bluuebunny/crossref_metadata_2025 huggingface dataset:
The isoflavonoid aglycone‐rich fractions of Chungkookjang, fermented unsalted soybeans, enhance insulin signaling and peroxisome proliferator‐activated receptor‐γ activity in vitro
Authors: Kwon Dae Young , Jang Jin Sun , Lee Ji Eun , Kim Yong‐Suk , Shin Dong‐Hwa , Park Sunmin
Published: December 2008

We investigated anti‐diabetic candidates and their mechanisms from the fractions of Chungkookjang (CKJ), a traditional fermented unsalted soybean, by investigating insulin signaling, peroxisome proliferator‐activated receptor (PPAR)‐γ activity and glucose‐stimulated insulin secretion, in vitro. Cooked soybeans (CSB) and CKJ, fermented predominantly with Bacillus subtilis, were extracted by 70% EtOH followed by an XAD‐4 column chromatography with a serial mixture of solvents comprised of MeOH and water. During fermentation, the contents of isoflavonoid aglycones were elevated, and the fractions enriched with aglycones enhanced insulin‐stimulated glucose uptake in 3T3‐L1 adipocytes. This increase in glucose uptake resulted from stimulating a translocation of the glucose transporter (GLUT)‐4 into the plasma membrane through the phosphorylation of insulin receptor substrate (IRS)‐1 and Akt. Especially, daidzein enriched fractions elevated insulin‐stimulated glucose uptake by acting as PPAR‐γ agonist up to levels exhibited when 10 nM insulin is administered. Fractions containing small peptides with low polarity in CKJ slightly increased glucose‐stimulated insulin secretion. The data suggest that an increase in isoflavonoid aglycones in CKJ, in comparison to CSB, enhances glucose utilization via activating insulin signaling and stimulates PPAR‐γ activity in adipocytes. In addition, CKJ contains small peptides improving glucose‐stimulated insulin secretion in insulinoma cells. Overall, CKJ is superior to CSB in anti‐diabetic action.
Regulation of Mg2+‐independent Ca2+‐ATPase by a low molecular mass protein purified from bovine brain
Authors: Ghoshal Srabasti , Sengupta Tanusree , Sen Parimal C.
Published: December 2008

The goat sperm microsomal membranes have been found to contain an Mg2+‐independent Ca2+‐ATPase, a low affinity but highly active enzyme sharing similarities with the SERCA family of ATPases. The present study reports the identification and characterization of a 14 kilodalton cytosolic protein from bovine brain which can act as an endogenous stimulator of the enzyme with an S50 (concentration producing 50% stimulation) of 0.8 μ molar. Kinetic analysis suggests that the stimulation is noncompetitive with respect to the substrate, and the binding site(s) of the stimulator and substrate are distinct. Binding of the stimulator to the enzyme is reversible. The stimulator increases the affinity of the enzyme for calcium as evident from a decrease in K0.5 of the enzyme for calcium in presence of the stimulator. Radioactive labeling of the enzyme with [γ;‐32P]‐ATP suggests that the stimulator enhances the rate of dephosphorylation of the phosphoenzyme intermediate without altering the phosphorylation reaction step. The stimulatory effect of the protein has been observed only for the Mg2+‐independent form of the enzyme, the Mg2+‐dependent form being unaffected.
Activation of redox‐systems of monocytes by hydrogen peroxide
Authors: Krjukov Alexei A. , Semenkova Galina N. , Cherenkevich Sergei N. , Gerein Valentin
Published: December 2008

In this work the influence of H2O2 on the ability of human blood monocytes to generate ROS upon stimulation of cells by adhesion to glass surface and fMLP was studied using the luminol‐dependent chemiluminescence (LDCL) method. Pretreatment of cells with H2O2 increased the adhesiveness of monocytes and ROS generation. Superoxide generation by cells in response to fMLP depended on the duration of pretreatment and the concentration of H2O2. The stimulatory effect on fMLP‐induced LDCL of cells further depended on the Ca2+ concentration in the medium and on the activities of phospholipase A2, cyclooxygenases, and Mek1/2.
High Resolution 1H NMR investigations of the oxidative consumption of salivary biomolecules by ozone: Relevance to the therapeutic applications of this agent in clinical dentistry
Authors: Grootveld Martin , Silwood Christopher J.L. , Lynch Edward
Published: December 2008

High resolution proton (1H ) nuclear magnetic resonance (NMR) spectroscopy was employed to simultaneously evaluate the oxidising actions of ozone (O3) towards a wide range of salivary biomolecules in view of its applications in dental practices, which may serve as a viable and convenient means for the treatment of dental caries. Treatment of supernatants derived from unstimulated human saliva specimens (n=12) with O3 (4.48 mmol) revealed that this reactive oxygen species gave rise to the oxidative consumption of pyruvate (generating acetate and CO2 as products), lactate (to pyruvate and sequentially acetate and CO2), carbohydrates in general (a process generating formate), methionine (giving rise to its corresponding sulphoxide), and urate (to allantoin). Further, minor O3‐induced modifications included the oxidation of trimethylamine and 3‐D‐hydroxybutyrate, the fragmentation of salivary glycosaminoglycans to NMR‐detectable saccharide fragments, and the conversion of polyunsaturated fatty acids to their ozonides. Moreover, evidence for the ability of O3 to induce the release of selected low‐molecular‐mass salivary biomolecules from macromolecular binding‐sites was also obtained. Since many of the oxidation products detectable in O3‐treated samples are identical to those arising from the attack of •OH radical on biofluid components, it appears that at least some of the modifications observed here are attributable to the latter oxidant (derived from O generated from the single electron reduction of O3).
Anti‐mitotic properties of resveratrol analog (Z)‐3,5,4′‐trimethoxystilbene
Authors: Chabert Philippe , Fougerousse André , Brouillard Raymond
Published: December 2008

(Z)‐3,5,4′‐Trimethoxystilbene is a natural polyphenol present in five different plants, Virola cuspidata, Virola elongata, Centipeda minima, Schoenus nigricans and Rheum undulatum. This molecule was prepared in a three‐step sequence in good overall yield. The isomerisation from the (E)‐ to (Z)‐isomer is performed using UV irradiation. Biological investigations were conducted on a human colon cancer cell line (Caco‐2) with anti‐mitotic activities. Growth was completely arrested at an added 0.4 μM level of (Z)‐3,5,4′‐trimethoxystilbene. This agent is 100‐fold more active than resveratrol or (E)‐3,5,4′‐trihydroxystilbene, and the mechanism of this process involves an inhibition of tubulin polymerisation in a dose dependent manner.
Thioredoxin‐binding protein‐2 (TBP‐2): Its potential roles in the aging process
Authors: Yoshida Toru , Kondo Norihiko , Oka Shin‐Ichi , Ahsan Md Kaimul , Hara Tomijiro , Masutani Hiroshi , Nakamura Hajime , Ab Junji Yodoi
Published: December 2008

Thioredoxin (TRX) binding protein‐2 (TBP‐2), a negative regulator of TRX, is involved in intracellular redox regulation and cellular growth. The expression of TBP‐2 is frequently lost in tumor cell lines and tissues, whereas the ectopic expression of TBP‐2 suppresses cellular proliferation along with cell cycle arrest at the G1 phase. TBP‐2 was also reported to be a cellular senescence‐associated gene. Besides the retardation of cellular growth, the reduction of white adipose, and alteration of the energy pathway are involved in several features of the aging process. We have generated TBP‐2 genetically modified mice and found that TBP‐2 is closely linked to lipid metabolism. Indeed, TBP‐2 has been suggesting to be related to familial combined hyperlipidemia analyzed by a spontaneous mutant mouse strain. As lipid metabolism is one of the most primitive sources of energy production, we discussed the possible roles of TBP‐2 in the regulation of energy utilization connected to the aging process.
Enhanced cardiovascular function and energy level by a novel chromium (III)‐supplement
Authors: Thirunavukkarasu Mahesh , Penumathsa Sureshvarma , Juhasz Bela , Zhan Lijun , Bagchi Manashi , Yasmin Taharat , Shara Michael A. , Thatte Hemant S. , Maulik Nilanjana
Published: December 2008

The impetus for the novel Energy Formula (EF) which combines the niacin‐bound chromium (III) (0.45%) (NBC), standardized extract ofWithania somniferaextracts (10.71%), caffeine (22.76%), D‐ribose (10.71%) and selected amino acids such as phenylalanine, taurine and glutamine (55.37%) was based on the knowledge of the cardioprotective potentials of theWithania somniferaextract, caffeine and D‐ribose as well as their abilities to increase energy levels and the abilities of amino acids to increase the muscle mass and energy levels. The effect of oral supplementation of EF on the safety, myocardial energy levels and cardioprotective ability were investigated in an ischemic‐reperfused myocardium model in both male and female Sprague‐Dawley rats over 90 days trial period. At the completion of 90 days, the EF‐treated male and female rats gained 9.4% and 3.1% less body weights, respectively, as compared to their corresponding control groups. No significant difference was found in the levels of lipid peroxidation and activities of hepatic Aspartate transaminase, Alanine transaminase and Alkaline phosphatase in EF treatment when compared with control animals. The male and female rat hearts were subjected to 30 min of global ischemia followed by 2 h of reperfusion at 30 and 90 days of EF treatment. Cardiovascular functions including heart rate, coronary flow, aortic flow, dp/dtmax, left ventricular developed pressure (LVDP) and infarct size were monitored. The levels of myocardial adenosine triphosphate (ATP), creatine phosphate (CP), phospho‐adenosine monophosphate kinase (p‐AMPK) levels, were analyzed at the end of 30 and 90 days of treatment. Significant improvement was observed in all parameters in the EF treatment groups as compared to their corresponding controls. Thus the niacin‐bound chromium (III) based energy formula is safe and effective supplement to boost energy levels and cardioprotection.
Croton lobatus, an African medicinal plant: Spectroscopic and chemical elucidation of its many constituents
Authors: Chabert Philippe , Attioua BarthÉLemy , Brouillard Raymond
Published: December 2008

The chemical constituents of the African medicinal plant Croton lobatus were elucidated and characterised using 1D and 2D‐NMR analysis and the application of the technique of High Resolution Electron Ionization Mass Spectrometry (HREIMS) and High Resolution Mass Spectrometry (HRMS). The novel triglyceride lobaceride or 3‐((6Z,9Z)dodeca‐6,9‐dienoyloxy)‐2‐octanoyloxypropyl (6Z,9Z)dodeca‐6,9‐dienoate, along with ten compounds were isolated from the stems and leaves of Croton lobatus.
Functional studies of an HIV‐1 encoded glutathione peroxidase
Authors: Zhao Lijun , Olubajo Babatunde , Taylor Ethan Will
Published: December 2008

In an alternate reading frame overlapping the viral envelope gene, HIV‐1 has been shown to encoded a truncated glutathione peroxidase (GPx) module. Essential active site residues of the catalytic core regions of mammalian GPx sequences are conserved in the putative viral GPx (vGPx, encoded by the env‐fs gene). Cells transfected with an HIV‐1 env‐fs construct show up to a 100% increase in GPx enzyme activity, and are protected against the loss of mitochondrial transmembrane potential and subsequent cell death induced by exogenous oxidants or mitochondrial reactive oxygen species. An intact vGPx gene was observed to be more common in HIV‐1‐infected long‐term non‐progressors, as compared to HIV‐1 isolates from patients developing AIDS. An antioxidant/antiapoptotic protective role of the vGPx is also consistent with the observation that −1 frameshifting induced by the HIV‐1 env‐fs sequence AAAAAGA (which contains a potential “hungry” arginine codon, AGA) increases during arginine deficiency, which has been associated with increased oxidative stress. Under arginine‐limited conditions, nitric oxide synthase generates superoxide, which rapidly combines with NO to form peroxynitrite, which can cause activated T‐cells to undergo apoptosis. Thus, biosynthesis of the HIV‐1 GPx as an adaptive response to low arginine conditions might delay oxidant‐induced apoptotic cell death, providing an enhanced opportunity for viral replication.
Determination of the illicit drug gamma‐hydroxybutyrate (GHB) in human saliva and beverages by 1H NMR analysis
Authors: Grootveld Martin , Algeo Deborah , Silwood Christopher J.L. , Blackburn John C. , Clark Anthony D.
Published: December 2008

High resolution 1H NMR spectroscopy has been employed to investigate the detection and quantification of the illicit ‘date‐rape’ drug gamma‐hydroxybutyrate (GHB) in both human saliva and a commonly‐consumed low‐alcohol beer product. Data acquired revealed that this multicomponent analytical technique provided unequivocal evidence for the detection of this agent by this technique in both of these matrices, i.e., all three of its resonances [those ascribable to the α‐CH2 (t, δ=2.25 ppm), β‐CH2 (tt, δ=1.81 ppm) and γ‐CH2 (t, δ=3.61 ppm) group protons] were present in spectra acquired on human saliva, and two of these (the α‐ and β‐CH2 group signals) in the beverage product examined, the latter observation attributable to overlap of the γ‐CH2 1H resonance with those of carbohydrates. Since good linear calibration relationships between the intensities of each of the NMR‐visible signals and added GHB concentration (the former normalised to that of an external 3‐trimethylsilyl [2,2,3,3‐2H4]‐ propionate standard present in a coaxial NMR tube insert) were observed, this illicit drug is also readily quantifiable in such multicomponent samples. Our data demonstrate the advantages offered by this technique when applied to the analysis of illicit drugs in multicomponent sample matrices such as human biofluids and beverage products.
Effects of quercetin and β‐carotene supplementation on azoxymethane‐induced colon carcinogenesis and inflammatory responses in rats fed with high‐fat diet rich in ω‐6 fatty acids
Authors: Choi Soo‐Yeon , Park Jung Han Yoon , Kim Jong‐Sang , Kim Mi Kyung , Aruoma Okezie I. , Sung Mi‐Kyung
Published: December 2008

Chronic inflammation in gastrointestinal tract has been suggested as a risk factor for tumor formation. The effect of dietary supplementation of quercetin or ß‐carotene on colon carcinogenesis and inflammatory response in rats fed with high‐fat diet rich in ω‐6 fatty acids was assessed. Animals were exposed to two weekly subcutaneous injections of AOM (azoxymethane) at a single dose of 15 mg/kg body weight. A portion of rats from each group was sacrificed at 8 weeks after the last AOM treatment to determine ACF (aberrant crypt foci) formation. Colonic mucosa expression of iNOS (inducible nitric oxide) and COX‐2 (cyclooxygenase‐2) protein, and blood PGE2 (prostaglandin E2) level were measured. The remaining groups of animals were sacrificed at 33 weeks after the last AOM treatment to examine colon tumor formation. Rats on high‐fat diet developed more aberrant crypt foci (P<0.05) compared with those of rats on regular diet. In the same vein, but in contrast to the effect seen with regular diet, the high‐fat diet induced a significant up‐regulation of iNOS expression. There was no significant change in the extent of COX‐2 expression or in the PGE2 levels. Quercetin or β‐carotene supplementation reduced the number of ACF only in animals fed high‐fat diet (p<0.05), however, no significant difference in tumor incidence was found. At week 33, the expression of iNOS was reduced by quercetin without a statistical significance, and COX‐2 expression was slightly reduced in rats on ß‐carotene supplementation. No change in PGE2 levels was observed. Whilst dietary antioxidants are considered as effective suppressors for precancerous lesion formation in colons exposed to high‐risk diet, it is clear that elucidating the role of individual antioxidants in colon tumor formation coupled with an understanding of the molecular mechanisms involved would benefit colon cancer prevention strategies.
Reversal of the TPA‐induced inhibition of gap junctional intercellular communication by Chaga mushroom (Inonotus obliquus) extracts: Effects on MAP kinases
Authors: Park Jung‐Ran , Park Joon‐Suk , Jo Eun‐Hye , Hwang Jae‐Woong , Kim Sun‐Jung , Ra Jeong‐Chan , Aruoma Okezie I. , Lee Yong‐Soon , Kang Kyung‐Sun
Published: December 2008

Chaga mushroom (Inonotus obliquus) has continued to receive attention as a folk medicine with indications for the treatment of cancers and digestive diseases. The anticarcinogenic effect of Chaga mushroom extract was investigated using a model system of gap junctional intercellular communication (GJIC) in WB‐F344 normal rat liver epithelial cells. The cells were pre‐incubated with Chaga mushroom extracts (5, 10, 20 μg/ml) for 24 h and this was followed by co‐treatment with Chaga mushroom extracts and TPA (12‐O‐tetradecanoylphorbol‐13‐acetate, 10 ng/ml) for 1 h. The inhibition of GJIC by TPA (12‐O‐tetradecanoylphorbol‐13‐acetate), promoter of cancer, was prevented with treatment of Chaga mushroom extracts. Similarly, the increased phosphorylated ERK1/2 and p38 protein kinases were markedly reduced in Chaga mushroom extracts‐treated cells. There was no change in the JNK kinase protein level, suggesting that Chaga mushroom extracts could only block the activation of ERK1/2 and p38 MAP kinase. The Chaga mushroom extracts further prevented the inhibition of GJIC through the blocking of Cx43 phosphorylation. Indeed cell‐to‐cell communication through gap junctional channels is a critical factor in the life and death balance of cells because GJIC has an important function in maintaining tissue homeostasis through the regulation of cell growth, differentiation, apoptosis and adaptive functions of differentiated cells. Thus Chaga mushroom may act as a natural anticancer product by preventing the inhibition of GJIC through the inactivation of ERK1/2 and p38 MAP kinase.
Activity of the dietary antioxidant ergothioneine in a virus gene‐based assay for inhibitors of HIV transcription
Authors: Xiao Lianchun , Zhao Lijun , Li Ting , Hartle Diane K. , Aruoma Okezie I. , Taylor Ethan Will
Published: December 2008

The “Long Terminal Repeat” (LTR) of HIV‐1 is the target of cellular transcription factors such as NF‐κB, and serves as the promoter‐enhancer for the viral genome when integrated in host DNA. Various LTR‐reporter gene constructs have been used for in vitro studies of activators or inhibitors of HIV‐1 transcription, e.g., to show that antioxidants such as lipoic acid and selenium inhibit NF‐κB‐dependent HIV‐1 LTR activation. One such construct is the pHIVlacZ plasmid, with the HIV‐1 LTR driving expression of the lacZ gene (encoding β‐galactosidase, β‐gal). Typically, for inhibitor screening, cells transfected with pHIVlacZ are activated using tumor necrosis factor‐α (TNF‐α), and the colorimetric o‐nitrophenol assay is used to assess changes in β‐gal activity. A variant of this assay was developed as described here, in which LTR activation was induced by pro‐fs, a novel HIV‐1 gene product encoded via a −1 frameshift from the protease gene. Cotransfection of cells with pHIVlacZ along with a pro‐fs construct produced a signifcant increase in β‐gal activity over controls. L‐ergothioneine dose dependently inhibited both TNF‐α‐mediated and pro‐fs‐mediated increases in β‐gal activity, with an IC50 of about 6 mM. Thus antioxidant strategy involving ergothioneine derived from food plants might be of benefit in chronic immunodeficiency diseases.
The effects of reactive oxygen and nitrogen species during yeast replicative ageing
Authors: Wilhelm Jiří , Fuksová Hana , Schwippelová Zuzana , Vytášek Richard , Pichová Alena
Published: December 2008

Free radicals are considered the most important cause of cellular ageing. We have investigated ageing process in the yeast Saccharomyces cerevisiae. We have compared the wild type strain with the mutant cells with constitutively active Ras oncogen, which generates increased amounts of free radicals. Increased generation of oxygen‐derived free radicals resulted in the Ras mutant cells accumulation of lipofuscin‐like pigments during ageing. Ageing wild type cells did not accumulate lipofuscin‐like pigments. This is quite unique feature among known biological models. It may be caused by increased concentration of alpha tocopherol (the most prominent lipophilic antioxidant) in the wild type cells. In contrast, the Ras mutant cells contained decreased levels of alpha tocopherol even in the young cells. This observation indicates that the increased free radical generation can overwhelm the endogenous antioxidant system. We have documented the involvement of nitrogen‐derived free radicals in the yeast metabolism. Protein nitrotyrosine, a marker of the reactive nitrogen species, has significantly increased in the senescent Ras mutant cells. The wild type cells contained basic level of nitrotyrosine corresponding to its concentration found in non‐activated mammalian macrophages.
Bio‐markers of lipid peroxidation in vivo: Hydroxyoctadecadienoic acid and hydroxycholesterol
Authors: Yoshida Yasukazu , Niki Etsuo
Published: December 2008

The biological role of lipid peroxidation products has continued to receive a great deal of attention not only for the elucidation of pathological mechanisms but also for their practical application to clinical use as bio‐markers. In the last fifty years, lipid peroxidation has been the subject of extensive studies from the viewpoints of mechanisms, dynamics, product analysis, involvement in diseases, inhibition, and biological signaling. Lipid hydroperoxides are formed as the major primary products, however they are substrates for various enzymes and they also undergo various secondary reactions. In this decade, F2‐isoprostanes from arachidonates and neuroprostanes from docosahexanoates have been proposed as bio‐markers. Although these markers are formed by a free radical‐mediated oxidation, the yields from the parent lipids are minimal. Compared to these markers, hydroperoxy octadecadienoates (HPODE) from linoleates and oxysterols from cholesterols are yielded by much simpler mechanisms from more abundant parent lipids in vivo. Recently, the method in which both free and ester forms of hydroperoxides and ketones as well as hydroxides of linoleic acid and cholesterol are measured as total hydroxyoctadecadienoic acid (tHODE) and 7‐hydroxycholesterol (t7‐OHCh), respectively, was proposed. The concentrations of tHODE and t7‐OHCh determined by GC‐MS analysis from physiological samples were much higher than that of 8‐iso‐prostagrandin F_{2α}. In addition to this advantage, hydrogen‐donor activity of antioxidants in vivo could be determined by the isomeric‐ratio of HODE (9‐ and 13‐(Z,E)‐HODE/9‐ and 13‐(E,E)‐HODE).
Mechanisms of adenosine‐induced cytotoxicity and their clinical and physiological implications
Authors: Seetulsingh‐Goorah Sharmila P.
Published: December 2008

Extracellular ATP (ATPo) and adenosine are cytotoxic to several cancer cell lines, suggesting their potential use for anticancer therapy. Adenosine causes cytotoxicity, either when added exogenously or when generated from ATPo hydrolysis, via mechanisms which are not mutually exclusive and which involve, adenosine receptor activation, pyrimidine starvation and/or increases in intracellular S‐adenosylhomocysteine: S‐adenosylmethionine ratio. Given that adenosine also appears to protect against cytotoxicity via mechanisms including immunity against damage by oxygen free radicals, an understanding of the contribution of adenosine to ATPo‐induced cytotoxicity is thus crucial, when considering any potential therapeutic use for these compounds. However, such an understanding has been largely hindered by the fact that many studies have not focused enough on the possibility that both ATPo and adenosine may mediate cytotoxicity in the same system. Such studies can benefit from use a range of ATPo concentrations when assessing the contribution of adenosine to ATPo‐induced cytotoxicity. Whilst future molecular and pharmacological studies are needed to establish the nature of the cytotoxic adenosine receptor, it is possible that more than just one adenosine receptor type is involved and that the cytotoxic receptor(s) type is more likely to have a low affinity for adenosine. Activation of the adenosine receptor(s) would thus lead to cytotoxicity only at relatively high adenosine concentrations, while lower adenosine concentrations mediate non‐cytotoxic physiological effects.
Bioavailability, antioxidant and immune‐enhancing properties of zinc methionine
Authors: Chien Xiaoming Xu , Zafra‐Stone Shirley , Bagchi Manashi , Bagchi Debasis
Published: December 2008

Although a large number of transition metals and cations remarkably induce oxidative deterioration of biological macromolecules including lipids, proteins and DNA, the trace element zinc acts as a novel dietary supplement and an essential micronutrient, and serves a wide range of biological functions in human and animal health. Zinc promotes antioxidant and immune functions, stabilizes and maintains the structural integrity of biological membranes, and plays a pivotal role in skin and connective tissue metabolism and repair. Zinc is an integral constituent of a large number of enzymes including antioxidant enzymes, and hormones including glucagon, insulin, growth hormone, and sex hormones. High concentrations of zinc are found in the prostate gland and choroids of the eye. Zinc deficiency leads to biochemical abnormalities including the impairments of growth, dermal, gastrointestinal, neurologic and immunologic systems. Given its superior bioavailability, antioxidant and immune‐enhancing properties, zinc methionine may serve as a novel dietary supplement to promote health benefits in humans and animals.
Role of oxygen‐derived free radicals in Helicobacter pylori water extract‐induced mouse skin carcinogenesis
Authors: Ishikawa Takeshi , Yoshida Norimasa , Tokuda Harukuni , Kokura Satoshi , Nakabe Nami , Kuchide Masashi , Ichiishi Eiichiro , Imaeda Atsumune , Ichikawa Hiroshi , Naito Yuji , Okanoue Takeshi , Yoshikawa Toshikazu
Published: December 2008

Helicobacter pylori (H. pylori) infection, the main cause of chronic gastritis, increases gastric cancer risk. The infection causes inflammatory cells to produce reactive oxygen metabolites that may damage DNA and promote carcinogenesis. However, its precise role in gastric carcinogenesis is as yet unknown. Recently we reported that H. pylori water extract (HPE) has an initiating activity on two‐stage mouse skin carcinogenesis. In this study, we investigated the effects of anti‐oxidants, ascorbic acid and a combination of superoxide dismutase (CuZnSOD)and catalase, on two‐stage mouse skin carcinogenesis. Ascorbic acid and CuZnSOD/catalase were given to mice during the period of HPE‐initiation. Both the ascorbic acid and CuZnSOD/catalase treatment attenuated the incidence of tumor formation. The present results suggest that HPE induces tumor formation via reactive oxygen species (ROS) production.
Influence of vitamin E on gastric mucosal injury induced by Helicobacter pylori infection
Authors: Sugimoto Naohito , Yoshida Norimasa , Nakamura Yasunari , Ichikawa Hiroshi , Naito Yuji , Okanoue Takeshi , Yoshikawa Toshikazu
Published: December 2008

We investigated the effect of vitamin E on gastric mucosal injury induced by Helicobacter pylori (H. pylori) infection. Male Mongolian gerbils were divided into 4 groups (normal group without H. pylori infection, vitamin E‐deficient, ‐sufficient and ‐supplemented groups with H. pylori infection). Following oral inoculation with H. pylori (ATCC43504 2 × 108 CFU), animals were fed diets α‐tocopherol 2 mg/100 g diet in the normal and vitamin E‐sufficient groups and α‐tocopherol 0.1 mg/100 g and 50 mg/100 g in the vitamin E‐deficient and ‐supplemented groups, respectively, for 24 weeks. Chronic gastritis was detected in all gerbils inoculated H. pylori. Gastric ulcer was detected in 2 of 7 gerbils only in the vitamin E‐deficient group. In the vitamin E‐deficient group, myeloperoxidase activity and mouse keratinocyte derived chemokine (KC) in gastric mucosa was significantly higher than in the vitamin E supplemented group. Subsequently, in an in vitro study expression of CD11b/CD18 on neutrophils was enhanced by H. pylori water extract. This effect was suppressed in a dose dependent manner by the addition of α‐tocopherol. These results suggest that vitamin E has a protective effect on gastric mucosal injury induced by H. pylori infection in gerbils, through the inhibition of accumulation of activated neutrophils.
Manganese depresses rat heart muscle respiration
Authors: Miller Kevin B. , Caton Joel S. , Finley John W.
Published: December 2008

It has previously been reported that moderately high dietary manganese (Mn) in combination with marginal magnesium (Mg) resulted in ultrastructural damage to heart mitochondria. Manganese may replace Mg in biological functions, including the role of enzyme cofactor. Manganese may accumulate and substitute for Mg during the condition of Mg‐deficiency. The objective of the current study was to determine whether high Mn alters heart muscle respiration and Mg‐enzyme activity as well as whole body Mn retention under marginal Mg. An additional objective was to determine whether high Mn results in increased oxidative stress. In experiment 1: forty‐eight rats were fed a 2 × 3 factorial arrangement of Mn (10, 100, or 1000 mg/kg) and Mg (200 or 500 mg/kg). In experiment 2: thirty‐two rats were fed one of four diets in a 2 × 2 factorial arrangement of Mn (10 or 250 mg/kg) and Mg (200 or 500 mg/kg). In experiment 3: thirty‐two rats were fed one of four diets in a 2 × 2 factorial arrangement of Mn (10 or 650 mg/kg) and Mg (200 or 500 mg/kg). In experiment 2, high Mn and marginal Mg reduced (P<0.05) oxygen consumption of left ventricle muscle. Marginal Mg, but not Mn, reduced (P<0.05) activity of sarcoplasmic reticulum calcium‐ATPase enzyme. Dietary Mg had no affect on54Mn kinetics, but high dietary Mn decreased (P<0.01) absorption, retention, and rate of excretion of54Mn. Neither cellular stress, measured by Comet assay, nor antioxidant activities were increased by high Mn. A strong interaction (P<0.001) between increasing Mn and adequate Mg on hematology was observed. These results confirm previous research in swine that high Mn alters myocardial integrity as well as function, but not as a result of altered calcium transport or oxidative stress.
Oligomeric hepatoproliferin disaggregates into an active monomer that was purified and forcefully dissociated into two different ionic species
Authors: Oosthuizen Mathys M.J. , Lambrechts Hugo
Published: December 2008

Hepatoproliferin (HPF), a liver regeneration factor, was isolated initially as an aggregated molecule (big‐HPF) and was purified into two homogeneous, bioactive species of 14 kDa and 18.5 kDa. These two big‐HPFs were disaggregated to completion into two monomeric forms (small‐HPFs) when incubated for 10 days in 0.15 M ammonium bicarbonate at 25°C. Both monomeric forms were purified to homogeneity as active entities, one with a molecular mass of 944 Da and one with a molecular mass of 1066 Da. Each of the two 35S‐labelled small‐HPFs was found, by enzymic analysis, to contain a charged sulfonated saccharide, which was neutralized by a specific amine. Monomeric HPF is therefore a stable ionic complex formed between these two ionic species. So strong was the electrostatic association that small‐HPF remained intact in solution and no amine was displaced by the ammonium ions of the buffer. Small‐HPF remained unimpaired during purification, since all activity was retained despite alternating acidic and basic conditions. However, when small‐HPF was brought into contact with either a cationic or an anionic resin, it was dissociated to completion when mixed continuously with the resin for 4 days. The ionic entity that was released had no bio‐activity and was either a pure radioactively labeled saccharide or a non‐labeled amine, depending on the kind of resin used. When incubated together, the separated counterions combine to regain full activity after 2 days of reassociation. However, with incubation for longer, this reassociated small‐HPF formed different oligomeric HPFs by aggregation.

Small‐HPF is therefore a new kind of growth enhancer, consisting of an acidic sulfonated saccharide and a basic amine assembled into a stable active ionic complex that has a tendency to aggregate.
Plasma membrane‐generated reactive oxygen intermediates and their role in cell growth of plants
Authors: Schopfer P. , Liszkay A.
Published: December 2008

Reactive oxygen species (ROS) produced as intermediates in the reduction of O2 to H2O (superoxide radical, hydrogen peroxide, hydroxyl radical), are generally regarded as harmful products of oxygenic metabolism causing cell damage in plants, animals and microorganisms. However, oxygen radical chemistry can also play useful roles if it takes place outside of the protoplast. In plants, the production of these ROS initiated by the plasma membrane NAD(P)H oxidase can be used for controlled polymer breakdown leading to wall loosening during extension growth. Backbone cleavage of cell wall polysaccharides can be accomplished by hydroxyl radicals produced from hydrogen peroxide and superoxide in a reaction catalyzed by cell wall peroxidase. Growing plant organs such as coleoptiles or roots of maize seedlings produce these ROS specifically in the apoplast of actively growing tissues, e.g. in the epidermis of the coleoptile and the growing zone of the root. Auxin promotes the release of hydroxyl radicals when inducing elongation growth. Experimental generation of hydroxyl radicals in the wall causes an increase in wall extensibility in vitro and replaces auxin in inducing growth. Auxin‐induced growth can be inhibited by scavengers of ROS or inhibitors interfering with the formation of these molecules in the cell wall. These results provide the experimental background for a novel hypothesis on the mechanism of plant cell growth in which the generation of hydroxyl radicals, initiated by the plasma membrane NAD(P)H oxidase, plays a central role.
ATP‐ and growth substance‐dependent cell‐free enlargement of plasma membrane vesicles from soybean
Authors: Auderseta Guy , Morré D.J.
Published: December 2008

Growth hormone‐responsive and nucleoside triphosphate‐dependent enlargement of inside‐out vesicles of plasma membranes from soybeans prepared by aqueous two‐phase partition and everted by freezing and thawing has been achieved in a cell‐free system. In the presence of 100 μM ATP in 40 mM HEPES buffer, pH 7, enlargement of isolated plasma membrane vesicles was accelerated by the synthetic plant growth factor, 2,4‐dichlorophenoxyacetic acid (2,4‐D), compared to ATP alone, 2,4‐D alone or no additions. After 20 min with 1 μM 2,4‐D, vesicles increased in diameter, 20% on average. Although vesicle diameters in the presence or absence of 2,4‐D overlapped, the means were clearly separated. The 20% increases in diameter corresponded to a doubling of vesicle volume. Both 100 μM ATP and 1 μM 2,4‐D were necessary to stimulate the cell‐free vesicle enlargement. In the presence of 1 μM 2,4‐D, enlargement observed with 100 μM ATP was greater than with either 10 μM ATP or 500 μM ATP alone. In the presence of 100 μM ATP, vesicle enlargement was proportional to the logarithm of 2,4‐D concentration. With the growth‐inactive 2,4‐D analog, 2,3‐D, no vesicle enlargement was observed either alone or in the presence of 100 μM ATP. Right side‐out vesicles did not enlarge in response to either ATP, 2,4‐D or the two in combination suggesting that the responsible ATP site was on the inside of the cell.
Inside‐out but not right side‐out plasma membrane vesicles from soybean enlarge when treated with ATP + 2,4‐D as determined by electron microscopy and light scattering: Evidence for involvement of a plasma membrane AAA‐ATPase
Authors: Hicks‐Bergera Carrie , Morreb D. James
Published: December 2008

The concept that the location of an AAA‐ATPase associated with the plant plasma membrane may be indicative of a functional relationship to growth or cell enlargement by analogy with roles in physical membrane displacements as proposed for AAA‐ATPases associated with internal membranes was tested. A plant growth hormone‐responsive and nucleoside triphosphate‐dependent enlargement of inside‐out vesicles of plasma membranes from soybeans was utilized in a completely cell‐free system. The rate of enlargement was accelerated by the synthetic plant growth factor 2,4‐dichlorophenoxyacetic acid (2,4‐D) in a log dose‐dependent manner and was increased approximately 2‐fold with the addition of 1 μM 2,4‐D plus 100 μM ATP compared to 100 μM ATP alone, 1 μM 2,4‐D alone or no additions. The cell‐free enlargement was inhibited by AAA‐ATPase‐specific antisera and by CoCl2, an inhibitor specific for AAA‐ATPases. The responsible ATP site appears to be on the inside of the cell, since right side‐out vesicles did not enlarge in response to either ATP, 2,4‐D or the two in combination.
ATP‐Dependent and drug‐inhibited vesicle enlargement reconstituted using synthetic lipids and recombinant proteins
Authors: Morre D.James , Kim Chinpal , Hicks‐Berger Carrie
Published: December 2008

A recombinant ECTO‐NOX (tNOX) and a recombinant plasma membrane associated AAA‐ATPase (ATPase Associated with Different Cellular Activities) were combined in stoichometric proportions into liposomes together with al‐bumin as a source of protein thiols. Large lamellar vesicles were formed from phosphatidylcholine, cholesterol and dicetyl phosphate in a molar ratio of 50:45:5, where the phosphatidylcholine was a 2:1 mixture of synthetic dimyristoyl and dipalmitoyl phosphatidylcholines. The lipids were dried to a film and reconstituted into vesicles by resuspension in buffer containing the recombinant proteins in equimolar ratios of 0.04 nmoles/mg lipid. In the presence of ATP, these vesicles enlarged in an ATP‐dependent manner based on light‐scattering measurements. Because the drug‐inhibited ECTO‐NOX protein, tNOX was utilized, the enlargement was inhibited by capsaicin, a quinone site tNOX inhibitor specific for tNOX. With the lipid vesicle systems, the recombinant ECTO‐NOX, the recombinant AAA‐ATPase, a source of protein thiols and ATP all were required. In control experiments, no ATP‐dependent vesicle enlargement was observed with the AAA‐ATPase or the ECTO‐NOX protein alone. Also addition of ATP was without any effect when only the single proteins were incorporated into the lipid vesicles. A model has been developed whereby the plasma membrane AAA‐ATPase is linked via disulfide bonds, formed and broken by the ECTO‐NOX protein, to membrane structural proteins. Binding of ATP and subsequent hydrolysis and release of ADP would advance the ATPase hexamer ratchet thereby both thinning the membrane and increasing the vesicle surface.
Effect of polyclonal antisera to recombinant tNOX protein on the growth of transformed cells
Authors: Chen Chun‐Feng , Huang Shing , Liu Shan‐Chi , Chueh Pin Ju
Published: December 2008

Previous reports have described a tumor‐associated NADH oxidase (tNOX) and its continuous activation in transformed culture cells. Certain anticancer drugs have been shown to inhibit preferentially both the tNOX activity and the growth of transformed culture cells and the cytotoxicity is associated with the induction of apoptosis. To investigate the biological function of tNOX protein, we have raised polyclonal antisera against bacterial expressed tNOX protein and the antisera are able to recognize protein bands in transformed cells but not the non‐transformed cells tested. With tNOX antisera treatment, the survival in transformed cell lines is decreased but not the non‐transformed cells. In addition, tNOX antisera‐induced cytotoxicity is accompanied by the induction of apoptosis. However, slightly higher amount of PARP cleavage and activation of caspase‐9 are observed in tNOX antisera treated HCT116 cells. Further experiments have demonstrated the activation of JNK and phosphorylation of p53 by treatment. In addition, tNOX antisera treatment leads to an impressive increase in reactive oxygen species in COS cells but not the control sera. Our data suggest that (a) tNOX antisera treatment may inhibit the growth of transformed cells by inducing apoptosis and (b) the apoptotic mechanism might be through modulating ROS production and JNK pathway.
A plasma membrane‐associated AAA‐ATPase from Glycine max
Authors: Hicks‐Berger Carrie , Sokolchik Irina , Kim Chinpal , Morré D. James
Published: December 2008

An AAA‐ATPase (ATPases Associated with a Variety of Cellular Activities) localized to the plasma membrane of soybean (Glycine max) was isolated, partially sequenced and cloned (SBPM AAA‐ATPase). The protein with an apparent monomer molecular mass of about 97 kDa was isolated using a combination of anion exchange, preparative SDS‐PAGE, reverse phase HPLC, and ATP affinity chromatography. The cDNA for the full‐length SBPM AAA‐ATPase was cloned by screening an expression library using an antibody against the highly conserved Walker B AAA‐ATP‐binding motif. Northern blot analysis detected one transcript of ∼2700 bp. The full‐length cDNA sequence was that previously obtained (GenBank Database; U20213) encoding a protein with two copies of the conserved AAA‐ATP‐binding motif and regions of sequence homology with other AAA‐ATPases. Electron microscopic preparations of the recombinant SBPM AAA‐ATPase revealed hexamers typically formed by these proteins. The cloned and expressed protein was identical to the protein isolated from the soybean plasma membrane as confirmed using antisera raised to a non‐conserved region of the derived protein sequence and by N‐terminal sequencing of peptides derived from the isolated protein.
Drastic effect of several caffeic acid derivatives on the induction of heme oxygenase‐1 expression revealed by quantitative real‐time RT‐PCR
Authors: Suzuki Keiko , Tanaka Izumi , Nakanishi Ikuo , Kurematsu Ayako , Yakumaru Haruko , Ikota Nobuo , Ishihara Hiroshi
Published: December 2008

Among antioxidative polyphenols, caffeic acid esters such as caffeic acid phenethyl ester (CAPE) and chlorogenic acid are contained in propolis, vegetables and coffee. In this study, we compared the efficacy of some polyphenols on the activation level of a cytoprotective heme oxygenase‐1 (HO‐1) gene in RAW264.7 mouse macrophage cells using quantitative real‐time RT‐PCR. The quantitative study revealed a variety of activation level of HO‐1 gene by the chemicals. CAPE and caffeic acid ethyl ester (CAEE) at the final concentration of 2 μM drastically activated the HO‐1 gene to 39.2‐fold and 20.1‐fold, respectively. Curcumin, structurally related with caffeic acid and an element of turmeric, induced the HO‐1 gene to 5.8‐fold. In contrast, no activation was observed by other caffeic acid esters such as chlorogenic acid and rosmarinic acid. Higher concentrations were necessary for the activation by an antioxidant cysteamine and the electrophile diethyl maleate. Although the inducible activities of CAPE and chlorogenic acid were distinctly different, they showed similar reductive capacities when determined by cyclic voltammetry. These results show that the drastic activation of HO‐1 gene by CAPE and CAEE is dependent upon their chemical structures, rather than the reductive activity of polyphenols, possibly reflecting the physiological effects of the nutritional elements.
Isoliquiritigenin (ISL) inhibits ErbB3 signaling in prostate cancer cells
Authors: Jung Jae In , Chung Eunkyung , Seon Mi Ra , Shin Hyun‐Kyung , Kim Eun Ji , Lim Soon Sung , Chung Won‐Yoon , Park Kwang‐Kyun , Park Jung Han Yoon
Published: December 2008

Isoliquiritigenin (ISL), a flavonoid found in licorice, shallot, and bean sprouts, has been identified as a potent anti‐tumor promoting agent. We previously demonstrated that ISL reduces cell proliferation and induces apoptosis in DU145 human prostate cancer cells and MAT‐LyLu (MLL) rat prostate cancer cells. Overexpression of members of the ErbB receptor family is a frequently observed event in several human cancers, and ErbB receptors currently constitute the primary targets of anticancer strategies. In order to elucidate the mechanisms underlying the ISL regulation of prostate cancer cell proliferation, the present study attempted to determine whether ISL inhibits heregulin (HRG)‐β‐induced ErbB3 signaling. DU145 and MLL cells were cultured in serum‐free medium with ISL and/or HRG‐β. Exogenous HRG‐β alone was shown to effect an increase in the numbers of viable cells, whereas HRG‐β did not counteract the ISL‐induced growth inhibition. ISL reduced the protein and mRNA levels of ErbB3 in a dose‐dependent manner, but exerted no effect on HRG protein levels. Immunoprecipitation/Western blot studies indicated that ISL inhibited the HRG‐β‐induced tyrosine phosphorylation of ErbB3, the recruitment of the p85 regulatory subunit of phosphatidylinositol 3‐kinase (PI3K) to ErbB3, and Akt phosphorylation in DU145 cells. These results indicate that ISL inhibits the proliferation of prostate cancer cells, at least in part, via the inhibition of ErbB3 signaling and the PI3K/Akt pathway.
Flavonoid content in leaf extracts of the fig (Ficus carica L.), carob (Ceratonia siliqua L.) and pistachio (Pistacia lentiscus L.)
Authors: Vaya Jacob , Mahmood Saeed
Published: December 2008

The total flavonoid content of leaf extracts (70% ethanol) from fig (Ficus carica L.), carob (Ceratonia siliqua L.) and pistachio (Pistacia lentiscus L.) plants were determined by using reverse phase high‐performance liquid chromatography (HPLC)‐and analyzed by UV/VIS array and electrospray ionization (ESI)‐mass spectrometry (MS) detectors. As a base for comparison, flavonoid type and level were also determined in extracts from soybeans and grape seeds. It was found that the major flavonoids in Ficus are quercetin and luteolin, with a total of 631 and 681 mg/kg extract, respectively. In Ceratonia leaves, nine different flavonoids were detected. The major one was myricetin (1486 mg/kg extract), with a similar level in Pistacia (1331 mg/kg extract, myricetin). The present study is the first to report the presence of the isoflavone genistein in the Pistacia leaf, which was discovered to consist of about a third of the genistein level detected in soybean.
Effect of atorvastatin withdrawal on circulating coenzyme Q10 concentration in patients with hypercholesterolemia
Authors: Chu Chih‐Sheng , Kou Hwang‐Shang , Lee Chun‐Jing , Lee Kun‐Tai , Chen Su‐Hwei , Voon Wen‐Chol , Sheu Sheng‐Hsiung , Lai Wen‐Ter
Published: December 2008

Statin therapy can reduce the biosynthesis of both cholesterol and coenzyme Q10 by blocking the common upstream mevalonate pathway. Coenzyme Q10 depletion has been speculated to play a potential role in statin‐related adverse events, and withdrawal of statin is the choice in patients developing myotoxicity or liver toxicity. However, the effect of statin withdrawal on circulating levels of coenzyme Q10 remains unknown. Twenty‐six patients with hypercholesterolemia received atorvastatin at 10 mg/day for 3 months. Serum lipid profiles and coenzyme Q10 were assessed before and immediately after 3 months and were also measured 2 and 3 days after the last day on the statin. After 3 months' atorvastatin therapy, serum levels of total cholesterol (TC), low‐density lipoprotein cholesterol (LDL‐C) and coenzyme Q10 (0.43 ± 0.23 to 0.16 ± 0.10 μg/mL) were all significantly reduced (all p < 0.001). On day 2 after the last atorvastatin, the coenzyme Q10 level was significantly elevated (0.37 ± 0.16 μg/mL) and maintained the same levels on day 3 (0.39 ± 0.18 μg/mL) compared with those on month 3 (both p < 0.001), while TC and LDL‐C did not significantly change within the same 3 days. These results suggest that statin inhibition of coenzyme Q10 synthesis is less strict than inhibition of cholesterol biosynthesis.