| RgeeneratingⅢγ(RegⅢγ),a new type of intestinal antimicrobial protein which belongs to C-type lectin,has mutilple functions including killing bacteria,inhibiting intestinal inflammation,promoting wound healing of intestinal tissue and cell proliferation.It is an indispensable factor in the process of intestinal innate immune defense.At the same time,the expression of RegⅢγ is inducible,and the intestinal mucosa can secrete RegⅢγafter being stimulated by bacteria,inflammation and injury,which has a good effect on the treatment of early weaned piglet diarrhea.Chitooligosaccharide(COS)is the second most abundant natural polymer after plant cellulose.It can be prepared by acetylation of chitin and chitosan under alkaline conditions or enzymatic degradation.Its structure is similar to peptidoglycan(PG).Therefore,COS was selected as the inducer to induce IPEC-J2 cells,pre weaning and post weaning mice to explore the effect of RegⅢγ expression.Methods and results are as follows⑴Effects of COS on viability of IPEC-J2: IPEC-J2 cells were treated with different concentrations of COS(0 μg/m L、25 μg/m L、50 μg/m L 、100 μg/m L、200 μg/m L)for24 h and the viability and morphology of IPEC-J2 cells was evaluated by MTT assay and microscopic observation respectively.The results showed that COS had less toxic effects on IPEC-J2 cells,the cells were granulosa spindle like,and the concentrations chosen in subsequent in vitro assays were within this range.⑵Effects of COS on RegⅢγ expression in IPEC-J2 cell: IPEC-J2 cells were treated with different concentrations of COS for 24 h,m RNA and protein of RegⅢγ was determined by q PCR and Western blot respectively.The results showed that different concentrations of COS could induce the expression of RegⅢγ.⑶Effects of COS on RegⅢγ expression in ICR mice intestinal: The mice were treated with different doses of COS solution(does: 200 mg/kg、600 mg/kg、1000 mg/kg)before and after weaning for 5 days to observe the changes of growth body mass index(BMI)of mice before and after weaning.The results showed that each dose of COS group had a certain degree of influence on the growth BMI of mice before and after weaning.Western blot and q PCR were used to detect the expression of RegⅢγ.The results showed that compared with the control group,COS could induce the expression of RegⅢγ in the intestine of mice(P <0.05),and the expression of RegⅢγ was significantly higher after weaning than before weaning(P < 0.05).⑷Morphological observation: The results of HE staining showed that before weaning,the villi in the control group were uniform in thickness,lamina propria was covered by mucosal epithelium,and there was no hyperemia.The villi height of each COS dose group was increased,and goblet cell(GC)increased significantly.After weaning,the villi in the control group were atrophied,damaged,and hyperemia.The villi height of each COS dose group was increased,and the damage was reduced The number of GC increased.The villus height,crypt depth,as well as villus height/crypt depth(V/C)ratio of small intestine in mice before and after weaning were detected.Compared with the control group,the villus height and V/C ratio were increased,while crypt depth was decreased.There was a significant difference,indicating that COS can repair villus damage caused by weaning stress,promote intestinal mucous secretion and form intestinal mucosal barrier.Conclusion:(1)COS induces RegⅢγ by both in vitro and in vivo assays expression.(2)COS can improve the morphology of intestinal villi before and after weaning,repair the injury of intestinal villi caused by weaning stress,promote the secretion of mucus and form intestinal mucosal barrier. |
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