Suppression of tumor promoter phorbolmyristate acetate-induced chromosome breakage by antioxidants and inhibitors of arachidonic acid metabolism

To gain insight into the mechanism of formation of chromosomal aberrations by the tumor promoter phorbolmyristate acetate (PMA) in human lymphocytes, we investigated the effect of antioxidants and inhibitors of arachidonic acid metabolism. Among the antioxidants bovine erythrocyte CuZn superoxide dismutase, glutathione peroxidase, mannitol (a scavenger of hydroxyl radicals), butylated hydroxytoluene and butylated hydroxyanisole were anticlastogenic while catalase and dimethylfuran (a scavenger of singlet oxygen) were inactive. These results show that the induction of aberrations by PMA occurs via indirect action, i.e. the intermediacy of superoxide and hydroxyl radicals. The following inhibitors of arachidonic acid metabolism were strongly anticlastogenic: the cyclo-oxygenase inhibitors indomethacin and flufenamic acid and the lipoxygenase inhibitor BN1015. Imidazole, nordihydroguaiaretic acid BN 1048 and 5,8,11,14-eicosatetraynoic acid were moderately active. The inhibitor of phospholipase A₂, fluocinolone acetonide, was also anticlastogenic.

We conclude that the oxidative metabolism of arachidonic acid is involved in the induction of chromosomal aberrations by PMA in human lymphocytes. However, because of the limited selectivity of these drugs, it is not yet possible to identify unambiguously the step(s) in the arachidonic acid cascade responsible for PMA clastogenicity.     

Effects of butylated hydroxyanisole,butylated hydroxytoluene and tertiary butylhydroquinone on growth and luteoskyrin production byPenicillium islandicum

Butylated hydroxyanisole (BHA), Butylated hydroxytoluene (BHT) and tertiary butylhydroquinone (TBHQ) alone in cultural media were tested for the inhibition of growth and luteoskyrin production by two toxigenic strains ofPenicillium islandicum UST-11 andP. islandicum HLT-6. In potato dextrose agar, the concentrations of BHA and TBHQ from 0.2 mg/disc, BHT from 5.0 mg/disc did affect the growth of both tested strains, but the initial concentrations of these antioxidants to reduced luteoskyrin production by UST-11 strain were BHA 0.5 mg/disc, BHT 1.0 mg/disc and TBHQ 0.4 mg/disc, while for HLT-6, BHA 0.4 mg/disc, BHT and TBHQ were 0.2 mg/disc, respectively. In grainy and powdery rice media, the effects of BHA, BHT and TBHQ on luteoskyrin production byP. islandicum UST-11 and HLT-6 were clearly demonstrated. The efficiency of the inhibitory effect was not only closely related to the concentration of antioxidants, but also completely inhibited the luteoskyrin production at a concentration of 200 mg/kg or higher. Also, the antioxidants at a concentration higher than 20 mg/kg reduced significantly the growth and luteoskyrin production by both strains ofP. islandicum.     

Nitrous acid mutagenesis of duplex DNA as a three-component system.

Purified native Hemophilus influenzae DNA is relatively insusceptible to nitrous acid (NA) mutagenesis in vitro, but is readily mutated following denaturation. NA mutagenicity for duplex DNA is significantly increased in the presence of various alcohols, glycols, phenols or primary amines. Phenol-extracted DNA contains dissociable contaminants of low molecular weight that enhance NA mutagenesis. Enhancement of NA mutagenesis by phenol and by spermine is due to the formation of unstable molecular species. We propose that reactive organic nitroso compounds are formed which then serve as delivery vehicles to promote mutagenicity of native DNA, perhaps via transnitrosation reactions. Similar reactions probably occur in vivo to promote NA-induced base substitution (but not frameshift) mutations in Salmonella typhimurium and in Escherichia coli. The possible significance of these observations to carcinogenesis is discussed.     

Effects of butylated hydroxyanisole on the aryl hydrocarbon hydroxylase of rats and mice

The effects of butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) on the aryl hydrocarbon hydroxylase (AHH) activities in the liver, lung and skin of rats and mice have been studied to examine the possible mechanisms of the anticarcinogenic actions of these compounds. Both compounds inhibit the hydroxylase activities of hepatic microsomes and nuclei, with BHA a more potent inhibitor than BHT. The AHH of lung microsomes is inhibited to a lesser extent by BHA and BHT than that of the liver. The AHH activities of both liver and lung microsomes become less susceptible to the inhibition after pretreatment of the animals with 3-methylcholanthrene (MC) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) but phenobarbital (PB) pretreatment does not produce such an effect. In skin homogenates, however, the AHH activities of control rats and mice are not inhibited by BHA and BHT. The only skin sample which is inhibited by BHA and BHT is that from TCDD-pretreated mice. It has been established that the extent of inhibition with different samples is related to the concentration of BHA in the incubation but not to the amounts or specific activities of microsomes used. Double reciprocal plots suggest that BHA exerts a mixed inhibition on the hydroxylase of liver microsomes with a K_i of 7.7 μM. Analysis of the metabolites of benzo[a]pyrene (BP) shows that BHA inhibits the formation of various metabolites uniformly without changing the regio-selectivity of the enzyme system. The mechanism of inhibition has also been studied with a reconstituted AHH system consisting of cytochrome P-450 (P-450), reductase and phospholipid. The system with P-450 isolated from PB-induced microsomes is inhibited to a much greater extent than that with MC-induced P-450. The results indicate that the inhibitory action of BHA is dependent on the species of the animal, tissue types and treatment with inducers.     

Iron chelation or anti-oxidants prevent renal cell damage in the rewarming phase after normoxic, but not hypoxic cold incubation

It has now been firmly established that, not only ischemia/reperfusion, but also cold itself causes damage during kidney transplantation. Iron chelators or anti-oxidants applied during the cold plus rewarming phase are able to prevent this damage. At present, it is unknown if these measures act only during the cold, or whether application during the rewarming phase also prevents damage. We aimed to study this after cold normoxic and hypoxic conditions. LLC-PK1 cells were incubated at 4 degrees C in Krebs-Henseleit buffer for 6 or 24h, followed by 18 or 6h rewarming, respectively. Cold preservation was performed under both normoxic (95% air/5% CO2) and hypoxic (95% N2/5% CO2) conditions. The iron chelator 2,2'-DPD (100 microM), anti-oxidants BHT (20 microM) or sibilinin (200 microM), and xanthine oxidase inhibitor allopurinol (100 microM) were added during either cold preservation plus rewarming, or rewarming alone. Cell damage was assessed by LDH release (n=3-9). Addition of 2,2'-DPD and BHT during cold hypoxia plus rewarming did, but during rewarming alone did not prevent cell damage. When added during rewarming after 6h cold normoxic incubation, BHT and 2,2'-DPD inhibited rewarming injury compared to control (p<0.05). Allopurinol did not prevent cell damage in any experimental set-up. Our data show that application of iron chelators or anti-oxidants during the rewarming phase protects cells after normoxic but not hypoxic incubation. Allopurinol had no effect. Since kidneys are hypoxic during transplantation, measures aimed at preventing cold-induced and rewarming injury should be taken during the cold.     

Cannabidiol as an emergent therapeutic strategy for lessening the impact of inflammation on oxidative stress

Oxidative stress with reactive oxygen species generation is a key weapon in the arsenal of the immune system for fighting invading pathogens and initiating tissue repair. If excessive or unresolved, however, immune-related oxidative stress can initiate further increasing levels of oxidative stress that cause organ damage and dysfunction. Targeting oxidative stress in various diseases therapeutically has proven more problematic than first anticipated given the complexities and perversity of both the underlying disease and the immune response. However, growing evidence suggests that the endocannabinoid system, which includes the CB₁ and CB₂ G-protein-coupled receptors and their endogenous lipid ligands, may be an area that is ripe for therapeutic exploitation. In this context, the related nonpsychotropic cannabinoid cannabidiol, which may interact with the endocannabinoid system but has actions that are distinct, offers promise as a prototype for anti-inflammatory drug development. This review discusses recent studies suggesting that cannabidiol may have utility in treating a number of human diseases and disorders now known to involve activation of the immune system and associated oxidative stress, as a contributor to their etiology and progression. These include rheumatoid arthritis, types 1 and 2 diabetes, atherosclerosis, Alzheimer disease, hypertension, the metabolic syndrome, ischemia-reperfusion injury, depression, and neuropathic pain.     

Toxicity of depleted uranium on isolated rat kidney mitochondria

Background

Kidney is known as the most sensitive target organ for depleted uranium (DU) toxicity in comparison to other organs. Although the oxidative stress and mitochondrial damage induced by DU has been well investigated, the precise mechanism of DU-induced nephrotoxicity has not been thoroughly recognized yet.

Methods

Kidney mitochondria were obtained using differential centrifugation from Wistar rats and mitochondrial toxicity endpoints were then determined in both in vivo and in vitro uranyl acetate (UA) exposure cases.

Results

Single injection of UA (0, 0.5, 1 and 2 mg/kg, i.p.) caused a significant increase in blood urea nitrogen and creatinine levels. Isolated mitochondria from the UA-treated rat kidney showed a marked elevation in oxidative stress accompanied by mitochondrial membrane potential (MMP) collapse as compared to control group. Incubation of isolated kidney mitochondria with UA (50, 100 and 200 μM) manifested that UA can disrupt the electron transfer chain at complex II and III that leads to induction of reactive oxygen species (ROS) formation, lipid peroxidation, and glutathione oxidation. Disturbances in oxidative phosphorylation were also demonstrated through decreased ATP concentration and ATP/ADP ratio in UA-treated mitochondria. In addition, UA induced a significant damage in mitochondrial outer membrane. Moreover, MMP collapse, mitochondrial swelling and cytochrome c release were observed following the UA treatment in isolated mitochondria.

General significance

Both our in vivo and in vitro results showed that UA-induced nephrotoxicity is linked to the impairment of electron transfer chain especially at complex II and III which leads to subsequent oxidative stress.     

Jacaric acid,a linolenic acid isomer with a conjugated triene system,has a strong antitumor effect in vitro and in vivo

In this study, we compared the cytotoxic effects of natural conjugated linolenic acids (CLnAs) on human adenocarcinoma cells (DLD-1) in vitro, with the goal of finding CLnA isomers with strong cytotoxic effects. The antitumor effect of the CLnA with the strongest cytotoxic effect was then examined in mice. The results showed that all CLnA isomers have strong cytotoxic effects on DLD-1 cells, with jacaric acid (JA) having the strongest effect. Examination of the mechanism of cell death showed that CLnAs induce apoptosis in DLD-1 cells via lipid peroxidation. The intracellular levels of incorporated CLnAs were measured to examine the reason for differences in cytotoxic effects. These results showed that JA was taken into cells efficiently. Collectively, these results suggest that the cytotoxic effect of CLnAs is dependent on intracellular incorporation and induction of apoptosis via lipid peroxidation. JA also had a strong preventive antitumor effect in vivo in nude mice into which DLD-1 cells were transplanted. These results suggest that JA can be used as a dietary constituent for prevention of cancer.     

气相色谱法测定食品中的抗氧化剂BHA，BHT，TBHQ

本文利用甲醇和石油醚直接提取油脂和含油脂食品中的抗氧化剂丁基羟基茴香醚(BHA),二丁基羟基甲苯(BHT),特丁基对苯二酚(TBHQ),建立同时测定这三种抗氧化剂的气相色谱方法。采用安捷伦HP-5毛细管柱(i.d.0.25μm×30.0m),氢火焰离子化检测器,外标法定量测量,经实验,样品加标回收率为95%-104%,检出限分别为BHA:0.22 mg/L,BHT:0.23 mg/L,TBHQ:0.33 mg/L。该方法具有前处理操作简便,灵敏度高,重现性好等优点。     

Glycyrrhetinic acid as inhibitor or amplifier of permeability transition in rat heart mitochondria

Glycyrrhetinic acid (GE), a hydrolysis product of glycyrrhizic acid, one of the main constituents of licorice root, is able, depending on its concentration, to prevent or to induce the mitochondrial permeability transition (MPT) (a phenomenon related to oxidative stress) in rat heart mitochondria (RHM). In RHM, below a threshold concentration of 7.5 μM, GE prevents oxidative stress and MPT induced by supraphysiological Ca²⁺ concentrations. Above this concentration, GE induces oxidative stress by interacting with a Fe-S centre of Complex I, thus producing ROS, and amplifies the opening of the transition pore, once again induced by Ca²⁺. GE also inhibits Ca²⁺ transport in RHM, thereby preventing the oxidative stress induced by the cation. However, the reduced amount of Ca²⁺ transported in the matrix is sufficient to predispose adenine nucleotide translocase for pore opening. Comparisons between observed results and the effects of GE in rat liver mitochondria (RLM), in which the drug induces only MPT without exhibiting any protective effect, confirm that it interacts in a different way with RHM, suggesting tissue specificity for its action. The concentration dependence of the opposite effects of GE, in RHM but not RLM, is most probably due to the existence of a different, more complex, pathway by means of which GE reaches its target. It follows that high GE concentrations are necessary to stimulate the oxidative stress capable of inducing MPT, because of the above effect, which prevents the interaction of low concentrations of GE with the Fe-S centre. The reported results also explain the mechanism of apoptosis induction by GE in cardiomyocytes.