Genotoxic and cell-transforming effects of titanium dioxide nanoparticles.

The in vitro genotoxic and the soft-agar anchorage independent cell transformation ability of titanium dioxide nanoparticles (nano-TiO2) and its microparticulated form has been evaluated in human embryonic kidney (HEK293) and in mouse embryonic fibroblast (NIH/3T3) cells. Nano-TiO2 of two different sizes (21 and 50 nm) were used in this study. The comet assay, with and without the use of FPG enzyme, the micronucleus assay and the soft-agar colony assay were used. For both the comet assay and the frequency of micronuclei a statistically significant induction of DNA damage, was observed at the highest dose tested (1000 µg/mL). No oxidative DNA damage induction was observed when the comet assay was complemented with the use of FPG enzyme. Furthermore, long-term exposure to nano-TiO2 has also proved to induce cell-transformation promoting cell-anchorage independent growth in soft-agar. Results were similar for the two nano-TiO2 sizes. Negative results were obtained when the microparticulated form of TiO2 was tested, indicating the existence of important differences between the microparticulated and nanoparticulated forms. As a conclusion it should be indicated that the observed genotoxic/tranforming effects were only detected at the higher dose tested (1000 µg/mL) what play down the real risk of environmental exposures to this nanomaterial.

In vivo genotoxic effects of four different nano-sizes forms of silica nanoparticles in Drosophila melanogaster.

Although the use of synthetic amorphous silica (SAS) is steady increasing, scarce information exists on its potential health risk. In particular few and conflictive data exist on its genotoxicity. To fill in this gap we have used Drosophila melanogaster as in vivo model test organism to detect the genotoxic activity of different SAS with different primary sizes (6, 15, 30 and 55 nm). The wing-spot assay and the comet assay in larvae haemocytes were used, and the obtained results were compared with those obtained with the microparticulated form (silicon dioxide). All compounds were administered to third instar larvae at concentrations ranging from 0.1 to 10mM. No significant increases in the frequencies of mutant spots were observed in the wing-spot assay with any of the tested compounds. On the other hand, significant dose-dependent increases in the levels of primary DNA damage, measured by the comet assay, were observed for all the SAS evaluated but mainly when high doses (5 and 10mM) were used. These in vivo results contribute to increase the database dealing with the potential genotoxic risk associated to SAS nanoparticles exposure.

Intraocular intrusion of suture material that was used as scleral buckling material.

PURPOSE: To report intrusion of encircling suture used for scleral buckling 20 years ago. METHODS: Observational case report of a patient who admitted with the complaint of floaters. The patient had surgery for retinal detachment 20 years ago. Fundus examination revealed an encircling suture material. In some locations, the encircling suture had eroded all the way through the sclera to the vitreous cavity. In these locations, it was passing through the vitreous cavity. There was no retinal detachment, inflammation, or hemorrhage. RESULTS: Prophylactic argon laser photocoagulation was performed. No complication occurred during 1-year follow-up. CONCLUSION: A conservative approach with close follow-up may be suitable in case of an intrusion of suture material if there are no accompanying complications.

Zinc oxide nanoparticles: genotoxicity, interactions with UV-light and cell-transforming potential.

The in vitro genotoxic and the soft agar anchorage independent cell transformation ability of zinc oxide nanoparticles (NPs) and its bulky forms have been evaluated in human embryonic kidney (HEK293) and in mouse embryonic fibroblast (NIH/3T3) cells, either alone or in combination with UVB-light. The comet assay, with and without the use of FPG and Endo III enzymes, the micronucleus assay and the soft-agar colony assay were used. For the comet assay a statistically significant induction of DNA damage, with and without the enzymes, were observed up of 100µg/mL. ZnO NPs were able to increase significantly the frequency of micronuclei, and similar results were observed in the cell transformation assay where such NPs were able to induce cell-anchorage independent growth. These effects were observed at doses up 100µg/mL. Although UVB-light was able to induce genotoxic damage and cell-anchorage growth, a significant antagonist interaction effect was observed in combination with ZnO NPs. These in vitro results, obtained with the selected cell lines, contribute to increase our genotoxicity database on the ZnO NPs effects as well as to open the discussion about their risk in photo-protection sun screens.

In vivo genotoxicity assessment of titanium, zirconium and aluminium nanoparticles, and their microparticulated forms, in Drosophila.

As in vivo system, we propose Drosophila melanogaster as a useful model for study the genotoxic risks associated with nanoparticle exposure. In this study we have carried out a genotoxic evaluation of titanium dioxide (TiO2), zirconium oxide (ZrO2) and aluminium oxide (Al2O3) nanoparticles and their microparticulated forms in D. melanogaster by using the wing somatic mutation and recombination assay. This assay is based on the principle that loss of heterozygosis and the corresponding expression of the suitable recessive markers, multiple wing hairs and flare-3, can lead to the formation of mutant clones in treated larvae, which are expressed as mutant spots on the wings of adult flies. Third instar larvae were feed with TiO2, ZrO2 and Al2O3 nanoparticles, and their microparticulated forms, at concentrations ranging from 0.1 to 10mM. Although a certain level of aggregation/agglomeration was observed in solution, it must be noted than the constant digging activity of larvae ensures that treated medium pass constantly through the digestive tract ensuring exposure. The results showed that no significant increases in the frequency of all spots (e.g. small single, large single, twin, total mwh and total spots) were observed, indicating that these nanoparticles were not able to induce genotoxic activity in the wing spot assay of D. melanogaster. Negative data were also obtained with the microparticulated forms. This indicates that the nanoparticulated form of the selected nanomaterials does not modify the potential genotoxicity of their microparticulated versions. These in vivo results contribute to increase the genotoxicity database on the TiO2, ZrO2 and Al2O3 nanoparticles.

Determination of TiO2, ZrO2, and Al2O3 nanoparticles on genotoxic responses in human peripheral blood lymphocytes and cultured embyronic kidney cells.

In this study a genotoxic evaluation of titanium dioxide (TiO2, 2.3 nm), zirconium oxide (ZrO2, 6 nm), aluminum oxide (Al2O3, 16.7 nm) nanoparticles (NP) and their ionic forms was conducted using human peripheral blood lymphocytes and cultured human embryonic kidney (HEK293) cells by means of a modified alkaline comet assay with/without the formamidopyrimidine-DNA glycosylase (Fpg) and endonuclease III (Endo III) enzymes. Modifications to the comet assay by using lesion-specific endonucleases, such as Endo III and Fpg, detect DNA bases with oxidative damage. Both human peripheral blood lymphocytes and cultured embryonic kidney cells were incubated with TiO2, ZrO2, or Al2O3 NP at concentrations of 1, 10, or 100 µg/ml. Our results showed no significant induction in DNA damage by the comet assay with/without the Endo III and Fpg enzymes at all concentrations of ZrO2 and Al2O3. In the case of TiO2 NP only the highest concentration of 100 µg/ml significantly induced a genotoxic response. Data thus indicate that both ZrO2 and Al2O3 NP were not genotoxic in our system and in the case of TiO2 the lowest-observed-adverse-effect level (LOAEL) for genotoxicity was 100 µg/ml. Evidence indicates that these metallic NP are considered safe in light of the fact that no genotoxicity was noted with ZrO2 and Al2O3 and that the highest TiO2 concentration is not environmentally relevant.

Is there a relation between histopathologic characteristics of pterygium and recurrence rates?

PURPOSE: To explore the interrelationships of histopathologic characteristics of pterygium and postoperative recurrence. METHODS: Consecutive patients with primary pterygium or recurrent pterygium treated in our tertiary center between January 2007 and January 2010 were included in the study. All the patients were surgically treated by limbal-conjunctival autograft transplantation and postoperatively followed up for at least 1 year. Histopathologic changes were classified as inflammation intensity, degree of vascularization, and fibrinoid change. The results of examination of histopathologic and clinical characteristics of pterygium were comparatively analyzed. A total of 101 consecutive patients were included in the study. Ninety eyes of 90 patients who had primary pterygium (PP group) were compared with 11 eyes of 11 patients who had recurrent pterygium (RP group). In the PP group, 7 of the 90 (7.8%) patients had evidence of recurrence, while in the RP group, 2 of the 11 (18.2%) patients had evidence of recurrence (p=0.254). RESULTS: No significant difference was found in inflammation intensity, degree of vascularization, and fibrinoid change between PP and RP groups (p>0.05). In the PP group, no significant difference was found for inflammation intensity, degree of vascularization, and fibrinoid change between patients with (7/90) and without recurrences (83/90) (p>0.05). In the RP group, no significant difference was detected for inflammation intensity, degree of vascularization, and fibrinoid change between patients with (2/11) and without recurrences (9/11) (p>0.05).
 CONCLUSIONS: No significant correlation between the histology of pterygium and recurrence rate could be established.

Mutagenic/recombinogenic effects of four lipid peroxidation products in Drosophila.

The human diet is an important factor in the development of different diseases. Lipid peroxidation during frying in edible vegetable liquid oils of food components is a mechanism leading to the formation of free radicals. Such radicals induce tissue damage and are implicated in diverse pathological conditions, including aging, atherosclerosis, brain disorders, cancer, lung disorders and various liver disorders. In the present study, we decided to investigate the genotoxic effects of four lipid peroxidation products in the in vivo Drosophila wing somatic mutation and recombination test. In this test, point mutation, chromosome breakage and mitotic recombination produce single spots; while twin spots are produced only by mitotic recombination. Drosophila is a suitable eukaryotic organism for mutagenicity studies and also its metabolism is quite similar to that of mammalians. Since conflicting data exist on the possible risk of several lipid peroxidation products for humans, we have selected four of them, namely acrolein, crotonaldehyde, 4-hydroxy-hexenal (4-HHE) and 4-oxo-2-nonenal (4-ONE). Especially at the highest concentrations tested all exert both mutagenic and recombinogenic effects in the Drosophila SMART assay, showing a direct dose-effect relationship. This is the first study reporting genotoxicity data in Drosophila for these compounds.