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R D Smith

Publications and source records attributed to R D Smith.

At least 91 records · Page 5Linked to original sources
PubMed2000

Specific frequencies of spontaneous Ca2+ transients upregulate GAD 67 transcripts in embryonic spinal neurons.

Spontaneous Ca2+ transients expressed prior to synaptogenesis regulate the developmental appearance of GABA in cultured Xenopus spinal neurons. We find that glutamic acid decarboxylase (GAD) immunoreactivity is also Ca(2+)-dependent and parallels the appearance of GABA. We show that xGAD 67 transcripts first appear in the embryonic spinal cord during the period in which these Ca2+ spikes are generated, in a pattern that is temporally and spatially appropriate to account for differentiation of GABAergic interneurons. RNase protection and competitive quantitative RT-PCR demonstrate that transcript levels are approximately threefold greater when neurons are cultured in the presence of extracellular Ca2+ that permits generation of transients than when cultured in its absence. The frequency of spontaneous Ca2+ spikes plays a crucial role in the regulation of transcripts, since reimposition of Ca2+ transients at the frequency generated in cultured neurons rescues normal expression. We conclude that naturally occurring low frequencies of these Ca2+ transients regulate levels of xGAD 67 mRNA in differentiating neurons.

Amino Acid Sequence↗
PubMed2000

Effects of losartan and candesartan monotherapy and losartan/hydrochlorothiazide combination therapy in patients with mild to moderate hypertension. Losartan Trial Investigators.

OBJECTIVE: The goal of this multicenter, double-blind, randomized, parallel-group study was to compare the effects of losartan potassium (hereafter referred to as losartan), candesartan cilexitil (hereafter referred to as candesartan), and losartan/hydrochlorothiazide (HCTZ) in patients with mild to moderate hypertension (sitting diastolic blood pressure [SiDBP] 95-115 mm Hg). METHODS: A total of 1161 patients were randomized in a 2:2:1 ratio to 12 weeks of treatment with losartan 50 mg QD, possibly titrated to 100 mg QD (n = 461); candesartan 8 mg QD, possibly titrated to 16 mg QD (n = 468); or losartan 50 mg QD, possibly titrated to losartan 50 mg plus HCTZ 12.5 mg QD (n = 232). At 6 weeks, the regimens of patients not reaching a goal SiDBP <90 mm Hg were titrated as described, whereas patients achieving this goal continued with low-dose monotherapy. The single primary end point at 12 weeks tested the equivalence of the 2 monotherapy regimens, predefined as a maximum between-treatment difference in the mean change from baseline trough SiDBP of 2.5 mm Hg. RESULTS: At 12 weeks, changes in SiDBP/sitting systolic blood pressure (SiSBP) of -12.4/-14.4 mm Hg with losartan 50 mg/100 mg and -13.1/-15.8 mm Hg with candesartan 8 mg/16 mg demonstrated equivalence between the 2 monotherapy regimens (95% CI for difference in SiDBP, -1.6 to 0.2). At 12 weeks, the losartan 50 mg/50 mg plus HCTZ 12.5 mg regimen had reduced SiDBP/SiSBP significantly more (-14.3/-18.0 mm Hg) than either the candesartan 8 mg/16 mg (SiDBP, P = 0.045; SiSBP, P = 0.017) or losartan 50 mg/100 mg regimen (SiDBP and SiSBP, P = 0.001). During the last 6 weeks, patients whose regimen had been titrated to losartan 50 mg plus HCTZ 12.5 mg (n = 114) showed a greater reduction in SiDBP/SiSBP (-14.5/ -18.7 mm Hg) than did those whose regimen had been titrated to either losartan 100 mg (-10.5/-12.3 mm Hg; n = 211) or candesartan 16 mg (-11.5/-13.2 mm Hg; n = 206), representing a clinically meaningful > or = 2.5-mm Hg) difference. All 3 treatments were well tolerated, with few patients experiencing drug-related adverse events (6.9% losartan 50 mg/100 mg, 7.5% candesartan 8 mg/16 mg, 3.0% losartan 50 mg/ 50 mg plus HCTZ 12.5 mg). Candesartan 8 mg/16 mg increased serum uric acid levels (0.13 mg/dL; 95% CI, 0.04 to 0.23), whereas losartan 50 mg/100 mg decreased them (-0.14 mg/dL; 95% CI, -0.24 to -0.04), and losartan 50 mg/50 mg plus HCTZ 12.5 mg left them unchanged (0.06 mg/dL; 95% CI, -0.07 to 0.20). CONCLUSIONS: Losartan 50 mg/100 mg and candesartan 8 mg/16 mg were comparable treatments in terms of blood pressure reduction. After titration, losartan 50 mg plus HCTZ 12.5 mg was superior to either candesartan 16 mg or losartan 100 mg in reducing hypertension. Losartan, but not candesartan, lowered serum uric acid levels and attenuated the expected increase in uric acid levels with HCTZ 12.5 mg.

Adult↗
PubMed2000

Obtaining more accurate Fourier transform ion cyclotron resonance mass measurements without internal standards using multiply charged ions.

Space-charge effects produce frequency shifts in Fourier transform ion cyclotron resonance (FTICR) mass spectrometry and correction for these shifts is necessary for obtaining accurate mass measurements. We report a novel method for obtaining accurate mass calibration to correct for space-charge induced mass shifts without the requirement for internal calibrants. The new approach is particularly well suited for electrospray ionization-FTICR mass spectra that contain multiple charge states of the same molecular species. This method, deconvolution of Coulombic affected linearity (DeCAL), is described and presented with several examples demonstrating the increased mass measurement accuracy obtained. DeCAL provides the basis for more routinely obtaining higher mass accuracy measurements in conjunction with chromatographic separations for complex mixture analysis, and obviates the need for internal calibration in many applications.

Algorithms↗
PubMed2000

Proteome analysis using selective incorporation of isotopically labeled amino acids.

A method is described for identifying intact proteins from genomic databases using a combination of accurate molecular mass measurements and partial amino acid content. An initial demonstration was conducted for proteins isolated from Escherichia coli (E. coli) using a multiple auxotrophic strain of K12. Proteins extracted from the organism grown in natural isotopic abundance minimal medium and also minimal medium containing isotopically labeled leucine (Leu-D10), were mixed and analyzed by capillary isoelectric focusing (CIEF) coupled with Fourier transform ion cyclotron resonance mass spectrometry (FTICR). The incorporation of the isotopically labeled Leu residue has no effect on the CIEF separation of the protein, therefore both versions of the protein are observed within the same FTICR spectrum. The difference in the molecular mass of the natural isotopic abundance and Leu-D10 isotopically labeled proteins is used to determine the number of Leu residues present in that particular protein. Knowledge of the molecular mass and number of Leu residues present can be used to unambiguously identify the intact protein. Preliminary results show the efficacy of this method for unambiguously identifying proteins isolated from E. coli.

Amino Acids↗
PubMed2000

Initial implementation of an electrodynamic ion funnel with Fourier transform ion cyclotron resonance mass spectrometry.

Fourier transform ion cyclotron resonance (FTICR) mass spectrometry has become a widely used method to study biopolymers. The method, in combination with an electrospray ionization (ESI) source has demonstrated the highest resolution and accuracy yet achieved for characterization of biomolecules and their noncovalent complexes. The most common design for the ESI interface includes a heated capillary inlet followed by a skimmer having a small orifice to limit gas conductance between a higher pressure (1 to 5 torr) source region and the lower pressure ion guide. The ion losses in the capillary-skimmer interface are large (estimated to be more than 90%) and thus reduce achievable sensitivity. In this work, we report on the initial implementation of a newly developed electrodynamic ion funnel in a 3.5 tesla ESI-FTICR mass spectrometer. The initial results show dramatically improved ion transmission as compared to the conventional capillary-skimmer arrangement. An estimated detection limit of 30 zeptomoles (approximately 18,000 molecules) has been achieved for the analysis of the proteins with molecular weights ranging from 8 to 20 kDa.

Cyclotrons↗
PubMed2000

A simple nanoelectrospray arrangement with controllable flowrate for mass analysis of submicroliter protein samples.

A simple arrangement for nanoelectrospray ionization using a conventional syringe pump connected to a pulled unmodified capillary has been evaluated. This arrangement avoids several disadvantages associated with metal-coated nanoelectrospray emitters. The relatively large orifice (approximately 9 microns) at the pulled capillary tip reduces sample clogging and the use of the pump minimizes spray disruption due to gas bubbles. Subattomole detection limit was achieved with nanomolar protein sample solutions at 5-10 nL/min flowrates using an LCQ mass spectrometer. Submicroliter samples can be loaded from the tip orifice and stored inside the capillary to virtually eliminate any dead volume, and then be electrosprayed for extended periods at well-controlled flowrates.

Mass Spectrometry↗
PubMed2000

Electrospray ionization Fourier transform ion cyclotron resonance mass spectrometric analysis of the recombinant human macrophage colony stimulating factor beta and derivatives.

The potential of electrospray ionization (ESI) Fourier transform ion cyclotron mass spectrometry (FTICR-MS) to assist in the structural characterization of monomeric and dimeric derivatives of the macrophage colony stimulating factor beta (rhM-CSF beta) was assessed. Mass spectrometric analysis of the 49 kDa protein required the use of sustained off-resonance irradiation (SORI) in-trap cleanup to reduce adduction. High resolution mass spectra were acquired for a fully reduced and a fully S-cyanylated monomeric derivative (approximately 25 kDa). Mass accuracy for monomeric derivatives was better than 5 ppm, after applying a new calibration method (i.e., DeCAL) which eliminates space charge effects upon high accuracy mass measurements. This high mass accuracy allowed the direct determination of the exact number of incorporated cyanyl groups. Collisionally induced dissociation using SORI yielded b- and y-fragment ions within the N- and C-terminal regions for the monomeric derivatives, but obtaining information on other regions required proteolytic digestion, or potentially the use of alternative dissociation methods.

Amino Acid Sequence↗
PubMed2000

Electrospray ionization Fourier transform ion cyclotron resonance analysis of large polymerase chain reaction products.

In previous work, we demonstrated the use of electrospray ionization to analyze small differences in size or sequence of relatively small polymerase chain reaction (PCR) products of 114 base pairs or less. The sequence information required to answer a biological question may be only a single nucleotide substitution or deletion. In many cases, the regions where these sequence variations can occur are several hundred base pairs in length, and the analysis of large PCR products is therefore desirable. Therefore, we have attempted to expand the size range of PCR products that can be analyzed by electrospray ionization (ESI) Fourier transform ion cyclotron resonance (FTICR) mass spectrometry. Previous work has shown that the difficulties associated with PCR product analysis increase with product size. A revised cleanup scheme was employed to target the removal of detergents with ethanol wash or precipitation steps, followed by additional desalting. Additionally, an in-trap cleanup to collisionally induce dissociation of noncovalent salt adducts was employed. This approach was extended to a 223 base pair PCR product yielding mass measurement accuracy within 26 ppM. The mass measurement accuracy obtained illustrates that a single base substitution could be identified at this size of PCR product with a 7 tesla ESI-FTICR.

Bacteriophage lambda↗
PubMed2000

Transmission Electron Microscope Specimen Preparation of Metal Matrix Composites Using the Focused Ion Beam Miller.

Transmission electron microscope samples of two types of metal matrix composites were prepared using both traditional thinning methods and the more novel focused ion beam miller. Electropolishing methods were able to produce, very rapidly, thin foils where the matrix was electron transparent, but the ceramic reinforcement particles remained unthinned. Thus, it was not possible in these foils to study either the matrix-reinforcement interface or the microstructure of the reinforcement particles themselves. In contrast, both phases in the composites prepared using the focused ion beam miller thinned uniformly. The interfaces in these materials were clearly visible and the ceramic reinforcement was electron transparent. However, microstructural artifacts associated with ion beam damage were also observed. The extent of these artifacts and methods of minimizing their effect were dependent on both the materials and the milling conditions used.

Journal Article↗
PubMed2000

Reduction and coordination of arsenic in Indian mustard.

The bioaccumulation of arsenic by plants may provide a means of removing this element from contaminated soils and waters. However, to optimize this process it is important to understand the biological mechanisms involved. Using a combination of techniques, including x-ray absorption spectroscopy, we have established the biochemical fate of arsenic taken up by Indian mustard (Brassica juncea). After arsenate uptake by the roots, possibly via the phosphate transport mechanism, a small fraction is exported to the shoot via the xylem as the oxyanions arsenate and arsenite. Once in the shoot, the arsenic is stored as an As(III)-tris-thiolate complex. The majority of the arsenic remains in the roots as an As(III)-tris-thiolate complex, which is indistinguishable from that found in the shoots and from As(III)-tris-glutathione. The thiolate donors are thus probably either glutathione or phytochelatins. The addition of the dithiol arsenic chelator dimercaptosuccinate to the hydroponic culture medium caused a 5-fold-increased arsenic level in the leaves, although the total arsenic accumulation was only marginally increased. This suggests that the addition of dimercaptosuccinate to arsenic-contaminated soils may provide a way to promote arsenic bioaccumulation in plant shoots, a process that will be essential for the development of an efficient phytoremediation strategy for this element.

Arsenic↗
PubMed2000

Babesiosis (Babesia bovis) stability in unstable environments.

Enzootic stability (herd immunity) in bovine babesiosis occurs when the rate of transmission (inoculation rate) of Babesia spp by the tick vector is sufficient to immunize a majority of susceptible calves before the loss of calfhood resistance. The effect of three tick (Boophilus microplus) control strategies (none, threshold, and strategic) on enzootic stability and the likelihood of babesiosis (Babesia bovis) outbreaks was studied using a spreadsheet age-class computer simulation model. The model was driven by weekly bovine tick counts from Brazil and Uruguay. The Eldorado do Sul, RS, Brazil bovine population (30 degrees 05' South latitude) was found to be in a naturally occurring state of enzootic stability, corresponding to an inoculation rate exceeding 0.005 throughout the year. Threshold dipping strategies should not increase the risk of babesiosis in cattle so managed. Strategic dipping resulted in an extended period of enzootic instability lasting 30 weeks, which requires protection of the herd through immunization. Because of the more prolonged low winter temperature conditions, the Tacuarembó, Uruguay bovine population (31 degrees 40' South latitude) was found to be in a naturally occurring state of enzootic instability, characterized by a 28 week period in which the inoculation rate was below 0.005. Strategic dipping should lead to eradication of the babesial parasite from tick and bovine populations, but would not result in eradication of the tick vector. This could lead to subsequent outbreaks if Babesia carrier animals were to be introduced into the herd. In both populations, strategic tick control could be accompanied by concurrent babesiosis vaccination.

Animals↗
PubMed2000

Homologous and heterologous phosphorylation of the AT(2) angiotensin receptor by protein kinase C.

The angiotensin AT(2) receptor is an atypical seven transmembrane domain receptor that is coupled to activation of tyrosine phosphatase and inhibition of MAP kinase, and does not undergo agonist-induced internalization. An investigation of the occurrence and nature of AT(2) receptor phosphorylation revealed that phorbol ester-induced activation of protein kinase C (PKC) in HA-AT(2) receptor-expressing COS-7 cells caused rapid and specific phosphorylation of a single residue (Ser(354)) located in the cytoplasmic tail of the receptor. Agonist activation of AT(2) receptors by angiotensin II (Ang II) also caused rapid PKC-dependent phosphorylation of Ser(354) that was prevented by the AT(2) antagonist, PD123177, and by inhibitors of PKC. In cells coexpressing AT(1) and AT(2) receptors, Ang II-induced phosphorylation of the AT(2) receptor was reduced by either PD123177 or the AT(1) receptor antagonist, DuP753, and was abolished by treatment with both antagonists or with PKC inhibitors. These findings indicate that the AT(2) receptor is rapidly phosphorylated via PKC during homologous activation by Ang II, and also undergoes heterologous PKC-dependent phosphorylation during activation of the AT(1) receptor. The latter process may regulate the counteracting effects of AT(2) receptors on growth responses to AT(1) receptor activation.

Animals↗
PubMed2000

Potentiation of rat brain sodium channel currents by PKA in Xenopus oocytes involves the I-II linker.

Functional modulation of voltage-gated sodium channels affects the electrical excitability of neurons. Protein kinase A (PKA) can decrease sodium currents by phosphorylation at consensus sites in the cytoplasmic I-II linker. Once the sites are phosphorylated, however, additional PKA activity can increase sodium currents by an unknown mechanism. When the PKA sites were eliminated by substitutions of alanine for serine, peak sodium current amplitudes were increased by 20-80% when PKA was activated in Xenopus oocytes either by stimulation of a coexpressed beta(2)-adrenergic receptor or by perfusion with reagents that increase cAMP. Potentiation required the I-II linker of the brain channel, in that a chimeric channel in which the brain linker was replaced with the comparable linker from the skeletal muscle channel did not demonstrate potentiation. Using a series of chimeric and deleted channels, we demonstrate that potentiation is not dependent on any single region of the linker and that the extent of potentiation varies depending on the total length and the residues throughout the linker. These data are consistent with the hypothesis that potentiation by PKA is an indirect process involving phosphorylation of an accessory protein that interacts with the I-II linker of the sodium channel.

Animals↗
PubMed2000

The Child-Friendly Healthcare Initiative (CFHI): Healthcare provision in accordance with the UN Convention on the Rights of the Child. Child Advocacy International. Department of Child and Adolescent Health and Development of the World Health Organization (WHO). Royal College of Nursing (UK). Royal College of Paediatrics and Child Health (UK). United Nations Children's Fund (UNICEF).

OBJECTIVE: Although modern medical technology and treatment regimens in well-resourced countries have improved the survival of sick or injured children, most of the world's families do not have access to adequate health care. Many hospitals in poorly resourced countries do not have basic water and sanitation, a reliable electricity supply, or even minimal security. The staff, both clinical and nonclinical, are often underpaid and sometimes undervalued by their communities. In many countries there continues to be minimal, if any, pain control, and the indiscriminate use of powerful antibiotics leads to a proliferation of multiresistant pathogens. Even in well-resourced countries, advances in health care have not always been accompanied by commensurate attention to the child's wider well-being and sufficient concerns about their anxieties, fears, and suffering. In accordance with the United Nations Convention on the Rights of the Child,(1) the proposals set out in this article aim to develop a system of care that will focus on the physical, psychological, and emotional well-being of children attending health care facilities, particularly as inpatients. DESIGN OF THE PROGRAM: To develop in consultation with local health care professionals and international organizations, globally applicable standards that will help to ensure that practices in hospitals and health centers everywhere respect children's rights, not only to survival and avoidance of morbidity, but also to their protection from unnecessary suffering and their informed participation in treatment. Child Advocacy International will liase closely with the Department of Child and Adolescent Health and Development of the World Health Organization (WHO) and the United Nations Children's Fund (UNICEF) in the implementation of the pilot scheme in 6 countries. In hospitals providing maternity and newborn infant care, the program will be closely linked with the Baby Friendly Hospital Initiative of WHO/UNICEF that aims to strengthen support for breastfeeding. United Nations Children's Fund, United Nations Convention on the Rights of the Child, child protection, breastfeeding, pain control, palliative care, child abuse.

Child↗
PubMed2000

Applications of informatics in veterinary medicine.

This study used the peer-reviewed biomedical literature to define the veterinary informatics knowledgebase and associated subspecialties, and assesses the level of activity in the field over the thirty-year period from 1966 through 1995. Grateful Med was used to search the MEDLINE bibliographic database for articles that shared one or more Medical Subject Headings (MeSH) keywords from the veterinary and medical informatics subject headings. Each of ninety-five MeSH medical informatics terms was assigned to one of twelve veterinary informatics subspecialties. The number of articles retrieved by each MeSH keyword and subspecialty was calculated. A total of 611 articles were retrieved, representing the contributions of 1,338 authors published in 153 journals. The field experienced slow growth over the twenty-year period from 1966 through 1985. In the following decade, the cumulative number of veterinary informatics articles almost tripled and the percentage of veterinary-related articles that included an informatics component increased almost two-and-one-half fold. Despite this recent growth, the number of veterinary-related articles with an informatics component has never exceeded 1% of either the veterinary or medical informatics literature over the past thirty years, and representation of veterinary subspecialties in the literature varied widely.

Animals↗