Genotoxicity evaluation of fried meat: A comprehensive review.

Some years ago, the IARC published the carcinogenic potential of processed and red meat. It is known that frying meat can produce genotoxic substances. A systematic review of the literature was conducted to evaluate in vitro and in vivo genotoxicity of fried meat. A total of 31 scientific articles were retrieved and analyzed. The meat extraction methods have been grouped into 6 types based on their similarity to an initially described method or on the general methodology used (solid-liquid extraction or others). The in vitro mutagenic results have been summarised by type of meat studied (beef, pork, others), cooking conditions (method, time and temperature), extraction method, and test used, with or without S9. Most articles assessed the mutagenicity of the extracts using the Ames test. Meat extracts were consistently positive in strains TA98/TA1538 with metabolic activation. In the in vitro studies with meat from restaurants, positive results were always found with variations in the number of His+ revertants between samples or between restaurants. The few in vivo studies retrieved show evidence of induced DNA damage in colon cells and chromosome aberrations in bone marrow cells after daily treatment with fried red meat for 4 weeks or longer.

Applying the comet assay to fresh vs frozen animal solid tissues: A technical approach.

The in vivo comet assay is usually performed in fresh tissues by processing cells immediately after collection, an approach that is not always possible from a logistical point of view. Although the comet assay has been applied to frozen rodent tissue samples on several occasions, there is currently no agreement on the best way to freeze and thaw them. We have tested two different thawing procedures and compared the levels of DNA strand breaks (SBs) and Fpg-sensitive sites in fresh and frozen (for up to year) liver, kidney and lung tissue samples, from untreated and methyl methanosulfonate treated rats. Tissues were snap frozen, stored at -80 °C and processed in such a way that the tissue remained frozen until the cells were in suspension. Our results showed that comparable levels of DNA SBs were detected in fresh and frozen liver and lung samples stored at -80 °C for up to 1 year and 3 months, respectively. In kidney, similar levels of SBs were detected either in fresh or in frozen tissues stored for up to 1 year. However, more studies are needed to control the variability observed in the Fpg-sensitive site levels in this tissue at the different freezing periods.

Genotoxic evaluation of poly(anhydride) nanoparticles in the gastrointestinal tract of mice.

Gantrez® AN 119-based NPs have been developed as oral drug carriers due to their strong bioadhesive interaction with components of the gastrointestinal mucosa and to their adaptable surface. The use of mannosamine to coat Gantrez® AN 119-based NPs results in a high mucus-permeable carrier, able to reach the gastrointestinal epithelium. Although their efficacy to transport a therapeutic agent has been demonstrated, their safety has not yet been thoroughly studied. They have proved to be non-cytotoxic, non-genotoxic and non-mutagenic in vitro; however, the in vivo toxicity profile has not yet been determined. In this study, the in vivo genotoxic potential of Gantrez® AN 119 NPs coated with mannosamine (GN-MA-NP) has been assessed using the in vivo comet assay in combination with the enzyme formamidopyrimidine DNA glycosylase in mice, following the OECD test guideline 489. To determine the relevant organs to analyse and the sampling times, an in vivo biodistribution study was also carried out. Results showed a statistically significant induction of DNA strand breaks and oxidized bases in the duodenum of animals exposed to 2000 mg/kg bw. However, this effect was not observed at lower doses (i.e. 500 and 1000 mg/kg which are closer to the potential therapeutic doses) or in other organs. In conclusion, GN-MA-NP are promising nanocarriers as oral drug delivery systems.

In vitro evaluation of the genotoxicity of poly(anhydride) nanoparticles designed for oral drug delivery.

In the last years, the development of nanomaterials has significantly increased due to the immense variety of potential applications in technological sectors, such as medicine, pharmacy and food safety. Focusing on the nanodevices for oral drug delivery, poly(anhydride) nanoparticles have received extensive attention due to their unique properties, such as their capability to develop intense adhesive interactions within the gut mucosa, their modifiable surface and their biodegradable and easy-to-produce profile. However, current knowledge of the possible adverse health effects as well as, toxicological information, is still exceedingly limited. Thus, we investigated the capacity of two poly(anhydride) nanoparticles, Gantrez® AN 119-NP (GN-NP) and Gantrez® AN 119 covered with mannosamine (GN-MA-NP), and their main bulk material (Gantrez® AN 119-Polymer), to induce DNA damage and thymidine kinase (TK+/-) mutations in L5178Y TK+/- mouse lymphoma cells after 24h of exposure. The results showed that GN-NP, GN-MA-NP and their polymer did not induce DNA strand breaks or oxidative damage at concentrations ranging from 7.4 to 600µg/mL. Besides, the mutagenic potential of these nanoparticles and their polymer revealed no significant or biologically relevant gene mutation induction at concentrations up to 600µg/mL under our experimental settings. Considering the non-genotoxic effects of GN-NP and GN-MA-NP, as well as their exceptional properties, these nanoparticles are promising nanocarriers for oral medical administrations.

Evaluation of the cytotoxicity, genotoxicity and mucus permeation capacity of several surface modified poly(anhydride) nanoparticles designed for oral drug delivery.

The main concerns with drugs designed for oral administration are their inactivation or degradation in the harsh conditions of the gastrointestinal tract, their poor solubility through the gastrointestinal mucus gel layer, the poor intestinal epithelium permeability that limits their absorption, and their toxicity. In this context, poly(anhydride) nanoparticles are capable of protecting the drug from the harsh environment, reduce the drug's toxicity and, by virtue of surface modification, to enhance or reduce their mucus permeability and the bioadhesion to specific target cells. The copolymer between methyl vinyl ether and maleic anhydride (commercialized as Gantrez® AN 119) are part of the poly(anhydride) nanoparticles. These biocompatible and biodegradable nanoparticles (NPs) can be modified by using different ligands. Their usefulness as drug carriers and their bioadhesion with components of the intestinal mucosa have been described. However, their toxicity, genotoxicity and mucus permeation capacity has not been thoroughly studied. The aim of this work was to evaluate and compare the in vitro toxicity, cell viability and in vitro genotoxicity of the bioadhesive empty Gantrez® AN 119 NPs modified with dextran, aminodextran, 2-hydroxypropyl-ß-cyclodextrin, mannosamine and poly-ethylene glycol of different molecular weights. Results showed that, in general, coated NPs exhibit better mucus permeability than the bare ones, those coated with mannosamine being the most permeable ones. The NPs studied did not affect cell metabolism, membrane integrity or viability of Caco-2 cells at the different conditions tested. Moreover, they did not induce a relevant level of DNA strand breaks and FPG-sensitive sites (as detected with the comet assay).

Toxicity evaluation of nanocarriers for the oral delivery of macromolecular drugs.

Oral administration is the most commonly used and accepted route for drug administration. However, two of the main concerns are the poor intestinal epithelium permeability and rapid degradation, which limit absorption of drugs. In this context, nanocarriers have shown great potential for oral drug delivery. Nevertheless, special importance should be given to the possible toxic effect of these nanocarriers, such as their bioaccumulation in different tissues of the body, as well as, the different physicochemical parameters influencing their properties and so their potential toxic effect. This review describes first some aspects related to the behavior of nanosystems within the gastrointestinal tract and then some aspects of nanotoxicology and its evaluation, including the most popular techniques and approaches used for in vitro and in vivo toxicity studies. It also reviews the physicochemical characteristics of polymeric nanoparticles that may influence the development of toxicological effects, and finally it summarizes the toxicity results that have been published regarding polymeric nanocarriers.

Ochratoxin A reduces aflatoxin B1 induced DNA damage detected by the comet assay in Hep G2 cells.

Mycotoxins aflatoxin B1 (AFB1) and ochratoxin A (OTA) can be present together in food commodities. These food contaminants are considered to be genotoxins, acting by different mechanisms. The aim of this work was to characterize combined genotoxic in vitro effects of both mycotoxins in Hep G2 cells. For this purpose, cytotoxicity was first determined in isolated and combined treatments in order to determine the dose range of genotoxicity studies. Co-exposure of cells to OTA+AFB1 for 24 h resulted in additive effects. Genotoxicity was determined in Hep G2 cells by the modified comet assay with restriction enzymes (endo III and FPG). Significant reactive oxygen species formation was detected in both single and combined treatments. AFB1 was genotoxic after 3 h with external metabolic activation (S9 mix) and after 24 h without metabolic activation. Co-exposure to OTA significantly decreased DNA damage induced by AFB1, not only in breaks and apurinic sites but also in FPG-sensitive sites. The apparent contradiction between additive cytotoxic effects and antagonic genotoxic effects may be explained if AFB1 and OTA compete for the same CYPs, yielding more ROS but less AFB1 adducts.

Anticancer effect of a new benzophenanthridine isolated from Zanthoxylum madagascariense (Rutaceline).

Fractionation of the cyclohexane extract from the stem bark powder of Zanthoxylum madagascariense led to the isolation of a new benzophenanthridine-type alkaloid, hydrochloride of 2,3-methylendioxy-8-hydroxy- 7-methoxy-benzo[C]phenanthridine (Rutaceline), characterized on the basis of its spectral data. Rutaceline was evaluated for its antiproliferative capacity on the human colorectal adenocarcinoma (Caco-2) and the African green monkey kidney (Vero) cell lines. The 50% inhibition of cell growth (IC50) obtained after 24 h incubation was similar for both cells lines (110-115 microg/ml, i.e. 269-281 microM), but at 48 h the IC50 value for the Caco-2 cells was lower than for the Vero cells (20 microg/lml, i.e. 49 microM versus 90 microg/ml, i.e. 220 microM) indicating a higher cell growth inhibitory effect on the colon adenocarcinoma cells. At the respective IC50 concentrations, Rutaceline did not significantly induce apoptosis but induced cell cycle arrest in the GO/G1 phase, as well as a decrease of cells in S phase. Rutaceline also induced DNA fragmentation in both cell lines, as revealed by agarose gel electrophoresis, and a dose-dependent clastogenic effect in both cell lines as revealed by the Comet assay.

Phenazine 5,10-dioxide derivatives as hypoxic selective cytotoxins: Part II. Structure-activity relationship studies.

The synthesis and evaluation as hypoxic selective cytotoxins of new derivatives of 2-amino or 2-hydroxyphenazine 5,10-dioxide are described. The compounds were developed as structural analogs of other bioreductive compounds and its in vitro cytotoxicities on V79 cells under hypoxic and aerobic conditions were determined. To gain insight into its mechanism of action electrochemical behavior, interaction with DNA experiments and QSAR studies were performed.

Design and evaluation of "3 + 1" mixed ligand oxorhenium and oxotechnetium complexes bearing a nitroaromatic group with potential application in nuclear medicine oncology.

The synthesis and evaluation of a series of oxotechnetium and oxorhenium complexes containing a nitroaromatic moiety as potential radiopharmaceuticals for targeting tumour hypoxia is presented. 99mTc labelling was performed in high yield (>85%) and radiochemical purity (>90%). Their structure was corroborated by means of the rhenium complexes. Reduction potentials were in the range for bioreducible compounds. 99mTc complexes III-VI were selected for "in vivo" experiments in view of the results of cytotoxicity studies. Biodistribution in normal animals was characterized by high initial blood, lung and liver uptake, fast blood and soft tissue depuration and preferential excretion via the hepatobiliary system. Initial tumour uptake was moderate but tumour/muscle ratios for complexes III and IV, were favourable at all time points. Although the results are encouraging further development is still necessary in order to achieve higher tumour uptake and lower gastrointestinal activity.

A comparative study on the gastroduodenal tolerance of different antianaemic preparations.

The most significant adverse effect of repeated oral administration of iron-containing antianaemic preparations is the gastroduodenal toxicity, attributable to a direct toxic effect of iron on the glandular epithelium. To assess gastroduodenal mucosal damage and the potential protective effect of different antianaemic preparations, a study was carried out to compare the gastroduodenal toxicity caused by three different types of antianaemic drugs in normal and anaemic rats administered at repeated therapeutic doses. Histological damage to the gastroduodenal mucosa was evaluated using light and electron microscopy. In both normal and anaemic rats, pathological changes were less marked in animals treated with ferrimannitol-ovoalbumin (TM/FMOA) than in those treated with iron protein succinylate or ferrous sulphate. Electron microscopic studies of duodenal mucosa in normal rats treated with iron protein succinylate and ferrous sulphate confirmed a severe ultrastructural alteration, whereas no changes were detected in animals treated with TM/FMOA. In anaemic rats, slight duodenal ultrastructural changes were noted with all three types of treatment. The effectiveness of the preparations in resolving the anaemia was similar in the three groups. It was concluded that TM/FMOA exerts a protective effect against the toxicity normally observed of the iron in other formulations in normal and anaemic rats, which was attributed to the fact that administration of iron bound to a protein core allows for gradual release of iron.