Mucin secretion induced by titanium dioxide nanoparticles

Nanoparticle (NP) exposure has been closely associated with the exacerbation and pathophysiology of many respiratory diseases such as Chronic Obstructive Pulmonary Disease (COPD) and asthma. Mucus hypersecretion and accumulation in the airway are major clinical manifestations commonly found in these diseases. Among a broad spectrum of NPs, titanium dioxide (TiO(2)), one of the PM10 components, is widely utilized in the nanoindustry for manufacturing and processing of various commercial products. Although TiO(2) NPs have been shown to induce cellular nanotoxicity and emphysema-like symptoms, whether TiO(2) NPs can directly induce mucus secretion from airway cells is currently unknown. Herein, we showed that TiO(2) NPs (<75 nm) can directly stimulate mucin secretion from human bronchial ChaGo-K1 epithelial cells via a Ca(2+) signaling mediated pathway. The amount of mucin secreted was quantified with enzyme-linked lectin assay (ELLA). The corresponding changes in cytosolic Ca(2+) concentration were monitored with Rhod-2, a fluorescent Ca(2+) dye. We found that TiO(2) NP-evoked mucin secretion was a function of increasing intracellular Ca(2+) concentration resulting from an extracellular Ca(2+) influx via membrane Ca(2+) channels and cytosolic ER Ca(2+) release. The calcium-induced calcium release (CICR) mechanism played a major role in further amplifying the intracellular Ca(2+) signal and in sustaining a cytosolic Ca(2+) increase. This study provides a potential mechanistic link between airborne NPs and the pathoetiology of pulmonary diseases involving mucus hypersecretion.     

Formation of potential titanium antigens based on protein binding to titanium dioxide nanoparticles

Degradation products of titanium implants include free ions, organo-metallic complexes, and particles, ranging from nano to macro sizes. The biological effects, especially of nanoparticles, is yet unknown. The main objective of this study was to develop Ti-protein antigens in physiological solutions that can be used in testing of cellular responses. For this purpose, 0.1% TiO2 nanoparticles less than 100 nm were mixed with human serum albumin (HSA), 0.1% and 1%, in cell culture medium (DMEM, pH 7.2). The Ti concentrations in the resulting solutions were analyzed by inductively coupled plasma mass spectrometry. The stability of the nanoparticles in suspension was analyzed by UV-vis spectrophotometer and Dynamic Light Scattering. The concentration of Ti in suspension was dependent on the presence and concentration of HSA. Albumin prevented high aggregation rate of TiO2 nanoparticles in cell culture medium. It is shown that nano TiO2-protein stable aggregates can be produced under physiological conditions at high concentrations, and are candidates for use in cellular tests.     

The toxic influence of silver and titanium dioxide nanoparticles on cultured ovarian granulosa cells

The application of metal nanoparticles in modern society is growing, but there is insufficient data concerning their influence on reproductive processes and comparison of their biological activity. The present experiments aimed to compare the effects of silver and titanium dioxide nanoparticles (AgNPs and TiO2NPs) on ovarian granulosa cell functions. AgNPs and TiO2NPs were added to culture of porcine granulosa cells at doses 0, 0.01, 0.1, 1 or 10 μg/mL. The mRNAs for proliferating cell nuclear antigen (PCNA), cyclin B1, bax and caspase 3 were quantified by RT-PCR; release of progesterone was analyzed by ELISA. It was shown that both AgNPs and TiO2NPs significantly reduced all the measured parameters. ED₅₀ of the inhibitory influence of AgNPs on the main ovarian cell parameters was higher than ED₅₀ of TiO2NPs. The ability of AgNPs and TiO2NPs to suppress ovarian granulosa cell functions should be taken into account by their application.     

Toxicity and interaction of titanium dioxide nanoparticles with microtubule protein

Titanium dioxide (TiO2) nanoparticles (NPs) are widely used in several manufactured products. The small size of NPs facilitates their uptake into cells as well as transcytosis across epithelial cells into blood and lymph circulation to reach different sites, such as the central nervous system. Different studies have shown the risks that TiO2 NPs in the neuronal system and other organs present. As membrane-bound layer aggregates or single particles, TiO2 NPs can enter not only cells, but also mitochondria and nuclei. Therefore these particles can interact with cytoplasmic proteins such as microtubules (MTs). MTs are cytoskeletal proteins that are essential in eukaryotic cells for a variety of functions, such as cellular transport, cell motility and mitosis. MTs in neurons are used to transport substances such as neurotransmitters. Single TiO2 NPs in cytoplasm can interact with these proteins and affect their crucial functions in different tissues. In this study, we showed the effects of TiO2 NPs on MT polymerization and structure using ultraviolet spectrophotometer and fluorometry. The fluorescent spectroscopy showed a significant tubulin conformational change in the presence of TiO2 NPs and the ultraviolet spectroscopy results showed that TiO2 NPs affect tubulin polymerization and decrease it. The aim of this study was to find the potential risks that TiO2 NPs pose to human organs and cells.     

Toxic effects of titanium dioxide nanoparticles on microbial activity and metabolic flux

The purpose of this research is to estimate and quantify the toxicity of titanium dioxide (TiO₂) nanoparticles in microorganisms. Nano-sized particles of TiO₂ were more toxic compared to micro-sized particles. Three microorganismal species, Escherichia coli, Bacillus subtilis, and Saccharomyces cerevisiae, were used to test TiO₂ antimicrobial effects. E. coli showed the lowest survival rate (36%), while S. cerevisiae showed the highest survival rate (71%). The antimicrobial effect of TiO₂ was also dependent on ultraviolet ray wavelength. The survival ratio of E. coli was 40% at a 254 nm wavelength and 80% at 365 nm. To observe the effect of TiO₂ on the intracellular metabolism, a metabolic flux analysis and the measurement of in vivo glucose-6-phosphate were performed. G6P concentration in cells exposed to TiO₂ increased, and glycolysis flux was also higher than the controls.     

The effect of cryoprotectant on kangaroo sperm ultrastructure and mitochondrial function

This study examined the effect of cryoprotectants (20% DMSO, a 10% DMSO/10% glycerol mixture, 20% glycerol and 1 M sucrose solution) on kangaroo sperm structure and function, along with the effect of varying concentrations of glycerol on sperm mitochondrial function. Eastern grey kangaroo cauda epididymidal spermatozoa were incubated for 10 min at 35 °C in each cryoprotectant and the plasma membrane integrity (PMI) and motility assessed using light microscopy. The same samples were fixed for TEM and the ultrastructural integrity of the spermatozoa examined. To investigate the effect of glycerol on the kangaroo sperm mitochondrial function, epididymidal spermatozoa were incubated with JC-1 in Tris–citrate media at 35 °C for 20 min in a range of glycerol concentrations (0%, 5%, 10%, 15% and 20%) and the mitochondrial membrane potential (MMP) and plasma membrane integrity determined. As expected, incubation of spermatozoa in 20% glycerol for 10 min resulted in a significant reduction in motility, PMI and ultrastructural integrity. Interestingly, incubation in 20% DMSO resulted in no significant reduction in motility or PMI but a significant loss of structural integrity when compared to the control spermatozoa (0% cryoprotectant). However, 20% DMSO was overall less damaging to sperm ultrastructure than glycerol, a combination of 10% glycerol and 10% DMSO, and sucrose. While all glycerol concentrations had an adverse effect on mitochondrial function, the statistical models presented for the relationship between MMP and glycerol predicted that spermatozoa, when added to 20% glycerol, would lose half of their initial MMP immediately at 35 °C and MMP would halve after 19.4 min at 4 °C. Models for the relationship between PMI and glycerol predicted that spermatozoa would lose half of their initial PMI after 1.8 min at 35 °C and PMI would halve after 21.1 min at 4 °C. These results suggest that if glycerol is to be used as a cryoprotectant for kangaroo spermatozoa then it is best administered at 4 °C and that mitochondrial function is more sensitive to glycerol than PMI. Future research should be directed at investigating strategies that reduce exposure of spermatozoa to glycerol during processing and that test the cryoprotective properties of 20% DMSO for kangaroo spermatozoa.     

Intratracheally administered titanium dioxide or carbon black nanoparticles do not aggravate elastase-induced pulmonary emphysema in rats

ABSTRACT: BACKGROUND: Titanium dioxide (TiO2) and carbon black (CB) nanoparticles (NPs) have biological effects that could aggravate pulmonary emphysema. The aim of this study was to evaluate whether pulmonary administration of TiO2 or CB NPs in rats could induce and/or aggravate elastase-induced emphysema, and to investigate the underlying molecular mechanisms. METHODS: On day 1, Sprague-Dawley rats were intratracheally instilled with 25 U kg1 pancreatic porcine elastase or saline. On day 7, they received an intratracheal instillation of TiO2 or CB (at 100 and 500 mug) dispersed in bovine serum albumin or bovine serum albumin alone. Animals were sacrificed at days 8 or 21, and bronchoalveolar lavage (BAL) cellularity, histological analysis of inflammation and emphysema, and lung mRNA expression of heme oxygenase-1 (HO-1), interleukin-1beta (IL-1beta), macrophage inflammatory protein-2, monocyte chemotactic protein-1, and matrix metalloprotease (MMP)-1, and -12 were measured. In addition, pulmonary MMP-12 expression was also analyzed at the protein level by immunohistochemistry. RESULTS: TiO2 NPs per se did not modify the parameters investigated, but CB NPs increased perivascular/peribronchial infiltration, and macrophage MMP-12 expression, without inducing emphysema. Elastase administration increased BAL cellularity, histological inflammation, HO-1, IL-1beta and macrophage MMP-12 expression and induced emphysema. Exposure to TiO2 NPs did not modify pulmonary responses to elastase, but exposure to CB NPs aggravated elastase-induced histological inflammation without aggravating emphysema. CONCLUSIONS: TiO2 and CB NPs did not aggravate elastase-induced emphysema. However, CB NPs induced histological inflammation and MMP-12 mRNA and protein expression in macrophages.     

Intratracheally administered titanium dioxide or carbon black nanoparticles do not aggravate elastase-induced pulmonary emphysema in rats

Background

The Influenza A H1N1 virus can be transmitted via direct, indirect, and airborne route to non-infected subjects when an infected patient coughs, which expels a number of different sized droplets to the surrounding environment as an aerosol. The objective of the current study was to characterize the human cough aerosol pattern with the aim of developing a standard human cough bioaerosol model for Influenza Pandemic control.

Method

45 healthy non-smokers participated in the open bench study by giving their best effort cough. A laser diffraction system was used to obtain accurate, time-dependent, quantitative measurements of the size and number of droplets expelled by the cough aerosol.

Results

Voluntary coughs generated droplets ranging from 0.1 - 900 microns in size. Droplets of less than one-micron size represent 97% of the total number of measured droplets contained in the cough aerosol. Age, sex, weight, height and corporal mass have no statistically significant effect on the aerosol composition in terms of size and number of droplets.

Conclusions

We have developed a standard human cough aerosol model. We have quantitatively characterized the pattern, size, and number of droplets present in the most important mode of person-to-person transmission of IRD: the cough bioaerosol. Small size droplets (< 1 ??m) predominated the total number of droplets expelled when coughing. The cough aerosol is the single source of direct, indirect and/or airborne transmission of respiratory infections like the Influenza A H1N1 virus.

Study design

Open bench, Observational, Cough, Aerosol study     

Effects of fixatives on function of pulmonary surfactant.

The structure of pulmonary surfactant films remains ill defined. Although plausible film fragments have been imaged by electron microscopy, questions about the significance of the findings and even about the true fixability of surfactant films by the usual fixatives glutaraldehyde (GA), osmium tetroxide (OsO(4)), and uranyl acetate (UA) have not been settled. We exposed functioning natural surfactant films to fixatives within a captive bubble surfactometer and analyzed the effect of fixatives on surfactant function. The capacity of surfactant to reach near-zero minimum surface tension on film compression was barely impaired after exposure to GA or OsO(4). Although neither GA nor OsO(4) prevented the surfactant from forming a surface active film, GA increased the equilibrium surface tension to above 30 mN/m, and both GA and OsO(4) decreased film stability as seen in the slowly rising minimum surface tension from 1 to ~5 mN/m in 10 min. In contrast, the effect of UA seriously impaired surface activity in that both adsorption and minimum surface tension were substantially increased. In conclusion, the fixatives tested in this study are not suitable to fix, i.e., to solidify, surfactant films. Evidently, however, OsO(4) and UA may serve as staining agents.     

Inhibition of pulmonary surfactant function by phospholipases

Previous studies have shown that respiratory failure associated with disorders such as acute pancreatitis correlates well with increased levels of phospholipase A2 (PLA2) in lung lavages and that intratracheal administration of PLA2 generates an acute lung injury. In addition, bacteria such as Pseudomonas have been shown to secrete phospholipase C (PLC). We studied the effects of these phospholipases on pulmonary surfactant activity using a pulsating bubble surfactometer. Concentrations greater than or equal to 0.1 unit/ml PLA2 destroyed surfactant biophysical activity, increasing surface tension at minimum bubble size from less than 1 to 15 mN/m. This surfactant inactivation was predominantly related to the effect of lysophosphatidylcholine on the surface film, although the fatty acids released with higher PLA2 concentrations also had a detrimental effect on surfactant function. Similarly, as little as 0.1 unit PLC increased the surface tension at minimal size of an oscillating bubble from less than 1 to 15 mN/m, an effect that could be mimicked by the addition of dipalmitin to surfactant in the absence of PLC. Moreover, lower, noninhibitory concentrations (0.01 unit/ml) of PLA2 and PLC increased the sensitivity of surfactant to other inhibitory agents, such as albumin. Thus, relatively low concentrations of PLC and PLA2 can cause severe breakdown of surfactant function and may contribute significantly to some forms of lung injury.     

Assessment of titanium dioxide nanoparticles toxicity via oral exposure in mice: effect of dose and particle size

Objective: The aim of the present work is to evaluate the toxicity of titanium dioxide nanoparticles (TiO₂NPs) according to their doses and particle sizes.

Materials and methods: The effect of five days oral administration of TiO₂NPs (21 and 80?nm) with different doses (50, 250 and 500?mg/kg body weight) was assessed in mice via measurement of oxidative stress markers; glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA) and nitric oxide (NO), liver function indices; aspartate and alanine aminotransferases (AST and ALT), chromosomal aberrations and liver histopathological pattern.

Results: The results revealed drastic alterations in all the measured parameters and showed positive correlation with the gradual dose increment. In addition, the smaller particle size of TiO₂NPS (21?nm) had more adverse effect in all the selected biochemical parameters, genetic aberrations and histological investigations.

Conclusions: Toxicity of TiO₂NPs increases in a dose-dependent manner and vice versa with particles size. The evaluated biomarkers are good indicators for TiO₂NPs toxicity. More detailed studies are required before the recommendation of TiO₂NP_S as food additives.     

Effect of artificial surfactant on pulmonary function in preterm and full-term lambs

We hypothesized that agents very different from surfactant may still support lung function. To test this hypothesis, we instilled FC-100, a fluorocarbon, and Tween 20, a detergent, which have higher minimum surface tensions and less hysteresis than surfactant, into 15 full-term and 14 preterm lambs. FC-100 and Tween 20 were as efficient as natural surfactant in improving gas exchange and compliance in preterm lambs with respiratory failure. Dynamic compliance correlated with the equilibrium surface tension of the alveolar wash in both full-term (P less than 0.02) and preterm (P less than 0.008) lambs. Functional residual capacity in full-term and preterm lambs was lower after treatment with the two test agents than with surfactant, findings consistent with qualitative histology. Oxygenation in full-term lambs correlated with mean lung volumes (P less than 0.003), suggesting that the hysteresis and/or low minimum surface tension of surfactant may improve mean lung volume, and hence oxygenation, by maintaining functional residual capacity. The effects of the test agents suggest that agents with biophysical properties different from surfactant may still aid lung expansion.     

Samarium doped titanium dioxide nanoparticles as theranostic agents in radiation therapy

PurposeTitanium dioxide nanoparticles (TiO₂ NPs) have been investigated for their role as radiosensitisers for radiation therapy. The study aims to increase the efficiency of these NPs by synthesising them with samarium.MethodsSamarium-doped TiO₂ NPs (Ti(Sm)O₂ NPs) were synthesised using a solvothermal method. Transmission electron microscopy (TEM), X-ray diffraction (XRD), and energy-dispersive X-ray spectroscopy (EDS) were performed for characterising of the Ti(Sm)O₂ NPs. The intracellular uptake and cytotoxicity were assessed in vitro using A549 and DU145 cancer cell lines. Furthermore, the effect of dose enhancement and generation of reactive oxygen species (ROS) in response to 6 MV X-rays was evaluated. Additionally, the image contrast properties were investigated using computed tomography (CT) images.ResultsThe synthesised Ti(Sm)O₂ NPs were about 13 nm in diameter as determined by TEM. The XRD pattern of Ti(Sm)O₂ NPs was consistent with that of anatase-type TiO₂. EDS confirmed the presence of samarium in the nanoparticles. At 200 μg/ml concentration, no differences in cellular uptake and cytotoxicity were observed between TiO₂ NPs and Ti(Sm)O₂ NPs in both A549 and DU145 cells. However, the combination of Ti(Sm)O₂ NPs and X-rays elicited higher cytotoxic effect and ROS generation in the cells than that with TiO₂ NPs and X-rays. The CT numbers of Ti(Sm)O₂ NPs were systematically higher than that of TiO₂ NPs.ConclusionsThe Ti(Sm)O₂ NPs increased the dose enhancement of MV X-ray beams than that elicited by TiO₂ NPs. Samarium improved the efficiency of TiO₂ NPs as potential radiosensitising agent.     

The impact of titanium dioxide nanoparticles on biological nitrogen removal from wastewater and bacterial community shifts in activated sludge

The potential impact of titanium dioxide nanoparticles (TiO₂ NPs) on nitrogen removal from wastewater in activated sludge was investigated using a sequencing batch reactor. The addition of 2–50 mg L^?1 of TiO₂ NPs did not adversely affect nitrogen removal. However, when the activated sludge was exposed to 100–200 mg L^?1 of TiO₂ NPs, the effluent total nitrogen removal efficiencies were 36.5 % and 20.3 %, respectively, which are markedly lower than the values observed in the control test (80 %). Further studies showed that the decrease in biological nitrogen removal induced by higher concentrations of TiO₂ NPs was due to an inhibitory effect on the de-nitrification process. Denaturing gradient gel electrophoresis profiles showed that 200 mg L^?1 of TiO₂ NPs significantly reduced microbial diversity in the activated sludge. The effect of light on the antibacterial activity of TiO₂ NPs was also investigated, and the results showed that the levels of TiO₂-dependent inhibition of biological nitrogen removal were similar under both dark and light conditions. Additional studies revealed that different TiO₂ concentrations had a significant effect on dehydrogenase activity, and this effect was most likely the result of decreased microbial activity.     

The in vitro effect of pirprofen on the surface ultrastructure and function of human platelets

The in vitro effect of pirprofen (Rengasil), an antiinflammatory agent, on the surface ultrastructure and function of human platelets was examined and compared with that of acetylsalicylic acid (Aspirin, ASA), and diclofenac sodium (Voltaren, DS). Incubation with pirprofen induced formation of long, needle-shaped pseudopodia, a phenomenon observed also after incubation of the cells with DS. In contrast with ASA and DS, pirprofen induced a marked increase in platelet protein synthesizing capacity. The drug decreased the platelet aggregation to a degree similar to that of ASA and DS. The release of platelet factors 3 and 4 and the level of beta-thromboglobulin following incubation with the drug remained unaltered.     

Role of calcium ions the structure and function of pulmonary surfactant

Pulmonary surfactant isolated by centrifugation in buffers containing ions contains at least three different morphologic structures. The presence of one of these, tubular myelin, is dependent on calcium ions, since chelation of the calcium ions causes disruption of this structure. Addition of EDTA also decreases the ability of the surfactant to absorb rapidly to air-water interfaces and lower surface tension. Titration with calcium ions (2.5 or 5 mM) restores rapid surface adsorption and restores the tubular myelin structural forms. Magnesium ions cannot substitute for calcium ions in these processes. The reversibility of structure and function induced by calcium ions and EDTA is also accompanied by reversible isopycnic density shifts probably related to aggregation and disaggregation of the lipid-protein complex with calcium ions and EDTA, respectively.     

The ultrastructure of multilamellar bodies and surfactant in the human lung

Summary Normal tissues from human lungs were dehydrated through Epon 812 resin to retain many of the lipids and carbohydrates in thin section. The three-dimensional structure of the multilamellar body was determined. The paired layer of phospholipid heads (PH) is 36Å thick; the layer of fatty-acid tails (FA) is 31Å, the same as reported previously for non-human primates and rodents. The human multilamellar body is apparently unique: the lamellae of the major focus divide into two or three lamellae; the matrix material of the core is without vesicular bodies and a projection core is present. When compared with those of the rat, human tissues contain a greater number of lamellar foci and fewer lamellae per focus. The presence of a peripheral layer of lamellae, an ever-present external limiting membrane, and the fusion of multilamellar bodies are also characteristic. Tubular myelin surfactant has the same appearance as in other mammals.Multilamellar bodies were observed in direct communication with Golgi vesicles. Their origin from multivesicular bodies and their maturation through secretion and exocytosis were demonstrated.Untransformed multilamellar bodies in the alveolar space demonstrated three periodicities (P): (1) compact regular lamellae, PH = 36Å, FA = 31Å, P = 66Å; (2) compact broad lamellae, PH = 72Å, FA = 22Å, P = 94Å; (3) loose lamellae, PH = 36Å, FA = 36Å, FA = 31Å with a variable interlamellar space.Appreciation is expressed to Nuket Olson and Phil Offenhauser for their technical assistance. Supported by a grant from the American Lung Association     

Exposure to titanium dioxide and other metallic oxide nanoparticles induces cytotoxicity on human neural cells and fibroblasts

The use of titanium dioxide (TiO₂) in various industrial applications (eg, production of paper, plastics, cosmetics, and paints) has been expanding thereby increasing the occupational and other environmental exposure of these nanoparticles to humans and other species. However, the health effects of exposure to TiO₂ nanoparticles have not been systematically assessed even though recent studies suggest that such exposure induces inflammatory responses in lung tissue and cells. Because the effects of such nanoparticles on human neural cells are unknown, we have determined the putative cytotoxic effects of these nanoparticles on human astrocytes-like astrocytoma U87 cells and compared their effects on normal human fibroblasts. We found that TiO₂ micro- and nanoparticles induced cell death on both human cell types in a concentration-related manner. We further noted that zinc oxide (ZnO) nanoparticles were the most effective, TiO₂ nanoparticles the second most effective, and magnesium oxide (MgO) nanoparticles the least effective in inducing cell death in U87 cells. The cell death mechanisms underlying the effects of TiO₂ micro- and nanoparticles on U87 cells include apoptosis, necrosis, and possibly apoptosis-like and necrosis-like cell death types. Thus, our findings may have toxicological and other pathophysiological implications on exposure of humans and other mammalian species to metallic oxide nanoparticles.     

Altered function of pulmonary surfactant in fatty acid lung injury

To determine whether acute fatty acid lung injury impairs pulmonary surfactant function, we studied anesthetized ventilated rabbits given oleic acid (55 mg/kg iv, n = 11) or an equivalent volume of saline (n = 8). Measurements of pulmonary mechanics indicated a decrease in dynamic compliance within 5 min of injury and a decrease in lung volume that was disproportionately large at low pressures, consistent with diminished surfactant activity in vivo. Bronchoalveolar lavage fluid obtained 1 h after injury had significantly increased erythrocytes and total leukocytes, largely polymorphonuclear cells. The phospholipid content and composition of the cell-free fraction had only minor changes from those of controls, but the protein content was increased 35-fold. Measurements of lavage surface activity in vitro showed an increase in average minimum surface tension from 1.3 +/- 0.4 (SE) dyn/cm in controls to 20.2 +/- 3.9 dyn/cm in injured animals. The alterations in static pressure-volume curves and decrease in lavage surface activity suggest a severe alteration of surfactant function in this form of lung injury that occurs despite the presence of normal amounts of surfactant phospholipids.