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Biomedical subjects

L Yang

Publications and source records attributed to L Yang.

At least 307 records · Page 17Linked to original sources
PubMed1999

Regulation of apoptosis in mature alphabeta+CD4-CD8- antigen-specific suppressor T cell clones.

The regulation of apoptosis in mature CD4+ or CD8+ alphabeta+ T cells has been well studied. How the survival and death is regulated in peripheral CD4-CD8- (double negative, DN) alphabeta+ T cells remains unknown. Recent studies suggest that peripheral DN T cells may play an important role in the regulation of the immune responses mediated by CD4+ or CD8+ T cells. Here, we used immunosuppressive DN T cell clones to elucidate the mechanisms involved in the regulation of death and survival of alphabeta+ DN T cells. The DN T cell clones were generated from the spleen cells of 2C transgenic mice, which express the transgenic TCR specific for Ld and permanently accepted Ld+ skin allografts after pretransplant infusion of Ld+ lymphocytes. We report that 1) the mature DN T cells are highly resistant to TCR cross-linking-induced apoptosis in the presence of exogenous IL-4; 2) Fas/Fas-ligand and TNF-alpha/TNFR pathways do not play an apparent role in regulating apoptosis in DN T cells; 3) the DN T cells constitutively express a high level of Bcl-xL, but not Bcl-2; 4) both Bcl-xL and Bcl-2 are up-regulated following TCR-cross-linking; and 5) IL-4 stimulation significantly up-regulates Bcl-xL and c-Jun expression and leads to mitogen-activated protein kinase phosphorylation in DN T cells, which may contribute to the resistance to apoptosis in these T cells. Taken together, these results provide us with an insight into how mature DN T cells resist activation-induced apoptosis to provide a long-term suppressor function in vivo.

Animals↗
PubMed1999

Inhibitory effects of beraprost on platelet aggregation: comparative study utilizing two methods of aggregometry.

We evaluated the inhibitory effects of beraprost, a stable prostacyclin analogue, on platelet aggregation, assessed by two methods of platelet aggregometry. The conventional aggregometry detects changes in light transmission (LT) of a platelet suspension, and a recently developed aggregometry based upon a particle counting principle detects light scattering (LS) generated by platelet aggregates. Since LS is more sensitive than LT in detecting platelet aggregates of small size, the minimal concentrations of agonists (ADP, epinephrine, collagen, and U46619) to induce detectable aggregate formation were consistently lower with LS (1/2 to 1/6) than with LT. The effects of beraprost were evaluated on platelet aggregation induced by the optimal concentrations of agonists thus determined for each sample. The IC50 values of beraprost on platelet aggregation, as assessed by LS, were 1/2 to 1/10 of those assessed by LT. In suppressing platelet aggregation assessed by LS, beraprost was especially potent with IC50 of 0.2-0.5 nM when platelets were activated by U46619, a thromboxane A2 analogue, or low concentrations of collagen which activates platelets through thromboxane A2 production. The IC50 values were 2-5 nM with ADP and epinephrine, which induce the formation of small aggregates independently of thromboxane A2 production. These findings suggest that LS can detect inhibitory effects of lower concentrations of antiplatelet agents, since it detects the formation of small aggregates induced by agonists in the lower concentration range than LT. It is also suggested that beraprost potently inhibits thromboxane A2-elicited initial signal transduction pathway, reflected by the formation of small aggregates.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1999

Time-resolved fluorescence anisotropy study of the refolding reaction of the alpha-subunit of tryptophan synthase reveals nonmonotonic behavior of the rotational correlation time.

Time-resolved fluorescence anisotropy of a bound extrinsic probe was studied in an effort to characterize dynamic properties of the transient partially folded forms that appear during the folding of the alpha-subunit of tryptophan synthase (alphaTS) from Escherichia coli. Previous studies have shown that alphaTS, a single structural domain, can be cleaved into autonomously folding amino- and carboxy-folding units comprising residues 1-188 and 189-268, respectively [Higgins, W., Fairwell, T., and Miles, E. W. (1979) Biochemistry 18, 4827-4835]. By use of a double-kinetic approach [Jones, B. E., Beechem, J. M., and Matthews, C. R. (1995) Biochemistry 34, 1867-1877], the rotational correlation time of 1-anilino-8-naphthalene sulfonate bound to nonpolar surfaces of folding intermediates was measured by time-correlated single photon counting at varying time delays following initiation of folding from the urea-denatured form by stopped-flow techniques. Comparison of the rotational correlation times for the full-length alphaTS and the amino-terminal fragment suggests that folding of the amino-terminal fragment and carboxy-terminal fragment is coordinated, not autonomous, on the milliseconds to seconds time scale. If a spherical shape is assumed, the apparent hydrodynamic radius of alphaTS after 5 ms is 26.8 A. The radius increases to 28.5 A by 1 s before decreasing to the radius for native alphaTS, 24.7 A, on a longer time scale (>25 s). Viewed within the context of the kinetic folding model of alphaTS [Bilsel, O., Zitzewitz, J. A., Bowers, K. E. , and Matthews, C. R. (1999) Biochemistry 38, 1018-1029], the initial collapse reflects the formation of an off-pathway burst-phase intermediate in which at least part of the carboxy folding unit interacts with the amino folding unit. The subsequent increase in rotational correlation time corresponds to the formation of an on-pathway intermediate that leads to the native conformation. The apparent increase in the radius for the on-pathway intermediate may reflect a change in the interaction of the two-folding units, thereby forming a direct precursor for the alpha/beta barrel structure.

Anilino Naphthalenesulfonates↗
PubMed1999

Incorporating qualitative knowledge in enzyme kinetic models using fuzzy logic.

Modeling of metabolic pathway dynamics requires detailed kinetic equations at the enzyme level. In particular, the kinetic equations must account for metabolite effectors that contribute significantly to the pathway regulation in vivo. Unfortunately, most kinetic rate laws available in the literature do not consider all the effectors simultaneously, and much kinetic information exists in a qualitative or semiquantitative form. In this article, we present a strategy to incorporate such information into the kinetic equation. This strategy uses fuzzy logic-based factors to modify algebraic rate laws that account for partial kinetic characteristics. The parameters introduced by the fuzzy factors are then optimized by use of a hybrid of simplex and genetic algorithms. The resulting model provides a flexible form that can simulate various kinetic behaviors. Such kinetic models are suitable for pathway modeling without complete enzyme mechanisms. Three enzymes in Escherichia coli central metabolism are used as examples: phosphoenolpyruvate carboxylase; phosphoenolpyruvate carboxykinase; and pyruvate kinase I. Results show that, with fuzzy logic-augmented models, the kinetic data can be much better described. In particular, complex behavior, such as allosteric inhibition, can be captured using fuzzy rules. The resulting models, even though they do not provide additional physical meaning in enzyme mechanisms, allow the model to incorporate semiquantitative information in metabolic pathway models.

Bacterial Proteins↗
PubMed1999

The superior olivary nucleus and its influence on nucleus laminaris: a source of inhibitory feedback for coincidence detection in the avian auditory brainstem.

Located in the ventrolateral region of the avian brainstem, the superior olivary nucleus (SON) receives inputs from nucleus angularis (NA) and nucleus laminaris (NL) and projects back to NA, NL, and nucleus magnocellularis (NM). The reciprocal connections between the SON and NL are of particular interest because they constitute a feedback circuit for coincidence detection. In the present study, the chick SON was investigated. In vivo tracing studies show that the SON projects predominantly to the ipsilateral NM, NL, and NA. In vitro whole-cell recording reveals single-cell morphology, firing properties, and postsynaptic responses. SON neurons are morphologically and physiologically suited for temporal integration; their firing patterns do not reflect the temporal structure of their excitatory inputs. Of most interest, direct stimulation of the SON evokes long-lasting inhibition in NL neurons. The inhibition blocks both intrinsic spike generation and orthodromically evoked activity in NL neurons and can be eliminated by bicuculline methiodide, a potent antagonist for GABAA receptor-mediated neurotransmission. These results strongly suggest that the SON provides GABAergic inhibitory feedback to laminaris neurons. We discuss a mechanism whereby SON-evoked GABAergic inhibition can influence the coding of interaural time differences for sound localization in the avian auditory brainstem.

Animals↗
PubMed1999

A new intragastric delivery system for the treatment of Helicobacter pylori associated gastric ulcer: in vitro evaluation.

A new strategy is proposed for the triple drug treatment (tetracycline, metronidazole and bismuth salt) of Helicobacter pylori associated peptic ulcers. The design of the delivery system was based on the swellable asymmetric triple layer tablet approach, with floating feature in order to prolong the gastric retention time of the delivery system. Hydroxypropylmethylcellulose and poly(ethylene oxide) were the major rate-controlling polymeric excipients. Tetracycline and metronidazole were incorporated into the core layer of the triple-layer matrix for controlled delivery, while bismuth salt could be included in one of the outer layers for instant release. The concentration of tetracycline and metronidazole released over time was determined simultaneously on a gradient high-performance liquid chromatography system. Results demonstrated that sustained delivery of tetracycline and metronidazole over 6-8 h can be easily achieved while the tablet remained afloat. The floating aspect was envisaged to extend the gastric retention time of the designed system to maintain effective localized concentration of tetracycline and metronidazole. Additionally, the developed HPLC method for the concurrent determination of tetracycline and metronidazole was proved to be rapid and accurate. The developed delivery system has potential to increase the efficacy of the therapy and improve patient compliance.

Algorithms↗
PubMed1999

Potent, orally bioavailable somatostatin agonists: good absorption achieved by urea backbone cyclization.

Backbone cyclization of urea-based somatostatin agonists resulted in novel, orally bioavailable agonists. Binding assays confirmed that the resulting conformationally constrained cyclic ureas retained the potency of their acyclic counterparts. SAR studies subsequently led to highly potent analogs, selective for receptor subtype 2, and having good oral bioavailability.

Administration, Oral↗
PubMed1999

An additional region of coactivator GRIP1 required for interaction with the hormone-binding domains of a subset of nuclear receptors.

Transcriptional coactivators of the p160 family (SRC-1, GRIP1, and p/CIP) associate with DNA-bound nuclear receptors (NRs) and help the NRs to recruit an active transcription initiation complex to the promoters of target genes. Previous studies have demonstrated the importance of the NR interaction domain (NID) of p160 proteins containing three NR box motifs (LXXLL) for the interaction with the hormone-binding domains of NRs. Here we report that, in addition to NID, another region of coactivator GRIP1 (amino acids 1011-1121), called the auxiliary NID (NIDaux), is required in vitro and in vivo for efficient interaction with a subset of NRs, including the glucocorticoid receptor (GR), androgen receptor, and retinoic acid receptor alpha. A second group of NRs, which includes the progesterone receptor, retinoid X receptor alpha, thyroid hormone receptor beta1, and vitamin D receptor, required only NID for efficient interaction. For binding to GR, the NID and NIDaux of GRIP1 must act in cis, but deletion of up to 144 amino acids between the two regions did not reduce binding efficiency. Amino acids 1011-1121 of GRIP1 also contain a p300 interaction domain, but mutational analysis indicated that the p300 interaction function within this region is separable from the ability to contribute to GR hormone-binding domain binding. SRC-1 lacks an NIDaux activity equivalent to that in GRIP1.

Amino Acid Sequence↗
PubMed1999

Nanocrystalline Silver Particles: Synthesis, Agglomeration, and Sputtering Induced by Electron Beam.

Nanocrystalline silver particles with an average size of 15 nm have been prepared by reducing Ag(NH3)+2 with N2H4. H2O, with sodium dodecylbenzene sulfate (DBS) used as the stabilizing agent. The influence of DBS in the reaction is discussed. When the so-prepared silver particles were observed in situ under a transmission electron microscope, we observed an agglomeration and sputtering phenomenon induced by the electron beam. The possible reason for agglomeration and sputtering is proposed. Silver crystals of hexagonal shape, ranging from 20- to 120-nm in size, were formed on a copper grid. This phenomenon may offer a useful means of forming small particles. Copyright 1999 Academic Press.

Journal Article↗
PubMed1999

Influence of 5-bromodeoxycytosine substitution on triplex DNA stability and conformation.

Three triple-helical hairpin DNAs with substitution of 5-bromocytosine for cytosine in different strands have been investigated by molecular mechanics and Raman spectroscopy. The stability of the three substituted triplexes were compared with the corresponding unsubstituted triplex DNA by the molecular mechanics method. Base stacking interactions and strand--strand interactions of each triplex were analyzed in detail. Sugar conformations in these triplexes have been determined by both vibrational spectroscopy and molecular dynamics simulation. The hairpin triplexes with substitution occurring in strand I or both in strands I and III have the main sugar conformation of C3'-endo, while the triplex with substitution occurring in strand III is the combination of C3'-endo and C2'-endo sugar conformation. Theoretical results are basically in agreement with experiments.

Base Sequence↗
PubMed1999

Membrane supports as the stationary phase in high-performance immunoaffinity chromatography.

The membrane with a composite of cellulose grafted with acrylic polymers formed by polymerizing a glycidyl methacrylate in the presence of dispersed cellulose fiber was prepared as the stationary phase; the column (40 x 4 mm i.d.) which was compatible with the HPLC instrument was packed with pieces of the cut membrane. Protein A and human IgG were immobilized on the membrane stationary phase. The column based on the membrane support provided us good reproducibility, high efficiency, and low back pressure. High-performance immunoaffinity chromatographic analysis of human IgG in serum and polyclonal antibody to human IgG raised in goat was performed within 2.5 min. The fast-speed immunoaffinity analysis was developed by increasing the flow rate of the mobile phase and decreasing the duration time for the switch of the mobile phase; an operation for immunoaffinity analysis of human IgG could be finished within 30 s.

Cellulose↗
PubMed1999

DNA damage and p21(WAF1/CIP1/SDI1) in experimental injury of the rat adrenal cortex and trauma-associated damage of the human adrenal cortex.

In vivo models are needed to study the reactions of tissues to DNA damage, such as the induction of the cyclin-dependent kinase inhibitor p21, indicating potential repair of the damage, versus apoptosis, indicating the elimination of the damaged cells. Damage to DNA occurs in tissues during shock, sepsis, and other critical medical conditions. Previous studies have found evidence of damage to the cortex of adrenal glands from organ donors who had undergone severe trauma prior to death. The present experiment studied rats under experimental interventions of clinical relevance to patients with conditions that put them at risk for damage to the adrenal glands. These interventions comprised ischaemia and reperfusion injury, sepsis following caecal ligation and puncture, acute pancreatitis, and administration of chemical agents (zymosan and acrylonitrile). All the interventions caused an increase in p21 mRNA as assessed by northern blotting and in situ hybridization. Increased nuclear p21 protein was shown by immunohistochemistry. All the interventions caused damage to DNA, as shown by labelling of available 3' termini of single-strand breaks with terminal transferase. The number of cells undergoing apoptosis, visualized by ligation of a hairpin oligonucleotide probe to double-strand breaks in DNA, was much lower. In rat adrenal glands, apoptotic cells were infrequent under all the conditions studied. They were more abundant in human organ donor adrenal glands that were previously shown to have extensive DNA damage accompanied by induction of p21. The similarity of the effects of a wide variety of surgical interventions and chemical agents suggest a common pathophysiological mechanism which is not specific to the initiating injury. Experimental injury of the rat adrenal cortex provides a model for investigating the role of organ DNA damage and of mediators of the response to DNA damage, such as p21.

Adrenal Cortex↗
PubMed1999

Hydration effects on the electrostatic potential around tuftsin.

The electrostatic potential and component dielectric constants from molecular dynamics (MD) trajectories of tuftsin, a tetrapeptide with the amino acid sequence Thr-Lys-Pro-Arg in water and in saline solution are presented. The results obtained from the analysis of the MD trajectories for the total electrostatic potential at points on a grid using the Ewald technique are compared with the solution to the Poisson-Boltzmann (PB) equation. The latter was solved using several sets of dielectric constant parameters. The effects of structural averaging on the PB results were also considered. Solute conformational mobility in simulations gives rise to an electrostatic potential map around the solute dominated by the solute monopole (or lowest order multipole). The detailed spatial variation of the electrostatic potential on the molecular surface brought about by the compounded effects of the distribution of water and ions close to the peptide, solvent mobility, and solute conformational mobility are not qualitatively reproducible from a reparametrization of the input solute and solvent dielectric constants to the PB equation for a single structure or for structurally averaged PB calculations. Nevertheless, by fitting the PB to the MD electrostatic potential surfaces with the dielectric constants as fitting parameters, we found that the values that give the best fit are the values calculated from the MD trajectories. Implications of using such field calculations on the design of tuftsin peptide analogues are discussed.

Computer Simulation↗
PubMed1999

The FANCA gene in Japanese Fanconi anemia: reports of eight novel mutations and analysis of sequence variability.

Fanconi anemia (FA), an autosomal recessive disorder characterized by a progressive pancytopenia associated with congenital anomalies and high predisposition to malignancies, is a genetically and clinically heterogeneous disease. At least eight complementation groups (FA-A to FA-H) have been identified with their relative prevalence varying among the ethnical backgrounds. Recently, responsible genes, FANCA and FANCC, have been cloned. This report describes mutations of the FANCA gene, which we studied by direct sequencing of cDNA with confirmation on genomic DNA in 15 unclassified Japanese FA patients. A total of 19 sequence alterations were identified, of which 10 (six missense and four silent alterations) were likely to be nonpathogenic polymorphism. The remaining nine alterations, of which eight were novel mutations, were assumed to be pathogenic and consisted of two missense mutations and seven mutations resulting in truncation of gene product, demonstrating a wide allelic heterogeneity. The pathogenic mutations were found in 12 patients (80%); they were either homozygous or compound heterozygous in 10 patients, apparently heterozygous in two patients and none in three patients. We conclude that the sequence variability is intrinsic to the FANCA gene and that the relative prevalence of the FA-A subtype is unusually high in Japanese FA patients.

DNA Mutational Analysis↗
PubMed1999

Immobilized iminodiacetic acid (IDA)-type Cu2+ -chelating membrane affinity chromatography for purification of bovine liver catalase.

A metal ion chelating membrane medium based on iminodiacetate-substituted modified short cotton cellulose was examined for the purification of bovine liver catalase (BLC). The effect of buffer pH, chelator surface density, initial concentration of crude enzyme and flow rate on BLC binding efficiency to the copper ion chelating membrane adsorbent were examined. Under the chromatographic conditions chosen, 67.7% recovery of BLC was attained with an overall 4.2-fold increase in specific activity in a single step. After performance of BLC purification, the chelating membrane adsorbent can be easily regenerated by imidazole or EDTA buffer with higher reviving effectiveness with the latter.

Animals↗
PubMed1999

Protein A tangential flow affinity membrane cartridge for extracorporeal immunoadsorption therapy.

Tangential flow affinity membrane cartridge (TFAMC) is a new model of immunoadsorption therapy for hemoperfusion. Recombinant Protein A was immobilized on the membrane cartridge through Schiff base formation for extracorporeal IgG and immune complex removal from blood. Flow characteristics, immunoadsorption capacity and biocompatibility of protein A TFAMC were studied. The results showed that the pressure drop increased with the increasing flow rate of water, plasma and blood, demonstrating reliable strength of membrane at high flow rate. The adsorption capacities of protein A TFAMC for IgG from human plasma and blood were measured. The cartridge with 139 mg protein A immobilized on the matrix (6 mg protein A/g dry matrix) adsorbed 553 mg IgG (23.8 mg IgG/g dry matrix) from human plasma and 499.4 mg IgG (21.5 mg IgG/g dry matrix) from human blood, respectively. The circulation time had a major influence on IgG adsorption capacity, but the flow rate had little influence. Experiments in vitro and in vivo confirmed that protein A TFAMC mainly adsorbed IgG and little of other plasma proteins, and that blood cell damage was negligible. The extracorporeal circulation system is safe and reliable.

Animals↗
PubMed1999

Enantiometric separation of amino acids by copper (II)-L-arginine ligand exchange capillary zone electrophoresis.

A novel method for the separation of amino acid enantiomers by ligand exchange capillary zone electrophoresis (CZE) using a chiral Cu(II)-L-arginine ligand was developed in this paper. The effects of the buffer concentration, pH, the molar ratio of L-arginine to Cu(II), and the total concentrations of Cu(II) and L-arginine on the migration times and resolutions of chiral amino acids were investigated. The optimum experimental conditions were 40 mmol/L of NH4OAc, pH 7.0, the 2:1 concentration ratio of L-arginine to Cu(II), 1 mmol/L CuSO4 x 5H2O and 2 mmol/L L-arginine. Using the proposed method, six different dansyl-amino acids were separated successfully with the resolution ranging from 1.1 to 2.3 in less than 15 min.

Amino Acids↗