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Biomedical subjects

A Brooks

Publications and source records attributed to A Brooks.

At least 91 records · Page 5Linked to original sources
PubMed1993

Identification of endogenous electrophiles by means of mass spectrometric determination of protein and DNA adducts.

Monitoring exposure to alkylating agents may be achieved by quantitatively determining the adduct levels formed with nucleic acids and/or proteins. One of the most significant results arising from the application of this approach has been the discovery in control populations of "background" levels of alkylated nucleic acid bases or alkylated proteins, in particular hemoglobin (Hb). In the case of Hb, a wide variety of such adducts have been detected and quantitated by mass spectrometric techniques, with methylated, 2-carboxyethylated, and 2-hydroxyethylated modifications being most abundant. Although the source of these alkylation products is unknown, both endogenous and exogenous sources may be proposed. We have recently confirmed the presence of the N-terminal hydroxyethylvaline adduct in control human Hb using tandem mass spectrometry (MS-MS) and have now established background levels using GC-MS in more than 70 samples. Smoking raises the levels of the adduct up to 10-fold and occupational exposure to ethylene oxide up to 300-fold. Background levels of alkylated nucleic acids may be studied by analysis of N7-alkylated guanine or N3-alkylated adenine, which are excised from nucleic acids after their formation and are excreted in urine. Although the presence of some of these urinary constituents may be accounted for by their natural occurrence in RNA or diet, the endogenous or exogenous source of others is unknown. Quantitative methods using MS-MS have now been developed for five of the observed urinary alkylguanines [N7-methyl-, N2-methyl-, N2-dimethyl-, N7-(2-hydroxyethyl)-, and N2-ethylguanine].(ABSTRACT TRUNCATED AT 250 WORDS)

Alkylating Agents↗
PubMed1991

Class II-restricted presentation of an endogenously derived immunodominant T-cell determinant of hen egg lysozyme.

An in vitro model was used to investigate the potential for different structural forms of endogenous antigen to be processed and presented by major histocompatibility complex class II molecules. For this purpose the class II-restricted presentation of an immunodominant epitope of hen egg lysozyme [HEL-(46-61)] was studied in class II-positive B-lymphoma cells (M12.C3) transfected with genes encoding HEL molecules either (i) secreted in high (hi) or low (lo) amounts as soluble antigen [sHEL(hi/lo)], (ii) localized within the endoplasmic reticulum (ER)/salvage compartment (ER-HEL), or (iii) anchored on the cell surface as an integral membrane protein (mHEL). The corresponding sHEL, ER-HEL, and mHEL gene products were expressed as predicted except that HEL determinants accumulated in the culture supernatant as well as on the cell membrane of mHEL-transfected cells. Class II-positive cells endogenously expressing all three forms of HEL antigen constitutively presented the immunodominant HEL-(46-61) determinant with differential efficiency (mHEL, sHEL greater than ERHEL) to a class II-restricted T hybridoma. A second T hybridoma recognized endogenous HEL-(46-61) determinants constitutively presented on sHEL(hi) and mHEL transfectants but not on sHEL(lo) or ERHEL transfectants. The formation of HEL-(46-61)/I-Ak complexes in the ERHEL and sHEL(lo) transfectants was therefore limiting. Mixing experiments with different antigen-presenting cells indicated that the HEL-(46-61) determinant was derived from endogenous antigen rather than by reuptake of shed or secreted HEL determinants. We conclude that MHC class II molecules can present some antigenic determinants derived from endogenous proteins that are sequestered in the ER/salvage compartment as well as distally transported in the form of secretory or membrane antigens.

Animals↗
PubMed1991

Genetic specification of life span and self-fertility in recombinant-inbred strains of Caenorhabditis elegans.

The genetic basis of life-span and age-specific fertility has been analysed using recombinant-inbred strains of the nematode Caenorhabditis elegans. Estimates of narrow-sense heritability range from 0.05 to 0.36 for life span and from 0.36 to 0.49 for total self-fertility. Positive phenotypic and genetic correlations for life span and total fertility were also observed, although in most cases the correlations were not significant. In general, age-specific hermaphrodite fertility was positively correlated with fertility on contiguous days but was negatively correlated with fertility on more distant days. We estimate that a minimum of two to three genes specify each of these traits in this genetic background. Three single-gene markers were used to generate strain distribution patterns, and two of these were found to be linked with loci that specify life span and/or fertility. We also saw evidence for a significant environmental component affecting self-fertility.

Animals↗
PubMed1990

Phasic volumetric coronary venous outflow patterns in conscious dogs.

We performed the present study to characterize phasic venous outflow patterns in conscious chronically instrumented dogs. An implantable ultrasonic transit-time flow probe capable of measuring volumetric venous outflow was placed on the coronary sinus (n = 9) and/or great cardiac vein (n = 5). Under resting conditions, great cardiac vein flow was predominantly systolic [systolic index 61 +/- 6%/beat (means +/- SD), diastolic index 39 +/- 6%/beat]. In contrast, coronary sinus volumetric flow was predominantly diastolic (systolic index 36 +/- 13%/beat, diastolic index 64 +/- 13%/beat). During adenosine vasodilation both great cardiac vein and coronary sinus flow patterns were predominantly systolic (systolic index 58 +/- 12%/beat for the coronary sinus and systolic index 75 +/- 6%/beat in great cardiac vein flow). In addition, the time at which peak venous flow occurred shifted from end systole to midsystole after vasodilation. The variation in coronary venous flow patterns with position suggests that outflow patterns are significantly influenced by a venous compliance between the great cardiac vein and distal coronary sinus. The lack of a predominantly systolic outflow pattern in the coronary sinus under resting conditions contrasts with previous findings in open-chest anesthetized animals and raises the possibility that cannulation of the coronary sinus and/or changes in vasomotor tone caused by anesthesia alter resting venous flow patterns.

Animals↗
PubMed1988

The electrophysiological effects of melatonin and a putative melatonin antagonist (N-acetyltryptamine) on rat suprachiasmatic neurones in vitro.

Several studies have implicated the suprachiasmatic nuclei (SCN) as a target for the action of melatonin in its regulation of seasonal and circadian behaviour. Single-unit activity from the SCN and adjacent paraventricular area and anterior hypothalamus was recorded using the in vitro rat hypothalamic slice preparation. Neurones were tested for responses to iontophoresed or pressure ejected melatonin (n = 62) and serotonin (5-HT, n = 80). The majority (n = 24-26) of melatonin-sensitive SCN neurones were inhibited by melatonin in a dose-dependent manner during the latter portion of the projected light phase of the circadian light-dark cycle. A putative melatonin antagonist, N-acetyltryptamine, exhibited concentration dependent mixed agonist-antagonist effects on melatonin-evoked responses.

Action Potentials↗
PubMed1988

Protein-bound ribulose bisphosphate correlates with deactivation of ribulose bisphosphate carboxylase in leaves.

Previous reports indicate that ribulose 1,5-bisphosphate (RuBP) binds very tightly to inactive ribulose bisphosphate carboxylase (rubisco) in vitro. Therefore, we decided to investigate whether there was evidence for tight binding of RuBP associated with deactivation of rubisco in vivo. We modified a technique for rapidly separating ;free' metabolites from those bound to high molecular compounds. Arabidopsis thaliana plants were illuminated at various irradiances before freezing the leaves in liquid N(2) and assaying rubisco activity and RuBP. The percentage activation of rubisco varied from 37% at low irradiance (45 micromoles quanta per square meter per second) to 100% at high irradiance (800 micromoles quanta per square meter per second). The total amount of RuBP did not vary much with irradiance, but bound RuBP changed from 36% of the total at low irradiance to none at high irradiance. Bound RuBP was significantly correlated with the estimated number of inactive rubisco sites, with a ratio of about 1:1. After a step increase in irradiance, rubisco activation increased and total RuBP increased transiently, but steady levels of both occurred by 10 minutes. The amount of bound RuBP decreased with a similar time course to the estimated decrease in inactive rubisco sites. After a step decrease in irradiance, rubisco deactivated slowly for at least 25 minutes. Bound RuBP increased gradually but did so more slowly than the estimated increase in inactive rubisco sites.

Journal Article↗
PubMed1988

Effects of Irradiance and Methyl Viologen Treatment on ATP, ADP, and Activation of Ribulose Bisphosphate Carboxylase in Spinach Leaves.

Since activation of ribulose bisphosphate carboxylase (rubisco) by rubisco activase is sensitive to ATP and ADP in vitro, we aimed to test the correlation between ATP level and rubisco activation state in intact leaves of Spinacia oleracea L. in response to changes in irradiance and after feeding the electron acceptor methyl viologen. Leaves were exposed to various irradiances for 45 minutes at atmospheric partial pressures of CO(2) and O(2). After measuring the rate of CO(2) assimilation, leaves were freeze-clamped in situ and the punched discs assayed for rubisco activity, and amounts of ribulose bisphosphate (RuBP), ATP, and ADP. The photosynthetic rate and the activation state of rubisco increased with increasing irradiance but the levels of RuBP, ATP, and ADP were not greatly affected. Methyl viologen fed leaves under low irradiance had rubisco activation states of 93% compared to 51% in control leaves. The ATP content of the leaves was also significantly higher and the ratio of ATP to ADP was 4.1 in methyl viologen fed leaves compared to 2.2 in control leaves. From these results and other published results we conclude that a correlation between ATP level and rubisco activation can be observed in intact leaves, but that during changes in irradiance some additional factors are involved in regulating rubisco activation.

Journal Article↗
PubMed1987

Algal and Bacterial Activities in Acidic (pH 3) Strip Mine Lakes.

Reservoir 29 and Lake B are extremely acid lakes (epilimnion pHs of 2.7 and 3.2, respectively), because they receive acidic discharges from coal refuse piles. They differ in that the pH of profundal sediments in Reservoir 29 increased from 2.7 to 3.8 during the period of thermal stratification, whereas permanently anoxic sediments in Lake B had a pH of 6.2. The pH rise in Reservoir 29 sediments was correlated with a temporal increase in H(2)S concentration in the anaerobic hypolimnion from 0 to >1 mM. The chlorophyll a levels in the epilimnion of Reservoir 29 were low, and the rate of primary production was typical of an oligotrophic system. However, there was a dense 10-cm layer of algal biomass at the bottom of the metalimnion. Production by this layer was low owing to light limitation and possibly H(2)S toxicity. The specific photosynthetic rates of epilimnetic algae were low, which suggests that nutrient availability is more important than pH in limiting production. The highest photosynthetic rates were obtained in water samples incubated at pH 2.7 to 4. Heterotrophic bacterial activity (measured by [C]glucose metabolism) was greatest at the sediment/water interface. Bacterial production (assayed by thymidine incorporation) was as high in Reservoir 29 as in a nonacid mesotrophic Indiana lake.

Journal Article↗
PubMed1983

Activation of antigen-specific suppressor T cells by B cells from mice immunized with type III pneumococcal polysaccharide.

The transfer of B lymphocytes from mice immunized with type III pneumococcal polysaccharide (SSS-III) results in antigen-specific suppression of the antibody response of recipients immunized with SSS-III. Such suppression shares many features associated with low-dose paralysis, a phenomenon mediated by suppressor T cells; it reaches maximal levels 3 d after the transfer of viable or irradiated immune B cells and can be eliminated by the depletion of SSS-III-binding cells from spleen cell suspensions before transfer. In a two-step cell transfer experiment, purified T lymphocytes, isolated from recipients previously given immune B cells, caused suppression upon transfer to other mice immunized with SSS-III. Also, B-cell-induced suppression could be abrogated in a competitive manner by the infusion of amplifier T lymphocytes, as was previously demonstrated in the case of low-dose paralysis. These findings suggest that B cell surface components, presumably the idiotypic determinants of cell-associated antibody specific for SSS-III, are instrumental in activating suppressor T cells involved in regulating the magnitude of the antibody response to SSS-III.

Animals↗
PubMed1978

Determination of water soluble imidazo-1,4-benzodiazepines in blood by electron- capture gas--liquid chromatography and in urine by differential pulse polaragraphy.

A sensitive and specific electron-capture gas--liquid chromatographic (GLC--ECD) assay was developed for the determination of 8-chloro-6-(2'-fluorophenyl)-1-methyl-4H-imidazo(1,5a)(1,4)benzodiazepine (I) or 8-chloro-1,4-dimethyl-6-(2'-fluorophenyl)-4H-imidazo (1,5a)(1,4)benzodiazepine (II) in blood. The assay for both compounds involves extraction into benzene--methylene chloride (9:1) from blood buffered to pH 12.6 The overall recovery of I and II from blood is 86% +- 5.0 (S.D.) and the sensitivity limit of detection is of the order of 2 to 3 ng of I or II per milliltre of blood. The major urinary metabolite of I is 8-chloro-6-(2'-fluorophenyl)-1-hydroxymethyl-4H-imidazo(1,5a)(1,4)benzodiazepine, (IA) present as a glucuronide conjugate while 8-chloro-6-(2'-fluorophenyl)-4-hydroxyl-1-methyl-4H-imidazo(1,5a)(1,4)benzodiazepine, (IB) and 8-chloro-6-(2'-fluorophenyl)-4-hydroxy-1-hydroxymethyl-4H-imidazo(1,5a)(1,4) benzodiazepine, (IC) are minor metabolites. The major metabolite IA is extracted into benzene--methylene chloride (9:1) from urine buffered to pH 11.0 (after incubation with glucuronidase--sulfatase as pH 5.0), and analyzed by differential pulse polarography (DPP) in 0.1 M phosphate buffer PH 3). The overall recovery of IA is 84 +- 3.0% (S.D.) with a sensitivity limit of 50 ng per millilitre of urine. The metabolites of compound II have not as yet been elucidated. The GLC--ECD and DPP assays were applied to the determination of blood levels and urinary excretion in dogs following single 10 mg/kg intravenous and oral doses of I and following single 6 mg/kg intravenous and 10 mg/kg oral doses of II. Blood levels of compound I were also evaluated in man following intravenous infusion of single 10 mg doses.

Animals↗