The Protective Effect Of Lactobacillus On Intestinal Barrier Function In Septic Rats

Background: In recent years,intestinal barrier dysfunction has been considered as the initiating factor for the occurrence of multiple organ failure in sepsis.The tight junction of intestinal epithelial cells is a component of the mechanical barrier of the intestinal mucosa.Once the tight junction is damaged,the permeability between the intestinal epithelial cells increases,and bacteria and endotoxins etc.can enter the systemic circulation through the intestinal barrier,which is related to the occurrence and development of various diseases.The occurrence and development of related,tight connections can be used to evaluate the function of the intestinal mucosal barrier.Objective: To investigate the effect of Lactobacillus plantarum on the expression of Claudin-1 and Occludin in the ileum of rats with sepsis using a lipopolysaccharide(LPS)-induced sepsis rat model,and then to investigate to the protective function of rat intestinal barrier function in sepsic rats.Methods: Ninety healthy male Sprague-Dawley rats were randomly divided into control group(C group),Lactobacillus plantarum pretreatment group(LP group),sepsis group(LPS group),and sepsis plus Lactobacillus plantarum intervention group(LPS+ LP group),Lactobacillus plantarum pretreatment plus sepsis group(LP + LPS group).Rat models of sepsis were induced by tail vein injection of LPS.LPS+LP groups were given Lactobacillus plantarum once a day after tail vein injection of LPS.In the LP+LPS group,LPS was injected once into the tail vein 3 days after continuous gavage.Six rats were taken from the experimental group at 6h,12 h and 24 h after treatment.Ultrastructure was closely observed by transmission electron microscopy.The expression of Claudin-1 and Occludin protein was detected by Western Blot.Immunohistochemistry and immunofluorescence were used to detect the expression of Claudin-1 and Occludin protein in ileum.Results: 1.Intervention of experimental animals: Group C rats had no effect after oral gavage,good living conditions,shiny hair,no abnormal activity response,sensitive to external sound and stimuli,and no death;LP group(Lactobacillus pretreatment group)Rats treated with gavage had no significant differences from group C and had normal activities.All rats in the LPS group(sepsis group)had significantly reduced activity after injecting LPS via the tail vein,and they either consumed little or did not eat water.Lustrous hair,diarrhea,fatigue,shortness of breath,unresponsiveness to external stimuli,and even death;LPS + LP group,LP + LPS group also appeared similar symptoms.2.Transmission electron microscope observation of the tight junction ultrastructure: In the C group,the LP group tight junction structure is normal,and the gap junction is not broken,the microvilli are arranged neatly,the cell gap does not increase;no abnormal tight junction protein structure is seen.In the LPS group,many abnormal and tightly connected structures were observed,such as discontinuous,swelling,etc.,and discontinuity in gap junctions.The lack of microvilli showed that sepsis had a greater impact on tight junctions.The LP+LPS group and the LP+LPS group also showed an abnormally tight junction structure,but they were less than the LPS group.3.Claudin-1 protein expression in small intestine: Claudin-1 had no difference in the LP group at 6h,12 h and 24 h compared with the C group.C group was higher than LPS group at 6h,12 h,24h(P<0.01).LP+LPS group was higher than LPS group at 6h,12 h,24h(P<0.05).The LPS+LP group was higher than the LPS group at 6h and 24h(P<0.05).There was no difference between the LPS+LP group and the LPS group at 12 h.4.Occludin protein expression in the small intestine: Occludin in the 6h,12 h,24h LP group compared with the control group(C)no difference,statistically meaningless.Compared with group C,there was no difference in statistical analysis at 6h between LPS group and CPS group at 12 h and 24h(P<0.01).There was no difference in LP+LPS group compared with LPS group at 6h,higher than LPS group at 12h(P<0.05)and higher than LPS group at 24h(P<0.01).Compared with LPS group,LP+LPS group had no difference between 6h and 12 h,and it was higher than LPS group at 24h(P<0.05).5.The immunohistochemical expression of Claudin-1 and Occludin protein in small intestine tissue: In group C and group LP,strong brown positive particles were distributed along the apical cell membrane epithelial membrane at 6h,12 h,and 24 h,all showing continuous distribution.In the LP+LPS group,the distribution along the epithelium of the gut mucosa was continuous and there was no obvious interruption.In the LPS+LP group and the LPS group,the brown positive particles were significantly reduced,the color was weakened,and there were multiple interruptions.6.Immunofluorescent expression of Claudin-1 and Occludin protein in small intestine: Claudin-1 and Occludin protein showed linear and dense particles in ileum mucosal epithelium in group C and group LP at 6h,12 h and 24 h.Most of the tight junction proteins in the LP+LPS group remained intact,but some fragmentation was still observed.There was no significant difference in fluorescence intensity between the LPS+LP group and the LPS group.There were many breakages in the two groups and the fluorescence was weak.Conclusion: 1.Rat model of sepsis can be induced successfully by tail vein injection of lipopolysaccharide.2.Lactobacillus plantarum can upregulate the expression of Claudin-1 and Occludin proteins and protect the intestinal barrier function in septic rats.