My opinion

By Dr. Csaba Varga
Corresponding Author Dr. Csaba Varga
Dept. Environmental Health, University of Pecs, - Hungary 7624
Submitting Author Dr. Csaba Varga

Particles, Genotoxicity, Asbestos Fibres, Carbon Nanotubes, Mesothelioma, Peloids

Varga C. Solid-Phase Environmental Genotoxicity: In Vivo Veritas!. WebmedCentral TOXICOLOGY 2012;3(1):WMC002932
doi: 10.9754/journal.wmc.2012.002932

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Submitted on: 26 Jan 2012 08:53:47 AM GMT
Published on: 26 Jan 2012 07:17:32 PM GMT


Specific genotoxicity of solid particles are discussed by using three different examples. Carcinogenic asbestos fibres have primary genotoxic effects as well, that can be studied in in vitro studies. Nevertheless, demonstration of their mesothelioma inducing effect is only possible in in vivo studies. Their carrier function seems to be essential for the biological activity. Carbon nanotubes do not show any genotoxic effects, but lack of their carcinogenicity is still debated. The size distribution is close to asbestos fibres and fibrils, after all, their biological behaviour are rather different. Medicinal muds used in balneotherapy and medical wellness - consisting of suspensions- cause dermal exposure of patients. Some kinds of these muds may have genotoxicity but probably not the particles themselves, rather their extractable chemical fractions. Ethical aspects of environmental health studies on particles are also discussed.


"The Stethoscope of Genetic Toxicology for the 21st Century." Claxton and co-workers [1] use this excellent metaphor in their most recent article for the Salmonella/Ames mutagenicity test. The Ames test is actually a useful tool for screening environmental chemicals, but is it also true for solid materials. The new challenge of the century, no doubt, is the broad spectrum of particulate matter involving fibres and especially the nanotech products.
What are the main philosophic and strategic differences of solid-phase genotoxicology in contrast to the traditional one. The answer involves completely different behaviour of particles as compared to chemical genotoxicants. While primary genotoxicity, as a result of surface properties and several related mechanisms [2], can be studied in cultured phagocytic cells, detecting secondary genotoxicity (e.g. permanent formation of reactive oxygen species from inflammation) requires in vivo systems. Chemicals can act either directly on the DNA or indirectly. In case of particles even entering into the cell is not simple. While chemical substances follow the classic toxicokinetics, it is completely different in case of particles. (The specific particle kinetics involves deposition, clearance, durability, overload, persistence of poorly soluble particles, etc.). Perhaps the most important feature is the carrier function, since the significance of transported chemicals cannot be overemphasized.
Genotoxicity studies appeared in the literature can be classified into 4 main groups. Non-cellular studies are restricted to the formation of lesions in the pure DNA. In vitro studies are considered as suitable tool to detect primary effects, excluding tests on non-mammalian cell lines. The few animal (in vivo) studies were performed on respiratory tract cells or whole lung tissue considered pulmonary system as main target. Transgenic animals were also involved in the recent investigations. The fewer human (biomarker) studies searched for relevant surrogate cells available in a non-invasive way [2].
Nowadays we are - more or less - aware of the possible mechanisms of primary genotoxicity of the different fibrous and non-fibrous particles. Intrinsic chemical properties, the possible contaminants, surface properties, size distribution and crystalline structure may have major role, but in some cases extractable chemical content significantly modifies the expected effects. The following examples demonstrate three different types of particles with different genotoxicity patterns. The examples prove the introductory statement: "in vivo veritas!".
Example #1: Asbestos fibres
Even after the hopeful complete banning of asbestos production, the environmental exposure to asbestos fibres will still cause persistent hazard to human health for long. In spite of the intensive research several open questions have remained till today. Asbestos fibres are archetypes of carcinogenic fibres, but the main drawback of animal and other laboratory studies is the almost exclusive application of UICC asbestos [3]. Use of these fibres is understandable due to the comparability, but humans are rather exposed to commercial size (non-respirable) ones. Vast majority of inhaled fibres are coughed up and ingested. That is, oral exposure may be the typical route in the whole population. Asbestos in drinking water means an additional source of ingested fibres. Therefore risk assessment should be based primarily on the in vivo effects of ingested asbestos considering the carrier function, as well [4]. Electron microscope studies proved that the fibres penetrate the gut wall, reach peritoneal cavity and the omentum accumulates them. Benzo[a]pyrene, as a model compound successfully adsorbed onto their surface. Comet-assay performed on intestinal, omental and peritoneal cells indicated weak genotoxic effects after oral exposure to pure amphibole fibres but significant potentiating effect of the polyaromatic compound carried was also clearly proved [5]. Mesothelioma as the very specific cancer caused by asbestos can only be studied in in vivo studies. Our experiments suggest that pollutants adsorbed by the fibres can cause specific cancers rather than fibres themselves. The drinking water-borne fibres can also adsorb organic compounds (e.g. chlorination by-products) of well-known biological activity (mutagenicity, carcinogenicity [4]). The importance of the route of exposure was also demonstrated by the in vivo genotoxicity studies with 1-nitropyrene, an air pollutant carcinogenic nitroarene. Results of oral administration significantly differed from other treatments. This fact also emphasize importance of the proper selection of ways of exposure in the risk assessment process [6].
Example #2: Single- and multiwalled carbon nanotubes (SW/MWCNTs)
These new nanoproducts are in scope of interest because their size parameters are very close to asbestos. Their primary genotoxicity was rejected evaluating results of in vitro Ames, cytogenetic and in vivo mutagenicity data. Only mitotic inhibition was observed with SW tubes [7]. The investigation on carbon nanotubes has accelerated after Poland and co-workers' scandalous announcement. An unethical media campaign was initiated accusing CNTs of mesothelioma inducing effect based on granuloma formation in a 5-day animal study [8]. Some recent studies delivered further but limited evidences while others rejected the hypothesis. In our rat model only granuloma but no mesothelioma formation was observed after 1-year direct peritoneal exposure to high doses of both SW- and MWCNTs [9].
Example #3: Peloids
Peloids (medicinal muds) are suspensions of inorganic and/or organic particulate matter. In spite of their extensive use in balneotherapy, the late toxicological consequences of dermal exposure have not been cleared; risk-benefit calculations cannot be performed. Two Hungarian peloids were tested in toto in the Ames test without any observable effect. Only extractable chemical fractions showed mutagenicity but it was fluctuating by time, perhaps depending on the microbial activity of the mud samples. After some successful ecotoxicological experiments with earthworms (Eisenia fetida) [10], the comet assay with their coelomocytes proved relevant test for a chronic in toto exposure study. Mud particles did not cause genotoxicity in contrast to soil contaminated with aromatics.


In conclusion, avoiding l' art pour l'art research on the health effects of particles we need (i) strategies considering the real world exposures, (ii) development of new tests/models or creative use of the existing ones, and (iii) ethical behaviour in handling data to avoid frightening the public. Media generate hyperboles but the responsibility of interpretation is of the science.


Supported by grant no. 34039 Faculty of Medicine, University of Pécs.


1. Claxton LD, Umbuzeiro GA, DeMarini DM. The Salmonella mutagenicity assay: The stethoscope of genetic toxicology for the 21st century. Environ Health Persp 2010; 118: 1515-22.
2. Schins RPF. Mechanisms of genotoxicity of particles and fibers. Inhalation Toxicol 2002; 14: 57-78.
3. Timbrell V, Rendall, REG. Preparation of the UICC standard reference samples of asbestos. Powder Technol 1971/72; 5: 279-87.
4. Varga C. Can one assess genotoxic and carcinogenic risk of asbestos without mentioning ingested fibres Mutation Res 2005; 572: 173-4.
5. Varga C, Horváth G, Timbrell V. On the mechanism of cogenotoxic action between ingested amphibole asbestos fibres and benzo(a)pyrene: II. Tissue specificity studies using comet assay. Cancer Lett 1999; 139: 173-6.
6. Varga C, Szendi K, Ember I. An in vivo model for testing genotoxicity of environmental fibre-associated nitroarenes. In Vivo 2006; 20: 539-42.
7. Szendi K, Varga C. Lack of genotoxicity of carbon nanotubes in a pilot study. Anticancer Res 2008; 28: 349-52.
8. Poland CA, Duffin R, Kinloch I, Maynard A, Wallace WAH, Seaton A, Stone V, Brown S, MacNee W, Donaldson K. Carbon nanotubes introduced into the abdominal cavity of mice show asbestos-like pathogenicity in a pilot study. Nature Nanotech 2008; 3: 423-8.
9. Varga C, Szendi K. Carbon nanotubes induce granulomas but not mesotheliomas. In Vivo 2010; 24: 153-6.
10. Gerencsér G, Szendi K, Murányi E, Varga C. Exotoxicological studies on Hungarian peloids (medicinal muds). Appl Clay Sci 2010; 50: 47-50.

End note

This paper is based on the lecture delivered in Pécs (Hungary) at the Collegium Ramazzini - University of Pécs Joint Symposium, November 17-18th, 2010.

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4 reviews posted so far

solid phase env genotoxicity
Posted by Dr. Marie Bourgeois on 22 Mar 2012 12:28:27 AM GMT

Accept completely
Posted by Prof. Amir Jalali on 08 Nov 2011 03:25:54 PM GMT

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