As nanotechnology is moving into producing tonnes of nanoparticles, Dr. Vyvyan Howard explains why harmless materials become dangerous when shrunk to the nanoscale.
The nano-technology industry has begun the bulk production of nanoparticles, especially ultrafine particles for a range of commercial applications, from titanium dioxide in sunscreens to carbon nanotubes for molecular electronics (see "Nanotubes highly toxic" and "Nanoshells cure or curse?" this series). Manufacturers are moving into production levels in excess of 100 tonnes per annum.
Particles that can be breathed in (respirable) are classified as: coarse (average diameter less than 10micron); fine (average diameter less than 2.5 micron); and ultrafine (average diameter less than one micron). One micron (m) is one millionth of a metre and 1 000 nanometres (nm).
We have two defence mechanisms in the lung to deal with particles breathed in. The first is a carpet of mucus that lines all but the most peripheral parts of the lung. This carpet moves slowly upwards, carrying particles that have landed on it, and is then swallowed. Particles that make it through this carpet of mucus, which tend to be fine and ultrafine, get into the alveolar spaces where gas exchange between the air and the blood takes place. The alveolar surfaces are patrolled by macrophages, scavenger cells that mop up particles. However, macrophages appear to have difficulty recognising particles less than 70nm in diameter as being foreign, and in addition, they can be easily overwhelmed by too many particles, a condition called overload .
It is illuminating to consider the types of particles we were exposed to throughout the course of evolution. These consisted mainly of suspended sand and soil particles and pollen grains; most of which are relatively coarse and are trapped in the mucus before getting to the alveoli. There have always been ultrafine particles (UFPs), mainly consisting of minute crystals of salt, which become airborne through the action of the sea waves . These salt particles are not toxic, however, because they are soluble in water. What has become clear is that, for particles less than 70 nm in diameter, there was nothing much in the air throughout our prehistory of particular concern until we harnessed fire for use in our everyday life.
Research is revealing that when normally harmless bulk materials are made into UFPs, they tend to become toxic. Generally, the smaller the particle, the more reactive and toxic it becomes. This should come as no surprise, because that is exactly how catalysts are prepared to enhance industrial chemical reactions. By making particles of just a few hundred atoms, you create an enormous amount of surface, which tends to become electrically charged and thus chemically reactive. The upper size limit for the toxicity of UFPs is not fully known, but is thought to lie between 65 and 200nm .
There is epidemiological evidence that chronic exposure to particulate aerosols leads to long-term health effects, primarily on the cardiovascular system [4, 5]. Most of these studies have used coarse particles to assess the effects. More studies are now tending to use fine particles, though the question of whether it is more predictive of harm than coarse particles is till being debated . There is also evidence that short term effects from poor air quality is due to particle overloading. The number of studies that have used UFPs is low, but there are indications that UFPs are more hazardous than fine particles .
The main questions on the safety of nanoparticles are:
Definitive answers to all these questions are still lacking, though research is underway. Evidence of potential harm associated with UFPs comes from in vivo studies on toxicology and absorption and fate of UFPs and in vitro studies on mechanisms of toxicity in cells and tissues.
First, it should be noted that there appears to be a natural passageway for nanoparticles to get into and subsequently around the body. This is through the caveolar openings in the natural membranes, which separate body compartments. These openings are between 40 and 100 nm in size and are thought to be involved in the transport of macromolecules such as proteins, and on occasion, viruses. They also happen to be about the right size for transporting UFPs. Most of the research on that has been performed by the pharmaceutical industry interested in finding ways of improving drug delivery to target organs. This is particularly so for the brain, protected by the blood brain barrier, which can be very restrictive . It appears that chemists are able to design UFPs that can hoodwink certain membranes into allowing piggybacking of novel chemicals across membranes that would not be possible otherwise. For example, poly(butylcyanoacrylate) nanoparticles precoated with polysorbate 80 can be used to enhance the delivery of apolipoproteins to the brain [8,9].
Although there are clear advantages to the intentional and controlled targeting of difficult organs, such as the brain, with nanoparticles to increase drug deliver, the obverse of this particular coin needs to be considered. When environmental UFPs (as from traffic pollution) gain unintentional entry to the body, it appears that there is a mechanism that can deliver them to vital organs . The body is then wide open to any toxic effects that they can exert. The probable reason why we have not built up any defences is that such environmental UFPs were not part of the prehistoric environment in which we evolved and therefore there was no need to develop defensive mechanisms against them.
There is considerable evidence that inhaled UFPs can gain access to the blood stream and are then distributed to other organs in the body [10, 11]. This has been shown for synthetically produced UFPs such as bucky-balls [12, 13], which accumulate in the liver.
Another possible portal of entry into the body is via the skin. A number of sunscreen preparations now available have incorporated nanoparticle titanium dioxide. Recent studies  have shown that particles of up to 1 m in diameter (within the category of UFPs) can get deep enough into the skin to be taken up into the lymphatic system, while particles larger than that were excluded. The implication is that UFPs can and will be assimilated into the body through the skin. The exact proportion of those deposited on the skin, which will be absorbed, remains unknown. Using post mortem human skin, it has been shown that dextran beads 0.5 to 1m can penetrate the rough outer layer (stratum corneum) of the skin when flexed. The penetration occurred in over 50 % of the samples if flexing was continued for 1 hour. In a small proportion of cases, the beads got as far as the dermis (inner layer of the skin).
In vivo studies on laboratory animals have looked at the ability of UFPS to produce inflammation in lungs after exposure to UFP aerosols [3, 14-16]. The degree to which UFPs appear to be able to produce inflammation is related to the smallness of the particles, the age of the aerosol and the level of previous exposure. It has been proposed  that the chronic inhalation of particles can set up a low grade inflammatory process that can damage the lining of the blood vessels, leading to arterial disease.
In vitro studies on living cells have confirmed the increased ability of UFPs to produce free radicals that cause cellular damage [18-20]. This damage can be manifested in different ways, including genotoxicity and altered rates of cell death [18, 19, 21, 22].
Early indications were that transitional metals might be more toxic as UFPs than other materials . Since then, other studies have shown very similar toxicities between different materials when presented as UFPs, for example latex and titanium dioxide . UFPs are able to transport transition metals, which have been implicated in the pro-inflammatory effects and toxicity of coarse particles . More recently, transition metals have been discounted as a source of oxidative stress , and attention is being concentrated on the effects of ultrafine carbon black. What seems clear from all the papers is that exposure of living systems to UFPs tends to increase oxidative stress, and therefore, the effect of small size is considerably more important for UFP toxicity than the actual composition of the material.
There is evidence that UFPs can gain entry to the body by a number of routes, including inhalation, ingestion and across the skin. There is considerable evidence that UFPs are toxic and therefore potentially hazardous. The basis of this toxicity is not fully established but a prime candidate for consideration is the increased reactivity associated with very small size. The toxicity of UFPs does not appear to be very closely dependent on the type of material from which the particles are made, although there is still much research to be done before this question is fully answered.
Dr. Vyvyan Howard is histo-toxicologist at University of Liverpool. A version of this article first appeared as annex to "No Small Matter II: The Case for a Global Moratorium" www.etcgroup.og
Article first published 04/12/03
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