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H Wang

Publications and source records attributed to H Wang.

At least 181 records · Page 10Linked to original sources
PubMed2004

Statistical methods for in situ hybridization: identification of autoradiographically labelled cells and structures.

In situ hybridization experiments frequently use autoradiography to identify labelled structures. Ideally, labelled cells will be overlain with a dense accumulation of particles, allowing one to discriminate them from unlabelled cells easily. However, if noise is high or the density of labelling is low, it can be difficult to distinguish bona fide labelling 'by eye'. In such situations, labelled cells could be overlooked. This paper evaluates two statistical solutions to this problem: (1) a parametric method proposed by Hashimoto and co-workers and (2) Wang & Wessendorf's non-parametric method using contingency testing (i.e. the chi-square or Fisher's exact tests). The Hashimoto method determines the mean and standard deviation of the density of background labelling, using sense-strand controls as the source of background levels. Cells labelled at densities greater than two standard deviations above the mean (P < 0.0455) are defined as significantly labelled. Contingency testing determines whether the grain density over a cell is significantly higher than that over the remainder of the image. When compared, the two methods gave similar results. The Hashimoto method may be more sensitive if most cells are labelled but contingency testing requires no assumptions about the uniformity of non-specific labelling.

Animals↗
PubMed2004

Identification of mariner elements from house flies (Musca domestica) and German cockroaches (Blattella germanica).

Full-length mariner elements were isolated and sequenced from house flies (Musca domestica) and German cockroaches (Blattella germanica). The amino acid sequence of the house fly mariner element (accession number: AF373028) showed 99.5% identity with Mos1 and peach elements, whereas the German cockroach mariner element (accession number: AF355143) showed 98.8% and 99.8% identity, respectively. Sequence analysis revealed that the mariner elements in house flies and German cockroaches differed from the active Mos1 mariner element by seven and 15 nucleotides, respectively. Four essential nucleotide substitutions at positions 64, 154, 305, and 1203, which have been proposed to contribute to the loss of activity of the inactive elements, were detected in the German cockroach mariner element. In contrast, although the mariner element in house flies contained substitutions at positions 64, 154, and 305, it retained T at position 1203, identical to active mariner elements. Mariner is present in approximately eight copies in the German cockroach genome.

Amino Acid Sequence↗
PubMed2004

Extracellular role of HMGB1 in inflammation and sepsis.

High mobility group box 1 (HMGB1), a 30 kDa nuclear and cytosolic protein widely studied as a transcription factor and growth factor, has recently been identified as a cytokine mediator of lethal systemic inflammation (e.g. endotoxaemia and sepsis), arthritis and local inflammation. It is released by activated macrophages, and serum levels increase significantly during endotoxaemia, sepsis and arthritis with significant delayed kinetics in comparison with tumour necrosis factor (TNF) and interleukin-1beta. Recently identified biological activities of HMGB1 include activation of macrophages/monocytes to release proinflammatory cytokines, upregulation of endothelial adhesion molecules, stimulation of epithelial cell barrier failure, and mediation of fever and anorexia. Passive immunization with anti-HMGB1 antibodies confers significant protection against lethal endotoxaemia, sepsis, arthritis and lipopolysaccharide-induced acute lung injury, even when antibody administration is delayed until after the early TNF responses have resolved. Strategies to inhibit HMGB1 activity and release are being investigated in these and other preclinical models of acute and chronic inflammation.

Amino Acid Sequence↗
PubMed2004

Terminal protein-induced stretching of bacteriophage phi29 DNA.

Stretching of DNA molecules helps to resolve detail during the fluorescence microscopy of both single DNA molecules and single DNA-protein complexes. To make stretching occur, intricate procedures of specimen preparation and manipulation have been developed in previous studies. By contrast, the present study demonstrates that conventional procedures of specimen preparation cause DNA stretching to occur, if the specimen is the double-stranded DNA genome of bacteriophage phi29. Necessary for this stretching is a protein covalently bound at both 5' termini of phi29 DNA molecules. Some DNA molecules are attached to a cover glass only at the two ends. Others are attached at one end only with the other end free in solution. The extent of stretching varies from approximately 50% overstretched to approximately 50% understretched. The understretched DNA molecules are internally mobile to a variable extent. In addition to stretching, some phi29 DNA molecules also undergo assembly to form both linear and branched concatemers observed by single-molecule fluorescence microscopy. The assembly also requires the terminal protein. The stretched DNA molecules are potentially useful for observing DNA biochemistry at the single molecule level.

Bacillus Phages↗
PubMed2004

Fluorescence microscopy of colour-tagged nanoparticles that are undergoing thermal motion.

To bypass limitations of conventional biochemical analysis, single-particle biochemical analysis is used. To improve single-particle biochemical analysis, procedures are needed to keep nanometre-sized particles in focus while the particles are undergoing thermal motion. A simple, inexpensive procedure is developed here for keeping particles in focus during the continuous observing/discriminating/recording of two different particles, both of which are undergoing thermal motion. This procedure concentrates the particles in a plane of solution that is in focus when the cover glass surface is in focus. An essential component of the procedure is the addition of molten, low-melt agarose to the specimen. Motionless binding to glass is inhibited by inclusion of anti-stick additives in the specimen. Both carrier protein (gelatin) and non-ionic detergent (Triton X-100) are anti-stick additives successfully used here. Intact bacteriophages T3 and T7 are used as model particles, in anticipation of the use of the procedures developed here for the analysis of the assembly of bacteriophages. Observing/discriminating/recording of colour-tagged bacteriophages T3 and T7 is achieved at video frame rate with image splitting to discriminate colours.

Acetic Acid↗
PubMed2004

Selective targeting of a laccase from Stachybotrys chartarum covalently linked to a carotenoid-binding peptide.

Atwo-step targeting strategy was used to identify improved laccases for bleaching carotenoid-containing stains on fabric. We first applied a modified phage display technique to identify peptide sequences capable of binding specifically to carotenoid stains and not to fabric. Prior deselection on the support on which the carotenoid was localized, increased stringency during the biopanning target selection process, and analysis of the phage peptides' binding to the target after acid elution and polymerase chain reaction (PCR) postacid elution, were used to isolate phage peptide libraries with increased binding selectivity and affinity. Peptide sequences were selected based on identified consensus motifs. We verified the enhanced carotenoid-binding properties of the peptide YGYLPSR and subsequently cloned and expressed C-terminal variants of laccase from Stachybotrys chartarum containing carotenoid-binding peptides YGYLPSR, IERSAPATAPPP, KASAPAL, CKASAPALC, and SLLNATK. These targeted peptide-laccase fusions demonstrate enhanced catalytic properties on stained fabrics.

Carotenoids↗
PubMed2004

Procoagulant surface exposure and apoptosis in rabbit platelets: association with shortened survival and steady-state senescence.

BACKGROUND: The signal(s) for removal of senescent platelets from the circulation are not fully understood; phosphatidylserine (PS) expression on platelets and another marker of apoptosis, loss of mitochondrial inner membrane potential (DeltaPsim), have been implicated in platelet clearance. OBJECTIVE: To investigate whether shortened platelet survival and steady-state platelet senescence are associated with increased surface exposure of PS and DeltaPsim collapse. METHODS: Survival of in-vitro biotinylated rabbit platelets treated with thrombin or Ca(2+)-ionophore A23187 was tracked by flow cytometry after injection. Steady-state platelet senescence was investigated by infusing biotin to label a platelet cohort. PS expression and DeltaPsim of in-vitro biotinylated platelets and of the aging platelet cohort biotinylated in-vivo were measured by flow cytometry using annexin V-FLUOS and the DeltaPsim-sensitive dye CMXRos, respectively. RESULTS: Although PS expression, DeltaPsim and survival of thrombin-degranulated platelets were similar to those of control platelets, increasing concentrations of A23187 caused increased surface exposure of PS and progressive shortening of platelet survival; only one-sixth of PS-expressing platelets also exhibited DeltaPsim loss. The cohort of senescent, biotinylated platelets remaining in the circulation at 96 h had increased exposure of PS and collapsed DeltaPsim; of the 17% of PS-expressing platelets, one-third did not exhibit DeltaPsim loss. There was also an increase in platelets with collapsed DeltaPsim but not expressing PS. CONCLUSIONS: Platelets with shortened survival and senescent platelets have increased surface exposure of PS, that may be involved in their clearance. PS expression can occur independently of DeltaPsim collapse and conversely, in aged platelets, DeltaPsim loss can occur independently of PS expression.

Animals↗
PubMed2004

Positive association of the DIO2 (deiodinase type 2) gene with mental retardation in the iodine-deficient areas of China.

BACKGROUND: Iodine deficiency is the commonest cause of preventable mental retardation (MR) worldwide. However, in iodine-deficient areas not everyone is affected and familial aggregation is common. This suggests that genetic factors may also contribute. Thyroid hormone (TH) plays an important role in fetal and early postnatal brain development. The pro-hormone T4 (3,3',5,5'-triiodothyronine) is converted in the brain to its active form, T3, or its inactive metabolite, reverse T3, mainly by the action of deiodinase type 2 (DIO2). METHODS: To investigate the potential genetic contribution of the DIO2 gene, we performed a case-control association study using three common SNPs in the gene (rs225014, rs225012, and rs225010) that were in strong linkage disequilibrium with each other. RESULTS: Single marker analysis showed a positive association of MR with rs225012 and rs225010. Particularly with rs255012 [corrected], CC [corrected] genotype frequency was significantly higher in MR cases than in controls (chi squared [corrected] = 9.18, p = 0.00246). When we compared the distributions of common haplotypes, we also found significant differences between mental retardation and controls in the haplotype combination of rs225012 and rs225010 (chi2 = 15.04, df 2, global p = 0.000549). This association remained significant after Bonferroni correction (p = 0.0016470). CONCLUSION: We conclude that allelic variation in the DIO2 gene may affect the amount of T3 available and in an iodine-deficient environment may partly determine overall risk of MR.

Adolescent↗
PubMed2004

Mechanisms of DNA double strand break repair and chromosome aberration formation.

It is widely accepted that unrepaired or misrepaired DNA double strand breaks (DSBs) lead to the formation of chromosome aberrations. DSBs induced in the DNA of higher eukaryotes by endogenous processes or exogenous agents can in principle be repaired either by non-homologous endjoining (NHEJ), or homology directed repair (HDR). The basis on which the selection of the DSB repair pathway is made remains unknown but may depend on the inducing agent, or process. Evaluation of the relative contribution of NHEJ and HDR specifically to the repair of ionizing radiation (IR) induced DSBs is important for our understanding of the mechanisms leading to chromosome aberration formation. Here, we review recent work from our laboratories contributing to this line of inquiry. Analysis of DSB rejoining in irradiated cells using pulsed-field gel electrophoresis reveals a fast component operating with half times of 10-30 min. This component of DSB rejoining is severely compromised in cells with mutations in DNA-PKcs, Ku, DNA ligase IV, or XRCC4, as well as after chemical inhibition of DNA-PK, indicating that it reflects classical NHEJ; we termed this form of DSB rejoining D-NHEJ to signify its dependence on DNA-PK. Although chemical inhibition, or mutation, in any of these factors delays processing, cells ultimately remove the majority of DSBs using an alternative pathway operating with slower kinetics (half time 2-10 h). This alternative, slow pathway of DSB rejoining remains unaffected in mutants deficient in several genes of the RAD52 epistasis group, suggesting that it may not reflect HDR. We proposed that it reflects an alternative form of NHEJ that operates as a backup (B-NHEJ) to the DNA-PK-dependent (D-NHEJ) pathway. Biochemical studies confirm the presence in cell extracts of DNA end joining activities operating in the absence of DNA-PK and indicate the dominant role for D-NHEJ, when active. These observations in aggregate suggest that NHEJ, operating via two complementary pathways, B-NHEJ and D-NHEJ, is the main mechanism through which IR-induced DSBs are removed from the DNA of higher eukaryotes. HDR is considered to either act on a small fraction of IR induced DSBs, or to engage in the repair process at a step after the initial end joining. We propose that high speed D-NHEJ is an evolutionary development in higher eukaryotes orchestrated around the newly evolved DNA-PKcs and pre-existing factors. It achieves within a few minutes restoration of chromosome integrity through an optimized synapsis mechanism operating by a sequence of protein-protein interactions in the context of chromatin and the nuclear matrix. As a consequence D-NHEJ mostly joins the correct DNA ends and suppresses the formation of chromosome aberrations, albeit, without ensuring restoration of DNA sequence around the break. B-NHEJ is likely to be an evolutionarily older pathway with less optimized synapsis mechanisms that rejoins DNA ends with kinetics of several hours. The slow kinetics and suboptimal synapsis mechanisms of B-NHEJ allow more time for exchanges through the joining of incorrect ends and cause the formation of chromosome aberrations in wild type and D-NHEJ mutant cells.

Androstadienes↗
PubMed2004

Antineutrophil autoantibodies and their target antigens in systemic lupus erythematosus.

Various autoantibodies have been identified in sera from patients with systemic lupus erythematosus (SLE) and autoantibodies against neutrophil have been reported. It was suggested that antineutrophil autoantibodies might be involved in the pathogenesis of neutrocytopenia; however, the role of autoantibodies against neutrophil precursors and their specific target autoantigen(s) remained further characterized. The objective was to investigate the target antigens and clinical associations of autoantibodies against neutrophils and neutrophil precursors in patients with SLE. Sera were collected from 92 patients with SLE and renal biopsy proven lupus nephritis. Cell lysates of peripheral neutrophils (as mature neutrophils) from a normal blood donor and white blood cells from a patient with blast crisis of chronic granulocytic leukemia (CGL) (as neutrophil precursors) were used as antigens in Western blot analysis to detect autoantibodies in sera from patients with SLE. The clinical significance of antineutrophil autoantibodies that recognized different antigens were further analysed. Using normal peripheral neutrophils as antigens, two bands could be blotted: 64 kD (33/92, 35.9%) and 50 kD (13/92, 14.1%). The prevalence of anti-64 kD autoantibody in patients with positive rheumatic factor was significantly higher than that in patients without (54.5 versus 18.8%, P < 0.05). Using CGL white cells as antigen, five bands could be blotted: 60 kD (34/92, 37.0%), 50 kD (32/92, 34.8%), 29 kD (27/92, 29.3%), 42 kD (19/92, 20.7%) and 18 kD (16/92, 17.4%). The prevalence of anti-60 kD autoantibody was significantly higher in patients with neutrocytopenia than that in patients without neutrocytopenia (100 versus 48.3%, P < 0.01). The prevalence of anti-29 kD autoantibody was significantly higher in patients with alopecia than that in patients without alopecia (45.8 versus 20.8%, P < 0.05). Furthermore, the prevalences of anti-60 kD, anti-50 kD and anti-42 kD autoantibodies were significantly higher in patients with anti-Ro autoantibody than those in patients without; the prevalences of anti-29 kD and anti-18 kD autoantibodies were significantly higher in patients with anti-Sm autoantibody than those in patients without. We conclude that there are heterogeneous autoantibodies against both neutrophils and their precursors in sera from patients with SLE. Different autoantibodies may have different clinical significance.

Antibodies, Heterophile↗
PubMed2004

International comparison of performance measures for screening mammography: can it be done?

OBJECTIVE: Published screening mammography performance measures vary across countries. An inter-national study was undertaken to assess the comparability of two performance measures: the recall rate and positive predictive value (PPV). These measures were selected because they do not require identification of all cancers in the screening population, which is not always possible. SETTING: The screening mammography programs or data registries in 25 member countries of the International Breast Cancer Screening Network (IBSN). METHODS: In 1999 an assessment form was distributed to IBSN country representatives in order to obtain information on how screening mammography was performed and what specific data related to recall rates and PPV were collected. Participating countries were then asked to provide data to allow calculation of recall rates, PPV and cancer detection rates for screening mammography by age group for women screened in the period 1997-1999. RESULTS: Twenty-two countries completed the assessment form and 14 countries provided performance data. Differences in screening mammography delivery and data collection were evident. For most countries, recall rates were higher for initial than for subsequent mammograms. There was no consistent relationship of initial to subsequent PPV, although PPV generally decreased as the recall rate increased. Recall rates decreased with increasing age, while PPV increased as age increased. CONCLUSION: Similar patterns for mammography performance measures were evident across countries.However, the development of a more standardized approach to defining and collecting data would allow more valid international comparisons, with the potential to optimize mammography performance. At present, international comparisons of performance should be made with caution due to differences in defining and collecting mammography data.

Breast Neoplasms↗
PubMed2004

Using atomic force microscopy to study nucleosome remodeling on individual nucleosomal arrays in situ.

In eukaryotes, genomic processes like transcription, replication, repair, and recombination typically require alterations in nucleosome structure on specific DNA regions to operate. ATP-dependent nucleosome remodeling complexes provide a major mechanism for carrying out such alterations in vivo. To learn more about the action of these important complexes, we have utilized an atomic force microscopy in situ technique that permits comparison of the same individual molecules before and after activation of a particular process, in this case nucleosome remodeling. This direct approach was used to look for changes induced by the action of the human Swi-Snf remodeling complex on individual, single-copy mouse mammary tumor virus promoter nucleosomal arrays. Using this technique, we detect a variety of changes on remodeling. Many of these changes are larger in scale than suggested from previous studies and involve a number of DNA-mediated events, including a preference for the removal of a complete turn (80 basepairs) of nucleosomal DNA. The latter result raises the possibility of an unanticipated mode of human Swi-Snf interaction with the nucleosome, namely via the 11-nm histone surface.

Adenosine Triphosphate↗
PubMed2004

Epigenetic regulation of inhibin alpha-subunit gene in prostate cancer cell lines.

Inhibin was first identified as a gonad-derived regulator of pituitary FSH; however, it has subsequently been shown to be a tumour suppressor in the gonad and adrenal glands. Whereas non-malignant regions of human primary prostate carcinomas express inhibin alpha-subunit (INHA), malignant tissues lack INHA transcript and protein, which is consistent with epigenetic regulation of the inhibin alpha-subunit gene (INHA) promoter. This study investigated whether methylation of the INHA promoter was responsible for inactivation of INHA transcription and translation in the prostate cancer cell lines, LNCaP, DU145 and PC3. Methylation of the promoter was revealed by bisulphite genomic sequencing and use of inhibitors of methylation and histone deacetylation resulted in reactivation of the INHA transcription and translation. Significant (P<0.05) downregulation of a luciferase reporter gene downstream from a methylated INHA promoter compared with unmethylated INHA promoter occurred in vitro. The data demonstrate that promoter methylation is associated with downregulation of the INHA gene in prostate cancer cell lines, which is consistent with its tumour suppressive role. Therefore INHA has a significant role in prostate tumorigenesis.

Base Sequence↗
PubMed2004

Trends in cancer incidence among Singapore Malays: a low-risk population.

INTRODUCTION: Inspection of trends in cancer incidence in ethnically or geographically diverse populations is important for the understanding of cancer patterns and also to provide clues for aetiologic studies. There is little information on cancer incidence among Malays, a low-risk population. The population-based cancer registration system in Singapore offers the opportunity in this regard. MATERIALS AND METHODS: A review of all newly diagnosed cancer cases among Singapore Malay residents in the period 1968 to 1997 was conducted to determine the time trends using data from Singapore Cancer Registry. Age-standardised incidence rates and average annual percentage change, using Poisson regression, were calculated. RESULTS: During the 30-year study period, a total of 9101 incident cases of cancer were diagnosed among Malays in Singapore. The 3 most common sites of cancer were lung, liver and colorectum in males; and breast, colorectum and ovary in females in the period 1983 to 1997. Leukaemia was among the top 10 cancer sites in both sexes. Malays generally experienced lower cancer incidence rates compared to Chinese, but rates have been increasing at 1.5% to 2% annually. Statistically significant increases in incidence were observed for most of the cancer sites. The incidence of nasopharyngeal carcinoma increased, while there were no significant changes in the general population. CONCLUSION: The relatively lower cancer risk among Singapore Malays might be attributed to genetic factors or closer adherence to a more traditional lifestyle. This study provides a baseline for comparisons with other populations with diverse cancer risk patterns in order to obtain a better understanding of possible aetiologic factors.

Asia, Southeastern↗
PubMed2003

Kinetic pathway of the bilayered-micelle to perforated-lamellae transition.

Using time-resolved small-angle neutron scattering, we have studied the kinetics of the recently observed bilayered-micelle (or so-called "bicelle") to perforated-lamellar transition in phospholipid mixtures. The data suggest that phase-ordering occurs via the early-time coalescence of bicelles into stacks of lamellae that then swell. Our measurements on this biomimetic system highlight the ubiquitous role of transient metastable states in the phase ordering of complex fluids.

Biophysics↗
PubMed2003

Early-stage compositional segregation in polymer-blend films.

The existence of a transient period during the surface enrichment of a binary polymer blend by one of its components has been suggested by previous theoretical and experimental studies as well as computer simulations. Taking advantage of the high depth resolution of neutron reflectivity and the slow dynamics of polymers near their glass transition, we investigate this early-stage surface compositional enrichment in a phase separating polymer blend for the first time. Two stages of surface enrichment layer growth are observed. A rapid local surface enrichment at the chain segmental level occurs first, followed by a slower growth of a diffuse layer having a scale on the order of the bulk correlation length and the radius of gyration of the surface enriching polymer chains.

Journal Article↗