Characterization of NR1J1 Paralog Responses of Marine Mussels: Insights from Toxins and Natural Activators.

The pregnane X receptor (PXR) is a nuclear hormone receptor that plays a pivotal role in regulating gene expression in response to various ligands, particularly xenobiotics. In this context, the aim of this study was to shed light on the ligand affinity and functions of four NR1J1 paralogs identified in the marine mussel Mytilus galloprovincialis, employing a dual-luciferase reporter assay. To achieve this, the activation patterns of these paralogs in response to various toxins, including freshwater cyanotoxins (Anatoxin-a, Cylindrospermopsin, and Microcystin-LR, -RR, and -YR) and marine algal toxins (Nodularin, Saxitoxin, and Tetrodotoxin), alongside natural compounds (Saint John's Wort, Ursolic Acid, and 8-Methoxypsoralene) and microalgal extracts (Tetraselmis, Isochrysis, LEGE 95046, and LEGE 91351 extracts), were studied. The investigation revealed nuanced differences in paralog response patterns, highlighting the remarkable sensitivity of MgaNR1J1γ and MgaNR1J1δ paralogs to several toxins. In conclusion, this study sheds light on the intricate mechanisms of xenobiotic metabolism and detoxification, particularly focusing on the role of marine mussel NR1J1 in responding to a diverse array of compounds. Furthermore, comparative analysis with human PXR revealed potential species-specific adaptations in detoxification mechanisms, suggesting evolutionary implications. These findings deepen our understanding of PXR-mediated metabolism mechanisms, offering insights into environmental monitoring and evolutionary biology research.

In vitro safety assessment of reduced graphene oxide in human monocytes and T cells.

Considering the increase in the use of graphene derivatives in different fields, the environmental and human exposure to these materials is likely, and the potential consequences are not fully elucidated. This study is focused on the human immune system, as this plays a key role in the organism's homeostasis. In this sense, the cytotoxicity response of reduced graphene oxide (rGO) was investigated in monocytes (THP-1) and human T cells (Jurkat). A mean effective concentration (EC50-24 h) of 121.45 ± 11.39 µg/mL and 207.51 ± 21.67 µg/mL for cytotoxicity was obtained in THP-1 and Jurkat cells, respectively. rGO decreased THP-1 monocytes differentiation at the highest concentration after 48 h of exposure. Regarding the inflammatory response at genetic level, rGO upregulated IL-6 in THP-1 and all cytokines tested in Jurkat cells after 4 h of exposure. At 24 h, IL-6 upregulation was maintained, and a significant decrease of TNF-α gene expression was observed in THP-1 cells. Moreover, TNF-α, and INF-γ upregulation were maintained in Jurkat cells. With respect to the apoptosis/necrosis, gene expression was not altered in THP-1 cells, but a down regulation of BAX and BCL-2 was observed in Jurkat cells after 4 h of exposure. These genes showed values closer to negative control after 24 h. Finally, rGO did not trigger a significant release of any cytokine at any exposure time assayed. In conclusion, our data contributes to the risk assessment of this material and suggest that rGO has an impact on the immune system whose final consequences should be further investigated.

Potential oestrogenic effects (following the OECD test guideline 440) and thyroid dysfunction induced by pure cyanotoxins (microcystin-LR, cylindrospermopsin) in rats.

Potential endocrine-disrupting properties of cyanotoxins, such as microcystin-LR (MC-LR) and cylindrospermopsin (CYN) are of concern due to their increasing occurrence, the scarcity of reports on the topic (particularly for CYN) and the impact of human's health at different levels. Thus, this work performed for the first time the uterotrophic bioassay in rats, following the Organization for Economic Cooperation and Development (OECD) Test Guideline 440, to explore the oestrogenic properties of CYN and MC-LR (75, 150, 300 µg/kg b.w./day) in ovariectomized (OVX) rats. Results revealed neither changes in the wet and blotted uterus weights nor in the morphometric study of uteri. Moreover, among the steroid hormones analysed in serum, the most remarkable effect was the dose-dependent increase in progesterone (P) levels in rats exposed to MC-LR. Additionally, a histopathology study of thyroids and serum levels of thyroids hormones were determined. Tissue affectation (follicular hypertrophy, exfoliated epithelium, hyperplasia) was observed, as well as increased T3 and T4 levels in rats exposed to both toxins. Taken together, these results point out that CYN and MC-LR are not oestrogenic compounds at the conditions tested in the uterotrophic assay in OVX rats, but, however, thyroid disruption effects cannot be discarded.

Identification of in vitro metabolites of an Allium organosulfur compound and environmental toxicity prediction as part of its risk assessment.

Propyl-propane-thiosulfonate (PTSO) is an organosulfur compound found inAllium spp. Due to its antioxidant and antimicrobial activities, PTSO has been proposed for applications in the agri-food sector, such as feed additive. However, its use with commercial purposes depends on its toxicity evaluation. The present work aimed to perform a pilot-study of toxicokinetic profile of PTSO combining in silico and in vitro techniques, important steps in the risk assessment process. In silico ecotoxicity studies were also performed considering the importance of the environmental impact of the compound before its commercial use. First, an analytical method has been developed and validated to determine the original compound and its metabolites by ultra-performance liquid chromatography-tandem mass spectrometry. The phase I and II metabolism of PTSO was predicted using Meta-Pred Web Server. For the phase I metabolism, rat (male and female) and human liver microsomes were incubated with PTSO and NADPH regeneration system. Furthermore, in the phase II, microsomes were incubated with PTSO and glutathione or uridine 5'- diphosphoglucuronic acid. The analysis revealed the presence of propylpropane thiosulfinate (PTS) originated by redox reaction in phase I, and two conjugates from the phase II: S-propylmercaptoglutathione (GSSP) and S-propylmercaptocysteine (CSSP). Additionally, considering the environmental fate of PTSO and its metabolites, the ADME parameters and the potential ecotoxicity were also predicted using in silico softwares. The results of the ecotoxicity in silico study evidenced that the metabolism induced the formation of detoxified metabolites from the parent compound, except for dimercaprol and 3-mercaptopropane1,2-diol. Further in vivo assays are needed to confirm this prediction.

Risk assessment and environmental consequences of the use of the Allium-derived compound propyl-propane thiosulfonate (PTSO) in agrifood applications.

The organosulfur compound propyl-propane thiosulfonate (PTSO), mainly found in Allium cepa, has a promising use in the agrifood industry. To confirm its safety for livestock, consumers, and environment, toxicological assessment is needed. In this regard, endocrine-disrupting chemicals (EDCs) are in the spotlight of research. Therefore, as part of the risk assessment of PTSO, in the present work, an in vivo study was performed in mice exposed to PTSO to investigate its potential reproductive toxicity considering fertility, genetic and endocrine endpoints. Five-weeks-old CD1 mice (80 males, 80 females) were exposed for 11 or 16 weeks (males or females, respectively) to different doses of PTSO (0, 14, 28 and 55 mg PTSO/kg b.w./day; 20 animals per group and sex) through the food pellets. No clinical observations or mortality and no changes in absolute organ weights and relative organ weights/body weight or brain ratios occurred during the study. The estrous cycle did not undergo any significant toxicologically relevant change. Most of the sex hormones displayed normal values. Some alterations in the expression of some genes related to reproduction is only observed in females, but they do not appear to have consequences in the development of sex organs. Docking results showed the impossibility of stable binding to estrogen and androgen receptors. Considering all the results obtained, the safe profile of PTSO can be confirmed for different agrifood applications at the conditions assayed.

A new method for the simultaneous determination of cyanotoxins (Microcystins and Cylindrospermopsin) in mussels using SPE-UPLC-MS/MS.

The aim of this study was to optimize the extraction conditions of Microcystin-LR (MC-LR), Microcystin-RR (MC-RR), Microcystin-YR (MC-YR) and Cylindrospermopsin (CYN) simultaneously from mussels by using response surface methodology (RSM) and to validate the method by a dual solid phase extraction (SPE) system combined with ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The optimal parameters were: 90% MeOH (% v/v) for the extraction, a solvent/sample ratio of 75 and 15% MeOH in the extract before loading onto SPE. Mussels were spiked at 10; 37.5 and 75 ng g-1 fresh weight (f.w) of the 4 toxins, showing linear ranges of 0.5-75 ng g-1 f.w; low values for the limits of detection (0.01-0.39 ng g-1 f.w.) and quantification (0.23-0.40 ng g-1 f.w.); acceptable recoveries (70.37-114.03%) and relative standard deviation (%RSDIP) values (2.61-13.73%). The method was successfully applied to edible mussels exposed to cyanobacterial extracts under laboratory conditions, and it could allow the monitoring of these cyanotoxins in environmental mussel samples.

Microcystin-RR: Occurrence, content in water and food and toxicological studies. A review.

Microcystins (MCs) are hepatotoxins, produced by various species of cyanobacteria, whose occurrence is increasing worldwide owing to climate change and anthropogenic activities. More than 100 variants have been reported, and among them MC-LR is the most extensively studied, but there are other MC congeners that deserve to be investigated. The need for data to characterize the toxicological profile of MC variants other than MC-LR has been identified in order to improve risk assessment in humans and wildlife. Accordingly, the aim of this study was to evaluate the information available in the scientific literature dealing with MC-RR, as this congener is the second most common cyanotoxin in the environment. The review focuses on aspects such as occurrence in water and food, and toxicity studies both in vitro and in vivo. It reveals that, although MC-RR is a real hazard with a high exposure potential in some countries, little is known yet about its specific toxicological properties that differ from those of MC-LR, and important aspects such as genotoxicity and chronic effects have not yet been sufficiently addressed.

Cytotoxic and morphological effects of microcystin-LR, cylindrospermopsin, and their combinations on the human hepatic cell line HepG2.

Cylindrospermopsin (CYN) and Microcystin-LR (MC-LR) are toxins produced by different cyanobacterial species, which are found mainly in freshwater reservoirs. Both of them can induce, separately, toxic effects in humans and wildlife. However, little is known about the toxic effects of the combined exposure, which could likely happen, taking into account the concomitant occurrence of the producers. As both cyanotoxins are well known to induce hepatic damage, the human hepatocellular HepG2 cell line was selected for the present study. Thus, the cytotoxicity of both pure cyanotoxins alone (0-5 µg/mL CYN and 0-120 µg/mL MC-LR) and in combination for 24 and 48 h was assayed, as long as the cytotoxicity of extracts from CYN-producing and nonproducing cyanobacterial species. The potential interaction of the combination was evaluated by the isobologram or Chou-Talalay's method, which provides a combination index as a quantitative measure of the two cyanotoxins interaction's degree. Moreover, a morphological study of the individual pure toxins and their combinations was also performed. Results showed that CYN was the most toxic pure cyanotoxin, being the mean effective concentrations obtained ≈4 and 90 µg/mL for CYN and MC-LR, respectively after 24 h. However, the simultaneous exposure showed an antagonistic effect. Morphologically, autophagy, at low concentrations, and apoptosis, at high concentrations were observed, with affectation of the rough endoplasmic reticulum and mitochondria. These effects were more pronounced with the combination. Therefore, it is important to assess the toxicological profile of cyanotoxins combinations in order to perform more realistic risk evaluations.

Use of micronucleus and comet assay to evaluate evaluate the genotoxicity of oregano essential oil (Origanum vulgare l. Virens) in rats orally exposed for 90 days.

Essential oils from Origanum spp. exhibit antioxidant and antimicrobial activities making them suitable for use as food additives. The incorporation of oregano essential oil in active food packaging is under study; however, it has been not authorized for this purpose thus far. In order to fulfill the requirements of the European Food Safety Authority (EFSA), the aim of the present study was to determine the genotoxic potential of oregano essential oil using both the micronucleus (MN) test and comet (standard and enzyme-modified) assays in Wistar rats treated with 50, 100, or 200 mg/kg body weight administered daily for 90 days. MN was performed in cells from the bone marrow and standard and enzyme-modified comet assays were conducted in stomach, liver and blood cells. The major compound detected in the analytical study of oregano essential oil from Origanum vulgare L. virens, was carvacrol (55.82%) followed by thymol (5.14%), as well as their precursors, γ-terpinene (16.39%), and ρ-cimne (4.71%). The results obtained in the genotoxicity assays indicated lack of effect in MN and standard comet assay under the conditions tested. Furthermore, no apparent oxidative damage was observed in the enzyme-modified comet assay in any of the tissues examined of rats exposed to oregano essential oil for 90 days. Therefore, this oregano essential oil appears to be safe in Wistar rats and might be considered as a potential active material in food packaging industry.

Effects of Chrysosporum (Aphanizomenon) ovalisporum extracts containing cylindrospermopsin on growth, photosynthetic capacity, and mineral content of carrots (Daucus carota).

Natural toxins produced by freshwater cyanobacteria, such as cylindrospermopsin, have been regarded as an emergent environmental threat. Despite the risks for food safety, the impact of these water contaminants in agriculture is not yet fully understood. Carrots (Daucus carota) are root vegetables, extensively consumed worldwide with great importance for human nourishment and economy. It is, therefore, important to evaluate the possible effects of using water contaminated with cyanotoxins on carrot cultivation. The aim of this work was to investigate cylindrospermopsin effects on D. carota grown in soil and irrigated for 30 days, with a Chrysosporum ovalisporum extract containing environmentally relevant concentrations of cylindrospermopsin (10 and 50 µg/L). The parameters evaluated were plant growth, photosynthetic capacity, and nutritional value (mineral content) in roots of carrots, as these are the edible parts of this plant crop. The results show that, exposure to cylindrospermopsin did not have a clear negative effect on growth or photosynthesis of D. carota, even leading to an increase of both parameters. However, alterations in mineral contents were detected after exposure to crude extracts of C. ovalisporum containing cylindrospermopsin. A general decline was observed for most minerals (Ca, Mg, Na, Fe, Mn, Zn, Mo, and P), although an increase was shown in the case of K and Cu, pointing to a possible interference of the cyanobacterial extract in mineral uptake. This study is the first to evaluate the effects of C. ovalisporum extracts on a root vegetable, however, more research is necessary to understand the effects of this toxin in environmentally relevant scenarios.

Dietary l-carnitine prevents histopathological changes in tilapia (Oreochromis Niloticus) exposed to cylindrospermopsin.

Cylindrospermopsin (CYN) is a cytotoxin highly water-soluble, which is easily taken up by several aquatic organisms. CYN acts as a potent protein and glutathione synthesis inhibitor, as well as inducing genotoxicity, oxidative stress, and histopathological alterations. This is the first study reporting the protective effect of a l-carnitine (LC) pretreatment (400 or 880 mg LC/kg bw fish/day, for 21 days) on the histopathological alterations induced by pure CYN or Aphanizomenon ovalisporum lyophilized cells (400 µg CYN/kg bw fish) in liver, kidney, heart, intestines, and gills of tilapia (Oreochromis niloticus) acutely exposed to the toxin by oral route. The main histopathological changes induced by CYN were disorganized parenchyma with presence of glycogen and lipids in the cytoplasm (liver), glomerulonephritis, glomerular atrophy, and dilatation of Bowman's capsule (kidney), myofibrolysis, loss of myofibrils, with edema and hemorrhage (heart), intestinal villi with necrotic enterocytes and partial loss of microvilli (gastrointestinal tract), and hyperemia and hemorrhage (gills). LC pretreatment was able to totally prevent those CYN-induced alterations from 400 mg LC/kg bw fish/day in almost all organs, except in the heart, where 880 mg LC/kg bw fish/day were needed. In addition, the morphometric study indicated that LC managed to recover totally the affectation in the cross sections of the proximal and distal convoluted tubules in CYN-exposed fish. © 2015 Wiley Periodicals, Inc. Environ Toxicol 32: 241-254, 2017.

Effects of depuration on histopathological changes in tilapia (Oreochromis niloticus) after exposure to cylindrospermopsin.

Cylindrospermopsin (CYN) is a highly water-soluble cytotoxin produced by several species of freshwater cyanobacteria and it is considered the second most studied cyanotoxin worldwide. CYN acts as a potent protein and glutathione synthesis inhibitor, as well as inducing genotoxicity, oxidative stress and histopathological alterations. Studies concerning the depuration of cyanobacterial toxins in aquatic organisms, especially in fish, are of great interest for fish economy and public health, but are scarce in the case of CYN. This is the first study reporting the ability of depuration (3 - 7 days) in reversing or ameliorating the histopathological lesions induced in liver, kidney, heart, intestines, and gills of tilapia (Oreochromis niloticus) due to exposure by immersion to repeated doses of a CYN-containing culture of A. ovalisporum for 14 days. The main histopathological changes induced by CYN were glucogenic degeneration and loss of the normal hepatic cord-structure (liver), hyperemia, dilated Bowman's capsule and cellular tumefaction (kidney), myofibrolysis, hemorrhages and edema (heart), necrosis and partial loss of microvilli (gastrointestinal tract), and hyperemia and inflammatory cells infiltrates (gills). After 3 days of depuration, gills were totally recovered, while the liver, kidney, and gastrointestinal tract required 7 days, and longer depuration periods may be needed for a full recovery of the heart. In addition, the morphometric study indicated that depuration managed to reverse the affectation in the hepatocytes nuclear diameters and cross sections of the proximal and distal convoluted tubules induced in CYN-exposed fish. In general, these results validate depuration as an effective practice for detoxification of fish contaminated with CYN. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1318-1332, 2017.

Vitamin E pretreatment prevents histopathological effects in tilapia (Oreochromis niloticus) acutely exposed to cylindrospermopsin.

Cylindrospermopsin (CYN) is a cyanotoxin frequently involved in blooms with a predominantly extracellular availability, which makes it easily taken up by a variety of aquatic organisms. CYN is a potent protein and glutathione synthesis inhibitor, and also induces genotoxicity, oxidative stress and several histopathological lesions. The present study investigates the protective role of a vitamin E pretreatment (700 mg vit E/kg fish bw/day, for 7 days) on the histopathological alterations induced in different organs of tilapia (Oreochromis niloticus) acutely exposed to a single oral dose of 400 µg pure CYN/kg bw fish. The major histological changes observed were degenerative glucogenic process and loss of the hepatic structure in the liver, glomerulopathy and tubular tumefaction in the kidney, myofibrolysis and edema in the heart, catarrhal enteritis and necrosis in the gastrointestinal tract, hyperemic processes in the gill lamellae, and high basophilia, degeneration and tumefaction of granular neurons in the brain. Vitamin E pretreatment was effective in preventing or ameliorating the abovementioned alterations induced by CYN. In addition, a morphometric study indicated that the average nuclear diameter of hepatocytes, and cross-sections of proximal and distal convoluted tubules, together with the cardiac fiber and capillaries diameters represent a useful tool to evaluate the damage induced by CYN. This is the first study reporting vitamin E prevention of histopathological damage in tissues (liver, kidney, heart, gastrointestinal tract, gills and brain) of fish intoxicated with CYN. Therefore, vitamin E can be considered a useful chemoprotectant in the treatment of histopathological changes induced in CYN-intoxicated fish. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1469-1485, 2016.

Molecular characterisation of a bio-based active packaging containing Origanum vulgare L. essential oil using pyrolysis gas chromatography-mass spectrometry.

BACKGROUND: Environmental, economic and safety challenges motivate shift towards safer materials for food packaging. New bioactive packaging techniques, i.e. addition of essential plant oils (EOs), are gaining attention by creating barriers to protect products from spoilage. Analytical pyrolysis gas chromatography-mass spectrometry (Py-GC-MS) was used to fingerprint a bioactive polylactic acid (PLA) with polybutylene succinate (PBS) (950 g kg(-1) :50 g kg(-1) ) film extruded with variable quantities (0, 20, 50 and 100 g kg(-1) ) of Origanum vulgare EO. RESULTS: Main PLA:PBS pyrolysis products were lactide enantiomers and monomer units from the major PLA fraction and succinic acid anhydride from the PBS fraction. Oregano EO pyrolysis released cymene, terpinene and thymol/carvacrol peaks as diagnostic peaks for EO. In fact, linear correlation coefficients better than 0.950R(2) value (P < 0.001) were found between the chromatographic area of the diagnostic peaks and the amount of oregano EO in the bioplastic. CONCLUSION: The pyrolytic behaviour of a bio-based active package polymer including EO is studied in detail. Identified diagnostic compounds provide a tool to monitor the quantity of EO incorporated into the PLA:PBS polymeric matrix. Analytical pyrolysis is proposed as a rapid technique for the identification and quantification of additives within bio-based plastic matrices. © 2015 Society of Chemical Industry.

Toxicological evaluation of clay minerals and derived nanocomposites: a review.

Clays and clay minerals are widely used in many facets of our society. This review addresses the main clays of each phyllosilicate groups, namely, kaolinite, montmorillonite (Mt) and sepiolite, placing special emphasis on Mt and kaolinite, which are the clays that are more frequently used in food packaging, one of the applications that are currently exhibiting higher development. The improvements in the composite materials obtained from clays and polymeric matrices are remarkable and well known, but the potential toxicological effects of unmodified or modified clay minerals and derived nanocomposites are currently being investigated with increased interest. In this sense, this work focused on a review of the published reports related to the analysis of the toxicological profile of commercial and novel modified clays and derived nanocomposites. An exhaustive review of the main in vitro and in vivo toxicological studies, antimicrobial activity assessments, and the human and environmental impacts of clays and derived nanocomposites was performed. From the analysis of the scientific literature different conclusions can be derived. Thus, in vitro studies suggest that clays in general induce cytotoxicity (with dependence on the clay, concentration, experimental system, etc.) with different underlying mechanisms such as necrosis/apoptosis, oxidative stress or genotoxicity. However, most of in vivo experiments performed in rodents showed no clear evidences of systemic toxicity even at doses of 5000mg/kg. Regarding to humans, pulmonary exposure is the most frequent, and although clays are usually mixed with other minerals, they have been reported to induce pneumoconiosis per se. Oral exposure is also common both intentionally and unintentionally. Although they do not show a high toxicity through this pathway, toxic effects could be induced due to the increased or reduced exposure to mineral elements. Finally, there are few studies about the effects of clay minerals on wildlife, with laboratory trials showing contradictory outcomes. Clay minerals have different applications in the environment, thus with a strict control of the concentrations used, they can provide beneficial uses. Despite the extensive number of reports available, there is also a need of systematic in vitro-in vivo extrapolation studies, with still scarce information on toxicity biomarkers such as inmunomodulatory effects or alteration of the genetic expression. In conclusion, a case by case toxicological evaluation is required taking into account that different clays have their own toxicological profiles, their modification can change this profile, and the potential increase of the human/environmental exposure to clay minerals due to their novel applications.

Genotoxic and cytotoxic effects and gene expression changes induced by fixed orthodontic appliances in oral mucosa cells of patients: a systematic review.

CONTEXT: The accumulation of chronic or severe acute DNA and cellular damage in oral mucosa cells is one of the main factors that help initiate a wide range of malignant lesions in the oral cavity. There has been considerable controversy in the literature about the effect of such sustained genotoxic and cytotoxic damage to oral mucosa cells. OBJECTIVE: The aim of this systematic review, reported in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, is to investigate the effects of such interventions. METHODS: Electronic and manual searches were performed (15 May 2015) for Randomized Clinical Trials/quasi-Randomized Clinical Trials that analyzed the genotoxic/cytotoxic effects of these types of oral appliances in humans. A primary outcome (cell/DNA damage) and a number of secondary outcomes were examined. Two reviewers carried out the study selection and performed a "risk of bias" assessment [Cochrane Collaboration's tool]. Wherever possible the meta-analysis was conducted on homogenous groups. RESULTS: From the electronic search (2797), 6 studies met the eligibility criteria. Most studies (5/6) observed significant differences in most comparisons at the short-term (1-3 months) and long-term (24-48 months) evaluations, with respect to critically acute genotoxic/cytotoxic effects. Some of the studies (2/3) concluded that the post-removable effects at DNA/cellular levels were not significant (p > 0.05) with respect to the controls. CONCLUSIONS: Acute DNA/cellular damage in oral mucosa cells is induced by orthodontic appliances. Nevertheless, even though these effects were no longer detected after removing the appliances, more rigorous RCTs are needed to explore the extent to which acquired damage can be observed in the oral mucosa.

Preliminary study of genotoxicity evaluation of orthodontic miniscrews on mucosa oral cells by the alkaline comet assay.

Miniscrew implants are widely used nowadays in orthodontic treatments due to their good results in clinical practice. However, data regarding the biocompatibility of commercially available orthodontic miniscrews and temporary devices are very scarce, and their role as genotoxicity inducers has been not previously evaluated with the alkaline comet assay. The aim of this study was to investigate the DNA damage in buccal cells of patients subjected to orthodontic treatments. The alkaline comet assay has been applied in oral mucosa cells from patients treated with conventional orthodontic treatment in comparison to patients treated additionally with miniscrews, non-treated volunteers (control) and smoking volunteers (positive control). The application of orthodontic appliances and miniscrews induced significant and similar (2-fold) increases of %DNA in tail in comparison to control group. Females experienced a significant increase in %DNA in all the treatments in comparison to the control group, whereas males showed significant damage only with the combined orthodontic and miniscrew treatment. In conclusion, conventional orthodontic appliances induced genotoxicity, and the incorporation of miniscrews assayed did not imply any additional increase of DNA damage.

In vivo evaluation of activities and expression of antioxidant enzymes in Wistar rats exposed for 90 days to a modified clay.

Although clays are wildly used in a range of applications, the toxicity assessment of these new materials is still scarce. In the present study, oxidative stress induced by Clay 1, a novel clay, was determined in rats after 90 d of oral exposure. The activities of antioxidant enzymes, namely, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione S-transferase (GST), were examined. In addition, genetic expressions of SOD and CAT and relative protein abundance of CAT were also determined. Data showed that most of the biomarkers assayed remained unaltered. Only CAT activity, as well as its genetic and protein expressions, appeared enhanced in the kidney. Therefore, further studies are needed to clarify the relevance and consequences of these findings to ensure the safety of this clay.

Toxic effects of a modified montmorillonite clay on the human intestinal cell line Caco-2.

The incorporation of the natural mineral clay montmorillonite into polymeric systems enhances their barrier properties as well as their thermal and mechanical resistance, making them suitable for a wide range of industrial applications, e.g., in the food industry. Considering humans could easily be exposed to these clays due to migration into food, toxicological and health effects of clay exposure should be studied. In the present work, the cytotoxic effects induced by two different clays (the unmodified clay Cloisite(®) Na(+) , and the organically modified Cloisite(®) 30B) on Caco-2 cells were studied after 24 and 48 h of exposure. The basal cytotoxicity endpoints assessed were total protein content, neutral red uptake and a tetrazolium salt reduction. Our results showed that only Cloisite(®) 30B induced toxic effects. Therefore, the effects of subcytotoxic concentrations of this clay on the generation of intracellular reactive oxygen species, glutathione content and DNA damage (comet assay) were investigated. Results indicate that oxidative stress may be implicated in the toxicity induced by Closite(®) 30B, in regards of the increases in intracellular reactive oxygen species production and glutathione content at the highest concentration assayed, while no damage was observed in DNA. The most remarkable morphological alterations observed were dilated cisternae edge in the Golgi apparatus and nucleolar segregation, suggesting impairment in the secretory functions, which could be related to inhibition in the synthesis of proteins.

Acute exposure to pure cylindrospermopsin results in oxidative stress and pathological alterations in tilapia (Oreochromis niloticus).

Cylindrospermopsin (CYN) is increasingly recognized as a potential threat to drinking water safety, due to its ubiquity. This cyanotoxin has been found to cause toxic effects in mammals, and although fish could be in contact with this toxin, acute toxicity studies on fish are nonexistent. This is the first study showing that single doses of CYN pure standard (200 or 400 µg CYN/kg fish bw) by oral route (gavage) generate histopathological effects in fish (Tilapia-Oreochromis niloticus) exposed to the toxin under laboratory condition. Among the morphological changes, disorganized parenchymal architecture in the liver, dilated Bowman's space in the kidney, fibrolysis in the heart, necrotic enteritis in the intestines, and hemorrhages in the gills, were observed. Moreover, some oxidative stress biomarkers in the liver and kidney of tilapias were altered. Thus, CYN exposure induced increased protein oxidation products in both organs, NADPH oxidase activity was significantly increased with the kidney being the most affected organ, and decreased GSH contents were also detected in both organs, at the higher dose assayed.