Acrylamide is an alkylating agent which reacts very slowly in direct reactions with DNA and is negative in the Ames test, but is carcinogenic in mice and rats. In order to explain the cancer-initiating properties of acrylamide we have studied DNA adduct formation in vitro with a metabolizing system and in vivo in mice and rats following i.p. administration of 14C-labeled acrylamide. A major adduct found in both species was N-7-(2-carbamoyl-2-hydroxy-ethyl)guanine, formed by reaction of the DNA with the epoxide metabolite glycidamide. The levels of this adduct were similar in the different organs of the two rodent species, which supports the notion that glycidamide is relatively evenly distributed among tissues and that the organ-specificity in acrylamide carcinogenesis cannot be explained by a selective accumulation of the DNA-reactive metabolite in target organs.
BackgroundCamelina sativa, an oilseed crop in the Brassicaceae family, has inspired renewed interest due to its potential for biofuels applications. Little is understood of the nature of the C. sativa genome, however. A study was undertaken to characterize two genes in the fatty acid biosynthesis pathway, fatty acid desaturase (FAD) 2 and fatty acid elongase (FAE) 1, which revealed unexpected complexity in the C. sativa genome.ResultsIn C. sativa, Southern analysis indicates the presence of three copies of both FAD2 and FAE1 as well as LFY, a known single copy gene in other species. All three copies of both CsFAD2 and CsFAE1 are expressed in developing seeds, and sequence alignments show that previously described conserved sites are present, suggesting that all three copies of both genes could be functional. The regions downstream of CsFAD2 and upstream of CsFAE1 demonstrate co-linearity with the Arabidopsis genome. In addition, three expressed haplotypes were observed for six predicted single-copy genes in 454 sequencing analysis and results from flow cytometry indicate that the DNA content of C. sativa is approximately three-fold that of diploid Camelina relatives. Phylogenetic analyses further support a history of duplication and indicate that C. sativa and C. microcarpa might share a parental genome.ConclusionsThere is compelling evidence for triplication of the C. sativa genome, including a larger chromosome number and three-fold larger measured genome size than other Camelina relatives, three isolated copies of FAD2, FAE1, and the KCS17-FAE1 intergenic region, and three expressed haplotypes observed for six predicted single-copy genes. Based on these results, we propose that C. sativa be considered an allohexaploid. The characterization of fatty acid synthesis pathway genes will allow for the future manipulation of oil composition of this emerging biofuel crop; however, targeted manipulations of oil composition and general development of C. sativa should consider and, when possible take advantage of, the implications of polyploidy.
The major degradation product of the volatile anesthetic sevoflurane, the haloalkene fluoromethyl-2,2-difluoro-1-(trifluoromethyl)vinyl ether (FDVE or "compound A"), is nephrotoxic in rats. FDVE undergoes complex metabolism and bioactivation, which mediates the nephrotoxicity. Nevertheless, the molecular and cellular mechanisms of FDVE toxification are unknown. This investigation evaluated the gene expression profile of kidneys in rats administered a nephrotoxic dose of FDVE. Male Fischer 344 rats (five per group) received 0.25 mmol/kg intraperitoneal FDVE or corn oil (controls) and were sacrificed after 24 or 72 h. Urine output and kidney histological changes were quantified. Kidney RNA was extracted for microarray analysis using Affymetrix GeneChip Rat Expression Array 230A arrays. Quantitative real-time PCR confirmed the modulation of several genes. FDVE caused significant diuresis and necrosis at 24 h, with normal urine output and evidence of tubular regeneration at 72 h. There were 517 informative genes that were differentially expressed >1.5-fold (p < 0.05) versus control at 24 h, of which 283 and 234 were upregulated and downregulated, respectively. Major classes of upregulated genes included those involved in apoptosis, oxidative stress, and inflammatory response (mostly at 24 h), and regeneration and repair; downregulated genes were generally associated with transporters and intermediary metabolism. Among the quantitatively most upregulated genes were kidney injury molecule, osteopontin, clusterin, tissue inhibitor of metalloproteinase 1, and TNF receptor 12, which have been associated with other forms of nephrotoxicity, and angiopoietin-like protein 4, glycoprotein nmb, ubiquitin hydrolase, and HSP70. Microarray results were confirmed by quantitative real-time PCR. FDVE causes rapid and brisk changes in gene expression, providing potential insights into the mechanism of FDVE toxification, and potential biomarkers for FDVE nephrotoxicity which are more sensitive than conventional measures of renal function.
Methoxyflurane nephrotoxicity seems to result from O-demethylation, which forms both fluoride and DCAA. Because their co-formation is unique to methoxyflurane compared with other volatile anesthetics and they are more toxic than fluoride alone, this suggests a new hypothesis of methoxyflurane nephrotoxicity. This may explain why increased fluoride formation from methoxyflurane, but not other anesthetics, is associated with toxicity. These results may have implications for the interpretation of clinical anesthetic defluorination, use of volatile anesthetics, and the laboratory methods used to evaluate potential anesthetic toxicity.
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