Ferric Nitrilotriacetate Research Articles

Curcumin attenuates oxidative damage in animals treated with a renal carcinogen, ferric nitrilotriacetate (Fe-NTA): implications for cancer prevention

Curcumin (diferuloylmethane), a biologically active ingredient derived from rhizome of the plant Curcuma longa, has potent anticancer properties as demonstrated in a plethora of human cancer cell lines/animal carcinogenesis model and also acts as a biological response modifier in various disorders. We have reported previously that dietary supplementation of curcumin suppresses renal ornithine decarboxylase (Okazaki et al. Biochim Biophys Acta 1740:357-366, 2005) and enhances activities of antioxidant and phase II metabolizing enzymes in mice (Iqbal et al. Pharmacol Toxicol 92:33-38, 2003) and also inhibits Fe-NTA-induced oxidative injury of lipids and DNA in vitro (Iqbal et al. Teratog Carcinog Mutagen 1:151-160, 2003). This study was designed to examine whether curcumin possess the potential to suppress the oxidative damage caused by kidney-specific carcinogen, Fe-NTA, in animals. In accord with previous report, at 1 h after Fe-NTA treatment (9.0 mg Fe/kg body weight intraperitoneally), a substantial increased formation of 4-hydroxy-2-nonenal (HNE)-modified protein adducts in renal proximal tubules of animals was observed. Likewise, the levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and protein reactive carbonyl, an indicator of protein oxidation, were also increased at 1 h after Fe-NTA treatment in the kidneys of animals. The prophylactic feeding of animals with 1.0% curcumin in diet for 4 weeks completely abolished the formation of (i) HNE-modified protein adducts, (ii) 8-OHdG, and (iii) protein reactive carbonyl in the kidneys of Fe-NTA-treated animals. Taken together, our results suggest that curcumin may afford substantial protection against oxidative damage caused by Fe-NTA, and these protective effects may be mediated via its antioxidant properties. These properties of curcumin strongly suggest that it could be used as a cancer chemopreventive agent.

Influence of inorganic and organic iron compounds on parameters of cell growth and survival in human colon cells

Increased risk for development of colon cancer is associated with red meat intake and iron toxicity is discussed for one underlying mechanism. Anyhow, for iron itself only limited evidence is found. In this study, effects of different iron compounds on proliferation of HT29 carcinoma and LT97 adenoma human colon cells were investigated. After treatment of cells with inorganic (ferrous sulfate: FeSO 4 and ferric nitrilotriacetate: FeNTA) and organic (hemoglobin and hemin) iron sources (24–72 h), number of cells and metabolic activity were measured. Under normal cell culture conditions, neither iron compound elevated cell growth in either cell line with the exception of FeNTA which induced LT97 cell growth significantly. Distinct inhibition of cell proliferation was measured for organic iron. Serum-free incubation of HT29 cells revealed growth promoting properties of iron under deficiency. Even though organic iron, especially hemin, was a potent growth factor, both substances showed also dose-dependent cytotoxic effects. In conclusion, these data emphasize that not iron itself, but merely organic iron may promote carcinogenic events. Since promotion of proliferation was only detectable under deficiency, cytotoxic properties of organic iron may be of more importance in colon carcinogenesis.

Guilandina bonduc L. Possesses Antioxidant Activity and Precludes Ferric Nitrilotriacetate (Fe-NTA) Induced Renal Toxicity and Tumor Promotion Response

Guilandina bonduc Linn. (Fabaceae), a prickly shrub that grows abundantly in tropical and subtropical regions of Southeast Asia, possesses an extensive medicinal value. The present communication reports ethanolic extract of G. bonduc fruit pods to possess potent antioxidant activity and significantly protect against Ferric Nitrilotriacetate (Fe-NTA)-induced renal oxi-dative stress, nephrotoxicity, and tumor promotion response. Intraperitoneal administration of Fe-NTA (9 mg Fe/Kg body weight) to Swiss albino mice depleted renal antioxidant armory (glutathione content and activities of antioxidant enzymes), and caused oxidative stress, induced inflammation (inducing the expression of inducible nitric oxide synthase and cyclooxygenase-2 and increasing the secretion of proinflammatory cytokines). It also induced hyperproliferation in kidney-enhancing ornithine decarboxylase (ODC) activity and [3H]-thymidine incorporation into DNA. Histopathological changes and levels of serum creatinine (SCr) and blood urea nitrogen (BUN) suggested Fe-NTA to afflict substantial damage to the kidney. Prophylactic treatment of mice with G. bonduc extract (100-200 mg /Kg body weight) for seven days not only restored renal antioxidant armory close to normal, but also significantly precluded Fe-NTA associated inflammation and hyperproliferation. Furthermore, pretreatment with extract markedly prevented renal oxidative damage and histopathological alterations, and restored the levels of renal damage markers close to control values. The data obtained in the present study indicate G. bonduc fruit pods to possess a potent antioxidant activity and avert chemically inflicted renal oxidative damage and tumor promotion response.

Iron increases HMOX1 and decreases hepatitis C viral expression in HCV-expressing cells

To investigate effects of iron on oxidative stress, heme oxygenase-1 (HMOX1) and hepatitis C viral (HCV) expression in human hepatoma cells stably expressing HCV proteins. Effects of iron on oxidative stress, HMOX1, and HCV expression were assessed in CON1 cells. Measurements included mRNA by quantitative reverse transcription-polymerase chain reaction, and protein levels by Western blots. Iron, in the form of ferric nitrilotriacetate, increased oxidative stress and up-regulated HMOX1 gene expression. Iron did not affect mRNA or protein levels of Bach1, a repressor of HMOX1. Silencing the up-regulation of HMOX1 nuclear factor-erythroid 2-related factor 2 (Nrf2) by Nrf2-siRNA decreased FeNTA-mediated up-regulation of HMOX1 mRNA levels. These iron effects were completely blocked by deferoxamine (DFO). Iron also significantly decreased levels of HCV core mRNA and protein by 80%-90%, nonstructural 5A mRNA by 90% and protein by about 50% in the Con1 full length HCV replicon cells, whereas DFO increased them. Excess iron up-regulates HMOX1 and down-regulates HCV gene expression in hepatoma cells. This probably mitigates liver injury caused by combined iron overload and HCV infection.

Genome-wide analysis identifies a tumor suppressor role for aminoacylase 1 in iron-induced rat renal cell carcinoma

A growing number of studies indicate a link between oxidative stress and cancer. We previously developed a rat model of renal cell carcinoma (RCC) induced by ferric nitrilotriacetate (Fe-NTA). Here, we performed a genome-wide analysis to study characteristics of genomic alteration and identify putative genes involved in the development of Fe-NTA-induced RCCs. Array-based comparative genomic hybridization analyses revealed a chromosomal loss spanning chromosome 8 in most of the RCCs studied, with a common deletion at 8q31-32, which was confirmed by loss of heterozygosity (LOH) analysis. Studies of gene expression in RCCs or following Fe-NTA treatment revealed globally decreased transcription levels of 34 genes derived from chromosome 8 that are expressed in the kidney. Among them, the aminoacylase 1 (Acy1) gene, which maps to 8q32 and is highly expressed in the kidney, displayed a significantly decreased level of expression in RCCs. Significant amounts of the Acy1 protein were detected in the cytoplasm as well as in the nuclei of renal proximal tubular cells of untreated rats. Transfection of Acy1 into RCC cell lines inhibited proliferation and colony formation on soft agar. An increased number of apoptotic cells were observed following Acy1 transfection. The rat 8q31-32 chromosomal region corresponds to human 3p21.31-24.1, a hot spot where LOH is frequently found in various human cancers. Thus, Fe-NTA-induced renal tumor model is ideal for studying the link between deletions within this region and tumor formation. Our data demonstrate that Acy1 functions as a tumor suppressor in this rat RCC model.

Effects of quercetin and curcumin on ferric nitrilotriacetate (Fe-NTA) induced renal and hepatic damage in rats

Effects of quercetin and curcumin on ferric nitrilotriacetate (Fe-NTA) induced renal and hepatic damage in rats

Protein N-Acylation: H2O2-Mediated Covalent Modification of Protein by Lipid Peroxidation-Derived Saturated Aldehydes

Various lines of evidence indicate that the oxidative modification of protein and the subsequent accumulation of the degenerated proteins have been found in cells and tissues during aging, oxidative stress, and in a variety of pathological states. The critical agents that give rise to this protein degeneration may be represented by aldehydes. Although the covalent modification of proteins by aldehydes alone has been well-studied, the effect of reactive oxygen species, such as H2O2, upon aldehyde modification of the protein has received little attention. We have now established a unique protein modification in which H2O2 and, to a lesser extent, alkyl hydroperoxides mediate the binding of alkanals to the lysine residues of protein to generate structurally unusual N-acylation products. Upon the reaction of a lysine-containing peptide, N(alpha)-benzoylglycyl-lysine, with hexanal in the presence of H2O2, a product containing one molecule of hexanal per peptide was detected. On the basis of the chemical and spectroscopic evidence, the product was identified to be the acylation product, N(epsilon)-hexanoyllysine. H2O2 mediated the N-acylation of the lysine derivative by the saturated aldehydes of 1-6 carbons in length. The H2O2-mediated acylation of the protein was immunochemically confirmed by reaction of the proteins with hexanal in the presence of H2O2. Furthermore, the enhanced N-acylations (N-acetylation and N-hexanoylation) were also observed in the kidney of rats exposed to ferric nitrilotriacetate, a well-characterized inducer of oxidative stress. Mechanistic studies using a phosphonium lysine derivative suggest a Baeyer-Villiger-like reaction proceeding through peroxide addition to the aldehyde Schiff base. These data suggest that the hydroperoxides, including H2O2, might be involved not only in the oxidative modification of protein but also in the covalent binding of the saturated aldehydes to proteins under oxidative stress.

Coordinated induction of Nrf2 target genes protects against iron nitrilotriacetate (FeNTA)-induced nephrotoxicity

The iron chelate, ferric nitrilotriacetate (FeNTA), induces acute proximal tubular necrosis as a consequence of lipid peroxidation and oxidative tissue damage. Chronic exposure of FeNTA leads to a high incidence of renal adenocarcinomas in rodents. NF-E2-related factor 2 (Nrf2) is a transcription factor that is activated by oxidative stress and electrophiles, and regulates the basal and inducible expression of numerous detoxifying and antioxidant genes. To determine the roles of Nrf2 in regulating renal gene expression and protecting against oxidative stress-induced kidney damage, wild-type and Nrf2-null mice were administered FeNTA. Renal Nrf2 protein translocated to the nucleus at 6h after FeNTA treatment. FeNTA increased mRNA levels of Nrf2 target genes, including NQO1, GCLC, GSTpi1/2, Mrp1, 2, and 4 in kidneys from wild-type mice, but not Nrf2-null mice. Protein expression of NQO1, a prototypical Nrf2 target gene, was increased in wild-type mice, with no change in Nrf2-null mice. FeNTA produced more nephrotoxicity in Nrf2-null mice than wild-type mice as indicated by higher serum urea nitrogen and creatinine levels, as more urinary NAG, stronger 4-hydroxynonenal protein adduct staining, and more extensive proximal tubule damage. Furthermore, pretreatment with CDDO-Im, a potent small molecule Nrf2 activator, protected mice against FeNTA-induced renal toxicity. Collectively, these results suggest that activation of Nrf2 protects mouse kidneys from FeNTA-induced oxidative stress damage by coordinately up-regulating the expression of cytoprotective genes.

The prophylactic protective effect of sesamol against ferric–nitrilotriacetate-induced acute renal injury in mice

The aim of this study was to examine the prophylactic protective effects of 3,4-methylenedioxyphenol (sesamol) on ferric–nitrilotriacetate (Fe–NTA)-induced acute renal damage in mice. We induced acute renal injury in mice by treating them with 4 mg/kg of Fe–NTA for 3 h. We used blood biochemistry, creatinine clearance, and histological examinations to assess renal function. With a high-performance chemiluminescence analyzer, we also determined the hydroxyl radical and superoxide anion levels (free radicals) generated. Renal xanthine oxidase activities were also assessed. Sesamol inhibited Fe–NTA-induced acute renal injury, renal lipid peroxidation, the levels of renal hydroxyl radical and superoxide anion generated, and the activity of xanthine oxidase in mice. Therefore, we concluded that sesamol protected mice against Fe–NTA-induced oxidative-stress-associated acute renal injury by at least partially inhibiting the production of reactive oxygen species.

Effect of melatonin, curcumin, quercetin, and resveratrol on acute ferric nitrilotriacetate (Fe-NTA)-induced renal oxidative damage in rats

The influence of melatonin, curcumin, quercetin, and resveratrol pretreatment on ferric nitrilotriacetate (Fe-NTA)-induced oxidative renal damage was studied. Male Wistar rats were treated orally once daily for 3 days with melatonin (10 mg/kg), curcumin (50 mg/kg), quercetin (15 mg/kg), and resveratrol (10 mg/kg). One hour after the last dose of antioxidants, a single dose of Fe-NTA was administered (8 mg of Fe/kg body weight, i.p.) to pre-treated animals. Twenty-four hours after Fe-NTA administration, the lipid peroxidation (LP), reduced glutathione (GSH), catalase (CAT), and glutathione peroxidase (GSH-Px) were estimated in kidney homogenates. Iron, zinc, and copper concentrations were estimated in kidney tissue. Administration of Fe-NTA to rats induced renal LP (170%, P < 0.001) and inhibited catalase (78%, P < 0.05) in the kidney. The oral pretreatment with melatonin, curcumin, quercetin, and resveratrol each one was effective in decreasing the Fe-NTA-induced LP (P < 0.001); however, it did not influence the FeNTA-induced inhibition of renal CAT activity. No changes were found in renal GSH level and GSH-Px activity compared to control animals. The pretreatment with antioxidants did not affect the increase in renal iron content, blood urea nitrogen/creatinine ratio, and relative kidney weight of FeNTA-intoxicated rats. The results indicate that the pretreatment with natural antioxidants, curcumin, melatonin, quercetin, and resveratrol, significantly and equally suppressed lipid peroxidation induced by Fe-NTA but had no effect on other markers of FeNTA nephrotoxicity and iron deposition in kidneys.

Asbestos induces doxorubicin resistance in MM98 mesothelioma cells via HIF-1

Human malignant mesothelioma (HMM), which is strongly related to asbestos exposure, exhibits high resistance to many anticancer drugs. Asbestos fibre deposition in the lung may cause hypoxia and iron chelation at the fibre surface. Hypoxia-inducible factor (HIF)-1alpha, which is upregulated by a decreased availability of oxygen and iron, controls the expression of membrane transporters, such as P-glycoprotein (Pgp), which actively extrude the anticancer drugs. The present study aimed to assess whether asbestos may play a role in the induction of doxorubicin resistance in HMM cells through the activation of HIF-1alpha and an increased expression of Pgp. After 24-h incubation with crocidolite asbestos or with the iron chelator dexrazoxane, or under hypoxia, HMM cells were tested for HIF-1alpha activation, Pgp expression, accumulation of doxorubicin and sensitivity to its toxic effect. Crocidolite, dexrazoxane and hypoxia caused HIF-1alpha activation, Pgp overexpression and increased resistance to doxorubicin accumulation and toxicity. These effects were prevented by the co-incubation with the cell-permeating iron salt ferric nitrilotriacetate, which caused an increase of intracellular iron bioavailability, measured as increased activity of the iron regulatory protein-1. Crocidolite, dexrazoxane and hypoxia induce doxorubicin resistance in human malignant mesothelioma cells by increasing hypoxia-inducible factor-1alpha activity, through an iron-sensitive mechanism.

Neuroprotective actions of deferiprone in cultured cortical neurones and SHSY‐5Y cells

Alzheimer's disease (AD) is a common neurodegenerative disorder, but the initiating molecular processes contributing to neuronal death are not well understood. AD is associated with elevated soluble and aggregated forms of amyloid beta (Abeta) and with oxidative stress. Furthermore, there is increasing evidence for a detrimental role of iron in the pathogenic process. In this context, iron chelation by compounds such as 3-hydroxypyridin-4-one, deferiprone (Ferriprox) may have potential neuroprotective effects. We have evaluated the possible neuroprotective actions of deferiprone against a range of AD-relevant insults including ferric iron, H(2)O(2) and Abeta in primary mouse cortical neurones. We have investigated the possible neuroprotective actions of deferiprone (1, 3, 10, 30 or 100 microM) in primary neuronal cultures following exposure to ferric iron [ferric nitrilotriacetate (FeNTA); 3 and 10 microM], H(2)O(2) (100 microM) or Abeta1-40 (3, 10 and 20 microM). Cultures were treated with deferiprone or vehicle either immediately or up to 6 h after the insult in a 24-well plate format. In order to elucidate a possible neuroprotective action of deferiprone against Parkinson's disease relevant insults another group of experiments were performed in the human neuroblastoma catecholaminergic SHSY-5Y cell line. SHSY-5Y cells were treated with MPP(+) iodide, the active metabolite of the dopaminergic neurotoxin MPTP and the neuroprotective actions of deferiprone evaluated. Cytotoxicity was assessed at 24 h by lactate dehydrogenase release, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium-bromide turnover (FeNTA and hydrogen peroxide) and morphometric analysis of cell viability by Hoechst 33324/propidium iodide (FeNTA, Abeta and MPP(+)) or 6-carboxyfluorescein diacetate and annexin V-Cy3 (Abeta). The present study demonstrates that deferiprone protects against FeNTA, hydrogen peroxide, MPP(+) and Abeta1-40-induced neuronal cell death in vitro, which is consistent with previous in vitro and in vivo studies that have demonstrated similar protection with other iron chelators.

A Possible Role of Nrf2 in Prevention of Renal Oxidative Damage by Ferric Nitrilotriacetate

To ascertain the possible roles of nuclear erythroid 2 p45-related factor 2 (Nrf2), a key transcription factor of phase 2 drug-metabolizing enzymes, in renal cellular defense against oxidative stress, wild-type and Nrf2-knockout -/- mice were treated with ferric nitrilotriacetate (Fe-NTA) at doses of 3 or 6 mg iron/kg body weight. After Fe-NTA treatment, Nrf2 -/- mice consistently showed lower levels of glutathione (GSH) in the kidney at the low dose and the liver at the high dose than the wild-type mice. Gamma-glutamylcysteine ligase (GCL) activity in the kidney and liver of Nrf2-/- mice was also consistently lower than in wild-type mice after the Fe-NTA treatment. Histopathological examination revealed that nephrotoxicity of Fe-NTA, reflected in necrosis of renal tubule epithelial cells following nuclear damage, was more severe in the Nrf2-/- mice than in their wild-type counterparts. Overall, the data suggest that Nrf2 -/- mice are unable to compensate for depletion of renal GSH because of oxidative stress, being more susceptible to Fe-NTA-induced nephrotoxicity. In conclusion, the present study showed that Nrf2 might play an important role in protecting cells from oxidative stress in the kidney through its regulation of antioxidant enzymes.

Royal Jelly Peptides Inhibit Lipid Peroxidation In Vitro and In Vivo

Royal jelly peptides (RJPx) isolated from hydrolysates of water-soluble royal jelly proteins prepared with protease P exhibited significantly stronger hydroxyl radical-scavenging activity (p<0.001), and antioxidant activity against lipid peroxidation (LPO, p<0.001), than did water-soluble royal jelly protein (WSRJP) in vitro. We also investigated the in vivo antioxidant activity of RJPx against ferric nitrilotriacetate (Fe-NTA)-induced LPO. Male Wistar rats were divided into a control group (Group C), an Fe-NTA group (Group Fe), and an Fe-NTA with RJPx group (Group Fe+R). Rats in Group Fe+R were fed RJPx (2 g/kg body weight) daily for 5 wk. Fe-NTA (8 mg Fe/kg body weight) was then intraperitoneally injected, and serum lipid levels were examined 2 h later. Serum total cholesterol (TC) levels were lower (p<0.05) while low-density lipoprotein (LDL) and LPO were significantly higher (p<0.01) in Group Fe than in Group C. TC (p<0.05) and LPO levels (p<0.01) were lower in Group Fe+R than in Group Fe. Our data suggest that RJPx may inhibit LPO both in vitro and in vivo.

Tissue-Targeted In Vivo Gene Transfer Coupled with Histone Deacetylase Inhibitor Depsipeptide (FK228) Enhances Adenoviral Infection in Rat Renal Cancer Allograft Model Systems

Although the adenoviral vector represents an efficient delivery system, hepatotropic accumulation often has detrimental effects on adenoviral vector-mediated cancer therapy. To overcome this disadvantage, we performed in vivo local gene transfer, in combination with the histone deacetylase inhibitor, depsipeptide (FK228), in a rat renal cancer model. Renal cancer cells induced by ferric nitrilotriacetate in ACI rats were used in this study. Adenoviral vectors containing luciferase cDNA were introduced into the tumor-burdened kidney by way of a catheter placed in the renal artery. Subcutaneous tumors were treated by herpes simplex virus thymidine kinase cDNA followed by intraperitoneal ganciclovir. The levels of Coxsackie-adenovirus receptor in various tissue were determined by quantitative reverse transcriptase-polymerase chain reaction. Depsipeptide (1 mg/kg) was intravenously administered 24 hours before adenoviral vector transduction. The catheter-based adenoviral vector delivery enabled strong gene transduction of the tumor-burdened kidney. Moreover, depsipeptide treatment before adenoviral vector injection significantly improved transgene expression at tumor sites. Quantitative reverse transcriptase-polymerase chain reaction analysis showed that depsipeptide increased the expression levels of the Coxsackie-adenovirus receptor in the renal tumor (13-fold), but not in other normal tissues. Furthermore, the use of herpes simplex virus thymidine kinase cDNA-expressing adenoviral vector followed by ganciclovir markedly inhibited the established tumor growth in combination with depsipeptide compared with herpes simplex virus thymidine kinase cDNA alone. The tissue-targeted in vivo gene transfer coupled with depsipeptide significantly enhanced adenoviral infection at tumor sites. Sensitization of tumor cells with depsipeptide can improve the efficacy of adenoviral vector-mediated suicide gene therapy. Thus, application of depsipeptide could be one of the beneficial adjunct for adenoviral vector-mediated cancer gene therapy.

Chronic Oxidative Stress Causes Amplification and Overexpression of ptprz1 Protein Tyrosine Phosphatase to Activate β-Catenin Pathway

Ferric nitrilotriacetate induces oxidative renal tubular damage via Fenton-reaction, which subsequently leads to renal cell carcinoma (RCC) in rodents. Here, we used gene expression microarray and array-based comparative genomic hybridization analyses to find target oncogenes in this model. At the common chromosomal region of amplification (4q22) in rat RCCs, we found ptprz1, a tyrosine phosphatase (also known as protein tyrosine phosphatase zeta or receptor tyrosine phosphatase beta) highly expressed in the RCCs. Analyses revealed genomic amplification up to eightfold. Despite scarcity in the control kidney, the amounts of PTPRZ1 were increased in the kidney after 3 weeks of oxidative stress, and mRNA levels were increased 16 approximately 552-fold in the RCCs. Network analysis of the expression revealed the involvement of the beta-catenin pathway in the RCCs. In the RCCs, dephosphorylated beta-catenin was translocated to nuclei, resulting in the expression of its target genes cyclin D1, c-myc, c-jun, fra-1, and CD44. Furthermore, knockdown of ptprz1 with small interfering RNA (siRNA), in FRCC-001 and FRCC-562 cell lines established from the induced RCCs, decreased the amounts of nuclear beta-catenin and suppressed cellular proliferation concomitant with a decrease in the expression of target genes. These results demonstrate that chronic oxidative stress can induce genomic amplification of ptprz1, activating beta-catenin pathways without the involvement of Wnt signaling for carcinogenesis. Thus, iron-mediated persistent oxidative stress confers an environment for gene amplification.

Protective effects of Phellinus linteus extract against Iron overload‐mediated oxidative stress in cultured rat hepatocytes

Phellinus linteus (PL) mushroom has been reported to possess antioxidant activity. The present study was designed to investigate whether an ethanol extract obtained from PL might ameliorate oxidative stress and enhance antioxidant enzyme activities in primary rat hepatocytes, which were overloaded with iron using ferric nitrilotriacetate (FeNTA) complex. FeNTA enables hepatocytes to accumulate substantially redox-active iron and stimulates the production of injurious hydroxyl radicals, which in turn, initiate oxidative stress-mediated cytotoxicity. The results showed that pretreatment of hepatocytes with PL extract (50, 100 and 200 microg/mL) for 24 h significantly reversed FeNTA-induced cell viability loss, lactate dehydrogenase leakage (LDH), lipid peroxidation (LPO) and protein carbonyl formation in a dose-dependent manner. It was further observed that PL extract produced an inhibitory effect on intracellular reactive oxygen species (ROS) formation caused by FeNTA. Concomitantly, the amount of GSH content and the activities of glutathione reductase (GSH Rd) and glutathione peroxidase (GSH Px) in hepatocytes pretreated with PL extract increased substantially compared with those treated with FeNTA alone. These results suggest that PL may be useful in protecting against FeNTA-induced oxidative damage and also be capable of attenuating cytotoxicity of other oxidants.

Association of microRNA‐34a overexpression with proliferation is cell type‐dependent

Recently Welch et al. reported that microRNA (miRNA)-34a functions as a potential tumor suppressor in neuroblastoma cells (Oncogene 26: 5017-22, 2007). Here, we conversely show that miRNA-34a supports cell proliferation in rat oxidative stress-induced renal carcinogenesis and is overexpressed in various types of human cancers. While searching for genetically unstable chromosomal areas in rat renal carcinogenesis, we found the miRNA-34 family reciprocally overexpressed in chromosomal areas with frequent allelic loss. By in situ hybridization and reverse transcription-polymerase chain reaction, cerebral neurons and Purkinje cells showed the highest expression of a major type, miRNA-34a, followed by a variety of endocrine cells and proliferating cells including germinal center lymphocytes and mouse embryonic fibroblasts and stem cells. In contrast, normal renal tubules, hepatocytes and myocardial cells showed faint expression. After 3 weeks of ferric nitrilotriacetate (Fe-NTA)-induced oxidative stress, regenerating renal proximal tubular cells showed high miRNA-34a expression. All of the Fe-NTA-induced rat renal carcinomas and an array of human cancers (151 positive cases of 177) showed high expression of miRNA-34a. Furthermore, knockdown of miRNA-34a with small interfering RNA significantly suppressed proliferation not only of renal carcinoma cells but also of HeLa and MCF7 cells. These results indicate that miRNA-34a overexpression, an acquired trait during carcinogenesis, supports cell proliferation in the majority of cancers suggesting an unexpected link in the cellular metabolism between cancer and neuronal and/or endocrine cells, which warrants further investigation.

Protective effect of olive oil minor polar components against oxidative damage in rats treated with ferric-nitrilotriacetate

The phenolic fraction of virgin olive oil exerts preventive effects against reactive oxygen species mediated degenerative diseases. To investigate its action as inhibitor of lipid peroxidation in vivo, we treated Wistar rats with olive oil minor polar components (MPC) (25–50 mg/kg bw) prior to the administration of a sublethal dose (15 mg Fe/kg bw) of ferric-nitrilotriacetate (Fe-NTA). Intraperitoneal injection (i.p.) of Fe-NTA lead to increased oxidative stress associated with extensive peroxidation of membrane lipids in plasma, kidney, and liver of treated rats. Fe-NTA treatment induced a significant decrease of the major oxidizable membrane lipids, α-tocopherol, fatty acids and cholesterol, together with an increase of fatty acids hydroperoxides (HP) and 7-ketocholesterol (7-keto). I.p. administration of MPC significantly inhibited fatty acids and cholesterol oxidation, and reduced the levels of HP and 7-keto. In summary, MPC administration protects organs against lipid peroxidation and conserves the endogenous α-tocopherol.

Iron Stimulates Urokinase Plasminogen Activator Expression and Activates NF-kappa B in Human Prostate Cancer Cells

: Urokinase-type plasminogen activator (uPA) on prostate cancer cell surfaces mediates pericellular proteolysis and destruction of extracellular matrix barriers to tumor invasion and metastasis. Increased expression of tumor-associated uPA leads to enhanced tumor dissemination and poor cancer outcomes in men with prostate cancer. Expression of uPA is regulated in part by the oxidant-sensitive transcription factor, NF-kappa B (NF-κB), which is activated by intracellular reactive oxygen intermediates (ROI). This study examined the effect of iron on the production of ROI, activation of NF-κB and expression of uPA in the human prostate cancer cell line, PC-3. Treatment of PC-3 cells with iron in the form of ferric nitrilotriacetate (FeNTA) in the absence of added transferrin resulted in a dose-dependent increase in cellular ferritin content in both the presence and absence of neutralizing antibody to the transferrin receptor. Cellular uptake of iron resulted in stimulation of intracellular ROI production, and increases in uPA mRNA, antigen, and activity. Concurrent treatment with the iron chelator, desferrioxamine (DFO) abrogated these effects, and treatment with DFO alone inhibited constitutive uPA production. Finally, we observed nuclear translocation, and therefore activation of NF-κB in response to iron exposure. We conclude that iron enters PC-3 cells via a non-transferrin dependent pathway and increases uPA expression. Our data indicate that one mechanism by which iron may stimulate uPA production is through the generation of intracellular ROI and activation of NF-κB-mediated signaling pathways.