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Biomedical subjects

N Stanford

Publications and source records attributed to N Stanford.

13 recordsLinked to original sources
PubMed1990

A single TLD dose algorithm to satisfy federal standards and typical field conditions.

Modern whole-body dosimeters are often required to accurately measure the absorbed dose in a wide range of radiation fields. While programs are commonly developed around the fields tested as part of the National Voluntary Accreditation Program (NVLAP), the actual fields of application may be significantly different. Dose algorithms designed to meet the NVLAP standard, which emphasizes photons and high-energy beta radiation, may not be capable of the beta-energy discrimination necessary for accurate assessment of absorbed dose in the work environment. To address this problem, some processors use one algorithm for NVLAP testing and one or more different algorithms for the work environments. After several years of experience with a multiple algorithm approach, the Dosimetry Services Group of Yankee Atomic Electric Company (YAEC) developed a one-algorithm system for use with a four-element TLD badge using Li2B4O7 and CaSO4 phosphors. The design of the dosimeter allows the measurement of the effective energies of both photon and beta components of the radiation field, resulting in excellent mixed-field capability. The algorithm was successfully tested in all of the NVLAP photon and beta fields, as well as several non-NVLAP fields representative of the work environment. The work environment fields, including low- and medium-energy beta radiation and mixed fields of low-energy photons and beta particles, are often more demanding than the NVLAP fields. This paper discusses the development of the algorithm as well as some results of the system testing including: mixed-field irradiations, angular response, and a unique test to demonstrate the stability of the algorithm. An analysis of the uncertainty of the reported doses under various irradiation conditions is also presented.

Algorithms↗
PubMed1979

Prevention of thrombosis in patients on hemodialysis by low-dose aspirin.

Since platelet cyclo-oxygenase is much more sensitive to inactivation by aspirin than is the enzyme in the arterial wall and low doses of aspirin may prevent thrombosis by blocking thromboxane synthesis, we conducted a randomized, double-blind trial of aspirin (160 mg per day) vs. placebo in 44 patients on chronic hemodialysis. The study was continued until there were 24 patients with thrombi and both groups had been under observation for a mean of nearly five months. Thrombi occurred in 18 of 25 (72 per cent) of patients given placebo and 16 of 19 (32 per cent) of those given aspirin (P less than 0.01). The incidence of thrombosis was reduced from 0.46 thrombi per patient month in the placebo group to 0.16 thrombi per patient month in the aspirin group (p less than 0.005). A dose of 160 mg of aspirin per day is an effective, nontoxic antithrombotic regimen in patients on hemodialysis.

Acetylation↗
PubMed1979

Diglyceride lipase: a pathway for arachidonate release from human platelets.

We provide evidence that the mechanism for arachidonate release from stimulated human platelets involves two enzymes: a phosphatidylinositol-specific phospholipase C (EC 3.1.4.10) and a diglyceride lipase. After incubation of platelets with thrombin for 15 seconds, 1.2 nmol of 1-stearoyl-2-arachidonoyl diglyceride per 10(9) platelets, was isolated. Arachidonate was released from this substrate by the action of diglyceride lipase located in the particulate fraction of platelets. The enzyme has a pH optimum of 7.0, is stimulated by calcium ions and reduced glutathione, and liberates 31 nmol of fatty acid per min per mg of platelet particulate protein. The diglyceride lipase has sufficient activity to account for the 5-10 nmol of arachidonate released per 10(9) platelets upon thrombin stimulation. That only arachidonate is released upon thrombin stimulation may be explained by the fact that the diglyceride substrate in platelets contains only arachidonate in the 2 position. The lipase activity found in platelet membranes can also hydrolyze the 1-position fatty acid. Stearate is not released when intact platelets are stimulated with thrombin, and the fate of this fatty acid remains to be elucidated.

Arachidonic Acids↗
PubMed1979

Deficiency of factor Xa-factor Va binding sites on the platelets of a patient with a bleeding disorder.

Factor V (Va) is essential for binding of factor Xa to the surface of platelets. After thrombin treatment, normal platelets release at least five times more factor Va activity than is required for maximal factor Xa binding. The concentration of factor V activity obtained after thrombin stimulation of 10(7) normal platelets is sufficient to allow half-maximal factor Xa binding to 10(8) platelets (10% normal, 90% factor-V deficient). Therefore, factor Va activity is not limiting in platelet-surface factor Xa binding and prothrombin activation in normal platelets; some other components limit the number of binding sites. We report studies of a patient (M.S.) with a moderate to severe bleeding abnormality whose platelets are deficient in the platelet-surface component required for the factor Va-factor Xa binding. The patient's platelet factor Va activity released after thrombin treatment is normal, but factor Xa binding is 20%-25% of control values at saturation. Abnormal prothrombin consumption in a patient with normal plasma coagulation factors and platelet function suggests a disorder in platelet-surface thrombin formation.

Binding Sites↗
PubMed1978

Sensitivity of fatty acid cyclooxygenase from human aorta to acetylation by aspirin.

The rate of acetylation of fatty acid cyclooxygenase (prostaglandin synthase, EC 1.14.99.1) by [acetyl-3H]-aspirin was measured in microsomes from human aortas and coronary arteries and intact and disrupted human platelets. We also measured the inhibition by aspirin of prostacyclin generation from exogenous arachidonic acid in shredded human aorta. Cyclooxygenase in human aorta and coronary artery microsomes is approximately 1/250th as sensitive to aspirin as enzyme in intact platelets, and 1/60th as sensitive to aspirin as enzyme measured in a platelet microsomal preparation. On the basis of the in vitro data presented, we predict that small oral doses of aspirin are sufficient to inhibit platelet prostaglandin production but are not sufficient to substantially affect aorta or coronary artery prostaglandin production.

Acylation↗
PubMed1978

Inhibition of platelet prostaglandin synthetase by oral aspirin.

Aspirin inhibits platelet function by permanently acetylating the cyclooxygenase that forms prostaglandins. We determined the sensitivity of platelets to aspirin in normal subjects by measuring [3H-acetyl]aspirin-susceptible cyclooxygenase in washed platelets obtained at various times after aspirin ingestion. A single 325-mg aspirin dose inactivated 89% of platelet cyclooxygenase. The inhibition persisted for 2 days suggesting that oral aspirin also inactivated megakaryocyte cyclooxygenase. Thereafter, active enzyme returned with a time-course reflecting platelet turnover (life-span 8.2+/-2 days). Single doses of 20-650 mg aspirin resulted in 34- greater than 95% inhibition after 24 h. Daily doses of 20-325 mg aspirin for brief periods produced 61- greater than 95% inactivation when measured 24 h after cessation of the drug. Platelet cyclooxygenase is more sensitive to inactivation by aspirin than enzyme in sheep seminal vesicles.

Acetylation↗
PubMed1977

Acetylation of prostaglandin synthetase by aspirin. Purification and properties of the acetylated protein from sheep vesicular gland.

We previously presented evidence that aspirin (acetylsalicylic acid) inhibits prostaglandin synthetase by acetylating and active site of the enzyme. In the current work, we have labeled the enzyme from an aceton-pentane powder of sheep vesicular gland using [acetyl-3H]aspirin and purified the [3H]acetyl-protein to near homogeneity. The final preparation contains protein of a single molecular weight (85 000) and an amino-terminal sequence of Asp-Ala-Gly-Arg-Ala. The [3H]acetyl-protein contained 0.5 mol of acetyl residues per mol of protein based on amino acid composition but only a single sequence was found.

Acetylation↗
PubMed1977

Lack of covalent modification of prostaglandin synthetase (cyclo-oxygenase) by indomethacin.

We have previously shown that aspirin irreversibly inhibits prostaglandin synthetase (cyclo-oxygenase) by acetylating the active site of the enzyme. By utilizing 14C-labeled indomethacin and a close analogue, we now show that indomethacin, unlike aspirin, does not covalently modify cyclo-oxygenase. Furthermore, indomethacin binding to the enzyme may be reversible since even though indomethacin can inhibit acetylation by aspirin, when enzyme inhibited by indomethacin (1 micronM) is treated with 200 micronM aspirin 3 times for 1 hour each, complete acetylation of cyclo-oxygenase is achieved.

Acetylation↗
PubMed1977

Comparative effects of aspirin and diflunisal on prostaglandin synthetase from human platelets and sheep seminal vesicles.

1. We have compared the effects of diflunisal on prostaglandin (PG) synthetase from human platelets and sheep seminal vesicles (SSV) by measuring malonaldehyde, PG products, and rabbit aorta contraction. 2. Aspirin inhibits PG synthetase by covalently acetylating the enzyme. Inhibition is dependent on time and aspirin concentration. Aspirin is 37-fold more potent in inhibiting enzyme in human platelets compared with enzyme from SSV. 3. Diflunisal inhibits PG synthetase from intact human platelets and SSV equally (50% inhibition at 3-5 micronM). The drug does not covalently modify the enzyme, and inhibition is not time dependent. Diflunisal probably acts at a site similar to aspirin since the drug will inhibit acetylation of the enzyme by aspirin. 4. The results suggest that diflunisal is relatively less inhibitory to platelet function than is aspirin.

Animals↗
PubMed1977

Cyclic adenosine 3',5'-monophosphate inhibits the availability of arachidonate to prostaglandin synthetase in human platelet suspensions.

When thrombin is added to washed human platelets, one of its actions results in activation of a phospholipase that hydrolyzes arachidonic acid from phospholipids. The arachidonate is converted to the cyclic endoperoxides (prostaglandin G2 and prostaglandin H2) by fatty acid cyclo-oxygenase. These compounds are then converted to thromboxane A2, also called rabbit aorta-contracting substance, by thromboxane synthetase. These labile, pharmacologically active compounds then break down to inactive products including thromboxane B2 and malonaldehyde. Incubation of platelets with either dibutyryl cyclic adenosine 3',5'-monophosphate (dBcAMP) or prostaglandin E1 (PGE1) before thrombin addition blocks the subsequent formation of oxygenated products of arachidonic acid including thromboxane A2, thromboxane B2, and malonaldehyde. In contrast, when arachidonic acid is added directly to platelets, prior incubation with dBcAMP or PGE1 does not inhibit production of the prostaglandins or their metabolites. Thrombin treatment of platelets also blocks the acetylation of cyclo-oxygenase by aspirin since the hydrolyzed arachidonic acid competes with aspirin for the active site on cyclo-oxygenase. Prior treatment of platelets with dBcAMP or PGE1 reverses the thrombin inhibition of the acetylation of cyclo-oxygenase. We conclude that agents which elevate platelet cAMP levels inhibit the hydrolysis of arachidonic acid from platelet phospholipids. We also find that prostaglandin synthesis can be dissociated, in part, from platelet aggregation and release, and that cAMP has separate actions on these processes. Higher thrombin concentrations are required to stimulate prostaglandin synthesis (0.05-2 U/ml) than are required to induce [14C]serotonin release (0.02-0.1 U/ml). Furthermore, dBcAMP and PGE1 both inhibit platelet aggregation induced by either arachidonic acid or prostaglandin H2 without affecting the production of prostaglandin metabolites from these compounds.

Arachidonic Acids↗
PubMed1975

Acetylation of prostaglandin synthase by aspirin.

When microsomes of sheep or bovine seminal vesicles are incubated with [acetyl-3H]aspirin (acetyl salicylic acid), 200 Ci/mol, we observe acetylation of a single protein, as measured by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The protein has a molecular weight of 85,000 and corresponds to a similar acetylated protein found in the particulate fraction of aspirin-treated human platelets. The aspirin-mediated acetylation reaction proceeds with the same time course and at the same concentration as does the inhibition of prostaglandin synthase (cyclo-oxygenase) (EC 1.14.99.1; 8,11,14-eicosatrienoate, hydrogen-donor:oxygen oxidoreductase) by the drug. At 100 muM aspirin, 50% inhibition of prostaglandin synthase and 50% of maximal acetylation are observed after 15 min at 37 degrees. Furthermore, the substrate for cyclo-oxygenase, arachidonic acid, inhibits protein acetylation by aspirin at concentrations (50% inhibition at 10-30 muM) which correlate with the Michaelis constant of arachidonic acid as a substrate for cyclooxygenase. Arachidonic acid analogues and indomethacin inhibit the acetylation reaction in proportion to their effectiveness as cyclo-oxygenase inhibitors. The results suggest that aspirin acts as an active-site acetylating agent for the enzyme cyclo-oxygenase. This action of aspirin may account for its anti-inflammatory and anti-platelet action.

Acetylation↗
PubMed1975

Thrombin-induced protein phosphorylation in human platelets.

Intact human platelets loaded with 32PO4 contain multiple phosphorylated proteins. Thrombin treatment of intact 32PO4-loaded platelets results in a 2-6-fold increase in phosphorylation of a platelet protein (designated "peak 7" protein) of approximately 40,000 mol wt as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis and by gel filtration on Sephadex G-150. A similar increase in phosphorylation was observed in a platelet protein (designated "peak 9" protein) of approximately 20,000 mol wt. The time for half-maximal phosphorylation of peak 7 and peak 9 protein was 10-14 s. The concentration of thrombin at half-maximal phosphorylation was 0.25 U/ml for both proteins. Prior incubation of platelets with dibutyryl cyclic adenosine 3',5'-monophosphate or prostaglandin E1 inhibited thrombin-induced peak 7 and peak 9 protein phosphorylation. The erythroagglutinating phytohemagglutinin of Phaseolus vulgaris, a non-proteolytic release-inducing agent, induced peak 7 and peak 9 protein phosphorylation. Thus, the characteristics of peak 7 and peak 9 protein phosphorylation are similar to those of the platelet release reaction, suggesting that the phosphorylation of these proteins may play a role in the platelet release reaction. When platelet sonicates or the supernatant fraction from platelet sonicates were incubated with [gamma-32P]ATP there was phosphorylation of both peak 7 and peak 9 proteins. This phosphorylation was unaffected by either added thrombin or adenosine 3',5'-cyclic monophosphate (cAMP) despite the presence of the phosphodiesterase inhibitor 1-methyl-3-isobutylxanthine. Thus, the thrombin-dependent phosphorylation depends upon intact platelets. When the supernatant fraction from platelet sonicates was fractionated by histone-Sepharose affinity chromatography, two distinct protein kinase enzymes were resolved, one a cAMP-dependent holoenzyme and the other a cAMP-independent enzyme. The isolated cAMP-dependent enzyme fraction catalyzed the cAMP-(but not thrombin-) stimulated phosphorylation of a protein that co-electrophoresed with peak 7 protein.

Animals↗