Fish Skin Gelatin Hydrolysate Inhibits Deoxynivalenol-Induced Intestinal Oxidative Damage And Its Mechanism

Vomiting toxin,also known as deoxynivalenol(deoxynivalenol,DON),is one of the most polluted mycotoxins in China.Deoxynivalenol poisoning can cause symptoms such as decreased appetite,weight loss,and intestinal bleeding.Studies have shown that the toxic mechanism of deoxynivalenol is to induce mitochondrial dysfunction,leading to decreased expression of intracellular antioxidant enzymes and induction of elevated reactive oxygen species,thereby causing cells to be in an oxidative stress state,leading to apoptosis.Therefore,effective relief of oxidative stress is of great significance in attenuating the toxic effects of deoxynivalenol.Fish Skin Gelatin Hydrolyzate 3(FSGH3)prepared in the laboratory showed high antioxidant activity under hydrogen peroxide-induced oxidative damage of porcine jejunal epithelium(IPEC-J2),but whether it can alleviate the oxidative damage induced by deoxynivalenol is not clear.Therefore,deoxynivalenol was used to challenge IPEC-J2 cells and C57BL/6 mice to study the protective effects of fish skin gelatin hydrolysate FSGH3 on intestinal epithelial oxidative damage and reveal its possible mechanism for alleviating DON-induced intestinal oxidative damage.This study consists of two parts.The main research contents and results are as follows:The first part : Effect of FSGH3 on oxidative damage induced by DON in IPEC-J2 cellsIn this experiment,IPEC-J2 cell was challenged using 1?g/ml DON.The effects of FSGH3 pretreatment on IPEC-J2 cell barrier damage,oxidative stress and Keap1-Nrf2 signaling pathway were studied.The main results are as follows:1.FSGH3 attenuate DON-induced IPEC-J2 cell barrier damage: Pretreatment with FSGH3 not only significantly increased the cell monolayer transmembrane resistance(TEER)value(P<0.05),but also significantly increased the protein expression of tight junction proteins Occludin and ZO-1 in intestinal epithelial cells(P<0.05).2.FSGH3 attenuate DON-induced oxidative stress in IPEC-J2 cells: FSGH3 pretreatment significantly reduced intracellular ROS levels and MDA content(P<0.05);significantly increased the mRNA expression levels of antioxidant enzymes CAT,SOD-1,GCLM and GCLC(P<0.05);significantly increased the protein expression levels of SOD-1 and GCLM;significantly increased the activities of antioxidant enzymes SOD-1 and GSH(P<0.05).3.The effect of FSGH3 on Keap1-Nrf2 signaling pathway: FSGH3 pretreatment significantly increased the mRNA and protein expression of nuclear transcription factor Nrf2(P<0.05);significantly increased the ratio of Nrf2/Keap1(P<0.01)and the protein expression of Nrf2 in the nucleus(P<0.05),for cytoplasm there was no improvement in the expression of Nrf2 protein(P>0.05).The second part : Effect of FSGH3 on DON-induced intestinal oxidative damage in miceIn this study,48 male C57BL/6 mice of 5 weeks old were randomly divided into 4 groups,12 in each group,for a total of 14 days.The specific groups were as follows:(I)The blank control group was intragastrically administered with PBS every day,and the sterilized water was intragastrically administered for 6 consecutive days in the late stage of the test;(II)The negative control group was intragastrically administered with PBS every day,and the DON model was administered for 6 consecutive days in the late stage of the test;(III)Treatment group 1 was treated with 100 mg/kg.BW FSGH3 per day,and DON was intragastrically administered for 6 consecutive days.(IV)Treatment group 2 was intragastrically administered with 200 mg/kg.BW FSGH3 every day,and DON was administered for 6 consecutive days.And then mice were slaughtered for sampling.The main results are as follows:1.Effect of FSGH3 on mouse phenotypic results: Compared with the negative control group of DON,the average daily weight gain of mice with 200mg/kg.BW FSGH3 was significantly increased(P<0.05),and there was no significant difference in average daily feed intake(P>0.05).The relative weight of the liver was significantly lower(P<0.05),and there was no significant difference in the relative weight of the kidney(P>0.05).The relative weight of the liver could be reduced to the level consistent with the blank control group.2.Effect of FSGH3 on plasma and jejunum antioxidant index in mice: Compared with the negative control group of DON,the MDA content in plasma and jejunum of mice treated with 200mg/kg.BW FSGH3 was significantly decreased(P<0.05),the plasma SOD activity was significantly increased(P<0.05),the jejunal GSH-Px activity was significantly increased(P<0.05)and it can be alleviated to the level consistent with the blank control group.3.Effect of FSGH3 on morphological changes of jejunum in mice: Compared with the negative control group of DON,the villus/crypt ratio in 200mg/kg.BW FSGH3 was significantly increased(P<0.05),and the length of jejunum and crypt depth has a tendency to ease.In summary,the main conclusion of this study is:1.Fish skin gelatin hydrolysate FSGH3 can protect intestinal epithelial cell integrity and alleviate DON-induced cell barrier damage.The mechanism is to increase the expression of the nuclear transcription factor Nrf2 and promote the entry of Nrf2 into the nucleus,thereby increasing the expression of antioxidant enzymes and effectively reducing the level of reactive oxygen species in the cells.2.Intravenous administration of 200 mg / kg BW FSGH3 for 14 days significantly increased the antioxidant capacity of C57 BL / 6 mice,maintained the structural integrity of jejunum villi,and relieved DON-induced growth performance decline in mice.