Lorella Severino

Effects of four Fusarium toxins (fumonisin B1, α-zearalenol, nivalenol and deoxynivalenol) on porcine whole-blood cellular proliferation

The in vitro effects of four Fusarium toxins, fumonisin B(1) (FB(1)), alpha-zearalenol (alpha-ZEA), nivalenol (NIV) and deoxynivalenol (DON), on mitogen-induced cell proliferation were determined in swine whole-blood cultures. Considering the lack of sufficient toxicological data both on single and in combination effects, in vitro studies may contribute to risk assessment of these toxins. Incubation with increasing concentrations of FB(1) did not produce any consequence on proliferation; in contrast alpha-ZEA, NIV and DON showed an inhibitory effect. Dose-response curves for each mycotoxin were generated. NIV was found to be the most potent toxin followed by DON and alpha-ZEA. The effects of both FB(1)+alpha-ZEA and NIV+DON mixtures were also analysed to investigate possible interactions. The results indicated that combination of FB(1)+alpha-ZEA produces a synergistic inhibition of porcine cell proliferation; whereas there is no interaction between DON and NIV on porcine whole-blood proliferation, at tested concentrations.

Modulation of macrophage activity by aflatoxins B1 and B2 and their metabolites aflatoxins M1 and M2.

Aflatoxins are natural contaminants frequently found both in food and feed. Many of them exert immunomodulatory properties in mammals; therefore, the aim of the current study was to investigate immune-effects of AFB1, AFB2, AFM1 and AFM2, alone and differently combined, in J774A.1 murine macrophages. MTT assay showed that AFB1, alone and combined with AFB2, possess antiproliferative activity only at the highest concentration; such effect was not shown by their hydroxylated metabolites, AFM1 and AFM2, respectively. However, the immunotoxic effects of the aflatoxins evaluated in the current study may be due to the inhibition of production of active oxygen metabolites such as NO. Cytofluorimetric assay in macrophages exposed to aflatoxins (10-100 μM) revealed that their cytoxicity is not related to apoptotic pathways. Nevertheless, a significant increase of the S phase cell population accompanied by a decrease in G0/G1 phase cell population was observed after AFB1 treatment. In conclusion, the results of the current study suggest that aflatoxins could compromise the macrophages functions; in particular, co-exposure to AFB1, AFB2, AFM1 and AFM2 may exert interactions which can significantly affect immunoreactivity.

Nivalenol and deoxynivalenol affect rat intestinal epithelial cells: a concentration related study.

The integrity of the gastrointestinal tract represents a crucial first level defence against ingested toxins. Among them, Nivalenol is a trichotecenes mycotoxin frequently found on cereals and processed grains; when it contaminates human food and animal feed it is often associated with another widespread contaminant, Deoxynivalenol. Following their ingestion, intestinal epithelial cells are exposed to concentrations of these trichothecenes high enough to cause mycotoxicosis. In this study we have investigated the effects of Nivalenol and Deoxynivalenol on intestinal cells in an in vitro model system utilizing the non-tumorigenic rat intestinal epithelial cell line IEC-6. Both Nivalenol and Deoxynivalenol (5–80 µM) significantly affected IEC-6 viability through a pro-apoptotic process which mainly involved the following steps: (i) Bax induction; (ii) Bcl-2 inhibition, and (iii) caspase-3 activation. Moreover, treatment with Nivalenol produced a significant cell cycle arrest of IEC-6 cells, primarily at the G0/G1 interphase and in the S phase, with a concomitant reduction in the fraction of cells in G2. Interestingly, when administered at lower concentrations (0.1–2.5 µM), both Nivalenol and Deoxynivalenol affected epithelial cell migration (restitution), representing the initial step in gastrointestinal wound healing in the gut. This reduced motility was associated with significant remodelling of the actin cytoskeleton, and changes in expression of connexin-43 and focal adhesion kinase. The concentration range of Nivalenol or Deoxynivalenol we have tested is comparable with the mean estimated daily intake of consumers eating contaminated food. Thus, our results further highlight the risks associated with intake of even low levels of these toxins.

Pro-apoptotic effects of nivalenol and deoxynivalenol trichothecenes in J774A.1 murine macrophages

Nivalenol (NIV) and deoxynivalenol (DON) are trichothecenes mycotoxins produced by Fusarium fungi that occur in cereal grains alone or in combination. Several studies have shown that exposure to high concentrations of these mycotoxins resulted in decreased cell proliferation; however, the molecular mechanism underlying their activities are still partially known. In this study, we evaluated the effects of NIV and DON, alone and in combination, on J7741.A macrophages viability. The results of the current study show that both NIV and DON (10-100 microM) significantly stimulate apoptosis in J774A.1 macrophages in a concentration-dependent manner; in particular, NIV results a stronger pro-apoptotic effect than DON on cultured J774A.1 murine macrophages. No interactive effects were observed by exposing J774A.1 cells to both NIV and DON simultaneously. Pro-apoptotic activity induced by both mycotoxins seems to be essentially mediated by caspase-3 and is associated with a cell cycle blocking in G0/G1 phase. Moreover, our results show that NIV and DON are able to influence apoptotic pathway by ERK, pro-apoptotic protein Bax, caspase-3 and poly-ADP-ribose synthase (PARP), DNA repairing enzyme.

Effect of fumonisin B1 on structure and function of macrophage plasma membrane

Fumonisin B1 (FB1), a mycotoxin produced by Fusarium moniliforme and related fungi, is nephrotoxic, neurotoxic, hepatotoxic, carcinogenic and immunosuppressive in animals and man. In this study we evaluate the modifications of fluidity, endocytosis and peroxidative damage of plasma membrane induced by FB1 in macrophage cell line J774A.1. In these immune cells FB1 (1-10 microM) enhances membrane fluidity and increases, time-dependently, the horseradish peroxidase (HRP) endocytosis. This effect is concentration-dependent, significant at 10 microM, and reverted by IFN-gamma (100 U/ml). Moreover, FB1 (1-10 microM) induces a membrane peroxidative damage as evident by the increase of malondialdehyde (MDA) production. All these mycotoxin effects provide additional insight into potential mechanism by which FB1, in macrophages, might enhance membrane damage and oxidative stress contributing to the pathogenesis of mycotoxin induced diseases.

Nivalenol induces oxidative stress and increases deoxynivalenol pro-oxidant effect in intestinal epithelial cells.

Mycotoxins are secondary fungal metabolites often found as contaminants in almost all agricultural commodities worldwide, and the consumption of food or feed contaminated by mycotoxins represents a major risk for human and animal health. Reactive oxygen species are normal products of cellular metabolism. However, disproportionate generation of reactive oxygen species poses a serious problem to bodily homeostasis and causes oxidative tissue damage. In this study we analyzed the effect of two trichothecenes mycotoxins: nivalenol and deoxynivalenol, alone and in combination, on oxidative stress in the non-tumorigenic intestinal epithelial cell line IEC-6. Our results indicate the pro-oxidant nivalenol effect in IEC-6, the stronger pro-oxidant effect of nivalenol when compared to deoxynivalenol and, interestingly, that nivalenol increases deoxynivalenol pro-oxidative effects. Mechanistic studies indicate that the observed effects were mediated by NADPH oxidase, calcium homeostasis alteration, NF-kB and Nrf2 pathways activation and by iNOS and nitrotyrosine formation. The toxicological interaction by nivalenol and deoxynivalenol reported in this study in IEC-6, points out the importance of the toxic effect of these mycotoxins, mostly in combination, further highlighting the risk assessment process of these toxins that are of growing concern.

Trichothecenes NIV and DON modulate the maturation of murine dendritic cells

Nivalenol (NIV) and Deoxynivalenol (DON), mycotoxins of the trichothecene family are considered very common food contaminants. In this work, we investigated whether the immunotoxic effects ascribed to these trichothecenes may be mediated by perturbations in the activity of dendritic cells (DCs). Murine bone marrow-derived DCs were used to evaluate the effects of NIV and DON on the LPS-induced maturation process. We found that the expression of the class II MHC and of the accessory CD11c molecules, but not of the costimulatory CD86 marker, was down-regulated by NIV and DON exposure in LPS-treated DCs, as well as nitric oxide (NO) production. Interestingly, NIV, but not DON, induced DC necrosis. Moreover, the analysis of the cytokine pattern showed that IL-12 and IL-10 expressions induced by LPS exposure were suppressed by both trichothecenes in a dose-dependent fashion. On the other hand, the secretion of the proinflammatory cytokine TNF-alpha was increased as a direct consequence of DON and NIV exposure. Taken together, our data indicated that the immunotoxicity of NIV and DON was related to the capacity of both trichothecenes to interfere with phenotypic and functional features of maturing DCs.

Phenolic Compounds from Olea europaea L. Possess Antioxidant Activity and Inhibit Carbohydrate Metabolizing Enzymes In Vitro

Phenolic composition and biological activities of fruit extracts from Italian and Algerian Olea europaea L. cultivars were studied. Total phenolic and tannin contents were quantified in the extracts. Moreover 14 different phenolic compounds were identified, and their profiles showed remarkable quantitative differences among analysed extracts. Moreover antioxidant and enzymatic inhibition activities were studied. Three complementary assays were used to measure their antioxidant activities and consequently Relative Antioxidant Capacity Index (RACI) was used to compare and easily describe obtained results. Results showed that Chemlal, between Algerian cultivars, and Coratina, among Italian ones, had the highest RACI values. On the other hand all extracts and the most abundant phenolics were tested for their efficiency to inhibit α-amylase and α-glucosidase enzymes. Leccino, among all analysed cultivars, and luteolin, among identified phenolic compounds, were found to be the best inhibitors of α-amylase and α-glucosidase enzymes. Results demonstrated that Olea europaea fruit extracts can represent an important natural source with high antioxidant potential and significant α-amylase and α-glucosidase inhibitory effects.

Heavy metals in canned tuna from Italian markets.

Fish is a good source of nutrients for humans but can pose a risk to human health because of the possible presence of some xenobiotics such as heavy metals and persistent organic contaminants. Constant monitoring is needed to minimize health risks and ensure product quality and consumer safety. The aim of the present study was to use atomic absorption spectrometry to determine the concentrations of some heavy metals (Hg, Pb, and Cd) in tuna packaged in different kinds of packages (cans or glass) in various countries (Italy and elsewhere). Concentrations of Cd and Hg were within the limits set by European Commission Regulation (EC) No 1881/2006 and in many samples were below the detection limit. Pb concentrations exceeded European limits in 9.8% of the analyzed samples. These results are reassuring in terms of food safety but highlighted the need to constantly monitor the concentrations of heavy metals in fish products that could endanger consumer health.

In vitro study of AFB1 and AFM1 effects on human lymphoblastoid Jurkat T-cell model.

Abstract Aflatoxin B1 (AFB1) is a mycotoxin produced by Aspergillus spp. that can occur as a natural contaminant in foods and feeds of vegetable origin. Post-ingestion, AFB1 can be metabolized in the liver of mammals into hydroxylated aflatoxin M1 (AFM1) that is excreted with milk. Although several studies have been carried out to evaluate effects of AFB1 on the immune system, studies regarding AFM1 are moreover lacking. The aim of the current study was to investigate effects of AFB1 and AFM1 on immune function using a lymphoblastoid Jurkat T-cell line as an experimental model. Both AFB1 and AFM1 produced significant decreases in Jurkat cell proliferation, whereas only minor effects were noted on interleukin (IL)-2 and interferon (IFN)-γ cytokines mRNA expression in stimulated cells that had been pre-incubated with AFB1 and AFM1. Particularly, AFB1, but not AFM1, at the highest concentration (50 µM) induced a marked increase in IL-8 mRNA expression. The results of the current study suggested the existence of a concentration threshold for AFB1 and AFM1 needed to exert biological activity on cell viability and innate immunity.