Research On Effects Of Eleutheroside B On Barrier Function In Intestinal Porcine Epithelial Cells

Acanthopanax has a good effect in the treatment of gastrointestinal diseases,and has the function of effectively promoting intestinal development of lactating sows and can promote the growth of piglets.In this study,the effects of Eleutheroside B on the barrier function of small intestinal epithelial cells were investigated in vitro using the piglet small intestinal epithelial cells as the model.Pre-test: Determination of the optimal concentration of Eleutheroside B.IPEC-J2 cells was cultured with the concentration of 0,0.05,0.1,0.2 mg/mL,and 0mg/mL was used as the control group.The proliferation of IPEC-J2 cells treated with different concentrations of Eleutheroside B was observed by xCELLigence RTCA DP system.A curve was used to determine the best concentration of Eleutheroside B.Based on the results,0.1mg/mL Eleutheroside B was selected as the best concentration.Experiment 1: Effect of Eleutheroside B on bacterial membrane permeability of small intestine of piglets.The porcine intestinal epithelial cells with the same growth status were used to measure the intercellular resistance(TEER)and the alkaline phosphatase(ALP)activity of the after measuring culture supernatant with 0 and 0.1mg/mL Eleutheroside B to search the effect of Eleutheroside B on cell membrane permeability.The results showed that the addition of Eleutheroside B to the cell culture medium significantly increased the TEER value and decreased the activity of ALP in the cell culture medium.This indicated that Eleutheroside B significantly reduced the permeability of IPEC-J2 cell membrane.Experiment 2: Effect of Eleutheroside B on the expression of tight junction proteins and its gene expression in small intestine epithelial cells of piglets.In this part,the effects of 0.1 mg/mL Eleutheroside B on the relative expression of Claudin-3,Occludin and ZO-1 were determined by real-time fluorescence quantitative PCR and protein Western blot analysis respectively.The results of real time quantitative PCR showed that the expression of Claudin-3,Occludin and ZO-1 mRNA was significantly increased in Eleutheroside B addition group.The results of Western blot were also consistent with the results.Experiment 3: Effect of Eleutheroside B on anti-inflammatory function of small intestinal epithelial cells in piglets.The relative expression of IL-6,IL-10,TNF-?,TGF-? and INF-? were measured by real-time fluorescence quantitative PCR.The results showed that the expression of IL-10 and TGF-? mRNA was increased and the mRNA expression of proinflammatory cytokines IL-6,TNF-? and INF-? were decreased.0.1mg/mL Eleutheroside B in vitro condition suggested that the Eleutheroside B can play a certain role in promoting cell proliferation,decreased the membrane permeability,tight junction expression and anti-inflammatory function in piglets intestinal epithelial cells.Thus,0.1mg/mL Eleutheroside B can play a protective role in the barrier function of small intestinal epithelial cells in piglets.