Gene Expression and Epigenetic Changes in Mice Following Inhalation of Copper(II) Oxide Nanoparticles.

We investigated the transcriptomic response and epigenetic changes in the lungs of mice exposed to inhalation of copper(II) oxide nanoparticles (CuO NPs) (8 × 105 NPs/m3) for periods of 3 days, 2 weeks, 6 weeks, and 3 months. A whole genome transcriptome and miRNA analysis was performed using next generation sequencing. Global DNA methylation was assessed by ELISA. The inhalation resulted in the deregulation of mRNA transcripts: we detected 170, 590, 534, and 1551 differentially expressed transcripts after 3 days, 2 weeks, 6 weeks, and 3 months of inhalation, respectively. Biological processes and pathways affected by inhalation, differed between 3 days exposure (collagen formation) and longer treatments (immune response). Periods of two weeks exposure further induced apoptotic processes, 6 weeks of inhalation affected the cell cycle, and 3 months of treatment impacted the processes related to cell adhesion. The expression of miRNA was not affected by 3 days of inhalation. Prolonged exposure periods modified miRNA levels, although the numbers were relatively low (17, 18, and 38 miRNAs, for periods of 2 weeks, 6 weeks, and 3 months, respectively). Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analysis based on miRNA-mRNA interactions, revealed the deregulation of processes implicated in the immune response and carcinogenesis. Global DNA methylation was not significantly affected in any of the exposure periods. In summary, the inhalation of CuO NPs impacted on both mRNA and miRNA expression. A significant transcriptomic response was already observed after 3 days of exposure. The affected biological processes and pathways indicated the negative impacts on the immune system and potential role in carcinogenesis.

Variability in the Clearance of Lead Oxide Nanoparticles Is Associated with Alteration of Specific Membrane Transporters.

Lead oxide nanoparticles (PbONPs), upon their entry into the lungs via inhalation, induce structural changes in primary and secondary target organs. The fate and ultrastructural localization of PbONPs in organs is known to be dependent on the specific organ. Here, we focused on the differences in the ability to clear the inhaled PbONPs from secondary target organs and on molecular and cellular mechanisms contributing to nanoparticle removal. Mice were exposed to PbONPs in whole-body inhalation chambers. Clearance of ionic lead and PbONPs (Pb/PbONPs) from the lungs and liver was very effective, with the lead being almost completely eliminated from the lungs and the physiological state of the lung tissue conspicuously restored. Kidneys exposed to nanoparticles did not exhibit serious signs of damage; however, LA-ICP-MS uncovered a certain amount of lead located preferentially in the kidney cortex even after a clearance period. The concentration of lead in femurs, as representatives of the axial skeleton, was the highest among studied organs at all designated time points after PbONP exposure, and the clearance ability of lead from the femurs was very low in contrast to other organs. The organ-specific increase of ABC transporters expression (ABCG2 in lungs and ABCC3 in the liver) was observed in exposed animals, suggesting their involvement in removing Pb/PbONPs from tissues. Moreover, the expression of caveolins and clathrin displayed a tissue-specific response to lead exposure. Our results uncovered high variability among the organs in their ability to clear Pb/PbONPs and in the transporters involved in this process.

Six-week inhalation of CdO nanoparticles in mice: The effects on immune response, oxidative stress, antioxidative defense, fibrotic response, and bones.

Due to the growing number of applications of cadmium oxide nanoparticles (CdO NPs), there is a concern about their potential deleterious effects. The objective of our study was to investigate the effect of CdO NPs on the immune response, renal and intestine oxidative stress, blood antioxidant defence, renal fibrotic response, bone density and mineral content. Six-week-old female ICR mice were exposed to CdO NPs for 6 weeks by inhalation (particle size: 9.82 nm, mass concentration: 31.7 µg CdO/m3, total deposited dose: 0.195 µg CdO/g body weight). CdO NPs increased percentage of thymus CD3e+CD8a+ cells and moderately enhanced splenocyte proliferation and production of cytokines and chemokines. CdO NPs elevated pro-fibrotic factors (TGF-ß2, α-SMA and collagen I) in the kidney, and concentrations of AGEs in the intestine. The ratio of GSH and GSSG in blood was slightly reduced. Exposure to CdO NPs resulted in 10-fold higher Cd concentration in tibia bones. No differences were found in bone mass density, mineral content, bone area values, bone concentrations of Ca, P, Mg and Ca/P ratio. Our findings indicate stimulation of immune/inflammatory response, oxidative stress in the intestine, starting fibrotic response in kidneys and accumulation of CdO NPs in bones of mice.

Inhalation of ZnO Nanoparticles: Splice Junction Expression and Alternative Splicing in Mice.

Despite the wide application of nanomaterials, toxicity studies of nanoparticles (NP) are often limited to in vitro cell models, and the biological impact of NP exposure in mammals has not been thoroughly investigated. Zinc oxide (ZnO) NPs are commonly used in various consumer products. To evaluate the effects of the inhalation of ZnO NP in mice, we studied splice junction expression in the lungs as a proxy to gene expression changes analysis. Female ICR mice were treated with 6.46 × 104 and 1.93 × 106 NP/cm3 for 3 days and 3 months, respectively. An analysis of differential expression and alternative splicing events in 298 targets (splice junctions) of 68 genes involved in the processes relevant to the biological effects of ZnO NP was conducted using next-generation sequencing. Three days of exposure resulted in the upregulation of IL-6 and downregulation of BID, GSR, NF-kB2, PTGS2, SLC11A2, and TXNRD1 splice junction expression; 3 months of exposure increased the expression of splice junctions in ALDH3A1, APAF1, BID, CASP3, DHCR7, GCLC, GCLM, GSR, GSS, EHHADH, FAS, HMOX-1, IFNγ, NF-kB1, NQO-1, PTGS1, PTGS2, RAD51, RIPK2, SRXN1, TRAF6, and TXNRD1. Alternative splicing of TRAF6 and TXNRD1 was induced after 3 days of exposure to 1.93 × 106 NP/cm3. In summary, we observed changes of splice junction expression in genes involved in oxidative stress, apoptosis, immune response, inflammation, and DNA repair, as well as the induction of alternative splicing in genes associated with oxidative stress and inflammation. Our data indicate the potential negative biological effects of ZnO NP inhalation.

Determination of short-term changes in levoglucosan and dehydroabietic acid in aerosols with Condensation Growth Unit - Aerosol Counterflow Two-Jets Unit - LC-MS.

Residential areas in urban agglomerations and also in the countryside are often burdened with high concentrations of aerosol in winter, this originating from local combustion sources. Aerosol sources can be identified by a monitoring of organic markers of biomass burning. Abundant markers of biomass and softwood burning are levoglucosan and dehydroabietic acid, respectively. The aim of this research was to develop an analytical method for the determination of levoglucosan and dehydroabietic acid in aerosol over short time periods involving aerosol sampling into liquid samples, quantitative pre-concentration of analytes, and their determination by liquid chromatography - mass spectrometry. A Condensation Growth Unit - Aerosol Counterflow Two-Jets Unit (CGU-ACTJU) sampler was used for the quantitative collection of aerosol directly into water. Dehydroabietic acid was pre-concentrated from the aqueous phase by solid phase extraction (C-18). Afterwards, levoglucosan in water samples was concentrated on a vacuum evaporator. The detection limits of levoglucosan and dehydroabietic acid were 28 ng m-3 and 5.5 ng m-3, respectively. The results obtained by the developed method were compared with an independent determination of both markers in aerosol by means of the sampling of aerosols on a filter and subsequent analysis by GC-MS. The developed method demonstrated sufficient agreement with the independent determination for generated standard aerosol as well as for urban aerosol over an eight-day winter campaign. The presented method allows the monitoring of concentration changes in biomass burning markers in 2-h intervals.

Aerosol sampler for analysis of fine and ultrafine aerosols.

A new aerosol sampler based on the original version of Aerosol Counterflow Two-Jets Unit (ACTJU) is described. The ACTJU collector, connected with a water-based Condensation Growth Unit (CGU) placed upstream of the ACTJU, accomplished the quantitative collection of fine and ultrafine aerosol particles down to a few nanometers in diameter. Condensation of water vapor in the CGU enlarges nanometer sized particles to larger sizes in the supermicrometer range and the formed droplets are then collected into water in the ACTJU collector. The continuous collection of aerosols with the CGU-ACTJU sampler allows for the time-resolved measurement of changes in the concentration of particulate constituents. Coupling of the CGU-ACTJU sampler with on-line detection devices allows in-situ automated analysis of water-soluble aerosol components with high time resolution of 1 s (e.g., FIA detection for nitrite or nitrate) or 1 h (e.g., IC detection with preconcentration step for inorganic anions). Under the optimum conditions (the air flow rate of 10 L min-1 and water flow rate of 1.5 mL min-1), the limit of detection (IC including the preconcentration) for particulate fluoride, chloride, nitrite, nitrate, sulphate and phosphate is 2.53, 6.64, 24.2, 16.8, 0.12 and 5.03 ng m-3, respectively. The apparatus is sufficiently robust for its application at routine monitoring of aerosol composition in real-time.

A portable device for fast analysis of explosives in the environment.

A novel portable device for fast and sensitive analysis of explosives in environmental samples is presented. The developed system consists of miniaturized microcolumn liquid chromatograph, photolytic converter and chemiluminescence detector. The device is able to determine selectively nitramine- and nitroester- and most of nitroaromates-based explosives as well as inorganic nitrates at trace concentrations in water or soil extracts in less than 8 min. The device allows to analyze various environmental samples such as soils or water materials without previous preconcentration. Because of internal power supply, the device ensures 12h of continuous operation. Limits of detection of compounds of interest are in the range of concentrations from 5.0 × 10(-9)M to 8.0 × 10(-5)M for a signal-to-noise ratio of 3. Limits of quantification are in the range of concentrations from 1.7 × 10(-8)M to 2.7 × 10(-4)M for a signal-to-noise ratio of 10. The repeatability of the method (RSD=2.9-5.6%) was determined by repeated injections (n=10) of the standard samples during 4h.