Increases in 4-hydroxynonenal and hexanal in bone marrow of rats subjected to total body X-ray irradiation: association with antioxidant vitamins.
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Radiation-induced lipid peroxidation and its association with antioxidant vitamins in the bone marrow (BM), of rats subjected to total body irradiation (TBI) of X-rays at a dose of 3 Gy was investigated. The concentration of vitamin C in the BM decreased at 4 h, and reached about 2% of the control level at 24 h after irradiation. The concentration of vitamin E in the BM also decreased to 43% at 24 h. Corresponding to the decrease in vitamin E concentration, the concentration of 4-hydroxynonenal (HNE) in the BM increased 2.5-fold at 24 h. Similarly, increases in the concentrations of hexanal and thiobarbituric acid-reactive substances (TBA-RS) were detected in the BM. In the plasma, these parameters of lipid peroxidation were unchanged up to 48 h, but were increased at 96 h after irradiation. Four days of vitamin E administration to rats (p.o. 460 mg/kg body weight) prior to the 3 Gy X-irradiation increased the vitamin E concentration in the BM to 1.3-fold the control level, but did not attenuate the increases in HNE and hexanal in the BM. The slight accumulation of vitamin E in the BM as a result of the vitamin E treatment may be partly related to this lack of vitamin E effect. (+info)
Protective function of chloroplast 2-cysteine peroxiredoxin in photosynthesis. Evidence from transgenic Arabidopsis.
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2-Cysteine peroxiredoxins (2-CPs) constitute a ubiquitous group of peroxidases that reduce cell-toxic alkyl hydroperoxides to their corresponding alcohols. Recently, we cloned 2-CP cDNAs from plants and characterized them as chloroplast proteins. To elucidate the physiological function of the 2-CP in plant metabolism, we generated antisense mutants in Arabidopsis. In the mutant lines a 2-CP deficiency developed during early leaf and plant development and eventually the protein accumulated to wild-type levels. In young mutants with reduced amounts of 2-CP, photosynthesis was impaired and the levels of D1 protein, the light-harvesting protein complex associated with photosystem II, chloroplast ATP synthase, and ribulose-1,5-bisphosphate carboxylase/oxygenase were decreased. Photoinhibition was particularly pronounced after the application of the protein synthesis inhibitor, lincomycin. We concluded that the photosynthetic machinery needs high levels of 2-CP during leaf development to protect it from oxidative damage and that the damage is reduced by the accumulation of 2-CP protein, by the de novo synthesis and replacement of damaged proteins, and by the induction of other antioxidant defenses in 2-CP mutants. (+info)
Antioxidants reversibly inhibit the spontaneous resumption of meiosis.
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We previously showed that the cell-permeant antioxidant 2(3)-tert-butyl-4-hydroxyanisole (BHA) inhibited germinal vesicle breakdown (GVBD) in oocyte-cumulus complexes (OCC) of the rat. The objective of the present studies was to assess other antioxidants and whether such inhibition was reversible. Spontaneous GVBD in OCC incubated for 2 h was significantly inhibited (P < 0.005) by nordihydroguaiaretic acid (NDGA; GVBD = 19.4%), BHA (GVBD = 25.7%), octyl gallate (OG; GVBD = 52.2%), ethoxyquin (EQ; GVBD = 58.8%), 2, 6-di-tert-butyl-hydroxymethyl phenol (TBHMP; GVBD = 59%), butylated hydroxytoluene (BHT; GVBD = 59.5%), and tert-butyl hydroperoxide (TBHP; GVBD = 60.0%). Other antioxidants that produced lower but significant (P < 0.05) inhibition of oocyte maturation included propyl gallate (PG; GVBD = 70.3%), 2,4,5-trihydroxybutrophenone (THBP; GVBD = 71.4%), and lauryl gallate (LG; GVBD = 71.4%). Antioxidants that had no effect on oocyte maturation at the same concentration (100 microM) included ascorbic acid, vitamin E, and Trolox. Inhibition of GVBD was evident for up to 8 h of incubation of OCC and denuded oocytes (DO) with BHA or NDGA and was reversed by washing. NDGA was less potent than BHA for inhibition of GVBD in DO, unlike that seen with OCC. Oocyte maturation was induced by incubation of follicles for 3 h with human chorionic gonadotropin (hCG), and this response was inhibited by BHA or NDGA. These findings support the conclusion that cell-permeant antioxidants inhibit spontaneous resumption of meiosis, which may implicate a role of oxygen radicals in oocyte maturation. (+info)
Abrogation of cyclin D1 expression predisposes lung cancer cells to serum deprivation-induced apoptosis.
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Cyclin D1 antisense (D1AS)-transfected lung epithelial cell lines were serum deprived and then analyzed for three hallmarks of apoptosis: appearance of single-strand DNA breaks, alteration of apoptosis-related protein expression, and induction of chromatin condensation. Single-strand DNA breaks appeared at significant levels 24 h after serum deprivation, whereas induction of chromatin condensation was observed after 72 h. The antioxidants dimethyl sulfoxide, ascorbate, and glutathione, as well as insulin-like growth factor-I, inhibited induction of DNA damage in this assay. Additionally, proliferating cell nuclear antigen expression is completely suppressed in the D1AS cells, indicating a mechanism to explain the reduced capacity for DNA repair. Increased expression of cyclin D1, which is a common lesion in lung cancer, may thus prevent induction of apoptosis in an oxidizing and growth factor-poor environment. Reducing cyclin D1 expression in lung cancer cells by expression of D1AS RNA disrupted these protective pathways. (+info)
BACKGROUND: Obstructive nephropathy leads to progressive renal tubular atrophy and interstitial fibrosis and is associated with sodium wasting and sodium depletion. Renal damage resulting from unilateral ureteral obstruction (UUO) may be aggravated by reactive oxygen species (ROS), which are produced by a variety of processes. Ideally, deleterious effects of ROS are attenuated by antioxidant enzymes, including the superoxide dismutases, glutathione peroxidases, catalase, and glutathione-S-transferases. The general paradigm is that tissue damage occurs when ROS production is greater than the protective capacity of the antioxidant enzymes. METHODS: This study was designed to investigate the response of renal antioxidant enzymes to UUO and sodium depletion. Adult, male Sprague-Dawley rats received normal-sodium or sodium-depleted siets and were subjected to UUO or sham operation. Obstructed (UUO), intact opposite, or sham-operated kidneys were harvested after 14 days, and antioxidant enzyme activities were measured in kidney homogenates. Thiobarbituric acid reactive substances were measured in these homogenates at 3 and 14 days after UUO or sham operation as an index of ROS production. RESULTS: Renal interstitial area, a measure of fibrosis, was increased by UUO and was doubled in sodium-depleted animals. Sodium depletion increased manganese superoxide dismutase, glutathione peroxidases, and glutathione-S-transferase activities in sham-operated kidneys but not in UUO kidneys. Relative to intact opposite kidneys, UUO kidneys had reduced activities of catalase, manganese superoxide dismutase, and glutathione-S-transferase in normal-sodium animals and all antioxidant enzymes tested in sodium-depleted animals. Renal thiobarbituric acid reactive substances were increased by three days of UUO and were increased further by 14 days of sodium depletion. CONCLUSION: In summary, sodium depletion increased several renal antioxidant enzymes, consistent with a stress response to increased ROS production. Further, UUO not only reduced antioxidant enzyme activities but also inhibited increases seen with sodium depletion. We conclude that suppression of renal antioxidant enzyme activities by UUO contributes to the progression of renal injury in obstructive nephropathy, a process exacerbated by sodium depletion. (+info)
Lp(a) and LDL induce apoptosis in human endothelial cells and in rabbit aorta: role of oxidative stress.
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BACKGROUND: Atherogenic lipoproteins cause injury to the vascular wall in the early phase of atherogenesis. We assessed the effects of native (nLDL) and oxidized (oxLDL) low-density lipoprotein (LDL) and lipoprotein (a) [Lp(a)] on O2- formation and cell death in cultured human umbilical vein endothelial cells (HUVECs) and rabbit aorta (RA). METHODS AND RESULTS: O2- formation of HUVECs and RA segments was not influenced by nLDL, but was dose dependently increased by oxLDL and was moderately increased by nLp(a). oxLp(a) was the most potent stimulus for O2- formation, increasing it in HUVECs by 356% at 5 micrograms/ml and in RA by 294% at 100 micrograms/ml. Apoptosis was detected by DNA fragmentation and Annexin assay in HUVECs and by TUNEL staining in RA. Incubation of HUVECs and RA with oxLDL, but not nLDL, dose and time dependently induced apoptosis with only a minimal effect on necrosis. nLp(a) elicited a small but significant effect on apoptosis, whereas oxLp(a) induced apoptosis more potently than oxLDL in HUVECs and RA and caused necrotic cell death in HUVECs. Induction of apoptosis by oxLDL and oxLp(a) in RA was enhanced by the superoxide dismutase (SOD) inhibitor, diethyl-dithio-carbamate, and was blunted by SOD and catalase in HUVECs and RA, suggesting that O2- formation was involved. The concentration of lysophosphatidylcholine, a lipoprotein oxidation product and stimulus for O2- formation, was significantly enhanced by factor 5 in oxLDL and by factor 7 in oxLp(a) compared with native lipoproteins. CONCLUSION: Atherogenic lipoproteins stimulate O2- formation and induction of apoptosis in HUVECs and RA, and may thereby influence the pathogenesis of atherosclerosis. (+info)
Diesel exhaust particles induce NF-kappa B activation in human bronchial epithelial cells in vitro: importance in cytokine transcription.
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Fine particles derived from diesel engines (diesel exhaust particles, DEP) have attracted attention, since their density in industrial countries seems related to the increased prevalence of pulmonary diseases. Previous studies have suggested that DEP have a potential to directly activate airway epithelial cells to produce and release inflammatory cytokines and mediators, and thus facilitate inflammatory responses in the lung. To elucidate the molecular mechanisms of their action, we studied here IL-8 gene expression, one of the important cytokines in inflammatory responses, by Northern blot analysis and run-on transcription assay. Suspended DEP (1-50 microgram/ml) increased the steady state levels of IL-8 mRNA, which was suggested to be largely due to increased transcriptional rates. Electrophoretic mobility shift assay demonstrated that DEP induced increased binding to the specific motif of NF-kappa B, but not of transcription factor AP-1. The luciferase reporter gene assay using wild-type and mutated NF-kappa B-binding sequences showed that DEP-induced NF-kappa B activation was involved in IL-8 transcription. Finally, both N-acetylcysteine and pyrrolidine dithiocarbamate attenuated the action of DEP on IL-8 mRNA expression, suggesting that oxidant-mediated pathway might be involved in its processes. These results suggested that DEP activate NF-kappa B, which might be an important mechanism of its potential to increase the expression of inflammatory cytokines in vitro. (+info)
The antioxidant N-acetylcysteine induces mesangial cells to create three-dimensional cytoarchitecture that underlies cellular differentiation.
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Prolonged culture of mesangial cells produces multifocal nodular structures, i.e., "hillocks," consisting of cells and extracellular matrix. Hillock formation is associated with induction of a differentiated phenotype of mesangial cells, with suppressed mitogenesis and downregulation of alpha-smooth muscle actin (alpha-SMA). Currently, little is understood regarding physiologically relevant factors that facilitate this cytodifferentiation. This study explores whether and how the cellular redox state modulates hillock formation. Exposure of confluent rat mesangial cells to the antioxidant N-acetylcysteine (NAC), an inducer of glutathione, dramatically facilitated hillock formation. This effect was mimicked by external addition of the reduced form of glutathione ethyl ester. In contrast, the oxidizing agents diamide and menadione inhibited the development of hillocks triggered by either NAC, glutathione, or prolonged culture. The induction of hillocks by NAC was correlated with downregulation of alpha-SMA as well as attenuated activity of the CArG box element (the cis-element relevant to the expression of the alpha-SMA gene and growth-associated genes). These results indicate that, by a redox-sensitive mechanism, NAC induces mesangial cells to create three-dimensional cytoarchitecture that underlies cellular differentiation. (+info)