| Aflatoxin B1(AFB1)is the most toxic mycotoxin,which can seriously damage the intestines of animals and cause serious effects on animal health and the breeding industry.Defensins are natural antimicrobial peptides produced by animals themselves and have the functions of repairing the damage and strengthening immunity.To investigate the effect of porcine beta-defensin-2(p BD-2)in intestinal injury induced by AFB1 on IPEC-J2 cells and KM mice,the available concentrations of AFB1 and p BD-2 were screened by CCK-8,and the influence of different concentrations of AFB1 on IPEC-J2 cells was confirmed by q PCR,and the model of AFB1 cell injury was established.Then we treated the cell injury model with different concentrations of p BD-2,and used q PCR,Western Blot,ROS fluorescence staining and flow cytometry to investigate the repair effect of p BD-2 on cell injury induced by AFB1.In animal experiment,AFB1 was used to treat SPF grade KM mice,and the injury model induced by AFB1 was established.The injury model of mice was treated with p BD-2,and the repair effect of p BD-2 on intestinal injury induced by AFB1 was studied by paraffin section,HE and immunofluorescence staining.The results showed that cell viability was significantly reduced by 60 mg/L and higher dosages of AFB1(P < 0.05),and p BD-2 exhibited cytotoxicity at 10 mg/L and significantly reduced cell viability(P < 0.05).Therefore,cells treated with AFB1 below 60 mg/L gradient concentration were tested for changes in cytokines and ROS.The influence of AFB1 on cells was most pronounced at 30 mg/L(P < 0.05).Subsequent AFB1 induced cell injury models were established at this concentration.Using different concentrations of p BD-2 to treat the injury model,the results showed that 2 and 4 mg/L p BD-2 could significantly reduce the induction of apoptosis induced by AFB1;4 and 6 mg/L p BD-2 could significantly alleviate the inflammatory reaction induced by AFB1,repair the damage of AFB1 to the tight junction,restore the expression of MUC2,and protect the intestinal mucosal barrier(P < 0.05);6 mg/L p BD-2 could effectively eliminate the accumulation of ROS induced by AFB1 and alleviate oxidative damage.In animal experiments,AFB1 significantly reduced the average daily gain of mice(P < 0.05),and recovered significantly after p BD-2 treatment(P < 0.05).Jejunal HE staining showed that AFB1 resulted in significant intestinal mucosal damage and the acidophily was enhanced,especially with severe intestinal wall thinning,intestinal villi atrophy and shedding.The ratio of villus height to crypt depth,the number of villus per unit distance and the thickness of intestinal wall were significantly decreased by AFB1(P < 0.05).Injury model of mice treated with p BD-2 showed significant recovery of intestinal structure(P < 0.05).The results of immunofluorescence showed that the inflammatory reaction and apoptosis of the jejunal mucosal epithelium were induced by AFB1,and the environmental homeostasis of the tight junction and the intestinal mucosal barrier were damaged.The above injury was significantly recovered after continued treatment with p BD-2(P < 0.05).In conclusion,AFB1 can induce inflammatory reaction and oxidative damage,promote apoptosis,break down tight junction,cause damage to intestinal mucosal epithelium,affect digestion and absorption,reduce immunity and daily weight gain of mice;p BD-2 can alleviate the influence of AFB1 on intestinal epithelial cells,reduce ROS accumulation,and repair intestinal mucosal structure,improve intestinal environment and intestinal mucosal barrier. |
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