Study Of Reproductive Toxicity Of Zearalenone And Ameliorative Effects Of Silymarin In Rats

Zearalenone?ZEN?is an oestrogenic mycotoxin commonly found in food and feed products and can affect reproduction and development in both humans and animals.Good reproductive performance is the basis for the development of swine industry.Maternal consumption of ZEN can reduce litter size and inhibit fetal development,furthermore,ZEN may cause toxicity on both mothers and offspring.Exploring the reproductive toxicities and long-term effects of ZEN on offspring is essential for the rational use of F1livestock breeding values.Meanwhile,myctoxins detoxification has always been the focus of public health research.Silymarin?Sily?,a natural plant extract,is reported to be antioxidative,antiinflammatory,and anticarcinogenic.Whatsmore,as a flavonoid,Sily has the potential to bind to and regulate the ESR and exert estrogenic agonistic or antagonistic activities,which can be related to the detoxification of ZEN.Therefore,our study aimed to determine the reproductive toxicities of ZEN on maternal rats and the F1offspring and to incestigate the ameliorative effects of dietary supplementation of silymarin on ZEN-induced hepatotoxic and reproductive toxicities.Experiment 1 Reproductive toxicities of ZEN on maternal rats and F1 offspringSixty-four pregnant rats were divided into 4 groups and exposed to feed contaminat-ed with ZEN?0,5,10,and 20 mg/kg feed?through gestational days?GDs?0-20.Serum and tissue samples were collected on GD 20,postnatal day?PND?21 and PND 63.The reproductive toxicities of ZEN on maternal rats,F1 female and F1 male offspring were studied.The main results are as follows:1.Gstational exposure of ZEN induced dose-dependant toxicities on pregnant rats.No effects were found in 5 mg/kg ZEN treated rats,while 20 mg/kg ZEN significantly decreased the newborn viability?P<0.05?.Meanwhile,20 mg/kg ZEN significantly increased serum alanine aminotransferase?ALT?and aspartate aminotransferase?AST?activities?27.9%and 80.7%?.In addition,10 and 20 mg/kg ZEN also significantly reduced serum glutathione transferase?GSH?and total superoxide dismutase?T-SOD?but increased malondialdehyde?MDA?levels?P<0.05?.ZEN treatment significantly increased maternal serum follicle stimulating hormone?FSH?and decreased estradiol?E₂?levels?P<0.05?.Moreover,10 and 20 mg/kg ZEN caused a decrease in placental trophoblast cells,and a significant increase in intercellular connections.These pathological changes of placenta were accompanied with a significant inhibition of Esr1but upregulation of 3?-HSD?P<0.05?.Besides,ZEN also decreased ABCb1 and ABCc1but increase ABCc5 gene expressions in placenta.2.Gestational exposure to ZEN also induced reproductive toxicities in F1 male and female offspring.20 mg/kg ZEN significantly?P<0.05?decreased the birth weight and viability of F1 newborn rats.Meanwhile,10 and/or 20 mg/kg ZEN exposure significantly reduced?P<0.05?Esr1,gonadotropin-releasing hormone receptor?GnRHr?,and ATP binding cassette transporters c5 in foetal and weaned F1 brains.In F1 adult female rats,10 and 20 mg/kg ZEN diets increased follicle-stimulating hormone concentrations but decreased?P<0.05?oestradiol in both maternal and F1 adult rats,and induced significant follicular atresia and a thinning uterine layer in F1 female adult rats.However for F1 male rats,adult testis weight increased with seminiferous tubules atrophy as well as decreased spermatocytes and mature sperms?35%and 31%?in ZEN-treated rats?P<0.05?.Furthermore,these impairments concurred with the inhibited mRNA and protein levels of oestrogen receptor-alpha?Esr1?,3?-hydroxysteroid dehydrogenase?HSD?and StAR in both mRNA and protein levels in weaned and adult reproductive organs?ovary,uterus and/or testis?.Experiment 2 Ameliorative effects of silymarin on hepatotoxicity and reproductive toxicity induced with zearalenone in ratsThirty-five 21-d-old female Sprague-Dawley rats?n=7/diet?were fed a control diet?Ctrl?or Ctrl plus 20 mg ZEN/kg or Ctrl plus 20 mg ZEN/kg with 100,200,or 500 mg silymarin/kg for 6 wk.Serum,livers,ovaries,and uterus were collected at week 6 for biochemistry,hormone,and redox status and selected gene and protein assays.The results showed that dietary Silymarin supplementation alleviates zearalenone-induced hepatotoxicity and reproductive toxicity in rats.The consumption of ZEN decreased?P<0.05?the final body weight by 17.9%,induced liver injury,increased?P<0.05?aspartate aminotransferase and alkaline phosphatase activities,and decreased?P<0.05?total protein and albumin concentrations in serum by 16.7–40.6%.ZEN also caused reproductive toxicity,including decreased?P<0.05?E₂ and increased?P<0.05?FSH concentrations in serum by 12.7–46.3%and induced histopathologic alterations in the liver,ovaries,and uterus.Interestingly,these alterations induced by ZEN were alleviated?P<0.05?by silymarin supplementation at 100,200,and 500 mg/kg.Moreover,silymarin supplementation at the 3 doses mitigated?P<0.05?ZEN-induced impairment in hepatic glutathione peroxidase activity,total antioxidant capacity,and MDA concentration by17.6–100%.Meanwhile,silymarin supplementation at all doses upregulated?P<0.05?phospho-ribosomal protein S6 kinase 1?p-p70S6K?and 3?-hydroxysteroid dehydrogenase?3?-HSD?by 43.0–121%but downregulated?P<0.05?AMP-activated protein kinase?AMPK?and 3?-hydroxysteroid dehydrogenase?3?-HSD?in the liver relative to the ZEN group by 11.2–40.6%.In addition,silymarin supplementation at all doses elevated?P<0.05?3?-HSD by 1.8-to 2.5-fold and decreased?P<0.05?Esr1,ABCc1,and ABCc5 in ovaries and the uterus by 10.7-63.2%.In summary,our studies come to the following conclusions:1.Gestational ZEN exposure not only decreases newborn viability,induce placental damages in maternal rats,but also can be transmitted to the fetus through placenta and induce F1 productive toxicity.ZEN disturbs the hormones?FSH,LH,E₂ and T?secretion and leading in pathological changes in ovary,uterus and testis.These reproductive dysfunctions are closely related to altered GnRHr,Esr1 and ABCc5 expressions in the fetal brain and down-regulation of 3?-HSD,StAR and Esr1 expressions in developing reproductive organs.2.Dietary supplementation of silymarin shows a good protection against ZEN induced hepatotoxicity and reproductive toxicity in rats.Silymarin could improve liver antioxidant enzymes and promote hepatic protein synthesis to alleviate ZEN-induced hepatic damages and oxidative injury in rats.Meanwhile,Sily regulates the HSDs to decrease the biotransformation of?-ZOL.Moreover,silymarin regulates hormone synthesis–related proteins to protect reproductive system.