One-pot hydrothermal synthesis of fluorophore-modified cerium oxide nanoparticles.

Cerium oxide nanoparticles (CeO2 NPs), which have powerful antioxidant properties, are promising nanomaterials for the treatment of diseases associated with oxidative stress. The well-developed surface of CeO2 NPs makes them promising for use as a multifunctional system for various biomedical applications. This work demonstrates a simple approach that allows the direct formation of a molecular fluorophore on the surface of CeO2 NPs using a simple one-pot hydrothermal synthesis. Thus, we were able to synthesize CeO2 NPs of ultra-small size ∼2 nm with a narrow distribution, highly stable fluorescence, and a quantum yield of ∼62%. UV-visible transmission studies revealed that the resulting CeO2 NPs exhibited fast autogenerative catalytic reduction. In vitro results showed high biocompatibility of CeO2 NPs; their internalization occurs mainly in the region of cell nuclei. Thus, the resulting NPs have the necessary parameters and can be successfully used in biovisualization and therapy.

An Experimental Model of Proton-Beam-Induced Radiation Dermatitis In Vivo.

Radiation dermatitis (RD) is one of the most common side effects of radiation therapy. However, to date, there is a lack of both specific treatments for RD and validated experimental animal models with the use of various sources of ionizing radiation (IR) applied in clinical practice. The aim of this study was to develop and validate a model of acute RD induced using proton radiation in mice. Acute RD (Grade 2-4) was obtained with doses of 30, 40, and 50 Gy, either with or without depilation. The developed model of RD was characterized by typical histological changes in the skin after irradiation. Moreover, the depilation contributed to a skin histology alteration of the irradiated mice. The assessment of animal vital signs indicated that there was no effect of proton irradiation on the well-being or general condition of the animals. This model can be used to develop effective therapeutic agents and study the pathogenesis of radiation-induced skin toxicity, including that caused by proton irradiation.

Hybrid Polyelectrolyte Capsules Loaded with Gadolinium-Doped Cerium Oxide Nanoparticles as a Biocompatible MRI Agent for Theranostic Applications.

Layer-by-layer (LbL) self-assembled polyelectrolyte capsules have demonstrated their unique advantages and capability in drug delivery applications. These ordered micro/nanostructures are also promising candidates as imaging contrast agents for diagnostic and theranostic applications. Magnetic resonance imaging (MRI), one of the most powerful clinical imaging modalities, is moving forward to the molecular imaging field and requires advanced imaging probes. This paper reports on a new design of MRI-visible LbL capsules, loaded with redox-active gadolinium-doped cerium oxide nanoparticles (CeGdO2-x NPs). CeGdO2-x NPs possess an ultrasmall size, high colloidal stability, and pronounced antioxidant properties. A comprehensive analysis of LbL capsules by TEM, SEM, LCSM, and EDX techniques was carried out. The research demonstrated a high level of biocompatibility and cellular uptake efficiency of CeGdO2-x-loaded capsules by cancer (human osteosarcoma and adenocarcinoma) cells and normal (human mesenchymal stem) cells. The LbL-based delivery platform can also be used for other imaging modalities and theranostic applications.

Metformin mitigates radiation toxicity exerting antioxidant and genoprotective properties.

The antidiabetic drug metformin (MF) exhibits redox-modulating effects in various pathologies associated with oxidative stress and mitigates ionizing radiation-induced toxicity, but the underlying mechanisms remain to be elucidated. Thus, we studied some radiomitigatory effects of MF and explored the possible mechanisms behind them. Highly sensitive luminescence methods and non-competitive enzyme-linked immunosorbent assay (ELISA) were used in in vitro studies, and in vivo the damage to bone marrow cells and its repair were assessed by the micronucleus test. In a solution, MF at concentrations exceeding 0.1 µM effectively intercepts •OH upon X-ray-irradiation, but does not react directly with H2O2. MF accelerates the decomposition of H2O2 catalyzed by copper ions. MF does not affect the radiation-induced formation of H2O2 in the solution of bovine gamma-globulin (BGG), but has a modulating effect on the generation of H2O2 in the solution of bovine serum albumin (BSA). MF at 0.05-1 mM decreases the radiation-induced formation of 8-oxoguanine in a DNA solution depending on the concentration of MF with a maximum at 0.25 mM. MF at doses of 3 mg/kg body weight (bw) and 30 mg/kg bw administered to mice after irradiation, but not before irradiation, reduces the frequency of micronucleus formation in polychromatophilic erythrocytes of mouse red bone marrow. Our work has shown that the radiomitigatory properties of MF are mediated by antioxidant mechanisms of action, possibly including its ability to chelate polyvalent metal ions.

Heavily Gd-Doped Non-Toxic Cerium Oxide Nanoparticles for MRI Labelling of Stem Cells.

Recently, human mesenchymal stem cells (hMSc) have attracted a great deal of attention as potential therapeutic agents in the treatment of socially significant diseases. Despite substantial advances in stem-cell therapy, the biological mechanisms of hMSc action after transplantation remain unclear. The use of magnetic resonance imaging (MRI) as a non-invasive method for tracking stem cells in the body is very important for analysing their distribution in tissues and organs, as well as for ensuring control of their lifetime after injection. Herein, detailed experimental data are reported on the biocompatibility towards hMSc of heavily gadolinium-doped cerium oxide nanoparticles (Ce0.8Gd0.2O2-x) synthesised using two synthetic protocols. The relaxivity of the nanoparticles was measured in a magnetic field range from 1 mT to 16.4 T. The relaxivity values (r1 = 11 ± 1.2 mM-1 s-1 and r1 = 7 ± 1.2 mM-1 s-1 in magnetic fields typical of 1.5 and 3 T MRI scanners, respectively) are considerably higher than those of the commercial Omniscan MRI contrast agent. The low toxicity of gadolinium-doped ceria nanoparticles to hMSc enables their use as an effective theranostic tool with improved MRI-contrasting properties.

Redox-Active Cerium Fluoride Nanoparticles Selectively Modulate Cellular Response against X-ray Irradiation In Vitro.

Ionizing radiation-induced damage in cancer and normal cells leads to apoptosis and cell death, through the intracellular oxidative stress, DNA damage and disorders of their metabolism. Irradiation doses that do not lead to the death of tumor cells can result in the emergence of radioresistant clones of these cells due to the rearrangement of metabolism and the emergence of new mutations, including those in the genes responsible for DNA repair. The search for the substances capable of modulating the functioning of the tumor cell repair system is an urgent task. Here we analyzed the effect of cerium(III) fluoride nanoparticles (CeF3 NPs) on normal (human mesenchymal stem cells-hMSC) and cancer (MCF-7 line) human cells after X-ray radiation. CeF3 NPs effectively prevent the formation of hydrogen peroxide and hydroxyl radicals in an irradiated aqueous solution, showing pronounced antioxidant properties. CeF3 NPs are able to protect hMSC from radiation-induced proliferation arrest, increasing their viability and mitochondrial membrane potential, and, conversely, inducing the cell death of MCF-7 cancer cells, causing radiation-induced mitochondrial hyperpolarization. CeF3 NPs provided a significant decrease in the number of double-strand breaks (DSBs) in hMSC, while in MCF-7 cells the number of γ-H2AX foci dramatically increased in the presence of CeF3 4 h after irradiation. In the presence of CeF3 NPs, there was a tendency to modulate the expression of most analyzed genes associated with the development of intracellular oxidative stress, cell redox status and the DNA-repair system after X-ray irradiation. Cerium-containing nanoparticles are capable of providing selective protection of hMSC from radiation-induced injuries and are considered as a platform for the development of promising clinical radioprotectors.

The Strong Protective Action of Ce3+/F- Combined Treatment on Tooth Enamel and Epithelial Cells.

We studied the toxic effects of cerium and fluoride species on human dental pulp stem cells and epithelial cells of Cercopithecus aethiops as a surrogate for the human oral mucosa. The sequential use of CeCl3 and NH4F solutions in equimolar sub-toxic concentrations enabled the possible toxic effects of individual components to be avoided, ensuring the preservation of the metabolic activity of the cells due to the formation of CeF3 nanoparticles. Cerium fluoride nanoparticles and terbium-doped cerium fluoride nanoparticles exhibited neither cytotoxicity nor genotoxicity to dental pulp stem cells, even at high concentrations (10-4 M). In millimolar concentrations (from 10-5-10-6 M), these nanoparticles significantly increased the expression of genes responsible for the cell cycle, differentiation and proliferation. The formation of cerium fluoride on the surface of the mucous membrane and teeth provided protection against the development of carious lesions, periodontitis, ROS attacks and other inflammatory diseases of the oral cavity. Luminescent CeF3: Tb nanoparticles enabled the visualization of tooth enamel microcracks.

Novel Antioxidant, Deethylated Ethoxyquin, Protects against Carbon Tetrachloride Induced Hepatotoxicity in Rats by Inhibiting NLRP3 Inflammasome Activation and Apoptosis.

Inflammation and an increase in antioxidant responses mediated by oxidative stress play an important role in the pathogenesis of acute liver injury (ALI). We utilized in silico prediction of biological activity spectra for substances (PASS) analysis to estimate the potential biological activity profile of deethylated ethoxyquin (DEQ) and hypothesized that DEQ exhibits antioxidant and anti-inflammatory effects in a rat model of carbon tetrachloride (CCl4)-induced ALI. Our results demonstrate that DEQ improved liver function which was indicated by the reduction of histopathological liver changes. Treatment with DEQ reduced CCl4-induced elevation of gene expression, and the activity of antioxidant enzymes (AEs), as well as the expression of transcription factors Nfe2l2 and Nfkb2. Furthermore, DEQ treatment inhibited apoptosis, downregulated gene expression of pro-inflammatory cytokines (Tnf and Il6), cyclooxygenase 2 (Ptgs2), decreased glutathione (GSH) level and myeloperoxidase (MPO) activity in rats with ALI. Notably, DEQ treatment led to an inhibition of CCl4-induced NLRP3-inflammasome activation which was indicated by the reduced protein expression of IL-1ß, caspase-1, and NLRP3 in the liver. Our data suggest that DEQ has a hepatoprotective effect mediated by redox-homeostasis regulation, NLRP3 inflammasome, and apoptosis inhibition, which makes that compound a promising candidate for future clinical studies.

Diet starch concentration and starch fermentability affect markers of inflammatory response and oxidant status in dairy cows during the early postpartum period.

Our objective was to evaluate the effects of diet starch concentration and starch fermentability on inflammatory response markers and oxidant status during the early postpartum (PP) period and its carryover effects. Fifty-two multiparous Holstein cows were used in a completely randomized block design experiment with a 2 × 2 factorial arrangement of treatments. Treatments were starch concentration and starch fermentability of diets; diets were formulated to 22% (low starch, LS) or 28% (high starch, HS) starch with dry-ground corn (DGC) or high-moisture corn (HMC) as the primary starch source. Treatments were fed from 1 to 23 d PP and then switched to a common diet until 72 d PP to measure carryover (CO) effects. Treatment period (TP) diets were formulated to 22% forage neutral detergent fiber and 17% crude protein. The diet for the CO period was formulated to 20% forage neutral detergent fiber, 17% crude protein, and 29% starch. Coccygeal blood was collected once a week during the TP and every second week during the CO period. Liver and adipose tissue biopsies were performed within 2 d PP and at 20 ± 3 d PP. Blood plasma was analyzed for concentrations of albumin, haptoglobin, reactive oxygen and nitrogen species (RONS), and antioxidant potential (AOP), with lipopolysaccharide-binding protein (LBP) and TNFα evaluated during the TP only. Oxidative stress index (OSi) was calculated as RONS/AOP. Abundance of mRNA from genes involved in inflammation and glucose metabolism in liver and genes involved in lipogenesis in adipose tissue were determined. Data were analyzed separately for the TP and CO periods. During the TP, treatments interacted to affect concentrations of TNFα, haptoglobin, and LBP, with HMC increasing their concentrations for HS (9.38 vs. 7.45 pg/mL, 0.45 vs. 0.37 mg/mL, and 5.94 vs. 4.48 µg/mL, respectively) and decreasing their concentrations for LS (4.76 vs. 12.9 pg/mL, 0.27 vs. 0.41 mg/mL, and 4.30 vs. 5.87 µg/mL, respectively) compared with DGC. Effects of treatments diminished over time for LBP and haptoglobin with no differences by the end of the TP and no main CO effects of treatment for haptoglobin. The opposite treatment interaction was observed for albumin, with HMC tending to decrease its concentration for HS (3.24 vs. 3.34 g/dL) and increase its concentration for LS (3.35 vs. 3.29 g/dL) compared with DGC, with no carryover effect. Feeding DGC increased the OSi during the first week of the TP compared with HMC, with this effect diminishing over time; during the CO period HMC increased OSi for HS and decreased it for LS compared with DGC, with this effect diminishing toward the end of CO. Feeding HMC increased the abundance of genes associated with inflammation and gluconeogenesis in liver for HS and decreased it for LS compared with DGC. Feeding HS increased the mRNA abundance of genes associated with adipose tissue lipogenesis compared with LS. Results during the TP suggest that feeding LS-DGC and HS-HMC elicited a more pronounced inflammatory response and induced an upregulation of genes associated with inflammation and gluconeogenesis in liver, without effects on OSi, but effects on plasma markers of inflammation diminished during the CO period.

Intracellular Delivery of Antioxidant CeO2 Nanoparticles via Polyelectrolyte Microcapsules.

Cerium oxide nanoparticles (nanoceria) are regarded as one of the most promising inorganic antioxidants for biomedical applications. Considering nanoceria as a potential therapeutic agent, we aimed to develop a robust system for its intracellular delivery using layer-by-layer polyelectrolyte microcapsules. We have shown that citrate-stabilized cerium oxide nanoparticles can be effectively incorporated into the structure of polyelectrolyte microcapsules made from biodegradable and nonbiodegradable polymers. The structure and morphology of synthesized microcapsules were investigated and analyzed using confocal laser scanning microscopy, scanning electron microscopy, transmission electron microscopy, energy-dispersive X-ray spectroscopy, and UV/vis spectroscopy. Results of experiments in vitro on B50 neuroblastoma cells confirmed nanoceria delivery into the cell while maintaining their antioxidant properties. The results presented confirm polyelectrolyte microcapsules to be an efficient intracellular delivery system for therapeutic nanoparticles.

Protection of mice against X-ray injuries by the post-irradiation administration of inosine-5'-monophosphate.

The aim of the present study was to investigate the radiation modulating properties of inosine-5'-monophosphate (IMP). Mice injected introperitoneally (i.p.) with IMP 15 minutes after irradiation with a lethal irradiation dose of 7 Gy have better survival rates comparative to irradiated mice non treated with IMP. The dose reduction factor of the IMP is 1.22. Using a hematologdical test we demonstrated that administration of IMP alleviates the symptoms of radiation-induced leukopenia and thrombocytopenia. The DNA damage in bone marrow and thymus cells of irradiated mice was measured by flow cytofluorometry and micronucleus test (MN-test). The tests show that i.p. administration of IMP to irradiated animals leads to a significant reduction of the DNA damage level. In this paper we show that IMP substantially modulates the damaging effects of ionizing radiation protecting irradiated mice and it is a promising agent for a treatment of leukopenia.

Ectopic Agouti protein overexpression increases stimulated corticosterone production without effect on adenylate cyclase activity in mouse adrenal cells.

OBJECTIVE: The antagonism of Agouti protein (AP) and Agouti-related protein on melanocortin receptors suggests an inhibitory role in the regulation of steroidogenesis. However, we have previously demonstrated that ectopic AP overexpression increased restraint-induced corticosterone release and adrenal reactivity to ACTH in mice. A high steroidogenic response to ACTH may be a consequence of a stimulatory AP action on the adenylate cyclase (AC) and/or intracellular steroidogenic enzymes. The aim of the present study was to estimate the effect of ectopic AP overexpression on the activity of AC and steroidogenic intracellular enzymes. METHODS: ACTH and forskolin were used for AC stimulation, and dibutyryl cAMP and progesterone were used for stimulation of intracellular steroidogenic enzymes in isolated adrenal cells in male C57Bl/6J mice of two Agouti genotypes: A(y)/a (ectopic AP overexpression) and a/a (absence of AP in all tissues). RESULTS: ACTH and forskolin increased cAMP accumulation to the same extent in both A(y)/a and a/a mouse adrenal cells (P<0.001; ANOVA), but resulted in higher corticosterone production in A(y)/a mice (P<0.001 for ACTH and P<0.01 for forskolin; ANOVA). Dibutyryl cAMP- and progesterone-induced corticosterone production was higher in A(y)/a mice than in a/a mice (P<0.001 for dibutyryl cAMP and P<0.01 for progesterone; ANOVA). CONCLUSIONS: Ectopic AP overexpression increased stimulated corticosterone production and intracellular steroidogenic enzyme reactivity to cAMP without an effect on AC activity.

Agouti yellow mutation increases adrenal response to ACTH in mice.

OBJECTIVE: Agouti protein (AP) and agouti-related protein with a similar sequence and action are endogenous antagonists of melanocortin receptors, implicated in the control of the hypothalamo-pituitary-adrenal (HPA) axis. Dominant mutation of the agouti gene (agouti yellow (A(y))) in heterozygous A(y)/a mice leads to ectopic overexpression of AP and produces an obese phenotype. The existing data on the HPA function in A(y)/a-mice are equivocal; therefore, the present study aimed to assess HPA function in 3-month-old male C57Bl/6J mice of two agouti genotypes: A(y)/a (ectopic AP overexpression) and a/a (absence of AP). DESIGN AND METHODS: In order to evaluate the HPA function, activating (15-min restriction, ACTH-induced corticosterone production in vitro) and inhibiting (i.p. injection of dexamethasone, 0.02 microg/g body weight) stimuli were employed. To estimate the effect of obesity on some HPA functions, A(y)/a males were subdivided into obese and non-obese groups. RESULTS: Basal plasma concentrations of ACTH and corticosterone; basal corticosterone production in vitro; and feedback inhibition of resting corticosterone levels by dexamethasone were similar in A(y)/a- and a/a-mice. Restraint-induced plasma corticosterone was greater in obese and non-obese A(y)/a-mice than in a/a-mice, whereas restraint-induced plasma ACTH levels were similar. Adrenal cell responses to ACTH (10(-13)-10(-10) M) were higher in obese and non-obese A(y)/a-mice than in a/a-mice. Dexamethasone, injected 3 h prior to stress, inhibited stress-induced corticosterone levels by a significantly greater amount in A(y)/a-mice than in a/a-mice. CONCLUSIONS: AP may have both stimulating and inhibiting influences on the HPA axis. AP overproduction increased the response of the HPA to short-restraint stress due to increased adrenal responsiveness to ACTH; this result was not effected by obesity development.