Jean-Pierre Kaiser

A comparison of acute and long-term effects of industrial multiwalled carbon nanotubes on human lung and immune cells in vitro

The close resemblance of carbon nanotubes to asbestos fibers regarding their high aspect ratio, biopersistence and reactivity increases public concerns on the widespread use of these materials. The purpose of this study was not only to address the acute adverse effects of industrially produced multiwalled carbon nanotubes (MWCNTs) on human lung and immune cells in vitro but also to further understand if their accumulation and biopersistence leads to long-term consequences or induces adaptive changes in these cells. In contrast to asbestos fibers, pristine MWCNTs did not induce overt cell death in A549 lung epithelial cells and Jurkat T lymphocytes after acute exposure to high doses of this material (up to 30 μg/ml). Nevertheless, very high levels of reactive oxygen species (ROS) and decreased metabolic activity were observed which might affect long-term viability of these cells. However, the continuous presence of low amounts of MWCNTs (0.5 μg/ml) for 6 months did not have major adverse long-term effects although large amounts of nanotubes accumulated at least in A549 cells. Moreover, MWCNTs did not appear to induce adaptive mechanisms against particle stress in long-term treated A549 cells. Our study demonstrates that despite the high potential for ROS formation, pristine MWCNTs can accumulate and persist within cells without having major long-term consequences or inducing adaptive mechanisms.

Contamination of nanoparticles by endotoxin: evaluation of different test methods

BackgroundNanomaterials can be contaminated with endotoxin (lipopolysaccharides, LPS) during production or handling. In this study, we searched for a convenient in vitro method to evaluate endotoxin contamination in nanoparticle samples. We assessed the reliability of the commonly used limulus amebocyte lysate (LAL) assay and an alternative method based on toll-like receptor (TLR) 4 reporter cells when applied with particles (TiO2, Ag, CaCO3 and SiO2), or after extraction of the endotoxin as described in the ISO norm 29701.ResultsOur results indicate that the gel clot LAL assay is easily disturbed in the presence of nanoparticles; and that the endotoxin extraction protocol is not suitable at high particle concentrations. The chromogenic-based LAL endotoxin detection systems (chromogenic LAL assay and Endosafe-PTS), and the TLR4 reporter cells were not significantly perturbed.ConclusionWe demonstrated that nanoparticles can interfere with endotoxin detection systems indicating that a convenient test method must be chosen before assessing endotoxin contamination in nanoparticle samples.

Is nanotechnology revolutionizing the paint and lacquer industry? A critical opinion.

Many paints for indoor and outdoor applications contain biocides and additives for protection against microbial, physical and chemical deterioration. The biocides should remain active as long as they are incorporated in the paint. Protection against microbial colonization should last at least a decade. Once the biocides are released they should degrade within a short time so that no accumulation in the environment can occur. The paint industry is not only focusing their research in producing better paint formulations with degradable biocides: they also consider using nanomaterials, such as nanosilver, nanocopper, nanozinc oxide, photocatalytic-active nanotitanium dioxide and nanosilica dioxide as additives for the protection of paints, against microbial degradation and physical and chemical deterioration. In the future nanomaterials should replace biodegradable biocides and improve the paint properties as well as impede colonization by microorganisms. At the time there is no guarantee that the nanomaterials in paints and façades will fulfill their task in the long run, since there are no long term studies available. From nanosilver doped paints it is known that silver is easily washed out by rain. Photocatalytic active nanotitanium dioxide adsorbs ultra violet light (UV-light) and generates hydroxyl radicals, which not only inhibit microbial growth but can also initiate or accelerate the photocatalytic degradation of the paint matrix. Thus at this time it is still unknown if it makes sense to incorporate nanomaterials into paints. Intensive research and development are still needed in order to find the answers.

Comparability of in Vitro Tests for Bioactive Nanoparticles: A Common Assay to Detect Reactive Oxygen Species as an Example

The release of reactive oxygen species (ROS) during the electron transport of mitochondrial aerobic respiration is the major source of ROS. However, contact between cells and nanoparticles (NPs) can also induce release of ROS, leading to an imbalance towards the pro-oxidative state. At low levels of ROS production, cells initiate a protective response to guarantee their survival, but an excess of ROS can damage cellular compounds such as membranes and various organelles, or directly cause genotoxicity. Thus an elevated level of ROS is an important indicator of cellular stress and an accurate recording of this parameter would be very informative. ROS can be measured by various assays, but all known assays measuring and quantifying ROS possess certain weaknesses. The problems and challenges of quantitatively detecting ROS in vitro using the 2′,7′-dichlorodihydrofluorescein (DCF) assay is discussed as an example. In addition, we debate the difficulties in finding a suitable and stable chemical reaction control for the DCF assay (or other ROS-detecting assays). As a conclusion, we believe that using 3-morpholinosydnonimine hydrochloride (Sin-1) as a ROS inducer in the DCF assay is feasible only qualitatively. However, a quantitative measurement of the absolute amount of ROS produced and a quantitative comparison between experiments is (at the moment) impossible.

Surface grafting of a thermoplastic polyurethane with methacrylic acid by previous plasma surface activation and by ultraviolet irradiation to reduce cell adhesion

The material performance, in a biological environment, is mainly mediated by its surface properties and by the combination of chemical, physical, biological, and mechanical properties required, for a specific application. In this study, the surface of a thermoplastic polyurethane (TPU) material (Elastollan(®)1180A50) was activated either by plasma or by ultra-violet (UV) irradiation. After surface activation, methacrylic acid (MAA) was linked to the surface of TPU in order to improve its reactivity and to reduce cell adhesion. Grafted surfaces were evaluated by X-ray photoelectron spectroscopy (XPS), by atomic force microscopy (AFM) and by contact angle measurements. Blood compatibility studies and cell adhesion tests with human bone marrow cells (HBMC) were also performed. If was found that UV grafting method led to better results than the plasma activation method, since cell adhesion was reduced when methacrylic acid was grafted to the TPU surface by UV.

Human Health Risk of Ingested Nanoparticles That Are Added as Multifunctional Agents to Paints: an In Vitro Study

Microorganisms growing on painted surfaces are not only an aesthetic problem, but also actively contribute to the weathering and deterioration of materials. A widely used strategy to combat microbial colonization is the addition of biocides to the paint. However, ecotoxic, non-degradable biocides with a broad protection range are now prohibited in Europe, so the paint industry is considering engineered nanoparticles (ENPs) as an alternative biocide. There is concern that ENPs in paint might be released in run-off water and subsequently consumed by animals and/or humans, potentially coming into contact with cells of the gastrointestinal tract and affecting the immune system. Therefore, in the present study we evaluated the cytotoxic effects of three ENPs (nanosilver, nanotitanium dioxide and nanosilicon dioxide) that have a realistic potential for use in paints in the near future. When exposed to nanotitanium dioxide and nanosilicon dioxide in concentrations up to 243 µg/mL for 48 h, neither the gastrointestinal cells (CaCo-2) nor immune system cells (Jurkat) were significantly affected. However, when exposed to nanosilver, several cell parameters were affected, but far less than by silver ions used as a control. No differences in cytotoxicity were observed when cells were exposed to ENP-containing paint particles, compared with the same paint particles without ENPs. Paint particles containing ENPs did not affect cell morphology, the release of reactive oxygen species or cytokines, cell activity or cell death in a different manner to the same paint particles without ENPs. The results suggest that paints doped with ENPs do not pose an additional acute health hazard for humans.

Nanomaterial cell interactions: how do carbon nanotubes affect cell physiology?

Nanoparticulate materials and, among them, carbon nanotubes (CNTs) are new types of material that are generating high expectations owing to their unique physical, chemical and optical properties. Owing to the predictably increasing production of various types of CNTs and other nanoparticle-containing products, it is expected that environmental and public exposure to engineered nanoparticles will also increase in parallel. If and how far CNTs are able to affect health is, at present, discussed controversially. In this article, we summarize how CNTs are produced and processed to identify critical parameters, which have to be included in the toxicological assessment. A special effort is made to address the adverse effects of CNTs on cell physiology. Furthermore, we report on CNTs in medical applications and we discuss two selected examples of prospective applications of CNTs in nanomedicine, which have realistic chances of achieving ready-to-market products in just a few years.

Cytotoxic effects of nanosilver are highly dependent on the chloride concentration and the presence of organic compounds in the cell culture media

BackgroundNanosilver shows great promise for use in industrial, consumer or medical products because of its antimicrobial properties. However, the underlying mechanisms of the effects of silver nanoparticles on human cells are still controversial. Therefore, in the present study the influence of the chloride concentration and different serum content of culture media on the cytotoxic effects of nanosilver was systematically evaluated.ResultsOur results show that nanosilver toxicity was strongly affected by the composition of the culture media. The chloride concentration, as well as the carbon content affected the silver agglomeration and the complex formation. But also the dissolution of nanosilver and the availability of free silver ions (Ag+) were severely affected by the compositions of the culture media. Cells, only exposed to silver particles in suspension and dissolved silver complexes, did not show any effects under all conditions. Nanosilver agglomerates and silver complexes were not very soluble. Thus, cells growing on the bottom of the culture dishes were exposed to sedimented nanosilver agglomerates and precipitated silver complexes. Locally, the concentration of silver on the cell surface was very high, much higher compared the silver concentration in the bulk solution. The cytotoxic effects of nanosilver are therefore a combination of precipitated silver complexes and organic silver compounds rather than free silver ions.ConclusionsSilver coatings are used in health care products due to their bacteriostatic or antibacterial properties. The assessment of the toxicity of a certain compound is mostly done using in vitro assays. Therefore, cytotoxicity studies of nanosilver using human cell cultures have to be undertaken under well controlled and understood cultivations conditions in order to improve the compatibility of different studies. Especially when eukaryotic versus prokaryotic systems are compared for the evaluation of the use of nanosilver as antibacterial coatings for implants in order to prevent bacterial colonization.

Release of copper-amended particles from micronized copper-pressure-treated wood during mechanical abrasion

BackgroundWe investigated the particles released due to abrasion of wood surfaces pressure-treated with micronized copper azole (MCA) wood preservative and we gathered preliminary data on its in vitro cytotoxicity for lung cells. The data were compared with particles released after abrasion of untreated, water (0% MCA)-pressure-treated, chromated copper (CC)-pressure-treated wood, and varnished wood. Size, morphology, and composition of the released particles were analyzed.ResultsOur results indicate that the abrasion of MCA-pressure-treated wood does not cause an additional release of nanoparticles from the unreacted copper (Cu) carbonate nanoparticles from of the MCA formulation. However, a small amount of released Cu was detected in the nanosized fraction of wood dust, which could penetrate the deep lungs. The acute cytotoxicity studies were performed on a human lung epithelial cell line and human macrophages derived from a monocytic cell line. These cell types are likely to encounter the released wood particles after inhalation.ConclusionsOur findings indicate that under the experimental conditions chosen, MCA does not pose a specific additional nano-risk, i.e. there is no additional release of nanoparticles and no specific nano-toxicity for lung epithelial cells and macrophages.

Improving cell adhesion: development of a biosensor for cell behaviour monitoring by surface grafting of sulfonic groups onto a thermoplastic polyurethane

The surface properties of a material in combination with the mechanical properties are responsible for the material performance in a biological environment as well as the behaviour of the cells which contact with the material. Surface properties such as chemical, physical, biological play an important role in the biomaterials filed. In this work, the surface of a thermoplastic polyurethane film (Elastollan®1180A50) was tailored with sulfonic groups by grafting [2-(methacryloxyl)ethyl]-dimethyl-(3-sulfopropyl)-ammonium hydroxide (SB) after a previous surface activation either by Argon plasma or by ultra-violet irradiation. This surface modification had the purpose of improving cell adhesion in order to develop a biosensor able to monitor cell behaviour. The surfaces were characterized by X-ray photoelectron spectroscopy, by atomic force microscopy and by contact angle measurements in order to evaluate the efficiency of the modification. Additionally, blood compatibility studies and cell adhesion tests with human bone marrow cells were performed. These methods allowed the grafting of SB and the results indicate that a higher density of grafting was achieved with previous surface plasma treatment than with UV irradiation. However, for both techniques, the presence of SB functional groups led to a decrease of hydrophobicity and roughness of the surface, together with an improvement of the materials biological performance.