.93 ?M) and was more potent than 5-Fu and Jaspine B against all three cancer cell lines. Further investigation of apoptosis assay and cell cycle analysis demonstrated that compound 14h caused cellular early and late apoptosis and arrested the cell cycle at G2/M phase in a concentration- and time-independent manner. This was the first report about the synthesis and in vitro cytotoxic evaluation of 1,2,3-triazole-Jaspine B hybrids.
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Comparison of reversed-phase liquid chromatography and hydrophilic interaction chromatography for the fingerprint analysis of Radix isatidis.
J Sep Sci
PUBLISHED: 02-21-2014
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Radix isatidis is a famous anti-influenza virus herbal medicine traditionally taken as a water decoction. However, the chemical fingerprint analysis of Radix isatidis is dominantly based on RPLC, from which it is difficult to obtain fingerprint information of hydrophilic compounds. Here, we developed the separation of Radix isatidis by RPLC and hydrophilic interaction chromatography, comparing the traditional RPLC fingerprint with the hydrophilic interaction chromatography fingerprint. Besides, an anti-viral assay of Radix isatidis was conducted to evaluate its efficacy. The fingerprint-efficacy relationships between the fingerprints and the anti-viral activity were further investigated with principal component regression analysis. The results showed that the anti-viral activity correlated better with the hydrophilic interaction chromatography fingerprint than with the RPLC fingerprint. This study indicates that hydrophilic interaction chromatography could not only be a complementary method to increase the fingerprint coverage of conventional RPLC fingerprint, but also can better represent the efficacy and quality of Radix isatidis.
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3-Phenylpropanoic acid-based phosphotyrosine (pTyr) mimetics: hit evolution to a novel orally active protein tyrosine phosphatase?1B (PTP1B) inhibitor.
ChemMedChem
PUBLISHED: 01-02-2014
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Protein tyrosine phosphatase?1B (PTP1B) is a promising therapeutic target for type?2 diabetes. Herein, we report the evolution of a previously identified 3-phenylpropanoic acid-based PTP1B inhibitor to an orally active lead compound. A series of 3-phenylpropanoic acid-based PTP1B inhibitors were synthesized, and three of them, 3-(4-(9H-carbazol-9-yl)phenyl)-5-(3,5-di-tert-butyl-4-methoxyphenyl)-5-oxopentanoic acid (9), 3-(4-(9H-carbazol-9-yl)phenyl)-5-(4'-bromo-[1,1'-biphenyl]-4-yl)-5-oxopentanoic acid (10) and 3-(4-(9H-carbazol-9-yl)-2-fluorophenyl)-5-(4-cyclohexylphenyl)-5-oxopentanoic acid (16), showed IC50 values at sub-micromolar level. Further in?vivo evaluation indicated the sodium salt of 9 not only exhibited significant insulin-sensitizing and hypoglycemia effects, but also decreased the serum levels of triglyceride and total cholesterol in high-fat-diet-induced insulin resistance model mice. Preliminary in?vivo pharmacokinetic studies on the sodium salt of 9 revealed its pharmacokinetic profile after oral administration in rats. These results provide proof-of-concept for the dual effects of PTP1B inhibitors on both glucose and lipid metabolisms.
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Binding pocket alterations in dihydrofolate synthase confer resistance to para-aminosalicylic acid in clinical isolates of Mycobacterium tuberculosis.
Antimicrob. Agents Chemother.
PUBLISHED: 12-23-2013
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The mechanistic basis for resistance of Mycobacterium tuberculosis to para-aminosalicylic acid (PAS), an important agent in the treatment of multi-drug resistant tuberculosis, has yet to be fully defined. As a substrate analog of the folate precursor para-aminobenzoic acid, PAS is ultimately bioactivated to hydroxydihydrofolate which inhibits dihydrofolate reductase and disrupts operation of folate-dependent metabolic pathways. As a result, mutation of dihydrofolate synthase, an enzyme needed for bioactivation of PAS, causes PAS resistance in M. tuberculosis H37Rv. Here, we demonstrate that various missense mutations within the coding sequence of the dihydropteroate (H2Pte) binding pocket of dihydrofolate synthase (FolC) confer PAS resistance in laboratory isolates of M. tuberculosis and M. bovis. From a panel of 85 multi-drug resistant M. tuberculosis clinical isolates, 5 were found to harbor mutations in folC within the H2Pte binding pocket resulting in PAS resistance. While these alterations in the H2Pte binding pocket resulted in reduced dihydrofolate synthase activity, they also abolished bioactivation of hydroxydihydropteroate to hydroxydihydrofolate. Consistent with this model for abolished bioactivation, introduction of a wild type copy of folC fully restored PAS susceptibility in folC mutant strains. Confirmation of this novel PAS resistance mechanism will be beneficial for development of molecular based diagnostics for M. tuberculosis clinical isolates and for further defining the mode of action of this important tuberculosis drug.
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[Changes of the hemodynamics and extravascular lung water after different-volume fluid resuscitation in a piglet model of endotoxic shock].
Zhonghua Er Ke Za Zhi
PUBLISHED: 12-17-2013
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Practice recommendations have evolved, and consensus now exists among leading organizations such as the American College of Critical Care Medicine (ACCM) and Surviving Sepsis Campaign that fluid infusion is best initiated with boluses of 20 ml/kg, commonly requires 40-60 ml/kg but can be as much as 200 ml/kg if the liver is not enlarged and/or rales are not heard. The present study aimed to investigate and compare the changes of the hemodynamics and extravascular lung water after higher volume fluid resuscitation in a piglet model of endotoxic shock.
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Rapid Detection of Mycobacterium tuberculosis and Pyrazinamide Susceptibility Related to pncA Mutations in Sputum Specimens through an Integrated Gene-to-Protein Function Approach.
J. Clin. Microbiol.
PUBLISHED: 11-13-2013
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Testing the pyrazinamide (PZA) susceptibility of Mycobacterium tuberculosis isolates is challenging. In a previous paper, we described the development of a rapid colorimetric test for the PZA susceptibility of M. tuberculosis by a PCR-based in vitro-synthesized-pyrazinamidase (PZase) assay. Here, we present an integrated approach to detect M. tuberculosis and PZA susceptibility directly from sputum specimens. M. tuberculosis was detected first, using a novel long-fragment quantitative real-time PCR (LF-qPCR), which amplified a fragment containing the whole pncA gene. Then, the positive amplicons were sequenced to find mutations in the pncA gene. For new mutations not found in the Tuberculosis Drug Resistance Mutation Database (www.tbdreamdb.com), the in vitro PZase assay was used to test the PZA resistance. This approach could detect M. tuberculosis within 3 h with a detection limit of 7.8 copies/reaction and report the PZA susceptibility within 2 days. In an initial testing of 213 sputum specimens, the LF-qPCR found 53 positive samples with 92% sensitivity and 97% specificity compared to the culture test for M. tuberculosis detection. DNA sequencing of the LF-qPCR amplicons revealed that 49 samples were PZA susceptible and 1 was PZA resistant. In the remaining 3 samples, with new pncA mutations, the in vitro PZase assay found that 1 was PZA susceptible and 2 were PZA resistant. This integrated approach provides a rapid, efficient, and relatively low-cost solution for detecting M. tuberculosis and PZA susceptibility without culture.
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Novel Chimeric Lysin with High-Level Antimicrobial Activity against Methicillin-Resistant Staphylococcus aureus In Vitro and In Vivo.
Antimicrob. Agents Chemother.
PUBLISHED: 11-04-2013
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The treatment of infections caused by methicillin-resistant Staphylococcus aureus (MRSA) is a challenge worldwide. In our search for novel antimicrobial agents against MRSA, we constructed a chimeric lysin (named as ClyH) by fusing the catalytic domain of Ply187 (Pc) with the non-SH3b-like cell wall binding domain of phiNM3 lysin. Herein, the antimicrobial activity of ClyH against MRSA strains in vitro and in vivo was studied. Our results showed that ClyH could kill all of the tested clinical isolates of MRSA with higher efficacy than lysostaphin as well as its parental enzyme. The MICs of ClyH against clinical S. aureus strains were found to be as low as 0.05 to 1.61 mg/liter. In a mouse model, a single intraperitoneal administration of ClyH protected mice from death caused by MRSA, without obvious harmful effects. The present data suggest that ClyH has the potential to be an alternative therapeutic agent for the treatment of infections caused by MRSA.
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Molecular characteristics of the HSP70 gene and its differential expression in female and male golden apple snails (Pomacea canaliculata) under temperature stimulation.
Cell Stress Chaperones
PUBLISHED: 10-11-2013
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Heat-shock protein 70 (HSP70) is one of the most important heat-shock proteins that helps organisms to modulate stress response via over-expression. The HSP70 gene from Pomacea canaliculata was cloned using the RACE approach; the gene is 2,767 bp in length and contains an open reading frame of 1,932 bp, which is encoded by a polypeptide of 643 amino acids. BLAST analysis showed that the predicted amino acid sequence of the P. canaliculata HSP70 gene shared a relatively high similarity with that of other known eukaryotic species that display conserved HSP characteristics. The phylogeny demonstrated a separate clustering of the apple snail HSP70 with other constitutive members from other mollusk species. Quantitative real-time RT-PCR was used to detect the differential expression of HSP70 in both sexes of P. canaliculata at different temperature conditions. These results showed that HSP70 transcript levels decreased slightly under cold shock and increased significantly under heat-shock conditions in both sexes compared to normal temperatures (26 °C). Under cold-shock treatment, the sex effect was not significant. With heat treatment, HSP70 expression could be induced at 36 °C in both females and males, and it peaked at 42 and 39 °C in females and males, respectively. In addition, a clear time-dependent HSP70 expression pattern of the apple snail exposed to the same high temperature (36 °C) was observed at different time points. The maximal induction of HSP70 expression appeared at 12 and 48 h in males and females after heat shock, respectively. The maximal induction in females was significantly higher compared to males under heat stimulus. Taken together, these results strongly suggested that males were more susceptible to heat than females and provided useful molecular information for the ecological adaptability of P. canaliculata against extreme environmental stress.
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Synthesis and biological evaluation of dehydroepiandrosterone-fused thiazole, imidazo[2,1-b]thiazole, pyridine steroidal analogues.
Steroids
PUBLISHED: 09-25-2013
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A series of steroidal[17,16-d]thiazole, steroidal[1,2-b]pyridine and steroidal[17,16-d]thiazole[2,1-b]imidazo products were synthesized through a convenient and productive method. Anti-proliferation activity against EC109 (human esophageal carcinoma), EC9706 (human esophageal carcinoma) and MGC-803 (human gastric carcinoma) cell lines was examined in vitro. Among the screened compounds, several highly potential compounds were located.
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CuI/1,10-phen/PEG promoted decarboxylation of 2,3-diarylacrylic acids: synthesis of stilbenes under neutral and microwave conditions with an in situ generated recyclable catalyst.
Org. Biomol. Chem.
PUBLISHED: 09-24-2013
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A series of trans- or cis-stilbenes have been synthesized in good to excellent yields via a functional group-dependent decarboxylation process from the corresponding 2,3-diaryl acrylic acids in a neutral CuI/1,10-phen/PEG-400 system under microwave conditions. The in situ generation of the recyclable catalytic complex, the use of environmentally benign solvent PEG-400, the operational simplicity, the short reaction times, as well as the functional group-dependent chemo- and stereo-selectivity have made the decarboxylation process a highly efficient and applicable protocol.
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Spatiotopic perceptual learning mediated by retinotopic processing and attentional remapping.
Eur. J. Neurosci.
PUBLISHED: 08-14-2013
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Visual processing takes place in both retinotopic and spatiotopic frames of reference. Whereas visual perceptual learning is usually specific to the trained retinotopic location, our recent study has shown spatiotopic specificity of learning in motion direction discrimination. To explore the mechanisms underlying spatiotopic processing and learning, and to examine whether similar mechanisms also exist in visual form processing, we trained human subjects to discriminate an orientation difference between two successively displayed stimuli, with a gaze shift in between to manipulate their positional relation in the spatiotopic frame of reference without changing their retinal locations. Training resulted in better orientation discriminability for the trained than for the untrained spatial relation of the two stimuli. This learning-induced spatiotopic preference was seen only at the trained retinal location and orientation, suggesting experience-dependent spatiotopic form processing directly based on a retinotopic map. Moreover, a similar but weaker learning-induced spatiotopic preference was still present even if the first stimulus was rendered irrelevant to the orientation discrimination task by having the subjects judge the orientation of the second stimulus relative to its mean orientation in a block of trials. However, if the first stimulus was absent, and thus no attention was captured before the gaze shift, the learning produced no significant spatiotopic preference, suggesting an important role of attentional remapping in spatiotopic processing and learning. Taken together, our results suggest that spatiotopic visual representation can be mediated by interactions between retinotopic processing and attentional remapping, and can be modified by perceptual training.
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Discovery of novel PTP1B inhibitors via pharmacophore-oriented scaffold hopping from Ertiprotafib.
Bioorg. Med. Chem. Lett.
PUBLISHED: 08-13-2013
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An integrated molecular design strategy combining pharmacophore recognition and scaffold hopping was exploited to discover novel PTP1B inhibitors based on the known PTP1B inhibitor Ertiprotafib. A composite pharmacophore model was proposed from the interaction mode of Ertiprotafib, and 21 diverse molecules from five distinct structural classes were designed and synthesized accordingly. New compounds with considerable inhibition against PTP1B were identified from each series, and the most active compound 3a showed IC50 value of 1.3 ?mol L(-1) against human recombinant PTP1B. Docking study indicated that the new inhibitors assumed binding modes similar to that of Ertiprotafib.
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Ketanserin improves cardiac performance after myocardial infarction in spontaneously hypertensive rats partially through restoration of baroreflex function.
Acta Pharmacol. Sin.
PUBLISHED: 07-23-2013
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Aim:Baroreflex dysfunction is associated with a higher rate of sudden death after myocardial infarction (MI). Ketanserin enhances baroreflex function in rats. The present work was designed to examine whether ketanserin improves the post-MI cardiac function and to explore the possible mechanism involved.Methods:Spontaneously hypertensive rats (SHR) were treated with ketanserin (0.3 mg·kg(-1)·d(-1)). Two weeks later, blood pressure and baroreflex function were measured, followed by a ligation of the left coronary artery. The expressions of vesicular acetylcholine transporter (VAChT) and ?7 nicotinic acetylcholine receptor (?7-nAChR) in ischemic myocardium, angiogenesis, cardiac function, and left ventricular (LV) remodeling were evaluated subsequently.Results:Ketanserin significantly improved baroreflex sensitivity (0.62±0.21 vs 0.34±0.12 ms/mmHg, P<0.01) and vagal tonic activity (heart rate changes in response to atropine, 54.8±16.2 vs 37.6±13.4 bpm, P<0.01) without affecting the blood pressure or basic heart rate in SHR. Treatment of SHR with ketanserin prominently improved cardiac function and alleviated LV remodeling, as reflected by increases in the ejection fraction, fractional shortening, and LV systolic pressure as well as decreases in LV internal diameter and LV relative weight. The capillary density, vascular endothelial growth factor expression, and blood flow in the ischemic myocardium were significantly higher in the ketanserin-treated group. In addition, ketanserin markedly increased the expression of VAChT and ?7-nAChR in ischemic myocardium.Conclusion:Ketanserin improved post-MI cardiac function and angiogenesis in ischemic myocardium. The findings provide a mechanistic basis for restoring baroreflex function using ketanserin in the treatment of MI.
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The Passive Leg Raise Test to Predict Fluid Responsiveness in Children -Preliminary Observations.
Indian J Pediatr
PUBLISHED: 06-27-2013
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To assess whether the passive leg raising (PLR) test can predict fluid responsiveness in pediatric patients.
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Stereoselective synthesis of novel antiproliferative steroidal (E, E) dienamides through a cascade aldol/cyclization process.
Steroids
PUBLISHED: 06-17-2013
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The stereoselective and metal-free protocol involving a cascade aldol/cyclization process for the synthesis of steroidal (E, E) dienamides from steroidal ?, ?-dicyanoalkene was reported. This protocol efficiently achieved the construction of C=C bond and selective conversion of cyano group into carboxamide in one-pot procedure under mild condition. Further biological evaluation showed that some of these compounds had moderate to excellent cytotoxic activities against all the tested cancer cell lines and were more potent than well-known drug 5-fluorouracil. Particularly, compound 3c represented excellent inhibitory effect against MCF-7 (IC50=0.76 ?M), which was about 10-fold more potent than 5-fluorouracil.
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A novel [1,2,4] triazolo [1,5-a] pyrimidine-based phenyl-linked steroid dimer: synthesis and its cytotoxic activity.
Eur J Med Chem
PUBLISHED: 06-14-2013
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A novel [1,2,4] triazolo [1,5-a] pyrimidine-based phenyl-linked steroid dimer was designed, synthesized and evaluated for its cytotoxic activity against five human cancer cell lines and the cytotoxicity against human normal liver cell L-02. Compound 3 showed excellent cytotoxic activity and good selectivity between cancer and normal cells. Further mechanistic studies revealed that treatment of EC109 cells with compound 3 caused an obvious G2/M arrest in a concentration- and time-dependent manner and induced apoptosis probably through the mitochondrial pathway accompanied with the decrease of mitochondrial membrane potential, activations of caspase-9/-3, cleavage of MDM2 as well as up-regulation of the expressions of p53 and Bax.
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Recognition and processing of double-stranded DNA by ExoX, a distributive 3-5 exonuclease.
Nucleic Acids Res.
PUBLISHED: 06-14-2013
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Members of the DnaQ superfamily are major 3-5 exonucleases that degrade either only single-stranded DNA (ssDNA) or both ssDNA and double-stranded DNA (dsDNA). However, the mechanism by which dsDNA is recognized and digested remains unclear. Exonuclease X (ExoX) is a distributive DnaQ exonuclease that cleaves both ssDNA and dsDNA substrates. Here, we report the crystal structures of Escherichia coli ExoX in complex with three different dsDNA substrates: 3 overhanging dsDNA, blunt-ended dsDNA and 3 recessed mismatch-containing dsDNA. In these structures, ExoX binds to dsDNA via both a conserved substrate strand-interacting site and a previously uncharacterized complementary strand-interacting motif. When ExoX complexes with blunt-ended dsDNA or 5 overhanging dsDNA, a wedge composed of Leu12 and Gln13 penetrates between the first two base pairs to break the 3 terminal base pair and facilitates precise feeding of the 3 terminus of the substrate strand into the ExoX cleavage active site. Site-directed mutagenesis showed that the complementary strand-binding site and the wedge of ExoX are dsDNA specific. Together with the results of structural comparisons, our data support a mechanism by which normal and mismatched dsDNA are recognized and digested by E. coli ExoX. The crystal structures also provide insight into the structural framework of the different substrate specificities of the DnaQ family members.
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A novel T-type overhangs improve the enzyme-free cloning of PCR products.
Mol. Biotechnol.
PUBLISHED: 06-05-2013
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PCR product cloning is the foundational technology for almost all fields in the life sciences. Numerous innovative methods have been designed during the past few decades. Enzyme-free cloning is the only one that avoids post-amplification enzymatic treatments, making the technique reliable and cost effective. However, the complementary staggered overhangs used in enzyme-free cloning tend to result in self-ligation of the vector under some circumstances. Here, we describe a "T-type" enzyme-free cloning method: instead of designing the complementary staggered overhangs used in conventional enzyme-free cloning, we create "T-type" overhangs that reduce the possibility of self-ligation and are more convenient for multi-vector cloning. In this study, we systematically optimize "T-type" enzyme-free cloning, compare its cloning background with that in conventional enzyme-free cloning, and demonstrate a promising application of this technique in multi-vector cloning. Our method simplifies post-amplification procedures and greatly reduces cost, offering a competitive option for PCR product cloning.
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[Effects of stubble-standing mode on the grain yield and water use efficiency of wheat and maize in wheat/maize intercropping system].
Ying Yong Sheng Tai Xue Bao
PUBLISHED: 05-28-2013
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Wheat/maize intercropping is the main intercropping pattern in the irrigation region of Hexi Oasis, Northwest China, but the traditional intercropping needs much water, making the regional water resource lacked increasingly. In 2010, a field experiment was conducted in the irrigation region of Shiyang River basin oasis, Gansu Province of Northwest China, aimed to study the effects of traditional stubble-burning, stubble-returning, and stubble-standing on the grain yield, water use efficiency (WUE), and economical benefits of wheat and maize in wheat/maize inter-cropping system. Compared with stubble-burning and stubble-returning, stubble-standing increased the grain yield of mono- and intercropped wheat by 7.2% and 5.1% , and 6.2%, 5.1%, and that of mono- and intercropped maize by 4.7% and 2.5%, and 7.2% and 3.3%, and increased the WUE of mono- and intercropped wheat by 20.4% and 16.2%, and 17.9% and 14.6%, and that of mono- and intercropping maize by 16.7% and 10.9%, and 11. 8% and 17.0%, respectively. As for the mono- and intercropped wheat and maize, their average net economical benefits under stubble-burning, stubble- returning, and stubble-standing were 10946, 11471, and 13454 RMB.hm-2, respectively. In considering the grain yield, WUE, and economic benefits, stubble- standing would be the optimal mode of wheat/maize intercropping in the oasis of Hexi irrigation region, Northwest China.
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Design and synthesis of novel D-ring fused steroidal heterocycles.
Steroids
PUBLISHED: 04-27-2013
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Using dehydroepiandrosterone as the starting material, we have synthesized a series of steroid analogs possessing a D-ring fused with heterocycles which are pyridine, imidazo [2,1-b]thiazoles or substituted thiazole imines. All the final structures are first reported and identified by NMR and MS spectroscopys, the yields of these products are moderate to good and the reaction conditions are mild. The cytotoxicity of the synthesized compounds against EC-109(human esophageal carcinoma), EC-9706(human esophageal carcinoma), MGC-803(human gastric carcinoma) were investigated.
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Efficient construction of novel D-ring modified steroidal dienamides and their cytotoxic activities.
Eur J Med Chem
PUBLISHED: 04-18-2013
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Two series of steroidal dienamides 4a-q and 5a-f were designed, synthesized and evaluated for cytotoxic activities against five human cancer cell lines (MGC-803, EC109, PC-3, SMMC-7721 and MCF-7). The protocol developed efficiently achieved the construction of carbon-carbon double bond and selective conversion of nitrile group into carboxamide in one-pot procedure. Besides, compounds 4a-q and 5a-f showed moderate to excellent cytotoxic activities with the IC?? values ranging from 0.1 to 40 ?M and most of them were more potent than 5-fluorouracil. Particularly, four compounds 4d, 4e, 4q and 5a showed excellent selectivity against MGC-803 with the IC?? values less than 1 ?M. Flow cytometry analysis demonstrated that compound 4c caused the cellular early apoptosis and cell cycle arrest in G2/M phase in a concentration-independent manner.
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Design, synthesis and antibacterial evaluation of novel AHL analogues.
Bioorg. Med. Chem. Lett.
PUBLISHED: 04-11-2013
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Two series of novel AHL analogues were designed, synthesized and evaluated for antibacterial activity under cell membrane conditions in vitro. Analogues 4a-c and 4g-m presented potent activity against Gram-positive bacteria. Especially the analogue 4l exerted the most potent inhibition against Bacillus subtilis with MIC50 value of 1.443?g/ml. To our surprise, analogues 6a-c and 6g showed weak inhibition against Gram-negative bacteria with MIC50 values ranging from 17.589 to 67.840?g/ml. This was the first report about synthesis and antibacterial evaluation in vitro of AHL analogues containing dithioester linkage.
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A structure-based design of new C2- and C13-substituted taxanes: tubulin binding affinities and extended quantitative structure-activity relationships using comparative binding energy (COMBINE) analysis.
Org. Biomol. Chem.
PUBLISHED: 03-28-2013
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Ten novel taxanes bearing modifications at the C2 and C13 positions of the baccatin core have been synthesized and their binding affinities for mammalian tubulin have been experimentally measured. The design strategy was guided by (i) calculation of interaction energy maps with carbon, nitrogen and oxygen probes within the taxane-binding site of ?-tubulin, and (ii) the prospective use of a structure-based QSAR (COMBINE) model derived from an earlier series comprising 47 congeneric taxanes. The tubulin-binding affinity displayed by one of the new compounds (CTX63) proved to be higher than that of docetaxel, and an updated COMBINE model provided a good correlation between the experimental binding free energies and a set of weighted residue-based ligand-receptor interaction energies for 54 out of the 57 compounds studied. The remaining three outliers from the original training series have in common a large unfavourable entropic contribution to the binding free energy that we attribute to taxane preorganization in aqueous solution in a conformation different from that compatible with tubulin binding. Support for this proposal was obtained from solution NMR experiments and molecular dynamics simulations in explicit water. Our results shed additional light on the determinants of tubulin-binding affinity for this important class of antitumour agents and pave the way for further rational structural modifications.
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Pharmacogenetic activation of midbrain dopaminergic neurons induces hyperactivity.
Neurosci Bull
PUBLISHED: 03-20-2013
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Dopaminergic neurons regulate and organize numerous important behavioral processes including motor activity. Consistently, manipulation of brain dopamine concentrations changes animal activity levels. Dopamine is synthesized by several neuronal populations in the brain. This study was carried out to directly test whether selective activation of dopamine neurons in the midbrain induces hyperactivity. A pharmacogenetic approach was used to activate midbrain dopamine neurons, and behavioral assays were conducted to determine the effects on mouse activity levels. Transgenic expression of the evolved hM3Dq receptor was achieved by infusing Cre-inducible AAV viral vectors into the midbrain of DAT-Cre mice. Neurons were excited by injecting the hM3Dq ligand clozapine-N-oxide (CNO). Mouse locomotor activity was measured in an open field. The results showed that CNO selectively activated midbrain dopaminergic neurons and induced hyperactivity in a dose-dependent manner, supporting the idea that these neurons play an important role in regulating motor activity.
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Insights into Stabilization of a Viral Protein Cage in Templating Complex Nanoarchitectures: Roles of Disulfide Bonds.
Small
PUBLISHED: 03-18-2013
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As a typical protein nanostructure, virus-based nanoparticle (VNP) of simian virus 40 (SV40), which is composed of pentamers of the major capsid protein of SV40 (VP1), has been successfully employed in guiding the assembly of different nanoparticles (NPs) into predesigned nanostructures with considerable stability. However, the stabilization mechanism of SV40 VNP remains unclear. Here, the importance of inter-pentamer disulfide bonds between cysteines in the stabilization of quantum dot (QD)-containing VNPs (VNP-QDs) is comprehensively investigated by constructing a series of VP1 mutants of cysteine to serine. Although the presence of a QD core can greatly enhance the assembly and stability of SV40 VNPs, disulfide bonds are vital to stability of VNP-QDs. Cysteine at position 9 (C9) and C104 contribute most of the disulfide bonds and play essential roles in determining the stability of SV40 VNPs as templates to guide assembly of complex nanoarchitectures. These results provide insightful clues to understanding the robustness of SV40 VNPs in organizing suprastructures of inorganic NPs. It is expected that these findings will help guide the future design and construction of protein-based functional nanostructures.
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Idiopathic left ventricular rupture in the absence of coronary artery disease.
J Card Surg
PUBLISHED: 03-12-2013
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Idiopathic cardiac rupture in the absence of coronary artery disease is rare. We describe a case of idiopathic left ventricular free wall rupture with successful surgical repair.
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Acute and long-term suppression of feeding behavior by POMC neurons in the brainstem and hypothalamus, respectively.
J. Neurosci.
PUBLISHED: 02-22-2013
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POMC-derived melanocortins inhibit food intake. In the adult rodent brain, POMC-expressing neurons are located in the arcuate nucleus (ARC) and the nucleus tractus solitarius (NTS), but it remains unclear how POMC neurons in these two brain nuclei regulate feeding behavior and metabolism differentially. Using pharmacogenetic methods to activate or deplete neuron groups in separate brain areas, in the present study, we show that POMC neurons in the ARC and NTS suppress feeding behavior at different time scales. Neurons were activated using the DREADD (designer receptors exclusively activated by designer drugs) method. The evolved human M3-muscarinic receptor was expressed in a selective population of POMC neurons by stereotaxic infusion of Cre-recombinase-dependent, adeno-associated virus vectors into the ARC or NTS of POMC-Cre mice. After injection of the human M3-muscarinic receptor ligand clozapine-N-oxide (1 mg/kg, i.p.), acute activation of NTS POMC neurons produced an immediate inhibition of feeding behavior. In contrast, chronic stimulation was required for ARC POMC neurons to suppress food intake. Using adeno-associated virus delivery of the diphtheria toxin receptor gene, we found that diphtheria toxin-induced ablation of POMC neurons in the ARC but not the NTS, increased food intake, reduced energy expenditure, and ultimately resulted in obesity and metabolic and endocrine disorders. Our results reveal different behavioral functions of POMC neurons in the ARC and NTS, suggesting that POMC neurons regulate feeding and energy homeostasis by integrating long-term adiposity signals from the hypothalamus and short-term satiety signals from the brainstem.
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Reversibly acetylated lysine residues play important roles in the enzymatic activity of Escherichia coli N-hydroxyarylamine O-acetyltransferase.
FEBS J.
PUBLISHED: 02-17-2013
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CobB is a bacterial NAD(+)-dependent protein deacetylase. Although progress has been made in functional studies of this protein in recent years, its substrates and biological functions are still largely unclear. Using proteome microarray technology, potential substrates of Escherichia coli CobB were screened and nine proteins were identified, including N-hydroxyarylamine O-acetyltransferase (NhoA). In vitro acetylation/deacetylation of NhoA was verified by western blotting and mass spectrometry, and two acetylated lysine residues were identified. Site-specific mutagenesis experiments showed that mutation of each acetylated lysine decreased the acetylation level of NhoA in vitro. Further analysis showed that variant NhoA proteins carrying substitutions at the two acetylated lysine residues are involved in both the O-acetyltransferase and N-acetyltransferase activity of NhoA. Structural analyses were also performed to explore the effects of the acetylated lysine residues on the activity of NhoA. These results suggest that reversible acetylation may play a role in the activity of Escherichia coli NhoA.
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Genome sequencing of 161 Mycobacterium tuberculosis isolates from China identifies genes and intergenic regions associated with drug resistance.
Nat. Genet.
PUBLISHED: 02-07-2013
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The worldwide emergence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis threatens to make this disease incurable. Drug resistance mechanisms are only partially understood, and whether the current understanding of the genetic basis of drug resistance in M. tuberculosis is sufficiently comprehensive remains unclear. Here we sequenced and analyzed 161 isolates with a range of drug resistance profiles, discovering 72 new genes, 28 intergenic regions (IGRs), 11 nonsynonymous SNPs and 10 IGR SNPs with strong, consistent associations with drug resistance. On the basis of our examination of the dN/dS ratios of nonsynonymous to synonymous SNPs among the isolates, we suggest that the drug resistance-associated genes identified here likely contain essentially all the nonsynonymous SNPs that have arisen as a result of drug pressure in these isolates and should thus represent a near-complete set of drug resistance-associated genes for these isolates and antibiotics. Our work indicates that the genetic basis of drug resistance is more complex than previously anticipated and provides a strong foundation for elucidating unknown drug resistance mechanisms.
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Use of hGluc/tdTomato pair for sensitive BRET sensing of protease with high solution media tolerance.
Talanta
PUBLISHED: 01-25-2013
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Due to the complicated media, monitoring proteases in real physiological environments is still a big challenge. Bioluminescence resonance energy transfer (BRET) is one of the promising techniques but its application is limited by the susceptibility to buffer composition, which might cause serious errors for the assay. Herein we report a novel combination of BRET pair with humanized Gaussia luciferase (hGluc) and highly bright red fluorescence protein tdTomato for sensitive and robust protease activity determination. As a result, the hGluc/tdTomato BRET pair showed much better tolerance to buffer composition, pH and sample matrices, and wide spectral separation (??:~110 nm). With the protease sensor built with this pair, the detection limit for enterokinase reached 2.1 pM in pure buffer and 3.3 pM in 3% serum. The proposed pair would find broad use in both in vitro and in vivo assays, especially for samples with complicated matrix.
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Characterization of shallow groundwater quality in the Lower St. Johns River Basin: a case study.
Environ Sci Pollut Res Int
PUBLISHED: 01-16-2013
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Characterization of groundwater quality allows the evaluation of groundwater pollution and provides information for better management of groundwater resources. This study characterized the shallow groundwater quality and its spatial and seasonal variations in the Lower St. Johns River Basin, Florida, USA, under agricultural, forest, wastewater, and residential land uses using field measurements and two-dimensional kriging analysis. Comparison of the concentrations of groundwater quality constituents against the US EPAs water quality criteria showed that the maximum nitrate/nitrite (NO x ) and arsenic (As) concentrations exceeded the EPAs drinking water standard limits, while the maximum Cl, SO 4 (2?-) , and Mn concentrations exceeded the EPAs national secondary drinking water regulations. In general, high kriging estimated groundwater NH 4 (+) concentrations were found around the agricultural areas, while high kriging estimated groundwater NO x concentrations were observed in the residential areas with a high density of septic tank distribution. Our study further revealed that more areas were found with high estimated NO x concentrations in summer than in spring. This occurred partially because of more NO x leaching into the shallow groundwater due to the wetter summer and partially because of faster nitrification rate due to the higher temperature in summer. Large extent and high kriging estimated total phosphorus concentrations were found in the residential areas. Overall, the groundwater Na and Mg concentration distributions were relatively more even in summer than in spring. Higher kriging estimated groundwater As concentrations were found around the agricultural areas, which exceeded the EPAs drinking water standard limit. Very small variations in groundwater dissolved organic carbon concentrations were observed between spring and summer. This study demonstrated that the concentrations of groundwater quality constituents varied from location to location, and impacts of land uses on groundwater quality variation were profound.
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Facile synthesis of novel D-ring modified steroidal dienamides via rearrangement of 2H-pyrans.
Steroids
PUBLISHED: 01-16-2013
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A simple and practical method for synthesis of the D-ring modified steroidal dienamides (4a-k) from the steroidal ?,?-dicyanoalkene 3 and aldehydes via vinylogous aldol reaction was first reported. By using NaOAc as a base, the desired products were obtained in moderate to good yields in ethanol under mild conditions. All the synthesized steroidal dienamides are new and are currently being evaluated for their biological activities.
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An update on antitumor activity of naturally occurring chalcones.
Evid Based Complement Alternat Med
PUBLISHED: 01-15-2013
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Chalcones, which have characteristic 1,3-diaryl-2-propen-1-one skeleton, are mainly produced in roots, rhizomes, heartwood, leaves, and seeds of genera Angelica, Sophora, Glycyrrhiza, Humulus, Scutellaria, Parartocarpus, Ficus, Dorstenia, Morus, Artocarpus, and so forth. They have become of interest in the research and development of natural antitumor agents over the past decades due to their broad range of mechanisms including anti-initiation, induction of apoptosis, antiproliferation, antimetastasis, antiangiogenesis, and so forth. This review summarizes the studies on the antitumor activity of naturally occurring chalcones and their underlying mechanisms in detail during the past decades.
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Mandibular reconstruction assisted by preoperative simulation and accurate transferring templates: preliminary report of clinical application.
J. Oral Maxillofac. Surg.
PUBLISHED: 01-05-2013
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This study investigated the application of a computer-aided design and manufacturing technique of defining tumor resection, fibula cutting, and positioning by surgical templates in mandibular reconstructive surgery.
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Design and synthesis of novel 1,2,3-triazole-dithiocarbamate hybrids as potential anticancer agents.
Eur J Med Chem
PUBLISHED: 01-04-2013
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A series of novel 1,2,3-triazole-dithiocarbamate hybrids were designed, synthesized and evaluated for anticancer activity against four selected human tumor cell lines (MGC-803, MCF-7, PC-3, EC-109). Majority of the synthesized compounds exhibited moderate to potent activity against MGC-803 and MCF-7. Among them, compounds 3a and 3c showed excellent broad spectrum anticancer activity with IC50 values ranging from 0.73 to 11.61 ?M and 0.49-12.45 ?M, respectively. Particularly, compound 3a was more potent than 5-fluorouracil against all tested human cancer cell lines. Flow cytometry analysis demonstrated that treatment of MGC-803 with 3c led to cell cycle arrest at G2/M phase accompanied by an increase in apoptotic cell death after 12 h.
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Imaging of mRNA-Protein Interactions in Live Cells Using Novel mCherry Trimolecular Fluorescence Complementation Systems.
PLoS ONE
PUBLISHED: 01-01-2013
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Live cell imaging of mRNA-protein interactions makes it possible to study posttranscriptional processes of cellular and viral gene expression under physiological conditions. In this study, red color mCherry-based trimolecular fluorescence complementation (TriFC) systems were constructed as new tools for visualizing mRNA-protein interaction in living cells using split mCherry fragments and HIV REV-RRE and TAT-TAR peptide-RNA interaction pairs. The new mCherry TriFC systems were successfully used to image RNA-protein interactions such as that between influenza viral protein NS1 and the 5 UTR of influenza viral mRNAs NS, M, and NP. Upon combination of an mCherry TriFC system with a Venus TriFC system, multiple mRNA-protein interactions could be detected simultaneously in the same cells. Then, the new mCherry TriFC system was used for imaging of interactions between influenza A virus mRNAs and some of adapter proteins in cellular TAP nuclear export pathway in live cells. Adapter proteins Aly and UAP56 were found to associate with three kinds of viral mRNAs. Another adapter protein, splicing factor 9G8, only interacted with intron-containing spliced M2 mRNA. Co-immunoprecipitation assays with influenza A virus-infected cells confirmed these interactions. This study provides long-wavelength-spectrum TriFC systems as new tools for visualizing RNA-protein interactions in live cells and help to understand the nuclear export mechanism of influenza A viral mRNAs.
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Quantum dot-induced viral capsid assembling in dissociation buffer.
Int J Nanomedicine
PUBLISHED: 01-01-2013
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Viruses encapsulating inorganic nanoparticles are a novel type of nanostructure with applications in biomedicine and biosensors. However, the encapsulation and assembly mechanisms of these hybridized virus-based nanoparticles (VNPs) are still unknown. In this article, it was found that quantum dots (QDs) can induce simian virus 40 (SV40) capsid assembly in dissociation buffer, where viral capsids should be disassembled. The analysis of the transmission electron microscope, dynamic light scattering, sucrose density gradient centrifugation, and cryo-electron microscopy single particle reconstruction experimental results showed that the SV40 major capsid protein 1 (VP1) can be assembled into ?25 nm capsids in the dissociation buffer when QDs are present and that the QDs are encapsulated in the SV40 capsids. Moreover, it was determined that there is a strong affinity between QDs and the SV40 VP1 proteins (KD=2.19E-10 M), which should play an important role in QD encapsulation in the SV40 viral capsids. This study provides a new understanding of the assembly mechanism of SV40 virus-based nanoparticles with QDs, which may help in the design and construction of other similar virus-based nanoparticles.
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Monofunctionalization of protein nanocages.
J. Am. Chem. Soc.
PUBLISHED: 11-23-2011
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Surface monofunctionalization of protein nanostructures will enable precise topological control over the protein-templated assembly of nanoscale motifs, however, this remains a formidable challenge. Here we demonstrated a novel strategy for this purpose with a protein nanocage, virus-based nanoparticle (VNP) of simian virus 40 as a model system. By simultaneously incorporating a function modality (cysteine) and a purification modality (polyhistidine tag) into the building block (VP1) of VNPs through rational design and genetic engineering, the monofunctionalized cysteine-VNPs are readily obtained through a routine affinity chromatography in virtue of the purification modality of polyhistidine tag, after the coassembly of the functional VP1 and the nonfunctional VP1 at an optimal ratio. This strategy has proved to be highly efficient in constructing monofunctionalized protein nanostructures as highlighted by the monofunctionalized-VNP-guided Au/QD-VNP nanostructures. These nanostructures could be utilized in a wide range of disciplines, including basic biological research, novel nanostructures, and nanodevices fabrication, etc.
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Multifunctional ferritin cage nanostructures for fluorescence and MR imaging of tumor cells.
Nanoscale
PUBLISHED: 11-14-2011
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Bionanoparticles and nanostructures have attracted increasing interest as versatile and promising tools in many applications including biosensing and bioimaging. In this study, to image and detect tumor cells, ferritin cage-based multifunctional hybrid nanostructures were constructed that: (i) displayed both the green fluorescent protein and an Arg-Gly-Asp peptide on the exterior surface of the ferritin cages; and (ii) incorporated ferrimagnetic iron oxide nanoparticles into the ferritin interior cavity. The overall architecture of ferritin cages did not change after being integrated with fusion proteins and ferrimagnetic iron oxide nanoparticles. These multifunctional nanostructures were successfully used as a fluorescent imaging probe and an MRI contrast agent for specifically probing and imaging ?(v)?(3) integrin upregulated tumor cells. The work provides a promising strategy for tumor cell detection by simultaneous fluorescence and MR imaging.
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Comparative analysis of mycobacterial NADH pyrophosphatase isoforms reveals a novel mechanism for isoniazid and ethionamide inactivation.
Mol. Microbiol.
PUBLISHED: 11-03-2011
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NADH pyrophosphatase (NudC) catalyses the hydrolysis of NAD(H) to AMP and NMN(H) [nicotinamide mononucleotide (reduced form)]. NudC multiple sequence alignment reveals that homologues from most Mycobacterium tuberculosis isolates, but not other mycobacterial species, have a polymorphism at the highly conserved residue 237. To elucidate the functional significance of this polymorphism, comparative analyses were performed using representative NudC isoforms from M. tuberculosis H37Rv (NudC(Rv)) and M. bovis BCG (NudC(BCG)). Biochemical analysis showed that the P237Q polymorphism prevents dimer formation, and results in a loss of enzymatic activity. Importantly, NudC(BCG) was found to degrade the active forms of isoniazid (INH), INH-NAD and ethionamide (ETH), ETH-NAD. Consequently, overexpression of NudC(BCG) in Mycobacterium smegmatis mc(2)155 and M. bovis BCG resulted in a high level of resistance to both INH and ETH. Further genetic studies showed that deletion of the nudC gene in M. smegmatis mc(2)155 and M. bovis BCG resulted in increased susceptibility to INH and ETH. Moreover, inactivation of NudC in both strains caused a defect in drug tolerance phenotype for both drugs in exposure assays. Taken together, these data suggest that mycobacterial NudC plays an important role in the inactivation of INH and ETH.
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[Study on rapid determination of nutrient composition in corn distillers dried grains with solubles by near-infrared reflectance spectroscopy].
Guang Pu Xue Yu Guang Pu Fen Xi
PUBLISHED: 10-31-2011
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The possibility of using near-infrared reflectance spectroscopy (NIRS) for quantitative determination of 8 important nutrient compositions, including moisture, crude protein, ether extract, ash, total phosphorus, neutral detergent fiber, acid detergent fiber and crude fiber in corn DDGS was investigated in the present study. Ninety-three samples were collected from 18 plants in China over a three years period. Calibrations were performed by modified partial least squared algorithm and 15 different derivatives plus scatter correction spectral pretreatments. The results showed that the second derivative mathematical treatment gave the best prediction performance for most constituents. The prediction performance of models developed using 93 calibration samples was better than that developed using 70 samples. The coefficients of determination for calibration (RSQcal), coefficients of determination for cross-validation (1-VR), and RPD(CV) of cross-validation in models developed using 93 samples were 0.94-0.99, 0.89-0.99, and 2.98-14.85, respectively. These results indicate that NIRS can be used as a quantitative method for rapid determination of nutrient composition in corn DDGS.
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Impact of epidemic rates of diabetes on the Chinese blood glucose testing market.
J Diabetes Sci Technol
PUBLISHED: 10-27-2011
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China has become the country with the largest diabetes mellitus population in the world since the 1990s. About 100 million diabetes cases have been diagnosed since 2008. Handheld blood glucose meters and test strips are urgently needed for daily patient measurement. The glucose monitor with a screen-printed carbon-based glucose electrode has been in commercial production since 1994. Since then, approximately 20 companies have been involved in manufacturing and marketing meters and test strips in China. The current market and production volume and updates on technology issues are discussed in this article.
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[Optimization of supercritical fluid extraction of ursolic acid from follum eriobotryae by response surface methodology].
Zhong Yao Cai
PUBLISHED: 10-25-2011
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To optimize the conditions for extracting ursolic acid from Follum eriobotryae with supercritical fluid extration (SFE).
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[Impact of Mikania micrantha invasion on soil meso- and micro-invertebrate community structure].
Ying Yong Sheng Tai Xue Bao
PUBLISHED: 10-20-2011
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Mikania micrantha, a notorious exotic weed of Asteraceae family, has invaded successfully in southern China, and caused serious damages to native ecosystems. In this paper, a field survey was conducted in the Huolushan Forest Park of Guangzhou, China, aimed to understand the impact of M. micrantha invasion on the soil meso- and micro-invertebrate community. Three sampling sites were installed, including M. micrantha-invaded site, ecotone, and native vegetation site. Through four samplings in 2009, a total of 5206 soil meso- and micro-invertebrate individuals were collected, belonging to 4 phyla, 10 classes, and 19 orders, among which, Nematoda was the dominant group, and Acarina, Collembolan, and Rotifera were the common groups. M. micrantha invasion altered the characteristics of soil meso- and micro-invertebrate community structure. Compared with those at the other two sampling sites, the numbers of total individuals, Nematoda, and Acarina at M. micrantha-invaded site increased significantly, but the groups of soil meso- and micro-invertebrates had less change. At M. micrantha-invaded site, the density-group index (DG) of soil meso- and micro-invertebrates was significantly higher, Margalef richness index (D) and Simpson dominance index (C) tended to ascend, but Pielou evenness index (E) and Shannon index (H) tended to descend. The similarity coefficient of soil meso- and micro-invertebrate community between M. micrantha-invaded site and ecotone was higher than that between M. micrantha-invaded site and native vegetation site. The changes of local climate conditions, plant litters, root secretions, and soil physical-chemical properties caused by M. micrantha invasion could be the major contributing factors that altered the community structure of soil meso- and micro-invertebrates at M. micrantha-invaded site.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.