Research On N- Acetyl-D- Galactosamine Weakening The Injury Of Intestinal Epithelial Cells Barrier Of Pig By Soybean Agglutinin

Soybean agglutinin is one of the main anti nutritional factors which has influence on the quality of soybean. Searching antagonist of Soybean agglutinin has a great influence on reducing the function of the anti-nutrition of soybean and increasing utilization. In this paper, I do the research on whether antagonist mediated soybean agglutinin could recede the damage function of mechanical barrier on epithelial cells of the intestine in piglet from Soybean agglutinin.Experiment 1: The effect of N--D-- gal attenuated soybean lectin on the permeability of intestinal epithelial cells in piglets. In this experiment,pre-add N- acetyl-D- galactosamine in 0,125×10-4,0.25×10-4,0.5×10-4,1.0×10-4,2.0×10-4mmol/l, add soybean agglutinin in 0.5mg/ml which should be reacted in 24 hours after reaction in 1,2,4,8 hours. Get the value of TEER and the activity of AP in cell medium. Which can reflect the change of the permeability and the integrality of the cells. The result shows that the N- acetyl-D- galactosamine decreases the decline of the value of TEER and the increase of activity of AP in cell medium.Experiment 2:The effects of N- acetyl-D- galactosamine attenuated soybean agglutinin on the relative expression quantity of the TJ protein and genes from Intestinal Epithelial Cells of Piglets. I test the change of relative expression of the TJ protein Occludin and Claudin-3 by Western blotting in the part of the relative expression quantity of TJ protein. I test the change of relative expression quantity of the TJ protein Occludin and Claudin-3 mRNA by the SYBR Green I fluorescent dye inlay in the Real Time Quantitative PCR in the part of the relative expression quantity of TJ genes. Comparing the change of the relative expression quantity of the TJ protein and genes through the three groups which are the blank group(without adding any stimulus), control group(only with soybean agglutinin) and the treatment group(add the agglutinin first and Soybean agglutinin next). The test results show that the change of relative expression quantity of the TJ protein Occludin and Claudin-3 has reduced by 17.8% and 43.5%(P<0.05)in the control group. The change of relative expression quantity of the TJ protein Occludin and Claudin-3 has reduced by 4.3% and 7.2%(P<0.05)in the treatment group. The expression quantity of the Occludin mRNA and the Claudin-3 mRNA has reduced by 17.3%(P<0.01) and 42%(P<0.01) in the control group. The expression quantity of the Occludin mRNA and the Claudin-3 mRNA has reduced by 1.6%(P>0.05) and 2.7%(P<0.01) in the treatment group.Conclusion: The N- acetyl-D- galactosamine could inhibit the effect of soybean agglutini n on permeability of epithelial cells in piglet intestine in the appropriate concentration in vitro c ultivation condition.The soybean agglutinin could inhibit the combination of the protein which connect tightly in the epithelial cell of intestine,the inhibition of the combination of the protein which connect tightly in the cell has been declined by adding the N- acetyl-D- galactosamine. The suitable concentration of N- acetyl-D- galactosamine can weaken the damage of the soybean lectin to the barrier function of the small intestine epithelial cells of piglets.