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Biomedical subjects

W A Scott

Publications and source records attributed to W A Scott.

At least 109 records · Page 6Linked to original sources
PubMed1983

Choice reaction time. A method of measuring postoperative psychomotor performance decrements.

Postoperative psychomotor performance changes were measured in three groups of thirteen subjects using a portable serial four-choice reaction time apparatus. Anaesthesia consisted of either thiopentone, nitrous oxide and halothane (thiopentone subjects) or methohexitone and fentanyl/nitrous oxide and incremental methohexitone (methohexitone subjects). Tests were performed on the day before operation and after 2, 4, 6, 8 and approximately 24 hours recovery on both groups of patients and at approximately the same times on nonoperated control patients. Control subjects' mean four-choice reaction time decreased steadily with practice such that the difference between first and last tests was statistically significant. Mean four-choice reaction times of thiopentone and methohexitone subjects increased from 515 to 550 ms and 552 to 600 ms respectively after 2 hours recovery. Subsequently four-choice reaction times decreased in both groups although methohexitone subjects were still significantly slower than controls after 6 hours. Thiopentone subjects were slower than controls at 2, 4 and 8 hours recovery. There was no difference between the three groups of subjects after 24 hours recovery.

Abortion, Induced↗
PubMed1983

Nuclease-sensitive sites in the two major intracellular simian virus 40 nucleoproteins.

Simian virus 40 nucleoprotein isolated from the nuclei of infected cells contains a nuclease-sensitive site adjacent to the viral origin of replication (between 0.66 and 0.73 map unit). Nuclear extracts were subfractionated by sucrose gradient centrifugation to yield provirions (200S) and simian virus 40 chromatin (80S). The 80S fraction was cleaved either by DNase I or by an endonuclease endogenous to BSC-1 cells with high preference for the 0.66 to 0.73 region. The 200S fraction was treated to release core particles that were sensitive to nuclease cleavage; however, DNase I showed little or no preference for the 0.66 to 0.73 region of the provirion core nucleoprotein.

Chromatin↗
PubMed1983

Chromatin structure of simian virus 40-pBR322 recombinant plasmids in COS-1 cells.

To study the nucleoprotein structure formed by recombinant plasmid DNA in mammalian cells, nuclei were isolated from COS-1 cells after transfection with a recombinant (pJI1) containing pBR322 sequences and a segment of simian virus 40 containing information for a nuclease-sensitive chromatin structure. The nuclei were incubated with DNase I. DNA fragments which were the size of linear pJI1 DNA were isolated, redigested with restriction enzymes, fractionated by electrophoresis, and detected by hybridization with nick-translated segments prepared from the plasmid DNA. Two DNase I-sensitive sites were detected in the simian virus 40 portion of the plasmid at the same sites that were DNase I sensitive in simian virus 40 chromatin prepared late after infection of African green monkey kidney (BSC-1) cells. One site extended from the viral origin of replication to approximately nucleotide 40. The 21-base pair repeated sequences were relatively DNase I resistant. A second site occurred over the single copy of the 72-base pair segment present in this plasmid. These results indicate that the nuclease-sensitive chromatin structure does not depend on the presence of viral structural proteins. In addition, late viral proteins added to pJI1-transfected COS-1 cells by superinfection with simian virus 40 caused no change in the distribution of DNase I-sensitive sites in plasmid chromatin. Analysis of transfected plasmid DNA may provide a general method applicable to the study of the chromatin structure of cloned segments of DNA.

Animals↗
PubMed1983

Leukotriene C release by macrophages.

Leukotriene C (LTC) and its metabolites leukotriene D and leukotriene E collectively make up the biological activity known as slow-reacting substance of anaphylaxis. Murine macrophages are potent sources of LTC (5(S)-hydroxy-6(R)-gamma-glutamylcysteinylglycyl-7,9-trans-11,14-cis-eicosatetr aenoic acid). Peritoneal and pulmonary tissue macrophages synthesize LTC and other arachidonic acid (20:4) metabolites in response to inflammatory stimuli such as unopsonized zymosan and IgG immune complexes. Peritoneal macrophages, in addition, release 20:4 when challenged with IgE immune complexes. These results suggest that macrophages may be a major source of leukotrienes in acute inflammation and also in immediate-type hypersensitivity reactions.

Animals↗
PubMed1982

Secretion of leukotriene C and other arachidonic acid metabolites by macrophages challenged with immunoglobulin E immune complexes.

Resident mouse peritoneal macrophages release the slow-reacting substance leukotriene C (LTC) on exposure to particulate IgE immune complexes. Because these cells lose their responsiveness to an IgE stimulus after 4 h in culture, maximum release of 20:4 metabolites is observed before this time. However, a similar diminution in 20:4 metabolism was not observed with a zymosan stimulus. Freshly explanted cells are deficient in intracellular glutathione (GSH) (12.4 +/- 0.4 pmol/micrograms cell protein), but GSH increases to a steady state value of 30-35 pmol/micrograms of cell protein between 3 and 9 h of culture. Because GSH is required for the synthesis of LTC and prostaglandin (PG)E2, cultures challenged immediately after explanation have a diminished capacity to synthesize these 20:4 metabolites and release prostacyclin as the major product. By 4-5 h in culture, macrophages form significant amounts of LTC and PGE2. Under optimum conditions of maximum responsiveness to an IgE stimulus and GSH content (after 4 h of culture), macrophages challenged with latex beads coated with IgE immune complexes synthesize 1.0 +/- 0.3 pmol of LTC/microgram cell protein (60 +/- 18 pmol/10(6) cells) in addition to prostacyclin (8.2 +/- 0.8 pmol/micrograms cell protein) and PGE2 (4.7 +/- 1.5 pmol/micrograms cell protein). These amounts are quantitatively similar to the arachidonic acid metabolites produced by macrophages challenged with IgG immune complex-coated latex beads or zymosan. These data demonstrate that macrophages produce large quantities of LTC and other 20:4 metabolites in response to particle-bound IgE and antigen, provided that the appropriate in vitro conditions are met. The macrophage might, therefore, be a major source of slow-reacting substance and other 20:4 metabolites generated during IgE-mediated reactions in vivo.

Animals↗
PubMed1982

Requirements of delta 9 and delta 12 fatty acid desaturation in Neurospora.

Microsomes prepared from the wild-type strain and lipid auxotrophs of Neurospora were analyzed for delta 9 - (stearoyl-CoA) and delta 12 - (oleoyl-CoA) desaturase activities. The wild-type delta 9-desaturase was found to have a 20-fold higher specific activity and 2-fold lower activation energy than the delta 12-desaturase. In addition, delta 12-desaturase had higher Km app values for oleoyl-CoA and for NADH than the equivalent values for delta 9-desaturase. These properties were correlated with a rate-limiting role of delta 12-desaturase in the production of 18:2, the major fatty acid of Neurospora. The delta 12-desaturase also exhibited a higher tolerance to pH changes and to cyanide than did the delta 9-desaturase. Both activities could be measured in the same reaction mixture using stearoyl-CoA as the substrate, indicating a coupling of the two enzymes. Enrichment of cellular membranes of the wild-type Neurospora with 18:0 and 18:1, 18:2, 18:3 fatty acids led to the conclusion that the presence of excess substrate in the membrane induces activation of the appropriate desaturase. These experiments also suggested that the membrane fluidity, as determined by the degree of unsaturation of membrane fatty acids, may influence the activities of the desaturating enzymes. Perturbation of the polar head groups of the membrane phospholipids indicated that the correct composition of anionic phospholipids is an absolute requirement for the function of both desaturases. These studies show that the activities of the delta 9-desaturase and the delta 12-desaturase are regulated by a variety of factors and that the delta 12-desaturase is subjected to less stringent controls than the delta 9-desaturase.

Cyanides↗
PubMed1982

Uptake and metabolism of monohydroxy-eicosatetraenoic acids by macrophages.

Within 5 min, resting macrophages metabolize microM quantities of exogenous arachidonic acid (20:4) to cyclooxygenase and lipoxygenase products. Mono-HETEs represent a major class of metabolites recovered from the medium. However, the quantity of mono-Hetes progressively decreases over a 60-min incubation period, with a concomitant increase in more polar lipoxygenase products, suggesting additional metabolic fates for these hydroxy acids. This was directly confirmed by exposing resident macrophage cultures to radiolabeled 15-, 12-, and 5-HETEs (1 microM). 12-30% of the recovered HETEs were cell-associated and predominantly esterified into phospholipid. High pressure liquid chromatography analyses of medium extracts indicated that 50% of each HETE was also converted to 10 or more metabolites over a 60-min time-course, a rate slower than for 20:4. The major metabolite generated from each mono-HETE had the elution characteristics of a di-HETE. The 5-HETE product has a triene spectrum similar to that of 5(S), 12(S)-di-HETE, whereas the 15- and 12-HETE products exhibited single ultraviolet absorption maxima, indicating a metabolic pathway for 5-HETE distinct from the other mono-HETEs. None of the stable cyclooxygenase products of 20:4 (6-keto PGF1 alpha, PGF2 alpha, PGE2, TXB2) nor polar metabolites of mono-HETEs are either incorporated or metabolized. The results indicate that macrophages have the capacity to specifically metabolize 20:4 and mono-HETEs to polar oxygenated products in the absence of a discernible trigger.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1982

Regulation of arachidonic acid metabolism by macrophage activation.

Levels of zymosan-induced arachidonic acid (20:4) metabolism by peritoneal macrophages elicited with inflammatory agents and resident macrophages were similar. Thyioglycollate (THIO)-elicited macrophages represented the exception; however, the diminished metabolism by these cells was reproduced by exposing resident cells to 5 mg/ml THIO broth in vitro. In contrast, reduced prostaglandin synthesis by macrophages from mice variously treated with the immunologic agents, Corynebacterium parvum or Bacille Calmette Guérin (BCG), closely correlated with enhanced antitoxoplasma activity, one measure of macrophage activation. This relationship, although not causative, suggested that the capacity for 20:4 metabolism is a function of the macrophage activation state. Modulation of macrophage 20:4 metabolism in vivo apparently required factors in addition to lymphocyte-derived products. Treatment of resident macrophages in vitro with BCG lymphokine was without effect on 20:4 release or prostaglandin synthesis. Activated macrophages from animals inoculated i.p. with C. parvum exhibited reduced 20:4 release and also failed to metabolize 70% of the 20:4 released in response to a zymosan stimulus. Consequently, the quantities of 20:4 metabolites formed were significantly less than expected from 20:4 release. These activated macrophages displayed greatly reduced synthesis of prostacylcin and leukotriene C compared with other 20:4 metabolites. It appeared that factors that regulate macrophage 20:4 metabolism influence the level of the inducible phospholipase and synthetic enzymes for specific 20:4 oxygenated products.

Animals↗
PubMed1982

Synthesis of leukotriene C and other arachidonic acid metabolites by mouse pulmonary macrophages.

Mouse resident pulmonary macrophages were subdivided into alveolar (PAM) and interstitial (PTM) populations on the basis of accessibility to pulmonary lavage, and zymosan-induced arachidonic acid (20:4) metabolism was examined in both populations labeled with [3H]20:4. Maximal phagocytic doses of unopsonized zymosan induced the specific release of 11% of phospholipid 20:4 by PTM and 4.6% by PAM. Direct fatty acid analysis of [3H]20:4-labeled PTM cultured in the presence or absence of zymosan indicated that the specific activity of the [3H]20:4 in cell phospholipid provided an accurate measure of 20:4 released by the cells, and could therefore be used to quantitate the synthesis of 20:4 metabolites by PTM in vitro. The single major 20:4 metabolite of PTM was the slow-reacting substance leukotriene C, which was synthesized in quantities of 3-4 pmol/microgram cell protein (280-370 pmol/10(6) cells), and comprised 20-25% of the released 20:4. PTM also synthesized prostaglandin E2, prostacyclin, thromboxane A2, and hydroxyeicosatetraenoic acids. In contrast, PAM produced leukotrienes D and E in addition to leukotriene C, prostaglandin E2, thromboxane A2, and hydroxyeicosatetraenoic acids. Prostacyclin formation by PAM was not observed. These studies define a set of experimental conditions for the study of 20:4 metabolism by pulmonary macrophages, and demonstrate that these cells are rich sources of LTC as well as other 20:4 oxygenated products.

Animals↗
PubMed1982

Glutathione metabolism in resting and phagocytizing peritoneal macrophages.

The steady state GSH content of cultured mouse resident peritoneal macrophages was 34 +/- 5 pmol/microgram of cell protein. Intracellular GSH content decreased concomitantly with zymosan ingestion. The half-life of GSH decreased from 1.9 h in resting cells to 0.58 h during phagocytosis as determined by inhibition of GSH synthesis with buthionine sulfoximine. The decrease in GSH half-life was directly related to the extent of particle uptake. In cytochalasin D-treated cells, attachment of zymosan to the macrophage plasma membrane in the absence of particle interiorization was sufficient to stimulate GSH turnover. Efflux was the major route of GSH loss in [35S]cystine-labeled macrophages, and was enhanced 3-fold by a zymosan challenge. GSH was lost intact since resident macrophages lack gamma-glutamyl transpeptidase (less than 1 pmol of L-gamma-glutamyl-p-nitroanilide/microgram of protein . h). Macrophages obtained from mice challenged in vivo with Corynebacterium parvum maintained higher intracellular GSH levels (50 +/- 5 pmol/microgram of cell protein) than did resident cells. The half-life of GSH in buthionine sulfoximine-treated C. parvum-elicited macrophages was 3.8 +/- 0.2 h while resting and 1.3 +/- 0.2 h during phagocytosis. C. parvum-elicited macrophages, in contrast to resident cells, contained sufficient levels of gamma-glutamyl transpeptidase activity to hydrolyze 55 pmol of L-gamma-glutamyl-p-nitroanilide/microgram of cell protein . h. These studies indicate that phagocytosis and cellular activation have profound effects on GSH metabolism in macrophages.

Animals↗
PubMed1982

Resting macrophages produce distinct metabolites from exogenous arachidonic acid.

Resident mouse peritoneal macrophages rapidly metabolize free arachidonic acid (20:4) in the absence of a discernible trigger. After a 20-min incubation in serumless medium, one-third of the fatty acid was found esterified in cell phospholipid and two-thirds was metabolized to oxygenated products which were recovered in the culture medium. The 20:4 oxygenated metabolites were identified by reverse-phase high performance liquid chromatography as hydroxyeicosatetraenoic acids (HETEs) and 6-keto prostaglandin F(1a) (6-ketoPGF(1a)), the stable form of prostacyclin, together with prostaglandin E(2) (PGE(2)) in proportions of 67:24:9. Inhibitor studies using indomethacin, nordihydroguaiaretic acid, and 5,8,11,14-eicosatetraenoic acid confirmed these metabolites to be lipoxygenase and cyclo-oxygenase products. The proportion of products differs considerably from those generated from phospholipid 20:4 in response to a phagocytic stimulus (HETEs:6-ketoPGF(1a):PGE(2):leukotriene C, 15:25:40: 15-20). Cornyebacterium parvum-elicited macrophages incorporated a higher percentage (70 percent) of exogenously supplied 20:4 and converted less than 20 percent of the fatty acid to oxygenated metabolites. Cyclo-oxygenase products (PGE(2), PGF(2a), TXB(2), and 6-ketoPGF(1a)) represented the major 20:4 metabolites (74 percent) synthesized by these activated macrophages. Esterification of 20:4 into cell phospholipids appeared not to be an initial obligatory step for synthesis of 20:4 oxygenated products by this route. To the contrary, incorporation of 20:4 into cell lipids and metabolism via the cyclo-oxygenase and lipoxygenase pathways represent distinct metabolic fates of exogenously supplied 20:4. These observations establish that resting macrophages contain high levels of cyclo-oxygenase and lipoxygenase activity and suggest macrophages can synthesize lipid mediators of inflammation in the absence of an inflammatory stimulus.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1982

IgE immune complexes stimulate arachidonic acid release by mouse peritoneal macrophages.

Resident mouse peritoneal macrophages were labeled with [3H]arachidonic acid and challenged with Sephadex beads coated with immune complexes of IgE and antigen. Arachidonic acid release by the cells was assessed by the quantity or radiolabel recovered from the culture medium. Freshly isolated macrophages responded to IgE immune complexes with a release of [3H]arachidonic acid that was linear for 1-2 hr. The magnitude of the response was dependent on both the number of immune complex-coated beads and on the degree of opsonization of the beads. Under conditions of maximal stimulation, macrophages challenged with IgE immune complex-coated Sephadex released 23 +/- 4.5% of their incorporated radiolabel. This is compared to values of 34.2 +/- 0.5% and 38.1 +/- 3.3% for cultures that received IgG immune complex-coated Sephadex or zymosan, respectively. Macrophages did not release arachidonic acid upon exposure to soluble IgE and antigen given sequentially or simultaneously, and soluble IgE did not inhibit the cells' response to IgE immune complexes. Incubation of macrophages for longer than 3 hr prior to challenge resulted in a selective loss in the cells' ability to respond to IgE immune complexes. After 16 hr of culture, macrophages released only 3.9 +/- 0.3% of their incorporated 3H on exposure to IgE immune complexes; however radiolabel release in response to zymosan (42.0 +/- 0.8%) was identical to that of freshly isolated cells. These data indicate that macrophages are capable of releasing arachidonic acid in response to preformed particulate immune complexes of IgE and antigen. Because Sephadex beads are too large to be interiorized by the cells, this response results from the interaction of the immune complexes with the macrophage plasma membrane.

Animals↗
PubMed1982

Human alveolar macrophages produce leukotriene B4.

Human alveolar macrophages obtained by bronchoalveolar lavage were labeled overnight with [3H]arachidonic acid. The cells were stimulated with calcium ionophore A23187, and the 20:4 oxygenated metabolites released into the culture medium were identified by reverse-phase HPLC. Leukotriene B4 was the major 20:4 metabolite produced by these cultures. Leukotriene B4 was identified by its reverse-phase HPLC elution time, its UV spectrum, and its chemotactic and chemokinetic activities for neutrophils. In addition, the macrophage- and neutrophil-derived leukotriene B4 free acids and methyl esters were found to have identical HPLC retention times.

Arachidonic Acid↗
PubMed1982

Arachidonic acid metabolism in glutathione-deficient macrophages.

Mouse resident peritoneal macrophages were treated with the glutathione (GSH) synthesis inhibitor buthionine sulfoximine to deplete intracellular GSH. The arachidonic acid metabolites released by the GSH-depleted macrophages in response to a zymosan challenge were analyzed by HPLC. Buthionine sulfoximine treatment resulted in inhibition of both prostaglandin E2 and leukotriene C synthesis that was directly related to the degree of GSH depletion. Macrophages in which GSH levels were reduced to 3% of normal exhibited reductions to 4% and 1%, respectively, in PGE2 and LTC formation. The total quantity of cyclooxygenase metabolites secreted by GSH-deficient macrophages was identical to that of control cells as a result of increased synthesis of prostacyclin and, to a lesser extent, 12-L-hydroxy-5,8,10-heptadecatrienoic acid. Total lipoxygenase products were decreased, however; increased formation of hydroxyicosatetraenoic acids only partially compensated for the deficit in leukotriene C production. These findings extent our earlier observations on the inhibition of leukotriene C synthesis in GSH-depleted macrophages and confirm with intact cells the previously suggested role of GSH in prostaglandin E2 formation.

Arachidonic Acid↗
PubMed1982

Deletion mutants which affect the nuclease-sensitive site in simian virus 40 chromatin.

A short segment of simian virus 40 (SV40) chromatin on the late side of the origin of replication is hypersensitive to nuclease cleavage. The role of DNA sequence information in this nuclease-sensitive feature was examined by constructing deletion mutations in this region. Deletions were introduced into the inserted segment of in(Or)-1411 (a viable, partially duplicated variant of SV40), and nuclease sensitivity of the inserted segment was compared with that of the unaltered sequences in their normal location in the viral genome. Extended deletions (118 to 161 base pairs) essentially abolished nuclease sensitivity within the inserted segment. Shorter deletions (21 to 52 base pairs) at separate locations retained the nuclease-sensitive feature. In some short-deletion mutants nuclease susceptibility was substantially reduced. We conclude that more than one genetic element in this region contributes to the organization of the nuclease-sensitive feature and that the SV40 72-base repeat is not, in itself, sufficient signal for this feature.

Animals↗
PubMed1981

Susceptibility of Entamoeba histolytica to oxygen intermediates.

To explore the susceptibility of the extracellular protozoan, Entamoeba histolytica, to toxic oxygen intermediates, trophozoites were exposed to fluxes of O2, H2O2, and OH. generated enzymatically by the glucose oxidase and xanthine oxidase reactions. HM-1 trophozoites were resistant to O2, but were readily killed by H2O2 alone. OH. and 1O2 were not required for effective amebicidal activity. The addition of peroxidase and halide enhanced trophozoite killing by H2O2. Sonicates of amebae contained virtually no catalase and little glutathione peroxidase activity which may contribute to susceptibility to H2O2. Coupled with our previous studies with Toxoplasma gondii and Leishmania spp. these observations indicate that there is a broad spectrum of susceptibility of intra- and extracellular pathogenic protozoa to killing by oxygen intermediates.

Animals↗
PubMed1981

Perinatal morbidity and mortality in small-for-dates babies: the relative importance of some maternal factors.

Data on 538 mothers and their small-for-dates babies were analysed to ascertain whether any specific maternal factors were associated with increased perinatal risks. There were 34 deaths; 21 (62%) of these babies had major congenital abnormalities. Among the survivors perinatal morbidity was also markedly increased when the baby was abnormal. No direct associations were found between abnormal babies and any maternal factors. Few differences were found in the incidence of mortality and morbidity factors according to maternal height, weight, weight gain in pregnancy, social class and smoking habits. There was a significantly higher death rate when the mother was pre-eclamptic, and the incidence of seven other morbidity factors was also increased. The only adverse effects of maternal hypertension without pre-eclampsia was a higher instrumental delivery rate, and more of these babies were tube-fed. There were only significantly more perinatal deaths among multiparous women whose previous babies had been of average birthweight. This was due to an excess of congenitally abnormal babies in this group. Only 35% small-for-dates babies escaped any perinatal problems.

Birth Weight↗