| Deoxynivalenol(DON)is the most widely contaminated mycotoxin in grain seeds and feed.DON can cause intestinal oxidative stress,inhibit intestinal cell proliferation,induce its apoptosis,lead to intestinal epithelial structure collapse and functional dysfunction,thus reduce animal growth performance and result in huge economic losses to the breeding industry.L-carnosine(LC)is a powerful antioxidant,which has the functions of scavenging oxygen free radicals and anti-apoptosis.However,it is unknown whether LC can alleviate DON-induced intestinal stem cell damage.To this study,the following experiments were designed: 40 4-week-old C57BL/6 male healthy mice with similar body weight were randomly divided into 4treatment groups,namely the control group(CON),the positive control group(LC),the attack group(DON)and the remission group(DON + LC).Each treatment group had10 replicates and one mice per repeat.The experiment lasted for 11 days.The mice in the CON group were given 0.3 mL phosphate buffered saline(PBS)for 1-10 days,and the mice in the LC group were given 0.3 mL 300 mg/kg BW LC for 1-10 days,DON group mice were given 0.3 mL PBS for 1-3 days and 9-10 days,0.3 mL 3 mg/kg BW DON by gavage for 4-8 days,DON + LC group mice were given 0.3 mL 300 mg/kg BW LC for 1-3 days and 9-10 days,0.15 mL 3 mg/kg BW DON + 0.15 mL 300 mg/kg BW LC by gavage for 4-8 days.On the 11 th day of the experiment,the eyeballs of the mice were removed,and the mice were sacrificed after the serum samples were collected.The changes of the food intake,water consumption and body weight of the mice during the whole period of the experiment were recorded.Then,the abdominal cavity of the mouse was dissected,and the jejunum of the mouse was separated and weighed,and calculated the weight of the jejunum per unit length.The jejunum sample is divided into 5 parts,one was used to determine the trans-epithelial electrical resistance(TEER)of intestinal epithelial cells through the Using Chamber;one was fixed in 4% paraformaldehyde for HE staining and tissue immunofluorescence staining;one was fixed in 2.5% glutaraldehyde for SEM observation;one was stored in liquid nitrogen for Western blotting to detect nuclear factor E2 p45-related factor 2(Nrf2)and signal transducer and activator of transcription 3(STAT3)signaling pathways,as well as the expression of proliferation,apoptosis and stem cell markers;one was used to isolate crypts and cultured ex vivo,and the efficiency of crypt cell expansion into enteroid and the enteroid budding efficiency in each group was calculated.The result is as follows:(1)Compared with the CON group,DON significantly reduced the average daily weight gain and average daily feed intake of mice(P<0.05).Compared with the DON group,the DON + LC group significantly increased the average daily weight gain of the mice(P<0.05),but it did not significantly increase the average daily feed intake and average daily water intake of the mice(P>0.05).(2)Compared with the CON group,the weight of jejunum per unit length and TEER value of the DON group were significantly reduced(P<0.05).Compared with the DON group,the weight of the jejunum per unit length in the DON +LC group increased(P=0.075),and the TEER value increased significantly(P<0.05).(3)The intestinal morphology of the jejunum villi in the CON,LC,and DON + LC groups was denser,while the jejunum villi in the DON group were disorderly arranged,with different degrees of ulceration at the top of the villi.The height of the villi and the villi-to-crypt ratio were significantly lower than those of the other three groups(P<0.05).(4)Compared with the CON group and the LC group,the diamine oxidase(DAO)activity in the serum of the DON group was significantly increased,while the DON + LC group significantly reduced the DON on serum DAO Increased(P<0.05);while the DAO activity in the jejunum tissue showed a completely opposite trend between the groups(P<0.05).And DON + LC group significantly increased the expression of cytoplasmic tight adhesion protein 1(Zonula occludens-1,ZO-1)and tight junction protein Claudin1 in the jejunum(P<0.05).(5)Compared with the CON group and the LC group,the activity of superoxide dismutase(SOD)in jejunum and serum of DON group was significantly reduced,and the activity of SOD in jejunum and serum of mice exposed to DON was significantly increased in DON + LC group(P <0.05).(6)Compared with the DON group,the CON,LC and DON + LC group can significantly increase the efficiency of stem cell expansion into enteroid(P<0.05)and enteroid budding efficiency(P<0.05).(7)Compared with the DON group,the CON,LC,and DON + LC groups all significantly increased the expression of Villin,MUC2,and LYZ in the jejunum(P<0.05).At the same time,the expression of the proliferation-related protein Ki67 and the proliferating cell nuclear antigen(PCNA)in the jejunum of other treatment groups were also significantly increased compared with the DON group(P<0.05).(8)Compared with the CON group,the DON group significantly inhibited the phosphorylation of Nrf2 and STAT3,and the DON + LC group significantly promoted the activation of Nrf2 and STAT3 under DON exposure(P<0.05),thereby improving the expression of the anti-apoptotic protein Bcl2 and reducing the expression of the pro-apoptotic protein Bax and the apoptosis executor Caspase3(P<0.05).Conclusion: DON can induce intestinal oxidative stress in mice,inhibit the proliferation and differentiation of intestinal stem cells,induce its apoptosis,and destroy intestinal barrier function,while adding LC can reactivate Nrf2 and STAT3,and improve the intestinal antioxidant and anti-withering of mice The ability to die,promote the expansion of intestinal stem cells,thereby alleviating DON damage to intestinal epithelial structure and function. |
