Biomedical subjects

S Chen

Publications and source records attributed to S Chen.

At least 505 records · Page 28Linked to original sources

PubMed2000

[In situ FTIR spectrocopic studies of CO and CN- adsorbed on electrode of nanometer thin film of palladium in alkaline solutions].

The adsorption and coadsorption of CO and CN- on electrode of nanometer thin film of palladium (nm-Pd/GC) was studied by using in situ FTIR spectroscopy. The results demonstrated that the nm-Pd/GC electrode exhibited abnormal infrared effects (AIREs) in alkaline solutions as in acid solutions. The characters of AIREs, i.e., the inversion of the direction of IR band, the augmentation of FWHM and the significant enhancement of IR absorption by adsorbed species were observed in all cases of adsorption and coadsorption of CO and CN-. It has been illustrated that strong interaction is existed between adsorbed CO and CN- on nm-Pd/GC surface. The study has contributed towards understanding the surface processes of chemisorption as well as to reveal the origin of nanometer size-effects of thin film materials.

English Abstract↗

PubMed2000

[In situ FTIRS studies of CO2 reduction on thin film alloy electrodes].

The nanometer thin film alloy electrode (Sb-Pb-Pt/GC) was prepared by electrochemical methods. Excellent catalytic effect was observed on these film alloy material in the electro-reduction of carbon dioxide (CO2). The reduction can take place at potentials as high as -0.4 V. The oxidation of reduced products of CO2 on Sb-Pb-Pt/GC yielded current peaks at arrowed -0.08, 0.2 and 0.3 V, while the oxidation of reduction products on a nm-Pt/GC electrode gave rise a current peak near 0.4 V. The in situ FTIR spectroscopic studies confirmed at a molecule level that the products of CO2 reduction on Sb-Pb-Pt/GC are mainly organic acid and alcoholic compounds, while on a nm-Pt/GC electrode the production of CO2 is mainly CO species.

Alloys↗

PubMed2000

[In situ FTIR studies of CO adsorption on palladium loading zeolite electrode].

Adsorption of CO on palladium loading zeolite electrode supported on Pt or GC substrate was studied by in situ MSFTIR spectroscopy. The results demonstrated that the geometry of faujasite supercages favors the formation of bridge-bonded CO species. The enhanced IR adsorption of CO adsorbed on Pd clusters located in the supercages of zeolite has been observed for the first time, which is a new phenomenon and differs from the abnormal infrared effects discovered by our group and the surface enhanced infrared absorption phenomenon reported in the literature.

English Abstract↗

PubMed2000

[In situ FTIRS studies of reduction of [OsVI(N)(NH3)4](CF3SO3)3 in acetonitrile].

In this paper, the electrochemical cyclic voltammetry, UV/Vis and in situ FTIR spectroscopy were used to study the electrochemical reduction of [OsVI(N)(NH3)4](CF3SO3)3 on GC and Pt electrode in acetonitrile. The results demonstrated that the N-N coupling can occur between [OsVI(N)(NH3)4]3+ and the osmium(V) species generated at the electrode. It has revealed for the first time, by in situ FTIR spectroscopy, that the IR absorption of the nu (N identical to N) mode of the mu-dinitrogen osmium[III, II] and [II, II] complex yields positive-going band at around 2,019 cm-1 and negative-going band near 1,970 cm-1, respectively. The results provided new insights in understanding the mechanism of the coupling reaction [OsV identical to N][OsVI identical to N]<=>[OsII-N identical to N-OsIII].

English Abstract↗

PubMed1999

Interdomain signaling in glutamine phosphoribosylpyrophosphate amidotransferase.

The glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase-catalyzed synthesis of phosphoribosylamine from PRPP and glutamine is the sum of two half-reactions at separated catalytic sites in different domains. Binding of PRPP to a C-terminal phosphoribosyltransferase domain is required to activate the reaction at the N-terminal glutaminase domain. Interdomain signaling was monitored by intrinsic tryptophan fluorescence and by measurements of glutamine binding and glutamine site catalysis. Enzymes were engineered to contain a single tryptophan fluorescence reporter in key positions in the glutaminase domain. Trp(83) in the glutamine loop (residues 73-84) and Trp(482) in the C-terminal helix (residues 471-492) reported fluorescence changes in the glutaminase domain upon binding of PRPP and glutamine. The fluorescence changes were perturbed by Ile(335) and Tyr(74) mutations that disrupt interdomain signaling. Fluoresence titrations of PRPP and glutamine binding indicated that signaling defects increased the K(d) for glutamine but had little or no effect on PRPP binding. It was concluded that the contact between Ile(335) in the phosphoribosyltransferase domain and Tyr(74) in the glutamine site is a primary molecular interaction for interdomain signaling. Analysis of enzymes with mutations in the glutaminase domain C-terminal helix and a 404-420 peptide point to additional signaling interactions that activate the glutamine site when PRPP binds.

Amidophosphoribosyltransferase↗

PubMed1999

Characterization of the molecular species of phosphatidylethanolamine from kidney of the fresh water snail Lymneae stagnalis by mass spectrometry.

The structural analysis of sixteen molecular species of diacyl glycerophosphoethanolamine from fresh water snail Lymneae stagnalis kidney using chromatography and mass spectrometry is described in this paper. 1-eicosadienoyl-2-eicosatetraenoyl-sn-glycero-3-phosphoethanolamine (PE 20:2-20:4), 1-eicosapentaenoyl-2-eicosadienoyl-sn-glycero-3-phosphoethanolamine (PE 20:5-20:2) and 1-eicosatrienoyl-2-eicosatetraenoyl-sn-glycero-3-phosphoethanolamine (PE 20:3-20:4) as well as 1-octadecanoyl-2-eicosatetraenoyl-sn-glycero-3-phosphoethanolamine (PE 18:0-20:4), 1-ocetadecenoyl-2-eicosatetraenoyl-sn-glycero-3-phosphoethanolamine (PE 18:1-20:4) and 1-octadecanoyl-2-eicosapentaneoyl-sn-glycero-3-phosphoethanolamine (PE 18:0-20:5) were found as the major molecular species, and the first three were tentatively identified as the novel species present in this biological material. The presence of a relatively high content of 1,2-dieicosenoyl-sn-glycero-3-phosphoethanolamine species (approximately 27% of total species) as well as the absence of 22-carbon fatty acid containing and plasmalogen PE molecular species are remarkable in healthy Lymneae stagnalis kidney. Copyright 1999 John Wiley & Sons, Ltd.

Journal Article↗

PubMed1999

Concomitant administration of sodium dichloroacetate and thiamine in west syndrome caused by thiamine-responsive pyruvate dehydrogenase complex deficiency.

We treated a female patient with West syndrome caused by thiamine-responsive pyruvate dehydrogenase complex (PDHC) deficiency. Infantile spasms occurred in association with elevated blood and CSF lactate concentrations; these symptoms disappeared when lactate concentrations had been lowered by treatment with concomitant sodium dichloroacetate (DCA) and high dose thiamine. Sequencing the patient's PDHC E(1)alpha subunit revealed a substitution of serine for glycine at position 89 in exon 3 (G89S). This mutation must be a de novo mutation because it was not found in either parents' genome DNA. To our knowledge, five previously described patients with PDHC deficiency have displayed the West syndrome. All six known patients, including our own, were female, even though an approximately equal number of males and females have been identified with PDHC deficiency and overall West syndrome occurs somewhat more frequently in males. These results indicated that West syndrome occurred more frequently in female patients with PDHC deficiency. It is suggested that lactate concentration should be measured in patients with West syndrome for potential PDHC deficiency, especially in females.

Amino Acid Substitution↗

PubMed1999

Hsp26: a temperature-regulated chaperone.

Small heat shock proteins (sHsps) are a conserved protein family, with members found in all organisms analysed so far. Several sHsps have been shown to exhibit chaperone activity and protect proteins from irreversible aggregation in vitro. Here we show that Hsp26, an sHsp from Saccharomyces cerevisiae, is a temperature-regulated molecular chaperone. Like other sHsps, Hsp26 forms large oligomeric complexes. At heat shock temperatures, however, the 24mer chaperone complex dissociates. Interestingly, chaperone assays performed at different temperatures show that the dissociation of the Hsp26 complex at heat shock temperatures is a prerequisite for efficient chaperone activity. Binding of non-native proteins to dissociated Hsp26 produces large globular assemblies with a structure that appears to be completely reorganized relative to the original Hsp26 oligomers. In this complex one monomer of substrate is bound per Hsp26 dimer. The temperature-dependent dissociation of the large storage form of Hsp26 into a smaller, active species and the subsequent re-association to a defined large chaperone-substrate complex represents a novel mechanism for the functional activation of a molecular chaperone.

Chromatography, High Pressure Liquid↗

PubMed1999

Identification and characterization of a cAMP-responsive element in the region upstream from promoter 1.3 of the human aromatase gene.

Aromatase converts androgens to estrogens. The expression of this enzyme is driven by multiple tissue-specific promoters which are differentially regulated. Aromatase expression in breast cancer and the surrounding adipose cells is directed mainly by promoters I.3 and II, while its expression in the normal breast adipose tissue is driven by promoter I.4. Like promoter II, promoters I.3 is thought to be a cAMP-driven promoter, demonstrated previously by cell culture experiments. In the present study, we have identified and characterized a cAMP-responsive element (CREaro) upstream from promoter 1.3. This positive element, TGAAGTCA, between -66 and -59 bp relative to the transcriptional start site of promoter 1.3 was identified by DNA deletion and mutation analyses. The sequence of CREaro is one base different from the consensus CRE sequence (CREpal; TGACGTCA), and the mutational analysis revealed that CREaro had a higher enhancer activity to promoter I.3 than CREpal. Nuclear proteins from both WS3TF breast tumor fibroblasts and SK-BR-3 breast cancer cells bound to this CREaro, as demonstrated by DNA mobility shift assay. The molecular weight of the major binding protein in fibroblasts was determined to be approximately 60 kDa, as shown by UV crosslinking, which is different from those of known CRE-binding proteins. It is thought that CREB1 is not expressed in tumor fibroblasts because the Western blot analysis using anti-CREB1 antibody was not able to detect CREB1 in the nuclear protein extract from these cells. DNA mobility shift analysis using a nuclear protein extract from SK-BR-3 cells revealed that at least two proteins bound to the CREaro and that one of these proteins was identified to be CREB1. These studies provide direct evidence that promoter I.3 is a cAMP-responsive promoter.

Aromatase↗

PubMed1999

Cloning of novel splice variants of mouse mGluR1.

Three splice variants of the mouse metabotropic glutamate receptor 1, mGluR1E55, mGluR1a and mGluR1b, have been isolated from mouse brain cDNA libraries. The sequences of mGluR1a and mGluR1b are similar to those from rat and human. mGluR1E55 is a novel splice variant. mGluR1E55 has two additional exons. One is 80-bp long at the 5' untranslational region. The other (E55) is 110-bp long at the cysteine-rich region after the ligand-binding domain and before the seven-transmembrane domain. Insertion of the E55 exon results in an inframe stop codon. The predicted protein product contains only the extracellular domain of the receptor and may be secreted.

Alternative Splicing↗

PubMed1999

Glucocorticoid receptor expression in the spinal cord after traumatic injury in adult rats.

Methylprednisolone (MP), a glucocorticoid, is the only effective therapeutic agent used in the clinical treatment of acute spinal cord injury (SCI). MP given within 8 hr after SCI significantly improves neurological function. Although the glucocorticoid receptor (GR) is suggested to mediate MP actions, limited knowledge is available on its expression and possible function after SCI. Presently, the expression of GR was studied in a weight-drop SCI model in adult rats. Immunohistochemistry and Western blot analysis revealed an increase in GR protein expression as early as 15 min after injury. GR expression sharply increased at 4 hr (22-fold), peaked at 8 hr (56-fold), rapidly declined at 1 d, and returned to the baseline level at and after 3 d. During its peak expression, GR was localized in neural somata and dendrites but not in axons and their terminals. GR immunoreactivity was also found in oligodendrocytes and astrocytes. Interestingly, other cell types, such as endothelial cells, were GR-negative. An increase in the binding activity of nuclear proteins to the glucocorticoid responsive element was also observed after SCI, demonstrating a functional element of GR activation. Finally, colocalization of GR and tumor necrosis factor alpha (TNF-alpha), an inflammatory cytokine, was observed in neurons and glial cells, consistent with MP regulation of TNF-alpha in this model. Thus, the transient expression of high levels of GR after SCI may provide new insights into the anti-inflammatory action of MP.

Animals↗

PubMed1999

Shape-selective separation of polycyclic aromatic hydrocarbons on protoporphyrin-silica phases. Effect of surface porphyrin distribution on column efficiency.

The chromatographic performance of various metalloprotoporphyrin-silica (MProP-silica) packing materials prepared using different porphyrin immobilization schemes is examined. Column efficiency and solute resolution for the shape-selective separation of polycyclic aromatic hydrocarbons (PAHs) can be improved significantly by preparing phases with lower porphyrin coverages and with a more homogeneous distribution of the porphyrin species on the surface. The latter is accomplished by spreading/diluting the number of aminopropyl reactive sites on the silica surface via mixing an inert methyltrimethoxysilane with 3-aminopropyltriethoxysilane during this preliminary reaction step. Subsequent covalent attachment of the ProP via amide bonds to the pendant amine sites results in a more even distribution of the porphyrins on the surface. Band shapes and retention times as a function of injected solute concentration as well as HPLC separation of various test mixtures of PAHs (including standard reference material SRM 869) are used to confirm the enhanced performance of these so-called "spread" phases. Changes in the nature of the immobilized porphyrin distribution on the silica surface are further probed by a coupled redox/UV-Vis absorbance method, and results suggest a decrease in the number of ProP species immobilized as aggregates on the surface.

Chromatography, High Pressure Liquid↗

PubMed1999

Three-dimensional measurement of cerebral microvascular plasma perfusion, glial fibrillary acidic protein and microtubule associated protein-2 immunoreactivity after embolic stroke in rats: a double fluorescent labeled laser-scanning confocal microscopic study.

Early astroglial response to post-ischemic microvascular hypoperfusion may contribute to progressive cerebral microcirculatory impairment and ischemic neuronal injury. Using laser-scanning confocal microscopy and three fluorescent probes, we measured in three-dimensions cerebral microvascular plasma perfusion, astrocytic reactivity, and neuronal injury assessed by fluorescein isothiocyanate (FITC)-dextran, GFAP immunoreactivity, and microtubule associated protein-2 (MAP2) immunoreactivity, respectively, in rats subjected to 2 h of middle cerebral artery occlusion. Three-dimensional quantitative analysis revealed that 2 h of embolic ischemia resulted in a significant (P<0.05) reduction of cerebral microvascular plasma perfusion in the ipsilateral cortex and subcortex. Tissue within the ipsilateral cortex and subcortex with low plasma perfusion exhibited a significant (P<0.05) increase in GFAP immunoreactivity compared with the homologous contralateral tissue. Three-dimensional re-constructed images showed that prominent GFAP immunoreactive astrocytes surrounded large vessels with decreased plasma perfusion in downstream capillaries in the ipsilateral MCA territory when compared to the vessels in the contralateral homologous tissue. Triple fluorescence probe-stained sections showed that tissue with decreased plasma perfusion and with increased GFAP immunoreactivity was accompanied by a reduction of MAP2 immunoreactivity. The present study demonstrates that an impairment of microvascular perfusion induces an early increase in GFAP immunoreactivity, and reactive astrocytes may contribute to a further reduction of cerebral microvascular plasma perfusion. The three-dimensional quantitative imaging analysis used in the present study provides a means to investigate parenchymal cellular responses to changes of cerebral microvascular plasma perfusion after MCA occlusion.

Animals↗

PubMed1999

Two organochlorine pesticides, toxaphene and chlordane, are antagonists for estrogen-related receptor alpha-1 orphan receptor.

Estrogen-related receptor (ERR) alpha-1 shares a high amino acid sequence homology with estrogen receptor alpha. Although estrogens are not ligands of ERR alpha-1, our recent results suggest that toxaphene and chlordane, two organochlorine pesticides with estrogen-like activity, behave as antagonists for this orphan nuclear receptor. The two compounds increased ERR alpha-1-mediated expression of the reporter enzyme beta-galactosidase in a yeast-based assay. The screen was developed by expressing the hERR alpha-1-yeast Gal 4 activation domain fusion protein in yeast cells carrying the beta-galactosidase reporter plasmid, which contains an ERR alpha-1-binding element. In transfection experiments using mammalian cell lines, such as the SK-BR-3 breast cancer cell line, the compounds were found to have an antagonist activity against ERR alpha-1-mediated expression of the reporter chloramphenicol acetyltransferase. In contrast to the findings with ERR alpha-1, the two compounds were found to slightly induce the estrogen receptor a-mediated expression of chloramphenicol acetyltransferase in SK-BR-3 cells. In a ligand-independent manner, the ERR alpha-1 activity in SK-BR-3 cells was induced 3-fold by cotransfection with the GRIP1 coactivator expression plasmid. Toxaphene was found to be capable of suppressing the GRIP1 coactivator-induced ERR alpha-1 activity in SK-BR-3 cells. In addition, a stable ERR alpha-1 expressing HepG2 hepatoma cell line was generated, and the aromatase activity in the transfected cell line was found to be twice that in the untransfected cell line. The enzyme aromatase converts androgens to estrogens, and aromatase expression in HepG2 cells is regulated in part by an ERR alpha-1-modulating promoter. A 24-h incubation of an ERR alpha-1-transfected HepG2 cell line with 10 microM toxaphene reduced its aromatase activity to the level in the untransfected cell line. Because toxaphene is not an inhibitor of aromatase, it is thought that the decrease of the aromatase activity in ERR alpha-1 transfected HepG2 cells following toxaphene treatment resulted from a suppression of the aromatase expression by toxaphene acting as the antagonist of ERR alpha-1. Toxaphene and chlordane are among the 12 persistent organic pollutants identified by the United Nations Environment Programme as requiring urgent attention. Their antagonistic effects on ERR alpha-1 should not be overlooked.

Breast Neoplasms↗

PubMed1999

Activities of constitutive promoters in Escherichia coli.

The in vivo activities of seven constitutive promoters in Escherichia coli have been determined as functions of growth rate in wild-type relA+ spoT+ strains with normal levels of guanosine tetraphosphate (ppGpp) and in ppGpp-deficient DeltarelADeltaspoT derivatives. The promoters include (i) the spc ribosomal protein operon promotor Pspc; (ii) the beta-lactamase gene promotor Pblaof plasmid pBR322; (iii) the PLpromoter of phage lambda; (iv) and (v) the replication control promoters PRNAIand PRNAIIof plasmid pBR322; and (vi) and (vii) the P1 and P2 promoters of the rrnB ribosomal RNA operon. Each strain carried an operon fusion consisting of one of the respective promoter regions linked to lacZ and recombined into the chromosome at the mal locus of a lac deletion strain. The amount of 5'-terminal lacZ mRNA and of beta-galactosidase activity expressed from these promoters were determined by standard hybridization or enzyme activity assays, respectively. In addition, DNA, RNA and protein measurements were used to obtain information about gene dosage, rRNA synthesis and translation rates. By combining lacZ mRNA hybridization data with gene dosage and rRNA synthesis data, the absolute activity of the different promoters, in transcripts/minute per promoter, was determined. In ppGpp-proficient (relA+ spoT+) strains, the respective activities of rrnB P1 and P2 increased 40 and fivefold with increasing growth rate between 0.7 and 3.0 doublings/hour. The activities of Pspc, PL, Pbla, and PRNAIincreased two- to threefold and reached a maximum at growth rates above 2.0 doublings/hour. In contrast, PRNAIIactivity decreased threefold over this range of growth rates. In ppGpp-deficient (DeltarelA DeltaspoT) bacterial strains, the activities of rrnB P1 and P2 promoters both increased about twofold between 1.6 and 3.0 doublings/hour, whereas the activities of Pspc, PL, Pbla, and PRNAI, and PRNAIIwere about constant. To explain these observations, we suggest that the cellular concentration of free RNA polymerase increases with increasing growth rate; for saturation the P1 and P2 rRNA promoters require a high RNA polymerase concentration that is approached only at the highest growth rates, whereas the other promoters are saturated at lower polymerase concentrations achieved at intermediate growth rates. In addition, the data indicate that the respective rrnB P1 and PRNAIIpromoters were under negative and positive control by ppGpp. This caused a reduced activity of rrnB P1 and an increased activity of PRNAIIduring slow growth in wild-type (relA+ spoT+) relative to ppGpp-deficient (DeltarelA DeltaspoT) bacterial strains.

Cell Division↗

PubMed1999

Tryptophan fluorescence monitors multiple conformational changes required for glutamine phosphoribosylpyrophosphate amidotransferase interdomain signaling and catalysis.

Single tryptophan residues were incorporated into each of three peptide segments that play key roles in the structural transition of ligand-free, inactive glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase to the active enzyme-substrate complex. Intrinsic tryptophan fluorescence and fluorescence quenching were used to monitor changes in a phosphoribosyltransferase (PRTase) "flexible loop", a "glutamine loop", and a C-terminal helix. Steady state fluorescence changes resulting from substrate binding were used to calculate binding constants and to detect the structural rearrangements that coordinate reactions at active sites for glutamine hydrolysis and PRTase catalysis. Pre-steady state kinetics of enzyme.PRPP and enzyme.PRPP.glutamine complex formation were determined from stopped-flow fluorescence measurements. The kinetics of the formation of the enzyme.PRPP complex were consistent with a model with two or more steps in which rapid equilibrium binding of PRPP is followed by a slow enzyme isomerization. This isomerization is ascribed to the closing of the PRTase flexible loop and is likely the rate-limiting step in the reaction of PRPP with NH(3). The pre-steady state kinetics for binding glutamine to the binary enzyme. PRPP complex could also be fit to a model involving rapid equilibrium binding of glutamine followed by an enzyme isomerization step. The changes monitored by fluorescence account for the interconversions between "end state" structures determined previously by X-ray crystallography and define an intermediate enzyme.PRPP conformer.

Amidophosphoribosyltransferase↗

PubMed1999

c-fos gene expression parallels auditory adaptation in the adult rat.

This study investigated the pattern of c-fos gene expression corresponding with auditory adaptation to novel sound. Using six groups of adult rats (naive control, 1 h, and 1, 2, 3, and 4 days of continuous stimulation), we quantified c-fos expressing cells in the dorsal and ventral cochlear nuclei and found a 54 fold increase in 1 h following novel sound stimuli. The number of reactive cells decreased sharply within 24 h and nearly disappeared by 96 h. Our results reveal that c-fos gene expression in the adult rat is attenuated in parallel with the expected auditory adaptation to novel sounds indicating an association with auditory learning and memory.

Acoustic Stimulation↗

PubMed1999

Characterization of transsynaptic tracing with central application of pseudorabies virus.

Although transsynaptic tracing with peripheral injection of pseudorabies virus (PRV) has been extensively characterized, several methodological issues related to central application of this tracer have not been addressed. In the present study, we addressed the following three issues by using microinjection of a cocktail containing PRV (Bartha strain) and cholera toxin subunit B (CTb) into different sites in the rat brain. First, we estimated PRV diffusion by examining injection sites at different times after application. Second, we tested whether PRV is taken up by fibers of passage following injections into the olivocerebellar pathway. Third, we developed criteria for leakage of PRV into cerebral ventricles. Our data indicate that (i) centrally injected PRV diffuses very little and produces focal injection sites; (ii) PRV is taken up and transported by fibers of passage, although less prominently than found for Ctb; (iii) PRV produces specific and easily identifiable ependymal cell as well as neuronal labeling following ventricular injection. This labeling can be used as a criterion for determining if labeling obtained was due to injected tracer leaking into brain ventricles. In summary, the present study provides new and important information about using PRV to trace central multisynaptic circuitry.

Animals↗