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Biomedical subjects

D Wang

Publications and source records attributed to D Wang.

At least 667 records · Page 37Linked to original sources
PubMed1997

Viral determinants of pea early browning virus seed transmission in pea.

Pea early browning virus (PEBV) is a member of the genus, Tobravirus. It is transmitted by soil-inhabiting trichodorid nematodes and through seeds from diseased plants. By introducing mutations into the PEBV genome, we have studied the viral determinants of seed transmission in pea. Neither deleting a portion of the genome containing the three nonstructural genes in RNA2 nor the interuption of any of the three genes individually prevented PEBV seed transmission. However, a comparison of two PEBV isolates indicated a minor role for RNA2 or its products. In contrast, the removal of the coding sequence of the 12K gene in RNA1 almost completely abolished viral seed transmission. The virus lacking the 12K gene caused more severe symptoms on leaves and pods, and accumulated to a higher level than the wild-type virus in both types of tissues. However, the 12K deletion mutant accumulated poorly in anthers and carpels, and could not be detected in pollen grains and ovules. These results suggest that the 12K gene is involved in the infection of the gametic cells and hence the seed transmission of PEBV in pea.

Pisum sativum↗
PubMed1997

Kinetics of oxidized cytosine repair by endonuclease III of Escherichia coli.

Endonuclease III of Escherichia coli excises a broad range of oxidized, hydrated and ring-fragmented pyrimidines from DNA. The kinetic parameters were compared for repair of three potentially mutagenic oxidized cytosine lesions: 5,6-dihydroxy-5, 6-dihydro-2'-deoxyuridine (uracil glycol or Ug), 5-hydroxy-2'-deoxycytidine (5-ohC), and 5-hydroxy-2'-deoxyuridine (5-ohU). Site-specifically modified 40-mer oligonucleotides containing each of the three lesions in the same sequence context were synthesized chemically or by a combination of chemical and enzymatic methods. Appropriately protected phosphoramidites of 5-ohC and 5-ohU were synthesized and incorporated into oligonucleotides by standard solid-phase synthetic methods. The lability of Ug made it necessary to use an alternative approach to prepare the analogous 40-mers containing Ug. An uracil containing pentamer oligonucleotide was oxidized with OsO4 to generate the corresponding Ug containing product, which was then ligated into an oligonucleotide scaffold to generate 40 base pair duplexes. Using 32P-labeled substrates and a gel electrophoresis based assay, the values of Km and Vmax for excision of 5-ohC, 5-ohU, and Ug were determined. In this experimental system, the order of repair efficiency is Ug >> 5-ohC >> 5-ohU based on ratios of Vmax/Km. Modest effects were observed when the base paired opposite the lesion was changed from G to A.

Base Composition↗
PubMed1997

Transcription factor AP-2 controls transcription of the human transforming growth factor-alpha gene.

The epidermal growth factor receptor is vital for normal development and plays a role in oncogenesis. The level of activation of this receptor by transforming growth factor-alpha (TGF-alpha) is controlled, in part, by the rate of transcription of the TGF-alpha gene. In the characterization of the proximal TGF-alpha promoter by DNase I footprinting, a 43-base pair element (-88 to -130 relative to the transcription start site), designated TalphaRE I, was found that was specifically protected by nuclear proteins from human mammary carcinoma MDA468 cells. TalphaRE I was essential for the maximal expression of the TGF-alpha gene as indicated by deletion and mutagenesis analyses. TalphaRE I consists of two cis-acting elements, a proximal regulatory element (PRE, -89 to -103) and a distal regulatory element (DRE, -121 to -128). Both elements were able to form specific complexes with protein from MDA468 cell nuclear extracts and are necessary for the full activity of the entire 1.1-kilobase pair TGF-alpha promoter. Competition and antibody studies determined that the DRE contains a binding site for the transcription factor AP-2, while the protein that binds to the PRE has yet to be identified. When linked upstream to the heterologous herpes simplex thymidine kinase promoter, the TalphaRE I enhanced transcription up to 11-fold in MDA468 cells. Cotransfection of an AP-2 expression vector was able to activate transcription from the TalphaREI-TK construct in a DRE-dependent manner. These results further our understanding of how TGF-alpha transcription is regulated.

Base Sequence↗
PubMed1997

Basic fibroblast growth factor transcriptional autoregulation requires EGR-1.

Basic fibroblast growth factor (bFGF) is an important growth factor for neuroectoderm- and mesoderm-derived cells. In addition bFGF is an important angiogenic factor and appears to contribute to tumorigenesis. This is exemplified by the fact that bFGF is expressed in a large majority of human gliomas and that bFGF expression is critical for the growth and tumorigenesis of these cells. It has been shown previously that bFGF can induce its own expression through an increase in bFGF mRNA. In this report, we show that bFGF leads to its own synthesis through an autoregulated transcriptional response that requires the transcription factor Egr-1 (also known as Krox24, Zif268 and NGFI-A). Egr-1 binds to two DNA elements in the bFGF promoter and positively regulates transcription. Mutation of these sites blocks the ability of bFGF to transcriptionally regulate the bFGF promoter. These data indicate a mechanism to explain how bFGF functions to autoregulate its expression and demonstrate that Egr-1 is as an essential transcription factor in this process.

Animals↗
PubMed1997

Image segmentation based on oscillatory correlation.

We study the image segmentation on the basis of locally excitatory, globally inhibitory oscillator networks (LEGION), whereby the phases of oscillators encode the binding of pixels. We introduce a lateral potential for each oscillators so that only oscillators with strong connections from their neighborhood can develop high potentials. Based on the concept of the lateral potential, a solution to remove noisy regions in an image is proposed for LEGION, so that it suppresses the oscillators corresponding to noisy regions but without affecting those corresponding to major regions. We show that the resulting oscillator network separates an image into several major regions, plus a background consisting of all noisy regions, and we illustrate network properties by computer stimulation. The network exhibits a natural capacity in segmenting images. The oscillatory dynamics leads to a computer algorithm, which is applied successfully to segmenting real gray-level images. A number of issues regarding biological plausibility and perceptual organization are discussed. We argue that LEGION provides a novel and effective framework for image segmentation and figure-ground segregation.

Algorithms↗
PubMed1997

GABAergic synapses on mu-opioid receptor-expressing neurons in the superficial dorsal horn: an electron microscope study in the cat spinal cord.

A double-immunocytochemical electron microscope study was performed in the cat to examine whether GABAergic axons might be in synaptic contact with spinal neurons expressing mu-opioid receptor (MOR) in laminae I and II of the spinal dorsal horn at the lumbar cord segments. Structures showing MOR-like immunoreactivity (-LI) and those showing GABA-LI were labeled, respectively, with diaminobenzidine/peroxidase-reaction products and immunogold particles. Approximately one-third of dendritic profiles with MOR-LI in laminae I and II were postsynaptic to axon terminals with GABA-LI; about one-fourth of somatic profiles with MOR-LI were also postsynaptic to axon terminals with GABA-LI. The results suggest that activation of MOR on postsynaptic neurons may modulate effects which are induced by GABA released from presynaptic neurons.

Animals↗
PubMed1997

Multiple interactions stabilize a single paused transcription intermediate in which hairpin to 3' end spacing distinguishes pause and termination pathways.

Transcription is delayed in the leader regions of the Escherichia coli trp and his operons by multipartite pause signals that consist of four components: a nascent RNA structure (the pause hairpin), the 10 or 11 nt 3'-proximal region between the pause hairpin and the RNA 3' end, the bases in the active site, and approximately 14 bp of duplex DNA downstream from the pause site. Results described in the accompanying paper suggest that the his pause hairpin slows nucleotide addition via interaction with an easily disordered surface on RNA polymerase. Here we report that the four pause signal components slow nucleotide addition in a single kinetic intermediate. Formation of the paused transcription complex, in contrast, involves synergistic effects of RNA and DNA sequences that select the wild-type pause site from among several adjacent possibilities. Extending the pause hairpin with one G x C base-pair reduces pausing, apparently by interfering with pause hairpin interaction; adding a second C x G base-pair that reduces the 3'-proximal RNA to 9 nt or less (within the 7 to 9 nt characteristic of rho-independent terminators) induces transcript release. We propose that escape from the pause is governed by a rate-limiting isomerization that may require substrate NTP binding to re-establish the active site geometry, whereas transcript release and termination ensue when the hairpin interaction is weakened and isomerization to an active conformation is blocked.

Bacterial Proteins↗
PubMed1997

Cloning and partial characterization of the mouse glutamine:fructose-6-phosphate amidotransferase (GFAT) gene promoter.

Glutamine:fructose-6-phosphate amidotransferase (GFAT) is the enzyme that is rate limiting in the synthesis of glucosamine and hexosamines. Glucosamine has been proposed to contribute to the glucotoxicity of diabetes. Evidence that the gene encoding GFAT is transcriptionally regulated prompted us to clone and characterize its promoter. The position of the mouse GFAT promoter relative to the translational start site was located by primer extension and found to be 149 bp upstream of the translational start site. A 1.9 kb SacI fragment of the GFAT gene was found to contain the promoter and 88 bp of sequence downstream of the transcriptional start site. This promoter segment could drive expression of a luciferase reporter gene, could confer correct transcriptional initiation to the reporter and could confer the EGF-responsiveness previously observed in the native gene. The mouse GFAT promoter lacks a canonical TATA box and has several GC boxes within a highly GC-rich region. Deletional analysis of the promoter indicated that a proximal element extending to -120 relative to the transcriptional start site could confer reporter expression at a level of 57% of the 1.9 kb construct. Detailed analysis of this proximal region by DNase I footprinting, electrophoretic mobility shift assays and site-directed mutagenesis indicated that Sp1 binds to three elements in this proximal promoter segment and plays a vital role in regulation of transcription from this gene.

3T3 Cells↗
PubMed1997

Integrin alpha8beta1 is critically important for epithelial-mesenchymal interactions during kidney morphogenesis.

We present genetic evidence that integrins regulate epithelial-mesenchymal interactions during organogenesis. Mice with a mutation in the alpha8 gene do not express the integrin alpha8 beta1 and exhibit profound deficits in kidney morphogenesis. In wild-type animals, inductive interactions between the ureteric epithelium and metanephric mesenchyme are essential for kidney morphogenesis. In alpha8 mutant homozygotes, growth and branching of the ureteric bud and recruitment of mesenchymal cells into epithelial structures are defective. Consistent with these phenotypes, alpha8 expression is induced in mesenchymal cells upon contact with the ureter. Since none of its previously identified ligands appears likely to mediate the essential functions of alpha8 beta1 in kidney morphogenesis, we have used an alpha8 beta1-alkaline phosphatase chimera to localize novel ligand(s) in the growing ureter. The distribution of these ligand(s) makes them strong candidates for regulators of kidney morphogenesis.

Alleles↗
PubMed1997

Nuclease cleavage of the upstream half of the nontemplate strand DNA in an Escherichia coli transcription elongation complex causes upstream translocation and transcriptional arrest.

We tested the susceptibility of nucleic acid strands in a halted transcription elongation complex to digestion by micrococcal nuclease (MN). The 16-nucleotide nascent RNA was protected within RNA polymerase. A 27-28-nucleotide template strand DNA fragment also was resistant to MN digestion. However, the upstream half of the nontemplate DNA within this region was digested rapidly by MN, suggesting that the nontemplate strand emerges from the RNA polymerase near the middle of the melted transcription bubble with the bases oriented away from the enzyme surface. MN cleavage of the exposed nontemplate DNA shifted polymerase backward, making it unable to extend the RNA chain. However, the MN-trimmed G16 complexes could be reactivated by GreB-stimulated cleavage of the nascent RNA. These results favor a model of transcriptional arrest involving upstream slippage of RNA polymerase along the RNA and DNA chains. They also suggest that the exposed segment of nontemplate DNA may directly or indirectly stabilize the lateral position of the transcription complex along the DNA.

Base Sequence↗
PubMed1997

Localization and up-regulation of mucin (MUC2) gene expression in human nasal biopsies of patients with cystic fibrosis.

Using digoxigenin-UTP-labelled human HAM-1 (92 bp) or SMUC41 (850 bp) cRNA probes, the expression and localization of MUC2 gene transcripts were determined by in situ hybridization in human nasal tissues obtained as biopsies from 12 patients with cystic fibrosis (CF): all had been part of a gene therapy trial in which CFTR cDNA-liposome complexes had been delivered by topical application to eight and liposome alone to four as a placebo control. For comparison, there were nasal tissues taken at surgical resection from four non-CF subjects and a further four biopsies taken from normal healthy volunteer controls. Both SMUC41 and HAM-1 probes provided a strong signal. MUC2 mRNA transcripts were present in serous and mucous acini of submucosal glands, ciliated and basal cells of the surface epithelium, and occasional mononuclear inflammatory cells. The percentages (mean +/- SEM) of serous and mucous acini showing positivity for MUC2 gene expression in the four samples surgically resected from non-CF subjects were 25.4 +/- 5.6 and 26.7 +/- 3.3 per cent, respectively. Compared with the non-CF subjects, the mean percentage of acini showing MUC2 gene expression in the four placebo-treated CF subjects was significantly higher for serous (80.5 +/- 12.7 per cent; P < 0.05, t-test), but not for mucous acini (53.1 +/- 16.8 per cent; P = 0.38). In CF and non-CF groups, where present, MUC2 positivity was strongly expressed and constituted approximately 84 per cent of the cell area in serous acini, whereas it was less obvious and was confined to the perinuclear area of cells in mucous acini. A significantly greater proportion of the surface epithelium was positive for MUC2 mRNA transcripts in the CF subjects (89.0, +/- 1.4 per cent) than in the surgically resected tissues of the four non-CF subjects (19.4, +/- 4.0 per cent) (P = 0.02). In the eight CFTR-cDNA-treated subjects, there was an overall trend to reduction, but no statistically significant alteration of MUC2 gene expression. It is concluded that the MUC2 gene is expressed at three- to four-fold higher levels in CF nasal mucosa than in non-CF nasal tissue and that it is expressed in a variety of cells additional to submucosal mucus-secreting glands.

Adult↗
PubMed1997

Germ cell nuclear antigen (GCNA1) expression does not require a gonadal environment or steroidogenic factor 1: examination of GCNA1 in ectopic germ cells and in Ftz-F1 null mice.

The germ cell lineage is first recognized as a population of mitotically proliferating primordial germ cells that migrate toward the gonadal ridge. Shortly after arriving at the gonadal ridge, the germ cells begin to initiate a commitment to gamete production in the developing gonad. The mechanisms controlling this transition are poorly understood. We recently reported that a mouse germ cell nuclear antigen 1 (GCNA1) is initially detected in both male and female germ cells as they reach the gonad at 11.5 days postcoitum (dpc). GCNA1 is continually expressed in germ cells through all stages of gametogenesis until the diplotene/dictyate stage of meiosis I. Since GCNA1 expression commences soon after primordial germ cells arrive at the gonadal ridge, we wanted to determine whether the gonadal environment was essential for induction of GCNA1 expression. By examining GCNA1 expression in germ cells that migrate ectopically into the adrenal gland, we determined that both the gonadal and adrenal gland environments allow GCNA1 expression. We also examined GCNA1 expression Ftz-F1 null mice, which were born lacking gonads and adrenal glands. During embryonic development in the Ftz-F1 null mice, the gonad and most germ cells undergo apoptotic degeneration at about 12.5 dpc. While most of the germ cells undergo apoptosis without expressing GCNA1, a few surviving germs cells, especially outside the involuting gonad clearly express GCNA1. Thus, although the Ftz-F1 gene is essential for gonadal and adrenal development, induction of GCNA1 expression in germ cells does not require Ftz-F1 gene products. The finding that germ cell GCNA1 expression is not restricted to the gonadal environment and is not dependent on the Ftz-F1 gene products suggests that GCNA1 expression may be initiated in the germ cell lineage by autonomous means.

Animals↗
PubMed1997

Experimental study on modified treatment and endothelialization of bovine pericardial valves.

The purpose of this study were to confirm whether the modified treatment with L-glutamic acid could attenuate the calcification of the GA-fixed valves and improve its biocompatibility. Pericardial valves were routinely treated with GA and valves were treated with GA and 8% L-glutamic acid. The valves treated with these methods were implanted subcutaneously in rats. Calcium deposits of the valves collected at the 7th, 21st, 60th, 90th day were assessed by atomic absorption spectroscopy, and the pathologic changes were examined by light and electron microscopy. Cultured endothelial cells (ECs) were seeded onto the valves. The cell counts were determined at the 1st, 4th, 7th, 10th day after seeding. PGI2 in culture medium was tested at the 10th day. Transmission and scanning electron microscopy were used to observe the growth of ECs on the valves. Results showed that subsequent treatment with L-glutamic acid could significantly mitigate calcification of bovine pericardial valves as compared with simple GA-fixed valves (P < 0.01). ECs seeded on the GA treated valves died within 4 days. On the valves treated by modified method, ECs could proliferate and release PGI2. It is concluded that treatment with L-glutamic acid can markedly inhibit the calcification and improve the biocompatibility of bioprosthetical valves.

Animals↗
PubMed1997

Treatment of traumatic defect of the tibia with two exposed fractured ends--reduction and lengthening at the proximal metaphysics of tibia.

From Jan. 1995 to June 1996, 6 cases of traumatic defect of the tibia were treated by remodeling and reduction of fractured ends and osteotomy and distraction at the proximal metaphysics of the tibia. In 6 patients, aged 19 to 54, with the limb-length discrepancy being at least 3 to 4.5 cm, the tibia was lengthened by 3 cm in 3 cases, 4 to 4.5 cm in 3 cases. The consolidation and maturation of the bone were accomplished within 3 to 6 months. The authors believe that this procedure is simple, less invasive and involves no bone-graft.

Adult↗
PubMed1997

Myoglobin and hemoglobin rotational diffusion in the cell.

The detection of the 1H NMR signal of myoglobin (Mb) in tissue opens an opportunity to examine its cellular diffusion property, which is central to its purported role in facilitating oxygen transport. In perfused myocardium the field-dependent transverse relaxation analysis of the deoxy Mb proximal histidyl NdeltaH indicates that the Mb rotational correlation time in the cell is only approximately 1.4 times longer than it is in solution. Such a mobility is consistent with the theory that Mb facilitates oxygen diffusion from the sarcoplasm to the mitochondria. The microviscosities of the erythrocyte and myocyte environment are different. The hemoglobin (Hb) rotational correlation time is 2.2 longer in the cell than in solution. Because both the overlapping Hb and Mb signals are visible in vivo, a relaxation-based NMR strategy has been developed to discriminate between them.

Animals↗
PubMed1997

Lactose hydrolysis and beta-galactosidase activity in sonicated fermentation with Lactobacillus strains.

Milk fermentation with four Lactobacillus strains, including L. delbrueckii subsp. bulgaricus B-5b, L. helveticus LH-17, L. delbrueckii subsp. lactis SBT-2080 and L. acidophilus SBT-2068, was carried out under sonicated conditions. Under continuous sonication, viable cell count decreased or grew less than in conventional fermentation, but it increased considerably in static incubation after sonication. Sonication caused beta-galactosidase release from lactic acid bacteria cells to the culture medium, thus resulting in higher total beta-galactosidase activity. However, lactose hydrolysis was enhanced only when beta-alactosidase was effectively released. With L. delbrueckii subsp. bulgaricus B-5b and L. helveticus LH-17. the degrees of lactose hydrolysis achieved were about 75% which is much higher than those in conventional fermentation (below 40%). Results presented here showed that a high viable cell count and a high degree of lactose hydrolysis could be simultaneously achieved by a suitable sonication method.

Culture Media↗