Biomedical subjects

G A Hamilton

Publications and source records attributed to G A Hamilton.

At least 19 recordsLinked to original sources

PubMed2001

Regulation of cell morphology and cytochrome P450 expression in human hepatocytes by extracellular matrix and cell-cell interactions.

The influence of extracellular matrix conditions and plating density on cell cytoarchitecture and the constitutive and chemically induced expression of cytochrome P450 3A4 (CYP3A4) was examined in primary cultures of human hepatocytes. Constitutive and drug-induced microsomal CYP3A4 expression occurred equally well in human hepatocyte cultures maintained on either a complex or simple substratum (Matrigel vs collagen, type I), or in a sandwich configuration (i.e., between two layers of extracellular matrix), despite the markedly different morphological properties exhibited by each condition. However, a density-dependent decrease in both the constitutive and induced levels of CYP3A4 was observed in hepatocytes maintained on a simple collagen substratum as plating density was reduced from 100% to 25%. Marked alterations in cell shape and cytoarchitecture were noted concomitant with decreases in the expression and localization of intercellular gap junctions and E-cadherin-mediated cell adhesions. In addition, the intracellular distribution of microtubules and microfilaments was altered substantially and the expression of immunoreactive actin and beta-tubulin increased as cell density was decreased. These effects were reversed to some extent by overlaying monolayers with extracellular matrix or by co-culturing with another cell type. Efforts to maintain normal cell shape and cytoskeletal distribution in hepatocytes at low cell density with a Matrigel substratum failed to restore normal basal levels of CYP3A4 expression or responsiveness to rifampicin (RIF). Likewise, E-cadherin and Cx-32 expression was again reduced, even though the distribution and expression of cytoskeletal elements returned to normal levels. These results suggest that cell-cell contacts, but not the extracellular matrix configuration or composition, play a critical role in determining normal responsiveness to chemical modulators in human hepatocytes.

Actins↗

PubMed2001

Safety and efficacy of fast track in patients undergoing coronary artery bypass surgery.

BACKGROUND: The incidence of coronary artery bypass surgery has been increasing annually with increasing pressure on the health care system. Fast track has been proposed as a means to increase efficiency and volume, without an increase in hospital resources. To date this approach has not been critically assessed in Canada. METHODS: We examined 617 consecutive patients undergoing isolated CABG surgery. The patients were divided into (1) fast track (FT) recovery (n = 219), without admission to an ICU, and (2) non-fast track (NFT) recovery (n = 398) with direct admission to the ICU. There were no differences in age, gender, timing of surgery, left main stenosis, preoperative myocardial infarction, renal failure, diabetes, peripheral vascular disease, or in the incidence of chronic obstructive pulmonary disease between the two groups. The NFT group had a higher proportion of patients with NYHA Class III/IV symptoms preoperatively (65.7% vs. 57.3%, p = 0.048), in patients with an ejection fraction < 40% (42.5% vs. 30.6%, p = 0.004), or in the number of individuals with an IABP inserted before surgery (13 vs. 1, p < 0.001). RESULTS: In the FT group the average period of aortic occlusion (40.7 +/- 15.2 min vs. 71.8 +/- 26.5 min, p < 0.001) and perfusion time (67.8 +/- 24.5 min vs. 117.5 +/- 40.2 min, p < 0.001) were significantly less than in the NFT group. The number of grafts per patient was 3.3 +/- 1.0 vs. 3.2 +/- 1.0, respectively (p = 0.38). Operative mortality was 0.9% in the FT group and 1.3% in the NFT group (p = 1.0). Significant differences were seen in the proportion of patients that suffered from postoperative ventilatory failure (3.2% in FT vs. 12.1% in NFT, p < 0.001), and the proportion of patients that suffered any postoperative complication was significantly higher in the NFT group (21.4%) than in the FT group (9.1%, p < 0.001). The differences in postoperative complications resulted in a shorter length of stay (LOS) in FT patients (5.6 +/- 4.1 days vs. 9.7 +/- 9.4 days NFT, p < 0.001). Only 4.1% of patients that entered the FT group failed and required admission to the ICU. Multivariate stepwise logistic regression analysis identified non-fast track recovery as an independent predictor of morbidity in CABG surgery patients. CONCLUSIONS: The data indicate it is possible to perform isolated CABG surgery, in a large proportion of the population, without the need for admission to an ICU for postoperative care.

Age Factors↗

PubMed2000

Comparative toxicity of trivalent and pentavalent inorganic and methylated arsenicals in rat and human cells.

Biomethylation is considered a major detoxification pathway for inorganic arsenicals (iAs). According to the postulated metabolic scheme, the methylation of iAs yields methylated metabolites in which arsenic is present in both pentavalent and trivalent forms. Pentavalent mono- and dimethylated arsenicals are less acutely toxic than iAs. However, little is known about the toxicity of trivalent methylated species. In the work reported here the toxicities of iAs and trivalent and pentavalent methylated arsenicals were examined in cultured human cells derived from tissues that are considered a major site for iAs methylation (liver) or targets for carcinogenic effects associated with exposure to iAs (skin, urinary bladder, and lung). To characterize the role of methylation in the protection against toxicity of arsenicals, the capacities of cells to produce methylated metabolites were also examined. In addition to human cells, primary rat hepatocytes were used as methylating controls. Among the arsenicals examined, trivalent monomethylated species were the most cytotoxic in all cell types. Trivalent dimethylated arsenicals were at least as cytotoxic as trivalent iAs (arsenite) for most cell types. Pentavalent arsenicals were significantly less cytotoxic than their trivalent analogs. Among the cell types examined, primary rat hepatocytes exhibited the greatest methylation capacity for iAs followed by primary human hepatocytes, epidermal keratinocytes, and bronchial epithelial cells. Cells derived from human bladder did not methylate iAs. There was no apparent correlation between susceptibility of cells to arsenic toxicity and their capacity to methylate iAs. These results suggest that (1) trivalent methylated arsenicals, intermediary products of arsenic methylation, may significantly contribute to the adverse effects associated with exposure to iAs, and (2) high methylation capacity does not protect cells from the acute toxicity of trivalent arsenicals.

Aged↗

PubMed2000

The use of liver spheroids as an in vitro model for studying induction of the stress response as a marker of chemical toxicity.

Stress protein induction has been advocated as a sensitive indicator of compound-induced toxicity. In monolayer cultures of primary hepatocytes, however, the two stress proteins, Hsp25 and Hsp72/3 are up-regulated, probably due to the effect of the isolation procedure and adaptation of the cells to the culture conditions. The aim of the current studies was to determine whether liver spheroids would provide an improved experimental model for the study of heat shock protein induction in vitro. Primary rat hepatocytes were cultured as liver spheroids and the expression of Hsp25 and Hsp72/3 measured along with the levels of ATP, GSH and albumin secretion. Hsp72/3 was initially increased in spheroid culture but returned to in vivo levels after 3 days of culture. Hsp25 was maintained at in vivo levels until day 6 of culture, after which levels increased slightly. The effects of the two hepatotoxins, hydrazine and cadmium chloride (CdCl(2)), were therefore measured on day 6 of spheroid culture. CdCl(2) had no effect on Hsp25 but increased Hsp72/3 at concentrations that affected other biochemical parameters. Hydrazine caused a rapid reduction in ATP levels and albumin secretion, but did not affect Hsp72/3. Hsp25 was slightly induced by hydrazine at later sampling times at concentrations, however, that affected other biochemical parameters. It can be concluded that liver spheroids provide a model for studying stress protein expression. However, the increase in stress proteins appears to be a relatively insensitive parameter compared to other more conventionally used toxicity endpoints and the response appears to vary with individual toxins under study.

Adenosine Triphosphate↗

PubMed2000

The pregnane X receptor: a promiscuous xenobiotic receptor that has diverged during evolution.

Transcription of genes encoding cytochrome P450 3A (CYP3A) monooxygenases is induced by a variety of xenobiotics and natural steroids. There are marked differences in the compounds that induce CYP3A gene expression between species. Recently, the mouse and human pregnane X receptor (PXR) were shown to be activated by compounds that induce CYP3A expression. However, most studies of CYP3A regulation have been performed using rabbit and rat hepatocytes. Here, we report the cloning and characterization of PXR from these two species. PXR is remarkably divergent between species, with the rabbit, rat, and human receptors sharing only approximately 80% amino acid identity in their ligand-binding domains. This sequence divergence is reflected by marked pharmacological differences in PXR activation profiles. For example, the macrolide antibiotic rifampicin, the antidiabetic drug troglitazone, and the hypocholesterolemic drug SR12813 are efficacious activators of the human and rabbit PXR but have little activity on the rat and mouse PXR. Conversely, pregnane 16alpha-carbonitrile is a more potent activator of the rat and mouse PXR than the human and rabbit receptor. The activities of xenobiotics in PXR activation assays correlate well with their ability to induce CYP3A expression in primary hepatocytes. Through the use of a novel scintillation proximity binding assay, we demonstrate that many of the compounds that induce CYP3A expression bind directly to human PXR. These data establish PXR as a promiscuous xenobiotic receptor that has diverged during evolution.

Amino Acid Sequence↗

PubMed1999

Metabolism of arsenic in primary cultures of human and rat hepatocytes.

The liver is considered a major site for methylation of inorganic arsenic (iAs). However, there is little data on the capacity of human liver to methylate iAs. This work examined the metabolism of arsenite (iAs(III)), arsenate (iAs(V)), methylarsine oxide (MAs(III)O), methylarsonic acid (MAs(V)), dimethylarsinous acid (DMAs(III)), and dimethylarsinic acid (DMAs(V)) in primary cultures of normal human hepatocytes. Primary rat hepatocytes were used as methylating controls. iAs(III) and MAs(III)O were metabolized more extensively than iAs(V) and MAs(V) by either cell type. Neither human nor rat hepatocytes metabolized DMAs(III) or DMAs(V). Methylation of iAs(III) by human hepatocytes yielded methylarsenic (MAs) and dimethylarsenic (DMAs) species; MAs(III)O was converted to DMAs. The total methylation yield (MAs and DMAs) increased over the range of 0.1 to 4 microM iAs(III). However, DMAs production was inhibited by iAs(III) in a concentration-dependent manner, and the DMAs/MAs ratio decreased. iAs(III) (10 and 20 microM) inhibited both methylation reactions. Inhibition of DMAs synthesis resulted in accumulation of iAs and MAs in human hepatocytes, suggesting that dimethylation is required for iAs clearance from cells. Methylation capacities of human hepatocytes obtained from four donors ranged from 3.1 to 35.7 pmol of iAs(III) per 10(6) cells per hour and were substantially lower than the methylation capacity of rat hepatocytes (387 pmol of iAs(III) per 10(6) cells per hour). The maximal methylation rates for either rat or human hepatocytes were attained between 0.4 and 4 microM iAs(III). In summary, (i) human hepatocytes methylate iAs, (ii) the capacities for iAs methylation vary among individuals and are saturable, and (iii) moderate concentrations of iAs inhibit DMAs synthesis, resulting in an accumulation of iAs and MAs in cells.

Animals↗

PubMed1998

Advanced practice nursing in the Nordic countries.

Changes in the delivery of health care and changes in population characteristics and health care requirements mandate changing requirements in nursing education. This is necessary to meet patient and family needs and to deliver quality health care. This paper describes the background to nursing education in the Nordic countries and gives an account of an initiative in Norway to prepare advanced practice nurses for clinical practice in this dynamic environment.

Curriculum↗

PubMed1996

Inhibition of proliferation and of IL-2 production and utilization in lymphocytes by S-oxalylglutathione.

Previously we have shown that S-oxalins (monothiolesters of oxalic acid) are ubiquitous mammalian metabolites whose concentrations decrease when lymphocytes are stimulated to proliferate. The present study was undertaken to further examine the role of S-oxalins in the proliferation process. When added to lymphocytes stimulated with concanavalin A, the S-oxalin, S-oxalylglutathione (GS-Ox), inhibited DNA synthesis by 50% when present at ca. 0.15 mM and virtually 100% at 0.5 mM. The inhibition was reversible. The presence of GS-Ox blocked IL-2 production, but addition of IL-2 did not permit DNA synthesis to proceed. GS-Ox also inhibited proliferation of an IL-2-dependent cell line, BT2. In primary lymphocytes GS-Ox reduced IL-2 receptor expression, but not in an IL-2-dependent blast cell line. Overall RNA synthesis and protein synthesis were not significantly altered by GS-Ox. Levels of the positive transcription factor, NF-kappaB, were decreased after incubation of lymphocytes with GS-Ox, but the amount of a negative transcription factor, NREA, was largely unchanged. The results not only provide further evidence that S-oxalins are small-molecule cell proliferation inhibitors, they also clarify to some extent the specific steps in the activation response modulated by S-oxalins.

Animals↗

PubMed1996

The S-oxalin, N-acetyl-S-oxalylcysteamine, inhibits lymphocyte proliferation, IL-2 production and utilization.

S-Oxalins are a recently described class of cell metabolites that appear to function as negative regulators of proliferation. Previously we have shown that exogenous S-oxalylglutathione (GS-Ox) inhibits the proliferation of lymphocytes by inhibiting the production and utilization of IL-2. In the present study the synthetic S-oxalin, N-acetyl-S-oxalylcysteamine (ACS-Ox), was utilized in similar experiments to determine whether GS-Ox itself, or possibly some metabolite formed following initial conversion of GS-Ox by gamma-glutamyltransferase (GGT), is responsible for the effects (ACS-Ox is not metabolized by GGT). ACS-Ox inhibited DNA synthesis in lymphocytes stimulated by concanavalin A similarly to GS-Ox. IL-2 production and utilization and IL-2R expression were inhibited as well. ACS-Ox also inhibited the proliferation of IL-2 dependent cells at the same concentration as GS-Ox. Because the effects of GS-Ox and ACS-Ox are so similar, presumably the S-oxalin itself, rather than some metabolite, is responsible for the observed effects. Transfer of oxalyl groups from S-oxalins to various proteins thiols is the most likely mechanism involved.

Animals↗

PubMed1996

Physical mapping of a centromere-proximal region of chromosome IV-L defines the placement of genes USO1, MBP1, PSA1 and SLC1.

A physical map of a 14.5 kb region close to the centromere on the left arm of chromosome IV of Saccharomyces cerevisiae is presented. This map has been constructed by restriction analysis of a clone from a YCp50 genomic library and by use of pre-existing and new sequence data from this region. The map reveals the following gene order (reading from the most centromere-distal to the most centromere-proximal locus): USO1/INT1-MBP1-PSA1-SLC1-YLA1 and defines the size of the open reading frames and intergenic regions.

Base Sequence↗

PubMed1991

Determination of oxalyl thiolesters, N-oxalylcysteine and N-oxalylcysteamine in biological materials.

A convenient method is described for the quantitative analysis of oxalyl thiolesters (OTEs), a newly discovered class of mammalian metabolites, in biological samples. By this particular technique the total concentration of all OTEs in the sample is determined. The method involves first reacting the biological material with cysteamine (2-aminoethanethiol) or cysteine under conditions that convert OTEs quantitatively to N-oxalylcysteamine (or N-oxalylcysteine), followed by reaction with monobromobimane to give a highly fluorescent derivative that is analyzed by reversed-phase ion-pair chromatography, with tetrabutylammonium ion as the counterion and N-(2-mercaptopropionyl)glycine as an internal standard. The method is capable of detecting as little as 0.6 pmol of the bimane derivative of the N-oxalyl compound in a single HPLC injection. The application of this method has led to the discovery that not only OTEs but also N-oxalylcysteine and N-oxalylcysteamine are normal mammalian metabolites. In various rat tissues the OTE concentration ranges up to 65 nmol/g (wet wt), the N-oxalylcysteine concentration is approximately 10 nmol/g, and the N-oxalylcysteamine concentration is 0-3 nmol/g.

Animals↗

PubMed1991

The importance of systematic inquiry in nursing research: An example; women and recovery from acute myocardial infarction.

A review of the literature related to recovery from acute myocardial after discharge from the hospital revealed that very little is known about the recovery period in women. The recovery period has been studied in men with the major focus on cardiac rehabilitation, return to work, and sexual activity. In this descriptive exploratory study women were compared to men in these three areas of recovery within 3 to 9 months after discharge from the hospital. There were statistically significant differences in the following areas: marital status, more women were widowed or single; women had more responsibility for household duties after acute myocardial infarction, a decrease in frequency of sexual activity, and more reports of chest pain during sexual activity. Subjects reported little or no counselling by nurses with regard to cardiac rehabilitation, return to work or household duties, and resumption of sexual activity.

Chest Pain↗

PubMed1990

Coronary heart disease risk factors in women.

Although men and women share a number of coronary risk factors including age, hypertension, cigarette smoking, diabetes, obesity, plasma lipoprotein concentration, and family history, the overall impact of these factors on the incidence and clinical manifestations of coronary heart disease (CHD) may differ. Additional risk factors which solely impact upon women include the use of oral contraceptives, menopause, and postmenopausal hormones. The impact of psychosocial and behavioral factors on CHD risk in women requires further investigation given that our current knowledge of traditional risk factors alone inadequately predicts all cases of CHD.

Coronary Disease↗

PubMed1990

Recovery from acute myocardial infarction in women.

This paper is a review of the literature on recovery from acute myocardial infarction in women. The topic has been subdivided into three areas for presentation: cardiac rehabilitation, return to work and sexual activity. The exploration of the literature revealed the paucity of research on women, but some comparisons could be made between men and women. Compared to men, women appear to utilize cardiac rehabilitation programs less frequently than men and have higher dropout rates, they return to work less frequently and after a longer period of time and resume sexual activity after a longer period of time reporting more symptoms during and after the activity. Investigation of the literature showed that the recovery period for women is incompletely explored and that there is a critical need for research.

Female↗

PubMed1989

Oxalyl thiolester concentrations decreased in lectin and phorbol ester-stimulated lymphocytes.

Oxalyl thiolesters (OTEs) are a newly discovered class of mammalian metabolites that are believed to function in controlling animal metabolism and possibly serve as intracellular mediators for some hormones. Previous correlations had suggested that the concentrations of OTEs might be decreased when cells are stimulated to proliferate, and in our research that was found to be the case. Thus, when bovine lymph node lymphocytes are stimulated either with concanavalin A (Con A) or with a combination of phytohemagglutinin (PHA) and the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), the concentration of OTEs in the lymphocytes decreases within 3 h by a factor of approximately two. With either PHA or TPA alone, the decrease in OTE concentration is considerably smaller. With Con A as stimulant, the OTE levels decrease within 1 h and remain low for at least 24 h. It was also noted that the concentration of OTEs in unstimulated isolated lymphocytes is significantly lower in lymphocytes obtained from 2-year-old animals than in lymphocytes obtained from older animals. The results of the current investigation, when considered in conjunction with other recent results, suggest that OTEs may be natural cell proliferation inhibitors.

Animals↗

PubMed1988

S-oxalylglutathione is a substrate for gamma-glutamyltransferase (GGT). Implications for the role of GGT in cell proliferation.

With glycylglycine or water as acceptor, bovine kidney gamma-glutamyltransferase catalyzes reactions of the known mammalian metabolite, S-oxalylglutathione, at rates comparable to those of L-gamma-glutamyl-p-nitroanilide, a known good substrate. N-Oxalyl-cysteinylglycine is the eventual product of the former reaction. Since oxalyl thiolesters are implicated as important cell proliferation inhibitors, it is proposed that this reaction plays a major role in controlling cell proliferation.

Animals↗

PubMed1987

Concerning the mechanism for transfer of D-glucuronate from myo-inositol oxygenase to D-glucuronate reductase.

The D-glucuronate product of myo-inositol oxygenase (EC 1.13.99.1) is efficiently reduced by NADPH in the presence of either purified D-glucuronate reductase (EC 1.1.1.19), or reductase that is part of a protein aggregate that also contains the oxygenase. This occurs despite the fact that the maximum concentration of D-glucuronate that could be formed by the oxygenase under the conditions used for the coupled enzyme experiments is 7 microM, and 10 microM externally supplied D-glucuronate (Km = 7.6 mM) does not support any detectable NADPH oxidation under the reaction conditions. The most likely explanation for the results is that the uncyclized aldehyde form of D-glucuronate is the product of the oxygenase reaction, and that it diffuses into solution and is captured by the reductase before it cyclizes to the more stable but less reactive hemiacetal form.

Carbohydrate Dehydrogenases↗

PubMed1987

Inhibition of malic enzyme by S-oxalylglutathione, a probable in vivo effector.

Various oxalyl thiol esters (RSCOCOO-), especially S-oxalylglutathione (GS-Ox), were found to be very effective inhibitors of chicken liver malic enzyme. When the conditions are similar to those encountered physiologically [high reduced nicotinamide adenine dinucleotide phosphate (NADPH) concentrations], inhibition is detectable with less than 1 microM concentrations of GS-Ox. The amount of inhibition is not reversed by excess glutathione, thus indicating that it is not due to oxalyl transfer to some enzymic thiol group with release of glutathione. Detailed kinetic studies show that the inhibition by GS-Ox can be treated as a simple reversible binding to the enzyme; the double reciprocal plot patterns indicate that the inhibition is linear noncompetitive (mixed type), vs. both L-malate in the oxidative decarboxylation reaction and pyruvate in the reverse reaction. At pH 7.4 and 25 degrees C in the presence of 100-200 microM NADPH, the Kis and Kii values for GS-Ox are 0.7 and 5 microM, respectively, and are the same for reactions run in either direction. The high specificity for GS-Ox is indicated by the observation that, under similar conditions, the Kis values for S-oxalyl coenzyme A and S-oxalyl-N-acetylcysteamine are 40 and 150 microM, respectively. Such high specificity indicates that the enzyme has evolved a specific binding site for the glutathione part of GS-Ox. The current results, when considered in conjunction with recent evidence that oxalyl thiol esters are present in animal tissues at concentrations up to 50 microM, imply that GS-Ox is an important in vivo regulator of malic enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗