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Biomedical subjects

R D Thompson

Publications and source records attributed to R D Thompson.

At least 19 recordsLinked to original sources
PubMed2001

Platelet-endothelial cell adhesion molecule-1 (PECAM-1)-deficient mice demonstrate a transient and cytokine-specific role for PECAM-1 in leukocyte migration through the perivascular basement membrane.

Studies with neutralizing antibodies have indicated roles for platelet-endothelial cell adhesion molecule-1 (PECAM-1) in leukocyte migration through the endothelium and the perivascular basement membrane. Because some of these findings have been contentious, this study aimed to explore the role of PECAM-1 in leukocyte migration by analyzing leukocyte responses in interleukin 1beta (IL-1beta)- and tumor necrosis factor-alpha (TNFalpha)-activated cremasteric venules of PECAM-1-deficient mice using intravital and electron microscopy. Although no differences in levels of leukocyte rolling flux or firm adhesion were observed, a delay in leukocyte transmigration in response to IL-1beta, but not TNFalpha, was detected in PECAM-1-deficient mice. Electron microscopy indicated that this delay occurred at the level of perivascular basement membrane. To address the cytokine specificity of PECAM-1 dependence, in vitro experiments demonstrated that TNFalpha, but not IL-1beta, could induce rapid adhesion of murine neutrophils to protein-coated surfaces, suggesting that TNFalpha elicited leukocyte transmigration in wild-type mice via direct stimulation of leukocytes. In summary, the results suggest a regulatory role for PECAM-1 in leukocyte migration through the perivascular basement membrane, a role that appears to be cytokine-specific and associated with the ability of the cytokine to stimulate rapid neutrophil adhesion.

Animals↗
PubMed2001

A transcript encoding a nucleic acid-binding protein specifically expressed in maize seeds.

A cDNA clone has been obtained for a low-abundance, seed-specific mRNA that encodes a polypeptide which defines a novel family of plant proteins with some similarities to the DnaJ class of molecular chaperones. The MEM1 (Maize Endosperm Motif binding protein) protein is capable of binding to the endosperm motif and activating transcription in the yeast one-hybrid system. Recombinant MEM1 was shown to bind in vitro to nucleic acids, with a preference for RNA over DNA. MEM1 is capable of forming homodimers, a property that is dependent on a domain close to the C-terminus of the protein. The protein is expressed in mid- to late-term endosperm cells. Subcellular fractionation and size fractionation under non-denaturing conditions indicate that the protein is present in the cytosol of endosperm cells. Possible roles of MEM1 in endosperm and protein body development are discussed.

Amino Acid Sequence↗
PubMed2001

Maize endosperm secretes a novel antifungal protein into adjacent maternal tissue.

A series of endosperm transfer layer-specific transcripts has been identified in maize by differential screening of a cDNA library of transcripts at 10 days after pollination. Sequence comparisons revealed among this class of cDNAs a novel, small gene family of highly diverged sequences encoding basal layer antifungal proteins (BAPs). The bap genes mapped to two loci on chromosomes 4 and 10. So far, bap-homologous sequences have been detected only in maize, teosinte and sorghum, and are not present in grasses outside the Andropogoneae tribe. BAP2 is synthesized as a pre-proprotein, and is processed by successive removal of a signal peptide and a 29-residue prodomain. The proprotein can be detected exclusively in microsomal membrane-containing fractions of kernel extracts. Immunolocalization reveals BAP2 to be predominantly located in the placentochalazal cells of the pedicel, adjacent to the basal endosperm transfer layer (BETL) cells, although the BAP2 transcript is found only in the BETL cells. The biological roles of BAP2 propeptide and mature peptide have been investigated by heterologous expression of the proprotein in Escherichia coli, and by tests of its fungistatic activity and that of the fully processed form in vitro. The mature BAP2 peptide exhibits potent broad-range activity against a range of filamentous fungi, including several plant pathogens.

Amino Acid Sequence↗
PubMed2000

Divergent effects of platelet-endothelial cell adhesion molecule-1 and beta 3 integrin blockade on leukocyte transmigration in vivo.

The final stage in the migration of leukocytes to sites of inflammation involves movement of leukocytes through the endothelial cell layer and the perivascular basement membrane. Both platelet-endothelial cell adhesion molecule-1 (PECAM-1/CD31) and the integrin alphavbeta3 have been implicated in this process, and in vitro studies have identified alphavbeta3 as a heterotypic ligand for PECAM-1. In the present study we have addressed the roles of these molecules by investigating and comparing the effects of PECAM-1 and alphavbeta3 blockade on leukocyte migration in vivo. For this purpose we have examined the effects of neutralizing Abs directed against PECAM-1 (domain 1-specific, mAb 37) and beta3 integrins (mAbs 7E3 and F11) on leukocyte responses in the mesenteric microcirculation of anesthetized rats using intravital microscopy. The anti-PECAM-1 mAb suppressed leukocyte extravasation, but not leukocyte rolling or firm adhesion, elicited by IL-1beta in a dose-dependent manner (e.g., 67% inhibition at 10 mg/kg 37 Fab), but had no effect on FMLP-induced leukocyte responses. Analysis by electron microscopy suggested that this suppression was due to an inhibition of neutrophil migration through the endothelial cell barrier. By contrast, both anti-beta3 integrin mAbs, 7E3 F(ab')2 (5 mg/kg) and F11 F(ab')2 (5 mg/kg), selectively reduced leukocyte extravasation induced by FMLP (38 and 46%, respectively), but neither mAb had an effect on IL-1beta-induced leukocyte responses. These findings indicate roles for both PECAM-1 and beta3 integrins in leukocyte extravasation, but do not support the concept that these molecules act as counter-receptors in mediating leukocyte transmigration.

Abciximab↗
PubMed2000

rgf1, a mutation reducing grain filling in maize through effects on basal endosperm and pedicel development.

The maize cob presents an excellent opportunity to screen visually for mutations affecting assimilate partitioning in the developing kernel. We have identified a defective kernel mutant termed rgf1, reduced grain filling, with a final grain weight 30% of the wild type. In contrast with most defective endosperm mutants, rgf1 shows gene dosage-dependent expression in the endosperm. rgf1 kernels possess a small endosperm incompletely filling the papery pericarp, but embryo development is unaffected and the seeds are viable. The mutation conditions defective pedicel development and greatly reduces expression of endosperm transfer layer-specific markers. rgf1 exhibits striking morphological similarities to the mn1 mutant, but maps to a locus approximately 4 cM away from mn1 on chromosome 2 of maize. Despite reduced starch accumulation in the mutant, no obvious lesion in starch biosynthesis has been detected. Free sugar levels are unaltered in rgf1 endosperm. Rates of sugar uptake, measured over short (8 h) periods in cultured kernels, are increased in rgf1 compared to the wild type. rgf1 and wild-type kernels, excised at 5 DAP and cultured in vitro also develop differently in response to variations in sugar regime: glucose concentrations above 1% arrest placentochalazal development of rgf1 kernels, but have no effect on cultured wild-type kernels. These findings suggest that either uptake or perception of sugar(s) in endosperm cells at 5-10 DAP determines the rgf1 kernel phenotype.

Base Sequence↗
PubMed2000

Analyte loss due to membrane filter adsorption as determined by high-performance liquid chromatography.

The phenomenon of membrane filter adsorption in high-performance liquid chromatography (HPLC) is investigated utilizing 16 brands of filters representing 3 polymeric materials: cellulose acetate (CA), nylon, and polyvinylidene difluoride in a variety of diameters (3, 4, 7, 13, and 25 mm). Sixteen compounds commonly encountered in drug preparations are selected as sample analytes and classified as acidic, basic, and neutral in chemical behavior. Six mobile phase/sample solvent mixtures are included: 3 with methanol-water and 3 with acetonitrile-water as major constituents. When using methanol as the mobile phase organic component, CA, nylon, and polyvinylidene difluoride (PVDF) filters exhibit negligible to moderate adsorption levels with regard to the neutral and basic drug compounds. The acidic drug test compounds are adsorbed by 50% of all 3 filter materials tested in methanol-water. In acetonitrile, neutral compounds are affected by 31.4%, basic compounds are affected by 47.0%, and acidic compounds are affected by 53.6% of the nylon and PVDF filters. CA is incompatible with acetonitrile and is excluded from the study with this solvent.

Absorption↗
PubMed1999

Evidence for factors regulating transfer cell-specific expression in maize endosperm.

In maize, a layer of basal endosperm cells adjacent to the pedicel is modified for a function in solute transfer. Three genes specifically expressed in this region, termed the basal endosperm transfer layer (BETL-2 to -4), were isolated by differential hybridization. BETL-2 to -4 are coordinately expressed in early and mid-term endosperm development, but are absent at later stages. BETL-2 to -4 coding sequences all predict small (< 100 amino acids), secreted, cysteine-rich polypeptides which lack close relatives in current database accessions. BETL-3 and BETL-1 display some sequence similarities with each other and to plant defensins. BETL-2 to -4 promoter regions were isolated and compared, revealing the presence of a promoter-proximal microsatellite repeat as the most highly conserved sequence element in each sequence. Electrophoretic mobility shift assays (EMSA) showed that specific BETL-2 to -4 promoter fragments competed for binding to the same DNA-binding activity in nuclear extracts prepared from maize endosperm. Although BETL-2 to -4 are only expressed in basal endosperm cells, the DNA-binding activities detected were of two types: distal endosperm-specific, or present in both basal and distal endosperm extracts. On the basis of these findings, a model to account for the coordinate regulation of BETL genes in endosperm cells is proposed.

Amino Acid Sequence↗
PubMed1999

Sensitivity and specificity of radioisotope right-left shunt measurements and pulse oximetry for the early detection of pulmonary arteriovenous malformations.

STUDY OBJECTIVES: To assess the effectiveness of pulse oximetry and radioisotope measurement of right-to-left (R-L) shunt for the early detection of pulmonary arteriovenous malformations (PAVMs) in patients with hereditary hemorrhagic telangiectasia (HHT). DESIGN: Patients with HHT had serial measurements of the following: (1) arterial oxygen saturation (SaO2) by pulse oximetry in erect and supine positions, and on maximal exercise using cycle ergometry; (2) quantitative radioisotope measurements of R-L shunt using IV 99mTc-labeled macroaggregates of albumin; and (3) routine pulmonary function. After percutaneous transcatheter embolization of all PAVMs with feeding vessel diameters > 3 mm, residual PAVMs were assessed with selective digital subtraction pulmonary angiography. Using postembolization angiography as the "gold standard," SaO2 and radioisotope shunt measurements after embolization were analyzed retrospectively using logistic regression to assess the ability of each test to predict for the presence of residual PAVMs. RESULTS: Of the 66 patients included, 40 had small PAVMs remaining postembolization. Using univariate logistic regression, radioisotope shunt and erect saturation showed a significant relationship with the presence of residual PAVMs (p=0.001, 0.005, respectively). Erect SaO2 < or = 96% had 73% sensitivity and 35% specificity for detecting PAVMs. Radioisotope shunt >3.5% of cardiac output had 87% sensitivity and 61% specificity for detecting PAVMs. CONCLUSIONS: These results confirm that noninvasive measurements are useful in the screening of patients with HHT for the presence of PAVMs without need for angiography and its associated risks, and that radionuclide scanning is better than pulse oximetry.

Angiography, Digital Subtraction↗
PubMed1998

Multiple elements of the S2-RNase promoter from potato (Solanum tuberosum L.) are required for cell type-specific expression in transgenic potato and tobacco.

A functional analysis of the promoter of the S2-RNase gene from potato was performed in transgenic potato and tobacco plants, using a deletion series of S2-RNase promoter GUS fusions. A detailed histochemical and quantitative analysis of the transgenic tobacco plants revealed that S2 promoter fragments ranging in size from 5.6 kb in length down to 0.2 kb mediate a weak developmentally regulated expression in the pistil, and strong ectopic expression in pollen. In the pistil, different expression patterns were seen depending on the transformant, the predominant one being characterized by expression in the stigma and the transmitting tract of the style, whereas a few plants showed expression exclusively either in the stigma or in the stylar transmitting tissue. All transformants also showed GUS expression in the placental epidermis of the ovary. Two sequences that are conserved between the potato S1-RNase and S2-RNase promoters, termed motif and motif III, are located in a fragment of the S2 promoter extending from position of -200 to bp -100, and motif II, located between by -498 and -480, was identified on the basis of sequence comparisons between pistil-specific promoters. Motif II was found to be dispensible for pistil-specific and for pollen-specific expression. Two submotifs, A and B, were identified with the motif I. Both were essential for expression in the pistil but only B was necessary for expression in pollen. Although motif III has a similar bipartite structure and sequence to motif I, it was not sufficient to confer-either pollen- or pistil-specific expression. However, deletion of motif III abolished pollen-specific expression in transient expression experiments, suggesting that an interaction between the two sequence motifs may be needed to specify cell type-specific expression. In transgenic potato the S2-RNase promoter also mediates expression in pollen and in the pistil; however, significantly fewer plants showed expression than in tobacco, with most plants also exhibiting GUS expression in other issues.

Enhancer Elements, Genetic↗
PubMed1998

Epidemiological study of Guillain-Barré syndrome in south east England.

OBJECTIVES: To determine the incidence, treatment, and outcome of Guillain-Barré syndrome in south east England. METHODS: Patients presenting with confirmed Guillain-Barré syndrome between 1 July 1993 and 30 June 1994 were recruited via a voluntary reporting scheme coordinated by the British Neurological Surveillance Unit, hospital activity data collected from acute admitting hospitals within the South East and South West Thames Regional Health Authorities, death certificates, and a contemporary research study of Guillain-Barré syndrome and Campylobacter jejuni infection. All patients were followed up for one year to determine outcome. RESULTS: Seventy nine patients were recruited, 35 (44%) male, 44 (56%) female, including three children (two boys, one girl). The crude (95% confidence interval (95% CI)) annual incidence was 1.2 (0.9-1.4) cases/100000 population and 1.5 (1.3-1.8)/100000 when adjusted for undetected cases. Twenty (25%) patients required ventilation for an average (SD) of 42 (64) days. Thirty six (46%) patients received intravenous human immunoglobulin, five (6%) received plasma exchange, 11 (14%) both treatments, three (4%) steroids, and 25 (32%) no immunomodulatory treatment. One year later, six patients (8%) had died, all of whom were older than 60, three (4%) remained bedbound or ventilator dependent, seven (9%) were unable to walk unaided, 14 (17%) were unable to run, and 49 (62%) had made a complete or almost complete recovery. Increasing age was significantly associated with a poorer outcome at one year. CONCLUSIONS: Despite the frequent use of modern immunomodulatory treatments Guillain-Barré syndrome still carries considerable morbidity and mortality.

Adolescent↗
PubMed1997

The activation domain of the maize transcription factor Opaque-2 resides in a single acidic region.

The maize (Zea mays L.) endosperm specific transcription factor, encoded by the Opaque-2(O2) locus, functions in vivo to activate transcription from its target promoters.O2 regulates the expression of a major storage protein class, the 22 kDa zeins, and of a type I ribosome inactivating protein, b-32, during maturation phase endosperm development. The coding sequence of O2, which indicates it to be a member of the basic region-leucine zipper (bZIP) class of DNA-binding proteins, contains a number of regions rich in either proline or acidic residues which are candidates for activation domains. In functional assays using tobacco mesophyll protoplasts, the level of transactivation conferred by a series of O2-deletion constructs was tested using as a reporter a fusion of the b-32 target promoter to beta-glucuronidase (GUS). The results indicate that O2 has a single acidic activation domain, located near the N-terminus of the protein (amino acids 41-91). The ability of a shorter part of this domain (amino acids 39-82) to confer transactivation was also demonstrated in domain swapping experiments, using fusions of the O2 polypeptide sequence to the DNA-binding domain of the parsley (Petroselinum crispum) transcription factor CPRF1.

DNA-Binding Proteins↗
PubMed1997

Lateralization of affective prosody in brain and the callosal integration of hemispheric language functions.

Although affective prosody appears to be a dominant function of the right hemisphere, its degre of lateralization has not yet been established since various publications have reported affective-prosodic deficits following left brain damage in association with aphasia. This paper explores the mechanisms underlying affective-prosodic deficits following left and right brain damage by testing the ability of subjects to repeat and comprehend affective prosody under progressively reduced verbal-articulatory conditions. The results demonstrate that reducing verbal-articulatory conditions robustly improves the performance of left but not right brain damaged patients, a finding that supports the supposition that affective prosody is strongly lateralized to the right hemisphere. However, the performance of left brain damaged patients was not correlated to the presence, severity, or type of aphasic deficit(s). Based on functional-anatomic correlations for spontaneous affective prosody and affective-prosodic repetition, deep white matter lesions located below the supplementary motor area that disrupt interhemispheric connections coursing through the mid-rostral corpus callosum may contribute to affective-prosodic deficits that are both additive and independent of any aphasic deficits. In light of these and other findings, various anatomical, functional, and maturational hierarchic relationships between the affective-prosodic and verbal-linguistic aspects of language are posited in order to help further explain discrepancies that exist in the literature regarding the neurology of affective prosody.

Adult↗
PubMed1997

A promoter directing high level expression in pistils of transgenic plants.

The promoter of the potato (Solanum tuberosum L.) SK2 gene, encoding a pistil-specific basic endochitinase, was cloned. Various fragments of the SK2-promoter, from 1 kb down to 0.23 kb in length, were fused to the GUS reporter gene. Chimaeric SK2 promoter-GUS fusion constructs were transformed into potato by Agrobacterium tumefaciens-mediated transformation. The SK2-GUS transgenic potato plants exhibited a highly specific GUS activity in the pistil. Expression in the pistil was shown to be developmentally regulated. In addition to the GUS activity in pistils, transgenic plants also showed a much weaker ectopic expression in anthers. In other tissues no systematic expression was detectable. All SK2 promoter fragments analysed conferred pistil-specific expression without significant qualitative or quantitative differences, demonstrating that the regulatory elements mediating this expression pattern are located within a 230 bp SK2 promoter fragment. The SK2 promoter may be used to engineer high levels of expression in pistils of transgenic plants.

Chitinases↗
PubMed1997

Nitrogen and hormonal responsiveness of the 22 kDa alpha-zein and b-32 genes in maize endosperm is displayed in the absence of the transcriptional regulator Opaque-2.

The maize (Zea mays L.) b-ZIP transcriptional activator Opaque-2(O2) regulates the synthesis of major endosperm proteins. In the o2 homozygote, 22 kDa zein prolamins and the b-32 ribosome-inactivating protein are greatly reduced in level. An in vitro endosperm culture system has been studied in which o2 endosperm synthesizes 22 kDa zein and b-32 in response to nitrogen supplements. An increase in 22 kDa zein mRNA concentration is also seen, implying an effect at the level of transcription or differential RNA turnover. The nitrogen-dependent induction of 22 kDa zein synthesis in cultured o2 endosperm was further investigated by analysing transient expression of reporter constructs. The highest response to nitrogen was exhibited by the intact 22 kDa zein promoter. Removal of individual O2 binding sites either reduced or increased overall promoter activity, but always decreased the nitrogen-dependent stimulation of activity. This effect was observed equally in wild-type and o2 mutant endosperm. It is concluded that a factor other then O2 is responsible for activating the 22 kDa zein promoter under high-nitrogen culture conditions. Despite its occurrence in the absence of O2 protein, the nitrogen response is mediated through binding at O2 binding sites. An induction of 22 kDa zein and b-32 synthesis in cultured o2 endosperm could also be achieved on nitrogen-free media by the addition of abscisic acid or methyl jasmonate.

Abscisic Acid↗
PubMed1996

The role of multiple binding sites in the activation of zein gene expression by Opaque-2.

Opaque-2 (O2) encodes a transcriptional activator of the basic domain-leucine zipper (bZIP) class, which controls the expression level in maize endosperm of the 22kD alpha-zeins and a number of non-storage proteins. The interaction of the O2 protein at three clustered binding sites on an isolated 22 kD zein gene promoter has been investigated. O2 is shown to transactivate transcription from these sites in tobacco mesophyll protoplasts as well as in maize endosperm cells transformed by particle bombardment. The binding sites have been mutated by base exchanges, singly or in different combinations, to determine their contribution to transactivation in vivo in both the leaf protoplast and the maize endosperm system. The effect of these mutations on binding of O2 in vitro was determined by electrophoretic mobility shift assays (EMSA), using O2 protein expressed in E. coli. Two of the sites seemed to be equally effective in responding to Opaque-2 in vivo in both cell types, although one of them does not contain an ACGT core sequence, and has a lower affinity for O2 in vitro than the ACGT-containing binding site. A third site, which has the lowest affinity of all three, confers no detectable O2-dependent promoter activation alone, but significantly increases activation in combination with either one of the other sites. Hence, weaker O2 binding sites can still mediate major O2-dependent effects when present in target promoters in vivo.

Binding Sites↗
PubMed1996

Regulation of cytosolic pyruvate, orthophosphate dikinase expression in developing maize endosperm.

Pyruvate orthophosphate dikinase (PPDK, E.C. 2.7.9.1) is an abundant enzyme in the leaves of C4 plants associated with the dicarboxylic acid pathways of CO2 fixation in the dark. PPDK activity has also been detected in the seeds of maize and other, non-C4 cereals, where its role has yet to be established. Using an anti-PPDK serum, two cross-reacting species of M(r) close to 90 000 were detected in developing maize endosperm of wild-type plants. In two independent opaque-2 mutant lines, one of the polypeptides was absent and the other was reduced in level. Similarly, endosperm PPDK mRNA levels were greatly reduced in the opaque-2 maize lines compared to wild type, suggesting that endosperm PPDK gene expression is under Opaque-2 control. However, a low level of PPDK mRNA could still be detected in these mutants, indicating that PPDK gene expression is not absolutely dependent on Opaque-2 but rather can be modulated by it. This interpretation was reinforced by the demonstration that the distribution of PPDK transcripts is not affected in o2 mutants, although the level is reduced, and that PPDK mRNA is detectable prior to 02 mRNA during the maturation of wild-type maize endosperm. Using oligonucleotides specific for the different maize PPDK genes, the o2 mutations were shown to affect only cyPPDKZml gene expression in maize line A69Y.

Base Sequence↗