Pulmonary toxicity of nanoparticles, influence on the barrier of the pulmonary epithelium

mso-ansi-language:EN-US" lang="EN-US">Nanotechnology is already being used in severalindustries for a very diverse range of products: personal care products,cosmetics, sunscreens, clothing, sporting goods, electronics, food and beverages,paints and coatings. The production of nanoparticles and their marketing willincrease even more in the coming years.mso-ansi-language:EN-US" lang="EN-US">It is generally assumed that the physical and chemicalcharacteristics that define nanoparticles (NPs) unique properties are alsoresponsible for the possible adverse effects. Due to their small size, which isin the range of biological molecules (proteins, lipids, DNA ), NPs mightdistribute to the smallest biological structures, access and interact with finecellular and molecular structures. mso-ansi-language:EN-US" lang="EN-US">Inhalation is considered to be the most importantroute of exposure, but the deleterious effects induced by inhalation of NPs arenot limited to the lungs, since the cardiovascular system may also be affected.Moreover, some human subpopulations may be more susceptible to the deleteriouseffects induced by nanoparticle inhalation, e.g. people withpre-existingcardiovascular disease and the elderly.mso-ansi-language:EN-US" lang="EN-US">The objectives of this project were to:line-height:150%;mso-list:l0 level1 lfo1">font-family:Symbol;mso-fareast-font-family:Symbol;mso-bidi-font-family:Symbol;mso-ansi-language:EN-US" lang="EN-US">· mso-bidi-font-size:11.0pt;line-height:150%;mso-ansi-language:EN-US" lang="EN-US">Optimize acoculture model of the lung-blood barrier to study the adverse pulmonary andextra-pulmonary effects after NP administration (to the pulmonary / apical compartment) line-height:150%;mso-list:l0 level1 lfo1">font-family:Symbol;mso-fareast-font-family:Symbol;mso-bidi-font-family:Symbol;mso-ansi-language:EN-US" lang="EN-US">· mso-bidi-font-size:11.0pt;line-height:150%;mso-ansi-language:EN-US" lang="EN-US">Study theadverse effects after pulmonary nanoparticle administration in two differentanimal models that reflect susceptible human subpopulations.mso-ansi-language:EN-US" lang="EN-US">The most important physical and chemicalcharacteristics of NPs were reviewed in detail (Chapter 1) and links betweenthese characteristics and the observed toxic effects were made where possible.It is generally assumed that the smaller a particle, the more toxic it is. Forother characteristics, a clear relation is less evident. Currently, we also lackclarity as to which characteristics are most important for determiningpotential hazards.mso-ansi-language:EN-US" lang="EN-US">A coculture model of the lung-blood barrier wasdeveloped where human bronchial epithelial cells (16HBE14o-) with monocytes(THP-1) and human microvascular endothelial cells (HLMVEC) were seeded onopposite sides of a permeable membrane, representing the lungs and pulmonarycirculation respectively. The roleof chemically-activated macrophage-likecells in the co-culture was investigated and it was observed that these cellsdisturbed the barrier integrity by disrupting the epithelial tight junctions(Chapter 2). Non-activated monocytes were shown to be activated by SiO2NPs and lipopolysaccharide (LPS), taken as a positive control, withoutdisturbing the barrier integrity. The toxicity of SiO2 and TiO2NPs at non-cytotoxic concentrations was assessed, with LPS and TBHP as positivecontrols for inflammation and oxidative stress (Chapter 3). Both NPs decreasedthe barrier integrity and total glutathione concentrations in the pulmonarycompartment. In a time-course experiment, oxLDL was decreased 24 hours afterexposure to the two types of nanoparticles. SiO2 NPs induced themost pronounced inflammatory response, which resembled that caused by LPS.mso-ansi-language:EN-US" lang="EN-US">Bmal1 (brain and muscle ARNT-like protein-1) knockout mice(Bmal1-/-) have a disturbed circadian rhythm, causing them toprematurely age, 13.0pt;line-height:150%;mso-ansi-language:EN-US" lang="EN-US">and have a mso-ansi-language:EN-US" lang="EN-US">procoagulant 12.0pt;mso-bidi-font-size:13.0pt;line-height:150%;mso-ansi-language:EN-US" lang="EN-US">phenotype(Chapter 4). These mice (Bmal1+/+and Bmal1-/-), togetherwith 18 month old mice (Chapter 5) were usedas animal models reflecting peoplewith pre-existing cardiovascular disease and the elderly, respectively and weresubacutely exposed to MWCNT or ZnO NPs and additionally TiO2 NPswere studied in the old mice. In the Bmal1 mice, tEN-US" lang="EN-US">he MWCNTs and ZnO NPs showed opposite pulmonary toxicity butsimilarprocoagulant effects. Moreover, the observed pulmonary effects correlated withthe hemostatic effects and these correlations were more apparent in the Bmal1-/-mice suggesting it is a sensitive model to study the pulmonary andextra-pulmonary toxicity after pulmonary administration ofNPs. In the oldmice, the MWCNTs and ZnO NPs induced pulmonary inflammation together with acuteprocoagulant effects. Moreover, histological analysis showed the presence ofMWCNTs in the heart and the induction of fibrosis induced by ZnO NPs.mso-ansi-language:EN-US" lang="EN-US"> mso-ansi-language:EN-US" lang="EN-US">In conclusion, we observed that inflammation andoxidative stress were the driving mechanisms in both normal">in vitro and in vivostudies causing the observed toxic effects. Moreover, we demonstrated thatpulmonary NP exposure evoked hemostatic toxicity, providing further evidencethat the lungs and the cardiovascular system are linked. 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How physico-chemical characteristics of nanoparticles cause their toxicity: complex and unresolved interrelations

The increased use of and interest in nanoparticles (NPs) have resulted in an enormous amount of NPs with different compositions and physico-chemical properties. These unique properties not only determine their utility for (bio-medical) applications, but also their toxicity. Recently, "nano-researchers" became aware of the importance of determining the characteristics since they might be predictors of their toxicity. Currently, we face a large set of (non-coordinated) experiments with miscellaneous objectives resulting in a large quantity of available (and often incomplete) data, which hamper the unraveling of the complex interrelated NP characteristics with experimental results. Here, we try to link different critical physico-chemical characteristics separately with toxicity observed in both in vitro and in vivo models.status: publishe

Not All Mouse Strains Respond Equal in a Model of Chemical-Induced Asthma

edition: 1ststatus: publishe

A coculture model of the lung–blood barrier: the role of activated phagocytic cells

We developed a coculture model of the lung–blood barrier using human bronchial epithelial cells(16HBE14o-), monocytes (THP-1) and human lung microvascular endothelial cells (HLMVEC) in which several parameters can be assessed simultaneously. The epithelial and endothelial cells were grown on opposite sides of a microporous membrane. Electron and confocal microscopic pictures show the presence of the cells in their appropriate compartment and both cell types do not show evidence of growing through the pores. Out of three endothelial cell types (EAhy.926, HUVEC and HLMVEC), the last was chosen as the most appropriate cell type, best resembling the pulmonary endothelium and allowing the expression of functional tight junctions in the 16HBE14o- monolayer with sufficiently high transepithelial electrical resistance (TEER) values. Finally, monocytes were added to the apical compartment. PMA-activated macrophages significantly affected barrier integrity (73% TEER reduction compared to control after 24 h) and disrupted the epithelial tight junctions as shown by redistribution of ZO-1 labeling. Alternatively, monocytes could be activated using lipopolysaccharide, at a sub-toxic level int he apical compartment and only induced a small, though significant, reduction in TEER.This coculture system is a representative model of the lung–blood barrier with barrier integrity as the main toxicity endpoint.status: publishe

Toxicity of nanoparticles embedded in paints compared to pristine nanoparticles, in vitro study

The unique physicochemical properties of nanomaterials has led to an increased use in the paint and coating industry. In this study, the in vitro toxicity of three pristine ENPs (TiO2, Ag and SiO2), three aged paints containing ENPs (TiO2, Ag and SiO2) and control paints without ENPs were compared. In a first experiment, cytotoxicity was assessed using a biculture consisting of human bronchial epithelial (16HBE14o-) cells and human monocytic cells (THP-1) to determine subtoxic concentrations. In a second experiment, a new coculture model of the lung-blood barrier consisting of 16HBE14o- cells, THP-1 and human lung microvascular endothelial cells (HLMVEC) was used to study pulmonary and extrapulmonary toxicity. The results show that the pristine TiO2 and Ag ENPs have some cytotoxic effects at relative high dose, while pristine SiO2 ENPs and all aged paints with ENPs and control paints do not. In the complex triculture model of the lung-blood barrier, no considerable changes were observed after exposure to subtoxic concentration of the different pristine ENPs and paint particles. In conclusion, we demonstrated that although pristine ENPs show some toxic effects, no significant toxicological effects were observed when they were embedded in a complex paint matrix.publisher: Elsevier articletitle: Toxicity of nanoparticles embedded in paints compared to pristine nanoparticles, in vitro study journaltitle: Toxicology Letters articlelink: http://dx.doi.org/10.1016/j.toxlet.2014.11.030 content_type: article copyright: Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.status: publishe

Choice of Mouse Strain Influences the Outcome in a Mouse Model of Chemical-Induced Asthma

BACKGROUND: The development of occupational asthma is the result of interactions between environmental factors and individual susceptibility. We assessed how our model of chemical-induced asthma is influenced by using different mouse strains. METHODOLOGY/PRINCIPAL FINDINGS: On days 1 and 8, male mice of 7 different strains (BALB/c, BP/2, A/J, C57Bl/6, DBA/2, CBA and AKR) were dermally treated with toluene-2,4-diisocyanate (TDI) (0.3%) or vehicle (acetone/olive oil, AOO, 2:3) on each ear (20 microl). On day 15, they received an oropharyngeal instillation of TDI (0.01%) or AOO (1:4). Airway reactivity to methacholine, total and differential cell counts in bronchoalveolar lavage (BAL) and total serum IgE and IgG(2a) levels were measured. Lymphocyte subpopulations in auricular lymph nodes and in vitro release of cytokines by ConA stimulated lymphocytes were assessed. In TDI-sensitized and challenged mice, airway hyper-reactivity was only observed in BALB/c, BP/2, A/J and AKR mice; airway inflammation was most pronounced in BALB/c mice; numbers of T-helper (CD4(+)), T-activated (CD4(+)CD25(+)), T-cytotoxic (CD8(+)) and B- lymphocytes (CD19(+)) were increased in the auricular lymph nodes of BALB/c, BP/2, A/J and CBA mice; elevated concentrations of IL-4, IL-10, IL-13 and IFN-gamma were detected in supernatant of lymphocytes from BALB/c, BP/2, A/J, C57Bl/6 and CBA mice cultured with concanavaline A, along with an increase in total serum IgE. CONCLUSION: The used mouse strain has considerable and variable impacts on different aspects of the asthma phenotype. The human phenotypical characteristics of chemically-induced occupational asthma were best reproduced in Th2-biased mice and in particular in BALB/c mice.status: publishe

Pulmonary and hemostatic toxicity of multi-walled carbon nanotubes and zinc oxide nanoparticles after pulmonary exposure in Bmal1 knockout mice

BackgroundPulmonary exposure to nanoparticles (NPs) may affect, in addition to pulmonary toxicity, the cardiovascular system such as procoagulant effects, vascular dysfunction and progression of atherosclerosis. However, only few studies have investigated hemostatic effects after pulmonary exposure.MethodsWe used Bmal1 (brain and muscle ARNT-like protein-1) knockout (Bmal1¿/¿) mice which have a disturbed circadian rhythm and procoagulant phenotype, to study the pulmonary and hemostatic toxicity of multi-walled carbon nanotubes (MWCNTs) and zinc oxide (ZnO) NPs after subacute pulmonary exposure. Bmal1¿/¿ and wild-type (Bmal1+/+) mice were exposed via oropharyngeal aspiration, once a week, during 5 consecutive weeks, to a cumulative dose of 32 or 128 ¿g MWCNTs or 32 or 64 ¿g ZnO NPs.ResultsMWCNTs caused a pronounced inflammatory response in the lung with increased cell counts in the broncho-alveolar lavage and increased secretion of interleukin-1ß and cytokine-induced neutrophil chemo-attractant (KC), oxidative stress (increased ratio of oxidized versus reduced glutathione and decreased total glutathione) as well as anemic and procoagulant effects as evidenced by a decreased prothrombin time with increased fibrinogen concentrations and coagulation factor (F)VII. In contrast, the ZnO NPs seemed to suppress the inflammatory (decreased neutrophils in Bmal1¿/¿ mice) and oxidative response (increased total glutathione in Bmal1¿/¿ mice), but were also procoagulant with a significant increase of FVIII. The procoagulant effects, as well as the significant correlations between the pulmonary endpoints (inflammation and oxidative stress) and hemostasis parameters were more pronounced in Bmal1¿/¿ mice than in Bmal1+/+ mice.ConclusionsThe Bmal1¿/¿ mouse is a sensitive animal model to study the procoagulant effects of engineered NPs. The MWCNTs and ZnO NPs showed different pulmonary toxicity but both NPs induced procoagulant effects, suggesting different mechanisms of affecting hemostasis. However, the correlation analysis suggests a causal association between the observed pulmonary and procoagulant effects.status: publishe

Nanoparticles in the lungs of old mice: Pulmonary inflammation and oxidative stress without procoagulant effects

Pulmonary exposure to nanoparticles (NPs) has been shown to induce pulmonary as well as cardiovascular toxicity. These effects might be enhanced in elderly subjects as a result of a compromised immunity and/or declined organ functions. To study the adverse in vivo effects of NPs in a model for the elderly, we exposed 18-month-old C75Bl/6 mice to multi-walled carbon nanotubes (MWCNTs) or ZnO NPs by intratracheal instillation once a week during 5 consecutive weeks. Pulmonary and hemostatic toxicity was determined 24 h (T1) and 8 weeks (T2) after the last administration. Both NP types significantly increased the pulmonary macrophages at both time points. The MWCNTs and ZnO NPs also induced a pulmonary influx of neutrophils, which was even larger at T2 compared to T1. All NPs induced only a modest increase of pulmonary IL-1β, IL-6 and KC levels. Both types of NPs also increased blood neutrophils. Red blood cells were not significantly affected. Both NPs significantly increased coagulation factor VIII levels at both time points. Histological analysis revealed the presence of MWCNTs in the alveolar macrophages up to 8 weeks after the last administration and the ZnO NPs induced a pronounced alveolar inflammation. In these 18-month-old mice, NPs caused pulmonary inflammation (without evidence of oxidative stress) accompanied by large increases in coagulation factor VIII up to 8 weeks after the last NP exposure. The persistence of the MWCNTs in the lungs resulted in translocation from the lungs to the left heart and the ZnO NPs induced a fibrosis-like pathology.status: publishe