Forsythoside A protects against Zearalenone-induced cell damage in chicken embryonic fibroblasts via mitigation of endoplasmic reticulum stress.

Zearalenone (ZEA) is a non-steroidal estrogenic mycotoxin that exerts its toxic effects through various damage mechanisms such as oxidative stress, endoplasmic reticulum stress (ERS), mitochondrial damage, cell cycle arrest, and apoptosis. At present, there are few studies on drugs that can rescue ZEA-induced chicken embryonic fibroblasts damage. Forsythoside A (FA) is one of effective ingredients of traditional Chinese medicine that plays a role in various biological functions, but its antitoxin research has not been investigated so far. In this study, in vitro experiments were carried out. Chicken embryo fibroblast (DF-1) cells was used as the research object to select the appropriate treatment concentration of ZEA and examined reactive oxygen species (ROS), mitochondrial membrane potential, ERS and apoptosis to investigate the effects and mechanisms of FA in alleviating ZEA-induced cytotoxicity in DF-1 cells. Our results showed that ZEA induced ERS and activated the unfolded protein response (UPR) leading to apoptosis, an apoptotic pathway characterized by overproduction of Lactate dehydrogenase (LDH), Caspase-3, and ROS and loss of mitochondrial membrane potential. We also demonstrated that FA help to prevent ERS and attenuated ZEA-induced apoptosis in DF-1 cells by reducing the level of ROS, downregulating GRP78, PERK, ATF4, ATF6, JNK, IRE1, ASK1, CHOP, BAX expression, and up-regulating Bcl-2 expression. Our results provide a basis for an in-depth study of the mechanism of toxic effects of ZEA on chicken cells and the means of detoxification, which has implications for the treatment of relevant avian diseases.

Enhanced glutathione production protects against zearalenone-induced oxidative stress and ferroptosis in female reproductive system.

Zearalenone (ZEN, a widespread fusarium mycotoxin) causes evoked oxidative stress in reproductive system, but little is known about whether this is involved in ferroptosis. Melatonin, a well-known antioxidant, has demonstrated unique anti-antioxidant properties in several studies. Here, this study was aimed to investigate whether ZEN-induced oxidative stress in female pig's reproductive system was involved in ferroptosis, and melatonin was then supplemented to protect against ZEN-induced abnormalities in vitro cell models [human granulosa cell (KGN) and mouse endometrial stromal cell (mEC)] and in vivo mouse model. According to the results from female pig's reproductive organs, ZEN-induced abnormalities in vulvar swelling, inflammatory invasion and pathological mitochondria, were closely linked with evoked oxidative stress. Using RNA-seq analysis, we further revealed that ZEN-induced reproductive toxicity was due to activated ferroptosis. Mechanistically, by using in vitro cell models (KGN and mEC) and in vivo mouse model, we observed that ZEN exposure resulted in oxidative stress and ferroptosis in a glutathione-dependent manner. Notably, these ZEN-induced abnormalities above were alleviated by melatonin supplementation through enhanced productions of glutathione peroxidase 4 and glutathione. Herein, the present results suggest that potential strategies to improve glutathione production protect against ZEN-induced reproductive toxicity, including oxidative stress and ferroptosis.

Protective effects of melanoidins from black garlic on zearalenone-induced toxicity in zebrafish embryonic developmental model.

Zearalenone (ZEN), a ubiquitous mycotoxin that widely occurs in grain and foodstuff may induce serious toxic effects after accumulation in vivo. Melanoidins (MLDs) have shown multiple bio-functional properties such as antioxidant, anti-bacterial and prebiotic activities. Black garlic exhibits several advantages over fresh garlic related to health improvement. In this study, the alleviative effects of black garlic MLDs on ZEN-induced toxicity and the potential mechanisms were studied using zebrafish embryonic developmental model. The results showed that MLDs restored the ZEN-induced adverse influences on zebrafish embryonic development, including delay in hatching time, morphological abnormality and the impairment of nervous development. Further studies showed that MLDs significantly inhibited the ZEN-induced production of reactive oxygen species (ROS) and enhanced the intrinsic antioxidant ability by increasing the activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and catalase (CAT) and the content of glutathione (GSH). In addition, co-exposure of MLDs significantly inhibited the ZEN-stimulated cellular apoptosis in zebrafish larvae through down-regulation of pro-apoptotic genes of bax, caspase-3 and caspase-9 and up-regulation of anti-apoptotic gene bcl-2. Moreover, MLDs inhibited the in vivo accumulation of ZEN in zebrafish larvae. To sum up, MLDs attenuated the ZEN-induced zebrafish embryonic developmental toxicity through suppression of the oxidative stress and intervention on mitochondria apoptosis pathway as well as inhibiting the absorption of ZEN in zebrafish embryos/larvae. The results suggest that black garlic MLDs have potential to be used as a functional ingredient against the adverse effects of exogenous toxins.

Cytoprotective, Antiproliferative, and Anti-Oxidant Potential of the Hydroethanolic Extract of Fridericia chica Leaves on Human Cancer Cell Lines Exposed to α- and ß-Zearalenol.

Fridericia chica (Bignoniaceae) is a Colombian Caribbean plant with numerous health benefits, including properties such as wound healing, immune system stimulation, and antioxidant capacity, among others. Mycotoxins alpha-zearalenol (α-ZEL) and beta-zearalenol (ß-ZEL) are phase I metabolites of zearalenone, a natural product involved in endocrine disruption and cell proliferation processes. This study aimed to investigate the cytotoxic potential of the hydroethanolic extract of F. chica leaves (HEFc) and determine their protective effects against proliferation induced by α-ZEL and ß-ZEL on human hepatoma HepG2, lung cancer Calu-1, and primary normal human epidermal keratinocytes, neonatal (HEKn). The cytotoxicity of HEFc was measured in a range from 4 to 1000 µg/mL and from 0.4 to 100 µM for both α-ZEL and ß-ZEL. Cell production of intracellular ROS was monitored using the H2-DCFDA probe. The cells exposed to HEFc presented IC50 of 128, 249, and 602 µg/mL for the HepG2, Calu-1, and HEKn cells, respectively. A greater selectivity was seen in HepG2 cells [selectivity index (SI) = 3.5] than in Calu-1 cells (SI = 2.4). Cells treated with mycotoxins remained viable during the first day, and cell proliferation increased at low tested concentrations (0.4-6.3 µM) in all three cell lines. However, after 48 h treatment, cells exposed to 50 and 100 µM of α-ZEL and ß-ZEL displayed decreased viability. HEFc at 16 µg/mL was able to give some protection against cytotoxicity induced by high concentrations of ß-ZEL in HepG2, reducing also cell proliferation elicited at low levels of α-ZEL and ß-ZEL. ROS production was not observed in cells treated with this HEFc concentration; however, it prevented ROS formation induced by treatment with 50 µM α-ZEL or ß-ZEL. In summary, HEFc isolated from plants grown in northern Colombia displayed promising results against cell proliferation and oxidative stress caused by mycotoxins.

Protective effect of glucosamine on zearalenone-induced reproductive toxicity and placental dysfunction in mice.

This study was conducted to determine the effects of glucosamine (GlcN) on zearalenone (ZEA)-induced reproductive toxicity and placental dysfunction in mice. The pregnant mice were randomly divided into one of the four groups, such as the control group, the ZEA group, the GlcN group, and the GlcN plus ZEA group. Reproductive toxicity was induced by consecutive gavages of ZEA at 5 mg/kg body weight during gestational days (GDs 0-14) and in the presence or absence of oral administration of GlcN (0.5 mM). The results showed that GlcN significantly alleviated the decrease of growth performance induced by ZEA exposure of pregnant mice. Meanwhile, ZEA ingestion significantly reduced the number and weight of fetuses, and reduction of placenta weight. Moreover, results of blood biochemical markers indicated that ZEA exposure led to increased oxidative stress levels in pregnant mice. Further analyses demonstrated that ZEA inhibited placental development, resulted in placental inflammation, increased the expression of pro-apoptotic proteins, and decreased the expression of placental tight junction proteins, which were reversed by the administration of GlcN. Results of western blot revealed that GlcN reversed ZEA-mediated phenotype by activating PI3K, while inhibiting MAPK signaling pathway. All these findings showed that GlcN was effective in the protection against ZEA-induced placental dysfunction and reproductive toxicity in pregnant mice. Supplementation of GlcN might be potential nutritional intervention with an ability to alleviate ZEA-induced toxicity in pregnant mice.

Toxicity of zearalenone and its nutritional intervention by natural products.

Zearalenone (ZEN) is a toxic secondary metabolite mainly produced by fungi of the genus Fusarium, and is often present in various food and feed ingredients such as corn and wheat. The structure of ZEN is similar to that of natural estrogen, and it can bind to estrogen receptors and has estrogenic activity. Therefore, it can cause endocrine-disrupting effects and promote the proliferation of estrogen receptor-positive cell lines. In addition, ZEN can cause oxidative damage, endoplasmic reticulum stress, apoptosis, and other hazards, resulting in systemic toxic effects, including reproductive toxicity, hepatotoxicity, and immunotoxicity. In the past few decades, researchers have tried many ways to remove ZEN from food and feed, but it is still a challenge to eliminate it. In recent years, natural compounds have become of interest for their excellent protective effects on human health from food contaminants. Researchers have discovered that natural compounds often used as dietary supplements can effectively alleviate ZEN-induced systemic toxic effects. Most of the compounds mitigate ZEN-induced toxicity through antioxidant effects. In this article, the contamination of food and feed by ZEN and the various toxic effects and mechanisms of ZEN are reviewed, as well as the mitigation effects of natural compounds on ZEN-induced toxicity.

Estrogenic in vitro evaluation of zearalenone and its phase I and II metabolites in combination with soy isoflavones.

Humans and animals are exposed to multiple substances in their food and feed that might have a negative health impact. Among these substances, the Fusarium mycoestrogen zearalenone (ZEN) and its metabolites α-zearalenol (α-ZEL) and α-zearalanol (α-ZAL) are known to possess endocrine disruptive properties. In a mixed diet or especially animal feed, these potential contaminants might be ingested together with naturally occurring phytoestrogens such as soy isoflavones. So far, risk assessment of potential endocrine disruptors is usually based on adverse effects of single compounds whereas studies investigating combinatorial effects are scarce. In the present study, we investigated the estrogenic potential of mycoestrogens and the isoflavones genistein (GEN), daidzein (DAI) and glycitein (GLY) as well as equol (EQ), the gut microbial metabolite of DAI, in vitro alone or in combination, using the alkaline phosphatase (ALP) assay in Ishikawa cells. In the case of mycoestrogens, the tested concentration range included 0.001 to 10 nM with multiplication steps of 10 in between, while for the isoflavones 1000 times higher concentrations were investigated. For the individual substances the following order of estrogenicity was obtained: α-ZEL > α-ZAL > ZEN > GEN > EQ > DAI > GLY. Most combinations of isoflavones with mycoestrogens enhanced the estrogenic response in the investigated concentrations. Especially lower concentrations of ZEN, α-ZEL and α-ZAL (0.001-0.01 nM) in combination with low concentrations of GEN, DAI and EQ (0.001-0.1 µM) strongly increased the estrogenic response compared to the single substances.

[Single-laboratory Validation of an Analytical Method for Detarmination of Fusarium toxins in Buckwheat and Job's Tears].

Fusarium species infect the major cereals consumed as food and feed, contaminating them with various toxic secondary metabolites known as toxins. Among these toxins, which include trichothecenes, zearalenone (ZEA), and fumonisins, the type-B trichothecene deoxynivalenol (DON) is generally considered as the most important. The present study evaluates an analytical method for the detection and quantification of multiple Fusarium toxins, namely, DON, acetyl forms of DON (3-Ac-DON and 15-Ac-DON), a glycoside form of DON (DON-3G), and other Fusarium toxins (nivalenol, an acetyl form of NIV (fusarenonX), T-2 and HT-2 toxins, diacetoxyscirpenol, and ZEA) in Job's tears and buckwheat.

Costus speciosus extract protects against the oxidative damage of zearalenone via modulation of inflammatory cytokines, Nrf2 and iNOS gene expression in rats.

Zearalenone (ZEN) is a non-steroidal estrogenic mycotoxin that induces severe health disturbances in humans and animals. This study aimed to determine the bioactive compounds in Costus speciosus extract (CSE) using GC-MS and evaluate its protective capability against ZEN-induced oxidative damage, genotoxicity, and cytotoxicity in rats. Six groups of male Sprague Dawley rats were treated orally for 15 days including the control group, CSE-treated groups at low (200 mg/kg b. w) or high (400 mg/kg b. w) dose, ZEN-treated group (40 µg/kg b. w), and the groups treated with ZEN plus the low or the high dose of CSE. Blood and tissue samples were collected for different assays and pathological analyses. The results of GC-MS indicated the identification of 6 compounds and Azulene was the major. Animals that received ZEN showed severe disturbances in serum biochemical, cytokines, oxidative stress indicators, mRNA expression of iNOS, Nrf2, and inflammatory-related genes. ZEN also increased micronucleated polychromatic erythrocytes (MNPCEs) and comet tail formation in bone marrow cells along with the disturbances in the histological architecture of the liver and kidney. Co-administration of CSE plus ZEN could normalize the majority of the tested parameters and the histological picture at a dose as low as 200 mg/kg b. w. Therefore, CSE protects against ZEN toxicity via its antioxidant activity, modulation of iNOS, inflammatory-related genes, and the Nrf2 pathway and it could be used in the endemic regions.

Effects of zearalenone on liver development, antioxidant capacity and inflammatory factors of prepubertal gilts.

Zearalenone (ZEA) is a kind of mycotoxin that pose great threat to the liver of human and livestock due to its toxicity to eukaryotic cells, however, its toxicity mechanism on prepubertal gilts liver development and function is not known. The study aimed to examine the effects of ZEA on liver development, antioxidant capacity and inflammatory factors of prepubertal gilts. Forty-eight prepubertal gilts (Landrace ×Yorkshire) were randomly divided into four groups: three treatment (T1, T2 and T3) groups and a control group. Prepubertal gilts in the control group were fed with basal diet, and those in T1, T2 and T3 groups were fed with basal diets supplemented with low, medium and high doses (200 µg/kg, 800 µg/kg and 1,600 µg/kg, respectively) of ZEA during the experiment period. The results showed that diets supplemented with ZEA significantly increased the activity of alanine aminotransferase of serum in the T3 group (p < 0.05). Besides, compared to the control group, the activities of total antioxidant capacity, superoxide dismutase, the content of tumour necrosis factor-alpha of liver in the T3 group and the relative expression level of manganese-superoxide dismutase mRNA of liver in the T2 group were significantly reduced (p < 0.05). We also performed correlation analysis among caecal microorganisms and antioxidant enzyme activities and inflammatory factor concentrations of liver. In conclusion, diets supplemented with ZEA has no obvious effect on liver development, but it can cause liver damage.

The Antagonistic Effect of Glutamine on Zearalenone-Induced Apoptosis via PI3K/Akt Signaling Pathway in IPEC-J2 Cells.

Zearalenone (ZEN) is a non-steroidal estrogen mycotoxin produced by Fusarium fungi, which inevitably exists in human and animal food or feed. Previous studies indicated that apoptosis seems to be a key determinant of ZEN-induced toxicity. This experiment aimed to investigate the protective effects of Glutamine (Gln) on ZEN-induced cytotoxicity in IPEC-J2 cells. The experimental results showed that Gln was able to alleviate the decline of cell viability and reduce the production of reactive oxygen species and calcium (Ca2+) induced by ZEN. Meanwhile, the mRNA expression of antioxidant enzymes such as glutathione reductase, glutathione peroxidase, and catalase was up-regulated after Gln addition. Subsequently, Gln supplementation resulted in the nuclear fission and Bad-fluorescence distribution of apoptotic cells were weakened, and the mRNA expression and protein expression of pro-apoptotic genes and apoptotic rates were significantly reduced. Moreover, ZEN reduced the phosphorylation Akt, decreased the expression of Bcl-2, and increased the expression of Bax. Gln alleviated the above changes induced by ZEN and the antagonistic effects of Gln were disturbed by PI3K inhibitor (LY294002). To conclude, this study revealed that Gln exhibited significant protective effects on ZEN-induced apoptosis, and this effect may be attributed to the PI3K/Akt signaling pathway.

Efficacy of lactic acid bacteria supplementation against Fusarium graminearum growth in vitro and inhibition of Zearalenone causing inflammation and oxidative stress in vivo.

Fusarium graminearum invasion and Zearalenone (ZEN)-mycotoxin contamination are considered the most global threat to food and feed. This study investigates the effect Lactobacillus plantarum MON03 viable cells (LPVC) and LP free cells supernatant (LPFCS) against Fusarium graminearum growth and ZEN production in vitro and evaluates if treatment with LP viable cells can counteract the negative effect of ZEN on inflammation and oxidative stress in mesenteric lymph nodes and serum biochemical parameters in mice. For the in vitro study, 7 days of LPVC, LPFCS and F. graminearum co-incubation at different concentrations was done in order to determine the antifungal activity and ZEN- production inhibition. Regarding the in vivo study, Balb/c mice were treated as following: Control, ZEN group, LP group and ZEN + LP group for 30 days. In vitro, LPVC showed an excellent antifungal activity after 7 days of co-incubation (103 CFU/ml). LPVC was succeeded also to inhibit ZEN production by the fungi. In vivo, ZEN has shown an important oxidative damage. As a result of the exposure to ZEN, an increase cytokines, as effectors of an inflammatory response, were observed in the mesenteric lymph nodes (MLN) of intoxicated mice. In parallel, a serum biochemical change was also observed. LPVC induced a reduction of ZEN-induced oxidative stress and counteracts also the biochemical parameters damage and the inflammatory markers increased by ZEN. LPVC can be valorized as an anti-cating agent in the vitro and in the gastro-intestinal tract to decrease ZEN-toxic effects.

Metaproteomics Reveals Alteration of the Gut Microbiome in Weaned Piglets Due to the Ingestion of the Mycotoxins Deoxynivalenol and Zearalenone.

The ingestion of mycotoxins can cause adverse health effects and represents a severe health risk to humans and livestock. Even though several acute and chronic effects have been described, the effect on the gut metaproteome is scarcely known. For that reason, we used metaproteomics to evaluate the effect of the mycotoxins deoxynivalenol (DON) and zearalenone (ZEN) on the gut microbiome of 15 weaned piglets. Animals were fed for 28 days with feed contaminated with different concentrations of DON (DONlow: 870 µg DON/kg feed, DONhigh: 2493 µg DON/kg feed) or ZEN (ZENlow: 679 µg ZEN/kg feed, ZENhigh: 1623 µg ZEN/kg feed). Animals in the control group received uncontaminated feed. The gut metaproteome composition in the high toxin groups shifted compared to the control and low mycotoxin groups, and it was also more similar among high toxin groups. These changes were accompanied by the increase in peptides belonging to Actinobacteria and a decrease in peptides belonging to Firmicutes. Additionally, DONhigh and ZENhigh increased the abundance of proteins associated with the ribosomes and pentose-phosphate pathways, while decreasing glycolysis and other carbohydrate metabolism pathways. Moreover, DONhigh and ZENhigh increased the abundance of the antioxidant enzyme thioredoxin-dependent peroxiredoxin. In summary, the ingestion of DON and ZEN altered the abundance of different proteins associated with microbial metabolism, genetic processing, and oxidative stress response, triggering a disruption in the gut microbiome structure.

Dietary garlic and chitosan enhanced the antioxidant capacity, immunity, and modulated the transcription of HSP70 and Cytokine genes in Zearalenone-intoxicated European seabass.

The present study was performed to evaluate the toxic effects of feed-born zearalenone (ZEN) on antioxidative status, immunity, transcriptomic responses of European seabass, and the modulating roles of dietary garlic and/or chitosan powders. Fish (30.7 ± 0.6 g) were randomly arranged in five experimental groups (in triplicates), whereas the first group was fed on the control diet only without any supplements (control), and the second group was fed on the basal diet contaminated with ZEN (0.725 mg/kg diet). Three other groups were fed on ZEN-contaminated diets and simultaneously supplemented with garlic powder (GP) (30 g/kg diet) (ZEN + GP), chitosan powder (CH) (10 g/kg diet) (ZEN + CH), and a mixture of GP and CH (ZEN + GP + CH). Fish were fed on the experimental diets thrice a day for 4 weeks. Two-way ANOVA revealed a gradual decline in serum superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) activities in the ZEN group reaching their lowest levels at the 4th week. Contrariwise, malondialdehyde levels were markedly higher in the ZEN group reaching their highest values at the end of the experiment. A significant decline of total immunoglobulins (P < 0.05) was observed in the serum of the ZEN group, especially after the 4th week. Moreover, significant down-regulation of interleukin-4 (IL-4) and interleukin 1 beta (IL-1ß) genes (P < 0.05) alongside significant up-regulation of tumor necrosis factor-alpha (TNF-α) and heat shock protein 70 (HSP70) genes (P < 0.05) in the liver and anterior kidney of ZEN-intoxicated group. Interestingly, dietary supplementation with GP and CH significantly attenuated ZEN-induced oxidative stress, immunosuppression, and modulated transcriptomic responses of ZEN-exposed fish. Moreover, combined dietary supplementation of both feed additives resulted in better effects than each one alone.

Modified hydrated sodium calcium aluminosilicate-supplemented diet protects porcine oocyte quality from zearalenone toxicity.

Zearalenone (ZEN) is one of the most common mycotoxins produced by fungus in contaminated feed. ZEN has multiple toxicities, including reproductive toxicity of domestic animals, particularly pigs. However, studies on the effects of ZEN on ovary/oocytes have been primarily based on in vitro experiments, and there is still no evidence from porcine in vivo models due to multiple limitations. Moreover, no report has investigated the effect of hydrated sodium calcium aluminosilicate (HSCAS) as a supplement on pig oocyte quality. In the present study, we fed pigs a 1.0 mg/kg ZEN-contaminated diet for 10 days. The results showed that pigs fed ZEN presented reduced oocyte-cumulus cell interactions, an increase in the number of denuded oocytes in ovaries, a decrease in the number of oocytes in each ovary, and an increase in the oocyte death rate. Oocytes from ZEN-exposed pigs exhibited a delayed cell cycle and abnormal cytoskeletal dynamics during meiotic maturation, which could be due to oxidative stress-induced autophagy. Moreover, we also show that supplementing the ZEN-contaminated diet with modified HSCAS effectively protected porcine oocyte quality. Taken together, our study provides in vivo data demonstrating the protective effects of HSCAS against ZEN toxicity in porcine oocytes.

Selenium Protects against Zearalenone-Induced Oxidative Stress and Apoptosis in the Mouse Kidney by Inhibiting Endoplasmic Reticulum Stress.

This study assessed the molecular mechanism of selenium (Se) protecting against kidney injury induced by zearalenone (ZEA) in mice. The experimental mice were divided into 4 groups including the control group, the Se group, the ZEA group, and the Se+ZEA group; ZEA and Se were administered orally for 28 days. The changes in renal biochemical index (BUN, UA, and CRE), biochemical change of kidney damage such as BUN, UA, and CRE, and oxidative damage such as MDA, T-SOD, and GSH-Px were investigated. Pathological sections and TUNEL staining were used to analyze renal pathological changes and cell apoptosis. qRT-PCR and Western blot were employed to detect the expression of genes and proteins which were related with endoplasmic reticulum stress. The results showed that ZEA increased the concentration of BUN, UA, and CRE and the content of MDA and decreased the activities of T-SOD and GSH-Px in the mouse kidneys. However, Se reversed above changes of the biochemical and antioxidant indexes of renal injury. Moreover, the results also showed that ZEA can increase the expression of Bax, caspase-12, caspase-3, Bip, CHOP, JNK protein, and mRNA and decrease the expression of Bcl-2 protein and mRNA. But Se reversed these proteins and genes related to endoplasmic reticulum stress and apoptosis. It can be concluded that Se protected against the kidney damage induced by ZEA. Se may protect the kidney from ZEA-induced apoptosis and oxidative stress by inhibiting ERS.

Zearalenone and Metabolites in Livers of Turkey Poults and Broiler Chickens Fed with Diets Containing Fusariotoxins.

Zearalenone (ZEN) and metabolites were measured in livers of turkeys and broilers fed a control diet free of mycotoxins, a diet that contained 0.5 mg/kg ZEN (ZEN diet), and a diet that contained 0.5, 5, and 20 mg/kg of ZEN, fumonisins, and deoxynivalenol, respectively (ZENDONFB diet). The feed was individually distributed to male Grade Maker turkeys from the 55th to the 70th day of age and to male Ross chickens from the 1st to the 35th day of age, without any signs of toxicity. Together, the free and conjugated forms of ZEN, α- and ß-zearalenols (ZOLs), zearalanone (ZAN), and α- and ß-zearalanols (ZALs) were measured by UHPLC-MS/MS with [13C18]-ZEN as an internal standard and immunoaffinity clean-up of samples. ZAN and ZALs were not detected. ZEN and ZOLs were mainly found in their conjugated forms. α-ZOL was the most abundant and was found at a mean concentration of 2.23 and 1.56 ng/g in turkeys and chickens, respectively. Consuming the ZENDONFB diet significantly increased the level of total metabolites in the livers of chickens. Furthermore, this increase was more pronounced for the free forms of α-ZOL than for the conjugated forms. An investigation of the presence of ZEN and metabolites in muscle with the methods validated for the liver failed to reveal any traces of these contaminants in this tissue. These results suggest that concomitant dietary exposure to deoxynivalenol (DON) and fumonisins (FB) may alter the metabolism and persistence of ZEN and its metabolites in the liver.

Melatonin alleviates defects induced by zearalenone during porcine embryo development.

Zearalenone (ZEA), which is produced by several fusarium mycotoxins, is found in animal feed and food products, and can exert estrogen-like activity. Melatonin (MT) is emerging as a supplement that can fight the toxic effects of mycotoxins. With a variety of physiological functions that play crucial roles in the development of animal germ cells and embryos, melatonin regulates circadian rhythms and has an anti-inflammatory and anti-oxidative role. This study investigated the protective effects of melatonin against ZEA in porcine early embryonic development. Our results showed that ZEA adversely affected this development, while melatonin supplementation ameliorated the toxic effects. ZEA exposure increased oxidative stress and impaired mitochondrial function, which may affect blastocyst formation. Moreover, we found that ZEA exposure promotes apoptosis, DNA damage, and autophagy in porcine blastocysts. The toxic effects of ZEA on early embryos may be the result of oxidative stress-mediated early apoptosis, while melatonin treatment significantly improved these phenotypes in ZEA-exposed porcine early embryos. Taken together, our results indicate that melatonin has a protective effect on defects caused by ZEA during early porcine embryonic development.

Compound probiotics alleviating aflatoxin B1 and zearalenone toxic effects on broiler production performance and gut microbiota.

In order to alleviate toxic effects of aflatoxins B1 (AFB1) and zearalenone (ZEA) on broiler production performance and gut microbiota, three kinds of compound probiotics (CP) were selected. The optimal ratios of Bacillus subtilis, Lactobacillus casei and Candida utilis in broiler diets were 7, 5 and 6 log CFU/g for ZEA biodegradation (CP1); 6, 7 and 7 log CFU/g for AFB1 biodegradation (CP2); 7, 6 and 7 log CFU/g for ZEA + AFB1 biodegradation (CP3). A total of 350 1-day-old Ross broilers were randomly divided into 7 groups. Group A was the basal diet, group B-G contained ZEA, AFB1, ZEA + AFB1, ZEA + CP1, AFB1+CP2, ZEA + AFB1+CP3, respectively. The experiment showed that AFB1 or AFB1+ZEA significantly decreased broiler production performance, damaged liver and jejunum, increased mycotoxin residues in broiler body; however, three kinds of compound probiotics additions could alleviate mycotoxin negative effects on the above parameters (p < 0.05). The gut microbiota analysis indicated that AFB1+ZEA increased jejunal microbial richness, but which were decreased to almost the same level as the control group by CP3 addition. CP3 addition significantly increased jejunal Firmicutes and Lactobacillus aviarius abundances. The correlative analysis showed that gut Lactobacillus aviarius abundance was positively correlated with average daily gain (ADG) of broilers (p < 0.05), while AFB1+ZEA addition decreased its relative abundance, indicating that CP3 addition increased broiler growth by increasing Lactobacillus aviarius abundance. AFB1 and ZEA residues in broiler body were negatively correlated with the gut beneficial bacterial abundances (p < 0.01), but positively correlated with the potentially harmful bacterial abundances (p < 0.05), which inferred that CP3 addition could decrease mycotoxin residues through positively regulating gut relative bacterial abundances. In conclusion, compound probiotics could keep gut microbiota stable, degrade mycotoxins, alleviate histological lesions, increase production performance and reduce mycotoxin toxicity for broilers.

Effects of zearalenone-induced oxidative stress and Keap1-Nrf2 signaling pathway-related gene expression in the ileum and mesenteric lymph nodes of post-weaning gilts.

Zearalenone (ZEA) contamination of feed affects animal husbandry and the human health. Currently, the molecular mechanism underlying small intestine-related diseases caused by ZEA-induced oxidative stress is not well understood. In this study, we aimed to identify the mechanisms involved in ZEA (0.5-1.5 mg/kg)-induced oxidative stress in the ileum and mesenteric lymph nodes (MLNs) and the role of the Kelch-like erythroid cell-derived protein with CNC homology-associated protein 1 (Keap1)-nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway in post-weaning gilts. Forty post-weaning gilts (Landrace × Yorkshire × Duroc) with an average body weight of 14.01 ± 0.86 kg were randomly allocated to four groups and fed a corn-soybean meal basal diet supplemented with < 0.1, 0.5, 1.0, or 1.5 mg/kg ZEA. The results showed that the activity of total superoxide dismutase and glutathione peroxidase decreased (p < 0.05) linearly and quadratically and that the content of malondialdehyde increased (p < 0.05) quadratically in the ileum and MLNs with increasing ZEA in the diet. Immunohistochemical analysis showed that the expression of Nrf2 and glutathione peroxidase 1 (Gpx1) immunoreactive proteins in the ileum and MLNs were significantly enhanced with increasing ZEA. The relative mRNA and protein expression of Nrf2, Gpx1, quinone oxidoreductase 1 (Nqo1), hemeoxygenase 1 (Ho1), modifier subunit of glutamate-cysteine ligase (Gclm), and catalytic subunit of glutamate-cysteine ligase (Gclc) increased (p < 0.05) linearly and quadratically, and the relative mRNA and protein expression of Keap1 decreased (p < 0.05) linearly and quadratically in the ileum with increasing ZEA concentrations in the diet. Further, the relative mRNA and protein expression of Nrf2 and Gpx1 increased (p < 0.05) linearly and quadratically, and the relative mRNA and protein expression of Nqo1, Ho1, and Gclm decreased (p < 0.05) quadratically in the MLNs as ZEA concentrations increased in the diet. Our results provide valuable genetic information on ZEA-induced oxidative stress in the ileum and MLNs of post-weaning gilts and have elucidated the key regulatory genes involved in the Keap1-Nrf2 signaling pathway. Results indicated that the Keap1-Nrf2 signaling pathway might be a key target to further prevent and treat ZEA-induced injury to the ileum in post-weaning gilts.