Chemopreventive activities of etodolac and oxyphenbutazone against mouse skin carcinogenesis

Previous cancer chemoprevention studies have demonstrated that the non-steroidal anti-inflammatory drugs (NSAIDs) can be effective in suppressing the development of various human malignancies. Recently we identified the possible anti-tumor promoting potentials of 14 new NSAIDs in the Epstein–Barr virus early antigen activation assay induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). In this study we report the inhibition of 7,12-dimethylbenz (a) anthracene (DMBA) induced two-stage mouse skin carcinogenesis by etodolac (ETD), one of the most potent NSAIDs identified in our in vitro cancer chemopreventive screening of this group of drugs. Topical administration of ETD at a very low dose of 85 nmol showed a significant decrease in both tumor incidence and burden. This effect is also accompanied by a delay in the tumor latency period. Since ETD showed potent chemopreventive activity in both in vitro and in vivo studies, it warrants prompt consideration for trial in humans as a potential cancer chemopreventive agent. We also investigated oxyphenbutazone (OPB) another commonly used NSAID for its cancer chemopreventive effect on peroxynitrite (PN) induced-TPA promoted skin tumors in the mouse. Following tumor initiation with 390 nmol of PN, the skin tumor promotion with 1.7 nmol of TPA was significantly inhibited by oral administration of 0.0025% OPB. The results demonstrate that OPB is a potent cancer chemopreventive agent in the highly sensitive in vivo mouse test model we used.     

Potentiation of tumour promotion by topical application of argemone oil/isolated sanguinarine alkaloid in a model of mouse skin carcinogenesis

Several incidences of adverse effects on human health have been reported in many countries, due to consumption of edible oil adulterated with argemone oil (AO). The clinical manifestation of the disease is commonly referred to as epidemic dropsy. Our prior studies have shown that AO and isolated sanguinarine alkaloid (SANG) possess genotoxic and tumour initiating activity. In this study, the effect of AO/SANG was investigated on the development of tumour formation in mice using 7,12-dimethylbenz (a) anthracene (DMBA) initiated followed by tetradecanoyl phorbol acetate (TPA)-promoted skin tumour protocol. Single application of AO (300 μl) or SANG (4.5 μmol) when used during initiation phase in DMBA/TPA group did not reveal substantial difference in tumourigenic response. However, twice weekly application of AO (100 μl) or SANG (1.5 μmol) during promotion phase (25 weeks) resulted in enhanced tumourigenic response by ≥30% in DMBA/TPA treated group along with significant decrease in dermal tyrosinase (45–49%), histidase (30–32%), superoxide dismutase (53–56%), catalase (41%), GSH reductase (37–40%) and GSH-peroxidase activity (29–33%) compared to control. Furthermore, significant decrease of epidermal GSH (64–66%) content and enhanced formation of lipid peroxides (96–121%) was noticed following AO or SANG treatment during promotion phase to DMBA/TPA induced animals indicating the modified pro-oxidant status in skin. Although dermal biochemical parameters were also altered by AO or SANG when used during initiation phase in DMBA/TPA treated animals, nonetheless, the response in these parameters were relatively more when AO or SANG were used during promotion phase in DMBA/TPA treated animals. These results clearly suggest that AO and SANG have the ability to enhance the tumourigenic response, which may have relevance to its carcinogenic potential.     

An association among iron,copper, zinc,and selenium,and antioxidative status in dyslipidemic pediatric patients with glycogen storage disease types IA and III

Dyslipidemia in patients with glycogen storage disease types Ia (GSD Ia) and III (GSD III) does not lead to premature atherosclerosis. The aim of this study was to investigate the association among serum copper (Cu), zinc (Zn), iron (Fe), and selenium (Se) concentrations, and their carrier proteins: ceruloplasmin, albumin, and related antioxidant enzyme activities [superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), paraoxonase (PON), and arylesterase (ARYL)] in 20 GSD Ia and 14 III patients compared to age and sex matched 20 healthy subjects. Erythrocyte oxidative stress was measured by erythrocyte thiobarbituric acid reactive substances (eTBARSs). Hypertriglyceridemia [333 (36–890) mg/dL] in GSD Ia and hypercholesterolemia with elevated LDL-cholesterol [188 (91–313) mg/dL] and decreased HDL-cholesterol [32(23–58) mg/dL] levels in GSD III were found. Serum Cu, Fe, and Zn showed no significant differences between groups. However, Se 60 (54–94), 81 (57–127) μg/L, ceruloplasmin 21 (10–90), 27 (23–65) μg/L, and albumin 2.4 (1.7–5.1), 2.8 (1.8–4.06) g/dL levels were decreased in GSD Ia and III groups, respectively, in comparison with the controls [Se 110 (60–136) μg/L, ceruloplasmin 72 (32–94) μg/L, and albumin 4.4 (4–4.8) g/dL)]. In spite of high oxidative stress in erythrocyte detected by elevated eTBARS/Hb levels in GSD group [674.8 (454.6–948.2) for GSD Ia, 636.3 (460.9–842.1) for GSD III, and 525.6 (449.2–612.6)], the activities of CAT, SOD, ARYL, and PON in GSD patients were not different from the controls. GPx activity was decreased in GSD Ia [3.7 (1.8–7.1) U/mL] and GSD III [4.2 (2.2–8.6) U/mL] compared with healthy controls [7.1 (2.9–16.2) U/mL].In conclusion, this study supplied the data for trace elements, their carrier, and antioxidative enzymes in the patients with GSD Ia and III. The trace elements and anti-oxidative enzyme levels in GSD patients failed to explain the atherosclerotic escape phenomenon reported in these patients.     

Copper/zinc and copper/selenium ratios,and oxidative stress as biochemical markers in recurrent aphthous stomatitis

ProjectRecurrent aphthous stomatitis (RAS) is a common oral mucosal disorder characterized by recurrent, painful oral aphthae, and oxidative stress presumably contributes to its pathogenesis. The aim of this study is to scrutinize the relationship between oxidative stress and serum trace elements (copper, Cu; zinc, Zn; selenium, Se), and to evaluate the ratios of Cu/Zn and Cu/Se in this disorder.ProcedurePatients with RAS (n = 33) and age- and sex-matched healthy control subjects (n = 30) were enrolled in this study. Malondialdehyde (MDA) concentrations in plasma and the activities of superoxide dismutase (SOD1; CuZnSOD), glutathione peroxidase (GPx) and catalase (CAT) in erythrocyte were determined as spectrophotometric. Also, the levels of Se, Zn and Cu in serum were determined on flame and furnace atomic absorption spectrophotometer using Zeeman background correction.Results and conclusionsOxidative stress was confirmed by the significant elevation in plasma MDA, and by the significant decrease in CAT, SOD1, and GPx (p < 0.05). When compared to controls, Zn and Se levels were significantly lower in patients, whereas Cu levels was higher in RAS patients than those in controls (p < 0.05). In addition, the correlation results of this study were firstly shown that there were significant and positive correlations between Se–CAT, Se–GPx, and Cu–MDA parameters, but negative correlations between Se–Cu, Se–MDA, Cu–CAT, Cu–SOD1 and Cu–GPx parameters in RAS patients. Furthermore, the ratios of Cu/Zn and Cu/Se were significantly higher in the patients than the control subjects (p < 0.05). Our results indicated that lipid peroxidation associated with the imbalance of the trace elements seems to play a crucial role in the pathogenesis of RAS. Furthermore, the serum Cu/Zn and Cu/Se ratios may be used as biochemical markers in these patients.     

A-ring modified betulinic acid derivatives as potent cancer preventive agents

Ten new 3,4-seco betulinic acid (BA) derivatives were designed and synthesized. Among them, compounds 7–15 exhibited enhanced chemopreventive ability in an in vitro short-term 12-O-tetradecanoylphorbol-13-acetate (TPA) induced Epstein–Barr virus early antigen (EBV-EA) activation assay in Raji cells. Specifically, analogs with a free C-28 carboxylic acid, including 7, 8, 11, and 13, inhibited EBV-EA activation significantly. The most potent compound 8 displayed 100% inhibition at 1 × 10³ mol ratio/TPA and 73.4%, 35.9%, and 8.4% inhibition at 5 × 10², 1 × 10², and 1 × 10 mol ratio/TPA, respectively, comparable with curcumin at high concentration and better than curcumin at low concentration. The potent chemopreventive activity of novel seco A-ring BAs (8 and 11) was further confirmed in an in vivo mouse skin carcinogenesis assay.     

Oxidative stress programming in a rat model of postnatal early overnutrition — role of insulin resistance

Postnatal early overfeeding (EO) is related to later development of overweight and other metabolic disorders. As oxidative stress is implicated in most human diseases, as obesity and diabetes, we decided to study some parameters related to oxidative stress and insulin signaling in liver from EO animals in adult life. To induce EO, litter size was reduced to three pups per litter (SL: small litter) and groups with normal litter size (NL:10 pups per litter) were used as control. After weaning, rats had free access to standard diet and water. Body weight and food intake were monitored daily and offspring were killed at 180 days-old. Significant differences had P<.05 or less. As expected, SL rats had hyperphagia, higher body weight and higher visceral fat mass at weaning and adulthood. In liver, postnatal EO programmed for lower catalase (? 42%), superoxide dismutase (? 45%) and glutathione peroxidase (? 65%) activities. The evaluation of liver injury in adult SL group showed lower nitrite content (? 10%), higher liver and plasma malondialdehyde content (+ 25% and 1.1-fold increase, respectively). No changes of total protein bound carbonyl or Cu/Zn superoxide dismutase protein expression in liver were detected between the groups. Regarding insulin signaling pathway in liver, SL offspring showed lower IRβ (? 66%), IRS1 (? 50%), phospho-IRS1 (? 73%), PI3-K (? 30%) and Akt1 (? 58%). Indeed, morphological analysis showed that SL rats presented focal areas of inflammatory cell infiltrate and lipid drops in their cytoplasm characterizing a microsteatosis. Thus, we evidenced that postnatal EO can program the oxidative stress in liver, maybe contributing for impairment of the insulin signaling.     

Characterization and efficacy of Muscodor cinnamomi in promoting plant growth and controlling Rhizoctonia root rot in tomatoes

Muscodor cinnamomi was selected and investigated for its in vitro ability to produce indole-3-acetic acid (IAA) to solubilize different toxic metal (Ca, Co, Cd, Cu, Pb, Zn)-containing insoluble minerals and tolerance to metals, herbicides and an insecticide. The results indicated that this fungus is able to produce IAA (45.36 ± 2.40 μg ml⁻¹) in liquid media. This phytohormone stimulated coleoptile elongation, and increased seed germination and root elongation of tested plants. The metal tolerance and solubilizing ability depended on the type of insoluble minerals. M. cinnamomi showed the highest growth tolerance on Ca-containing media at 150 mM, followed by Zn-containing media at 100 mM and Cd-containing media at 10 mM. This fungus tolerated the three herbicides (2,4-d-dimethylammonium, glyphosate and paraquat dichloride) and an insecticide (methomyl) at the recommended dosages for field application. Moreover, M. cinnamomi completely controlled Rhizoctonia solani AG-2 root rot in tomato plants, and increased root length, shoot dry weight and root dry weight. This is the first report of in vitro IAA production, solubilization of insoluble metal minerals, and tolerance to herbicides, an insecticide and metals as well as the plant growth promoting ability of M. cinnamomi.     

A combination of ascorbic acid and α-tocopherol or a combination of Mg and Zn are both able to reduce the adverse effects of lindane-poisoning on rat brain and liver

The purpose of this study, carried out on male Wistar rats, was to evaluate the beneficial effects of supplementation with ascorbic acid (Vit C) and α-tocopherol (Vit E) or with Mg and Zn upon lindane-induced damages in liver and brain. Under our experimental conditions, lindane poisoning (5 mg/kg body weight per day for 3 days) resulted in (1) an increased level of plasma glucose, cholesterol and triglycerides, (2) an increased activity of LDH, ALP, AST, ALT, (3) an oxidative stress in liver and brain as revealed by an increased level of lipids peroxidation (TBARS) and a decrease of glutathione-peroxidase, superoxide dismutase and catalase activities in liver and brain. In conclusion, both Vit C + E or Mg + Zn treatments display beneficial effects upon oxidative stress induced by lindane treatment in liver and brain.     

Characterization of the major plasma protein of the eastern oyster,Crassostrea virginica,and a proposed role in host defense

The major plasma protein of the eastern oyster, Crassostrea virginica, was purified, characterized and named dominin. SDS-PAGE analyses revealed that dominin consistently made up more than 40% of eastern oyster plasma and extrapallial fluid proteins. Three different forms of dominin were observed under non-reducing conditions. PCR and RACE primers designed from partial amino acid sequences obtained by tandem mass spectrometry of purified dominin identified 720 bp of complete cDNA encoding 192 amino acid residues. Based on the deduced amino acid sequence of mature dominin, its molecular mass was calculated to be 19,389 Da and was lower than the molecular mass of purified dominin measured by MALDI. This difference is likely due to post-translational modifications of dominin as the purified protein was found to be glycolysated, phosphorylated and likely sulfated. The amino acid sequence showed high similarity to the major plasma protein of the Pacific oyster (Crassostrea gigas), cavortin, and of the green-lipped mussel (Perna canaliculus), pernin, and to a recently described protein labeled as an extracellular superoxide dismutase from the Sydney rock oyster Saccostrea glomerata. While dominin was found to possess a Cu/Zn superoxide dismutase (SOD) domain, the domain was not completely conserved which explained why purified dominin lacked SOD activity. Dominin mRNA was detected in hemocytes by in situ hybridization and its expression measured by quantitative real time RT-PCR was significantly higher in winter than summer. Although the function(s) of dominin and homologous proteins is uncertain, the reported ability of cavortin to sequester iron and possibly limit the availability of this essential metal to pathogens suggests a potential role in host defense for this group of dominant plasma proteins. Other possible functions of dominin in antioxidation, wound repair, metal transport and shell mineralization are discussed leading us to conclude that dominin is likely a multifunctional protein.     

Regulation of the synthesis of superoxide dismutases in rat lungs during oxidant and hyperthermic stresses

Heat shock proteins are induced at normal temperatures by oxidants and during reoxygenation following hypoxia. We now report cyanide-resistant O2 consumption increased 30-50% in rat lungs exposed to heat shock or reoxygenation following hypoxia. The synthesis of Cu,Zn superoxide dismutase, but not Mn superoxide dismutase, was increased in rat lung slices by in vivo hyperthermia (39 degrees C), by in vitro heat shock (41 degrees C), and during incubation of lung slices with the Cu chelator diethyldithiocarbamate, which decreased the activity of Cu,Zn superoxide dismutase. The heat shock-induced increase in Cu,Zn superoxide dismutase developed 2 h later than the induction of heat shock proteins and was not blocked by actinomycin D. The rates of synthesis of both superoxide dismutases were decreased 50% by hypoxia and failed to increase during reoxygenation. During hypoxia the activity of Cu,Zn superoxide dismutase decreased about 50%, but the activity of Mn superoxide dismutase remained unchanged. We conclude that hyperthermia increases the synthesis of Cu,Zn superoxide dismutase, the synthesis of Cu,Zn superoxide dismutase and Mn superoxide dismutase are not coordinately regulated by hyperthermia or by the oxidant stress produced by lowering the activity of Cu,Zn superoxide dismutase, and the synthesis of heat shock proteins and Cu,Zn superoxide dismutase are regulated at different levels of gene expression.     

Copper,zinc superoxide dismutase of Curcuma aromatica is a kinetically stable protein

This study details on cloning and characterization of Cu,Zn superoxide dismutase (Ca–Cu,Zn SOD) from a medicinally important plant species Curcuma aromatica. Ca–Cu,Zn SOD was 692 bp with an open reading frame of 459 bp. Expression of the gene in Escherichia coli cells followed by purification yielded the enzyme with K_m of 0.047 ± 0.008 μM and V_max of 1250 ± 24 units/mg of protein. The enzyme functioned (i) across a temperature range of −10 to +80 °C with temperature optima at 20 °C; and (ii) at pH range of 6–9 with optimum activity at pH 7.8. Ca–Cu,Zn SOD retained 50% of the maximum activity after autoclaving, and was stable at a wide storage pH ranging from 3 to 10. The enzyme tolerated varying concentrations of denaturating agent, reductants, inhibitors, trypsin, was fairly resistant to inactivation at 80 °C for 180 min (k_d, 6.54 ± 0.17 × 10⁻³ min⁻¹; t_1/2, 106.07 ± 2.68 min), and had midpoint of thermal transition (T_m) of 70.45 °C. The results suggested Ca–Cu,Zn SOD to be a kinetically stable protein that could be used for various industrial applications.     

Copper and zinc in Elodea canadensis from rivers with various pollution levels

The anthropogenic impact of xenobiotics contributes to environmental risk for the aquatic environment and thus, must be controlled. Elodea canadensis, a cosmopolitan aquatic macrophyte with an important role in the ecology of many littoral zones, may provide an integrated record of pollution. Therefore, it was interesting to investigate the accumulation of Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb and Zn in this species and in water and bottom sediments collected from rivers with various levels of contamination. Of these rivers one control and one polluted was selected for the collection of E. canadensis for an experiment to compare the ability of this species to accumulate Cu and Zn. These elements were supplemented at concentrations (mg L⁻¹) of 0.01, 0.02, 0.03, 0.05, 0.08 and 0.14 as CuSO₄·5H₂O, and 0.4, 0.6, 0.9, 1.4, 2.03 and 3.04 as ZnSO₄·7H₂O and in a mixture containing (mg L⁻¹) 0.01Cu + 0.4Zn, 0.02Cu + 0.6Zn, 0.03Cu + 0.9Zn, 0.05Cu + 1.4Zn, 0.08Cu + 2.03Zn and 0.14Cu + 3.04Zn. After the experiment, E. canadensis from the polluted river contained significantly higher Cu and Zn concentrations when applied separately and also significantly higher Cu and Zn concentrations when applied as a mixture compared to the control river. These higher concentrations in E. canadensis from the polluted river were found in all combinations in the experiment. Thus, E. canadensis habituated in polluted sites to the exposure, and long-term influence of elevated metal levels appeared to be better adapted, and it also exhibited a higher increase in biomass than plants from the control river in all the experimental Cu and Zn solutions. Younger leaves of E. canadensis were more resistant to the effects of Cu and Zn than older leaves. Both Cu and Zn negatively affected the cell structure of older leaves, although the influence of Cu on plasma membrane integrity and chloroplast distribution was stronger than that of Zn. The influence of the Cu + Zn mixture on E. canadensis resulted in less pronounced cell disintegration than the influence of Cu added separately.The explanation of differences in the E. canadensis biomass increase and metal concentrations under the binary Cu and Zn impact needs further examination.     

Cypermethrin exposure leads to regulation of proteins expression involved in neoplastic transformation in mouse skin

Cypermethrin, a synthetic pyrethroid insecticide is shown to exert carcinogenic effects in rodents; however, its underlying mechanism remains elusive. Here, we showed the effect of cypermethrin on protein expression involved in neoplastic transformation in mouse skin. Comparative protein expression profiles between untreated control and cypermethrin-treated mouse skin were explored using 2-DE. A total of 27 spots that were statistically significant (p<0.05) and differentially expressed in response to cypermethrin exposure were identified by MALDI-TOF/TOF and LC-MS/MS. Among them, six up-regulated proteins (carbonic anhydrase 3 (Ca 3), Hsp-27, S100A6, galectin-7, S100A9, S100A11) and one down-regulated protein (superoxide dismutase [Cu-Zn] (Sod 1)) are associated with cancer-related key processes. These selected dysregulated proteins were further validated in 2-DE gels of mouse skin treated with known tumorigens (benzo-[a]-pyrene, 12-O-tetradecanoyl-phorbol-13-acetate and mezerein), respectively. Comparative studies showed that Ca 3, S100A6, S100A9, S100A11 and Sod 1 are specific for stages of development and progression of tumors whereas Hsp-27 and galectin-7 are specific for tumor promotion stage by cypermethrin in mouse skin. Furthermore, these chosen proteins confirmed by Western blotting and immunofluorescence staining were consistent with changes in 2-DE check. This proteomic investigation for the first time provides key proteins that will contribute in understanding the mechanism behind cypermethrin-induced neoplastic transformation.     

Subcellular fractionation of Cu exposed oysters,Crassostrea virginica,and Cu accumulation from a biologically incorporated Cu rich oyster diet in Fundulus heteroclitus in fresh and sea water

In order to examine the effect of salinity on Cu accumulation from a naturally incorporated diet, oysters (Crassostrea virginica) were exposed in sea water for 96 days to four waterborne [Cu]: 2.9 ± 0.7 (control), 4.3 ± 0.6, 5.4 ± 0.5, and 10.7 ± 1.0 µg L^? 1. After 96 days, the control whole body [Cu] increased from 2.1 ± 0.5 to 9.1 ± 1.1 µg g^? 1 w.w. and the highest [Cu] was 163.4 ± 27.1 µg g^? 1 w.w. in the oysters. Despite large differences in tissue [Cu], there was no effect on the fraction of trophically available metal in the oyster suggesting that trophic transfer will correlate well with tissue [Cu]. The control and highest [Cu] oysters became diet for killifish (Fundulus heteroclitus) in fresh and seawater for 40 days. The two diets contained 84.7 ± 5.1 and 850.5 ± 8.8 µg Cu g^? 1 d.w. Fish were fed a combined diet of oyster and a pellet supplement (20.5 ± 1.0 µg Cu g^? 1 d.w.) both at 5% body mass day^? 1. In killifish, Cu increased ~ 7% in gills and 100% in intestines after 6 weeks of exposure to the high Cu diet. No other tissues accumulated Cu above control levels. An 11-fold difference free Cu²⁺ concentrations was predicted in intestinal fluid between fresh and sea water, but there was no corresponding effect of salinity on intestinal Cu accumulation suggesting that Cu is not accumulated as the free ion.     

Synthesis and evaluation of copper-64 labeled benzofuran derivatives targeting β-amyloid aggregates

In vivo imaging of β-amyloid (Aβ) aggregates consisting of Aβ(1–40) and Aβ(1–42) peptides by positron emission tomography (PET) contributes to the diagnosis and therapy for Alzheimer’s disease (AD). Because ⁶⁴Cu (t_1/2 = 12.7 h) is a radionuclide for PET with a longer physical half-life than ¹¹C (t_1/2 = 20 min) and ¹⁸F (t_1/2 = 110 min), it is an attractive radionuclide for the development of Aβ imaging probes that are suitable for routine use. In the present study, we designed and synthesized two novel ⁶⁴Cu labeled benzofuran derivatives and evaluated their utility as PET imaging probes for Aβ aggregates. In an in vitro binding assay, 6 and 8 showed binding affinity for Aβ(1–42) aggregates with a K_i value of 33 and 243 nM, respectively. In addition, these probes bound to Aβ plaques deposited in the brain of an AD model mouse in vitro. In a biodistribution experiment using normal mice, these probes showed low brain uptake (0.33% and 0.36% ID/g) at 2 min post-injection. Although refinement to enhance brain uptake is needed, [⁶⁴Cu]6 and [⁶⁴Cu]8 demonstrated the feasibility of developing novel PET probes for imaging Aβ aggregates.     

Inhibitory effects of chlorophyllin, hemin and tetrakis(4-benzoic acid)porphyrin on oxidative DNA damage and mouse skin inflammation induced by 12-O-tetradecanoylphorbol-13-acetate as a possible anti-tumor promoting mechanism

Reactive oxygen species (ROS) from both endogenous and exogenous sources can cause oxidative DNA damage and dysregulated cell signaling, which are involved in the multistage process of carcinogenesis such as tumor initiation, promotion and progression. A number of structurally different anticarcinogenic agents inhibit inflammation and tumor promotion as they reduce ROS production and oxidative DNA damage. Evidence suggests that porphyrins can interfere with the actions of various carcinogens and mutagens by forming face-to-face complexes and their antimutagenic or antigenotoxic effects may also be attributed to their antioxidant activities. However, little is known regarding the anti-tumor promoting potential and mechanism of the porphyrin compounds. Based on our previous results on the inhibitory effects of chlorophyllin (CHL), hemin and tetrakis(4-benzoic acid)porphyrin (TBAP) against two-stage mouse skin carcinogenesis, we have investigated their anti-tumor promoting mechanisms. In the present work, CHL, hemin and TBAP reduced superoxide anion generation by 12-O-tetradecanoylphorbol-13-acetate (TPA) in differentiated HL-60 cells and the production of hydroxyl radicals by Fenton reaction. Porphyrins exert a dose-related inhibition of his(+) reversion in Salmonella typhimurium TA102 induced by tert-butylhydroperoxide (t-BOOH). DNA strand breaks by ROS derived from H(2)O(2)/Cu(II) and the formation of 8-hydroxydeoxyguanosine (8-OH-dG) in calf thymus DNA treated with H(2)O(2)/UV also were inhibited markedly by porphyrins in a concentration-dependent manner. Furthermore, CHL, hemin and TBAP decreased myeloperoxidase (MPO) activity and H(2)O(2) formation as well as epidermal ornithine decarboxylase (ODC) activity in mouse skin treated with TPA. These results demonstrate that the antioxidative properties of porphyrins are important for inhibiting TPA-induced tumor promotion.     

Liposome membrane can act like molecular and metal chaperones for oxidized and fragmented superoxide dismutase

A mechanism for liposome-recruited activity of oxidized and fragmented superoxide dismutase (Fr.-SOD) [Tuan LQ, Umakoshi H, Shimanouchi T, Kuboi R. Liposome-recruited activity of oxidized and fragmented superoxide dismutase. Langmuir 2008;24:350–4] was further investigated, focusing on the secondary structure of Fr.-SOD. Liposome membrane was found to assist the conformational change of Fr.-SOD and reactivate the enzymatic activity, like molecular and metal chaperones. The loss of SOD activity and its secondary structure was observed during 6 h oxidation in 2 mM hydrogen peroxide. The contents of the α-helix and β-sheet structures in the oxidized and fragmented SOD (2 μM) were increased only in the presence of 10 μM Cu²⁺ and Zn²⁺ together, or in the presence of 2 mM POPC liposomes. The mixture of all of these elements (fragmented SOD and POPC liposomes with Cu²⁺ and Zn²⁺) gave not only the increase of the α-helix and β-sheet contents but also the mediation of the high SOD-like activity.     

Salinity-induced physiological and proteomic changes in Anabaena doliolum

This study is the first to offer information on salinity-induced inhibition of physiological variables, changes in proteome, and induction of glycolate metabolism in Anabaena doliolum. A significant reduction in O₂-evolution, carbon fixation, chlorophyll and NADPH/NADH level and increase in intracellular Na⁺ and respiration were observed following 150 mM NaCl treatment for 1 and 24 h. Interestingly, ATP content registered significant decrease after 1 h and recovery after 24 h treatment of 150 mM NaCl. Two-dimensional gel electrophoresis and MALDI-TOF MS detected a set of six proteins showing significant reproducible alterations, and homology with iron superoxide dismutase, superoxide dismutase (imported), phycocyanin alpha chain, elongation factor-Tu (EF-Tu), ribulose 1,5-bisphosphate carboxylase/oxygenase and phosphoribulokinase of Nostoc PCC7120. Increased RuBisCO and decreased carbon fixation suggested operation of glycolate metabolism. This was confirmed by accumulation of free and phospho-glyceric acid, increase in glycolate oxidase activity, glycine, serine and ammonium contents. Since peroxide generated in this pathway cannot be scavenged due to sensitivity of catalase to NaCl the organism fails to acclimatize under salt stress.     

Lack of contribution of covalent benzo[a]pyrene-7,8-quinone–DNA adducts in benzo[a]pyrene-induced mouse lung tumorigenesis

Benzo[a]pyrene (B[a]P) is a potent human and rodent lung carcinogen. This activity has been ascribed in part to the formation of anti-trans-7,8-dihydroxy-7,8-dihydroB[a]P-9,10-epoxide (BPDE)-DNA adducts. Other carcinogenic mechanisms have been proposed: (1) the induction of apurinic sites from radical cation processes, and (2) the metabolic formation of B[a]P-7,8-quinone (BPQ) that can form covalent DNA adducts or reactive oxygen species which can damage DNA. The studies presented here sought to examine the role of stable BPQ-DNA adducts in B[a]P-induced mouse lung tumorigenesis. Male strain A/J mice were injected intraperitoneally once with BPQ or trans-7,8-dihydroxy-7,8-dihydroB[a]P (BP-7,8-diol) at 30, 10, 3, or 0 mg/kg. Lungs and livers were harvested after 24 h, the DNA extracted and subjected to ³²P-postlabeling analysis. Additional groups of mice were dosed once with BPQ or BP-7,8-diol each at 30 mg/kg and tissues harvested 48 and 72 h later, or with B[a]P (50 mg/kg, a tumorigenic dose) and tissues harvested 72 h later. No BPQ or any other DNA adducts were observed in lung or liver tissues 24, 48, or 72 h after the treatment with 30 mg/kg BPQ. BP-7,8-diol gave BPDE-DNA adducts at all time points in both tissues and B[a]P treatment gave BPDE-DNA adducts in the lung. In each case, no BPQ-DNA adducts were detected. Mouse body weights significantly decreased over time after BPQ or BP-7,8-diol treatments suggesting that systemic toxicity was induced by both agents. Model studies with BPQ and N-acetylcysteine suggested that BPQ is rapidly inactivated by sulfhydryl-containing compounds and not available for DNA adduction. We conclude that under these treatment conditions BPQ does not form stable covalent DNA adducts in the lungs or livers of strain A/J mice, suggesting that stable BPQ-covalent adducts are not a part of the complex of mechanisms involved in B[a]P-induced mouse lung tumorigenesis.