Chronic exposure to TiO2 micro- and nano particles: A biochemical and histopathological experimental study.

The aim of this work was to analyze the effects of long-term exposure to titanium dioxide (TiO2) micro- (MPs) and nanoparticles (NPs) (six and 12 months) on the biochemical and histopathological response of target organs using a murine model. Male Wistar rats were intraperitoneally injected with a suspension of TiO2 NPs (5 nm; TiO2-NP5 group) or MPs (45 µm; TiO2-NP5 group); the control group was injected with saline solution. Six and 12 months post-injection, titanium (Ti) concentration in plasma and target organs was determined spectrometrically (ICP-MS). Blood smears and organ tissue samples were evaluated by light microscopy. Liver and kidney function was evaluated using serum biochemical parameters. Oxidative metabolism was assessed 6 months post-injection (determination of superoxide anion by nitroblue tetrazolium (NBT) test, superoxide dismutase (SOD) and catalase (CAT), lipid peroxidation, and paraoxonase 1). Titanium (Ti) concentration in target organs and plasma was significantly higher in the TiO2-exposed groups than in the control group. Histological evaluation showed the presence of titanium-based particles in the target organs, which displayed no structural alterations, and in blood monocytes. Oxidative metabolism analysis showed that TiO2 NPs were more reactive over time than MPs (p < .05) and mobilization of antioxidant enzymes and membrane damage varied among the studied organs. Clearance of TiO2 micro and nanoparticles differed among the target organs, and lung clearance was more rapid than clearance from the lungs and kidneys (p < .05). Conversely, Ti concentration in plasma increased with time (p < .05). In conclusion, neither serum biochemical parameters nor oxidative metabolism markers appear to be useful as biomarkers of tissue damage in response to TiO2 micro- and nanoparticle deposits at chronic time points.

Decreased immune response in undernourished rats after air pollution exposure.

Children are highly vulnerable subpopulation to malnutrition and air pollution. We investigate, in a rat nutritional growth retardation (NGR) model, the impact of Residual Oil Fly Ash (ROFA) on the lung immune response using in vitro and ex vivo methods. In vitro: Alveolar macrophages (AM) were isolated from Control (C) and NGR animals, cultured and treated with ROFA (1-100 µg/ml) for 24 h. Ex vivo: C and NGR rats were intranasally instilled with ROFA (1 mg/kg BW) or PBS. 24 h post-exposure AM were isolated and cultured. ROFA-treatment increased superoxide anion production and TNFα secretion in C-AM in vitro, though for NGR-AM this response was lower. A similar pattern was observed for TNFα and IL-6 secretion in ex vivo experiments. Regarding the antioxidant response, although NGR-AM showed increased Nrf2, after ROFA instillation an attenuated activation was observed. To conclude, chronic undernutrition altered AM response to ROFA affecting immune responsiveness to air pollutants.

Exfoliated oral mucosa cells as bioindicators of short- and long-term systemic titanium contamination.

BACKGROUND: Humans are exposed to exogenous sources of titanium-containing particles that can enter the body mainly by inhalation, ingestion, or dermal absorption. Given the widespread use of biomaterials in medicine, the surface of a titanium (Ti) biomedical device is a potential endogenous source of Ti ions and/or Ti-containing particles, such as TiO2 micro-(MPs) and nano-particles (NPs), resulting from biotribocorrosion processes. Ti ions or Ti-containing particles may deposit in epithelial cells of the oral mucosa, and the latter may therefore serve as bioindicators of short and long-term systemic Ti contamination. The aim of the present study was to histologically and quantitatively evaluate the presence of Ti traces in cells exfoliated from the oral mucosa as possible bioindicators of systemic contamination with this metal at short and long-term experimental time points METHODS: Thirty Wistar rats were intraperitoneally injected with a suspension of titanium dioxide (TiO2) (0.16 g/100 g body weight of TiO2 in 5 ml of NaCl 0.9%) using 5 nm NPs (Group: TiO2-NP5; n = 10), 45 µm MPs (Group: TiO2-MP45; n = 10), or vehicle alone (Control group; n = 10). At one and six months post-injection, right-cheek mucosa cells were obtained by exfoliative cytology using a cytobrush; they were spray fixed and stained using Safranin or the Papanicolaou technique. The smears were cytologically evaluated (light microscopy) to determine the presence of particulate material, which was also analyzed microchemically (SEM-EDS). Left-cheek mucosa cells were similarly obtained and re-suspended in 5 ml of PBS (pH: 7.2-7.4); the samples corresponding to each group were pooled together and analyzed spectrometrically (ICP-MS) to determine Ti concentration in each of the studied groups. Blood samples were obtained for histological determination of the presence of particulate material on Safranin-stained blood smears and determination of plasma concentration of Ti by ICP-MS RESULTS: Different size and shape metal-like particles were observed inside and outside epithelial cells in TiO2-NP5 and TiO2-MP45 cytological smears at both one and six months post-injection. EDS analysis showed the presence of Ti in the particles. ICP-MS revealed higher Ti concentrations in both TiO2 injected groups compared to the control group. In addition, Ti concentration did not vary with time or particle size. Monocytes containing particles were observed in blood smears of TiO2-exposed animals one- and six-months post-injection. Plasma levels of Ti were significantly higher in TiO2-NP5- and TiO2-MP45- exposed animals than in controls (p < 0.05), and Ti concentration was significantly higher at one month than at six months in both TiO2-exposed groups (p < 0.05). CONCLUSIONS: Cells exfoliated from the oral mucosa could be used as bioindicators of short- and long-term systemic contamination with Ti. Exfoliative cytology could be used as a simple, non-invasive, and inexpensive diagnostic method for monitoring biotribocorrosion of Ti implants and patient clinical follow-up.

Systemic effect of TiO2 micro- and nanoparticles after acute exposure in a murine model.

The surface of a biomedical implant can be a potential endogenous source of release of microparticles (MPs) and nanoparticles (NPs) into the biological environment. In addition, titanium particles from exogenous sources can enter the body through inhalation, ingestion, or dermal contact. The aim of this work was to evaluate the biological response of the lung, liver, and kidneys to acute exposure to titanium dioxide (TiO2 ). Male Wistar rats were intraperitoneally injected with a suspension of 45 µm or 5 nm TiO2 particles. One month post-exposure, titanium concentration was determined spectrometrically (ICP-MS) in plasma and target organs. Blood smears and organ tissue samples were examined histopathologically, and oxidative metabolism was analyzed (superoxide anion by nitro blue tetrazolium (NBT) test; superoxide dismutase (SOD) and catalase (CAT); lipid peroxidation; paraoxonase 1). Liver (aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase) and kidney (urea, creatinine) function was evaluated using serum biochemical markers. Microchemical and histological analysis revealed the presence of particles, though no structural alterations, in TiO2 -exposed groups. NBT test showed an increase in the percentage of reactive cells and antioxidant enzyme consumption in lung samples in the 45 µm and 5 nm TiO2 -exposed groups. Only the 5 nm particles caused a decrease in SOD and CAT activity in the liver. No changes in renal oxidative metabolism were observed in either of the TiO2 -exposed groups. Determination of serum biochemical markers and analysis of oxidative metabolism are not early bioindicators of tissue damage caused by TiO2 MPs and NPs.

Changes in extrapulmonary organs and serum enzyme biomarkers after chronic exposure to Buenos Aires air pollution.

Urban air pollution is a serious environmental problem in developing countries worldwide, and health is a pressing issue in the megacities in Latin America. Buenos Aires is a megacity with an estimated moderate Air Quality Index ranging from 42 to 74 µg/m3. Exposure to Urban Air Particles from Buenos Aires (UAP-BA) induces morphological and physiological respiratory alterations; nevertheless, no studies on extrapulmonary organs have been performed. The aim of the present study was to explore the health effects of chronic exposure to UAP-BA (1, 6, 9, and 12 months) on the liver, heart, and serum risk biomarkers. BALB/c mice were exposed to UAP-BA or filtered air (FA) in inhalation chambers, and liver and heart histopathology, oxidative metabolism (superoxide dismutase, SOD; catalase, CAT; lipoperoxidation, TBARS), amino transaminases (AST, ALT) as serum risk biomarkers, alkaline phosphatase (ALP), paraxonase-1 (PON-1), and lipoprotein-associated phospholipase A2 (Lp-PLA2) were evaluated. Chronic exposure to real levels of UAP in Buenos Aires led to alterations in extrapulmonary organs associated with inflammation and oxidative imbalance and to changes in liver and heart risk biomarkers. Our results may reflect the impact of the persistent air pollution in Buenos Aires on individuals living in this Latin American megacity.

Neurotoxicity mediated by oxidative stress caused by titanium dioxide nanoparticles in human neuroblastoma (SH-SY5Y) cells.

BACKGROUND: Titanium is widely used in biomedicine. Due to biotribocorrosion, titanium dioxide (TiO2) nanoparticles (NPs) can be released from the titanium implant surface, enter the systemic circulation, and migrate to various organs and tissues including the brain. A previous study showed that 5 nm TiO2 NPs reached the highest concentration in the brain. Even though TiO2 NPs are believed to possess low toxicity, little is known about their neurotoxic effects. The aim of the study was to evaluate in vitro the effects of 5 nm TiO2 NPs on a human neuroblastoma (SH-SY5Y) cell line. METHODS: Cell cultures were divided into non-exposed and exposed to TiO2 NPs for 24 h. The following were evaluated: reactive oxygen species (ROS) generation, apoptosis, cellular antioxidant response, endoplasmic reticulum stress and autophagy. RESULTS: Exposure to TiO2 NPs induced ROS generation in a dose dependent manner, with values reaching up to 10 fold those of controls (p < 0.001). Nrf2 nuclear localization and autophagy, also increased in a dose dependent manner. Apoptosis increased by 4- to 10-fold compared to the control group, depending on the dose employed. CONCLUSIONS: Our results show that TiO2 NPs cause ROS increase, induction of ER stress, Nrf2 cytoplasmic translocation to the nucleus and apoptosis. Thus, neuroblastoma cell response to TiO2 NPs may be associated with an imbalance of the oxidative metabolism where endoplasmic reticulum-mediated signal pathway seems to be the main neurotoxic mechanism.

Strong inhibition of replicative DNA synthesis in the developing rat cerebral cortex and glioma cells by roscovitine.

The effects of the cyclin-dependent kinase inhibitors roscovitine and olomoucine on DNA synthesis rate during normal rat brain development were studied by using short time (90 min) incubation. Both purine analogues at 100 microM concentration decreased the DNA synthesis of rat cerebral cortex in an age-dependent manner. The maximum inhibitory effect (approximately 90% for roscovitine, approximately 60% for olomoucine) occurred in rats of 2-13 days postnatal age. In adult rats (> 60 days postnatal age), the effect of both purine analogues was low. Roscovitine even at 200 microM concentration did not inhibit the fraction of DNA synthesis insensitive to hydroxyurea (unscheduled DNA synthesis (UDS)). In addition, in the RG2 rat glioma model, roscovitine produced a strong inhibition of DNA synthesis in glioma cells when compared to adult normal tissue. Since in adult rat brain more than 60% of DNA synthesis is related to DNA repair, usually measured as UDS, our results indicate that roscovitine strongly blocks ongoing DNA synthesis connected with replicative processes.

Characterization and biological effect of Buenos Aires urban air particles on mice lungs.

Exposure to increased levels of ambient air particulate matter (PM) is associated with increased cardiopulmonary morbidity and mortality. Its association with adverse health effects and the still unclear mechanisms of action are of concern worldwide. Our objective was to analyze air PM from downtown Buenos Aires (UAP-BA), and evaluate its biological impact on normal airways. We studied the inflammatory response to intranasal instillation of UAP-BA in a short-term-exposure mouse model. We analyzed UAP-BA morphology by scanning electron microscopy and characterized particle chemical composition by energy dispersive X-ray analysis and capillary gas chromatography. We evaluated lung changes by histomorphometry and histochemical methods. Regarding size, surface area and distribution, UAP-BA proved to be small spherical ultrafine particles: free, in clusters and associated to a matrix. The particles contained polycyclic aromatic hydrocarbons, polychlorinated biphenyls and almost no metal traces. Histologically, UAP-BA induced the recruitment of phagocytes, a reduction in air spaces, an increase in mucous PAS positive cells and weak incomplete elastic fiber network. Our results demonstrate that UAP-BA causes adverse biological effects on the respiratory tract generating inflammation that, in turn, may cause tissue injury or organ dysfunction and may contribute to the pathogenesis of lung diseases.

Age-dependent change in reactive oxygen species and nitric oxide generation by rat alveolar macrophages.

Immunosenescence is an age-associated dysregulation of the immune function, which contributes to increased susceptibility to disease in the elderly. Alveolar macrophages (AM) are known phagocytes that generate reactive oxygen species (ROS) and nitric oxide (NO), essential mediators for host defence. We studied phagocytosis, ROS and NO production in AM obtained from young, adult and senescent rats (1-2, 9-12 and 18-24 months old, respectively) after exposure to lipopolysaccharide (LPS, 0.1-10 microg mL(-1)), 12-O-tetradecanoylphorbol 13-acetate (TPA, 0.1 microg mL(-1)) or LPS + TPA in culture. Phagocytosis was significantly lower in control AM from adult rats than in AM from young animals. Nevertheless, AM from adult animals pretreated with LPS exhibited higher phagocytic capacity than AM from younger animals. ROS was identified by the NBT test at single cell level and quantified by automated image analysis. When TPA was added to all three populations, AM from adult and senescent animals responded more than AM from young animals. All LPS-stimulated AM produce more NO than controls. However, NO production increased three-, four- and two-fold in young, adult and senescent animals, respectively. Our results demonstrate that AM from young, adult and senescent animals display differential responsiveness to inflammatory mediators. Therefore, aging processes markedly affect AM metabolic functions and may further compromise the lung immune defence response, increasing adverse long-term health effects.