葡萄糖氧化酶诱导肝细胞氧化应激模型的建立

目的：研究不同浓度葡萄糖氧化酶（GO）对人肝细胞L02氧化应激水平的影响,以确定建立肝细胞氧化应激模型的合适浓度。方法：用不同浓度GO干预L02肝细胞2h,MTT法检测细胞的存活率,流式细胞术检测细胞内活性氧簇（ROS）,荧光强度（FI）来表示ROS水平。分光光度法检测检测细胞MDA、GSH,速率法检测细胞培养液LDH、AST和ALT的水平。结果：①随GO浓度增加,肝细胞的存活率逐渐降低,其中75U/L、100U/L和125U/L组存活率显著低于对照组（P〈0.05）。②随GO浓度增加,MDA含量逐渐增高,其中50U/L、75U/L、100U/L、125U/L组MDA水平较对照组显著增高（P〈0.05）。GSH水平随GO浓度增高而逐渐减低,各干预组较对照组均显著降低（P〈0.05）。GO各干预组FI均较对照组显著降低（P〈0.05）。③各干预组LDH活性均显著高于对照组（P〈0.05）,50U/L、75U/L、100U/L、125U/L干预组AST与ALT水平均较对照组显著增高（P〈0.05）。结论：GO能引起的肝细胞氧化应激损伤有剂量依赖性,100U/L是建立肝细胞氧化应激的合适浓度。     

Crosstalk between calcium and reactive oxygen species signaling in cancer

The interplay between Ca²⁺ and reactive oxygen species (ROS) signaling pathways is well established, with reciprocal regulation occurring at a number of subcellular locations. Many Ca²⁺ channels at the cell surface and intracellular organelles, including the endoplasmic reticulum and mitochondria are regulated by redox modifications. In turn, Ca²⁺ signaling can influence the cellular generation of ROS, from sources such as NADPH oxidases and mitochondria. This relationship has been explored in great depth during the process of apoptosis, where surges of Ca²⁺ and ROS are important mediators of cell death. More recently, coordinated and localized Ca²⁺ and ROS transients appear to play a major role in a vast variety of pro-survival signaling pathways that may be crucial for both physiological and pathophysiological functions. While much work is required to firmly establish this Ca²⁺-ROS relationship in cancer, existing evidence from other disease models suggests this crosstalk is likely of significant importance in tumorigenesis. In this review, we describe the regulation of Ca²⁺ channels and transporters by oxidants and discuss the potential consequences of the ROS-Ca²⁺ interplay in tumor cells.     

Nitric oxide signaling: classical, less classical, and nonclassical mechanisms

Although nitric oxide (NO) was identified more than 150 years ago and its effects were clinically tested in the form of nitroglycerine, it was not until the decades of 1970-1990 that it was described as a gaseous signal transducer. Since then, a canonical pathway linked to cyclic GMP (cGMP) as its quintessential effector has been established, but other modes of action have emerged and are now part of the common body of knowledge within the field. Classical (or canonical) signaling involves the selective activation of soluble guanylate cyclase, the generation of cGMP, and the activation of specific kinases (cGMP-dependent protein kinases) by this cyclic nucleotide. Nonclassical signaling alludes to the formation of NO-induced posttranslational modifications (PTMs), especially S-nitrosylation, S-glutathionylation, and tyrosine nitration. These PTMs are governed by specific biochemical mechanisms as well as by enzymatic systems. In addition, a less classical but equally important pathway is related to the interaction between NO and mitochondrial cytochrome c oxidase, which might have important implications for cell respiration and intermediary metabolism. Cross talk trespassing these necessarily artificial conceptual boundaries is progressively being identified and hence an integrated systems biology approach to the comprehension of NO function will probably emerge in the near future.     

Cross talk between mitochondria and NADPH oxidases

Reactive oxygen species (ROS) play an important role in physiological and pathological processes. In recent years, a feed-forward regulation of the ROS sources has been reported. The interactions between the main cellular sources of ROS, such as mitochondria and NADPH oxidases, however, remain obscure. This work summarizes the latest findings on the role of cross talk between mitochondria and NADPH oxidases in pathophysiological processes. Mitochondria have the highest levels of antioxidants in the cell and play an important role in the maintenance of cellular redox status, thereby acting as an ROS and redox sink and limiting NADPH oxidase activity. Mitochondria, however, are not only a target for ROS produced by NADPH oxidase but also a significant source of ROS, which under certain conditions may stimulate NADPH oxidases. This cross talk between mitochondria and NADPH oxidases, therefore, may represent a feed-forward vicious cycle of ROS production, which can be pharmacologically targeted under conditions of oxidative stress. It has been demonstrated that mitochondria-targeted antioxidants break this vicious cycle, inhibiting ROS production by mitochondria and reducing NADPH oxidase activity. This may provide a novel strategy for treatment of many pathological conditions including aging, atherosclerosis, diabetes, hypertension, and degenerative neurological disorders in which mitochondrial oxidative stress seems to play a role. It is conceivable that the use of mitochondria-targeted treatments would be effective in these conditions.     

Effect on lycopene, β-carotene,ascorbic acid and phenolic content in tomato fruits infected by Alternaria alternata and its toxins (TeA,AOH and AME)

Tomato is considered as one of the most important sources of nutrients such as lycopene, β-carotene, flavonoids, ascorbic acid (vitamin C) and hydroxyl-cinnamic acid derivatives. The quality and quantity of nutrients in tomato fruits were decreased during the severe infection of Alternaria alternata. The present study deals with the estimation of lycopene, β-carotene, phenolic and ascorbic acid content in tomato fruits which were infected with A. alternata and its toxins such as tenuazonic acid (TeA), alternariol (AOH) and alternariol monomethyl ether (AME). The lycopene, β-carotene, ascorbic acid and phenolic content were found lowest in pathogen-infected fruits i.e. (0.66 ± 0.03 mg/g), (0.14 ± 0.01 mg/g), (1.89 ± 0.2 mg/g) and (0.58 ± 0.05 mg/g), respectively, followed by toxins-treated samples as compared to the control. The results concluded that A. alternata mostly affects the nutritional values of tomato fruits due to the combined effect of the toxins.     

Morphological variability and genetic identity in Rhinoestrus spp. causing horse nasal myiasis

Larvae of Rhinoestrus purpureus (Brauer) and Rhinoestrus usbekistanicus Gan (Diptera: Oestridae) cause nasal myiases of equids. During a recent epidemiological survey in southern Italy some morphological and taxonomical doubts arose concerning the identification of Rhinoestrus third stage larvae on the basis of the features of the posterior spiracles and the distribution of dorsal spines on the third segment. Four different morphotypes were retrieved: R. usbekistanicus-like, R. purpureus-like and two morphotypes with shared features. The genes encoding for the mitochondrial cytochrome oxidase I (COI) and for the ribosomal subunits 16S and 28S of the four morphotypes of Rhinoestrus were investigated to determine whether they belonged to a single taxon or they displayed genetic differences indicative of more than one species. The three genes showed a very low level of sequence variation (COI 0-0.43%, 16S 0-1.45%, 28S 0-0.23%) falling within the intraspecific ranges previously described for Oestridae species. Finally, the peritreme features and the spinulation of the third segment of the four morphotypes examined could not be used to differentiate the two species.     

Variation in the activity of some enzymes of photorespiratory metabolism in C4 grasses

BACKGROUND AND AIMS: Photorespiration occurs in C4 plants, although rates are small compared with C3 plants. The amount of glycine decarboxylase in the bundle sheath (BS) varies among C4 grasses and is positively correlated with the granal index (ratio of the length of appressed thylakoid membranes to the total length of all thylakoid membranes) of the BS chloroplasts: C4 grasses with high granal index contained more glycine decarboxylase per unit leaf area than those with low granal index, probably reflecting the differences in O2 production from photosystem II and the potential photorespiratory capacity. Thus, it is hypothesized that the activities of peroxisomal enzymes involved in photorespiration are also correlated with the granal development. METHODS: The granal development in BS chloroplasts was investigated and activities of the photorespiratory enzymes assayed in 28 C4 grasses and seven C3 grasses. KEY RESULTS: The NADP-malic enzyme grasses were divided into two groups: one with low granal index and the other with relatively high granal index in the BS chloroplasts. Both the NAD-malic enzyme and phosphoenolpyruvate carboxykinase grasses had high granal index in the BS chloroplasts. No statistically significant differences were found in activity of hydroxypyruvate reductase between the C3 and C4 grasses, or between the C4 subtypes. The activity of glycolate oxidase and catalase were smaller in the C4 grasses than in the C3 grasses. Among the C4 subtypes, glycolate oxidase activities were significantly smaller in the NADP-malic enzyme grasses with low granal index in the BS chloroplasts, compared with in the C4 grasses with substantial grana in the BS chloroplasts. CONCLUSIONS: There is interspecies variation in glycolate oxidase activity associated with the granal development in the BS chloroplasts and the O2 production from photosystem II, which suggests different potential photorespiration capacities among C4 grasses.     

Simultaneous determination of ascorbic acid and dehydroascorbic acid in cultures of C3H/10T1/2 cells

Summary A reproducible method is described for the separation and quantification of ascorbic acid and dehydroascorbic acid by ion-pairing reverse-phase high performance liquid chromatography and detection by absorbance at 232 nm. Lowest detectable concentrations with a linear response of detection were 5 nmol for ascorbic acid and 50 nmol for dehydroascorbic acid. This method was applied to the analysis of C3H/10T1/2 cells and culture medium after influx or efflux experiments and single or multiple treatments with ascorbic acid. Subsequent measurement of the radioactivity in the eluted fractions increased the detectability of both ascorbic acid and dehydroascorbic acid to 10 to 20 pmol. This research was supported by grant CA 09320 and CA 31574 from the National Cancer Institute, Bethesda, MD, and grant BC441 from The American Cancer Society.     

Respiration-dependent proton translocation and the mechanism of protonmotive force generation in Nitrobacter winogradskyi

Abstract Proton translocation associated with electron flow to oxygen has been observed with cells of Nitrobacter winogradskyi in the presence of either potassium ferrocyanide or isoascorbate plus N , N , N ', N ' tetramethyl- p -phenylenediamine. The data are consistent with a proton pumping function for the terminal oxidase, cytochrome aa ₃, in this organism as the mechanism for generating a protonmotive force. The failure of previous work with Nitrobacter [4] to detect proton translocation linked to oxidation of nitrite, the physiological substrate, is discussed.