Biomedical subjects

Z Zhang

Publications and source records attributed to Z Zhang.

At least 217 records · Page 12Linked to original sources

PubMed2001

[Frequent somatic mutations of the von Hippel-Lindau tumor suppressor gene in primary sporadic human renal clear cell carcinomas].

OBJECTIVE: To investigate the mutation of Von Hippel-Lndau (VHL) tumor suppressor gene in patients with primary sporadic human renal cell carcinoma (RCC). METHODS: DNA samples from 20 primary sporadic renal clear carcinoma patients were analyzed by polymerase chain reaction, single strand conformational polymorphism analyses (PCR-SSCP) and direct sequencing. RESULTS: Somatic mutations of the VHL gene were detected in 11 (55%) of the 20 clear cell renal carcinomas, including 6 deletions, 2 insertions, and 3 missense mutations. These mutations mainly occurred in the last one-third region of exon 1, 2 and 3 of the VHL gene. CONCLUSION: The VHL gene may frequently mutate in Chinese patients with primary sporadic renal clear cell carcinoma. The VHL gene may be useful as a marker gene for the diagnosis of RCC and as a target gene for molecular therapy.

Adult↗

PubMed2001

The role of the M6-M7 loop (L67) in stabilization of the phosphorylation and Ca(2+) binding domains of the sarcoplasmic reticulum Ca(2+)-ATPase (SERCA).

The amino acid sequence (L67) intervening between the M6 and M7 transmembrane segments of the Ca(2+) transport ATPase was subjected to mutational analysis. Mutation of Pro(820) to Ala interferes with protein expression even though transcription occurs at normal levels. Single mutations of Lys(819) or Arg(822) to Ala, Phe, or Glu allow good expression, but produce strong inhibition of ATPase activity. The main defect produced by these mutations is strong interference with enzyme phosphorylation by ATP in the presence of Ca(2+), and also by P(i) in the absence of Ca(2+). The Lys(819) and Arg(822) mutants undergo slight and moderate reduction of Ca(2+) binding affinity, respectively. Reduction of overall steady state ATPase velocity is then due to inhibition of phosphorylated intermediate formation. On the other hand, a cluster of conservative mutations of Asp(813), Asp(815), and Asp(818) to Asn interferes strongly with enzyme activation by Ca(2+) binding and formation of phosphorylated enzyme intermediate by utilization of ATP. Enzyme phosphorylation by Pi in the absence of Ca(2+) undergoes slight or no inhibition by the triple aspartate mutation. Therefore, the triple mutation interferes mainly with the calcium-dependent activation of the ATPase. The effect of the triple mutation can be to a large extent reproduced by single mutation of Asp(813) (but not of Asp(815) or Asp(818)) to Asn. Functional and structural analysis of the experimental data demonstrates that the L67 loop plays an important role in protein folding and function. This role is sustained by linking the cytosolic catalytic domain and the transmembrane Ca(2+) binding domain through a network of hydrogen bonds.

Animals↗

PubMed2001

Opposite effect of NF-kappa B and c-Jun N-terminal kinase on p53-independent GADD45 induction by arsenite.

Cell cycle checkpoint, a major genomic surveillance mechanism, is an important step in maintaining genomic stability and integrity in response to environmental stresses. Using cells derived from human bronchial epithelial cells, we demonstrate that NF-kappaB and c-Jun N-terminal kinase (JNK) reciprocally regulate arsenic trioxide (arsenite)-induced, p53-independent expression of GADD45 protein, a cell cycle checkpoint protein that arrests cells at the G(2)/M phase transition. Inhibition of NF-kappaB activation by stable expression of a kinase-mutated form of IkappaB kinase caused increased and prolonged induction of GADD45 by arsenite. In contrast, the induction of GADD45 by arsenite was transient and less potent in cells where the NF-kappaB activation pathway was normal. Analysis of the cell cycle profile by flow cytometry indicated that NF-kappaB inhibition potentiates arsenite-induced G(2)/M cell cycle arrest. Abrogation of JNK activation, on the other hand, decreased GADD45 expression induced by arsenite, suggesting a role for JNK activation in GADD45 induction. These results indicate a molecular mechanism by which NF-kappaB and JNK may differentially contribute to cell cycle regulation in response to arsenite.

Arsenites↗

PubMed2001

Wnt-1 signaling inhibits apoptosis by activating beta-catenin/T cell factor-mediated transcription.

Wnt signaling plays a critical role in development and oncogenesis. Although significant progress has been made in understanding the downstream signaling cascade of Wnt signaling, little is known regarding Wnt signaling modification of the cell death machinery. Given that numerous oncogenes transform cells by providing cell survival function, we hypothesized that Wnt signaling may inhibit apoptosis. Here, we report that cells expressing Wnt-1 were resistant to cancer therapy-mediated apoptosis. Wnt-1 signaling inhibited the cytochrome c release and the subsequent caspase-9 activation induced by chemotherapeutic drugs, including both vincristine and vinblastine. Furthermore, we found that Wnt-1-mediated cell survival was dependent on the activation of beta-catenin/T cell factor (Tcf) transcription. Inhibition of beta-catenin/Tcf transcription by expression of the dominant-negative mutant of Tcf-4 blocked Wnt-1-mediated cell survival and rendered cells sensitive to apoptotic stimuli. These results provide the first demonstration that Wnt-1 inhibits cancer therapy-mediated apoptosis and suggests that Wnt-1 may exhibit its oncogenic potential through a mechanism of anti-apoptosis.

Animals↗

PubMed2001

A new approach to predict the helix/strand content of globular proteins.

An improved multiple linear regression method has been proposed to predict the content of alpha-helix and beta-strand of a globular protein based on its primary sequence and structural class. The amino acid composition and the auto-correlation functions derived from the hydrophobicity profile of the primary sequence have been taken into account. However, only the compositions of a part of the amino acids and a part of the auto-correlation functions are selected as the regression terms, which lead to the least prediction error. The resubstitution test shows that the average absolute errors are 0.052 and 0.047 with the standard deviations 0.050 and 0.047 for the prediction of helix/strand content, respectively. A rigorous cross-validation test, the jackknife test shows that the average absolute errors are 0.058 and 0.053 with the standard deviations 0.057 and 0.053 for the prediction of helix/strand content, respectively. Both tests indicate the self-consistency and the extrapolating effectiveness of the new method. The high prediction accuracy means that the method is suitable for practical applications.

Amino Acid Sequence↗

PubMed2001

Study of Surface-Enhanced Infrared Spectroscopy.

The dependence of surface-enhanced infrared absorption (SEIRA) on structure and vibration modes of chemisorbates was studied in a reflection-absorption mode by employing self-assembled monolayers of 4-pyridinethiol and 4-nitrothiophenol formed on Au island films deposited on glass substrates. The present work shows that both the electromagnetic and chemical effects contribute to the total SEIRA enhancement, with the former playing a predominant role. Moreover, the bands due to different vibration modes give rise to distinctly different magnitudes of enhancement, suggesting the presence of a chemical origin in the enhanced IR absorption. Copyright 2001 Academic Press.

Journal Article↗

PubMed2001

Study of Surface-Enhanced Infrared Spectroscopy.

In this paper we report a versatile and effective strategy to attain strong surface-enhanced infrared absorption by employing a sandwich system consisting of metal island films and self-assembled monolayers (SAMs) of 4-pyridinethiol. The observed larger enhancement factor stems from coupling of the electric fields induced by excitation of the surface plasmon resonance of the overlayer and underlayer Au island films, and from enhanced chemical interactions of the Au island films and the pyridine molecule in the sandwiched structures, compared to the corresponding SAM-Au configuration. Copyright 2001 Academic Press.

Journal Article↗

PubMed2001

In vitro biocompatibility study of electrically conductive polypyrrole-coated polyester fabrics.

This study investigated the basic biocompatibility aspects of two types of polypyrrole (PPy)-coated polyester fabrics for possible use as vascular prostheses. These PPy-coated fabrics, PPy-Phos and PPy-Plas, were sterilized with ethylene oxide (EO) and the following characterizations were performed: surface morphology by scanning electron microscope, EO residuals analysis by the headspace method, acute systemic toxicity in the mouse model, hemolysis, blood coagulation time, viability and proliferation of endothelial cells measured with the WST-1 method, and activation of polymorphonuclear (PMN) cells indicated by the specific expression of interleukin 8 mRNA measured by reverse transcription polymerase chain reaction. Virgin polyester fabrics, expanded poly(tetrafluoroethylene) (ePTFE), and medical-grade Bionate 80A poly(carbonate urethane) were used as references in the cell culture experiments. The PPy-coated fabrics revealed different surface morphologies by showing more PPy lamina and clusters on the PPy-Plas. Neither of the PPy-coated fabrics had an adverse effect on hemolysis and coagulation time, and they did not cause any acute systemic toxicity. The EO residual level was as low as 5 ppm or less, which is considered quite acceptable. Although exhibiting a relatively low initial cell adhesion at 24 h, the two PPy-coated samples showed no cytotoxicity at 72 and 168 h. Bionate 80A and ePTFE recorded cytotoxicity at 72 and 168 h, respectively. The virgin fabrics also demonstrated a decrease of viable cells at 72 h that was not significant. The activation of PMN cells induced by both PPy-coated fabrics, the ePTFE, and the negative control was significantly lower than that induced by their respective tumor necrosis factor-alpha controls. These results therefore highlighted the potential of PPy-coated fabrics for use as cardiovascular prostheses. It was suggested that cell adhesion moieties should be incorporated into the PPy/fabric composite to increase cell adhesion and subsequent cell proliferation.

Animals↗

PubMed2001

Manipulation of protein fingerprints during on-column fluorescent labeling: protein fingerprinting of six Staphylococcus species by capillary electrophoresis.

Bacterial proteomes were analyzed by use of electrophoretically mediated microanalysis (EMMA) and field-enhanced stacking. A water-soluble protein fraction was injected onto a capillary. Next, a fluorogenic reagent was injected and allowed to react with the protein mixture, producing fluorescent products that were separated by submicellar capillary electrophoresis and detected by laser-induced fluorescence. By use of a low-ionic strength sample buffer and a brief electrophoretic step, slow moving anionic proteins were stacked at the reagent-sample interface and were preferentially labeled. By reversing the order of sample injection and labeling reagent, fast moving cationic proteins were preferentially labeled. By adjustment of the sample buffer pH, proteins with different isoelectric points were selectively labeled. Electrophoresis fingerprints were generated for the water-soluble protein fraction from six Staphylococcus species. The protein patterns produced were species-specific and were used to construct a phylogenetic tree.

Bacterial Proteins↗

PubMed2001

Decreased Ras-mitogen-activated protein kinase signaling may cause DNA hypomethylation in T lymphocytes from lupus patients.

OBJECTIVE: Previous studies have shown that inhibiting T cell DNA methylation causes a lupus-like disease by modifying gene expression. T cells from patients with lupus exhibit diminished levels of DNA methyltransferase (MTase) enzyme activity, hypomethylated DNA, and changes in gene expression similar to those exhibited by T cells treated with methylation inhibitors, suggesting that DNA hypomethylation may contribute to human lupus. Since it is known that DNA MTase levels are regulated by the ras-mitogen-activated protein kinase (MAPK) pathway, this study sought to determine whether decreased ras-MAPK signaling could account for the DNA hypomethylation in lupus T cells. METHODS: DNA MTase messenger RNA (mRNA) from lupus patients and from healthy controls was quantitated by Northern analysis, and ras-MAPK signaling was determined by immunoblotting with antibodies to the activated forms of extracellular receptor-associated kinase (ERK). Results were compared with those in T cells in which ras-MAPK signaling was inhibited with a soluble inhibitor of MAPK ERK I (MEK1). RESULTS: T cells from patients with active lupus had diminished DNA MTase mRNA levels and decreased signaling through the ras-MAPK pathway. Inhibiting signaling through the ras-MAPK pathway with the MEK1 inhibitor decreased DNA MTase mRNA and enzyme activity to the levels seen in lupus T cells, and resulted in DNA hypomethylation resembling that seen in lupus T cells. CONCLUSION: These results suggest that a decrease in signaling through the ras-MAPK pathway may be responsible for the decreased MTase activity and DNA hypomethylation in patients with lupus.

Adolescent↗

PubMed2001

Activation of a fibroblast-specific enhancer of the proalpha2(I) collagen gene in tight-skin mice.

OBJECTIVE: Reporter transgenes were introduced into the type 1 tight-skin (Tsk1/+) mouse model of scleroderma to test the hypothesis that fibroblast-specific genetic programs are activated in fibrosis. METHODS: Transgenes harboring upstream fragments of the 5' flanking region of the mouse proalpha2(I) collagen gene (Col1a2), linked to a 400-bp minimal Col1a2 promoter driving an Escherichia coli beta-galactosidase (LacZ) reporter gene, were introduced into Tsk1/+ mice by breeding. Expression of these transgenes, which function as lineage-specific markers of fibroblast differentiation, was compared between the Tsk-LacZ mice and non-Tsk littermates. Responsiveness of these constructs to the profibrotic cytokine, transforming growth factor beta1 (TGFbeta1), was investigated by transient transfection of reporter constructs in tissue-culture cells. RESULTS: There was significant activation of reporter genes harboring the upstream enhancer in Tsk1/+ mice starting from 1 week of age. This was maximal at 6 weeks old (mean +/- SD 237 +/- 24% of non-Tsk controls; P= 0.001). Recombinant TGFbeta1 significantly activated reporter genes regulated by the upstream enhancer in transient transfection, and Tsk-LacZ fibroblasts showed elevated LacZ expression in tissue culture. CONCLUSION: These data suggest that activating signals in Tsk1/+ mice may act via fibroblast-specific regulatory elements within the murine Col1a2 gene. Although TGFbeta has been implicated in the pathogenesis of fibrosis, and reporter genes regulated by the upstream enhancer appear to be TGFbeta responsive in vitro, our results suggest that fibroblast-specific pathways may also be involved.

Animals↗

PubMed2001

Expression of recombination-activating genes and terminal deoxynucleotidyl transferase and secondary rearrangement of immunoglobulin kappa light chains in rheumatoid arthritis synovial tissue.

OBJECTIVE: Lymphocytic infiltrates in rheumatoid arthritis (RA) synovium often resemble lymphoid follicles and contain clonally related Ig transcripts, suggesting in situ antigen-dependent B cell selection. Recent reports have shown expression of recombination-activating genes (RAGs) and concurrent secondary rearrangement of Ig genes in normal peripheral lymphoid organs (receptor revision). We sought to determine if RAG-mediated receptor revision of Ig kappa light chains occurs in B cells within the RA synovium. Because we previously reported enhanced N-region addition at V(L)-J(L) joins in clonally expanded light-chain transcripts from RA synovium, we also sought expression of terminal deoxynucleotidyl transferase (TdT), which is normally expressed only in B cell precursors or immature B cells. METHODS: Reverse transcription-polymerase chain reaction (PCR) was used to detect RAG and TdT transcripts from unselected and B cell-enriched synovial and peripheral blood mononuclear cells obtained from 12 RA patients. Activity of RAG protein was sought using ligation-mediated PCR to detect recombination intermediates, and immunohistochemistry was performed to identify RAG+ cells within synovia. RESULTS: We found evidence of RAG-mediated secondary Ig kappa light chain rearrangements in about one-third of RA synovia. TdT expression was found in several samples, but did not correlate with RAG expression. CONCLUSION: RAG-mediated secondary Ig rearrangements of kappa light chains may contribute to the local production of antibodies to autoantigens (e.g., rheumatoid factor) or exogenous antigens, or it may represent a failed attempt at immune tolerance. TdT expression suggests the presence of immature B cells in RA synovia. These findings have important implications for the local generation of antibodies in RA and other chronic inflammatory diseases.

Adult↗

PubMed2001

Multiple separations facilitate identification of protein variants by mass spectrometry.

Identification of variant proteins from complex biological samples promises to contribute much to our understanding the etiology of pathological states. Characterization of variants, either due to genetic mutations in protein sequences or to post-translational modifications, is considerably more difficult than the simple protein identifications typical of most current proteomic investigations. Identification of a few peptides by database retrieval is not adequate when the goal is to have a complete understanding of the modifications of the protein. Although one advantage of mass spectrometry is its ability to obtain specific responses to several components, the complexity of biological samples is often overwhelming, resulting in spectra lacking useful information. For complex mixtures, isolation procedures before mass spectrometric analysis may need to include a variety of chromatographic and electrophoretic separation techniques. In this report, we illustrate how several preparative steps were essential for obtaining information about modified human lens beta-crystallins. The preparative techniques prior to mass spectrometry included size exclusion chromatography, reversed phase chromatography, two-dimensional gel electrophoresis, in situ digestion of the proteins and peptide trapping and washing before a final reversed phase high performance liquid chromatographic separation on-line to the mass spectrometer. This approach for isolation and analysis, when customized for other proteins, should find application in many studies where protein variants of complex mixtures are to be identified.

Chromatography, Gel↗

PubMed2001

A nitric oxide donor induces neurogenesis and reduces functional deficits after stroke in rats.

The adult rodent brain is capable of generating neuronal progenitor cells in the subventricular zone, and in the dentate gyrus of the hippocampus, throughout the life of the animal. Signals that regulate progenitor cell proliferation, differentiation, and migration are not well known. We report that administration of a nitric oxide donor, (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl) aminio]diazen-1-ium-1,2-diolate (DETA/NONOate), to young adult rats significantly increases cell proliferation and migration in the subventricular zone and the dentate gyrus. Treatment with DETA/ NONOate also increases neurogenesis in the dentate gyrus. Furthermore, administration of DETA/NONOate to rats subjected to embolic middle cerebral artery occlusion significantly increases cell proliferation and migration in the subventricular zone and the dentate gyrus, and these rats exhibit significant improvements of neurological outcome during recovery from ischemic stroke. Administration of DETA/NONOate significantly increases cortical levels of guanosine monophosphate both in ischemic and nonischemic rats, supporting the role of nitric oxide in promoting cell proliferation and neurogenesis. Thus, our data indicate that nitric oxide is involved in the regulation of progenitor cells and neurogenesis in the adult brain. This suggests that nitric oxide delivered to the brain well after stroke may have therapeutic benefits.

Animals↗

PubMed2001

The Acute toxicity of alpha-branched phenylsulfonyl acetates in Photobacterium phosphoreum test.

The acute toxicity (15-min EC(50)) of 20 alpha-substituted phenylsulfonyl acetates was measured using the Microtox test with Photobacterium phosphoreum. On the basis of TLSER, quantitative structure-activity relationships (QSARs) were calculated for the inhibition of bioluminescence. With the parameters molecular volume (V(mc)), dipolarity/polarizability (pi*), and the covalent contribution to Lewis basicity (epsilon(b)), a squared correlation coefficient (r(2)(adj)) of 0.868 and standard error of 0.094 for log EC(50) were obtained. By introducing a structural indicator variable (I), the regressive quality was improved (correlation coefficient of 0.948 and standard error of 0.059); the descriptors V(mc), pi*, and the electrostatic basicity contribution (q(-)) were significant. -NO(2)-- and -SO(2)--groups are active centers of these compounds. The strong hydrogen bonding effects between the compound active centers and FMNH(2) play an important role in the greater toxicity of these chemicals.

Luminescent Measurements↗

PubMed2001

Resistance of human betaB2-crystallin to in vivo modification.

Post-translational modifications and/or structural changes induced by modifications are likely causes of the decrease in crystallin solubility associated with aging and the development of cataract. Characterization of human lens crystallins by mass spectrometry has demonstrated that betaB2-crystallin undergoes less modification than any of the other crystallins. As the lens ages, betaB2-crystallin retains its hydrophilic N-terminus while the hydrophilic C-termini of alpha-crystallins and large portions of the N-termini of betaA3/A1 and betaB1 are truncated. The hydrophilic terminal regions of crystallins contribute to their solubility. Furthermore, deamidation and disulfide bond formation, other modifications that may affect solubility by altering conformation, are less extensive in betaB2 than in the other crystallins. This resistance to modification results in higher levels of betaB2 compared with the other crystallins in the water-soluble fraction of older lenses. The solubility of betaB2 and its propensity to form non-covalent associations with less soluble beta-crystallins may contribute to the solubility of the other beta-crystallins. A current hypothesis is that the chaperone-like properties of alpha-crystallins contribute to lens crystallin solubility, particularly in younger lenses. In older lenses, where most of the alpha-crystallins have become water-insoluble, betaB2-crystallins may play a dominant role in lens crystallin solubility.

Adolescent↗

PubMed2001

Semi-automated icosahedral particle reconstruction at sub-nanometer resolution.

Electron cryomicroscopy of large macromolecular complexes is becoming an increasingly powerful tool for revealing three-dimensional structures without the need for crystallization. The execution of image processing, however, requires experience and is error-prone due to the need for a human operator to carry out interactive and repetitive processes. We have designed an approach which is intended to make image processing simple and rapid, both for experts and for novice users. We demonstrate this approach using the well-established reconstruction scheme for icosahedral particles. Finally, we implement semi-automated virus reconstruction (SAVR), an expert system that integrates the most CPU intensive and iterative steps using the scripting language Python. SAVR is portable across platforms and has been parallelized to run on both shared and distributed memory platforms. SAVR also allows the incorporation of new algorithms and facilitates the management of the increasingly large data sets needed to achieve higher resolution reconstructions. The package has been successfully applied to several data sets and shown capable of generating icosahedral reconstructions to sub-nanometer resolutions (7-10 A ).

Automation↗

PubMed2001

Pharmacological MRI mapping of age-associated changes in basal ganglia circuitry of awake rhesus monkeys.

While the pathophysiological changes induced by the loss of dopamine innervation in the basal ganglia by Parkinson's disease (PD) are well studied, little is known about functional changes in the neural circuitry of this area during normal aging. Here we report the first survey of age-associated changes in the basal ganglia of behaviorally characterized, awake rhesus monkeys, using pharmacological MRI to map responses to dopaminergic stimulation. Apomorphine, a mixed D(1)/D(2) dopamine receptor agonist, evoked little change in the substantia nigra (SN) of aged animals while significantly reducing activation in young adult monkeys. Compared to young animals, both apomorphine and d-amphetamine (which increases synaptic dopamine levels) significantly increased activation of the aged rhesus globus pallidus externa (GPe). In addition, the aged animals showed decreased activity in the putamen in response to d-amphetamine administration. Although the responses in the SN and putamen of the aged monkeys differed from those in animal models of PD, the apomorphine-evoked activation of their GPe corresponded with apomorphine-induced increases in neuronal activity seen in Parkinson's patients and animal models. Given the major role of the GPe in regulating motor behavior, the altered responses in the aged GPe may contribute significantly to the motor slowing and movement dysfunctions characterizing advanced age.

Aging↗