| Deoxynivalenol(DON)is one of the most polluted mycotoxins in the world and a common inducer of intestinal damage in livestock.The DON destroys the intestinal mucosal structure,weakens the intestinal mucosal barrier,digestion and absorption functions,leading to the growth slow of livestock and poultry,which brings enormous economic losses to the livestock industry.In this study,a model of intestinal injury induced by DON was used to investigate whether it can reduce the damage of DON to intestinal tract by hydrolyzed wheat gluten(HWG),and further analyze its mechanism.The potential value of HWG provides theoretical guidance and technical support for its promotion and application in livestock production.A total of 72 C57BL/6 male healthy mice of similar 4-week-old age were randomly divided into 6 treatments,12 replicates per treatment,and 1 mouse per replicate.Three days before the attack,the mice were given HWG(dissolved in physiological saline)and the same volume of normal saline.The control group(CON)and the deoxynivalenol group(DON)were administered with normal saline,low-dose HWG(LH),low-dose HWG+DON(LH+D),and high-dose HWG(HH),high-dose HWG+DON(HH+D)respectively,1000,2000 mg/kg body weight(BW)HWG;Attack period 7 d.The mice in the CON group were given normal saline,the LH group and the HH group were intragastrically administered with 1000 and 2000 mg/kg BW HWG,the DON group was intragastrically administered with 2 mg/kg BW DON,the LH+D group and the HH+D group were administered with 2 mg/kg BW DON and 1000 mg/kg BW HWG,2 mg/kg BW DON and 2000 mg/kg BW HWG,respectively.After the attack: the stomach was the same as before the attack,and the mice were sacrificed on the third day after the attack.The average daily feed intake(ADFI),average daily water intake(ADWI)and average daily gain(ADG)of the whole period of mice were counted,and the duodenum,jejunum and ileum weight were weighed.The jejunal crypts were sampled and cultured ex vivo,and the efficiency and area of enteroid were statistically analyzed.Observation of jejunal morphology by HE staining and scanning electron microscopy.Diamine oxidase(DAO)and lipopolysaccharide(LPS)in jejunum and serum were detected by ELISA.Western blotting and automated capillary western blotting were used to evaluate Wnt/β-catenin signaling pathway,proliferation and differentiation markers and tight junction-associated protein expression in jejunum,crypt and enteroid.The results were as follows:(1)Compared with the DON group,the ADG and ADWI of the mice were significantly increased after HWG(P<0.05).There was no significant difference in ADFI(P>0.05).(2)Compared with the CON group,the intestinal weight of the jejunum unit of the DON group was significantly decreased(P<0.05).After intragastric administration of HWG,compared with the DON group,the intestinal weight of jejunum was significantly increased(P<0.05),but had no significant effect on duodenum and ileum weight(P>0.05).(3)After HWG was administered,the jejunum villi height,crypt depth and the ratio of villi height to crypt depth were significantly greater than the DON group(P<0.05),and the effect of the LH group was better than that of the HH group.Therefore,the jejunum crypts were separated by the CON group,the DON group,the LH group and the LH+D group,and the molecular mechanism of stem cell level was studied.(4)Compared with the DON group,DAO activity and LPS level in jejunum of the LH+D group were significantly increased(P<0.05),and DAO activity and LPS level in serum were opposite to jejunal tissue.Serum DAO activity was significantly greater in the DON group than in the CON group and the LH+D group(P<0.05),and there was an increasing trend compared with the LH group(P=0.078).The serum LPS level of the DON group was significantly greater than that of the CON group,the LH group and the LH+D group(P<0.05).(5)Most of the crypts in the DON group are rod-shaped,and most of the crypts in the CON group,the LH group and the LH+D group are Y-shaped.DON reduced the formation efficiency of enteroid(P<0.05)and decreased the area of enteroid(P<0.05).Compared with the DON group,HWG increased the efficiency of enteroid(P<0.05)and increased intestinal area.(P<0.05),HWG can eliminate the inhibitory effect of DON on the expansion of stem cells to form enteroid.(6)Compared with the CON group,DON significantly inhibited Wnt/β-catenin signaling pathway and down-regulated the protein expression of β-catenin and TCF4 in jejunum,crypt and enteroid(P<0.05),while HWG could reverse the inhibition of DON(P<0.05).(7)Compared with the CON group,the protein expression of the stem cell marker Lgr5,the proliferating cell marker Ki67 and PCNA in the jejunum,crypt and intestinal tract of the DON group was significantly decreased(P<0.05),and the expression of the cell differentiation marker KRT20 was significantly decreased(P<0.05).MUC2+(goblet cell marker)and LYZ+(paneth cell marker)cells in the jejunum were significantly decreased(P<0.05);HWG was able to withstand DON-induced stem cell activity,proliferation and differentiation decrease.Conclusion: HWG up-regulates Wnt/β-catenin signaling pathway to alleviate the inhibition of DON on intestinal stem cell activity,promote stem cell expansion,improve intestinal epithelial cell proliferation,differentiation,and intestinal barrier function,and the effect is better in 1000 mg/kg BW HWG than in 2000 mg/kg BW. |
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