zearalenone and Necrosis

The purpose of this study was to determine the effects of single and combined administrations of deoxynivalenol (DON) and zearalenone (ZEN) on the histology and ultrastructure of pig liver. The study was performed on immature gilts, which were divided into four equal groups. Animals in the experimental groups received DON at a dose of 12 μg/kg body weight (BW) per day, ZEN at 40 μg/kg BW per day, or a mixture of DON (12 μg/kg BW per day) and ZEN (40 μg/kg BW). The control group received vehicle. The animals were killed after 1, 3, and 6 weeks of experiment. Treatment with mycotoxins resulted in several changes in liver histology and ultrastructure, including: (1) an increase in the thickness of the perilobular connective tissue and its penetration to the lobules in gilts receiving DON and DON + ZEN; (2) an increase in the total microscopic liver score (histology activity index (HAI)) in pigs receiving DON and DON + ZEN; (3) dilatation of hepatic sinusoids in pigs receiving ZEN, DON and DON + ZEN; (4) temporary changes in glycogen content in all experimental groups; (5) an increase in iron accumulation in the hepatocytes of gilts treated with ZEN and DON + ZEN; (6) changes in endoplasmic reticulum organization in the hepatocytes of pigs receiving toxins; (7) changes in morphology of Browicz-Kupffer cells after treatment with ZEN, DON, and DON + ZEN. The results show that low doses of mycotoxins used in the present study, even when applied for a short period, affected liver morphology.

Topics: Animals; Chemical and Drug Induced Liver Injury; Female; Glycogen; Iron; Liver; Necrosis; Sus scrofa; Trichothecenes; Zearalenone

Zearalenone (ZEA) is a nonsteroidal estrogenic mycotoxin produced mainly by Fusarium. ZEA causes reproductive disorders and is both cytotoxic and genotoxic in animals; however, little is known regarding the molecular mechanism(s) leading to ZEA toxicity. Sertoli cells are somatic cells that support the development of spermatogenic cells. The objective of this study was to explore the effects of ZEA on the proliferation, apoptosis, and necrosis of rat Sertoli cells to uncover signaling pathways underlying ZEA cytotoxicity. ZEA reduced the proliferation of rat Sertoli cells in a dose-dependent manner, as indicated by a CCK8 assay, while flow cytometry revealed that ZEA caused both apoptosis and necrosis. Immunoblotting revealed that ZEA treatment increased the ratio of Bax/Bcl-2, as well as the expression of FasL and caspases-3, -8, and -9, in a dose-dependent manner. Collectively, these data suggest that ZEA induced apoptosis and necrosis in rat Sertoli cells via extrinsic and intrinsic apoptotic pathways. This study provides new insights into the molecular mechanisms by which ZEA exhibits cytotoxicity. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1731-1739, 2016.

Topics: Animals; Apoptosis; Caspase 3; Caspase 8; Caspase 9; Cells, Cultured; Estrogens, Non-Steroidal; Fas Ligand Protein; Male; Necrosis; Proto-Oncogene Proteins c-bcl-2; Rats, Sprague-Dawley; Sertoli Cells; Signal Transduction; Zearalenone

This study was designed to evaluate the effect of zearalenone (ZEA) on cell cycle checkpoints cyclin D1 and E2f1 expression at mRNA level and to analyze the estrogen like impact of ZEA on male reproductive system. Thirty mature male rats were randomly assigned into five groups as; control (received 0.5 mL saline normal, i.p.), ZEA-received groups (1, 2 and 4 mg/kg, b.w., i.p.) and 17 β-estradiol-received group (0.1 mg/kg, i.p.). All animals received chemicals for 28 days. Cyclin D1 expression was evaluated both by using PCR and immunohistochemistry staining. The mRNA level of E2f1 was also assessed by PCR. Cellular apoptosis was evaluated by using TUNEL and DNA laddering tests. ZEA, at low and medium doses increased the cellular apoptosis and DNA fragmentation. However, at high dose-received animals, severe necrosis was revealed in germinal epithelium. The medium dose of ZEA exhibited the same phenotype as 17 β-estradiol-showed. Accordingly, the medium dose of ZEA-and 17 β-estradiol significantly up-regulated the expression of cyclin D1 and E2f1 in the testis. In contrast, high dose of ZEA down regulated the expression of both genes at mRNA level. The histomorphometric analyses showed that ZEA in medium and high doses remarkably lowered the tubular differentiation and spermiogenesis indices. In conclusion, our data suggest that ZEA enhances the cellular apoptosis likely via oppositional roles of cyclin D1 and E2F1 checkpoint machineries for cell cycles in testicular tissue. Moreover, at medium dose level ZEA exerts the same pathological phenotype as 17 β-estradiol induces.

Topics: Animals; Apoptosis; Cyclin D1; DNA Fragmentation; Down-Regulation; E2F1 Transcription Factor; Estradiol; Immunohistochemistry; In Situ Nick-End Labeling; Male; Necrosis; Organ Size; Phenotype; Polymerase Chain Reaction; Random Allocation; Rats; Rats, Wistar; RNA, Messenger; Spermatogenesis; Testis; Zearalenone

Zearalenone is a mycotoxin produced mainly by Fusarium. There are numerous incidences of mycotoxicosis in laboratory and domestic animals, especially in pigs. However, little is known about molecular mechanisms of zearalenone toxicity. Granulosa cells are the maximal cell population in follicles, and they play an essential role in the development and maturation of follicles. The objective of this study was to explore the effect of zearalenone at high concentrations on proliferation and apoptosis of porcine granulosa cells and uncover signaling pathway underlying the cytotoxicity of zearalenone. We found that zearalenone reduced the proliferation of porcine granulosa cells in a dose-dependent manner as shown by the MTT assay and zearalenone resulted in an obvious apoptosis and necrosis in porcine granulosa cells as determined by the TUNEL analysis and flow cytometry. In addition, TUNEL assay with caspase inhibitors showed that zearalenone triggered a caspase-3- and caspase-9-dependent apoptotic process in porcine granulosa cells. Fluorescence spectrophotometer displayed that zearalenone led to a loss of mitochondrial transmembrane potential of porcine granulosa cells but enhanced reactive oxygen species (ROS) levels of the cells. Notably, Western blots revealed that caspase-3 and caspase-9 were activated by zearalenone in porcine granulosa cells. Collectively, our results suggest that zearalenone induces apoptosis and necrosis of porcine granulosa cells in a dose-dependent manner via a caspase-3- and caspase-9-dependent mitochondrial pathway. This study thus offers a novel insight into molecular mechanisms by which zearalenone has adverse cytotoxicity on reproduction.

Topics: Animals; Antioxidants; Apoptosis; Caspase 3; Caspase 9; Cell Proliferation; Cells, Cultured; Chromans; Dose-Response Relationship, Drug; Estrogens, Non-Steroidal; Female; Granulosa Cells; In Situ Nick-End Labeling; Mitochondria; Necrosis; Reactive Oxygen Species; Signal Transduction; Swine; Zearalenone

Zearalenone (ZEN) is commonly found in many food commodities and is known to cause reproductive disorders and genotoxic effects. However, the mode of ZEN-induced cell death of macrophages and the mechanisms by which ZEN causes cytotoxicity remain unclear. The present study shows that ZEN treatment reduces viability of RAW264.7 cells in a dose-dependent manner. ZEN causes predominantly necrotic and late apoptotic cell death. ZEN treatment also results in the loss of mitochondrial membrane potential (MMP), mitochondrial changes in Bcl-2 and Bax proteins, and cytoplasmic release of cytochrome c and apoptosis-inducing factor (AIF). Pre-treatment of the cells with either z-VAD-fmk or z-IETD-fmk does not attenuate ZEN-mediated cell death, whereas catalase suppresses the ZEN-induced decrease in viability in RAW264.7 cells. Treating the cells with c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (MAPK), or p53 inhibitor prevented ZEN-mediated changes, such as MMP loss, cellular reactive oxygen species (ROS) increase, and cell death. JNK or p38 MAPK inhibitor inhibited mitochondrial alterations of Bcl-2 and Bax proteins with attendant decreases in cellular ROS levels. Knockdown of AIF via siRNA transfection also diminished ZEN-induced cell death. Further, adenosine triphosphate was markedly depleted in the ZEN-exposed cells. Collectively, these results suggest that ZEN induces cytotoxicity in RAW264.7 cells via AIF- and ROS-mediated signaling, in which the activations of p53 and JNK/p38 play a key role.

Topics: Adenosine Triphosphate; Animals; Apoptosis; Apoptosis Inducing Factor; bcl-2-Associated X Protein; Cell Line; Cytochromes c; JNK Mitogen-Activated Protein Kinases; MAP Kinase Signaling System; Membrane Potential, Mitochondrial; Mice; Mitochondria; Necrosis; p38 Mitogen-Activated Protein Kinases; Proto-Oncogene Proteins c-bcl-2; Reactive Oxygen Species; Tumor Suppressor Protein p53; Zearalenone

Zearalenone is a mycotoxin that is widespread in cereal food. We questioned whether this mycotoxin, administered during known critical exposure periods such as the fetal period and the first days of life, at doses compatible with mean daily intake in humans, could have an effect on mammary gland development in rodents. Wistar female rats were exposed to zearalenone (0.2 μg/kg to 5mg/kg) during the last 14 days of fetal life and the first 5 post-natal days (PND). The mammary tissue was examined for development and maturation by morphologic analyses and immunochemistry. At PND 30, the mean length of terminal buds was significantly enhanced in all of the zearalenone-exposed females (p<0.05). The mammary tissue, as evaluated by scoring of tissue slides, was significantly more differentiated in the 1mg/kg treated group than in controls (p<0.05). At PND 180, mammary tissue was more differentiated in all of the zearalenone treated groups (p<0.05). At six months, 4 of 18 females exposed to 5mg/kg of zearalenone presented mammary hyperplasia lesions. The induction of phenotypic alterations by zearalenone administered in utero and in the neonatal period at doses as low as 0.2 μg/kg suggests that zearalenone could contribute to the induction of breast endocrine disorders.

Topics: Animals; Animals, Newborn; Apoptosis; Cell Differentiation; Cell Proliferation; Endocrine Disruptors; Endothelial Cells; Female; Fetus; Immunohistochemistry; In Situ Nick-End Labeling; Mammary Glands, Animal; Mycotoxins; Necrosis; Phenotype; Pregnancy; Rats; Rats, Wistar; Zearalenone

The mycotoxin zearalenone (ZEA) is a common contaminant of all major cereal grains worldwide with estrogenic and anabolic activity. We investigated the in vitro cytopathic effects of ZEA on freshly isolated human peripheral blood mononuclear cells (PBMC) in relation to proliferation and cell death patterns of untreated and mitogen-activated cells. The higher concentration of 30microg/ml ZEA was found to totally inhibit T and B lymphocyte proliferation from the stimulation with phytohemagglutinin and pokeweed mitogen. The inhibitory effects of ZEA were further related to cell necrosis/apoptosis. Flow cytometry analysis showed a distinct necrotic effect on PBMC, irrespective of mitogen stimulation, whereas apoptotic activity was less evident. Necrosis was observed in both the lymphocyte and monocyte/granulocyte gates. Measurements of ZEA-induced intracellular calcium ion (Ca(2+)) mobilization showed an increase of both Ca(2+) levels and the number of cells with high Ca(2+) only in the monocyte/granulocyte gated cells. Using phenylmethyl sulfonyl fluoride (PMSF), a serine protease inhibitor, and ammonium chloride (NH(4)Cl), a lysosomal inhibitor, both associated with cell necrosis inhibition, we showed that PMSF at 0.05mM and NH(4)Cl at 1 and 10mM reduced the cytopathic effects induced by 30microg/ml ZEA, whereas apoptosis was less affected. Expose of PBMC to 1microg/ml ZEA did not alter the viability of the cells. Our results suggest that high ZEA concentrations in the blood may well exert cytotoxic effects that merit further investigation.

Topics: Ammonium Chloride; Apoptosis; Calcium; Cell Proliferation; Cells, Cultured; Estrogens, Non-Steroidal; Humans; Leukocytes, Mononuclear; Necrosis; Phenylmethylsulfonyl Fluoride; Time Factors; Zearalenone

The efficacy of dietary tamoxifen (TAM) to alleviate the hyperestrogenic effects of the mycotoxin zearalenone (ZEN) was assessed by pathologic examination of the reproductive organs of female mink (Mustela vison). Mink were fed 20 mg/kg ZEN, 10 mg/kg TAM, or 20 mg/kg ZEN + 10 mg/kg TAM from about 2 mo prior to breeding until the kits reached 3 w of age. All female mink fed ZEN mated, but only 25% whelped. No mink fed TAM or TAM + ZEN mated. Postmortem examination revealed moderate to severely distended uteri filled with caseated necrotic substances in the TAM, ZEN and ZEN + TAM fed mink. Histologic examination revealed mild to severe endometrial hyperplasia to uterine atrophy, endometritis, metritis and pyometra. Ovarian follicles were atrophied and degenerated. TAM was not effective in alleviating the hyperestrogenic effects of ZEN but was a potent estrogen agonist in mink.

Topics: Animals; Dose-Response Relationship, Drug; Drug Interactions; Endometrial Hyperplasia; Endometritis; Estrogen Antagonists; Estrogens, Non-Steroidal; Female; Mink; Necrosis; Ovarian Follicle; Random Allocation; Tamoxifen; Uterus; Zearalenone

An outbreak of zearalenone mycotoxicosis occurred between early March and mid-April involving 62 suckling piglets of both sexes. The clinical picture was characterized by edematous swelling and reddening of the vulva, sometimes associated with reddening and/or necrosis of the tail. Six female piglets had congenital lesions of the external genitalia while in the remainder clinical signs appeared 2 to 3 d after birth. No sows ingesting the contaminated feed had signs of hyperestrogenism. The distribution of affected litters showed a correlation with poor hygienical conditions. Zearalenone residues were detected only in feed samples from mangers where the hyperestrogenic syndrome occurred.

Topics: Animal Feed; Animals; Animals, Suckling; Chromatography, High Pressure Liquid; Disease Outbreaks; Eating; Estrogens, Non-Steroidal; Female; Food Contamination; Hyperemia; Male; Mycotoxicosis; Necrosis; Swine; Swine Diseases; Tail; Vulva; Zearalenone

Male weanling rats fed diets containing fermented and unfermented tannin-free sorghum meals naturally contaminated with traces of ochratoxin A, zearalenone, and an unidentified toxic substance developed anorexia. Mean changes in body weight over a 28-day feeding period for rats fed fermented and unfermented sorghum meal and ANCR casein diets of 8% protein were -2.8, +13.8, and +100.5 g, respectively. Rats fed fermented sorghum meal developed alopecia; hematological findings showed a decreased mean corpuscular volume, hypochromic microcytosis, balanced leukopenia, and hypoproteinuria . Histological findings showed testicular hypoplasia of the germinal epithelium and no mature spermatozoa. Necrosis and mineralization in Henle's loop were also observed. No damage was apparent to the liver, cerebellum, cerebrum, spleen, or adrenal glands.

Topics: Animal Feed; Animals; Blood Cell Count; Cats; Cattle; Dogs; Edible Grain; Fermentation; Food Contamination; Guinea Pigs; Kidney; Male; Mycotoxins; Necrosis; Ochratoxins; Rats; Rats, Inbred Strains; Testis; Zearalenone