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Biomedical subjects

Y Qu

Publications and source records attributed to Y Qu.

At least 91 records · Page 5Linked to original sources
PubMed1998

[Qualitative and quantitative analyses of telomerase activity in cultured cells].

In this study, we established a qualitative PCR-TRAP assay by using SYBR Green stain instead of EB stain; this modification based on Kim's method raised the sensitivity of telomerase detection by 25-100 fold. Besides, a quantitative assay was established by us using 32P-ATP labeled TS primer and the internal control TSK1. We detected the telomerase activity of 12 cultured cells by means of these two assays. The results showed that the telomerase activity could be detected in all the cells, but the activity levels of the cells differed prominently (from 23 to 652 TPG units). The establishment of PCR-TRAP assay and especially the establishment of quantitative assay have enabled us to evaluate the telomerase activity of cells more accurately, and they can be used in our further studies of tumor gene therapy by reducing the telomerase activity of tumor cells.

Breast Neoplasms↗
PubMed1998

Modulation of L-type calcium current kinetics by sarcoplasmic reticulum calcium release in ferret isolated right ventricular myocytes.

The gigaohm seal patch clamp (whole cell configuration) and an internal perfusion technique were used to study the effects of sarcoplasmic reticulum (SR) calcium release on L-type calcium current (ICa,L) in ferret enzymatically isolated right ventricular myocytes. ICa,L (22 to 24 degrees C) was isolated by using various sodium- and potassium-free salines, which eliminated or greatly minimized activation of the sodium-calcium exchanger and calcium-activated cation and anion currents. When calcium was the charge carrier, inactivation of ICa,L was nonmonotonic in many myocytes; after an early rapid phase of inactivation, a secondary inward 'hump' component was frequently observed between -40 to -10 mV. The hump component was not present when barium replaced calcium but was observed when calcium carried the current in low intracellular (aspartate) and extracellular (methanesulphonate) chloride solutions. When BAPTA 10 mM was perfused internally the amplitude of ICa,L increased, the kinetics of inactivation slowed and the hump component disappeared. Both caffeine 10 mM and ryanodine 10 microM increased the amplitude of ICa,L in the hyperpolarized range of potentials (negative to 0 mV), slowed the kinetics of ICa,L inactivation and caused the hump component to disappear. Under current clamp mode, both caffeine and ryanodine significantly prolonged the duration of the action potential. Taken in aggregate, preliminary data demonstrate that, in ferret right ventricular myocytes, a secondary inward hump component can be frequently observed after the early rapid phase of inactivation of ICa,L, causing the net inward current to display biphasic, nonmonotonic behaviour. This secondary inward hump current is only present when calcium is the charge carrier, is absent when BAPTA is used as an intracellular calcium chelator and SR calcium release is disrupted by either caffeine or ryanodine, and is not due to activation of either the sodium-calcium exchanger or various putative calcium-activated cation or anion channels. Rather, preliminary results strongly suggest that this secondary inward hump current component is due to modulation of ICa,L by SR calcium release. Possible physiological and theoretical implications of the results are discussed briefly.

Action Potentials↗
PubMed1997

Determination of serotonin, catecholamines and their metabolites by direct injection of supernatants from chicken brain tissue homogenate using liquid chromatography with electrochemical detection.

An isocratic liquid chromatographic method with electrochemical detection for the determination of L-3,4-dihydroxyphenylalanine, dopamine, norepinephrine, epinephrine, serotonin, and their major metabolites, 3,4-dihydroxyphenylacetic acid, 4-hydroxy-3-methoxyphenylacetic acid and 5-hydroxyindole-3-acetic acid in chicken brain tissue is described. Chickens were killed at different ages, the brains were quickly frozen and 300-microm cryostat sections were made. From these sections, two to six tissue micropunches (1 mm in diameter) were punched out from 20 different areas of the hypothalamus and homogenated in 100 microl 0.1 M perchloric acid which included 0.01% cysteine as antioxidant. Fifty-microl supernatants were injected directly onto the LC system, separated on a 3-microm Phase II ODS column (100 x 3.2 mm I.D.) and detected by an electrochemical detector at a potential of +0.75 V. Standard curves, recoveries, analytical precision and detection limits were investigated for each monoamine neurotransmitter and its metabolites. The method was applied to study the influence of food restriction on the concentration of monoamine neurotransmitters in different brain areas, known to be involved in feeding and reproductive behaviour of female broiler chickens. Over 1000 micropunched tissue samples from ad libitum fed and food-restricted female broiler chickens were analyzed. Our results provide a possible role for catecholamines and indolamines in the altered feeding and reproductive behaviour of the broiler chicken.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1997

Effect of geometric isomerism in dinuclear platinum antitumour complexes on the rate of formation and structure of intrastrand adducts with oligonucleotides.

The dinuclear platinum complexes [[trans -PtCl (NH3)2]2[mu]-[NH2(CH2) n NH2]](NO3)2[1,1/t,t ( n = 4,6)] and [[cis-PtCl(NH3)2]2[mu];-[NH2(CH2) n NH2](NO3) 2[1,1/c,c ( n = 4,6)] exhibit antitumour activity comparable with cisplatin. 1,1/c,c complexes do not form 1,2 GG intrastrand adducts, the major adduct of cisplatin, with double-stranded DNA. This 1H NMR spectroscopy study shows that, in the absence of a complementary strand, 1,1/c,c ( n = 4,6) form a 1,2 GG (N7, N7) intrastrand adduct with r(GpG), d(GpG) and d(TGGT). Initial binding to r(GpG) (and also reaction with GMP) at 37 degrees C was slower for 1,1/c,c compared with 1,1/t,t, whereas the second binding step (adduct closure) was faster for 1,1/c,c. However, the 1H NMR spectra of the 1,1/c,c adducts at 37 degrees C show two H8 signals, one of which is broad and becomes sharper on increasing the temperature, indicating restricted rotation around the Pt-N7 bond. For the d(GpG)-1,1/c,c ( n = 4) adduct, 2D NMR spectroscopy assigned the broad H8 signal to the 3' G, which has syn base orientation and 60% S-type/40% N-type sugar conformation. The 5' G has anti base orientation and S-type sugar conformation. Apart from the restricted rotation around the 3' G, the structure is similar to that of 1,2 GG intrastrand adducts of 1,1/t,t. This steric hindrance may explain the inability of 1,1/c,c complexes to form 1,2 GG intrastrand adducts with sterically more demanding double-stranded DNA.

Acetylcysteine↗
PubMed1997

Proton nuclear magnetic resonance studies on huwentoxin-I from the venom of the spider Selenocosmia huwena: 2. Three-dimensional structure in solution.

The three-dimensional structure in aqueous solution of native huwentoxin-I, a neurotoxin from the venom of the spider Selenocosmia huwena, has been determined from two-dimensional H NMR data recorded at 500 and 600 MHz. Structural constraints consisting of interproton distances inferred from NOEs and dihedral angles from spin-spin coupling constants were used as input for distance geometry calculation with the program XPLOR 3.1. The best 10 structures have NOE violations < 0.3 A, dihedral violations < 2 degrees, and pairwise root-mean-square differences of 1.08 (+/- 0.20) A over backbone atoms (N, C alpha, C). The molecule adopts a compact structure consisting of a small triple-stranded antiparallel beta-sheet and five beta-turns. A small hydrophobic patch consisting of Phe 6, Trp 28, and Trp 31 is located on one side of the molecule. All six lysine residues are distributed on the molecular surface. The three disulfide bridges are buried within the molecule. The structure contains an "inhibitor cystine knot motif" which is adopted by several other small proteins, such as omega-conotoxin, agatoxin IVA, and gurmarin.

Amino Acid Sequence↗
PubMed1997

[Observation on postmortem tracheal cilia of drowning by scanning electron microscope (SEM)].

The changes of tracheal cilia of drowning rat were observed by SEM at 0, 24, 48, 72, 96 h after death. The authors found: the tracheal cilia lost their normal arrangement in some area, and the opening increased on the top of the Goblet cell. There were some thing resembling cotton, or looking like network on the surface of tracheal cilia. At different times after death, the tracheal cilia appeared adhering, downing, and destroying, Goblet cell membrane creasing, the top of cell flattening and the cell collapsing. Observing this changes, we could estimate the time of death.

Animals↗
PubMed1997

A method for comparing positive rates of two blood culture systems.

McNemar's test is used to compare two marginal positive rates from an independent sample of paired binary data. When the pairs are not mutually independent, the McNemar's test may not be valid. In this paper, we propose a random-effects regression model for comparing two marginal probabilities from nonindependent matched pairs data with covariates. An example of comparing positive rates of two blood culture systems illustrates this method. In this example, there is no external gold standard, the paired data are clustered, the data with negative results from both systems are not available, and one culture-specific covariate is involved. The computing method for the maximum likelihood estimation is efficient.

Biometry↗
PubMed1997

Effect on product formation in recombinant Saccharomyces cerevisiae strains expressing different levels of xylose metabolic genes.

The XYL1 and XYL2 genes from Pichia stipitis encoding xylose reductase (XR) and xylilitol dehydrogenase (XDH), respectively, were transformed into Saccharomyces cerevisiae. These two genes were placed in different directions under the control of the alcohol dehydrogenase I (ADHI) and phosphoglycerate kinase (PGK) promoters and inserted into the E. coli-yeast shuttle plasmid YEp24. Different recombinant S. cerevisiae strains were constructed with different specific activities of XR and XDH. The highest XR or XDH activities were obtained when the expressed gene was controlled by the PGK promoter and located downstream after the ADHI promoter-gene-terminator sequence. The XR/XDH ratio (ratio of specific enzyme activities of XR and XDH) in these recombinant S. cerevisiae strains varied from 17.5 to 0.06. In order to enhance xylose utilization, in the XYL1, XYL2 containing S. cerevisiae strains, the native TKL1 gene encoding transketolase and the TALI gene encoding transaldolase were also overexpressed, which showed considerably good growth on the xylose plate. Fermentation of the recombinant S. cerevisiae strains containing XYL1, XYL2, TKL1, and TAL1 were studied with mixtures of glucose and xylose. The strain with XR/XDH ratio of 0.06 consumed 3.25 g/L xylose and formed no xylitol and less glycerol and acetic acid, but more ethanol compared with the strains with a higher XR/XDH ratio.

Aldehyde Reductase↗
PubMed1996

Site-specific d(GpG) intrastrand cross-links formed by dinuclear platinum complexes. Bending and NMR studies.

The novel platinum drugs [{trans-PtCl(NH3)2}2H2N(CH2)nNH2]2+ (1,1/t,t) are currently undergoing preclinical development. The bifunctional DNA binding of these agents allows comparison with that of cisplatin [Farrell et al. (1995) Biochemistry, 34, 15480]. The major DNA lesion of cisplatin, the 1,2-d(GpG) intrastrand adduct, produces a rigid, directed bend 30-35 degrees into the major groove of DNA. We have now completed a structural analysis of the corresponding adduct formed with the dinuclear complexes. Gel retardation assays on 15-22 bp oligonucleotides containing a central d(TG*G*T) site show that the (Pt,Pt)-intrastrand adducts result in a flexible nondirectional bend. This bend is essentially independent of chain length (n = 2, 4, 6). Chemical reactivity assays indicated a hypersensitivity of the thymine 5' to the adduct and an enhanced sensitivity of the 3'-thymine to OsO4. 2D 1H NMR studies on a d(TG1G2T) adduct of [{trans-PtCl(NH3)2}2H2N(CH2)6NH2]2+ have delineated the structural features responsible for these observations. In contrast to the cisplatin adduct, which displays a 100% N-type sugar of the 5'-G and an anti base conformation of the platinated bases in both solid state and solution, the dinuclear adduct does not display the typical N-type sugar pucker. The base orientations are anti (5'-T), anti (G1), anti/syn (G2), and anti (3'-T) while the sugar conformations are N, S/N, N, and S, respectively. The 5'-T remains stacked with its guanine neighbor while the 3'-T becomes unstacked, a reverse of the situation observed for cis-DDP.

Antineoplastic Agents↗
PubMed1996

Molecular determinants of high affinity binding of alpha-scorpion toxin and sea anemone toxin in the S3-S4 extracellular loop in domain IV of the Na+ channel alpha subunit.

alpha-Scorpion toxins and sea anemone toxins bind to a common extracellular site on the Na+ channel and inhibit fast inactivation. Basic amino acids of the toxins and domains I and IV of the Na+ channel alpha subunit have been previously implicated in toxin binding. To identify acidic residues required for toxin binding, extracellular acidic amino acids in domains I and IV of the type IIa Na+ channel alpha subunit were converted to neutral or basic amino acids using site-directed mutagenesis, and altered channels were transiently expressed in tsA-201 cells and tested for 125I-alpha-scorpion toxin binding. Conversion of Glu1613 at the extracellular end of transmembrane segment IVS3 to Arg or His blocked measurable alpha-scorpion toxin binding, but did not affect the level of expression or saxitoxin binding affinity. Conversion of individual residues in the IVS3-S4 extracellular loop to differently charged residues or to Ala identified seven additional residues whose mutation caused significant effects on binding of alpha-scorpion toxin or sea anemone toxin. Moreover, chimeric Na+ channels in which amino acid residues at the extracellular end of segment IVS3 of the alpha subunit of cardiac Na+ channels were substituted into the type IIa channel sequence had reduced affinity for alpha-scorpion toxin characteristic of cardiac Na+ channels. Electrophysiological analysis showed that E1613R has 62- and 82-fold lower affinities for alpha-scorpion and sea anemone toxins, respectively. Dissociation of alpha-scorpion toxin is substantially accelerated at all potentials compared to wild-type channels. alpha-Scorpion toxin binding to wild type and E1613R had similar voltage dependence, which was slightly more positive and steeper than the voltage dependence of steady-state inactivation. These results indicate that nonidentical amino acids of the IVS3-S4 loop participate in alpha-scorpion toxin and sea anemone toxin binding to overlapping sites and that neighboring amino acid residues in the IVS3 segment contribute to the difference in alpha-scorpion toxin binding affinity between cardiac and neuronal Na+ channels. The results also support the hypothesis that this region of the Na+ channel is important for coupling channel activation to fast inactivation.

Amino Acid Sequence↗
PubMed1996

Molecular prenatal diagnosis of glycogen storage disease type Ia.

Glycogen storage disease type Ia (GSD Ia, von Gierke disease) is an autosomal recessive inborn error of metabolism caused by the deficiency of D-glucose-6-phosphatase (G6Pase). Since this enzyme is expressed primarily in hepatocytes, couples at risk for GSD type Ia relied on fetal liver biopsy for prenatal diagnosis. The recent isolation of the G6Pase gene and identification of several disease-causing mutations have permitted molecular prenatal diagnosis using amniocytes or chorionic villi. Chorionic villus sampling (CVS) was performed in an Ashkenazi Jewish family in whom a previous child was homoallelic and both parents were heterozygous for the R83C mutation. Molecular analysis revealed that the fetus was not affected. The prenatal diagnosis was confirmed postnatally by biochemical and molecular studies. Thus, the molecular prenatal diagnosis of GSD type Ia can be safely and accurately made in the first trimester.

Base Sequence↗
PubMed1996

Adaptive motor unit action potential clustering using shape and temporal information.

An adaptive algorithm is described that groups motor unit action potentials (MUAPs), detected in a composite EMG signal during signal decomposition, and creates partial motor unit action potential trains (MUAPTs). Data-driven MUAP shape and motor unit firing-pattern based criteria are used to form the clusters. An algorithm for estimating MUAPT temporal parameters, which provides accurate estimates even for partially defined trains, is used to obtain firing-pattern information. No a priori knowledge is required regarding the number of clusters or the distribution of their template shapes. The clustering algorithm when applied to real concentric-needle detected MUAP data provides accurate and useful clustering results. Compared to a classical leader-based algorithm, it provides more robust performance, is better able to estimate the true number of motor units represented in a set of detected MUAPs, and obtains more complete and accurate MUAPTs.

Action Potentials↗
PubMed1996

Robust method for estimating motor unit firing-pattern statistics.

An error-filtered estimation (EFE) algorithm for estimating the mean and standard deviation of a set of time intervals between consecutive motor unit firing times (inter-pulse intervals (IPIs)) is described. As the input IPI data are filtered and only valid IPIs are used to estimate mean and standard deviation values, the EFE algorithm provides accurate estimates even when the data defining the train of motor unit firing times are only partially complete or have several erroneous firing times. The algorithm has been evaluated using both simulated and real motor unit firing time data, and has been found to provide accurate and unbiased mean and standard deviation estimates, even when up to 70% of the IPI data are incorrect.

Electromyography↗
PubMed1996

A circular dichroism study of ethidium bromide binding to Z-DNA induced by dinuclear platinum complexes.

Dinuclear bis(platinum) complexes have been shown previously to induce the B-->Z transition in synthetic DNAs (Nucleic Acids Res. 7, 1697-1703, J. Inorganic Biochem. 54, 207-220). In this paper, the reversibility of the Z conformation back to the B form was assessed by treatment of the induced Z form in poly(dG-dC).poly(dG-dC) with ethidium bromide (Etd). Z-DNA induced by the tetra-amine cations [{Pt(NH3)3}2(H2N(CH2)nNH2)]4+, which are capable of only electrostatic interactions with the polynucleotide, was readily reversible. The spectroscopic data mirrored that of ethidium bromide/poly(dG-dC).poly(dG-dC) in the presence of 4.4 M NaCl. In contrast, Z-DNA induced by the bifunctional complexes [{trans-PtCl(NH3)2}2(H2N(CH2)nNH2)]2+ did not produce spectra typical of Etd intercalation and reversal to B-form DNA. The original Z-form CD spectra of DNA treated with the bifunctional complexes could be reobtained following removal of Etd by extensive dialysis. The bifunctional complexes are very effective interstrand cross-linking agents. The data suggest that interstrand cross-linking by dinuclear complexes can stabilize or "lock" the Z-conformation prohibiting its reversal to the B-form. The implications for the biological activity of the dinuclear complexes are briefly discussed.

Circular Dichroism↗
PubMed1996

A SAS macro for stepwise correlated binary regression.

Several regression methods have been proposed for the analysis of correlated binary data, but none deals with the selection of covariates when there exist a large number of potentially relevant covariates. We present a SAS macro based on a stepwise selection procedure for the analysis of correlated binary data. Using regression methods based on generalized estimating equations originally proposed by Liang and Zeger and extended by Prentice, we describe a score test for forward selection, a Wald's test for backward elimination, and a test for model adequacy based on generalized scores. The methodology and the accompanying computer macro program written in SAS IML are illustrated with data from a prospective study of functional decline in the activities of daily living in a group of elderly patients.

Activities of Daily Living↗
PubMed1996

Phosphorylation of S1505 in the cardiac Na+ channel inactivation gate is required for modulation by protein kinase C.

Inactivation of both brain and cardiac Na+ channels is modulated by activation of protein kinase C (PKC) but in different ways. Previous experiments had shown that phosphorylation of serine 1506 in the highly conserved loop connecting homologous domains III and IV (LIII/IV) of the brain Na+ channel alpha subunit is necessary for all effects of PKC. Here we examine the importance of the analogous serine for the different modulation of the rH1 cardiac Na+ channel. Serine 1505 of rH1 was mutated to alanine to prevent its phosphorylation, and the resulting mutant channel was expressed in 1610 cells. Electrophysiological properties of these mutant channels were indistinguishable from those of wild-type (WT) rH1 channels. Activation of PKC with 1-oleoyl-2-acetyl-sn-glycerol (OAG) reduced WT Na+ current by 49.3 +/- 4.2% (P < 0.01) but S1505A mutant current was reduced by only 8.5 +/- 5.4% (P = 0.29) when the holding potential was -94 mV. PKC activation also caused a -17-mV shift in the voltage dependence of steady-state inactivation of the WT channel which was abolished in the mutant. Thus, phosphorylation of serine 1505 is required for both the negative shift in the inactivation curve and the reduction in Na+ current by PKC. Phosphorylation of S1505/1506 has common and divergent effects in brain and cardiac Na+ channels. In both brain and cardiac Na+ channels, phosphorylation of this site by PKC is required for reduction of peak Na+ current. However, phosphorylation of S1506 in brain Na+ channels slows and destabilizes inactivation of the open channel. Phosphorylation of S1505 in cardiac, but not S1506 in brain, Na+ channels causes a negative shift in the inactivation curve, indicating that it stabilizes inactivation from closed states. Since LIII/IV containing S1505/S1506 is completely conserved, interaction of the phosphorylated serine with other regions of the channel must differ in the two channel types.

Animals↗
PubMed1996

Random effects models in latent class analysis for evaluating accuracy of diagnostic tests.

When the results of a reference (or gold standard) test are missing or not error-free, the accuracy of diagnostic tests is often assessed through latent class models with two latent classes, representing diseased or nondiseased status. Such models, however, require that conditional on the true disease status, the tests are statistically independent, an assumption often violated in practice. Consequently, the model generally fits the data poorly. In this paper, we develop a general latent class model with random effects to model the conditional dependence among multiple diagnostic tests (or readers). We also develop a graphical method for checking whether or not the conditional dependence is of concern and for identifying the pattern of the correlation. Using the random-effects model and the graphical method, a simple adequate model that is easy to interpret can be obtained. The methods are illustrated with three examples from the biometric literature. The proposed methodology is also applicable when the true disease status is indeed known and conditional dependence could well be present.

AIDS Serodiagnosis↗