In vitro comet and micronucleus assays do not predict morphological transforming effects of silica particles in Syrian Hamster Embryo cells.

Crystalline silica particles and asbestos have both been classified as carcinogenic by the International Agency for Research on Cancer (IARC). However, because of the limited data available, amorphous silica was not classifiable. In vitro, the carcinogenic potential of natural crystalline and amorphous silica particles has been revealed by the Syrian Hamster Embryo (SHE) cell transformation assay. On the other hand, the genotoxic potential of those substances has not been investigated in SHE cells. And yet, genotoxicity assays are commonly used for hazard evaluation and they are often used as in vitro assays of reference to predict a possible carcinogenic potential. The main objective of this study was to compare the genotoxic potential and the carcinogenic potential of different crystalline and amorphous silica particles in SHE cells. Three silica samples of different crystallinity were used: natural amorphous silica, partially crystallized silica and quartz silica particles. Their genotoxicity were tested through the in vitro micronucleus assay and the comet assay in SHE, and their carcinogenic potential through the SHE transformation assay. In addition, silica samples were also tested with the same genotoxicity assays in V79 hamster-lung cells, a common in vitro model for particle exposure. Results obtained in the micronucleus and the comet assays show that none of the silica was capable of inducing genotoxic effects in SHE cells and only the amorphous silica induced genotoxic effects in V79 cells. However in the SHE cell transformation assays, the partially crystallized and quartz silica were able to induce morphological cell transformation. Together, these data suggest that, in vitro, the short-term genotoxic assays alone are not sufficient to predict the hazard and the carcinogenic potential of this type of particles; SHE transformation assay appears a more reliable tool for this purpose and should be included in the "in vitro battery assays" for hazard assessment.

Surface reactivity, cytotoxic, and morphological transforming effects of diatomaceous Earth products in Syrian hamster embryo cells.

In order to evaluate the effect of thermal treatments on the surface reactivity and carcinogenic potential of diatomaceous earth (DE) products, the physicochemical features of some specimens--derived by heating the same original material--were compared with their cytotoxic and transforming potency. The samples were an untreated DE (amorphous) progressively heated in the laboratory at 900 degrees C (DE 900) and 1200 degrees C (DE 1200) and a commercial product manufactured from the same DE (Chd) from which the finer fraction (< 10-microm diameter) was separated (Chd-F). Quartz (Min-U-Sil 5) and a vitreous silica (amorphous) smoothed up with hydrofluoric acid and were used as positive and negative controls, respectively. All samples were analyzed for their degree of crystallization, for their ability to release free radicals and reactive oxygen species, and for their cytotoxic and transforming potencies in Syrian hamster embryo (SHE) cells. X-ray diffractometry showed that DE 900, like DE, was still amorphous, whereas DE 1200 as well as the commercial product (Chd) were partially crystallized into cristobalite. The ability of the dust to release hydroxyl (*OH) radicals in the presence of hydrogen peroxide, as revealed by the spin-trapping technique, was as follows: Chd-F, DE 1200 > Chd > DE 900 > DE, suggesting that on heating, the surface acquires a higher potential for free radical release. Most of the silica samples generated COO* radicals from the formate ion, following homolytic rupture of the carbon-hydrogen bond, in the presence of ascorbic acid. A concentration-dependent decrease in cell proliferation and colony-forming efficiency was observed in SHE cultures treated with Chd-F, Chd, and DE. Heating abolished DE cytotoxicity but conferred a transforming ability to thermal treated particles. DE was the only sample that did not induce morphological transformation of cells. According to their transformation capacity, the samples were classified as follows: Chd-F > Chd, DE 1200 > DE 900 >> DE. Taken together, the reported results suggest that (1) the transforming potential of a biogenic amorphous silica is related to the thermal treatment that transforms the original structure in cristobalite and generates surface active sites; (2) the reactivity of samples in releasing *OH radicals correlates to their transforming ability; (3) the finer fraction of the commercial product is significantly more toxic and transforming than the coarse dust; and (4) opposite to silica dusts of mineral origin, which loose both cytotoxicity and transforming ability upon heating, heated diatomite acquires a cell-transforming potency. DE products should be thus considered a set apart of silica-based potentially toxic materials.

Surface reactivity, cytotoxicity, and transforming potency of iron-covered compared to untreated refractory ceramic fibers.

Untreated and iron-coated refractory ceramic fibers (RCFs) 1, 3, and 4 were examined for their potential to generate free radicals and to catalyze hydrogen peroxide decomposition in cell-free assays and were compared for cytotoxic and transforming potencies in Syrian hamster embryo (SHE) cell system. Coating with a high quantity of iron increased the capability of RCFs to generate hydroxyl radicals and to catalyze the decomposition of hydrogen peroxide. In the SHE cells, the untreated RCFs had varying ability to induce inhibition of cell proliferation, cytotoxicity (as measured by the colony-forming efficiency, CE) and morphological transformation, in a concentration-dependent manner. According to cytotoxic and transforming potencies, they ranged as follows: RCF3 > RCF1 > RCF4. The lethal concentration 50 (LC50; decrease of CE to 50% of controls after 7 d of treatment) expressed per number of RCF3 and RCF1/cm(2) of culture dish was 2.5 x 10(4) and 3.7 x 10(4), respectively, whereas RCF4 was not cytotoxic up to the highest concentration tested (23.7 x 10(4) fibers/cm(2)). At LC50, RCF3 was 1.4-fold more transforming than RCF1, and the weakest, RCF4, induced less than 1% transformation. Iron coating of RCF1 and RCF3 markedly attenuated their cytostatic, cytotoxic, and transforming potencies without a linear concentration-transformation relationship. In contrast, iron coating of RCF4 affected slightly its low transforming potency, although the growth inhibitory effect was reduced. The observed decrease rather than increase in the cytotoxic and transforming potencies of the active samples RCF1 and RCF3 by their coating with large amounts of ferric iron suggests that it is not the quantity or any form of iron on the surface of fibers but the iron, even in trace, in a particular redox and coordinate state that might play a role in the fiber's surface reactivity with regard to the biological material. Surface chemical functions involved in the interaction with the cell could be inactivated by the deposition of a high quantity of Fe(III) on the surface of fibers. Physicochemical studies correlated to biological effects is an approach for understanding the properties of solids related to a given biological response and for elucidating the cellular and molecular mechanisms.