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Biomedical subjects

X Xu

Publications and source records attributed to X Xu.

At least 325 records · Page 18Linked to original sources
PubMed2000

A 500-kb region on chromosome 16p13.1 contains the pseudoxanthoma elasticum locus: high-resolution mapping and genomic structure.

We have recently mapped the genetic defect underlying pseudoxanthoma elasticum (PXE), an inherited disorder characterized by progressive calcification of elastic fibers in skin, eye, and cardiovascular system, to chromosome 16p 13.1. Here we report further data on the fine-mapping and genomic structure of this locus. Haplotype analysis of informative PXE families narrowed the locus to an interval of less than 500 kb located between markers D16B9621 and D16S764. Three overlapping YAC clones were found to cover this region through YAC-STS content mapping. An overlapping BAC contig was then constructed to cover this interval and the surrounding region. About 80% of this chromosomal region has been fully sequenced using the BAC shotgun technique. Gene content and sequence analysis predicted four genes (MRP1, MRP6, PM5, and a novel transcript) and two pseudogenes (ARA and PKDI) within this interval. By screening a somatic cell hybrid panel we were able to precision-map the breakpoint of Cy185 and the starting point of a chromosomal duplication within 20 kb of BAC A962B4. The present data further refine the localization of PXE, provide additional physical cloning resources, and will aid in the eventual identification of the genetic defect causing PXE.

Adult↗
PubMed2000

Analysis of expression of the binary toxin genes from Bacillus sphaericus in Anabaena and the potential in mosquito control.

Anabaena strains expressing the binary toxin genes of Bacillus sphaericus produce high larvicidal activity with living cells. Western blot analysis showed that the 51-kDa and 42-kDa toxin proteins were stable in Anabaena. When a DNA fragment upstream of the 51-kDa protein gene was deleted, the toxicity was reduced by over a hundred-fold, whereas deletions at the coding regions showed that the cooperation of the two proteins expressed in Anabaena is essential for the larvicidal activity. Outdoor tests showed that the genetically altered Anabaena could keep containers with natural water from being inhabited by Culex larvae for over 2 months.

Anabaena↗
PubMed2000

A high-resolution linkage map of the vicinity of the rice submergence tolerance locus Sub1.

Resistance to submergence stress is an important breeding objective in areas where rice cultivars are subjected to complete inundation for a week or more. The present study was conducted to develop a high-resolution map of the region surrounding the submergence tolerance gene Sub1 in rice, which derives from the Indian cultivar FR13A. Submergence screening of 8-day-old plants of F3 families kept for 14 days submerged in 60 cm of water allowed an accurate classification of Sub1 phenotypes. Bulked segregant analysis was used to identify AFLP markers linked to Sub1. A population of 2950 F2 plants segregating for Sub1 was screened with two RFLP markers flanking the Sub1 locus, 2.4 and 4.9 cM away. Submergence tolerance was measured in the recombinant plants, and AFLP markers closely linked to Sub1 were mapped. Two AFLP markers cosegregated with Sub1 in this large population, and other markers were localized within 0.2 cM of Sub1. The high-resolution map should serve as the basis for map-based cloning of this important locus, as it will permit the identification of BAC clones spanning the region.

Chromosome Mapping↗
PubMed2000

In vivo expression of the novel CXC chemokine BRAK in normal and cancerous human tissue.

Using differential display, we cloned a gene with reduced expression in short-term explants of head and neck squamous cell carcinoma (HNSCC) tumors compared to cultured normal oral epithelial cells. The differentially expressed gene was identical to the recently cloned CXC chemokine BRAK, which is ubiquitously expressed in normal tissue extracts but is absent from many tumor cell lines in vitro. To define the cell populations expressing BRAK in vivo, in situ mRNA hybridization was performed on normal and cancerous tissues from six different histological sites. The predominant normal cell type constitutively expressing BRAK in vivo was squamous epithelium. Expression in tumors was heterogeneous, with the majority of HNSCCs and some cervical squamous cell carcinomas (SCCs) showing loss of BRAK mRNA. Although absent in unstimulated peripheral blood mononuclear cells, high levels of BRAK were consistently found in infiltrating inflammatory cells (with lymphocyte morphology) in nearly all cancers examined. Furthermore, BRAK expression was demonstrated in B cells and monocytes, after stimulation of peripheral blood mononuclear cells with lipopolysaccharide. This study demonstrates for the first time up-regulation of BRAK mRNA by inflammatory cells in the tumor microenvironment and lost expression from certain cancers in vivo. The data suggest that BRAK may have a role in host-tumor interactions.

Amino Acid Sequence↗
PubMed2000

Molecular cloning of cDNA encoding a drosophila ryanodine receptor and functional studies of the carboxyl-terminal calcium release channel.

Ryanodine is a plant alkaloid that was originally used as an insecticide. To study the function and regulation of the ryanodine receptor (RyR) from insect cells, we have cloned the entire cDNA sequence of RyR from the fruit fly Drosophila melanogaster. The primary sequence of the Drosophila RyR contains 5134 amino acids, which shares approximately 45% identity with RyRs from mammalian cells, with a large cytoplasmic domain at the amino-terminal end and a small transmembrane domain at the carboxyl-terminal end. To characterize the Ca(2+) release channel activity of the cloned Drosophila RyR, we expressed both full-length and a deletion mutant of Drosophila RyR lacking amino acids 277-3650 (Drosophila RyR-C) in Chinese hamster ovary cells. For subcellular localization of the expressed Drosophila RyR and Drosophila RyR-C proteins, green fluorescent protein (GFP)-Drosophila RyR and GFP-Drosophila RyR-C fusion constructs were generated. Confocal microscopic imaging identified GFP-Drosophila RyR and GFP-Drosophila RyR-C on the endoplasmic reticulum membranes of transfected cells. Upon reconstitution into the lipid bilayer membrane, Drosophila RyR-C formed a large conductance cation-selective channel, which was sensitive to modulation by ryanodine. Opening of the Drosophila RyR-C channel required the presence of microM concentration of Ca(2+) in the cytosolic solution, but the channel was insensitive to inhibition by Ca(2+) at concentrations as high as 20 mM. Our data are consistent with our previous observation with the mammalian RyR that the conduction pore of the calcium release channel resides within the carboxyl-terminal end of the protein and further demonstrate that structural and functional features are essentially shared by mammalian and insect RyRs.

Amino Acid Sequence↗
PubMed2000

Purification and characterization of anticoagulation factors from the venom of Agkistrodon acutus.

Two anticoagulants from five-pace snake (Agkistrodon acutus) venom, anticoagulation factor I (ACF I) and anticoagulation factor II (ACF II), have been purified by a multiple-step chromatography procedure of anion-exchange chromatography, gel permeation chromatography and cation-exchange chromatography. Each of them is shown to be homogeneous as judged by PAGE, SDS-PAGE and mass spectrometry. In vitro, both proteins show equivalent anticoagulant activity, and are devoid of proteolytic, esterolytic, L-amino acid oxidase, phospholipase A, thrombin-like, fibrinolytic, hemorrhagic and lethal activities. They have similar amino acid compositions with similar absorption coeffecients (A(1%)(280)) (30.5 for ACF I and 30.0 for ACF II). Both are disulfide-linked consisting of two 14.7 kD chains for ACF I and two 14.6 kD chains for ACFII. ACF I has a molecular mass of 29,604+/-8 atomic mass units (amu) compared to 29,468+/-6 amu for ACF II, determined by mass spectrometry. The isoelectric points of ACF I and ACF II are 5.7 and 7.0, respectively. We conclude that the two isoforms possess equivalent biological activities with similar amino acid compositions and molecular masses, but different isoelectric points.

Agkistrodon↗
PubMed2000

Adaptation of E. coli cell method for micro-scale nitrate measurement with the Griess reaction in culture media.

The E. coli cell method for nitrate measurement consists of two-steps: nitrate reduction by the E. coli cell usually under anaerobic conditions and subsequently nitrite measurement with the Griess reaction. It was found that the E. coli DSM 498k wildtype cell can reduce nitrate to nitrite under aerobic conditions. Therefore, the E. coli method for nitrate measurement was adapted to be performed under aerobic conditions in a microtiter plate. The adapted method is simpler than the original E. coli method and other nitrate methods such as those with inorganic reductants and with purified enzymes. Furthermore, it was found that for the Griess reaction the pH values of samples after addition of the Griess reagent A should be lower than 1.8 for a stable absorbance at 540 nm to be reached. It is important to add the two Griess reagents separately and to read the absorbance twice consecutively in a microtiter plate. The adapted E. coli method was successfully applied to measure the traces of nitrate in MRS and other medium components by measuring the standard curve of a dilution of each individual medium component. It was found that many organic medium components contain traces of nitrate, while none of them contain detectable nitrite. Among these, the extract of meat and yeast extract contain relatively high amounts of nitrate: 217 mg N/kg and 99 mg N/kg respectively. MRS broth contains nitrate from 0.3 to 0.6 mg N/l depending on the batch numbers of the product. The adapted E. coli can also be used for nitrate measurement in other matrices.

Culture Media↗
PubMed2000

Nociceptin/orphanin FQ in spinal nociceptive mechanisms under normal and pathological conditions.

Nociceptin and its receptor are present in dorsal spinal cord, indicating a possible role for this peptide in pain transmission. The majority of functional studies using behavioral and electrophysiological studies have shown that nociceptin applied at spinal level produces antinociception through pre- and post-synaptic mechanisms. The spinal inhibitory effect of nociceptin is not sensitive to antagonists of opioid receptors such as naloxone. Thus, nociceptin-induced antinociception is mediated by a novel mechanism independent of activation of classic opioid receptors. This has raised the possibility that agonists of the nociceptin receptor may represent a novel class of analgesics. Supporting this hypothesis, several groups have shown that intrathecal nociceptin alleviated hyperalgesic and allodynic responses in rats after inflammation or partial peripheral nerve injury. Electrophysiological studies have also indicated that the antinociceptive potency of spinal nociceptin is maintained or enhanced after nerve injury. It is concluded that the predominant action of nociceptin in the spinal cord appears to be inhibitory. The physiological role of nociceptin in spinal nociceptive mechanisms remains to be defined. Moreover, further evaluation of nociceptin as a new analgesic calls the development of non-peptide brain penetrating agents.

Analgesics↗
PubMed2000

Effects of tobacco smoking on topographic EEG and Stroop test in smoking deprived smokers.

1. Quantitative electroencephalography (EEG) was used to measure human brain electrical changes produced by tobacco deprivation and smoking. Sixteen scalp cortical recording sites monitored regional changes in brain activity. 2. The quantitative EEG was subdivided into delta, theta, alpha1, alpha2, beta1 and beta2 bands for topographic mapping. A demanding version of the Stroop test was used to determine tobacco smoking effects on attention. The version used was more difficult than that used in previously reported studies. 3. Healthy drug and substance free adult male and female volunteers were divided into nonsmoker (n=7) and smoker (n=7) groups according to their smoking status. They were instructed to abstain from tobacco products for at least 12 hr overnight before the next morning's experiment. 4. EEG was recorded before and after smoking either a fake placebo cigarette for nonsmokers or the cigarette of their choice for smokers. Subjects were also asked to perform the Stroop test before and after smoking the placebo or tobacco cigarette. 5. The results showed that tobacco smoking significantly depressed delta and increased alpha2 and beta1 activity and slightly increased the Stroop effect. Although smoking one cigarette stimulated brain electrical activity of smoking deprived smokers, it did not improve performance on a difficult Stroop test.

Adult↗
PubMed2000

Inhibition of 1,2-dimethylhydrazine-induced oxidative DNA damage in rat colon mucosa by black tea complex polyphenols.

The effect of black tea polyphenols on 1,2-dimethylhydrazine (DMH)-induced oxidative DNA damage in rat colon mucosa has been investigated. Fischer 344 rats were treated orally with thearubigin (TR) or theafulvin (TFu) for 10 days (40 mg/kg), injected ip with DMH (20 mg/kg) or saline and sacrificed 24 hr after DMH administration. The levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) were measured in colonic mucosa DNA and expressed as a ratio relative to 2'-deoxyguanosine (2dG). Control rat mucosa had 8-OHdG values of 1.12 +/- 0.14/10(5) dG (mean +/- SEM, n=11), whereas DMH-treated rats significantly higher values (1.52 +/- 0.14/10(5) dG, n=26, P<0.05). Pretreatment of rats with TR had significantly inhibited DMH-induced oxidative DNA damage 0.99 +/- 0.09/10(5) dG, n=10, P<0.05) and a similar, although less marked, effect was observed with TFu (1.15 +/- 0.19/10(5), n=9, P=0.06). These findings confirm that DMH causes oxidative DNA damage in the colon mucosa of rats and demonstrate that this effect is prevented by the consumption of complex polyphenols from black tea.

1,2-Dimethylhydrazine↗
PubMed2000

Chiral analysis of selected dopamine receptor antagonists in serum using capillary electrophoresis with cyclodextrin additives.

Dopamine D-2 receptor antagonists' eticlopride and sulpiride were determined in serum using capillary electrophoresis with cyclodextrin additives. Chiral resolution of S(-) and R(+) sulpiride and eticlopride were achieved using 2% sulfated-beta-cyclodextrin (S-beta-CD) in 20 mM citrate run buffer (pH 2.90). A 72-cm fused silica capillary operated in the reversed polarity mode voltage of 20 kV was used for the analysis. The analytes of interest were isolated from serum using a solid phase extraction procedure with recoveries in excess of 85% for all four enantiomers. The D-2 receptor antagonist (-) butaclamol was used as internal standard. The limits of detection were 0.3 and 0.1 microg/ml for S(-) and R(+) eticlopride and for S(-) and R(+) sulpiride, respectively, in 1 ml of serum. The limits of quantitation were 2 and 1 microg/ml for S(-) and R(+) eticlopride, and for S(-) and R(+) sulpiride, respectively. Calibration curves were linear over the 2-20 micro/ml range for eticlopride and 1-20 microg/ml range for sulpiride. The coefficients of determination were greater than 0.99 (n = 12 for eticlopride and n = 15 for sulpiride). Precision and accuracy of the method were 0.27-6.38 and 0.20-3.60% for S(-) eticlopride, 2.33-4.28 and 0.80-5.73%, for R(+) eticlopride, 3.46-6.84 and 0.80-4.26%, for S(-) sulpiride; and 4.71 -6.47 and 2.00-6.67%, for R(+)-sulpiride, respectively.

Buffers↗
PubMed2000

Synthesis of structured triacylglycerols by lipase-catalyzed acidolysis in a packed bed bioreactor.

Structured triacylglycerols (ST) from canola oil were produced by enzymatic acidolysis in a packed bed bioreactor. A commercially immobilized 1,3-specific lipase, Lipozyme IM, from Rhizomucormiehei, was the biocatalyst and caprylic acid the acyl donor. Parameters such as substrate flow rate, substrate molar ratio, reaction temperature, and substrate water content were examined. High-performance liquid chromatography was used to monitor the reaction and product yields. The study showed that all of the parameters had effects on the yields of the expected di-incorporated (dicaprylic) ST products. Flow rates below 1 mL/min led to reaction equilibrium, and lower flow rates did not raise the incorporation of caprylic acid and the product yield. Incorporation of caprylic acid and the targeted di-incorporated ST was increased by approximately 20% with temperature increase from 40 to 70 degrees C. Increasing the substrate molar ratio from 1:1 to 7:1 increased the incorporation of caprylic acid and the product yield slightly. Water content in the substrate also had a mild influence on the reaction. Water content at 0.08% added to the substrate gave the lowest incorporation and product yield. The use of solvent in the medium was also studied, and results demonstrated that it did not increase the reaction rate at 55 degrees C when 33% hexane (v/v) was added. The main fatty acids at the sn-2 position of the ST were C(18:1), 54. 7 mol %; C(18:2), 30.7 mol %; and C(18:3), 11.0 mol %.

Acids↗
PubMed2000

Isolation of a novel protein tyrosine phosphatase inhibitor, 2-methyl-fervenulone, and its precursors from Streptomyces.

High-throughput screening identified an extract from Streptomyces sp. IM 2096 with inhibitory activity toward several protein tyrosine phosphatases (PTPs). Four 1,2,4-triazine compounds 2096A-D (1-4) were isolated from this extract and their structures elucidated by interpretation of spectroscopic data and confirmed by degradation and synthesis. The novel glycocyamidine derivatives 1 and 2 are diastereomers and may interconvert. Both are inactive in the PTP inhibition assay. Compounds 1 and 2 are unstable and partially decompose to 3 and glycocyamidine (5) at room temperature. Compound 3, known as MSD-92 or 2-methyl-fervenulone, is a broad-specificity PTP inhibitor with comparable potency to vanadate. The imidazo[4, 5-e]-1,2,4-triazine (4), inactive in the PTP-inhibition assay, may be a degradation product of 3.

Enzyme Inhibitors↗
PubMed2000

Anchored multiplex amplification on a microelectronic chip array.

We have developed a method for anchored amplification on a microchip array that allows amplification and detection of multiple targets in an open format. Electronic anchoring of sets of amplification primers in distinct areas on the microchip permitted primer-primer interactions to be reduced and distinct zones of amplification created, thereby increasing the efficiency of the multiplex amplification reactions. We found strand displacement amplification (SDA) to be ideal for use in our microelectronic chip system because of the isothermal nature of the assay, which provides a rapid amplification system readily compatible with simple instrumentation. Anchored SDA supported multiplex DNA or RNA amplification without decreases in amplification efficiency. This microelectronic chip-based amplification system allows multiplexed amplification and detection to be performed on the same platform, streamlining development of any nucleic acid-based assay.

Aromatase↗
PubMed2000

The direction of microsatellite mutations is dependent upon allele length.

Microsatellites, comprising tandemly repeated short nucleotide sequences, are ubiquitous in eukaryotic genomes. Mutations within microsatellites are frequent, altering their overall length by insertion or deletion of a small number of repeat units, with a rate as high as 10(-3) in humans. Despite their high mutability, stable allele frequency distributions are typically observed for microsatellites in humans as well as other primates, although the mechanism maintaining these stable distributions remains unclear. Previous studies have suggested that microsatellite mutations occur more frequently in longer alleles and favour expansion. Generalizing these results has been hindered because the sample sizes were small, only a small subset of alleles for any marker was studied and the direction of mutation (expansion or contraction) was not rigorously determined. Here we examine 236 mutations at 122 tetranucleotide repeat markers and find that the rate of contraction mutations increases exponentially with allele size, whereas the rate of expansion mutations is constant across the entire allele distribution. The overall rate of expansion mutations does not differ from that of contractions. Our findings offer an explanation for the stationary allele distribution of microsatellites.

Alleles↗