University of Konstanz, Germany
Manmade or engineered nano- materials, usually defined as being under 100 nanometer in one dimension, are increasingly appearing in our environment and our daily consumer products. They are made in all kinds of shapes and materials. A nanotube, for example, can have a diameter of 1 nanometer, which is 100,000 times smaller than the diameter of a hair. Over 1000 consumer products already contain nanomaterials, such as textiles, food items, cosmetics, electronics, optics, as well as coatings for facades and cars. In discussion is also the use of nanomaterials in medical devices, treatments and diagnostics, as well as for soil and water remediation. Akin to the promises of stem cell technologies to revolutionize our concept of human health and life, nanotechnology holds great promises for the future. Fueled by government and industry funding, nanotechnology enabled products emerge at an increasing pace in the marketplace, with a forecast market share of around 1 trillion US dollars by 2015. However, as with all great human ‘inventions’ that leapfrogged the development of human kind, there is also a potential for harm.
Due to their small size and high surface area, nanomaterials are more aggressive than larger forms of the same material. Importantly, nanoparticles elicit the formation of reactive oxygen species in cells. For example, silver is relatively inert as a metal and not toxic at fine particle size. At nanosize however, silver generates reactive oxygen species and oxidative stress in cells, damages DNA, and is thus toxic to cells. And while nanoparticles incorporated into materials and coatings are not as aggressive as free nanoparticles, leaching cannot be excluded in the long term, and accumulation in the environment and the food chain is likely. The future use of nanoparticles in remediation of soil and water will also contribute to accumulation of nanoparticles in the environment. Moreover, the use of nanomaterials in medicine as medical devices, new diagnostic tools, drug delivery and improved cancer therapies will entail a direct injection into the body. It is thus to be expected that human exposure will increase. Surprising then is the scarcity of human studies investigating potential hazardous effects of these new materials already widely in use.
There is however a plethora of data from animal studies demonstrating toxicity of nanoparticles. It has been shown that inhaled nanoparticles penetrate the blood circulation and reach most organs, including the brain. In dogs exposed to high levels of air pollution, which contains a significant proportion of nanoparticulate combustion materials, particles could be detected in the brain. There was DNA damage in nasal and brain tissues, neuronal activation of the pro-inflammatory transcription factor NFkB and chronic brain inflammation, with diffuse plaques and Alzheimer’s pathology. Toxicological studies in rodents point to health risks for humans, including cardiovascular disease, pulmonary diseases and impairment of brain function. Potential outcomes of nanomaterial exposure can be inferred from numerous human epidemiological studies in high air pollution areas and studies investigating outcomes after inhalation exposure to diesel exhaust particles. Air pollution and diesel exhausts contain a high proportion of nanosized particles. Epidemiological studies have shown that diesel exhausts cause pulmonary complications such as asthma and COPD (chronic obstructive pulmonary disease) and cardio-vascular disease. Moreover, inhalation studies of diesel exhaust particles in healthy volunteers showed short-term changes in the lung, with systemic inflammation, changes in vascular function and brain activity. Cellular studies showed that diesel exhaust particles are specifically toxic to dopaminergic neurons, the neurons that are lost in Parkinson’s disease.
Mechanisms of nanotoxicity are various, including induction of oxidative stress, integration into mitochondria and ensuing damage, activation of inflammation and immune responses, changes in receptor or channel function by bound or incorporated NPs, interaction with enzymes, DNA damage, cell cycle changes and more (1, 2). The effects differ between the different NPs and depend on chemical composition, shape, size and surface properties of the nanoparticle. The same material can have dramatically different effects on cells with changing shape or size. Nanoparticles can enter the cells by simple diffusion, or are being taken up actively by the cells, by endocytosis or macropinocytosis. Or they can enter through membrane channels, and depending on their size block or alter the function of these channels. A peripheral neuronal cell, for example, with altered channel function can have wide-ranging effects in the body. Another organ that may be susceptible to such effects is the heart.
Nanoparticles also bind to receptors and can thus function as cytokines or growth factors and trigger signaling cascades that are otherwise not active. Also, nanoparticles coat themselves with a protein corona when in contact with biological fluids, which may increase cellular uptake, depending on which proteins are bound. Furthermore, the proteins found in the coronas differ from nanoparticle to nanoparticle, with a difference in bound proteins, when shape is changed for the same material. It was also demonstrated that as nanoparticles interact with proteins they enhance the rate of protein fibrillation, that is aggregation, suggesting that they could contribute to amyloid fibril formation in the brain, similar to the diffuse plaques observed in the brains of air pollution-exposed dogs. Interestingly, our brain naturally contains biogenic magnetite nanoparticles, which have been shown to be increased in the case of Alzheimer’s disease.
One example of a widely used and well characterized nanoparticle is titanium dioxide. In cultured cells titanium dioxide nanoparticles elicit oxidative stress. Long-term cell cultures showed a decreased mitochondrial activity, morphological changes of the cells, and an increase in S, G2/M phase. A recent study investigated the properties of the food additive E171 (titanium dioxide), which is freely used by the food and cosmetic industry. E171 is not labeled as nano-particulate titanium dioxide but nevertheless contains a significant amount of particles under 100 nm in diameter (3). E171 is added to a variety of food items, ranging from tooth paste to white candy coatings as well as pharmaceuticals. Titanium dioxide nanoparticles are also in most sunscreens and many personal health care products. As many candy coatings and products consumed by children contain E171, exposure is higher for children than adults. In rodent studies inhalation of titanium dioxide nanoparticles elicited pulmonary inflammation. The nanoparticles crossed cellular membranes by nonphagocytic mechanisms and ended in all major compartments of the lung, in the cells and in capillaries.
It is known that nanoparticles can cross the placenta and the blood brain barrier (without specific functionalization) and can thus reach the brain of the developing fetus. Concerning is the finding that titanium dioxide nanoparticles (which we ingest in our daily food) cause complications during pregnancy in mice and elicit mild behavioral alterations in the offspring as well as changes in their gene expression in the brain. Genes that were affected in the offspring of exposed mice play a role in brain development and psychiatric disease (4, 5). Nothing is known yet of the effects of gestational or early childhood exposure to nanoparticles in humans. We can however draw on past poisoning instances. For example, gestational or early childhood exposure to mercury or lead, has resulted in children with mild to severe malformation and altered behavior and intelligence, while the mothers suffered no harm (6). During development, the human nervous system is very susceptible to low levels of toxicity. The developing cells have to go through a finely tuned and highly regulated process of proliferation, specification, differentiation, migration, maturation and proper integration into neuronal circuits. If during these highly complex and dynamic processes which need to proceed at the right time and place, nanoparticles interfere with cellular components, by either binding to them, triggering membrane receptors, blocking channels or inhibiting enzyme function, cells cannot divide or differentiate properly and may soon not function adequately in the three-dimensional and fast developing fetus. This can then lead to inappropriately located neurons, or not the right proportions of neuronal subtypes or neurons and glial cells, and thus will impede proper function, if not lead to severe malformations. It is estimated that lead exposure alone has decreased intelligence by several IQ points in the general population in the US.
We still have only scarce information about effects of long-term exposure to nanomaterials. Yet there are sufficient instances in the past where chemicals were used widely without any prior consideration of potential long-term harm, and of which we are still assessing the damages. One prominent example are pesticides, which in the early days of pesticide usage were deemed harmless to humans. Now, some 60 years later we know that the 12 most toxic persistent organic pollutants include 8 pesticides. Persistent organic pollutants cause allergies, hypersensitivity, nervous system damage, reproductive and immune dysfunction, neurobehavioral and developmental disorders, endocrine disruption and cancer. While most of these highly toxic pesticides have been banned in most countries, bioaccumulation and persistence in the environment still affects health and almost daily reports are published demonstrating toxicity of pesticides. Exposure to environmental pesticides has been shown to be associated with increased risk for Alzheimer’s and Parkinson’s disease, and multiple sclerosis. A recent study even showed changes in brain anatomy in children that were exposed to higher levels of pesticide than the control group with lower exposure levels (7).
Similarly, almost daily reports on toxicity of nanomaterials are published. Unfortunately, most reports are on short-term effects. However, this is changing, and hopefully we will know more about nanotoxicty before these, otherwise highly attractive, materials have conquered too much of our daily lives.
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About the Author
Suzanne Kadereit is a stem cell biologist who obtained her PhD at the Pasteur Institute in Paris. She worked on umbilical cord blood immune and stem cells in the US, prior to working with human embryonic stem cells in Singapore. While in Singapore she became founding head of the Singapore stem cell bank. She is now group leader of the stem cell group at the Chair for in vitro Toxicology and Biomedicine at the University of Konstanz, developing stem cell-based assay systems from human embryonic stem cells for disease modelling and toxicological studies of nanoparticles.
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