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Biomedical subjects

G Wang

Publications and source records attributed to G Wang.

At least 361 records · Page 20Linked to original sources
PubMed1999

Accelerated solvent extraction and gas chromatography/mass spectrometry for determination of polycyclic aromatic hydrocarbons in smoked food samples.

Accelerated solvent extraction (ASE) is a new sample extraction method offering a number of advantages such as low pe -extraction cost, reduced solvent and time consumption, and simplified extraction protocols. In this study, the ASE method was applied to the extraction of polycyclic aromatic hydrocarbons (PAHs) from biological samples. For recovery studies, fish tissues and ground pork were used as sample matrices. Sample aliquots fortified with 16 PAHs were extracted by ASE, and the extracts were treated with sulfuric acid and Florisil, followed by gas chromatography-mass spectrometry analysis. The PAH recoveries by the ASE method were found to be comparable with or better than those by Soxhlet extraction. The extraction and quantitation method was then applied to the determination of PAHs in several smoked meat samples obtained from a local market. Up to 12 PAHs were found to be present at concentrations ranging from 3 to 52 ng/g wet sample.

Food Handling↗
PubMed1999

Enhanced antitumor activity of sarCNU in comparison to BCNU in an extraneuronal monoamine transporter positive human glioma xenograft model.

A novel analogue of nitrosoureas, 2-chloroethyl-3-sarcosinamide-1-nitrosourea (SarCNU), has demonstrated increased anticancer effects in vitro and in vivo. Our previous work suggested that SarCNU enters cells via the extraneuronal monoamine transporter (EMT), that contributes to its enhanced cytotoxicity. In the present study, comparative activities of SarCNU and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) were evaluated in an EMT positive human glioma xenograft model. Athymic nude mice implanted subcutaneously or intracranially with human glioma SHG-44, a cell line that has been confirmed EMT positive by using reverse-transcription polymerase chain reaction (RT-PCR) assay, were treated with SarCNU at an optimal dose of 167 mg/kg, or BCNU at 20 mg/kg or 30 mg/kg, q4d x 3 intraperitoneally (i.p.). In 17 animals with subcutaneous tumor grafts treated with SarCNU, 9 animals became tumor free and 8 demonstrated tumor regression. While in the BCNU treated group, there were only 2 out of 10 mice in the 20 mg/kg group and 2 out of 7 in the 30 mg/kg group, which demonstrated some tumor regression. There were 4 drug related deaths in the BCNU (30 mg/kg) group, while there were no drug related deaths in the SarCNU group. In the intracranially implanted mice, the median survival time in the SarCNU group was more than 130 days, while in the BCNU treated group it was only 22 days which was similar to the control group (18 days). This is the first demonstration that SarCNU, in comparison to BCNU, has enhanced anticancer activity in an EMT positive human glioma xenograft model.

Animals↗
PubMed1999

Molecular genetic basis for the variable expression of Lewis Y antigen in Helicobacter pylori: analysis of the alpha (1,2) fucosyltransferase gene.

Helicobacter pylori lipopolysaccharides (LPS) express human oncofetal antigens Lewis X and Lewis Y. The synthesis of Lewis Y involves the actions of alpha (1,3) and alpha (1,2) fucosyltransferases (FucTs). Here, we report the molecular cloning and characterization of genes encoding H. pylori alpha (1,2) FucT (Hp fucT2) from various H. pylori strains. We constructed Hp fucT2 knock-out mutants and demonstrated the loss of Lewis Y production in these mutants by enzyme-linked immunosorbent assay (ELISA) and immunoelectron microscopy. The Hp fucT2 gene contains a hypermutable sequence [poly (C) and TAA repeats], which provides a possibility of frequent shifting into and out of coding frame by a polymerase slippage mechanism. Thus, the Hp fucT2 gene displays two major genotypes, consisting of either a single full-length open reading frame (ORF; as in the strain UA802) or truncated ORFs (as in the strain 26695). In vitro expression of Hp fucT2 genes demonstrated that both types of the gene have the potential to produce the full-length protein. The production of the full-length protein by the 26695 fucT2 gene could be attributed to translational-1 frameshifting, as a perfect translation frameshift cassette resembling that of the Escherichia coli dnaX gene is present. Examination of the strain UA1174 revealed that its fucT2 gene has a frameshifted ORF at the DNA level, which cannot be compensated by translation frameshifting, accounting for its Lewis Y off phenotype. In another strain, UA1218, the fucT2 gene is apparently turned off because of the loss of its promoter. Based on these data, we proposed a model for the variable expression of Lewis Y by H. pylori, in which regulation at the level of replication slippage (mutation), transcription and translation of the fucT2 gene may all be involved.

Amino Acid Sequence↗
PubMed1999

A nonNMDA antagonist, GYKI 52466 improves microscopic O2 balance in the cortex during focal cerebral ischemia.

This study was performed to test whether GYKI 52466, a nonNMDA receptor antagonist, would improve microregional oxygen supply and consumption balance in the focal cerebral ischemic area. Rats were anesthetized with 1.4% isoflurane. For the GYKI Group (n = 8), 10 min before middle cerebral artery (MCA) occlusion, a bolus of 5 mg kg-1 of GYKI 52466 i.v. was administered and was followed by an infusion of 5 mg kg-1 h-1. For the Control Group (n = 8), the same volume of the vehicle was administered. One hour after MCA occlusion, regional cerebral blood flow (rCBF) was measured using the 14C-iodoantipyrine autoradiographic technique. Microscopic arterial and venous oxygen saturations were determined using microspectrophotometry. In the cortex contralateral to MCA occlusion, the average rCBF and the average O2 consumption were lower in the GYKI Group than in the Control Group (rCBF: GYKI 65.5 +/- 24.1 ml 100 g-1 min-1, Control 97.7 +/- 33.4 ml 100 g-1 min-1; O2 consumption: GYKI 3.9 +/- 1.2 ml O2 100 g-1 min-1, Control 6.2 +/- 2.5 ml O2 100 g-1 min-1) without a significant difference in the number of veins with SvO2 < 50%. In the ischemic cortex, the number of veins with SvO2 < 50% was significantly smaller in the GYKI Group (21 veins out of 63) than in the Control Group (45 out of 59) without a significant difference in the average rCBF (GYKI 44.9 +/- 17.7, Control 29.7 +/- 10.4) or regional O2 consumption between these two groups (GYKI 3.3 +/- 1.4, Control 2.7 +/- 1.2). Our data demonstrated that GYKI 52466 was effective in improving microscopic O2 balance in the focal ischemic cortical area of the brain and it decreased O2 consumption in the non-ischemic cortex.

Animals↗
PubMed1999

p53 status, cellular recovery and cell cycle arrest as prognosticators of in vitro radiosensitivity in human pancreatic adenocarcinoma cell lines.

PURPOSE: To investigate the factors contributing to the in vitro radiosensitivity of four human pancreatic adenocarcinoma cell lines differing in p53 status, and the basis for the lack of post-irradiation G1 arrest in the two cell lines that have retained a wild-type p53 allele. MATERIALS AND METHODS: Cells were X-irradiated and the parameters related to radiosensitivity, as well as the modulation of gene products linked to regulation of cell cycle transit (p53, p21/WAF1/CIP1, pRb) or DNA replication and repair (DNA topoisomerase I and II), were determined. RESULTS: Both cell lines expressing either mutant (mt) R248W or R273H p53 proteins were more radioresistant. All the cell lines arrested in G2. None of the cell lines arrested in G1 and this was linked to the inability to upregulate p21/WAF1/CIP1. There were no correlations between p53 status and the magnitude or time of maximum G2 arrest. However, there was a negative correlation between a protracted arrest in G2 and the ability to recover from potentially lethal damage (PLD). CONCLUSIONS: Variation in radiosensitivity is related to p53 status, but the survival advantage conferred by having mutant p53 status is not readily explained neither by recovery from PLD nor by cell cycle arrest kinetics. There is no p53-independent pathway for the recruitment of p21 in these cell lines following irradiation.

Adenocarcinoma↗
PubMed1999

Polygalacturonase and polygalacturonase inhibitor protein: gene isolation and transcription in Glycine max-Heterodera glycines interactions.

The cell wall acts as the first line of defense during pathogen invasion. Polygalacturonases (PGs) are a class of cell-wall-modifying enzymes with precise temporal and organ-specific expression. A 350-bp fragment with high homology to PGs was identified by differential display (DD) analysis of soybean cyst nematode (SCN) race 3 resistant PI 437654 and susceptible cultivar Essex. The fragment was strongly expressed in Essex, 2 days after inoculation (DAI). Complete coding sequences of two PG cDNAs, PG1 and PG2, were isolated by 3' and 5' rapid amplification of cDNA ends polymerase chain reaction (RACE PCR). PI 437654 and Essex had identical PG1 and PG2 sequences. A transversion from A to C created a PstI restriction site in the PG2 cDNA that was used to distinguish the two PG cDNAs by cleaved amplified polymorphic sequence (CAPS) analysis. A cDNA encoding a polygalacturonase-inhibitor protein (PGIP) that is 89% identical to the Phaseolus vulgaris PGIP was isolated from soybean roots by reverse transcription (RT)-PCR. Steady-state levels of PG and PGIP were investigated by RNA gel blot analysis in roots 1 to 5 DAI and in hypocotyls and leaves. Differences in the constitutive levels of PG mRNAs were observed in roots of different soybean genotypes. Steady-state levels of PG mRNAs were enhanced during compatible interactions with SCN and reduced in incompatible interactions and in mechanically wounded roots. Enhanced PGIP transcription was observed in response to mechanical wounding in both PI 437654 and Essex, but only in compatible interactions with SCN, suggesting uncoupling of PGIP functions in developmental and stress cues. Constitutive expression in incompatible interactions shows PGIP is not a factor in SCN resistance. Thus, the up-regulation of endogenous PG transcription in soybean roots early after SCN infection could facilitate successful parasitism by SCN.

Amino Acid Sequence↗
PubMed1999

Comparative sequence analysis and predictions for the envelope glycoproteins of foamy viruses.

The foamy viruses (FVs) are a genus of complex retroviruses that has recently been found to possess several novel molecular features. There is increasing interest in the development of FVs as novel vectors for gene delivery. As there are remarkably few published studies of FV proteins, these recent findings prompted us to predict the structural features of FV glycoproteins with the aid of computer programs. We analysed all seven available FV Env sequences, a greater number of sequences than in previously published analyses. The relative rates of change for FV structural proteins were Pol < Env < Gag in increasing order, which differs from all other retroviruses. We determined that this difference is primarily caused by a higher relative rate of change for FV Gag proteins. We analysed the functional domains of FV glycoproteins and found that their structural organization was generally similar to other retroviruses. Putative structures were identified for the signal peptide, cleavage site, fusion peptide, membrane-spanning domain and the unique endoplasmic reticulum retrieval signal. Based on the predicted secondary structure of the transmembrane glycoprotein (TM) subunit, gp47, we also identified a unique prolonged central 'sheets and loops' region as the dominant feature of an unusually lengthy TM ectodomain. This lengthy central domain was flanked at each end by alpha-helices. The predictions reported here will stimulate and facilitate experimental approaches to better understand the structure and function of FV glycoproteins, and should assist in the planning and development of FV vectors.

Amino Acid Sequence↗
PubMed1999

The effect of pitch in multislice spiral/helical CT.

The purpose of this study is to understand the effect of pitch on raw data interpolation in multislice spiral/helical computed tomography (CT) and provide guidelines for scanner design and protocol optimization. Multislice spiral CT is mainly characterized by the three parameters: the number of detector arrays, the detector collimation, and the table increment per x-ray source rotation. The pitch in multislice spiral CT is defined as the ratio of the table increment over the detector collimation in this study. In parallel to the current framework for studying longitudinal image resolution, the central fan-beam rays of direct and opposite directions are considered, assuming a narrow cone-beam angle. Generally speaking, sampling in the Radon domain by the direct and opposite central rays is nonuniform along the longitudinal axis. Using a recently developed methodology for quantifying the sensitivity of signal reconstruction from nonuniformly sampled finite points, the effect of pitch on raw data interpolation is analyzed in multislice spiral CT. Unlike single-slice spiral CT, in which image quality decreases monotonically as the pitch increases, the sensitivity of raw data interpolation in multislice spiral CT increases in an alternating way as the pitch increases, suggesting that image quality does not decrease monotonically in this case. The most favorable pitch can be found from the sensitivity-pitch plot for any given set of multislice spiral CT parameters. An example for four-slice spiral CT is provided. The study on the effect of pitch using the sensitivity analysis approach reveals the fundamental characteristics of raw data interpolation in multislice spiral CT, and gives insights into interaction between pitch and image quality. These results may be valuable for design of multislice spiral CT scanners and imaging protocol optimization in clinical applications.

Image Processing, Computer-Assisted↗
PubMed1999

Multiplex sequence analysis demonstrates the competitive growth advantage of the A-to-G mutants of clarithromycin-resistant Helicobacter pylori.

Clarithromycin resistance in Helicobacter pylori is due to point mutation within the 23S rRNA. We examined the growth rates of different types of site-directed mutants and demonstrated quantitatively the competitive growth advantage of A-to-G mutants over other types of mutants by a multiplex sequencing assay. The results provide a rational explanation of why A-to-G mutants are predominantly observed among clarithromycin-resistant clinical isolates.

Alanine↗
PubMed1999

Molecular typing of Borrelia burgdorferi sensu lato: taxonomic, epidemiological, and clinical implications.

Borrelia burgdorferi sensu lato, the spirochete that causes human Lyme borreliosis (LB), is a genetically and phenotypically divergent species. In the past several years, various molecular approaches have been developed and used to determine the phenotypic and genetic heterogeneity within the LB-related spirochetes and their potential association with distinct clinical syndromes. These methods include serotyping, multilocus enzyme electrophoresis, DNA-DNA reassociation analysis, rRNA gene restriction analysis (ribotyping), pulsed-field gel electrophoresis, plasmid fingerprinting, randomly amplified polymorphic DNA fingerprinting analysis, species-specific PCR and PCR-based restriction fragment length polymorphism (RFLP) analysis, and sequence analysis of 16S rRNA and other conserved genes. On the basis of DNA-DNA reassociation analysis, 10 different Borrelia species have been described within the B. burgdorferi sensu lato complex: B. burgdorferi sensu stricto, Borrelia garinii, Borrelia afzelii, Borrelia japonica, Borrelia andersonii, Borrelia valaisiana, Borrelia lusitaniae, Borrelia tanukii, Borrelia turdi, and Borrelia bissettii sp. nov. To date, only B. burgdorferi sensu stricto, B. garinii, and B. afzelii are well known to be responsible for causing human disease. Different Borrelia species have been associated with distinct clinical manifestations of LB. In addition, Borrelia species are differentially distributed worldwide and may be maintained through different transmission cycles in nature. In this paper, the molecular methods used for typing of B. burgdorferi sensu lato are reviewed. The current taxonomic status of B. burgdorferi sensu lato and its epidemiological and clinical implications, especiallly correlation between the variable clinical presentations and the infecting Borrelia species, are discussed in detail.

Bacterial Typing Techniques↗
PubMed1999

Molecular cloning and characterization of rat genes encoding homologues of human beta-defensins.

beta-Defensins are cationic peptides with broad-spectrum antimicrobial activity that may play a role in mucosal defenses of several organs. They have been isolated in several species, and in humans, two beta-defensins have been identified. Here, we report the identification of two genes encoding beta-defensin homologues in the rat. Partial cDNAs were found by searching the expressed-sequence-tag database, and primers were designed to generate full-length mRNA coding sequences. One gene was highly similar to the human beta-defensin-1 (HBD-1) gene and mouse beta-defensin-1 gene at both the nucleic acid and amino acid levels and was termed rat beta-defensin-1 (RBD-1). The other gene, named RBD-2, was homologous to the HBD-2 and bovine tracheal antimicrobial peptide (TAP) genes. The predicted prepropeptides were strongly cationic, were 69 and 63 residues in length for RBD-1 and RBD-2, respectively, and contained the six-cysteine motif characteristic of beta-defensins. The beta-defensin genes mapped closely on rat chromosome 16 and were closely linked to the alpha-defensins genes, suggesting that they are part of a gene cluster, similar to the organization reported for humans. Northern blot analysis showed that both RBD-1 and RBD-2 mRNA transcripts were approximately 0.5 kb in length; RBD-1 mRNA was abundantly transcribed in the rat kidney, while RBD-2 was prevalent in the lung. Reverse transcription-PCR indicated that RBD-1 and RBD-2 mRNAs were distributed in a variety of other tissues. In the lung, RBD-1 mRNA expression localized to the tracheal epithelium while RBD-2 was expressed in alveolar type II cells. In conclusion, we characterized two novel beta-defensin homologues in the rat. The rat may be a useful model to investigate the function and contribution of beta-defensins to host defense in the lung, kidney, and other tissues.

Amino Acid Sequence↗
PubMed1999

Comparison of molecular methods for typing Vibrio parahaemolyticus.

An outbreak of Vibrio parahaemolyticus gastroenteritis on Canada's west coast in 1997 emphasized the need to develop molecular methods for differentiation and typing of these organisms. Isolates were analyzed by enterobacterial repetitive intergenic consensus sequence (ERIC) PCR, detection of restriction fragment length polymorphisms (RFLP) in rRNA genes (ribotyping), pulsed-field gel electrophoresis (PFGE), and RFLP analysis of the genetic locus encoding the polar flagellum (Fla locus RFLP analysis). ERIC PCR and ribotyping were the most informative typing methods, especially when used together, while Fla locus RFLP analysis was the least discriminatory. PFGE exhibited good discrimination but suffered from a high incidence of DNA degradation. ERIC PCR and ribotyping will be useful for the evaluation of genetic and epidemiological relationships among V. parahaemolyticus strains.

Bacterial Typing Techniques↗
PubMed1999

Phenotypic and genetic characterization of a novel Borrelia burgdorferi sensu lato isolate from a patient with lyme borreliosis.

Borrelia burgdorferi sensu lato A14S was cultured from a skin biopsy specimen of a patient with erythema migrans in The Netherlands. This isolate had a unique DNA fingerprint pattern compared to 135 other B. burgdorferi sensu lato isolates. In this study, the isolate A14S was further characterized by protein analysis with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and reactivity with various monoclonal antibodies. In addition, the 16S rRNA, ospA, and ospC genes, as well as the 5S-23S rRNA intergenic spacer DNA, were amplified by PCR, cloned, and sequenced. SDS-PAGE protein profiles and phylogenetic analysis based on all of the analyzed genes confirmed that B. burgdorferi sensu lato A14S was phenotypically and genetically different from the three human pathogenic species B. burgdorferi sensu stricto, Borrelia garinii, and Borrelia afzelii, as well as from other B. burgdorferi sensu lato species. Our findings indicate that Borrelia genomic groups or isolates other than the three well-known human pathogenic species may also cause human Lyme borreliosis.

Antigens, Bacterial↗
PubMed1999

An endoplasmic reticulum retrieval signal partitions human foamy virus maturation to intracytoplasmic membranes.

Among all retroviruses, foamy viruses (FVs) are unique in that they regularly mature at intracytoplasmic membranes. The envelope glycoprotein of FV encodes an endoplasmic reticulum (ER) retrieval signal, the dilysine motif (KKXX), that functions to localize the human FV (HFV) glycoprotein to the ER. This study analyzed the function of the dilysine motif in the context of infectious molecular clones of HFV that encoded mutations in the dilysine motif. Electron microscopy (EM) demonstrated virion budding both intracytoplasmically and at the plasma membrane for the wild-type and mutant viruses. Additionally, mutant viruses retained their infectivity, but viruses lacking the dilysine signal budded at the plasma membrane to a greater extent than did wild-type viruses. Interestingly, this relative increase in budding across the plasma membrane did not increase the overall release of viral particles into cell culture media as measured by protein levels in viral pellets or infectious virus titers. We conclude that the dilysine motif of HFV imposes a partial restriction on the site of viral maturation but is not necessary for viral infectivity.

Animals↗
PubMed1999

Functional domains of the Rsp5 ubiquitin-protein ligase.

RSP5, an essential gene of Saccharomyces cerevisiae, encodes a hect domain E3 ubiquitin-protein ligase. Hect E3 proteins have been proposed to consist of two broad functional domains: a conserved catalytic carboxyl-terminal domain of approximately 350 amino acids (the hect domain) and a large, nonconserved amino-terminal domain containing determinants of substrate specificity. We report here the mapping of the minimal region of Rsp5 necessary for its essential in vivo function, the minimal region necessary to stably interact with a substrate of Rsp5 (Rpb1, the large subunit of RNA polymerase II), and the finding that the hect domain, by itself, is sufficient for formation of the ubiquitin-thioester intermediate. Mutations within the hect domain that affect either the ability to form a ubiquitin-thioester or to catalyze substrate ubiquitination abrogate in vivo function, strongly suggesting that the ubiquitin-protein ligase activity of Rsp5 is intrinsically linked to its essential function. The amino-terminal region of Rsp5 contains three WW domains and a C2 calcium-binding domain. Two of the three WW domains are required for the essential in vivo function, while the C2 domain is not, and requirements for Rpb1 binding and ubiquitination lie within the region required for in vivo function. Together, these results support the two-domain model for hect E3 function and indicate that the WW domains play a role in the recognition of at least some of the substrates of Rsp5, including those related to its essential function. In addition, we show that haploid yeast strains bearing complete disruptions of either of two other hect E3 genes of yeast, designated HUL4 (YJR036C) and HUL5 (YGL141W), are viable.

Alleles↗