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Are cysteines present at the active site of glycogen phosphorylase?

Periodate and the anions of the transitional elements inactivate glycogen phosphorylase with resolution of the coenzyme from the active site. The effects of the ionic strength and pH of the incubation mixture on the rate of inactivation allow to discriminate between the actions of these chemicals, suggesting that they recognize different regions at the enzyme active site. This conclusion is in agreement with the identification of cysteine as the target amino acid for the inactivation by periodate while arginine was reported to be responsible for the vanadate mediated inactivation.

Arginine↗

Synergism in insulin-like effects of molybdate plus H2O2 or tungstate plus H2O2 on glucose transport by isolated rat adipocytes.

The effect of molybdate, tungstate, molybdate plus H2O2 or tungstate plus H2O2 on 3-O-methylglucose (3-O-MG) uptake was studied in isolated rat adipocytes to investigate whether these agents possess an insulin-like action. High concentrations (10-30 mM) of molybdate or tungstate significantly stimulated the uptake of 3-O-MG while 1 mM of the metaloxides did not. The combination of 1 mM molybdate and 1 mM H2O2, or 1 mM tungstate and 1 mM H2O2 induced striking stimulation of the uptake of 3-O-MG in a synergistic manner, whereas 1 mM H2O2 alone showed only a small effect. The effect of metaloxides plus H2O2 (1 mM) and the effect of insulin (20 nM) were not additive, and both effects were ATP or energy dependent based on experiments using KCN. These results indicate that a weak insulin-like effect of molybdate or tungstate is potentiated synergistically with H2O2, presumably by producing peroxocompounds. Based on the present findings, these new agents may be useful for investigating the mechanism of insulin action and may indicate a new class of drugs for diabetes mellitus.

3-O-Methylglucose↗

Epileptogenic action of tungstic acid gel on cat lumbar motoneurons.

Injection of tungstic acid gel (but not pH adjusted saline) into the cat lumbar ventral horn results in spontaneous, epileptiform activity consisting of waves of repetitive, high frequency, action potentials in motoneurons surrounding the injection site. In most motoneurons the action potentials are grouped in high frequency bursts composed of action potentials triggered from the delayed depolarization of the preceding action potential. The same kind of bursting can be triggered by intracellular current pulses, indicating that altered neuronal membrane properties are associated with the bursting activity. This type of bursting differs markedly from that seen in motoneurons during penicillin or strychnine-induced spinal seizures.

Action Potentials↗

Subcellular localization of HPA-23 in different rat organs: electron microprobe study.

The anti-viral drug HPA-23 (ammonium 21-tungsto-9-antimonate) has been proposed for use in the combat against AIDS. The two elements tungsten (W) and antimony (Sb) in the molecule enable the intracellular localization and possible breakdown of the product to be studied using electron probe microanalysis methods. Such studies have been carried out after intravenous injection of different doses of HPA-23 in the rat followed by removal of the liver, kidney, thymus, spleen, bone marrow, and lung. HPA-23 was concentrated in the lysosomes and localized in the macrophages of different tissues (thymus, spleen, and bone marrow). The W/Sb ratio was identical in these macrophages. This localization is perhaps relevant to the mechanism of action of HPA-23.

Animals↗

Gastric mucosal injury in the rat. Role of iron and xanthine oxidase.

Recent studies have implicated oxygen free radicals in ischemia-reperfusion injury to the gastric mucosa. The aims of the present study were to test the hypothesis that the enzyme xanthine oxidase is the source of the oxygen radicals in the ischemic stomach and determine the importance of the iron-catalyzed Haber-Weiss reaction in generating the cytotoxic oxygen radicals. Gastric mucosal clearance of 51Cr-labeled red blood cells was measured during a 30-min control period, a 30-min ischemic period (hemorrhage to 25 mmHg arterial pressure), and a 60-80-min reperfusion period (reinfusion of shed blood). In untreated (control) rats, a dramatic rise (100-fold) in the leakage of 51Cr-labeled red blood cells into the gastric lumen was observed only during the reperfusion period. After the reperfusion period, gastric mucosal damage was further assessed using gross lesion area and histology. Rats were placed on a sodium tungstate diet (to inactivate xanthine oxidase), or treated with either deferoxamine (an iron chelating agent) or superoxide dismutase (a superoxide scavenger). All three interventions substantially reduced 51Cr-labeled red blood cell clearance and gross lesion area relative to untreated rats. However, tissue injury assessed histologically was similar in both treated and untreated animals. The results of this study support the hypothesis that oxygen free radicals mediate the hemorrhagic shock-induced extravasation of red blood cells. The data also indicate that xanthine oxidase is the source of the oxy-radicals and that the iron-catalyzed Haber-Weiss reaction is largely responsible for hydroxyl radical generation in this model.

Animals↗

Some observations of a correlation between the symmetry of large heavy-atom complexes and their binding sites on proteins.

Two large, electron dense heavy-metal complexes were found to bind to crystals of heavy riboflavin synthase. The crystallographic analysis by difference Fourier methods shows that the tungsten cluster compound [(W3O2(O2CCH3)6]2+, which has trigonal symmetry, binds to a site on the 3-fold axis of the protein. The heteropolytungstate compound [NaP5W30O110]14-, which has pentagonal symmetry, binds to a site on the 5-fold axis of the protein.

Binding Sites↗

Systematized procedure of crown preparation.

The theoretical considerations of the marginal fit of crowns are discussed and the conventional approach in tooth-cutting procedures is addressed. Gingival marginal surface irregularities described as "peak and valley" and "stepping" effects can leave a cement line in excess of 80 microns after cementation. A regimented method of tooth reduction for metal ceramic crowns, using specially designed rotary cutting instruments, is described. The suggested technique accomplishes the crown preparation more rapidly with a consistent standard of excellence.

Carbon↗

Studies on dental high-speed cutting with carbide burs used on bovine dentin.

Bovine dentin was cut by hand with carbide burs and two cutting procedures while the rotational handpiece speed and the applied load were simultaneously monitored. It became evident that the maximum applied load during intermittent cutting varied from approximately 30 to 40 gm, whereas that during successive cutting ranged from approximately 20 to 60 gm. It showed that intermittent cutting produced a greater cutting effectiveness (for example, cutting volume per cut contact time) when carbide burs were used on dentin, compared with continuous cutting. It is recommended that the intermittent cutting procedure be used during work on patients.

Analysis of Variance↗

Physicochemical characterization and transformation of the cytosolic glucocorticoid receptor from rabbit liver.

Characterization of the glucocorticoid receptor from rabbit liver cytosol was studied in vitro. Binding of [3H]-dexamethasone showed a high affinity (KD = 2.4 . 10(-9)M) and a concentration of binding sites of 0.3 . 10(-12) mol/mg proteins. Association and dissociation rate constants for [3H]-dexamethasone were respectively 1.6 . 10(5)M-1 . min-1 and 2.5 . 10(-4) min-1. Competition experiments showed that dexamethasone and triamcinolone acetonide were the most effective competitors while progesterone and aldosterone competed very poorly. The stabilities of bound and unbound forms were investigated with and without molybdate and other oxyanions. Calibrated gel filtration gave a stokes radius of 5.6 nm and an apparent mol. wt of 280,000. The untransformed complex sedimented at 9 s in 5-20% sucrose density gradients and the transformed species sedimented near 4 s. The behaviour of the three forms of receptor, untransformed, transformed and molybdate-stabilized was studied on ionic exchangers. Transformation of the [3H]-dexamethasone--receptor complex was shown to occur both with high ionic strength and elevated temperature. The transformation step was almost completely inhibited by tungstate and molybdate while vanadate revealed a very slight inhibitory effect.

Animals↗

Influence of phosphatase inhibitors and nucleotides on [3H]dexamethasone binding in cytosol of human placenta.

The purpose of this investigation was to establish the properties of [3H]dexamethasone binding sites in cytosol of human placenta at term. Cytosol containing 20 mM sodium molybdate (MoO4Na2) was incubated for 120 min at 20 degrees C with 40 nM [3H]dexamethasone. The following properties were observed: (a) a single population of binding sites of high affinity and low capacity was measured by Scatchard analysis; (b) potent glucocorticoids such as dexamethasone and cortisol displaced the tritiated ligand, progesterone showed an intermediate activity, whereas cortisone, testosterone and 17 beta-estradiol were ineffective competitors; (c) ultracentrifugation on 16-41% glycerol gradients containing 20 mM MoO4Na2 yielded sedimentation values of 10.25 +/- 0.35 S (n = 4 placentas); (d) the binding sites could be differentiated from the enzyme 11 beta-hydroxysteroid dehydrogenase, as the activity of the former, but not that of the latter, was greatly dependent on the presence of MoO4Na2 in the incubation medium. Inactivation of binding sites labelled with [3H]dexamethasone by incubation at 20 degrees C was prevented by phosphatase inhibitors such as 20 mM MoO4Na2 (P less than 0.01), 20 mM sodium tungstate (WO4Na2) (P less than 0.01) and to a lower extent by 5 mM ATP and cAMP (P less than 0.05). 50 mM NaF, 5 mM GTP or cGMP had no effect. The protection afforded by MoO4Na2 and WO4Na2 was correlated with a significant inhibition of the activity of acid phosphatase, but not alkaline phosphatase. Neither ATP nor cAMP modified phosphatase activity. It is suggested that binding sites for [3H]dexamethasone in cytosol of human placenta showed properties similar to those described for glucocorticoid receptors in target cells, and that these binding sites are regulated by phosphorylation and dephosphorylation mechanisms.

Acid Phosphatase↗

Self-sterilization of the electrosurgery electrode.

Sterilization of the electrode used in electrosurgery has been assumed to occur at the instant of contact of the energized electrode tip to a grounded source. This study evaluated whether the time of electrode activation or the amount of energy delivered is important in the self-sterilization process. Standard bacteriologic culturing techniques were employed to assess sterility of the electrode tip following activation. Total time of application of the current and the energy used were recorded on audiotape and later tabulated. The data indicate that the time of energizing and the energy units used are critical in the sterilization of the electrode tip. These results confirm that self-sterilization of the electrosurgery electrode tip occurs, but full confidence is not realized with the time intervals commonly employed in clinical practice. The electrode tip must be considered a potential source of bacterial contamination of electrosurgically managed dental procedures.

Electric Conductivity↗

Comparative study of the acute lung toxicity of pure cobalt powder and cobalt-tungsten carbide mixture in rat.

Alveolitis progressing to lung fibrosis has been reported in workers exposed to cobalt containing dust (e.g., tungsten carbide-cobalt mixture as produced by the hard metal industry) but rarely following exposure to pure cobalt dust (e.g., in cobalt-producing factories). We have previously demonstrated that tungsten carbide-cobalt mixture is more toxic toward rat alveolar macrophages in vitro than pure cobalt metal powder. The present study was undertaken to compare in female rats the acute pulmonary response (lung weight, lung histology, cellular and biochemical analyses of bronchoalveolar lavage fluid, and mortality) following the intratracheal instillation of pure cobalt (Co) particles (median particle size, d50:4 microns), pure tungsten carbide (WC) particles (d50:2 microns), tungsten carbide-cobalt (WC-Co) powder (d50:2 microns; cobalt 6.3%, tungsten 84%, carbon 5.4%) and crystalline silica (d50 less than 5 micron) used as pneumotoxic reference material. WC alone (15.67 mg/100 g body wt) behaves as an inert dust producing only a mild accumulation of macrophages in the alveolar duct walls. Co alone (1.0 mg/100 g) only causes a moderate inflammatory response. An identical amount of Co given as WC-Co mixture (16.67 mg/100 g; corresponding to 1.0 mg Co/100 g) produces a severe alveolitis and fatal pulmonary edema. Cellular and biochemical characteristics of bronchoalveolar lavage fluid collected 24 hr after the intratracheal instillation of WC (1.0 mg/100 g) or Co (0.06 mg/100 g) are not significantly different from those of control animals instilled with sterile saline. On the contrary, bronchoalveolar lavage fluid changes following administration of the WC-Co mixture (1.0 mg/100 g; corresponding to 0.06 mg Co/100 g) are very similar to those induced by crystalline silica (1.0 mg/100 g). The amount of cobalt excreted in urine is significantly higher when the animals are exposed to WC-Co powder as compared to an equivalent amount of pure cobalt particles, suggesting an increased bioavailability of cobalt metal when combined with tungsten carbide. This study demonstrates that the acute lung toxicity of tungsten carbide-cobalt mixture is much higher than that of each individual component and may explain why lung fibrosis is rarely if ever induced by exposure to pure cobalt dust.

Animals↗

A combination fluorescence assay and Folin-Ciocalteau phenol reagent assay for the detection of paralytic shellfish poisons.

Paralytic shellfish poison (PSP) profiles of crude shellfish extracts were determined by linear gradient elution from a Bio-Rad AG-50-X4 strong cation-exchange resin mini-column. STX, GTX2 and GTX3 were detected by fluorescence assay. NeoSTX and GTX1/GTX4 were detected with a Folin-Ciocalteau phenol reagent assay. The major toxicity associated with extracts of Mytilus edulis and Mya arenaria collected during a 1972 red tide off Hampton, New Hampshire, was due to the presence of GTX1/GTX4, with some activity associated with neoSTX, GTX2 and GTX3. STX was also present. Correlations to mouse toxicity are provided.

Amino Acids↗

Preliminary data on hard metal workers exposure to tungsten oxide fibres.

The first personal exposure data for tungsten oxide fibres, in two hard metal manufacturing industries is presented. Occupational exposure was studied by static and personal measurements during tungsten metal production. Three different initial materials were used, ammonium-paratungstate (APT), blue oxide and tungsten trioxide. In both factories, airborne tungsten oxide fibres were detected both in static and personal samples. Fibre exposure varies for different activities. Charging of raw material, changing the hammer, cleaning-work on the cyclone and the micro-filter were found to be among the most high dusty operations. However, as workers use respirators during cleaning operations, these filters could not be related to personal exposure. The calcination of APT to the blue oxide generates fibrous dust. The raw material imported as blue oxide is also fibrous material and both charging it into the calcination furnace and re-charging it into the reduction furnace generates unnecessarily additional dusty periods. A single reduction-stage is, therefore, preferable. Furthermore, the tungsten trioxide raw material is non-fibrous, therefore, calcination of APT to tungsten trioxide and its reduction to tungsten metal is preferable with respect to minimising workers exposure to tungsten oxide fibres.

Air Pollutants, Occupational↗