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Biomedical subjects

R W Bryant

Publications and source records attributed to R W Bryant.

At least 55 records · Page 3Linked to original sources
PubMed1986

Standardizing evaluations of the clinical marginal fracture of amalgam.

In this study, we investigated both inter-examiner and intra-examiner differences in the evaluation of the marginal fracture characteristics of dental amalgam restorations. Evaluations were made by matching intra-oral photographs of the occlusal surfaces of clinical restorations to a reference set of clinical photographs illustrating increasing amounts of marginal fracture. Four amalgam alloys and five examiners provided the data base. Differences in marginal fracture indices were found among examiners and between evaluation times for the same examiners. However, these differences were reduced significantly after application of a standardization procedure.

Dental Alloys↗
PubMed1985

Lipoxygenase products of arachidonic acid modulate biosynthesis of platelet-activating factor (1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) by human neutrophils via phospholipase A2.

When human neutrophils, previously labeled in their phospholipids with [14C]arachidonate, were stimulated with the Ca2+-ionophore, A23187, plus Ca2+ in the presence of [3H]acetate, these cells released [14C]arachidonate from membrane phospholipids, produced 5-hydroxy-6,8,11,14-[14C]eicosatetraenoic acid (5-HETE) and 14C-labeled 5S,12R-dihydroxy-6-cis,8,10-trans, 14-cis-eicosatetraenoic acid ([14C]leukotriene B4), and incorporated [3H]acetate into platelet-activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine). Ionophore A23187-induced formation of these radiolabeled products was greatly augmented by submicromolar concentrations of exogenous 5-hydroperoxy-6,8,11,14-eicosatetraenoic acid (5-HPETE), 5-HETE, and leukotriene B4. In the absence of ionophore A23187, these arachidonic acid metabolites were virtually ineffective. Nordihydroguaiaretic acid (NDGA) and several other lipoxygenase/cyclooxygenase inhibitors (butylated hydroxyanisole, 3-amino-1-(3-trifluoromethylphenyl)-2-pyrazoline and 1-phenyl-2-pyrazolidinone) caused parallel inhibition of [14C]arachidonate release and [3H]PAF formation in a dose-dependent manner. Specific cyclooxygenase inhibitors, such as indomethacin and naproxen, did not inhibit but rather slightly augmented the formation of these products. Furthermore, addition of 5-HPETE, 5-HETE, or leukotriene B4 (but not 8-HETE or 15-HETE) to neutrophils caused substantial relief of NDGA inhibition of [3H]PAF formation and [14C]arachidonate release. As opposed to [3H]acetate incorporation into PAF, [3H]lyso-PAF incorporation into PAF by activated neutrophils was little affected by NDGA. In addition, NDGA had no effect on lyso-PAF:acetyl-CoA acetyltransferase as measured in neutrophil homogenate preparations. It is concluded that in activated human neutrophils 5-lipoxygenase products can modulate PAF formation by enhancing the expression of phospholipase A2.

Acetates↗
PubMed1985

Leukotriene formation by a purified reticulocyte lipoxygenase enzyme. Conversion of arachidonic acid and 15-hydroperoxyeicosatetraenoic acid to 14, 15-leukotriene A4.

The purified lipoxygenase of rabbit reticulocytes converts arachidonic acid at 0 degrees C to 15-hydroperoxyeicosatetraenoic acid (15-HPETE) and to 12-hydroperoxyeicosatetraenoic acid (12-HPETE) via reactions which involve hydrogen abstraction at C-13 and C-10, respectively. At 37 degrees C the enzyme converts arachidonic acid to additional products which were identified as 13-hydroxy-14,15-epoxy-5,8,11-eicosatrienoic acid, 8,15-dihydroperoxy-5,9,11,13- and 5,15-dihydroperoxy-6, 6,8,11,13-eicosatetraenoic acids (8,15-diHPETE and 5,15-HPETE, respectively) and diastereoisomers of 8,15-dihydroxy-5,9,11,13-eicosatetraenoic acid (8,15-diHPETEs). The 8,15- and 5,15-diHPETEs were formed by double lipoxygenation since each incorporated 2 molecules of 18O2 and since their synthesis from 15-HPETE was blocked under anaerobic conditions. The 8,15-diHETEs each incorporated 18O from 18O2 at C-15 and were found to arise from nonenzymatic hydrolysis of an epoxytriene which was identified as 14,15-leukotriene A4 by trapping in acidic methanol. This compound was a major product of 15-HPETE in anaerobic incubations. The conversion of 15-HPETE to 14,15-leukotriene A4 was inhibited by the lipoxygenase inhibitors nordihydroguairetic acid and 5,8,11,14-eicosatetraynoic acid. The 14,15-leukotriene A4 synthase and 15-lipoxygenase activities were inhibited by 5,8,11,14-eicosatetraynoic acid in a similar time-dependent manner. The results support a mechanism whereby 14,15-leukotriene A4 is synthesized from 15-HPETE by a further enzymatic step carried out by the reticulocyte 15-lipoxygenase via hydrogen abstraction at C-10 and a redox cycle of the non-heme iron atom of the enzyme.

Animals↗
PubMed1985

Ionophore-stimulated lipoxygenase activity and histamine release in a cloned murine mast cell, MC9.

A cloned murine mast cell line designated MC9 expresses a 5-lipoxygenase activity when stimulated with the ionophore A23187. Upon addition of 0.5 microM ionophore, MC9 cells produce 270 +/- 43 pmoles 5-HETE, 74 +/- 40 pmoles 5,12 diHETEs and 65 +/- 31 pmoles LTC4/10(6) cells from 37 microM exogenously added [1-14C]arachidonic acid in two minutes. 5-HETE and 5,12-diHETES, including LTB4 were identified by GC/MS whereas LTC4 was confirmed by HPLC mobility, bio-assay, RIA and enzymatic transformation. The principal cyclooxygenase products were PGD2 and TxB2 (8.5 +/- 2.4 and 5.4 +/- 1.2 pmoles/10(6) cells respectively). Prostanoids were identified by comigration with authentic standards on two-dimensional thin layer chromatograms. Production of arachidonic acid lipoxygenase metabolites stimulated with ionophore proved relatively insensitive to removal of extracellular Ca+2 and chelation by EGTA. In addition, MC9 5-lipoxygenase required only low micromolar amounts of exogenous arachidonic acid for maximal activity. Whereas production of arachidonic acid metabolites lasted only two to five minutes, histamine release stimulated with ionophore was not initiated until 5 minutes (12 +/- 3% cellular histamine) and continued for 30 minutes (37 +/- 7% cellular histamine). Although these cells metabolize arachidonic acid differently from the classic peritoneal-derived mast cell, they resemble subpopulations found in certain tissues (such as mucosa) and should be useful in understanding the biochemistry of mast cell mediator release.

Animals↗
PubMed1985

Aspirin enhances the sensitivity of human platelet 12-lipoxygenase to inhibition by 15-HETE, an endogenous regulator.

Human platelets metabolize arachidonic acid via cyclooxygenase (E.C. 1.14.99.1) to thromboxane A2 and the 12-lipoxygenase to 12-hydroxyeicosatetraenoic acid (12-HETE). Aspirin inhibits cyclooxygenase while the neutrophil product 15-Hydroxyeicosatetraenoic acid (15-HETE) is a selective inhibitor of platelet 12-lipoxygenase. The unexpected observation was made that the platelet 12-lipoxygenase of individuals who had ingested aspirin showed up to a twenty-fold increase in sensitivity to inhibition by 15-HETE. This observation was confirmed in platelets treated with aspirin in vitro. Aspirin pretreatment consistently resulted in a decrease in the I50 for 15-HETE from an average of 21.5 microM to only 5.2 +/- 1.5 microM, indicating a probable interaction between the cyclooxygenase and lipoxygenase pathways.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1985

The reaction zone around experimental amalgam alloy particles of Ag-Cu eutectic composition.

An electron microprobe was used to investigate the composition and microstructure of the reaction zone occurring around experimental particles of approximately Ag-Cu eutectic composition in two experimental dental amalgams. This study formed the basis for the examination, described in a subsequent paper, of the reaction zones in several commercially available amalgams. Techniques of phase composition analysis were further developed in order to identify and quantify the component phases in the regions of the reaction zone. The clumped, outer, more darkly shaded region of the zone contained predominantly eta 1 (Cu6Sn5) phase. The inner, more lightly shaded region of the reaction zone was found to contain substantial quantities of eta 1, gamma 1, a Cu-Hg phase and a Ag-Cu component.

Chemical Phenomena↗
PubMed1985

The reaction zone around commercially available amalgam alloy particles of Ag-Cu eutectic composition.

An electron microprobe was used to investigate the composition and microstructure of regions in the reaction zones around particles of approximately Ag-Cu eutectic composition in commercially available and experimental dispersion modified amalgams. Two distinct regions were identified. Techniques of phase composition analysis were further developed in order to identify and quantify the component phases present in the two regions. The outer, somewhat clumped region contained principally a network of strands and particles of eta 1 (Cu6Sn5) composition and some gamma 1 phase. After phase composition analysis, the inner region was found to contain eta 1 phase and substantial quantities of gamma 1 phase, a Cu-Hg phase (probably Cu4Hg3 formulation) and unreacted Ag-Cu. The two regions may be best described as eta 1-rich and eta 1-sparse, respectively.

Chemical Phenomena↗
PubMed1984

Microprobe analysis of dental amalgam alloy particles of Ag-Cu eutectic composition.

An electron microprobe was used to investigate the composition and microstructure of commercially available, and experimental dental amalgam alloy particles of approximately Ag-Cu eutectic composition. The method of alloy and amalgam specimen preparation and the technique used in microprobe analysis have been briefly outlined. The dispersant particles in one commercial alloy were of Ag-Cu eutectic composition (28.1% Cu), in another commercial alloy they were Cu-rich (31% Cu). Although the dispersant particles were predominantly of uniform appearance, there was evidence in some particles of an internal structure pattern, small spherical inclusions or voids. The possible composition of the Cu-rich inclusions was investigated using particles from an experimental alloy of approximately Ag-Cu eutectic composition.

Chemical Phenomena↗
PubMed1984

Inhibition of phytohemagglutinin-induced lymphocyte mitogenesis by lipoxygenase metabolites of arachidonic acid: structure-activity relationships.

Phytohemagglutinin (PHA)-induced mitogenesis of mixed mouse spleen lymphocyte populations, measured by [3H]thymidine incorporation, was completely inhibited by micromolar concentrations of certain hydroxyeicosatetraenoic acids (HETE's). These are lipoxygenase metabolites of arachidonic acid which are synthesized in considerable concentrations by macrophages, lymphocytes, and other components of the immune system when appropriately stimulated. In the studies described here, the structural requirements for the maximum antimitogenic activities were examined. A series of monohydroxylated HETE's were prepared using a singlet oxygen photochemical procedure or by enzymatic synthesis from arachidonic acid substrate, and isolated by HPLC. Isomers containing different numbers of double bonds were synthesized using the appropriate unsaturated fatty acid as substrate, and the functional importance of the OH and carboxylic functions was tested using various acetoxy- and carbomethoxy derivatives. A serum-free mitogenesis assay system was used for testing, which minimized binding of the fatty acids by serum proteins and increased the inhibitory potency of the various HETE's several-fold. It was found that inhibition of cell proliferation was related to: hydroxyl proximity to the center of the eicosatetraenoic acid, decreasing in the order: 9 greater than 11 greater than 12 greater than 15 greater than 8 much greater than 5; the number of double bonds in the fatty acid chain, decreasing in the order: 15-OH, 20:4 greater than 15-OH 20:3 much greater than 15-OH, 20:2 much greater than 15-OH, 20:0; and the 15-position functional group as well as the 1-carboxylic group, decreasing in the order: 15-hydroxy, 1-carboxylic greater than 15-acetoxy, 1-carboxylic much greater than 15-hydroxy, 1-carbomethoxy greater than 15-acetoxy, 1-carbomethoxy.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Hydroperoxy fatty acid formation in selenium deficient rat platelets: coupling of glutathione peroxidase to the lipoxygenase pathway.

After incubation with [1-14C]-arachidonic acid, washed platelets from selenium deficient rats produced a sevenfold greater amount of 12-hydroperoxytetraenoic acid than platelets from control animals. When stimulated with either arachidonic acid or t-butyl-hydroperoxide, antimycin-A1 treated platelets from the deficient rats also converted markedly lower amounts of [1-14C]-glucose to [14C]-CO2 than platelets from control rats. These results indicate a significant role for platelet selenium-dependent glutathione peroxidase in the enzymatic reduction of platelet-produced hydroperoxides.

Animals↗
PubMed1983

Development of a radioimmunoassay for 15-HETE and its application to 15-HETE production by reticulocytes.

Mono-hydroxy-eicosatetraenoic acids (HETE's) are frequently the principal lipoxygenase-derived products in a number of cell types. This paper describes the development of a selective and sensitive radioimmunoassay procedure for 15-HETE, a metabolite which has previously been shown to be both an activator and inhibitor of leukotriene formation in various cells. Initially, rabbits were immunized with 15-HETE conjugated to bovine serum albumin. After seven months, the anti-plasma showed significant binding of tritiated 15-HETE (40-45% binding with a 1:600 dilution of the anti-plasma) which was displaceable by cold 15-HETE. The sensitivity of the assay was approximately 20 pg. of 15-HETE. The anti-plasma exhibited very little (less than 1%) cross-reactivity with arachidonic acid, 5-, 8-, 9-, 11- and 12-HETE's, HHT, TXB2, PGE2 and 6-Keto-PGF1 alpha. Significant cross-reactivity was observed with 5,15-diHETE (53%), 8, 15-diHETE (6.6%), and several other 15-hydroxy-eicosanoids. Rabbit reticulocytes have a very active 15S-lipoxygenase and converted arachidonic acid (final concentration 7 microM) principally to 15-HETE. Unstimulated reticulocytes were found to release negligible amounts of 15-HETE as determined by radioimmunoassay. Treatment of these cells with the calcium ionophore A23187 (0.16 to 4.0 micrograms/ml) elicited a level of 15-HETE release (8 - 14 ng/ml) that was twenty to forty times less than that obtained with exogenous arachidonic acid (2.5 micrograms/ml). The radioimmunoassay reported here may be useful for identifying factors which stimulate cellular release of 15-HETE and other 15-hydroxy-eicosanoids from endogenous arachidonic acid.

Animals↗