Antioxidant survey to assess antagonism to redox stress using a prokaryotic and an eukaryotic system

Using a prokaryote (Escherichia coli) and a metazoa-resembling eukaryote (Ochromonas danica), we surveyed antioxidants which might overcome redox stress imposed by menadione sodium bisulphite (MD) and buthionine sulphoximine (BSO). BSO oxidant stress was evident only inO. danica; MD oxidant stress was evident in both organisms. Glutathione, its precursors, e.g. cysteine, homocysteine, and 2-oxo-4-thiazolidine carboxylic acid, and red blood cells, emerged as prime antioxidants for relieving BSO and MD oxidant stress. BSO and MD oxidant activity and antioxidant-annulling effect inO. danica were judged comparable to those found in animal cells whereas the resultsE. coli were not entirely equivalent. TheO. danica system emerged as a practical, rapid, and useful system for pinpointing oxidant stressors and antioxidants, and shows promise for studies with mammalian systems.     

(−)-Epigallocatechin gallate,a polyphenolic tea antioxidant,inhibits peroxynitritemediated formation of 8-oxodeoxyguanosine and 3-nitrotyrosine

Reaction with peroxynitrite at pH 7.4 and 37°C was found to increase the 8-oxodeoxyguanosine levels in calf thymus DNA 35-38-fold. This oxidation of deoxyguanosine, as well as the peroxynitrite-mediated nitration of tyrosine to 3-nitrotyrosine, was significantly inhibited by ascorbic acid, glutathione and (–)-epigallocatechin gallate, a polyphenolic antioxidant present in tea. For 50% inhibition of the oxidation of deoxyguanosine to 8-oxodeoxyguanosine, 1.1, 7.6 of 0.25 mM ascorbate, glutathione or (–)-epigallocatechin gallate, respectively, was required. For 50% inhibition of tyrosine nitration, the respective concentrations were 1.4, 4.6 or 0.11 mM. Thus, (–)-epigallocatechin gallate is a significantly better inhibitor of both reactions than either ascorbate or glutathione. Reaction of (–)-epigallocatechin gallate with peroxynitrite alone resulted in the formation of a number of products. Ultraviolet spectra of two of these suggest that the tea polyphenol and/or its oxidation products are nitrated by peroxynitrite.     

Complex reorganization and predominant non-homologous repair following chromosomal breakage in karyotypically balanced germline rearrangements and transgenic integration

We defined the genetic landscape of balanced chromosomal rearrangements at nucleotide resolution by sequencing 141 breakpoints from cytogenetically interpreted translocations and inversions. We confirm that the recently described phenomenon of 'chromothripsis' (massive chromosomal shattering and reorganization) is not unique to cancer cells but also occurs in the germline, where it can resolve to a relatively balanced state with frequent inversions. We detected a high incidence of complex rearrangements (19.2%) and substantially less reliance on microhomology (31%) than previously observed in benign copy-number variants (CNVs). We compared these results to experimentally generated DNA breakage-repair by sequencing seven transgenic animals, revealing extensive rearrangement of the transgene and host genome with similar complexity to human germline alterations. Inversion was the most common rearrangement, suggesting that a combined mechanism involving template switching and non-homologous repair mediates the formation of balanced complex rearrangements that are viable, stably replicated and transmitted unaltered to subsequent generations.     

ZG35CrMnSiRe热处理的试验研究

ZG35CrMnSiRe铸钢经不同的热处理,对其强度、硬度、塑性和冲击韧性等机械性能进行测定,实验结果表明其综合机械性能良好。     

两种前处理方法测定菜籽油中6种金属元素含量的比较

目的:比较不同前处理方法对菜籽油中6种金属元素含量测定的影响.方法:分别使用干灰化法和湿消化法进行前处理,并用火焰原子吸收分光光度法分别测定15个菜籽油样品中铅、镉、铜、铁等6种金属元素的含量.结果:干灰化法和湿消化法处理后所测菜籽油样品中Pb、Cu含量差异无统计学意义(P>0.05),Fe、Mn含量差异有统计学意义(P<0.05).各元素的R2均大于0.99,干灰化法的加标回收率81.90%~107.45%,湿消化法的加标回收率99.90%~108.05%.均满足痕量分析要求,相比之下,干灰化法与湿消化法的加标回收率比较差异有统计学意义(P<0.05),且湿消化法-火焰原子吸收分光光度法测定油样的回收率相对干灰化法的要高.结论:两种方法都能获得准确的测定结果;湿消化法要优于干灰化法.     

The p66shc adaptor protein controls oxidative stress response and life span in mammals

Gene mutations in invertebrates have been identified that extend life span and enhance resistance to environmental stresses such as ultraviolet light or reactive oxygen species. In mammals, the mechanisms that regulate stress response are poorly understood and no genes are known to increase individual life span. Here we report that targeted mutation of the mouse p66shc gene induces stress resistance and prolongs life span. p66shc is a splice variant of p52shc/p46shc (ref. 2), a cytoplasmic signal transducer involved in the transmission of mitogenic signals from activated receptors to Ras. We show that: (1) p66shc is serine phosphorylated upon treatment with hydrogen peroxide (H2O2) or irradiation with ultraviolet light; (2) ablation of p66shc enhances cellular resistance to apoptosis induced by H2O2 or ultraviolet light; (3) a serine-phosphorylation defective mutant of p66shc cannot restore the normal stress response in p66shc-/- cells; (4) the p53 and p21 stress response is impaired in p66shc-/- cells; (5) p66shc-/- mice have increased resistance to paraquat and a 30% increase in life span. We propose that p66shc is part of a signal transduction pathway that regulates stress apoptotic responses and life span in mammals.     

Podocalyxin enhances the adherence of cells to platelets

Podocalyxin (PODXL) is a mucin protein of the CD34 family expressed in kidney glomerular podocytes, vascular endothelium, progenitor bone marrow and tumor cells. It is assumed that PODXL plays an anti-adherent role in kidney podocytes. CHO cells stably expressing human PODXL (CHO-PODXL) or human tumor cells (Tera-1) inherently expressing PODXL showed increased adherence to platelets. The adherence of cells was inhibited (70%) by blockers of platelet P-selectin, prevented by the soluble ectodomain of human PODXL (PODXL-Δ) or by the arginine-glycine-aspartate (RGDS) peptide and partially impeded by inhibition of integrin αVβ3/αVβ5, suggesting a coordinated action of P-selectin and integrins. Colocalization of platelet P-selectin and PODXL expressed on CHO cells was demonstrated by confocal immunofluorescence. No adherence to platelets was observed when PODXL was expressed in glycomutant CHO cells deficient in sialic acid. Received 14 August 2007; received after revision 12 September 2007; accepted 13 September 2007     

Germline gain-of-function mutations in SOS1 cause Noonan syndrome

Noonan syndrome, the most common single-gene cause of congenital heart disease, is characterized by short stature, characteristic facies, learning problems and leukemia predisposition. Gain-of-function mutations in PTPN11, encoding the tyrosine phosphatase SHP2, cause approximately 50% of Noonan syndrome cases. SHP2 is required for RAS-ERK MAP kinase (MAPK) cascade activation, and Noonan syndrome mutants enhance ERK activation ex vivo and in mice. KRAS mutations account for <5% of cases of Noonan syndrome, but the gene(s) responsible for the remainder are unknown. We identified missense mutations in SOS1, which encodes an essential RAS guanine nucleotide-exchange factor (RAS-GEF), in approximately 20% of cases of Noonan syndrome without PTPN11 mutation. The prevalence of specific cardiac defects differs in SOS1 mutation-associated Noonan syndrome. Noonan syndrome-associated SOS1 mutations are hypermorphs encoding products that enhance RAS and ERK activation. Our results identify SOS1 mutants as a major cause of Noonan syndrome, representing the first example of activating GEF mutations associated with human disease and providing new insights into RAS-GEF regulation.