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Biomedical subjects

J M Smith

Publications and source records attributed to J M Smith.

At least 271 records · Page 15Linked to original sources

Distribution of motor cortical neuron synaptic terminals on monkey parvocellular red nucleus neurons.

We determined the location of 54 horseradish peroxidase (HRP)-labeled motor cortical neuron synaptic terminals on 17 parvocellular neurons in the monkey red nucleus. Synaptic terminals and their postsynaptic elements were identified and reconstructed, using light- and electron-microscopic techniques, from serial thick and thin sections. Terminals were found on proximal and distal dendrites of small and medium-sized parvocellular neurons, where they formed excitatory synapses. Some were 180 microns from cell somata. Approximately half of the labeled terminals, aside from those located at dendritic origins, were situated strategically at or near dendritic branch points. Since monkey parvocellular neurons show little activity during movement, the obvious next question is this: How and in what way does motor cortex influence these cells?

Animals↗

Data collection in the Great Plains Genetics Service Network: using limited funds to collect data from centers with varying resources.

The Data Committee of the Great Plains Genetics Service Network (GPGSN) coordinates the collection of data relating to delivery of genetic services in eight states. These states are Iowa, Missouri, Arkansas, Oklahoma, Kansas, Nebraska, South Dakota and North Dakota. The funds allocated to this project by the GPGSN are limited. The distance between genetics service sites is great and the population density in the regions being served is low. The local resources available to the genetics services sites participating in data collection vary from robust to "bare-bones". The approach to solving the problem involved the following. First the committee the data items to be collected were identified and defined. Second, a standard format for transmitting the data to the GPGSN regional coordinating center in Iowa City was developed. Third, the services sites and their resources for collecting data were identified. Fourth, resources were allocated to different sites in a manner that seemed most able to help that center to contribute data to the regional center. Fifth, data were aggregated at the regional center and aggregated data reports were returned to collecting sites. Finally, items were modified in response to the feedback received from the genetics services sites. Although the philosophy is that data collection should be a by-product of providing quality genetic services, the region recognizes that service sites will need help to conform with regional standards. Therefore the region encourages each service site to develop its own method to collect data, and provides assistance to it in getting the data into the regional transmission format.(ABSTRACT TRUNCATED AT 250 WORDS)

Data Collection↗

Active-site mapping and site-specific mutagenesis of glycinamide ribonucleotide transformylase from Escherichia coli.

The affinity reagent N10-(bromoacetyl)-5,8-dideazafolate has previously been shown to inactivate glycinamide ribonucleotide transformylase (EC 2.1.2.2) from Escherichia coli in an active-site-directed manner with a 1:1 stoichiometry [Inglese et al. (1990) Biochemistry 29, 1436-1443]. After a series of mild proteolytic digestions, the dideazafolate label was localized to an active-site peptide attached by an ester linkage to the highly conserved residue Asp 144. Subsequent site-specific mutagenesis of Asp 144 to Asn 144 resulted in a catalytically inactive enzyme that retained the ability to bind substrates and inhibitors. The Asn 144 mutant could be further labeled with the affinity reagent in an active-site-directed stoichiometric fashion; however, the site of modification in this case was His 119. These results imply that Asp 144 may function as a general base within the catalytic center of the transformylase and is in close proximity to His 119 in the folded protein.

Acyltransferases↗

Bilateral renal dysplasia with nephron hypoplasia in a foal.

Bilateral renal dysplasia and nephron hypoplasia was diagnosed in a Quarter Horse foal with clinical signs of lethargy, convulsions, and diarrhea. Laboratory evaluation revealed anemia, hypoproteinemia, leukopenia, hyponatremia, hypochloremia, and hyposmolality. The foal also had high concentrations of serum creatinine, BUN, and phosphorus. Evaluation of urinary indices revealed a high ratio of urinary gamma-glutamyl-transferase activity to concentration of creatinine, as well as a high fractional clearance ratio of sodium and potassium. Intravenous treatment with saline solution (0.9% NaCl) and antimicrobials provided only temporary resolution of some of the abnormalities. Diagnosis was partly established by histologic evaluation of renal tissue obtained via an ultrasonographically guided biopsy and was confirmed at necropsy. Pathologic changes in the kidney were unique in that the size of the kidneys, along with the appearance and number of glomeruli, were essentially normal despite marked hypoplasia of nephron tubules in the medulla.

Animals↗

Contempo '90.

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Medicine↗

Subcloning, characterization, and affinity labeling of Escherichia coli glycinamide ribonucleotide transformylase.

Glycinamide ribonucleotide transformylase (GAR TFase; EC 2.1.2.2) has been purified 70-fold to apparent homogeneity from Escherichia coli harboring an expression vector encoding the purN gene product, GAR TFase. The protein is a monomer of Mr 23,241 and catalyzes a single reaction. Steady-state kinetic parameters for the enzyme have been obtained. The structural requirements for cofactor utilization have been investigated and found to parallel those of the multifunctional avian enzyme. The enzyme was inactivated with the affinity label N10-(bromoacetyl)-5,8-dideazafolate in a stoichiometric and active-site-specific manner. The ionization state of the cofactor analogue in the enzyme-cofactor complex appears to require the dissociation of the proton at N3 of the pyrimidine within the complex.

Acyltransferases↗

Identification of a novel transforming growth factor-beta (TGF-beta 5) mRNA in Xenopus laevis.

A novel transforming growth factor-beta (TGF-beta) mRNA of about 3.0 kilobases, which encodes a putative protein of 382 amino acids, has been identified in amphibians by cDNA cloning. This mRNA, which we designate as TGF-beta 5, is developmentally regulated and highly expressed beginning at early neurula (stage 14) and in many adult tissues in Xenopus laevis. Following the first methionine, the putative precursor protein has a hydrophobic region, approximately 22 amino acids long, which probably represents a signal sequence, similar to that found in TGF-beta s 1-3. The precursor also has potential sites for glycosylation, integrin binding (RGD), and a tetrabasic amino acid (RKKR) site for potential cleavage of the precursor peptide to a biologically active protein. The putative mature protein consists of 112 amino acids with 9 cysteines and has 76, 66, 69, and 72% identity to TGF-beta s 1-4, respectively.

Amino Acid Sequence↗

Glycinamide ribonucleotide synthetase from Escherichia coli: cloning, overproduction, sequencing, isolation, and characterization.

The purD gene of Escherichia coli encoding the enzyme glycinamide ribonucleotide (GAR) synthetase, which catalyzes the conversion of phosphoribosylamine (PRA), glycine, and MgATP to glycinamide ribonucleotide, MgADP, and Pi, has been cloned and sequenced. The protein, as deduced by the structural gene sequence, contains 430 amino acids and has a calculated Mr of 45,945. Construction of an overproducing strain behind a lambda pL promoter allowed a 4-fold purification of the protein to homogeneity. N-Terminal sequence analysis and comparison of the sequence with those of other GAR synthetases confirm the amino acid sequence deduced from the gene sequence. Initial velocity studies and product and dead-end inhibition studies are most consistent with a sequential ordered mechanism of substrate binding and product release in which PRA binds first followed by MgATP and then glycine; Pi leaves first, followed by loss of MgADP and finally GAR. Incubation of [18O]glycine, ATP, and PRA results in quantitative transfer of the 18O to Pi. GAR synthetase is very specific for its substrate glycine.

Amino Acid Sequence↗

Effect of oltipraz on the susceptibility of adult Schistosoma mansoni to killing by mouse peritoneal exudate cells.

Incubation of the adult Schistosoma mansoni with the anti-schistosomal compound oltipraz (OPZ) (40 nM) resulted in a significant decrease in schistosome-reduced glutathione (GSH), a thiol compound which may have a role in protection against oxidant-mediated damage. A significant proportion (20-47%) of worms treated with OPZ became susceptible to in vitro killing by zymosan-stimulated peritoneal exudate cells from mice infected with S. mansoni or inoculated with Bacillus Calmette Guérin (BCG). Killing of the worms was partially inhibited by the addition to the assay system of exogenous glutathione peroxidase with GSH but not by superoxide dismutase. These results suggested that killing of parasites exposed to the drug was partly mediated by cell-generated hydrogen peroxide. They indicate also that depletion of schistosome GSH levels could render the parasites susceptible to killing by oxidative mechanisms, and suggest that there is potential in exploiting schistosome oxidant defense systems and reactive oxygen byproducts in the treatment of schistosomiasis. Inhibition of schistosome oxidant defense systems with drugs may render the parasites susceptible to killing by reactive oxygen byproducts of effector cells.

Animals↗

Schistosoma mansoni: levels of antioxidants and resistance to oxidants increase during development.

The effects of cell-free generated oxidants on migrating and developing stages of Schistosoma mansoni were investigated and the levels of antioxidant enzymes and of glutathione were determined for each stage. Schistosomula and 2-week-old parasites recovered from the livers of infected mice showed similar susceptibility to killing by added hydrogen peroxide and t-butylhydroperoxide. However, when glucose (0.5 mM)-glucose oxidase (2.5 mU ml-1) and xanthine (0.5 mM) or hypoxanthine (0.5 mM)-xanthine oxidase (5.0 mU ml-1) systems were used to generate hydrogen peroxide and oxygen free-radicals, schistosomula were more susceptible to oxidative killing than the 2-week-old parasites. The 4- and 8-week-old worms were more resistant to oxidants than all of the younger stages. High levels of superoxide dismutase (16.2-24.8 U mg-1 protein) were present in all stages. Catalase was not detected. Glutathione peroxidase activity with cumene hydroperoxide as substrate was not detectable in the schistosomula but the activity was present in the 2-week-old parasites. However, hydrogen peroxide-sensitive glutathione peroxidase activity was present in all the stages with a threefold difference in activity between schistosomula and the adult stages. Glutathione-s-transferase activity was significantly lower in the schistosomula, lung stages, and the 2-week-old parasites than in the older stages. Progressive increases in the levels of glutathione reductase and glutathione were also observed with development. The differences in the levels of antioxidants between different stages of development may partly explain the increase in resistance to oxidant-mediated damage as the parasite develops.

Animals↗

Screening procedure for assessment of ototoxicity in the common marmoset.

Detection of drug-induced ototoxicity in safety evaluation studies of novel chemical entities is rarely attempted. Where such examinations are included, they usually rely on reflex testing. The Brainstem Auditory Evoked Response can be measured with the use of externally positioned electrodes, and it monitors electrophysiologic responses to sound from the cochlear nerve and associated structures of the 8th cranial nerve. These responses have been reproducibly measured in sedated marmosets and the method shown to be a sensitive detector of hearing loss caused by loop diuretics or aminoglycoside antibiotics. additionally, where hearing damage is reversible, recovery can also be monitored. It is proposed that where a sensitive marker for ototoxicity is considered necessary as part of a multifaceted study investigating in vivo drug safety, this procedure may prove advantageous over existing methods.

Animals↗