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In an effort to examine signaling pathway of inflammation of the mouse liver caused by intragastric administration of titanium dioxide nanoparticles (NPs), we assessed Toll-like receptor-2 (TLR2), TLR-4, IκB kinase (IKK-α, IKK-ß), IκB nucleic factor-κB (NF-κB), NF-κBP52, NF-κBP65, tumor necrosis factor-α (TNF-α), NF-κB-inducible kinase (NIK), interleukin-2 (IL-2), biochemical parameters of liver functions, and histopathological changes and liver ultrastructure in the TiO(2) NPs-treated mice. The results showed the titanium accumulation in liver, histopathological changes and hepatocytes apoptosis of mice liver, and the liver function damaged by TiO(2) NPs. The real-time quantitative reverse transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay analyses showed that TiO(2) NPs can significantly increase the mRNA and protein expression of TLR2 and TLR4 and several inflammatory cytokines, including IKK1, IKK2, NF-κB, NF-κBP52, NF-κBP65, TNF-α, and NIK, and TiO(2) NPs can significantly decrease the mRNA and protein expression of IκB and IL-2. The results of this study added to our understanding of TiO(2) NPs-induced liver toxicity. It implied that the signaling pathway of liver injury in the TiO(2) NPs-stimulated mouse liver sequentially might occur via activation of TLRs→NIK→IκB kinase→NF-κB→TNF-α→inflammation→apoptosis→liver injury.

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It has been proven that higher dose of lanthanoid (Ln) can induce liver toxicities, but the mechanisms and the molecular pathogenesis are still unclear. In this study, LaCl3, CeCl3, and NdCl3 at a higher dose of 20 mg/kg body weight was injected into the abdominal cavity of ICR mice for 14 consecutive days, and the inflammatory responses of liver of mice were investigated by histopathological test, real-time quantitative reverse transcription polymerase chain reaction (RT-PCR), and enzyme-linked immunosorbent assay (ELISA) methods. The results showed the significant accumulation of Ln in the liver results in liver histopathological changes and, therefore, liver malfunctions. The real-time quantitative RT-PCR and ELISA analyses showed that Ln could significantly alter the mRNA and protein expressions of several inflammatory cytokines, including nucleic factor-κB, macrophage migration inhibitory factor, tumor necrosis factor-α, interleukin-1ß, interleukin-6, cross-reaction protein, interleukin-4, and interleukin-10. Our results also implied that the inflammatory responses and liver injury likely are caused by 4f shell and alterable valence properties of Ln-induced liver toxicity.

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Some recent studies have been previously suggested that nanoparticulate titanium dioxide (TiO(2)) damaged liver function and decreased immunity of mice, but the spleen injury and its oxidative stress mechanism are still unclear. To understand the spleen injury induced by intragastric administration of nanoparticulate anatase TiO(2) for consecutive 30 days, the spleen pathological changes, the oxidative stress, and p38 and c-Jun N-terminal kinase signaling pathways, along with nuclear factor-κB and nuclear factor-E2-related factor-2 (Nrf-2), were investigated as the upstream events of oxidative stress in the mouse spleen from exposure to nanoparticulate TiO(2). The results suggested that nanoparticulate TiO(2) caused congestion and lymph nodule proliferation of spleen tissue, which might exert its toxicity through oxidative stress, as it caused significant increases in the mouse spleen reactive oxygen species accumulations, subsequently leading to the strong lipid peroxidation and the significant expression of heme oxygenase-1 via the p38-Nrf-2 signaling pathway. The studies on the mechanism by which nanoparticulate TiO(2) induced the p38-Nrf-2 signaling pathway are helpful to a better understanding of the nanoparticulate TiO(2)-induced oxidative stress and reduction of immune capacity.

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In order to study the mechanisms underlying the effects of lanthanoid (Ln) on the liver, ICR mice were injected with LaCl3, CeCl3, and NdCl3 at a dose of 20 mg/kg BW into the abdominal cavity daily for 14 days. We then examined oxidative stress-mediated responses in the liver. The increase of lipid peroxide in the liver produced by Ln suggested an oxidative attack that was activated by a reduction of antioxidative defense mechanisms as measured by analyzing the activities of superoxide dismutase, catalase, and ascorbate peroxidase, as well as antioxidant levels such as glutathione and ascorbic acid, which were greatest in Ce(3+) treatment, medium in Nd(3+), and least in La(3+). Our results also implied that the oxidative stress in the liver caused by Ln likely is Ce(3+) > Nd(3+) >La(3+), but the mechanisms need to be further studied in future.

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While the hepatocyte apoptosis induced by TiO(2) nanoparticles (NPs) has been demonstrated, very little is known about the molecular mechanisms underlying this mouse liver apoptosis. In order to understand the hepatocyte apoptosis induced by intragastric administration of TiO(2) NPs for consecutive 60 days, the hepatocyte apoptosis, various oxidative stress parameters and the stress-related gene expression levels were assayed for the mouse liver. 60 days of TiO(2) NPs exposure, hepatocyte apoptosis in the liver could be observed, which was followed by increased reactive oxygen species accumulation, and decreased the stress-related gene expression levels of superoxide dismutase, catalase, glutathione peroxidase, metallothionein, heat shock protein 70, glutathione S transferase, P53, and transferrin; and the significant enhancement of the cytochrome p450 1A expression level. It implied that hepatocyte apoptosis, oxidative stresses, and alteration of expression levels of the genes related with TiO(2) NPs detoxification/metabolism regulation and radical scavenging action. Therefore, the application of TiO(2) NPs and exposure effects especially on human liver for long-term and low-dose treatment should be cautious.

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Titanium dioxide nanoparticles (TiO(2) NPs) are now in daily use including popular sunscreens, toothpastes, and cosmetics. However, the effects of TiO(2) NPs on human body, especially on the central nervous system, are still unclear. The aim of this study was to determine whether TiO(2) NPs exposure results in persistent alternations in nervous system function. ICR mice were exposed to TiO(2) NPs through intragastric administration at 0, 5, 10 and 50 mg/kg body weight every day for 60 days. The Y-maze test showed that TiO(2) NPs exposure could significantly impair the behaviors of spatial recognition memory. To fully investigate the neurotoxicological consequence of TiO(2) NPs exposure, brain elements and neurochemicals were also investigated. The contents of Ca, Mg, Na, K, Fe and Zn in brain were significantly altered after TiO(2) NPs exposure. Moreover, TiO(2) NPs significantly inhibited the activities of Na(+)/K(+)-ATPase, Ca(2+)-ATPase, Ca(2+)/Mg(2+)-ATPase, acetylcholine esterase, and nitric oxide synthase; the function of the central cholinergic system was also noticeably disturbed and the contents of some monoamines neurotransmitters such as norepinephrine, dopamine and its metabolite 3, 4-dihydroxyphenylacetic acid, 5-hydroxytryptamine and its metabolite 5-hydroxyindoleacetic acid were significantly decreased, while the contents of acetylcholine, glutamate, and nitric oxide were significantly increased. These first findings indicated that exposure to TiO(2) NPs could possibly impair the spatial recognition memory ability, and this deficit may be possibly attributed to the disturbance of the homeostasis of trace elements, enzymes and neurotransmitter systems in the mouse brain. Therefore, the application of TiO(2) NPs and exposure effects especially on human brain for long-term and low-dose treatment should be cautious.

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Nanoparticulate titanium dioxide (TiO(2)) has been demonstrated to decrease immunity of mice, but very little is known about the injury of spleen involved immunomodulation and its molecular mechanism. In order to understand the spleen injury induced by intraperitoneal injection of TiO(2) nanoparticules (NPs) for consecutive 45 days, the spleen pathological changes, apoptosis, the expression levels of the apoptotic genes and their proteins, and oxidative stress in the mouse spleen were investigated. The results demonstrated that TiO(2) NPs had obvious accumulation in the mouse spleen, leading to congestion and lymph nodule proliferation of spleen tissue, and splenocyte apoptosis. TiO(2) NPs effectively activated caspase-3 and -9, decreased the Bcl-2 the levels of gene and protein, and increase the levels of Bax, and cytochrome c genes and their protein expression, promoted ROS accumulation. Taken together, this study indicated that TiO(2) NPs-induced apoptosis in the mouse splenocyte via mitochondrial-mediated pathway. These findings provide strong evidence that the TiO(2) NPs can induce the spleen pathological changes, apoptosis, leading to the reduction of immunity of mice.

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In an effort to examine liver injury, immune response, and other physiological effects in mice caused by intragastric administration of nanoparticulate anatase titanium dioxide (5nm), we assessed T lymphocytes, B lymphocyte and NK lymphocyte counts, hematological indices, biochemical parameters of liver functions, and histopathological changes in nanoparticulate titanium dioxide -treated mice. Indeed, mice treated with higher dose nanoparticulate titanium dioxide displayed a reduction in body weight, an increase in coefficients of the liver and histopathological changes in the liver. Specifically, in these nanoparticulate titanium dioxide -treated mice, interleukin-2 activity, white blood cells, red blood cells, haemoglobin, mean corpuscular haemoglobin concentration, thrombocytes, reticulocytes, T lymphocytes (CD3(+), CD4(+), CD8(+)), NK lymphocytes, B lymphocytes, and the ratio of CD4 to CD8 of mice were decreased, whereas NO level, mean corpuscular volume, mean corpuscular haemoglobin, red (cell) distribution width, platelets, hematocrit, mean platelet volume of mice were increased. Furthermore, liver functions were also disrupted, as evidenced by the enhanced activities of alanine aminotransferase, alkaline phosphatase, aspartate aminotransferase, lactate dehydrogenase and cholinesterase, an increase of the total protein, and the reduction of ratio of albumin to globulin, the total bilirubin, triglycerides, and the total cholesterol levels. These results suggested that the liver function damage observed in mice treated with higher dose nanoparticulate titanium dioxide is likely associated with the damage of haemostasis blood system and immune response. However, low dose nanoparticulate anatase TiO(2) has little influences on haemostasis blood system and immune response in mice.

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This study was performed with the objective of assessing the antioxidant response of the lung of mice to different rare earths. LaCl(3), CeCl(3), and NdCl(3) at a higher dose of 20 mg/kg body weight were injected into the nasal cavity of ICR mice for consecutive 14 days, respectively. The increase of pulmonary lipids peroxide produced by Ln suggested an oxidative attack that was activated by a reduction of antioxidative defense mechanisms as measured by analyzing the activities of superoxide dismutase, catalase, ascorbate peroxidase, and total antioxidant capacity, as well as antioxidant levels such as glutathione and ascorbic acid, which were greatest in Ce(3+) treatment, medium in Nd(3+), and least in La(3+). It implied that the antioxidative responses of lung may be involved in 4f shell and alternable valence properties of Ln-induced lung toxicity.

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In order to study the mechanisms underlying the effects of TiO(2) nanoparticles on the brain, ICR mice were injected with nanoparticulate anatase TiO(2) (5 nm) of various doses into the abdominal cavity daily for 14 days. We then examined the coefficient of the brain, the brain pathological changes and oxidative stress-mediated responses, and the accumulation of nanoparticulate anatase TiO(2) and levels of neurochemicals in the brain. The results showed that high-dose nanoparticulate anatase TiO(2) could induce some neurons to turn into filamentous shapes and others into inflammatory cells. The concentration of nanoparticulate anatase TiO(2) in the brain was increased as increases in nanoparticulate anatase TiO(2) dosages used. The oxidative stress and injury of the brain occurred as nanoparticulate anatase TiO(2) appeared to trigger a cascade of reactions such as lipid peroxidation, the decreases of the total anti-oxidation capacity and activities of antioxidative enzymes, the excessive release of nitric oxide, the reduction of glutamic acid, and the downregulated level of acetylcholinesterase activities. We concluded that TiO(2) nanoparticles injected at the abdominal cavity could be translocated into the brain and in turn caused the brain injury.

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In order to study the mechanisms underlying the effects of TiO(2) nanoparticles on lactate dehydrogenase (LDH, EC1.1.1.27), Institute of Cancer Research region mice were injected with nanoparticulate anatase TiO(2) (5 nm) of various doses into the abdominal cavity daily for 14 days. We then examined LDH activity in vivo and in vitro and direct evident for interaction between nanoparticulate anatase TiO(2) and LDH using spectral methods. The results showed that nanoparticulate anatase TiO(2) could significantly activate LDH in vivo and in vitro; the kinetics constant (Km) and Vmax were 0.006 microM and 1,149 unit mg(-1) protein min(-1), respectively, at a low concentration of nanoparticulate anatase TiO(2), and 3.45 and 0.031 microM and 221 unit mg(-1) protein min(-1), respectively, at a high concentration of nanoparticulate anatase TiO(2). By fluorescence spectral assays, the nanoparticulate anatase TiO(2) was determined to be directly bound to LDH, and the binding constants of the binding site were 1.77 x 10(8) L mol(-1) and 2.15 x 10(7) L mol(-1), respectively, and the binding distance between nanoparticulate anatase TiO(2) and the Trp residue of LDH was 4.18 nm, and nanoparticulate anatase TiO(2) induced the protein unfolding. It was concluded that the binding of nanoparticulate anatase TiO(2) altered LDH structure and function.

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The activity of alanine aminotransferase (ALT; E.C. 2.6.1.2) is often changed upon inflammatory responses in animals. Rare earths was shown to provoke various inflammatory responses both in rats and mice; however, the molecular mechanism by which rare earths exert its toxicity has not been completely understood, especially, we know little about the mechanism of the interaction between CeCl(3) and ALT. In this report, we investigated the mechanisms of CeCl(3) on ALT activity in vivo and in vitro. Our results showed that Ce(3+) could significantly activate ALT in vivo and in vitro; the kinetics constant (Km) and Vmax were 0.018 microM and 1,380 unit mg(-1) protein min(-1), respectively, at a low concentration of Ce(3+), and 0.027 microM and 624 unit mg(-1) protein min(-1), respectively, at a high concentration of Ce(3+). By UV absorption and fluorescence spectroscopy assays, the Ce(3+) was determined to be directly bound to ALT; the binding site of Ce(3+) to ALT was 1.72, and the binding constants of the binding site were 4.82 x 10(8) and 9.05 x 10(7) L mol(-1). Based on the analysis of the circular dichroism spectra, it was concluded that the binding of Ce(3+) altered the secondary structure of ALT, suggesting that the observed enhancement of ALT activity was caused by a subtle structural change in the active site through the formation of the complex with Ce(3+).

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Magnesium is one of the essential elements for plant growth and cerium is a beneficial element for plant growth. However, the effects of the fact that cerium improves the nitrogen metabolism of plants under magnesium deficiency is poorly understood. The main aim of the study was to determine the role of cerium in the amelioration of magnesium-deficiency effects in spinach plants. Spinach plants were cultivated in Hoagland's solution. They were subjected to magnesium deficiency and to cerium chloride administered in the magnesium-present media and magnesium-deficient media. Spinach plants grown in the magnesium-present media and magnesium-deficient media were measured for key enzyme activities involved in nitrogen metabolism such as nitrate reductase, nitrite reductase, glutamate dehydrogenase, glutamate synthase, urease, glutamic­pyruvic transaminase, and glutamic­oxaloace protease transaminase. As the nitrogen metabolism in spinach was significantly inhibited by magnesium deficiency, it caused a significant reduction of spinach plant weight, leaf turning chlorosis. However, cerium treatment grown in magnesium-deficiency media significantly promoted the activities of the key enzymes as well as the contents of the free amino acids, chlorophyll, soluble protein, and spinach growth. It implied that Ce3+ could partly substitute for magnesium to facilitate the transformation from inorganic nitrogen to organic nitrogen, leading to the improvement of spinach growth, although the metabolism needs to be investigated further.

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The activity of lactate dehydrogenase (LDH, EC1.1.1.27) is often changed upon inflammatory responses in animals. Lanthanoid (Ln) was shown to provoke various inflammatory responses both in rats and mice; however, the molecular mechanism by which Ln3+ exert its toxicity has not been completely understood, especially that we know little about the mechanism of the interaction between Ln with 4f electron shell and alternation valence and LDH. In this report, we investigated the mechanisms of LaCl3, CeCl3, and NdCl3 on LDH activity in vivo and in vitro. Our results showed that La3+, Ce3+, and Nd3+ could significantly activate LDH in vivo and in vitro; the order of activation was Ce3+>Nd3+> La3+>control. The affinity of LDH for Ce3+ was higher than Nd3+ and La3+; the saturated binding sites for Ce3+ on the LDH protein were 1.2 and for La3+ and Nd3+ 1.55. Ln3+ caused the reduction of exposure degree of cysteine or tryptophan/tyrosine of LDH, the increase of space resistance, and the enhancement of α-helix in secondary structure of LDH, which was greatest in Ce3+ treatment, medium in Nd3+ treatment, and least in La3+ treatment. It implied that the changes of structure-function on LDH caused by Ln3+ were closely related to the characteristics of 4f electron shell and alternation valence in Ln.

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Nano-TiO2 and superoxide dismutase (SOD, EC 1.15.1.1) have been added to cosmetics and used to prevent injury of skin from UV-radiation, which might be related to the decrease of oxidative damage of skin. In previous studies we had proven that nano-anatase could increase the activity of SOD and decrease the oxidative damage in vivo. The mechanisms by which nano-anatase promoted SOD activity, however, are still not clearly understood. In the present work, nano-anatase in various concentrations was added to SOD from rat erythrocytes in vitro to gain insight into the mechanism of molecular interactions between nano-anatase and SOD by various spectral methods, suggesting that the reaction between SOD and nano-anatase was two-order, which meant that the SOD activity was greatly increased by low concentration of nano-anatase and inhibited by high concentration of nano-anatase. The spectroscopic assays suggested that the nano-anatase was determined to directly bind to SOD; the binding site of nano-anatase to SOD was 0.256 and the binding constants were 6.54 x 10(5) and 3.6 x 10(5)Lmol(-1); Ti was bound with three oxygen or nitrogen atoms and a sulfur atoms of amino acid residues at the Ti-O(N) and Ti-S bond lengths of 1.86 and 2.37 A, respectively, the binding nano-anatase entirely altered the secondary structure of SOD. It implied that the nano-anatase coordination created a new metal ion-active site form in SOD, thus leading to an enhancement in SOD activity.

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Lactate dehydrogenase (LDH, EC1.1.1.27), widely expressed in the heart, liver, and other tissues, plays an important role in glycolysis and glyconeogenesis. The activity of LDH is often altered upon inflammatory responses in animals. Nano-TiO(2) was shown to provoke various inflammatory responses both in rats and mice; however, the molecular mechanism by which TiO(2) exerts its toxicity has not been completely understood. In this report, we investigated the mechanisms of nano-anatase TiO(2) (5 nm) on LDH activity in vitro. Our results showed that LDH activity was greatly increased by low concentration of nano-anatase TiO(2), while it was decreased by high concentration of nano-anatase TiO(2). The spectroscopic assays revealed that the nano-anatase TiO(2) particles were directly bound to LDH with mole ratio of [nano-anatase TiO(2)] to [LDH] was 0.12, indicating that each Ti atom was coordinated with five oxygen/nitrogen atoms and a sulfur atoms of amino acid residues with the Ti-O(N) and Ti-S bond lengths of 1.79 and 2.41 A. We postulated that the bound nano-anatase TiO(2) altered the secondary structure of LDH, created a new metal ion-active site for LDH, and thereby enhanced LDH activity.

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Nano-TiO(2) was shown to cause various toxic effects in both rats and mice; however, the molecular mechanism by which TiO(2) exerts its toxicity is poorly understood. In this report, an interaction of nano-anatase TiO(2) with liver DNA from ICR mice was systematically studied in vivo using ICP-MS, various spectral methods and gel electrophoresis. We found that the liver weights of the mice treated with higher amounts of nano-anatase TiO(2) were significantly increased. Nano-anatase TiO(2) could be accumulated in liver DNA by inserting itself into DNA base pairs or binding to DNA nucleotide that bound with three oxygen or nitrogen atoms and two phosphorous atoms of DNA with the Ti-O(N) and Ti-P bond lengths of 1.87 and 2.38 A, respectively, and alter the conformation of DNA. And gel electrophoresis showed that higher dose of nano-anatase TiO(2) could cause liver DNA cleavage in mice.

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Although it is known that nano-TiO2or other nanoparticles can induce liver toxicities, the mechanisms and the molecular pathogenesis are still unclear. In this study, nano-anatase TiO2(5 nm) was injected into the abdominal cavity of ICR mice for consecutive 14 days, and the inflammatory responses of liver of mice was investigated. The results showed the obvious titanium accumulation in liver DNA, histopathological changes and hepatocytes apoptosis of mice liver, and the liver function damaged by higher doses nano-anatase TiO2. The real-time quantitative RT-PCR and ELISA analyses showed that nano-anatase TiO2can significantly alter the mRNA and protein expressions of several inflammatory cytokines, including nucleic factor-κB, macrophage migration inhibitory factor, tumor necrosis factor-α, interleukin-6, interleukin-1ß, cross-reaction protein, interleukin-4, and interleukin-10. Our results also implied that the inflammatory responses and liver injury may be involved in nano-anatase TiO2-induced liver toxicity.