Effect Of RegⅢγ On Proliferation Of IPEC-1 And Its Expression And Regulation

Reg Ⅲγ,a member of regenerating gene family with 149 amino acid,belongs to C-type lectin IV family.It is mainly expressed in mammalian skin and intestinal epithelium,with a variety of biological functions,such as antimicrobial function,antiinflammation,cell proliferation and wound healing.It plays a vital role in the immune process after pathogens invading.In view of the multiple functions of this protein,Reg Ⅲγ has a better prospect in the prevention and control of disease of newborn weaned piglets.We determined the expression of Reg Ⅲγ in different parts of the intestine of healthy newborn piglets by immunohistochemistry,analyzed the proliferation effect of recombinant porcine Reg Ⅲγ on IPEC-1 cell and detected the expression of Reg Ⅲγ in IPEC-1 cell after induced by inactivated bacteria: 1.The localization of Reg Ⅲγ protein in different parts of neonatal pigletsIn order to identify Reg Ⅲγ expression in different section and different developmental stage of the newborn piglets intestine,the samples from duodenum,jejunum,ileum,and colon from 0 to 5 weeks of healthy piglets were collected and fixed in 35% formalin,and then transferred to 75% of the alcohol for long-term preservation.The cell type of Reg Ⅲγ expressed in different intestinal segments and different growth periods were detected by immunohistochemistry.The results showed that the Reg Ⅲγ in different intestinal tract of piglets expressed in all three major cell type: Paneth’s cell,intestinal absorption epithelia cells and goblet cell.2.The effect of porcine recombinant Reg Ⅲγ on IPEC-1 cell proliferationSome reports turned out that Reg protein can stimulate the cell proliferation.In order to determine whether Reg Ⅲγ promote intestinal epithelial cell proliferation,IPEC-1 cell were tested by CCK-8 method after incubated with porcine recombinant Reg Ⅲγ.The results showed that Reg Ⅲγ can promote IPEC-1 cell proliferation significantly.3.Determination of the expression of Reg Ⅲγ in IPEC-1 line induced by inactivated bacteriaThe Q-PCR and Western blotting were used to quantify the Reg Ⅲγ m RNA and protein expression in IPEC-1 cell after induced by inactivated bacteria.The results showed that inactivated S.aureus(≥108)and E.coli(≥105)can stimulate the expression of Reg Ⅲγ m RNA and protein expression in IPEC-1 cell line.