A device for the accurate production of tailored excitation pulse trains in NMR spectroscopy.
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Biomedical subjects
Publications and source records attributed to J Field.
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A system capable of in vivo volume selected 1H NMR spectroscopy of voxels as small as 0.2 cm3 is described. Signal-to-noise ratio improvements with probe design and a novel signal steering device are detailed. A high-resolution, image-directed proton spectrum from 0.2 cm3 of a rat's brain at 200 MHz obtained using the SUBMERGE/SPACE pulse sequence is presented. Single-scan voxel shimming was implemented to improve spectral resolution.
Large deletion and small insertion mutations in the adenylyl cyclase gene of Saccharomyces cerevisiae were used to map regions required for activation by RAS protein in vitro. The amino-terminal 605 amino acids were found to be dispensable for responsiveness to RAS protein. All other deletions in adenylyl cyclase destroyed its ability to respond to RAS. Small insertion mutations within the leucine-rich repeat region also prevented RAS responsiveness, while other insertions did not.
We report a class of interfering mutants of the human H-ras gene capable of inhibiting phenotypes arising from the expression of the activated RAS2 gene, RAS2val19, in the yeast Saccharomyces cerevisiae. All these mutants encode unprocessed H-ras proteins that remain in the cytoplasm. One of the mutants, H-rasarg186, was examined in detail. H-rasarg186 protein is a competitive inhibitor of RAS2val19 protein. It does not interfere with processing and membrane localization of RAS2val19, nor does it appear to compete with RAS protein for its proposed regulator, the CDC25 protein. By several criteria the RAS2val19 adenylate cyclase interaction is unaffected by H-rasarg186. We infer from our results that H-rasarg186 protein interferes with an alternative function of RAS2val19.
The combination of a frequency nonselective excitation suppression method (1331 sequence) with selective excitation followed by gradient-induced dephasing of water transverse magnetization yielded suppression ratios of greater than 10,000:1. The need for gradient preemphasis and correction of B0 field shifts is discussed. The suppression efficiency of this method compared favorably to results obtained using the CPMG spin-echo technique to observe metabolite resonances in a urine sample.
Successful in vivo NMR spectroscopy requires a combination of techniques to address the problems of volume selection, water suppression, and resolution. All this needs to be done in the very heterogeneous environment found in living organisms. Previously published techniques are used to obtain 1H spectra from a dog brain, observing metabolites with concentrations below 1 mM. Measurements of spin-lattice relaxation times (T1) are also presented. The 1H relaxation times are long (T1 greater than 1.0 s) yielding information about the fluidity of the molecular environment. Comments are made concerning the achievable linewidth in vivo and the deficiencies that phase-encoding spectroscopic methods may have in obtaining high-resolution 1H spectra.
We cloned the adenylyl cyclase gene from the fission yeast Schizosaccharomyces pombe using low-stringency hybridization to the Saccharomyces cerevisiae adenylyl cyclase gene. The Sc. pombe gene encodes a 1692-amino acid-residue protein. The identity of this gene was confirmed by studies of its expression in Sa. cerevisiae. Expression of the carboxyl-terminal region of the Sc. pombe adenylyl cyclase protein will suppress a temperature-sensitive mutation in the Sa. cerevisiae adenylyl cyclase gene. Furthermore, Sa. cerevisiae that lack their endogenous adenylyl cyclase gene and express the carboxyl-terminal region of the Sc. pombe adenylyl cyclase protein have measurable adenylyl cyclase activity. The carboxyl-terminal region of this protein has strong homology with the catalytic domain of the Sa. cerevisiae adenylyl cyclase. Also, Sc. pombe adenylyl cyclase, like Sa. cerevisiae adenylyl cyclase, contains a tandemly repeated motif rich in leucine. Neither yeast protein is particularly homologous to the recently cloned Gs-responsive mammalian adenylyl cyclase [Krupinski, J., Coussen, F., Bakalyar, H. A., Tang, W.-J., Feinstein, P. G., Orth, K., Slaughter, C., Reed, R. R. & Gilman, A. G. (1989) Science 244, 1558-1564].
A formulary covering 10 drug groups and over 50% of prescribing was devised in a general practice and doctors' attitudes to the idea to a formulary were assessed before and after the study. The prescribing of formulary drugs rose from 72% to 81% over two years, and the general practitioners were significantly more positive towards formularies in the practice which devised the formulary but not in three control practices. A group of patients receiving repeat prescriptions was interviewed in three practices in three consecutive years, and there was no difference in satisfaction with drugs between the formulary practice and the two control practices. However, an association between changing a drug and dissatisfaction was noted, and there was a trend of decreasing satisfaction with prescribing and with information given about drugs over the three years in all the practices. Overall 51% of patients felt that they had either too little or no information given to them about their drugs. Improving information given to both doctors and patients about drugs may be important in improving prescribing without causing discontentment.
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We developed a method for immunoaffinity purification of Saccharomyces cerevisiae adenylyl cyclase based on creating a fusion with a small peptide epitope. Using oligonucleotide technology to encode the peptide epitope we constructed a plasmid that expressed the fusion protein from the S. cerevisiae alcohol dehydrogenase promoter ADH1. A monoclonal antibody previously raised against the peptide was used to purify adenylyl cyclase by affinity chromatography. The purified enzyme appeared to be a multisubunit complex consisting of the 200-kilodalton adenylyl cyclase fusion protein and an unidentified 70-kilodalton protein. The purified protein could be activated by RAS proteins. Activation had an absolute requirement for a guanine nucleoside triphosphate.
B mode ultrasound was used to assess and map the long saphenous vein in 20 limbs prior to femorodistal bypass. The assessment was compared with operative findings. Eighteen of 19 adequate veins and 8 of 9 anatomical abnormalities or major divisions were correctly identified. B mode ultrasound allows accurate marking of the vein, facilitating dissection, alerts the surgeon to possible difficulties and is an ideal non-invasive technique for preoperative assessment of the long saphenous vein.
In vivo high resolution volume-selected 1H magnetic resonance spectroscopy of human tibia has been undertaken using spatial coordinates obtained from magnetic resonance images. Adult tibial marrow has a 1H spectrum rich in fatty acid resonances and is readily distinguished from the 1H spectra of surrounding leg muscle. In all four leukemic patients examined, infiltration of fat cells of tibial marrow by proliferating cells rich in mobile H2O protons was evident by magnetic resonance imaging. Selective examination of volumes of tibial marrow (1 cm3) by 1H magnetic resonance spectroscopy confirmed marked differences in the 1H spectra of marrow from these patients. Increases in the H2O peak of the 1H spectra were correlated with infiltration of blast cells and lack of control of the neoplastic disease. These studies are the first to report the use of volume selected magnetic resonance spectroscopy to selectively monitor leukemia in humans.
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Induction of iron overload in mice using 1% (w/w) dietary carbonyl iron resulted in marked decreases in 1H and 31P NMR relaxation times. Natural-abundance deuterium (2H) NMR spectroscopy has been used to measure 2H T1 values in vivo in the presence of body paramagnetic iron. This procedure offers a method for noninvasive determination of body iron stores.
Four of the techniques proposed for in vivo volume-selected NMR spectroscopy have been compared using a simple phantom with a large background water signal which was outside the region of interest. The methods VSE, SPACE, SPARS, and DIGGER all use pulsed field gradients for spatial encoding and were tested with a symmetric and an asymmetric phantom. SPACE was used to obtain volume-selected 1H NMR spectra of a human leg, demonstrating excellent discrimination between bone marrow and muscle.
Volume selection using SPACE has been combined with water suppression techniques to provide high-resolution 1H NMR spectra from aqueous solutions. The technique developed was used to obtain spectra from a tumor growing on the hind leg of a rat. Water suppression factors of between 1000 and 2000 were achieved simultaneously with excellent volume selection.
A novel method for reducing the rf power requirements of selective pulses employed for z magnetization slice inversion in localized NMR spectroscopy is presented. Following slice preparation by the use of frequency-incremented sinc pulses, the slice gradient is reduced, which narrows the frequency width of the slice. This allows the use of relatively low-power selective inversion pulses, an important consideration for in vivo applications.