Search PubMed⌕ Search

Biomedical subjects

Y Wu

Publications and source records attributed to Y Wu.

At least 379 records · Page 21Linked to original sources
PubMed2000

Osteopontin is required for full expression of the transformed phenotype by the ras oncogene.

The secreted phosphoprotein osteopontin (OPN) is strongly associated with the process of neoplastic transformation, based both on its pattern of expression in vivo and in vitro and on functional analyses. We have used 3T3 cells derived from wildtype and OPN-deficient mice and transformed by transfection with oncogenic ras to assess the role of OPN in transformation in vitro and in tumorigenesis in vivo. There was no effect of an absence of OPN on the ability of the cells to undergo immortalization or to form morphologically transformed foci following ras transfection. Wildtype and OPN-deficient cell lines were established from such foci, and lines with similar ras mRNA levels selected for further analysis. Ras-transformed cell lines from both wildtype and OPN-deficient mice could form colonies in soft agar indicating that this process can occur in the absence of OPN. However, the ability of the OPN-deficient cell lines to form colonies was reduced as compared to wildtype cell lines. Tumorigenesis in syngeneic and nude mice was assessed for a subset of cell lines that formed colonies efficiently in soft agar. Cell lines unable to make OPN formed tumors in these mice much more slowly than wildtype cells, despite similar growth of the cells on plastic and in soft agar. Taken together, these results indicate that maximal transformation by ras requires OPN expression, and implicate increased OPN expression as an important effector of the transforming activity of the ras oncogene.

3T3 Cells↗
PubMed2000

Orientation of membrane-bound melittin studied by a combination of HPLC and liquid secondary ion mass spectrometry (LSIMS).

Combining two analytical techniques, HPLC and liquid secondary ion mass spectrometry, the orientation of liposomal membrane-bound melittin was analyzed through its trypsin-digested products. We found that trypsin can access all proteolytic sites of the membrane-bound melittin when the liposomes have no transmembrane potential, whereas the proteolytic site near the N terminus of melittin is blocked when the liposomes have a negative transmembrane potential. The results suggest that the negative transmembrane potential may induce the melittin molecules to insert into the membrane perpendicularly, whereas melittin lies flat on the membrane surface in the absence of a negative potential.

Amino Acid Sequence↗
PubMed2000

Dynamic splitting of large intensity-modulated fields.

The aims of this paper are to describe a method of splitting large intensity-modulated fields that cannot be delivered as a single field and to verify the accuracy of our method. Some multi-leaf collimators may be operated in the dynamic mode to deliver intensity-modulated radiation treatments (IMRT) using the 'sliding window' technique. In this technique each pair of leaves sweeps over the treatment field while the beam is on. However, there are limitations on the width of the field that can be treated due to the limited length of the leaves. For instance, the leaf length of the Varian MLC is 14.5 cm. Since each leaf pair must travel from the left boundary to the right boundary of the beam aperture, the maximum width of the field aperture that can be accommodated in one sweep of leaves is also limited to 14.5 cm, in fact to a slightly smaller value. It has been shown that IMRT is more efficient when used to plan and deliver the large and boost fields simultaneously. In such situations, the fields must be large enough to cover simultaneously the volumes of the gross tumour, microscopic disease and electively treated regions. Such field sizes are often larger than 14.5 cm wide. In this paper, we present a dynamic 'feathering' technique to split the large intensity-modulated fields into smaller fields. In this technique, the component beams overlap each other by a small amount, and the intensity in the overlap region gradually decreases for one field component and increases for the other. The sum of intensities remains the same as for the original field. This method eliminates the field matching problems associated with the conventional step 'break' for static fields. The splitting process is integrated into the IMRT treatment procedure and the entire planning process is automated. Comparison of dose distributions calculated and measured in a phantom showed good agreement. Such a method can be applied to the 'step and shoot' technique as well. IMRT fields of widths up to 25 cm can be delivered by splitting only once, which is adequate for most treatments.

Algorithms↗
PubMed2000

Adaptation of roots to low water potentials by changes in cell wall extensibility and cell wall proteins.

It is common for the root/shoot ratio of plants to increase when water availability is limiting. This ratio increases because roots are less sensitive than shoots to growth inhibition by low water potentials. The physiological and molecular mechanisms that assist root growth under drought conditions are reviewed, with a focus on changes in cell walls. Maize seedlings adapt to low water potential by making the walls in the apical part of the root more extensible. In part, this is accomplished by increases in expansin activity and in part by other, more complex changes in the wall. The role of xyloglucan endotransglycosylase, peroxidase and other wall enzymes in root adaptation to low water potential is evaluated and some of the complications in the field of study are listed.

Adaptation, Physiological↗
PubMed2000

Multiple paths of sugar-sensing and a sugar/oxygen overlap for genes of sucrose and ethanol metabolism.

The two-fold purpose of this work is, first, to review current hypotheses for multiple paths of sugar-sensing in an oxygen-responsive context, and second, to present evidence for the extent of sugar/oxygen overlap regulating genes for sucrose and ethanol metabolism. Current data indicate that sugar signals in plants may be initiated by (a) hexokinases, (b) membrane sensors, (c) acetate and/or respiratory metabolites, and (d) other signals and/or crosstalk. Responses may also involve concurrent input along transduction paths by effectors such as energy charge, P status, and phytohormones. Prime candidates for initiation and/or integration of such signal integration include SNF1- and SCF-like, multi-enzyme complexes. In addition, different paths of sugar signal transduction may be linked to contrasting roles of responsive genes during feast, famine or pathogen attack. Oxygen can potentially alter sugar signals at several points, so its influence on feast and famine responses was initially tested with genes for sucrose metabolism in maize root tips. The Sus1 and Sh1 sucrose synthases in maize (typically up-regulated by carbohydrate abundance and deprivation, respectively) showed parallel responses to hypoxia (3% O2 [0.03l l-1 O2]) and anoxia (0% O2 [0l l-1 O2]) that were consistent with involvement of similar signals. In contrast, the differential sugar-responses of the lvr1 and lvr2 invertases were not evident under low oxygen, and both genes were rapidly repressed. A third response was evident in the marked, sugar-regulation of an oxygen-responsive Adh1 gene for alcohol dehydrogenase, which was sensitive to sugar availability from deficit to abundance, regardless of oxygen status (anaerobic to fully aerobic [40% O2 (0.04l l-1 O2)]. A clear interface is thus evident between sugar and oxygen signals, but this varies markedly with the genes involved and probable differences in respective transduction paths.

Carbohydrate Metabolism↗
PubMed2000

Regulation of intestinal apolipoprotein B mRNA editing levels by a zinc-deficient diet and cDNA cloning of editing protein in hamsters.

This study was conducted to investigate the influence of dietary zinc on intestinal apoB mRNA editing in hamsters. Apolipoprotein B-48 (apoB-48) is synthesized from the same gene as apoB-100 by a post-transcriptional, site-specific cytidine deamination, a process known as apoB mRNA editing. A cDNA encoding the hamster apoB mRNA editing enzyme was obtained by reverse transcriptase-polymerase chain reaction (RT-PCR) and the deduced amino acid sequence was found to possess high amino acid sequence identity to apoB mRNA editing enzymes from several other species. Editing activity was detected in the small intestine and colon but, like humans, none was detected in the liver. Analysis by RT-PCR indicated that the small intestine possessed the highest expression of editing enzyme mRNA abundance, whereas both liver and small intestine expressed relatively high levels of apoB mRNA. The influence of dietary zinc on intestinal apoB mRNA editing levels was examined in Golden Syrian hamsters (7 wk old) assigned to one of the following three dietary treatments: Zn-adequate (ZA, 30 mg Zn/kg diet), Zn-deficient (ZD, <0. 5 mg Zn/kg diet), or Zn-replenished (ZDA, ZD hamsters receiving ZA diet for last 2 d) for 7 wk. Hamsters consuming the ZD diet had modestly but significantly lower intestinal editing activity than ZA hamsters. Intestinal editing activity in the ZDA group was not different from that of ZA hamsters. Data derived from these studies contribute to the understanding of lipoprotein metabolism in hamsters, a suitable model for the study of atherosclerosis.

Amino Acid Sequence↗
PubMed2000

Pathogenesis of axonal degeneration: parallels between Wallerian degeneration and vincristine neuropathy.

Peripheral neuropathies and Wallerian degeneration share a number of pathological features; the most prominent of which is axonal degeneration. We asked whether common pathophysiologic mechanisms are involved in these 2 disorders by directly comparing in vitro models of axonal degeneration after axotomy or exposure to the neurotoxin vincristine. Embryonic rat dorsal root ganglia (DRG) were allowed to extend neurites for 5 days in culture, and then were either axotomized or exposed to 0.01 microM vincristine. Neurites universally degenerated by 3 days after axotomy or after 6 days of vincristine exposure. The neuroprotective effects of a low calcium environment or pharmacologic inhibition of the cysteine protease calpain were compared in these 2 models of axonal degeneration. Addition of EGTA or growth in zero-calcium media provided significant protection against axonal degeneration after either axotomy or vincristine exposure. Treatment with the experimental calpain inhibitor AK295 was equally protective in both models. Chronic exposure to AK295 was not toxic to the cultures. These data suggest that common mechanisms involving calcium and calpains are involved in both axotomy-induced and vincristine-induced axonal degeneration. In addition, calpain inhibition may provide a strategy for preventing axonal degeneration and preserving neurologic function in a variety of PNS and CNS disorders.

Animals↗
PubMed2000

Bombesin recovers gut-associated lymphoid tissue and preserves immunity to bacterial pneumonia in mice receiving total parenteral nutrition.

OBJECTIVE: To study the ability of bombesin (BBS) to recover gut-associated lymphoid tissue (GALT) and preserve immunity in a lethal model of Pseudomonas aeruginosa (Ps) pneumonia in mice receiving total parenteral nutrition (TPN). SUMMARY BACKGROUND DATA: TPN causes depression of mucosal immunity compared with enterally fed animals, which may explain the increased incidence of pneumonia in parenterally fed trauma patients. BBS prevents this TPN-induced GALT atrophy, depressed gastrointestinal and respiratory tract IgA levels, and impaired antiviral IgA-mediated mucosal immunity. The authors examined whether some supplement could be added to TPN to avoid this GALT atrophy and lower the incidence of infectious complications in the parenterally fed animal. METHODS: Male mice were randomized to chow or intravenous (IV) TPN. After 5 days of IV TPN, mice received 0, 1, 2, or 3 days of BBS IV three times a day and then were killed to harvest Peyer's patch, intraepithelium, and lamina propria for cell yields. Gastrointestinal and respiratory tract IgA levels were analyzed by enzyme-linked immunosorbent assay. Next, mice underwent intranasal inoculation with liposomes alone (nonimmune) or liposome-containing Ps polysaccharide. Ps immune mice were catheterized and randomized to chow, IV TPN, or IV TPN + BBS. The liposome group received chow but no IV catheter. These mice were given an LD90 dose of intratracheal Ps, and death rates were recorded. RESULTS: GALT and gastrointestinal and respiratory tract IgA levels improved to those in chow-fed mice after 3 days of BBS. Immunization reduced the death rate from 92% in chow-fed liposome-only animals to 20% in immunized animals. TPN-fed animals lost their mucosal immunity, with a death rate of 86% compared with 21% in the TPN + BBS group. CONCLUSION: The results demonstrate that BBS reverses TPN-induced changes in GALT and preserves mucosal immunity. Ps immunization reduces the death rate in a gram-negative pneumonia model and maintains gastrointestinal and respiratory immunity in Ps immune mice receiving IV TPN.

Animals↗
PubMed2000

Administration of selective endothelin receptor type A antagonist Ro 61-1790 does not improve outcome in focal cerebral ischemia in cat.

The authors examined the effect of selective endothelin (ET) receptor type A (ET(A)) antagonism on histological and functional recovery in cat at 24 hours after reversible middle cerebral artery occlusion (MCAO). A novel and specific ET(A) antagonist, Ro 61-1790 [5-methylpyridine-2-sulfonic acid-6-(2-hydroxyethoxy)-5-(2-methoxyphenoxy)-2-(2-1H-tetrazol-5-y l-pyridin-4-yl)-pyrimidin-4-ylamide sodium salt (1:2)] (Roche, Basel, Switzerland), was used at doses that produced steady-state plasma concentrations and abolished ET-induced pial arteriolar vasoconstriction. In a cranial window preparation, 8 nmol/L ET constricted pial arterioles by 33 +/- 18% (mean +/- SD), but this response was ablated by intravenous Ro 61-1790 treatment (10-mg/kg bolus, 4-mg/kg/h infusion). In additional animal cohorts, halothane-anesthetized cats were treated with 90 minutes of MCAO and 24 hours of reperfusion. Animals received Ro 61-1790 infusion beginning at the onset of reperfusion and continuing for 6 or 24 hours (n = 41). Control cats were treated with 0.9% saline by intravenous infusion throughout reperfusion. There was no difference in injury volume or neurologic evaluation score in saline-treated cats (n = 11; caudate 24 +/- 28%, cortical injury 7.5 +/- 5% of ipsilateral structure; score 52 +/- 8) versus the results in cats treated with Ro 61-1790 for either 24 hours (n = 6; caudate 22 +/- 23%, cortex 6 +/- 5%, injury volume of ipsilateral structure; score 55 +/- 3) or 6 hours (n = 11; caudate 33 +/- 30%, cortex 12 +/- 14%, injury volume of ipsilateral structure; score 50 +/- 10). Mortality was greatest in the 24-hour drug treatment group. These data suggest that blockade of ET(A) receptor activity is not beneficial to tissue or functional outcomes from experimental stroke in cat.

Animals↗
PubMed2000

Overall cardiac functional effect of positive inotropic drugs with differing effects on relaxation.

Recent interest in so-called calcium-sensitizing positive inotropic drugs has highlighted the potential problem of a positive effect on force development being offset, at least partially, by the negative effect that many of these drugs have on relaxation. The purpose of this study was to examine the interplay of contraction and relaxation in determining the overall cardiac effect of different positive inotropic drugs. Using a buffer-perfused isolated rabbit heart preparation, we studied four drugs (calcium, dobutamine, EMD 57033, and CGP 48506) that were given at doses sufficient to increase similarly left ventricular pressure-generating capability by approximately 20%. We show that, even though they produce equivalent changes in pressure-generating capability, these four agents produce dissimilar changes in relaxation capability, with dobutamine speeding relaxation, EMD 57033 slowing relaxation, and calcium and CGP 48506 having little effect of relaxation. Similar relative effects were observed for drug-induced changes in the timing of pressure-generation events. These effects combine to produce different drug-induced changes in overall cardiac pump function judged by the relation between cardiac output and heart rate. Dobutamine shifted the maximal cardiac output to a higher heart rate. In contrast, both calcium sensitizers shifted the maximum in cardiac output to a lower heart rate, whereas calcium had no effect. Thus even though positive inotropic drugs may have similar effects on left ventricular pressure generation, the overall benefit of such drugs on ventricular pump function will depend on how the drug also affects ventricular relaxation and ejection capabilities.

Animals↗
PubMed2000

KORRIGAN, an Arabidopsis endo-1,4-beta-glucanase, localizes to the cell plate by polarized targeting and is essential for cytokinesis.

The formation of the cell plate, a unique structure in dividing plant cells, is pivotal for cytokinesis. A mutation in the Arabidopsis KORRIGAN (KOR) gene causes the formation of aberrant cell plates, incomplete cell walls, and multinucleated cells, leading to severely abnormal seedling morphology. The mutant, designed kor1-2, was identified as a stronger allele than the previously identified kor1-1, which appears to be defective only in cell elongation. KOR1 encodes an endo-1,4-beta-d-glucanase with a transmembrane domain and two putative polarized targeting signals in the cytosolic tail. When expressed in tobacco BY2 cells, a KOR1-GFP (green fluorescence protein) fusion protein was localized to growing cell plates. Substitution mutations in the polarized targeting motifs of KOR1 caused the fusion proteins to localize to the plasma membrane as well. Expression of these mutant genes in kor1-2 plants complemented only the cell elongation defect but not the cytokinesis defect, indicating that polarized targeting of KOR1 to forming cell plates is essential for cytokinesis. Our results suggest that KOR1 plays a critical role during cytokinesis.

Amino Acid Sequence↗
PubMed2000

Identification of G protein-coupled receptor kinase 2 phosphorylation sites responsible for agonist-stimulated delta-opioid receptor phosphorylation.

Agonist-induced receptor phosphorylation is an initial step in opioid receptor desensitization, a molecular mechanism of opioid tolerance and dependence. Our previous research suggested that agonist-induced delta-opioid receptor (DOR) phosphorylation occurs at the receptor carboxyl terminal domain. The current study was carried out to identify the site of DOR phosphorylation during agonist stimulation and the kinases catalyzing this reaction. Truncation (Delta15) or substitutions (T358A, T361A, and S363G single or triple mutants) at the DOR cytoplasmic tail caused 80 to 100% loss of opioid-stimulated receptor phosphorylation, indicating that T358, T361, and S363 all contribute and are cooperatively involved in agonist-stimulated DOR phosphorylation. Coexpression of GRK2 strongly enhanced agonist-stimulated phosphorylation of the wild-type DOR (WT), but Delta15 or mutant DOR (T358A/T361A/S363G) failed to show any detectable phosphorylation under these conditions. These results demonstrate that T358, T361, and S363 are required for agonist-induced and GRK-mediated receptor phosphorylation. Agonist-induced receptor phosphorylation was severely impaired by substitution of either T358 or S363 with aspartic acid residue, but phosphorylation of the T361D mutant was comparable with that of WT. In the presence of exogenously expressed GRK2, phosphorylation levels of T358D and S363D mutants were approximately half of that of WT, whereas significant phosphorylation of the T358/S363 double-point mutant was not detected. These results indicate that both T358 and S363 residues at the DOR carboxyl terminus are capable of serving cooperatively as phosphate acceptor sites of GRK2 in vivo. Taken together, we have demonstrated that agonist-induced opioid receptor phosphorylation occurs exclusively at two phosphate acceptor sites (T358 and S363) of GRK2 at the DOR carboxyl terminus. These results represent the identification of the GRK phosphorylation site on an opioid receptor for the first time and demonstrate that GRK is the prominent kinase responsible for agonist-induced opioid receptor phosphorylation in vivo.

Amino Acid Sequence↗
PubMed2000

Molecular cloning and mutagenesis of a DNA locus involved in lipooligosaccharide biosynthesis in Haemophilus somnus.

Haemophilus somnus undergoes antigenic and structural phase variation in its lipooligosaccharide (LOS). A gene (lob-1) containing repetitive 5'-CAAT-3' sequences that may, in part, contribute to phase variation was cloned and sequenced (T. J. Inzana et al., Infect. Immun. 65:4675-4681, 1997). We have now identified another putative gene (lob-2A) immediately upstream from lob-1. Lob-2A contained homology to several LOS biosynthesis proteins of the family Pasteurellaceae and the LgtB and LgtE galactosyltransferases of Neisseria meningitidis and N. gonorrhoeae. Unlike lob-1, lob-2A contained 18 to 20 5'-GA-3' repeats 141 bp upstream of the termination codon as determined by PCR amplification of DNA from individual colonies. Twenty repeats were most common, but when 19 5'-GA-3' repeats were present a stop codon would occur 1 bp after the last 5'-GA-3' repeat. A 630-bp SalI-BsgI fragment within lob-2A was deleted, and a kanamycin resistance (Km(r)) gene was inserted into this site to create pCAATDeltalob2A. Following electroporation of pCAATDeltalob2A into H. somnus 738, several allelic exchange mutants were isolated. The LOS electrophoretic profile of one mutant, strain 738-lob2A1::Km, was altered, and the phase variation rate was reduced but phase variation was not eliminated. A variant with 19 5'-GA-3' repeats in lob-2A had an LOS profile similar to that of 738-lob2A1::Km, suggesting that lob-2A was turned off in this phase. Nanoelectrospray mass spectrometry (nES-MS) and nuclear magnetic resonance spectroscopy showed that 738-lob2A1::Km was deficient in the terminal betaGal(1-3)betaGlcNAc residue present in parent strain 738. Mutant 738-lob2A1::Km was significantly more sensitive to the bactericidal action of normal bovine serum and was less virulent in mice than was parent strain 738. When H. somnus 129Pt was electrotransformed with shuttle vector pLS88 containing lob-2A, its LOS electrophoretic profile was modified and additional N-acetylhexosamine residues were present, as determined by nES-MS analysis. These results indicated that lob-2A may be an N-acetylglucosamine transferase involved in LOS biosynthesis and phase variation and that LOS structure is important to H. somnus virulence.

Animals↗
PubMed2000

Identification and characterization of genes required for early Myxococcus xanthus developmental gene expression.

Starvation and cell density regulate the developmental expression of Myxococcus xanthus gene 4521. Three classes of mutants allow expression of this developmental gene during growth on nutrient agar, such that colonies of strains containing a Tn5 lac Omega4521 fusion are Lac(+). One class of these mutants inactivates SasN, a negative regulator of 4521 expression; another class activates SasS, a sensor kinase-positive regulator of 4521 expression; and a third class blocks lipopolysaccharide (LPS) O-antigen biosynthesis. To identify additional positive regulators of 4521 expression, 11 Lac(-) TnV. AS transposon insertion mutants were isolated from a screen of 18,000 Lac(+) LPS O-antigen mutants containing Tn5 lac Omega4521 (Tc(r)). Ten mutations identified genes that could encode positive regulators of 4521 developmental expression based on their ability to abolish 4521 expression during development in the absence of LPS O antigen and in an otherwise wild-type background. Eight of these mutations mapped to the sasB locus, which encodes the known 4521 regulators SasS and SasN. One mapped to sasS, whereas seven identified new genes. Three mutations mapped to a gene encoding an NtrC-like response regulator homologue, designated sasR, and four others mapped to a gene designated sasP. One mutation, designated ssp10, specifically suppressed the LPS O-antigen defect; the ssp10 mutation had no effect on 4521 expression in an otherwise wild-type background but reduced 4521 developmental expression in the absence of LPS O antigen to a level close to that of the parent strain. All of the mutations except those in sasP conferred defects during growth and development. These data indicate that a number of elements are required for 4521 developmental expression and that most of these are necessary for normal growth and fruiting body development.

Bacterial Proteins↗
PubMed2000

Sialylation of the host receptor may modulate entry of demyelinating persistent Theiler's virus.

Theiler's murine encephalomyelitis virus (TMEV) is a picornavirus of the Cardiovirus genus. Certain strains of TMEV may cause a chronic demyelinating disease, which is very similar to multiple sclerosis in humans, associated with a persistent viral infection in the mouse central nervous system (CNS). Other strains of TMEV only cause an acute infection without persistence in the CNS. It has been shown that sialic acid is a receptor moiety only for the persistent TMEV strains and not for the nonpersistent strains. We report the effect of sialylation on cell surface on entry and the complex structure of DA virus, a persistent TMEV, and the receptor moiety mimic, sialyllactose, refined to a resolution of 3.0 A. The ligand binds to a pocket on the viral surface, composed mainly of the amino acid residues from capsid protein VP2 puff B, in the vicinity of the VP1 loop and VP3 C terminus. The interaction of the receptor moiety with the persistent DA strain provides new understanding for the demyelinating persistent infection in the mouse CNS by TMEV.

Animals↗
PubMed2000

Altered L-type Ca(2+) channel currents in vascular smooth muscle cells from experimental diabetic rats.

Vascular complications of diabetes are associated with abnormal Ca(2+) handling by vascular smooth muscle cells (SMCs) in which the alteration in L-type voltage-dependent Ca(2+) channel (VDCC) currents may play an important role. In the present study, the characteristics of L-type VDCC currents in tail artery SMCs from streptozotocin-induced diabetic rats were examined. The densities, but not the voltage dependence, of L-type VDCC currents were reduced as diabetes progressed from 1 wk to 3 mo. The inhibitory effect of dibutyryl-cAMP on L-type VDCC currents was greater in diabetic SMCs than in age-matched control cells (P < 0.01). Both the stimulatory effect of BAY K 8644 and the inhibitory effect of nifedipine on L-type VDCC currents were significantly enhanced in diabetic cells. The diabetes-related abnormalities in L-type VDCC currents were mimicked by culturing SMCs with a high concentration of glucose. Our results suggest that the properties of L-type VDCC in diabetic vascular SMCs were significantly altered, partially related to the increased L-type VDCC sensitivity to cAMP and hyperglycemia.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗