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Biomedical subjects

J L Stevens

Publications and source records attributed to J L Stevens.

106 records · Page 6Linked to original sources
PubMed1981

Effect of cysteine, diethyl maleate, and phenobarbital treatments on the hepatotoxicity of [1H]chloroform.

The effects of cysteine, diethyl maleate and phenobarbital treatments and 2H-substitution on the hepatotoxicity of chloroform were investigated. Time course studies of covalent binding and hepatoxicity in phenobarbital-treated rats showed that covalent binding of 14C-label from [14C]chloroform was maximal at 6 h after chloroform administration while hepatotoxicity reached a peak at 18 h. Cysteine treatment reduced both covalent binding and hepatotoxicity, while diethyl maleate and phenobarbital treatments increased both the hepatotoxicity of chloroform and the covalent binding of chloroform metabolites to hepatic proteins. A deuterium isotope effect was present on chloroform-induced hepatotoxicity in diethyl maleate-treated rats suggesting that the previously reported inhibition of haloform metabolism by diethyl maleate occurs at a step in the reaction mechanism after phosgene production. These data support the concept that phosgene is the toxic intermediate in chloroform metabolism.

Alanine Transaminase↗
PubMed1980

Halogenated methanes: metabolism and toxicity.

Dihalomethanes are metabolized to carbon monoxide (CO) both in vivo and in vitro. The reaction is catalyzed by the hepatic microsomal cytochrome P-450 dependent mixed function oxidase system. Reaction mechanism studies suggest an initial oxygen insertion reaction followed by rearrangement to a formyl halide intermediate, which in turn decomposes to yield CO. In vitro studies show that [14C]dichloromethane becomes covalently bound to both microsomal protein and lipid. The similar characteristics of metabolism to CO and covalent suggest that a common intermediate, perhaps the formyl halide, may be involved. Dihalomethanes are also metabolized for formaldehyde, formic acid, and inorganic halide. A glutathione transferase located in hepatic cytosol fractions appears to be involved. Reaction mechanism studies suggest that a S-hydroxymethyl glutathione intermediate may yield formaldehyde or be diverted via formaldehyde dehydrogenase/S-formyl glutathione hydrolase to yield formic acid. Haloforms are also metabolized to carbon monoxide both in vivo and in vitro by a hepatic microsomal cytochrome P-450 dependent mixed function oxidase system. In vitro, this reaction is markedly stimulated by sulfhydryl compounds. Reaction mechanism studies suggest an initial oxygen insertion reaction followed by rearrangement to a dihalocarbonyl intermediate, which in turn reacts with sulfhydryl reagents to yield a thiol-S-formyl halide. Subsequent attack by other sulfhydryl compounds would result in the formation of CO and a disulfide.

Biotransformation↗
PubMedDate not supplied

Intraocular pressure and hemodynamic changes following tracheal intubation in children.

STUDY OBJECTIVE: To determine the optimal time in which to make intraocular pressure (IOP) measurements in children following tracheal intubation. DESIGN: Randomized, controlled trial. SETTING: Operating rooms of a tertiary-care children's hospital. PATIENTS: Thirteen healthy children undergoing elective strabismus correction surgery under halothane and nitrous oxide (N2O) endotracheal anesthesia. INTERVENTIONS: Following induction of anesthesia, patients were randomly assigned to receive stable end-tidal halothane concentrations of 0.5% or 1.0% in 66% N2O. MEASUREMENTS AND MAIN RESULTS: Baseline (preintubation) IOP, heart rate (HR), and mean arterial pressure (MAP) were recorded after 10 minutes of steady-state end-tidal concentrations. These measurements were repeated at 1-minute intervals following tracheal intubation, which was facilitated with atracurium. HR and MAP changes were found to be good predictors of IOP changes. IOP returned to baseline (preintubation) values when HR and MAP returned to preintubation levels. However, IOP measurements under anesthesia may not reflect awake values. CONCLUSIONS: We recommend that IOP be measured only after HR and MAP have returned to preintubation levels in children who have undergone tracheal intubation during halothane and N2O anesthesia.

Anesthesia, Endotracheal↗
PubMedDate not supplied

Formation of mitochondrial phospholipid adducts by nephrotoxic cysteine conjugate metabolites.

Nephrotoxic cysteine conjugates derived from a variety of halogenated alkenes are enzymatically activated via the beta-lyase pathway to yield reactive sulfur-containing metabolites which bind covalently to cellular macromolecules. Mitochondria contain beta-lyase enzymes and are primary targets for binding and toxicity. Previously, mitochondrial protein and/or DNA have been considered as molecular targets for cysteine conjugate metabolite binding. We now report that metabolites of nephrotoxic cysteine conjugates form covalent adducts with rat kidney mitochondrial phospholipids. Rat kidney mitochondria were incubated with the 35S-labeled conjugates S-(1,1,2,2-tetrafluoroethyl)-L-cysteine (TFEC), S-(2-chloro-1,1,2-trifluoroethyl)-L-cysteine (CTFC), S-(1,2-dichlorovinyl)-L-cysteine, and S-(1,2,3,4,4-pentachlorobutadienyl)-L-cysteine. Quantitation of metabolite binding to whole mitochondria and to mitochondrial protein and lipid fractions revealed that as much as 42% of the 35S-label associated with the mitochondria was found in the lipid fraction. Total lipids were also extracted from 35S-treated mitochondria and separated by thin-layer chromatography. 35S-Containing metabolites were found in the lipid fractions from mitochondria treated with each of the conjugates. Lipids from both [35S]CTFC- and [35S]-TFEC-treated mitochondria contained major 35S-labeled lipid adducts which had similar mobility by thin-layer chromatography. Fatty acid analysis, 19F and 31P NMR spectroscopy, and mass spectrometric analyses confirmed that the major TFEC and CTFC adducts are thioamides of phosphatidylethanolamine.

Animals↗
PubMedDate not supplied

Formation, characterization, and immunoreactivity of lysine thioamide adducts from fluorinated nephrotoxic cysteine conjugates in vitro and in vivo.

Fluorinated nephrotoxic cysteine conjugates undergo bioactivation via the beta-lyase pathway to thionoacetyl fluorides (TAF), the putative reactive intermediates. The TAF derived from S-(1,1,2,2,-tetrafluoroethyl)-L-cysteine (TFEC) difluorothionoacetylates amine nucleophiles found in proteins and lipids. A specific antisera, raised against (trifluoroacetamido)lysine adducts formed in vivo after halothane treatment, has previously been used to localize TFEC-derived protein adducts immunohistochemically, and a good correlation between adduction and toxicity was demonstrated. Interestingly, thioamide formation is facilitated by acyl-transfer catalysts such as imidazoles and phenols. However, although putative lysine adducts have been reported to be formed from the related TAF derived from S-(2-chloro-1,1,2-trifluoroethyl)-L-cysteine (CTFC), protein adducts derived from CTFC metabolism have not been completely characterized. In the present investigation we characterize (chlorofluorothionacetamido)lysine (CFTAL) adduct formation during S-(2-chloro-1,1,2-trifluoroethyl)-L-cysteine (CTFC) metabolism, both in vitro and in vivo. Our data indicate that formation of CTFC-derived lysine thioamides was not as dependent on nucleophilic catalysis as observed for TFEC, and this appears to be due to an apparent greater reactivity of the TAF resulting in a higher trapping efficiency in the absence of catalyst. Also, qualitative and quantitative differences in the structures and time course of CTFC versus TFEC adduct breakdown were observed. Antibodies raised against the halothane metabolite protein adduct (trifluoroacetamido)lysine cross-react with specific mitochondrial proteins from the kidneys of TFEC-treated rats. Using this antibody, we have found that the pattern of adducted proteins from TFEC- and CTFC-treated Fischer rats was similar, but the intensity was considerably lower after treatment with equimolar concentrations of CTFC in vivo.

Animals↗
PubMedDate not supplied

Data-driven analysis approach for biomarker discovery using molecular-profiling technologies.

High-throughput molecular-profiling technologies provide rapid, efficient and systematic approaches to search for biomarkers. Supervised learning algorithms are naturally suited to analyse a large amount of data generated using these technologies in biomarker discovery efforts. The study demonstrates with two examples a data-driven analysis approach to analysis of large complicated datasets collected in high-throughput technologies in the context of biomarker discovery. The approach consists of two analytic steps: an initial unsupervised analysis to obtain accurate knowledge about sample clustering, followed by a second supervised analysis to identify a small set of putative biomarkers for further experimental characterization. By comparing the most widely applied clustering algorithms using a leukaemia DNA microarray dataset, it was established that principal component analysis-assisted projections of samples from a high-dimensional molecular feature space into a few low dimensional subspaces provides a more effective and accurate way to explore visually and identify data structures that confirm intended experimental effects based on expected group membership. A supervised analysis method, shrunken centroid algorithm, was chosen to take knowledge of sample clustering gained or confirmed by the first step of the analysis to identify a small set of molecules as candidate biomarkers for further experimentation. The approach was applied to two molecular-profiling studies. In the first study, PCA-assisted analysis of DNA microarray data revealed that discrete data structures exist in rat liver gene expression and correlated with blood clinical chemistry and liver pathological damage in response to a chemical toxicant diethylhexylphthalate, a peroxisome-proliferator-activator receptor agonist. Sixteen genes were then identified by shrunken centroid algorithm as the best candidate biomarkers for liver damage. Functional annotations of these genes revealed roles in acute phase response, lipid and fatty acid metabolism and they are functionally relevant to the observed toxicities. In the second study, 26 urine ions identified from a GC/MS spectrum, two of which were glucose fragment ions included as positive controls, showed robust changes with the development of diabetes in Zucker diabetic fatty rats. Further experiments are needed to define their chemical identities and establish functional relevancy to disease development.

Algorithms↗
PubMedDate not supplied

Dichoptic luminance beat visual evoked potentials in the assessment of binocularity in children.

Direct evidence of a distinct cortical binocular pathway has been provided by the production of nonlinear (difference) beats from dichoptic luminance stimulation in stereonormal adults and the absence or diminution of these beats in stereoblind subjects. We have investigated a clinically useful application of this technique in a pediatric population with potentially abnormal binocular vision. We recorded dichoptic luminance beat visual evoked potentials (VEPs) from 20 children (ages 7 months to 8 years) with abnormal binocular ability secondary to strabismus and/or amblyopia and compared this to a control group of 20 children with normal binocularity. Stereoblind children generated significantly lower dichoptic signal-to-noise ratios than stereonormal children (P < .001). Responses to monoptic multifrequency flicker were not significantly different between the two groups (P = .936). This dichoptic VEP can be performed quickly and easily on young children and gives a quantitative assessment of cortical binocularity that may not be determinable by standard clinical methods. This technique may also prove useful for the preoperative gradation of binocular potential and prediction of postoperative binocular fusion.

Amblyopia↗
PubMedDate not supplied

Quantitation of multiple pathways for the metabolism of nephrotoxic cysteine conjugates using selective inhibitors of L-alpha-hydroxy acid oxidase (L-amino acid oxidase) and cysteine conjugate beta-lyase.

In this study, we have established the selectivity of inhibitors for rat kidney cysteine conjugate beta-lyase and L-alpha-hydroxy acid oxidase (L-amino acid oxidase) and have used these inhibitors to explore the relative roles of these two enzymes in the metabolism of nephrotoxic cysteine conjugates by rat kidney homogenate. In addition, we have investigated the relationship between structure and the metabolism of toxic cysteine conjugates by purified rat kidney L-alpha-hydroxy acid oxidase. With purified enzyme, S-(1,2,3,4,4-pentachlorobutadienyl)-L-cysteine (PCBDC) was about four times more active than S(1,2-dichlorovinyl)-L-cysteine (DCVC). Three alkyl conjugates were less active than DCVC. Purified L-alpha-hydroxy acid oxidase was not inhibited by the beta-lyase inhibitor aminooxyacetic acid but was inactivated by 2-hydroxy-3-butynoate. PCBDC metabolism in rat kidney homogenate was inhibited 74% by aminooxyacetic acid and 42% by 2-hydroxy-3-butynoate, whereas DCVC metabolism was inhibited 77% by aminooxyacetic acid and 28% by 2-hydroxy-3-butynoate. However, only aminooxyacetic acid inhibited the binding of 35S label from [35S]DCVC. Based on these results we have reached three conclusions. First, L-alpha-hydroxy acid oxidase plays a significant role in the metabolism of some cysteine conjugates. Second, metabolism of DCVC by L-alpha-hydroxy acid oxidase does not contribute directly to covalent binding. Third, as much as 65% of DCVC may be metabolized to its corresponding alpha-keto acid. The results are discussed with regard to the nephrotoxicity of cysteine conjugates.

Amino Acid Oxidoreductases↗
PubMedDate not supplied

Metabolism of haloforms to carbon monoxide. II. In vivo studies.

Administration of haloforms (trihalomethanes) to rats led to substantial elevations in blood carbon monoxide levels. The administration of 13C-bromoform led to the formation of similarly enriched 13CO. A dose-dependent relationship between bromoform dose and CO production was observed. It was found that phenobarbital, but not 3-methylcholanthrene, treatment increased the blood CO levels seen after the administration of bromoform as compared to saline-treated controls. Lower blood CO levels were found in rats given 2H-bromoform as compared to rats given bromoform. Furthermore, SKF 525-A significantly inhibited the in vivo metabolism of bromoform to CO. Administration of either diethyl maleate or D-penicillamine did not alter the blood CO levels produced in response to bromoform administration. The in vivo metabolism of haloforms to CO followed the halide order; thus, administration of iodoform yielded the highest blood CO levels, whereas chloroform yielded the lowest levels.

Animals↗