Ethanol-Induced Cell Death by Lipid Peroxidation in PC12 Cells

Free radical generation is hypothesized to be the cause of alcohol-induced tissue injury. Using fluorescent cis-parinaric acid and TBARS, lipid peroxidation was shown to be increased in the presence of trace amounts of free ferrous ion in PC12 cells. This increase in lipid peroxidation was enhanced by ethanol in a dose dependent manner and also correlated with loss of cell viability, as measured by increased release of lactate dehydrogenase (LDH). Resveratrol, a potent antioxidant, had a protective effect against lipid peroxidation and cell death. These findings strongly suggest that ethanol-induced tissue injury and cell death is a free radical mediated process, and may be important in alcohol-related premature aging and other degenerative diseases.     

Lipid Peroxidation and Antioxidant Systems in Rat Brain: Effect of Chronic Alcohol Consumption

The effect of chronic ethanol exposure, in a liquid diet, on lipid peroxidation and some antioxidant systems of rat brain was investigated. Chronic ethanol administration induced a greater susceptibility to iron/ascorbate-induced lipid peroxidation, estimated as thiobarbituric reactive substances (TBARS) production, in the microsomal fraction, but a lower lipid peroxidation in the total homogenate. Glutathione (GSH) levels as well as GSH peroxidase and GSH reductase were unaffected, while the activity of Cu-Zn superoxide dismutase was decreased and that of catalase increased. Lipid peroxidation experiments performed in the presence of some hydroxyl radical scavengers suggested that a greater OH^· generation may be responsible of the greater TBARS production in the microsomal fraction of ethanol treated rats; differently, in total homogenate of control and ethanol rats a relationship was found between the redox state of iron and TBARS production, suggesting that the lower lipid peroxidation in treated rats may depend on a different modulation of the iron redox state.     

Lipid Requirements and Lipid Uptake by Giardia lamblia Trophozoites in Culture

To better understand the lipid requirements of Giardia lamblia trophozoites and the mechanisms of lipid uptake, we supplemented serum-free TYI-S-33 medium with lipids incorporated into different lipid carriers. We found that serum lipoproteins, β-cyclodextrins, and bile salts are able to supply cholesterol and phospholipids to Giardia and to support the multiplication of the parasite in vitro. The growth rates obtained with different lipoproteins or bile salts and lipid mixtures were similar to that in standard culture medium containing serum. Pulse labelling experiments using fluorescent lipid analogs demonstrated that Giardia can take up lipids from lipoproteins, β-cyclodextrins, or bile salt micelles, but with different kinetics, and that bile salts greatly facilitated lipid transfer from lipoproteins and cyclodextrins to the parasite surface. The binding of different radioiodinated lipoprotein classes to the trophozoite surface, inhibition of lipoprotein interiorization at 4°C or by cytochalasin D, and incorporation studies using fluorescent LDL suggested that a small component of lipid uptake by trophozoites was likely due to endocytosis of lipoproteins.     

高温与香烟联合对大鼠早期胚胎致畸作用的研究

为了探讨高温与香烟联合对胚胎的毒性和致畸作用,用高温（４０、４１、４２、４３℃）及香烟烟不溶４物联合处理怀孕的大鼠。结果表明,高温与香烟联合可加强胚胎的致畸作用,并与温度、香烟浓度和高温持续时间明显相关,当持续３分钟的高温与香烟联合,胚胎致畸率达７０％以上,与单独高温组、单独给烟组有明显差异（Ｐ〉０．０１）,并具有协同效应。结果提示,持续高温与香烟联合有明显的胚胎毒性和致畸作用。     

Antioxidant Properties of New Chalcogenides Against Lipid Peroxidation in Rat Brain

Ebselen (2-phenyl- 1,2-benzisoselenazole-3 (2H)-one) is a seleno-organic compound with antioxidant properties, and anti-inflammatory actions. Recently, ebselen improved the outcome of acute ischemic stroke in humans. In the present study, the potential antioxidant capacity of organochalcogenide compounds diphenyl diselenide (PhSe)₂, diphenyl ditelluride (PhTe)₂, diphenyl disulfide (PhS)₂, p-Cl-diphenyl diselenide (pCl-PhSe)₂, bis-[S-4-isopropyl 2-phenyl oxazoline] diselenide (AA-Se)₂, bis-[S-4-isopropyl 2-phenyl oxazoline] ditelluride (AA-Te)₂ and bis-[S-4-isopropyl 2-phenyl oxazoline] disulfide (AA-S)₂ was compared with that of ebselen (a classical antioxidant). Spontaneous and quinolinic acid (QA)- (2 mM) and sodium nitroprusside (SNP)- (5 M)-induced thiobarbituric reactive species (TBARS) production by rat brain homogenates was determined colorimetrically. TBARS formation was reduced by ebselen, (PhSe)₂, (PhTe)₂, (AA-Se)₂, (AA-S)₂ and (pCl- PhSe)₂ to basal rates. The concentrations of these compounds needed to inhibit TBARS formation by 50% (lC₅₀) are 1.71 M, 3.73 M, 1.63 M, 9.85 M, > 33.3 M, 23.2 M and 4.83 M, respectively for QA. For TBARS production induced by SNP the lC₅₀ was 2.02 M, 12.5 M, 2.80 M, > 33.3 M, 24.5 M and 7.55 M, respectively. The compounds (AA-Te)₂ and (PhS)₂ have no antioxidant activity and pro-oxidant activity, respectively. These results suggest that (AA-Se)₂ and (AA-S)₂ can be considered as potential pharmaceutical antioxidant agents.     

Inhibitory Effects of Ebselen on Lipid Peroxidation in Rat Liver Microsomes

The effects of ebselen(2-pheny1-1,2-benzoisoselenazol-3(2H)-one), a synthetic seleno-organic compound with glutathione peroxidase-like activity were investigated on lipid peroxidation in rat liver microsomes. Ebselen inhibited malondialdehyde production coupled to the lipid peroxidation stimulated by either ADP-iron-ascorbate or CC1₄. The inhibitory activity of ebselen on each system was strongly increased by a 5-min preincubation with liver microsomes; the IC₅₀ values against ADP-Fe-ascorbate-stimulated and CC1₄-stimulated lipid peroxidation were 1.6/jM and 70 μM respectively. Ebselen also inhibited the endogenous lipid peroxidation with a NADPH-generating system, but it slightly stimulated the endogenous activity of ADP-Fe-ascorbate-stimulated lipid peroxidation (without a NADPH-generating system). Furthermore, ebselen inhibited oxygen uptake coupled to the lipid peroxidation by ADP-Fe-ascorbate and NADPH-ADP-iron; the IC₅₀ values were 2.5μM AND 20.3 μM respectively. Ebselen also prolonged the lag-time of onset of ADP-Fe-ascorbate-stimulated lipid peroxidation significantly, but not that observed with NADPH-ADP-Fe-stimulated lipid peroxidation.     

Curcumin Prevents Bile Canalicular Alterations in the Liver of Hamsters Infected with Opisthorchis viverrini

Opisthorchis viverrini infection causes inflammation and liver injury leading to periductal fibrosis. Little is known about the pathological alterations in bile canaliculi in opisthorchiasis. This study aimed to investigate bile canalicular alterations in O. viverrini-infected hamsters and to examine the chemopreventive effects of curcumin on such changes. Hamsters were infected with O. viverrini and one group of animals was fed with 1% dietary curcumin supplement. Animals were examined during the acute infection phase, days 21 and 30 post-infection (PI) and chronic infection phase (day 90 PI). Scanning electron microscopy revealed that in the infected group fed with a normal diet, bile canaliculi became slightly tortuous by 30 day PI and more tortuous at day 90 PI. Transmission electron microscopy showed a reduction in microvilli density of canaliculi starting at day 30 PI, with a marked loss of microvilli at day 90 PI. These ultrastructral changes were slightly seen at day 21 PI, which was similar to that found in infected animals fed with 1% curcumin-supplemented diet. Notably, curcumin treatment prevented the reduction of microvilli density, reduced the dilation of bile canaliculi, and decreased the tortuosity of the bile canaliculi relative to non-infected animals on a normal diet at days 30 and 90 PI. These results suggest that curcumin reduces alteration of bile canaliculi and may be a promising agent to prevent the onset of bile duct abnormalities induced by O. viverrini infection.     

Increase of Lipid Hydroperoxides in the Rat Liver and Kidney after Administering Ferric Nitrilotriacetate

Two hours after an intraperitoneal injection of ferric nitrilotriacetate to rats at a dose of 15 mg of Fe/kg of body weight, the level of lipid hydroperoxides, which was determined by a specific method involving chemical conversion into 1-naphthyldiphenylphosphine oxide and HPLC, had increased significantly in the liver (from 190.1±58.8 of the control to 467.1±175.8 pmol/mg of protein) and kidney (from 181.8±52.3 of the control to 405.9±22.7 pmol/mg of protein). These results demonstrate that oxidative stress was transiently increased by an iron overload in these tissues.     

Effect of Fish Oil and Coconut Oil on Antioxidant Defence System and Lipid Peroxidation in Rat Liver

Diets high in fish oil containing polyunsaturated fatty acids of the n-3 family. have been suggested to decrease the risk of cardiovascular disease. However these lipids are highly susceptible to oxidative deterioration. In order to investigate the influence of n-3 fatty acids on oxidative status, the effect of feeding rats with fish oil or cocunut oil diets was studied by measuring different parameters related to an oxidative free radical challenge. Synthetic diets containing 15% (w/v) fish oil or coconut oil were used to feed growing rats for 4 weeks. As compared to control diet, the fish oil containing diet produced a significant decrease of cholesterol and triglyceride concentration in serum. however there was a significant increase in lipid peroxidation products. In addition, in fish oil fed animals, there was also a decrease in vitamin E and A concentration. Furthermore, the rate of lipid peroxidation in isolated microsomes was three fold higher in rats fed fish oil as compared to rats with coconut oil diet. No significant differences between the two experimental groups were observed in superoxide dismutase (SOD) and phospholipid hydroperoxide glutathione peroxidase (PHGPX) activities. However, there was a decrease in glutathione peroxidase (GPX) activity. These results suggest that fish oil feeding at an amount compatible with human diet, although decreasing plasma lipids, actually challenge the antioxidant defence system, thus increasing the susceptibility of tissues to free radical oxidative damage.     

Antimalarial Drugs Exacerbate Rat Liver Microsomal Lipid Peroxidation in the Presence of Oxidants

The study was undertaken to evaluate the effect of prior treatment of rats with the antimalarial drugs amodiaquine (AQ) mefloquine (MQ) and halofantrine (HF) on rat liver microsomal lipid peroxidation in the presence of 1 mM FeSO4, 1 mM ascorbate and 0.2 mM H₂O₂ (oxidants). Ingestion of -tocopheral, a radical chain-breaking antioxidant was also included to assess the role of antioxidants in the drug treatment. In the presence of oxidants AQ, MQ and HF elicited 288%, 175% and 225% increases in malondialdehyde (MDA) formation while the drugs induced 125%, 63% and 31% increases in the absence of oxidants respectively. Similarly, AQ, MQ and HF induced lipid hydroperoxide formation by 380%, 256%, 360% respectively in the presence of oxidants and 172%, 136% and 92% in the absence of exogenously added oxidants respectively. -tocopherol reduced AQ, MQ and HF-induced MDA formation by 40%, 55% and 52% respectively and lipid hydroperoxide formation by 53%, 59% and 54% respectively. Similarly, -tocopherol attenuated the AQ, MQ and HF-induced MDA formation by 49%, 51% and 51% in the presence of oxidants and lipid hydroperoxide formation by 61%, 62% and 47% respectively. The results indicate that rat liver microsomal lipid peroxidation could be enhanced by antimalarial drugs in the presence of reactive oxygen species and this effect could be ameliorated by treatment with antioxidants.     

Effect of Fish Oil and Coconut Oil on Antioxidant Defence System and Lipid Peroxidation in Rat Liver

Diets high in fish oil containing polyunsaturated fatty acids of the n-3 family. have been suggested to decrease the risk of cardiovascular disease. However these lipids are highly susceptible to oxidative deterioration. In order to investigate the influence of n-3 fatty acids on oxidative status, the effect of feeding rats with fish oil or cocunut oil diets was studied by measuring different parameters related to an oxidative free radical challenge. Synthetic diets containing 15% (w/v) fish oil or coconut oil were used to feed growing rats for 4 weeks. As compared to control diet, the fish oil containing diet produced a significant decrease of cholesterol and triglyceride concentration in serum. however there was a significant increase in lipid peroxidation products. In addition, in fish oil fed animals, there was also a decrease in vitamin E and A concentration. Furthermore, the rate of lipid peroxidation in isolated microsomes was three fold higher in rats fed fish oil as compared to rats with coconut oil diet. No significant differences between the two experimental groups were observed in superoxide dismutase (SOD) and phospholipid hydroperoxide glutathione peroxidase (PHGPX) activities. However, there was a decrease in glutathione peroxidase (GPX) activity. These results suggest that fish oil feeding at an amount compatible with human diet, although decreasing plasma lipids, actually challenge the antioxidant defence system, thus increasing the susceptibility of tissues to free radical oxidative damage.     

红葡萄酒对大鼠肝脏抗氧化酶和脂质过氧化的影响

选用雄性SD大鼠,分别灌胃红葡萄酒、酒精及水。实验90 d中每隔30 d处死一批动物,测定大鼠肝脏匀浆组织中的超氧化物歧化酶(Superoxide dismutase SOD)、过氧化氢酶(Catalase CAT)、谷胱甘肽过氧化物酶(Glutathione peroxidase GSH-Px)活性和脂质过氧化产物丙二醛(Malondialdehyde MDA)含量变化。观察摄入红葡萄酒后大鼠肝脏抗氧化酶活性变化及对肝脏脂质过氧化的影响。结果表明,红葡萄酒能提高SOD活性,且SOD活性与灌胃时间、剂量有一定关系;长期红葡萄酒和酒精摄入可诱导CAT活性增强,加剧肝脏的脂质过氧化(LPO)作用;红葡萄酒组、酒精组0.63、1.25 g/kg剂量GSH-Px活性均明显升高(P<0.05),酒精组1.88 g/kg剂量有极显著性差异(P<0.01);试验初期,红葡萄酒大剂量显著降低肝脏中MDA的含量。试验中期,红葡萄酒中大剂量显著降低MDA含量。试验末期,红葡萄酒大剂量和酒精中大剂量显著升高肝脏中MDA含量。     

Ageing-Induced Alterations in Lipid/Phospholipid Profiles of Rat Brain and Liver Mitochondria: Implications for Mitochondrial Energy-Linked Functions

Effects of ageing on the lipid/phospholipid profile of brain and liver mitochondria from rats were examined. In the brain mitochondria the contents of total phospholipid (TPL) and cholesterol (CHL) increased with simultaneous increase in the TPL/CHL (mole:mole) ratio. The proportion and contents of lysophospholipid (Lyso), sphingomyelin (SPM), phosphatidylinositol (PI), phosphatidylserine (PS) and diphosphatidylglycerol (DPG) components increased, with maximal increases seen for PS and PI; phosphatidylcholine (PC) and phosphatidylethanolamine (PE) components registered decrease. In the liver mitochondria contents of TPL and CHL increased. However, the TPL/CHL (mole:mole) ratio was not altered. Lyso, PI and PS increased. However, the magnitude of increase was competitively lower; PE and DPG decreased. SPM and PC did not change as a consequence of ageing. These changes altered the contents of individual phospholipids in the two membrane systems. Respiration with glutamate, pyruvate + malate, succinate and ascorbate + N,N,N’,N’-tetramethyl-p-phenylenediamine was significantly impaired in brain mitochondria from old animals. For liver mitochondria the respiratory activity declined with glutamate and succinate. Correlation studies by regression analysis revealed that the lipid/phospholipid classes regulate respiratory function differently in the mitochondria from the two tissues. The respiration-related parameters in the brain mitochondria were dependent on multiple lipid/phospholipid components, and the process of regulation was complex compared to the liver mitochondrial functions.     

Two Mechanisms of Cc14-Induced Fatty Liver: Lipid Peroxidation Or Covalent Binding Studied in Cultured Rat Hepatocytes

With cultured hepatocytes it was studied whether CCl₄-induced inhibition of secretion of VLDL and HDL from liver cells is a consequence of covalent binding of CC1₄ metabolites (i.e. CO,; CC1,00) to cell constituents or of membrane damage by lipid peroxidation. Comparing the kinetics of inhibition of lipoprotein secretion with that of CCl₄-bioactivation it was found, that covalent binding of (^HC)-CC1₄ occurred at early time points (5 min) after CC1₄ administration and inhibited the lipoprotein secretion. At 100μM CC1₄ it was depressed by 53% within 60min. Incubations of CC1₄-treated cells with increasing concentrations of vitamin E blocked lipid peroxidation, but lipoprotein secretion was still inhibited. Piperonyl butoxid, a radical scavenger, protected against CCl₄-induced inhibition of lipoprotein section, lipid peroxidation and covalent binding.

These results show that during the early phases of CC1₄ poisoning fat accumulation is the consequence of covalent binding of CC1₄ metabolities to cell structures.     

Alternative Roles of STAT3 and MAPK Signaling Pathways in the MMPs Activation and Progression of Lung Injury Induced by Cigarette Smoke Exposure in ACE2 Knockout Mice

Inflammation-mediated abnormalities in the renin-angiotensin system (RAS) and expression of matrix metalloproteinases (MMPs) are implicated in the pathogenesis of lung injury. Angiotensin converting enzyme II (ACE2), an angiotensin converting enzyme (ACE) homologue that displays antagonist effects on ACE/angiotensin II (Ang II) axis, could also play a protective role against lung diseases. However, the relationship between ACE2 and MMPs activation in lung injury is still largely unclear. The purpose of this study is to investigate whether MMPs activity could be affected by ACE2 and which ACE2 derived signaling pathways could be also involved via using a mouse model with lung injury induced by cigarette smoke (CS) exposure for 1 to 3 weeks. Wild-type (WT; C57BL/6) and ACE2 KO mice (ACE2^-/-) were utilized to study CS-induced lung injury. Increases in the resting respiratory rate (RRR), pulmonary immunokines, leukocyte infiltration and bronchial hyperplasia were observed in the CS-exposed mice. Compared to WT mice, more serious physiopathological changes were found in ACE2^-/- mice in the first week of CS exposure. CS exposure increased pulmonary ACE and ACE2 activities in WT mice, and significantly increased ACE in ACE2^-/- mice. Furthermore, the activity of pulmonary MMPs was decreased in CS-exposed WT mice, whereas this activity was increased in ACE2^-/- mice. CS exposure increased the pulmonary p-p38, p-JNK and p-ERK1/2 level in all mice. In ACE2^-/- mice, a significant increase p-STAT3 signaling was detected; however, no effect was observed on the p-STAT3 level in WT mice. Our results support the hypothesis that ACE2 deficiency influences MMPs activation and STAT3 phosphorylation signaling to promote more pulmonary inflammation in the development of lung injury.     

Effects of Cigarette Smoke Extracts on the Growth and Senescence of Skin Fibroblasts In Vitro

Epidemiological studies have shown that cigarette smoke (CS), a very common environmental factor, plays an important role in skin aging. Although some in vivo studies have suggested that CS affects skin aging, the detailed effects of CS on skin cells in vitro remain largely unknown. In this study, we investigated the effects of cigarette smoke extract (CSE) on the growth, proliferation, and senescene of skin fibroblasts and the possible mechanism underlying these effects. Primary cultured human fibroblasts were exposed to a range of concentrations of CSE. Cell viability and cell proliferation after CSE exposure were analyzed with the methyl thiazolyl tetrazolium (MTT) assay and bromodeoxyuridine incorporation assay, respectively. Growth curves of fibroblasts exposed to different concentrations of CSE were developed and prolonged CSE-exposed cells were observed. Morphological and ultrastructural changes in fibroblasts were assessed by inverted light microscopy and transmission electron microscopy (TEM). Dying cells were stained with senescence-associated β-galactosidase (SA β-gal). Intracellular reactive oxygen species (ROS) levels, superoxide dismutase (SOD) activity, and glutathione peroxidase (GSH-Px) activity were determined by a colorimetric method. We found that proliferative capacity and growth were inhibited by CSE exposure in a dose- and time-dependent manner. Fibroblasts exposed to even low concentrations of CSE for a long period of time (5 passages) showed significantly increased SA β-gal activity and typical features of aging cells. Meanwhile, CSE inhibited superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities and augmented ROS levels. Our observations suggest that CSE exposure impairs fibroblast growth and proliferation and leads to features similar to those seen in senescent cells. Oxidative stress injury and inhibition of antioxidant defense activity may be involved in CSE-induced fibroblast senescence.     

Curcumin prevents maleate-induced nephrotoxicity: Relation to hemodynamic alterations,oxidative stress,mitochondrial oxygen consumption and activity of respiratory complex I

Abstract

The potential protective effect of the dietary antioxidant curcumin (120 mg/Kg/day for 6 days) against the renal injury induced by maleate was evaluated. Tubular proteinuria and oxidative stress were induced by a single injection of maleate (400 mg/kg) in rats. Maleate-induced renal injury included increase in renal vascular resistance and in the urinary excretion of total protein, glucose, sodium, neutrophil gelatinase-associated lipocalin (NGAL) and N-acetyl β-D-glucosaminidase (NAG), upregulation of kidney injury molecule (KIM)-1, decrease in renal blood flow and claudin-2 expression besides of necrosis and apoptosis of tubular cells on 24 h. Oxidative stress was determined by measuring the oxidation of lipids and proteins and diminution in renal Nrf2 levels. Studies were also conducted in renal epithelial LLC-PK1 cells and in mitochondria isolated from kidneys of all the experimental groups. Maleate induced cell damage and reactive oxygen species (ROS) production in LLC-PK1 cells in culture. In addition, maleate treatment reduced oxygen consumption in ADP-stimulated mitochondria and diminished respiratory control index when using malate/glutamate as substrate. The activities of both complex I and aconitase were also diminished. All the above-described alterations were prevented by curcumin. It is concluded that curcumin is able to attenuate in vivo maleate-induced nephropathy and in vitro cell damage. The in vivo protection was associated to the prevention of oxidative stress and preservation of mitochondrial oxygen consumption and activity of respiratory complex I, and the in vitro protection was associated to the prevention of ROS production.     

姜黄素对抗高脂饮食诱导SD大鼠血脂水平升高及改善血管内皮依赖性舒张功能的研究

目的：观察高脂饮食对SD大鼠血脂水平及胸主动脉环舒张功能的影响及姜黄素对以上改变的影响。方法：健康SD大鼠30只,分高脂饮食组（10只）、正常饮食对照组、高脂饮食＋姜黄素组（10只）,大鼠行适应性饲养1周后分别给予高脂饮食及正常饮食;于实验开始时、10周及实验结束前测各组大鼠体重,20周后取血测定血清血脂浓度,取胸主动脉测定血管环舒张功能。结果：①高脂饮食喂养的大鼠体重明显高于其他各组大鼠,姜黄素可明显对抗高脂饮食导致的体重升高。②与对照组比较,高脂饮食组TC、TG、LDL-C明显升高（P〈0.01和P〈0.05）;③与正常对照组及姜黄素对照组比较,高脂饮食组胸主动脉环的内皮依赖性舒张功能显著减弱（P〈0.05）。结论：①SD大鼠给予高脂饮食后使大鼠血脂水平明显升高,胸主动脉环内皮依赖性舒张功能显著减弱。②姜黄素具有防治高脂饮食导致的血脂升高及改善高脂饮食导致的血管内皮依赖性的舒张功能减退。