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  NANOMEDICINE  -  Toxicity under nano-surveillance

The ‘nano’ prefix can cause confusion, insofar as it can relate to the dimension in which a new kind of scientific activity is conducted (nanotechnologies) or the depth of analysis of a phenomenon being studied. Thus, when one speaks of research on the toxicity of nanoparticles, the reference is not to a danger posed by nanotechnologies specifically – although this could be the case – but to the mechanisms at work when one analyses the specific toxic effects of all kinds of particles (of anthropic as well as natural origin) on the nano scale. This is a new field and it is only now being explored.

Technicians and a robot producing oligonucleotides. © P. Stroppa, CEA
Technicians and a robot producing oligonucleotides.
© P. Stroppa, CEA
"It is not because of all the current excitement surrounding nanosciences that we are looking at the toxicity of nanoparticles,” stresses Ken Donaldsen, Scientific Director of Edinburgh University’s Inflammation Research Centre (Scotland). For this toxicologist and pillar of the British Association for Lung Research, there are two main reasons for scientific interest in the risks of exposure to nanoparticles.

The first is awareness of the dangers of transport-related pollution which, according to the WHO, results in 80 000 premature deaths a year in Europe. It is now clear that the pollutants that are usually studied (ozone, nitrogen or sulphur monoxide, etc.) do not in themselves explain this mortality, and that small suspended particles, known as PM10s (1), which are released by diesel engines in particular, also play a role. 

The second relates to what was a huge failure on the part of toxicologists in being unable to warn of the health risks of asbestos, as a result of which tens of thousands of people suffered. In this latter case too, it is minute suspended particles, just a few micrometres in length, which are responsible for the damage. The smaller the particles the easier it is for them to penetrate the respiratory system. So, if microparticles such as those linked to air or asbestos pollution present such serious risks to health, there would seem to be good reason to be suspicious of particles that are a thousand times smaller. 

Risk = danger x exposure
Toxicologists are not starting from scratch in their exploration of the toxicity of nanoparticles. They already have a conceptual framework summed up by the equation: risk = danger x exposure. The danger is an intrinsic characteristic of a substance, while the exposure varies depending on behaviour. As Ken Donaldsen explains, “the danger of using a chainsaw is of cutting oneself, but the risk is very different depending on whether it is used by a forester wearing all the protective gear or in a juggling act!”.   

In practice, it is difficult to quantify these two elements. The danger depends on the nature of the nanoparticle, its size and active surface area, the individual who absorbs it, the organ studied, and often varies depending on whether the exposure is isolated or regular. As to the exposure, that is a factor that has to be reconstructed after the event with all the unknowns that this implies. 

Effective toxicity
DNA analysis by polymerase chain reaction (PCR). © P. Stroppa, CEA
DNA analysis by polymerase chain reaction (PCR).
© P. Stroppa, CEA
The toxicology of nanoparticles is still largely unknown territory for the good reason that we know next to nothing about the two factors in the risk equation. To measure the danger, one must know what pathology to study: mesothelioma, in the case of asbestos; silicosis, in the case of carbon particles; and asthma, in the case of the famous family of airborne dust particles known as PM10s. But as yet we are unaware of what could be the other pathologies caused by nanoparticles and that – if they exist – would depend on their chemical nature.

As a result of this inability to evaluate the danger we are unable to estimate the exposure. The latter must be expressed in a unit of measurement that reflects what the specialists call the ‘toxicity effect’, that is, the principal component of the toxicity. For the PM10s, this is the mass and they are therefore quantified in micrograms/ m3; for asbestos, it is the fibres, and the unit of measurement is the number of fibres per m3.

And for nanoparticles? It is impossible to say at present. The only point on which researchers agree is that their surface area must be taken into account. For geometric reasons, 1 000 particles of 100 nanometres in diameter have a bigger surface area than a single particle measuring 1 micrometre in diameter, hence an increase in the possibilities for contact with biological tissue. There is therefore an urgent need to reach a consensus on the unit of measurement because, as Rob Aitken of Edinburgh University’s Institute of Occupational Medicine points out, “the rare data we have at present for measuring the toxicity of nanoparticles cannot be compared the one with the other as they are not expressed in the same units”.

Ingesting, touching, inhaling
To be able to determine the two key variables of nanoparticles – danger and exposure – toxicologists therefore have no alternative but to return to physiology to determine the path they take in the body. Theoretically, there are three possible means of access to the body’s interior and its vital organs: by means of ingestion, through the intestinal mucosa; by means of simple contact, through the skin; or by means of inhalation, with subsequent transport through the blood, through the pulmonary alveoli, or directly into the brain through the neurones and olfactive mucosa. 

It is only the latter of these three means of penetration that has so far been confirmed. As Günter Oberdörster, of Rochester University (United States), a pioneer in this field points out, “we have known since 1914 that when administered to a chimpanzee, a poliomitis virus, which measures no more than 30 nanometres, reaches the brain at a speed of 2.4 millimetres an hour by way of the olfactive neurones”.  

This observation has been repeated on many occasions, to the point of being used as a method for the study of olfactive neurone connections. Oberdöster’s work has also shown that nanoparticles could follow this path to quickly reach the upper centres of the brain: the cortex, thalamus and cerebellum. What are the consequences of their presence here? “Our preliminary results, among healthy volunteers, show that inhalation of nanoparticles emitted by the diesel engine changes the electroencephalogram rate,” explains Paul Borm of the Centre for Expert Assessment in Life Science in An Herlen (the Netherlands). And in the longer term? “That is one of the questions to be resolved in the coming years, in particular by studying the possibility of neurodegeneration,” admits Günter Oderdöster.

Much less is known about the mechanism for absorption by the lungs or intestine. The only certainty here is that exposure to certain nanoparticles can cause cardiac problems in animals. Some believe this is because the cells of the pulmonary epithelium are able to absorb these nanoparticles and transport them to the bloodstream, from where they spread throughout the body and thus also to the heart. Others see it as an indirect effect linked to the inflammation of the pulmonary mucosa through contact with the nanoparticles. Even less is known about possible absorption through the skin. The European NANOderm project was one of the first to tackle this question, through the in vivo and in vitro study, on human and pig skin cultures, of the passage of titanium oxide nanoparticles through the skin. 

This research is not without its economic significance as these particles are widely used in the cosmetic industry and are present in many sun lotions. After two years of research, Project Coordinator Tilman Butz of Leipzig University (Germany) is reassuring: ”Titanium oxide nanoparticles remain blocked in the upper layers of the epidermis and almost never penetrate to the dermis, except along the follicular cells that generate hairs.” The NANOderm consortium is nevertheless to continue its research to find out if damaged skin (burns, psoriasis, etc.) continues to act as an effective barrier.  

(1) PM10 refers to the fine suspended dust particles of an aerodynamic diameter of under ten micrometres. Formed of both primary and secondary pollutants, of natural or anthropic origin (soot, geological matter, abrasion dust, biological matter, etc.), this dust has a very variable composition (heavy metals, sulphates, nitrates, ammonium, organic carbon, aromatic polycyclic hydrocarbons, dioxins and furans). 
  The safety of nanoapplications  
 
The safety of nanoapplications
It will not be possible to develop nanotechnologies on an industrial scale without acquiring a better knowledge of the toxicity of nanoparticles, both for industry workers and the consumer. Hence the decision to set up the consortium of 23 academic and industrial laboratories from seven European countries, known as NANOSAFE2. With EU financing of €7 million, the consortium aims to compile a single database of current knowledge of nanoparticles commonly used in industry, such as carbon nanotubes, zinc oxide or silicon crystals. The database will draw on existing knowledge available in the specialist literature but which will now be pooled in a standard format, and the production of new knowledge. On this basis, NANOSAFE2 will study the potential sources of nanoparticles in various industrial sectors (aeronautics, energy, construction, etc.) and what happens to particles when they enter the atmosphere. Resolutely focused on applications, the research should make it possible to improve nanoparticle detection devices and define a scientific basis for rules of health and safety in the workplace. 		 

  Let’s talk nano-society  
  "This is the first time that scientists are turning to the media and general public to explain what they are doing at a very early stage in their research,” observes Ottilia Saxl, Director of the Stirling Institute of Nanotechnologies (UK). This institute plays an active role in promoting nanotechnologies – by increasing public awareness as well as supporting the sector’s industrialists and academics – and was instrumental in organising the 2005 EuroNanoForum, in September 2005.  

Communication with the general public is a priority of European nanotechnology policy, giving rise to diverse initiatives for dialogue between researchers and the public. There are information portals – the Nanoforum site, hosted by the aforementioned Stirling Institute, for example, is a genuine showcase for European nanosciences – and travelling exhibitions offering the very best of European research in the field, such as the NanoDialogue project piloted by the Naples City of Science (IT). Exchanges between the layman and the experts are structured and organised by sociologists, for example through the Nanologue project, piloted by the Wuppertal Institut (DE), one of the aims of which is to "help researchers to understand social expectations so they can take them into account when conducting their research”. One rather surprising initiative is a pack of cards, known as Democs. Developed with the aid of the New Economic Foundation in London, these offer factual information and ethical considerations and are designed to provide a entertaining way of increasing awareness. 

  

  GMOs and nanos  
 
Plasma membrane and proteins viewed through a scanning force microscope. © H. Oberleithner, University of Münster
Plasma membrane and proteins viewed through a scanning force microscope.
© H. Oberleithner, University of Münster
Is the same contrast not found, in both debates, between the utopian promises of the advocates and the apocalyptic prophesies of the detractors? Joyce Tait, a sociologist at Edinburgh University, believes that “while it is clear that the controversy over GMOs has influenced the way nanotechnologies are perceived, the comparison does not stand up to analysis as it supposes that nanotechnologies form a coherent whole that can be judged by citizens in full knowledge of the facts”. In fact, nanotechnologies cover a mixed bag of research and technological applications, some of which could be accepted and others rejected. “What is more,” stresses Joyce Tait, “nanotechnologies are still in their infancy and a lot of things could change, especially as a result of initiatives for dialogue between researchers and citizens.”