Biomedical subjects

J Armstrong

Publications and source records attributed to J Armstrong.

At least 55 records · Page 3Linked to original sources

PubMed2000

Sequence and analysis of chromosome 5 of the plant Arabidopsis thaliana.

The genome of the model plant Arabidopsis thaliana has been sequenced by an international collaboration, The Arabidopsis Genome Initiative. Here we report the complete sequence of chromosome 5. This chromosome is 26 megabases long; it is the second largest Arabidopsis chromosome and represents 21% of the sequenced regions of the genome. The sequence of chromosomes 2 and 4 have been reported previously and that of chromosomes 1 and 3, together with an analysis of the complete genome sequence, are reported in this issue. Analysis of the sequence of chromosome 5 yields further insights into centromere structure and the sequence determinants of heterochromatin condensation. The 5,874 genes encoded on chromosome 5 reveal several new functions in plants, and the patterns of gene organization provide insights into the mechanisms and extent of genome evolution in plants.

Animals↗

PubMed2000

Radiation induces upregulation of cyclooxygenase-2 (COX-2) protein in PC-3 cells.

PURPOSE: To investigate the impact of gamma-irradiation on cyclooxygenase-2 (COX-2) expression and its enzymatic activity in PC-3 cells. Cell cycle redistribution, viability, and apoptosis were quantitated in control and irradiated cells with or without the COX-2 inhibitor NS-398. METHODS AND MATERIALS: Western blot analysis was used to assess COX-2 protein expression. Prostaglandin (PGE(2)) was measured after addition of arachidonic acid (AA) using a Monoclonal Immunoassay Kit. Cell cycle and apoptosis were assessed using flow cytometry. RESULTS: We observed a dose-dependent increase in COX-2 of 37.0%, 79.7%, and 97.5% following irradiation with 5, 10, and 15 Gy, respectively. The PGE(2) level of irradiated cells was higher than in controls (1512 +/- 157.5 vs. 973.7 +/- 54.2 rhog PGE(2)/mL; p < 0.005, n = 4) while cells irradiated in the presence of NS-398 had reduced PGE(2) levels (218.8 +/- 80.1 rhog PGE(2)/mL; p < 0.005; n = 4). We found no differences in cell cycle distribution or apoptosis between cells irradiated in the presence or absence of NS-398. CONCLUSIONS: COX-2 protein is upregulated and enzymatically active after irradiation, resulting in elevated levels of PGE(2). This effect can be suppressed by NS-398, which has clinical implications for therapies combining COX-2 inhibitors with radiation therapy.

Apoptosis↗

PubMed2000

Membrane traffic between genomes.

Proteins of the Rab and SNARE families target vesicles to their intracellular destinations. A comparison of these families from the budding yeast, fission yeast, nematode and fruitfly genomes has implications for the organization of membrane traffic in different organisms.

Animals↗

PubMed2000

In vitro evaluation of c7E3-Fab (ReoPro) eluting polymer-coated coronary stents.

OBJECTIVE: Stent thrombosis and in-stent restenosis remain problematic in certain patient sub-groups. c7E3-Fab (ReoPro, abciximab) inhibits the platelet glycoprotein IIb/IIIa receptor as well as the smooth muscle cell alpha(v)beta3 receptor, and thus may influence both processes, especially if high local concentrations could be achieved. We have studied the adsorption and elution characteristics of c7E3-Fab on commercially available polymer-coated stents. We have also investigated the effect of such antibody binding on platelet deposition in vitro, and on antibody deposition into ex vivo human saphenous vein wall to assess whether such stents may influence stent thrombosis and restenosis. METHODS AND RESULTS: Adsorption was measured using a radioisotope technique after immersing segments of polymer-coated stents in c7E3-Fab solutions. Uptake was dependent on antibody concentration and duration of immersion of wire in the solution. After 22 h (at 5 mg ml(-1)), 1146+/-101 ng cm(-1) wire was adsorbed. In an in vitro perfusion circuit, the antibody eluted slowly, with 53% remaining after 12 days washing. To determine the value that such stents might have in clinical practise, adsorption to balloon-mounted stents was assessed at room temperature, using commercially available c7E3-Fab (2 mg ml(-1)). Efficacy of eluting c7E3-Fab was determined by measuring deposition of 111-Indium platelets. Immersing stents in c7E3-Fab for 20 min inhibited platelet deposition by 82.3% compared to controls (P=0.018). Deployment of treated stents in ex vivo saphenous vein resulted in the deposition of c7E3-Fab in the intima and media. CONCLUSIONS: c7E3-Fab can be passively adsorbed onto polymer-coated stents. It elutes slowly and in a predictable manner, significantly inhibiting platelet deposition in vitro. These studies pave the way to developing stent-based delivery of a potent anti-platelet agent that may additionally affect smooth muscle cell activity.

Abciximab↗

PubMed2000

The impact of three-dimensional radiation on the treatment of non-small cell lung cancer.

PURPOSE: Non-small cell lung cancer (NSCLC) patients with locally advanced unresectable disease have a grim prognosis. Radiotherapeutic strategies are necessary to improve the permanent eradication of thoracic disease. The poor results achieved with conventional external beam radiation therapy reflect in part, the inadequacy of such therapy in achieving its primary objective of achieving local control. The impact of three-dimensional conformal radiation therapy (3-DCRT) on local disease eradication and its potential role in improving survival is assessed. DESIGN: This review addresses aspects of the software and hardware technology of 3-DCRT, the clinical and technical aspects of target volume definition, the use of 3-DCRT to predict radiation pneumonitis, strategies for dose escalation in NSCLC, and analyses the clinical results to date. RESULTS: Initially investigators compared the best treatment techniques devised with conventional planning techniques to those devised with 3-DCRT. These analyses showed that 3-DCRT had the potential to deliver high dose radiation (>70 Gy) with minimal underdosing and with a concomitant relative sparing of normal tissues. This technical demonstration of enhanced therapeutic ratio is the basis for the evolving clinical utilization of 3-DCRT for NSCLC. Software and hardware developments continue to develop and have the potential to solve evolving clinical issues. Dose-volume-histograms have been used to accurately quantify lung dose and derived parameters have the potential to predict the risk of pneumonitis for individual patients before treatment. Initial clinical results have been promising and strategies for further dose escalation are emerging. CONCLUSION: Preliminary experience has resulted in promising survival following three-dimensional conformal radiation therapy alone for locally advanced NSCLC. More follow-up and experience will determine late toxicity, maximum dose, and efficacy of dose escalation with three-dimensional conformal radiation therapy. Strategies should be developed to integrate this modality into the combined treatment of locally advanced non-small cell lung cancer.

Carcinoma, Non-Small-Cell Lung↗

PubMed2000

Mutation analysis of 16S rRNA in patients with Rett syndrome.

Rett syndrome (RTT) is a progressive neurodevelopmental disorder that affects one in 10,000-15,000 females. RTT is mainly sporadic; familial cases have an estimated frequency of less than 1%. Before the recent identification of de novo dominant mutations in the X-linked MECP2 gene, many other hypotheses had been proposed to explain the particular pattern of inheritance and the phenotypic expression of the disease. The involvement of mitochondrial DNA had been investigated because of the structural and functional mitochondrial abnormalities evident in the patients. In 1997 the finding of mutations at 16S rRNA in several affected RTT females and their mothers was reported, suggesting that mitochondrial DNA might play a key role in the pathogenesis of RTT. To investigate the relevance of such mutations, we used the same methodologic approach to analyze RTT mitochondrial DNA in our series. No 16S rRNA alterations were evident in 27 Spanish patients with classic RTT.

Adolescent↗

PubMed2000

How do Rab proteins function in membrane traffic?

The Rabs are a group of GTP-binding proteins implicated for some time in the targeting of different transport vesicles within the cell, but it has been unclear how they function, or how they relate to a second group of targeting proteins, the SNAREs. Recent work, discussed in this review, has used biophysical, biochemical and genetic approaches to begin to answer these questions for Rab3, Rab5 and the yeast protein Sec4p. However, the results from these three Rabs lead to a surprising conclusion: the different Rabs seem to function via highly diverse target proteins.

Fungal Proteins↗

PubMed2000

A panel of monoclonal antibodies targeting the rabies virus phosphoprotein identifies a highly variable epitope of value for sensitive strain discrimination.

A recombinant rabies virus phosphoprotein fusion product (GST-P) was used to generate a series of monoclonal antibodies (MAbs) with anti-P reactivity. Competitive binding assays classified 27 of these MAbs into four groups (I to IV), and 24 of them were deemed to recognize linear epitopes, as judged by their reaction in immunoblots. The linear epitope recognized in each case was mapped by using two series of N- and C-terminally deleted recombinant phosphoproteins. Assessment of the reactivities of representative MAbs to a variety of lyssavirus isolates by an indirect fluorescent antibody test indicated that group I MAbs, which recognized a highly conserved N-terminal epitope, were broadly cross-reactive with all lyssaviruses assayed, while group III MAbs, which reacted with a site overlapping that of group I MAbs, exhibited variable reactivities and group IV MAbs reacted with most isolates of genotypes 1, 6, and 7 only. In contrast, group II MAbs, which recognized an epitope located within a highly divergent central portion of the protein, were exquisitely strain specific. These anti-P MAbs are potentially useful tools for lyssavirus identification and discrimination.

Animals↗

PubMed2000

Alterations in cardiac adrenergic terminal function and beta-adrenoceptor density in pacing-induced heart failure.

Congestive heart failure is associated with cardiac adrenergic nerve terminal changes and beta-adrenoceptor density downregulation. To study the temporal sequence of these changes, we performed studies in rabbits at 2, 4, and 8 wk of cardiac pacing (360 beats/min) and at 1, 2, and 4 wk after cessation of pacing. Rapid pacing produced left ventricular (LV) dysfunction and an increase in plasma norepinephrine (NE) in 1-2 wk. At week 2, NE uptake activity, NE uptake-1 density, and adenylyl cyclase responses to isoproterenol, 5'-guanylyl imidodiphosphate [Gpp(NH)p], and forskolin reduced. However, immunostained tyrosine hydroxylase profile, beta-adrenoceptor density, and NE histofluorescence did not reduce until 4-8 wk of pacing. After cessation of cardiac pacing, LV function normalized quickly, followed by return of tyrosine hydroxylase and NE profiles in 1 wk and adenylyl cyclase responses to agonists and NE uptake activity in 2 wk. Myocardial beta-adrenoceptor density returned to normal by 4 wk after cessation of pacing. Our results suggest that there is no permanent structural neuronal damage in the myocardium within the first 8 wk of rapid cardiac pacing. Abnormal myocardial NE reuptake mechanism may play an important pathophysiological role in heart failure.

Adenylyl Cyclases↗

PubMed2000

Treatment margins for prostate brachytherapy.

The purpose of this article was to determine what planned treatment margin (TM) would allow for implant-related prostate volume changes and still achieve an adequate periprostatic cancercidal dose. Twenty consecutive, unselected patients who underwent (125)I implantation (144 Gy prescription dose) were studied. The treated volume (TV) was calculated as the volume encompassed by the 144 Gy isodose distribution. A post-implant computed tomography scan was obtained the following day, using 5-mm images at every 5 mm. The distances between the prostate margin (GTV) and TV were determined by measuring the distance between the ultrasound-defined prostatic margin and the prescription isodose, perpendicular to the prostatic margin. The lateral, anterior, and posterior TM margins were determined at the base, mid-level, and apex of the prostate. The pre-implant TV was nearly twice as large as the GTV, ranging from 36 to 199 mL (median, 73 mL). The anterior, lateral, and posterior planned TMs varied substantially between patients, due to lack of a consistent policy the magnitude of the CTV and the acceptable CTV-to-TV distance. For all measurement points, the median planned treatment margin was 3 mm (range, -16 mm to 14 mm). Overall, there was only a loose correlation between pre- and post-implant treatment margins primarily due to variable, implant-related prostatic dimensional changes. Patients with a greater implant-related volume increase tended to have smaller post-implant treatment margins. The post-implant TMs were negatively correlated with dimensional changes, and the negative correlation was most marked for the anterior and posterior TMs due to predominant anterior-posterior dimensional increase. As expected, the post-implant target coverage was higher when larger planning TMs were used, but the correlation was loose due to the unpredictable, highly variable degree of implant-related volume increase. We currently are using 5-mm TMs around the GTV, as identified on pre-implant transrectal ultrasonography or computed tomography. However, the poor correlation between planned and actual post-implant TMs call into question any attempt to make a rational recommendation regarding optimal TMs.

Brachytherapy↗

PubMed1999

Dose escalation of chart in non-small cell lung cancer: is three-dimensional conformal radiation therapy really necessary?

PURPOSE: To evaluate, preclinically, the potential for dose escalation of continuous, hyperfractionated, accelerated radiation therapy (CHART) for non small-cell lung cancer (NSCLC), we examined the strategy of omission of elective nodal irradiation with and without the application of three-dimensional conformal radiation technology (3DCRT). METHODS AND MATERIALS: 2D, conventional therapy plans were designed according to the specifications of CHART for 18 patients with NSCLC (Stages Ib, IIb, IIIa, and IIIb). Further plans were generated with the omission of elective nodal irradiation (ENI) from the treatment portals (2D minus ENI plans [2D-ENI plans]). Both sets were inserted in the patient's planning computed tomographies (CTs). These reconstructed plans were then compared to alternative, three-dimensional treatment plans which had been generated de novo, with the omission of ENI: 3D minus elective nodal irradiation (3D-ENI plans). Dose delivery to the planning target volumes (PTVs) and to the organs at risk were compared between the 3 sets of corresponding plans. The potential for dose escalation of each patient's 2D-ENI and 3D-ENI plan beyond 54 Gy, standard to CHART, was also determined. RESULTS: PTV coverage was suboptimal in the 2D CHART and the 2D-ENI plans. Only in the 3D-ENI plans did 100% of the PTV get > or = 95% of the dose prescribed (i.e., 51.5 Gy [51.3-52.2]). Using 3D-ENI plans significantly reduced the dose received by the spinal cord, the mean and median doses to the esophagus and the heart. It did not significantly reduce the lung dose when compared to 2D-ENI plans. Escalation of the dose (minimum > or = 1 Gy) with optimal PTV coverage was possible in 55.5% of patients using 3D-ENI, but was possible only in 16.6% when using the 2D-ENI planning strategy. CONCLUSIONS: 3DCRT is fundamental to achieving optimal PTV coverage in NSCLC. A policy of omission of elective nodal irradiation alone (and using 2D technology) will not achieve optimal PTV coverage or dose escalation. 3DCRT with omission of ENI can achieve true escalation of CHART in 55.5% of tumors, depending on their site and N-stage.

Aged↗

PubMed1999

Alterations in c-fos expression, cell proliferation and apoptosis in pressure distended human saphenous vein.

OBJECTIVES: Saphenous vein graft failures, resulting from thrombosis and the abnormal proliferation, migration and apoptosis of vascular smooth muscle cells (VSMC) are major limitations of coronary artery bypass surgery. We investigated whether surgical trauma of human saphenous vein induces the early response gene c-fos and causes alterations in rates of proliferation and apoptosis. METHODS: Surgically prepared human vein consisted of distended (at 350 mmHg for 2 min) or non-distended segments of vein maintained in serum free RPMI at 37 degrees C and 5% CO2 for various time intervals. c-fos expression was detected by Northern analysis. Cell proliferation and apoptosis were determined by [3H]thymidine incorporation combined with proliferation cell nuclear antigen (PCNA) immunostaining and TUNEL, respectively. Labelling indices for proliferation and apoptosis were correlated with vessel was thicknesses. RESULTS: Control, freshly isolated vein expressed no c-fos. Surgically prepared vein synthesized c-fos 1 h following harvesting. There was a significant increase in c-fos in distended compared to non-distended vein. c-Fos protein increased in surgically prepared vein 24 h after harvesting. There was a significant increase in vascular cell proliferation in the non-distended compared to the distended vein: mean (S.E.M.) 1279 (218) vs. 863 (155) dpm/microgram DNA, P < 0.05, n = 6. In addition, the apoptotic index was significantly lower in the media of non-distended vs. distended vein 0.82 (0.2) vs. 5.5 (1.5), P < 0.05, n = 5. CONCLUSIONS: These findings demonstrate that surgical preparation of human saphenous vein increases expression of c-fos mRNA and apoptosis and reduces proliferation when compared with non-distended vein. These changes may influence the failure of saphenous vein grafts.

Adult↗

PubMed1999

Chemoradiation for inoperable non small cell lung cancer: a phase II study using a regimen with acceptable toxicity.

Over the past few years there have been numerous schedules of combined modality therapy proposed as being useful in the management of inoperable non-small cell lung cancer (NSCLC). These have generally involved the use of high dose radiation therapy to doses of the order of 60 Gy combined with chemotherapy given prior to or concurrently with the radiation. Concurrent chemotherapy has been given with the intention of being both active in NSCLC and with the role of being a possible radiosensitiser. The most commonly employed drugs have been cisplatin, etoposide, 5-fluorouracil, vindesine and mitomycin. Although response rates of the primary tumour to the combined therapy have been optimistic, there has not been a great survival benefit with the median survival in most series remaining at just over 12 months. In this study we have prospectively treated a group of patients with non-metastatic inoperable NSCLC with a regimen of known acceptable toxicity. These patients were inoperable because they were unfit for surgery or had locally advanced disease. The local radiological response rate was 86% and the median survival for the whole group was 13 months. Adenocarcinomas appeared to do significantly worse than squamous cell carcinomas. Toxicity was acceptable and lower than reported in other similar series. There was one treatment related death. We feel that this combination of radiation therapy and chemotherapy is a reasonable compromise for a disease which still has a very poor outlook.

Adult↗

PubMed1999

Although calnexin is essential in S. pombe, its highly conserved central domain is dispensable for viability.

In mammalian cells, the calnexin/calreticulin chaperones play a key role in glycoprotein folding and its control within the endoplasmic reticulum (ER), by interacting with folding intermediates via their monoglucosylated glycans. This lectin activity has been mapped in mammalian calnexin/calreticulin chaperones to the central region, which is a highly conserved feature of calnexin/calreticulin molecules across species. The central domain has also been implicated in Ca(2+) binding, and it has been proposed to be involved in the regulation of calcium homeostasis in the ER. Herein, we show that although the Schizosaccharomyces pombe calnexin is essential for viability, cells lacking its 317-amino-acid highly conserved central region are viable under normal growth conditions. However, the central region appears to be necessary for optimal growth under high ER-stress, suggesting that this region is important under extreme folding situations (such as DTT and temperature). The minimal length of calnexin required for viability spans the C-terminal 123 residues. Furthermore, cells with the central domain of the protein deleted were affected in their morphology at 37 degrees C, probably due to a defect in cell wall synthesis, although these mutant cells exhibited the same calcium tolerance as wild-type cells at 30 degrees C.

Amino Acid Sequence↗

PubMed1999

Memory for psychophysical scaling judgments.

In three experiments, we examined memory for responses and stimuli experienced in a single direct psychophysical scaling session in which subjects made absolute magnitude estimations of the loudnesses of pure tones. Recall of scaling responses was found to be accurate for the softest and loudest stimuli, but systematically greater than actual judgments for the intermediate stimulus amplitudes, yielding distorted psychophysical functions for the recall data which nonetheless had the same power function exponent as that for the judged stimuli. Also, memory for the range of stimulus amplitudes was fairly accurate, but subjects could not distinguish between judged and nonjudged amplitudes within that range. The results are consistent with the role of extreme stimuli as anchors for judgment, and indicate that memories for these stimuli and responses made to them can be expected to influence future scaling judgments. These results also are consistent with the uncertainty hypothesis of mnemophysics.

Analysis of Variance↗

PubMed1998

Calnexin and BiP interact with acid phosphatase independently of glucose trimming and reglucosylation in Schizosaccharomyces pombe.

The association of newly synthesized glycoproteins with the ER molecular chaperones calnexin and immunoglobulin binding protein (BiP) has been well documented in a variety of higher eukaryotes. Here we report that Cnx1p, the calnexin homologue in Schizosaccharomyces pombe, associates with newly synthesized molecules of the secreted glycoprotein acid phosphatase. Unlike ligand binding to mammalian calnexin, glucose trimming and reglucosylation of acid phosphatase by UDP-Glc:glycoprotein glucosyltransferase were shown to be dispensable for its binding to Cnx1p. Thus, despite the essentiality of Cnx1p for S. pombe viability, the glucose trimming and reglucosylation cycle does not appear to be required for protein folding in the fission yeast. The association of core-glycosylated acid phosphatase with Cnx1p after exposure of cells to heat shock or to DTT was shown to be reversible. However, Cnx1p stably associated with unglycosylated acid phosphatase after treatment with the core-glycosylation inhibitor tunicamycin. BiP was found to coprecipitate with Cnx1p, under normal and stress conditions, and following inhibition of protein synthesis by cycloheximide. We postulate that Cnx1p and BiP are part of a complex that is involved in the folding of both core-glycosylated trimmed ligands and unglycosylated proteins.

Acid Phosphatase↗

PubMed1998

Byr4 and Cdc16 form a two-component GTPase-activating protein for the Spg1 GTPase that controls septation in fission yeast.

BACKGROUND: Spatial and temporal control of cytokinesis ensures the accurate transmission of genetic material and the correct development of multicellular organisms. An excellent model system in which to study cytokinesis is Schizosaccharomyces pombe because there are similarities between cytokinesis in S. pombe and mammals and because genes involved in S. pombe cytokinesis have been characterized. In particular, formation of the septum is positively regulated by the Spg1 GTPase and its effector, the Cdc7 kinase. Septation is negatively regulated by Cdc16, a protein similar to GTPase-activating proteins (GAPs) for Ypt GTPases, and by Byr4, a protein of unknown biochemical function. This study investigates the relationship between Byr4, Cdc16, and Spg1. RESULTS: Genetic interactions were observed between byr4, cdc16, and spg1 mutants. Byr4 bound to Cdc16 and Spg1 in yeast two-hybrid assays and in coprecipitations in vitro and in yeast. Byr4 inhibited the dissociation and hydrolysis of GTP bound to Spg1, but when Byr4 and Cdc16 were combined together they displayed Spg1GAP activity in vitro; Cdc16 alone had no detectable GAP activity. The binding of Byr4 to Spg1 and the Byr4-Cdc16 Spg1GAP activity were specific because Byr4 and Cdc16 did not bind to or affect the GTPase activities of the seven known S pombe Ypt family GTPase. CONCLUSIONS: Byr4 and Cdc16 form a two-component GAP for the Spg1 GTPase. Byr4 and Cdc16 appear to negatively regulate septation in S. pombe by modulating the nucleotide state of Spg1 possibly in a spatially or temporally controlled manner.

Cell Cycle Proteins↗

PubMed1998

Regulated vacuole fusion and fission in Schizosaccharomyces pombe: an osmotic response dependent on MAP kinases.

BACKGROUND: The budding yeast Saccharomyces cerevisiae uses two mitogenactivated protein (MAP) kinase cascades, the Hog1p and the Mpk1p pathways, to signal responses to hypertonic and hypotonic stress, respectively. Mammalian cells and the fission yeast Schizosaccharomyces pombe have functional homologues of Hog1p - p38/RK/CSBP and Sty1 - which, unlike Hog1p, also mediate other responses. We have investigated the involvement of S. pombe MAP kinase pathways in signalling a newly described response to osmotic stress - that of vacuole fusion and fission. RESULTS: When S. pombe is placed into water, its vacuoles rapidly fuse into larger structures enclosing a greater proportion of the cell's volume. Under some conditions, its vacuoles can slowly fragment in response to salt. Fission requires the Sty1 pathway and also Pmk1, the homologue of S. cerevisiae Mpk1p. Fusion requires Pmk1, Ypt7 - the homologue of a protein involved in S. cerevisiae vacuole fusion - and part of the Sty1 pathway, although Sty1 phosphorylation is unaffected by hypotonic conditions. CONCLUSIONS: Vacuole fusion and fission appear to be homeostatic mechanisms that restore the concentration of the cytosol. Vacuole fusion, like stimulated secretion in higher eukaryotes, is a rapid and specific process of membrane fusion in response to an external stimulus. The Sty1 pathway, in addition to its role in responding to hypertonic stress, is required at a basal level for the expression of factors required to respond to hypotonic stress - a mechanism that may allow the cell to use a common pathway for different responses.

Amino Acid Sequence↗