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A J Ward

Publications and source records attributed to A J Ward.

61 records · Page 4Linked to original sources
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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

Evaluation of moisture penetration through skin protectant barriers by paper chromatography.

Currently, there is no convenient and safe experimental method in the literature to evaluate skin protectant formulations for barrier properties. A novel and simple experimental method, based on paper chromatography, was developed to screen a broad spectrum of skin protectants available for moisture penetration. The method involves the use of paper chromatography for determining the Moisture Penetration Rate through a thin barrier film of a given skin protectant product. It is simple, inexpensive, convenient, and safe and was used to evaluate 12 products currently used as skin protectants. This method can be used by formulators in their development efforts and by caregivers to determine the barrier properties of various products in a short period of time without expensive instrumentation or extensive time commitments. Skin irritation resulting from fecal and urinary incontinence is a common problem for geriatric patients (Fiers & Siebert, 1993, Talbot, 1994). Various products on the market today are used as protective barriers to prevent skin irritation from moisture penetration. However, from the caregiver's perspective, there is a need for identifying the best product or the relative ranking of products based on suitable test methods. There is a lack of suitable in vitro test methods to screen skin protectant barriers (Guillemin, Murset, Lob, & Riquez, 1974; Pigatto, Bigardi, Legori, Altomare, & Finzi, 1993) on the market. For any method to be acceptable it must be convenient, simple, reproducible, and safe for use on human volunteers. More importantly, it must have clinical relevance.

Administration, Cutaneous↗