Bifidobacterium Modulate The TLR2,TLR4,TLR9Expression In Intestinal Tissuse Of A Neonatal Rat Model Of Necrotizing Enterocolitis

Objective: To study the role of bifidobacteria on the clinicalbehaviour of the neonatal rat model of necrotizing enterocolitis(NEC) andthe morphologic appearance of the gut. TO provide the preclinical medicalevidengce applying bifidobacteria to cure NEC in newborn.Method：1．Animal and group: The pups which are24hours after birth are randomdivided into3groups,12pups in every group.There are controlgroup,experimental group and therapy group.Control group:fed withoutbifidobacteria and modeled. Experimental group: built the NEC modeled.Therapy group: fed the bifidobacteria and modeled.2. The way of adding bifidobacteria: Adding viable bifidobacteria intothe formula milk, the concentration of bifidobacteria is1×108cfu/ml,dose is twice one day.3.Setting up the NEC model: The NEC modeled pups were gavage fed withFormular milk (six times daily).Subjected to90sec of hypoxia wtith100%Nitrogen gas and4℃cold exposure for10min twice a day during3consecutive days.4.Tissue harvest:All the pups were sacrificed on the finished modeled.Nearly0.5cm to cecum,about0.5cm of terminal ileum wereexcised,fixed in neutral formalin and paraffin embedded forhistological blades.5. Experimental index: Observe the changes of growth indexs,comparedthe difference of the neonates’clinical behaviour,the morbility of NEC,evaluated the histological scored of each guts tissues.6.Statistics and method: The data were calculated bySPSS17.0.Measurement datas were presented by mean±standaeddeviation,Wilcoxon rank sum test was used for group comparison,Chi squarewas used for rate comparison,it was considered significant differencewith P<0.05.Results:1.The weight increasing of experiment group was lower than control groupand therapy group (﹣0.28±0.05VS2.16±0.28VS0.56±0.07),the differenceof weight increasing of these three groups were significant(P<0.05).Thebody length increasing of the experiment group（0.35±0.14） are slowerthan the other groups, the therapy group（0.71±0.02） are higher thanexperiment group but smaller than control group（1.23±0.06）,there weresignificant difference in these three groups(P<0.05).2.The incidence of abdominal distention and hematochezia of NECclinical behaviour in experiment group was50%, the therapy group’s was20%,the incidence of experiment group higher than therapy group’s.Thecomparence were significant difference.3.The experiment group’s NEC incidence was58.33%,control group was0%, therapy group was33.33%, the incidence of therapy group decrease thanexperiment group(P<0.05).The average intestinal pathology score of thesethree group was experiment group：1.75±0.75，therapy group：0.83±0.7，control group：0.The highest pathology hurt was the experiment group. Conclusions:1. Bifidobacteria therapy could decrease the growth retardation in NEC.2. Bifidobacteria therapy could decrease the NEC clinical behavor likehematochezia and abdominal distention.3. The way of adding bifidobacteria could reduce the incidence of NEC andthe injury degree of intestinal tissuse. Objective: To explore the role of adding bifidobacteria in expressionof TLR2、TLR4、TLR9in intestinal tissuse of a neonatal rat model of NECthough quantitation real time PCR(qRT-PCR).To provide the molecularbiological evidence for the protection role of bifidobacteria tharepy.Method:The three group neonatal rat intestinal tissue from the partone were tested the gene expression of TLR2、TLR4、TLR9by qRT-PCR,thencompared their difference expression. The data were calculated bySPSS17.0.Measurement datas, it was considered significant differencewith P<0.05.Results:As compared with the control group, the gene expression of TLR2in experiment group was0.48times,the therapy’s was2.49times, the geneexpression of therapy group higher than experiment group;The expressionof TLR4in experiment group was3.48times greater than control group,thevalue of therapy group was1.25times as great as control group, theexpression of TLR4in experiment group highter than the therapy group;TLR9expression in therapy group was4.00times greater than control group,theexperiment group was0.47times as greater as control group,the therapygroup have higher TLR9expression than the other groups.The geneexpression compare was significant difference between the three groups.Conclusion: Bifidobacteria therapy could decrease the incidence of NEC may be related that bifidobacteria could increase the gene expression ofTLR2、TLR9and decrease the gene expression of TLR4. Object: To observed the role of bifidobacteria affected the expressionof intestinal tissuse TLR2、TLR4、TLR9precede histological injury,andto investigate the mechanism of bifidobacteria therapy could prevent NEC.Method:Fourty five within24hours pups after birth are random dividedinto3groups:control group(n=15),experiment group(n=15),therapy group(n=15).All the group treat with the same to the part I,then5rats fromevery group were euthanized on24h,48h,and72h.Every intestinal tissusewere tested the expression of TLR2、TLR4、TLR9by qRT-PCR,and compare tothe difference time of gene expression. The data were calculated bySPSS17.0.Measurement datas, it was considered significant differencewith P<0.05.Results:0n24hours,TLR2expression in every group was not significantdifference(P>0.05);on48hours, TLR2mRNA of the experiment group was1.07times greater than control group,the therapy was3.75times as greateras control heigher than experiment group(P<0.05); on72hours,TLR2expression of therapy group (5.97times as greater as control) greaterthan experiment group(0.72times as greater as control group). Theexpression of TLR4：on24hours, there were not significant differencein each groups;48hours,the value of experiment group was3.45timescompare to control group,the therapy group was1.33times as control group,experiment group heighest expression TLR4in three groups(P<0.05);72hours, experiment group was8times compare to control group heigher thanthe other groups (P<0.05).The TLR9expression was not obviously increasedin every group on the24hours (P<0.05);48hours later,the TLR9expression of therapy group began to increased,the value of therapy group was3.44times greater than control group,and heigher than experimentgroup(P<0.05); The expression of TLR9on72hours, therapy group was5.56times as greater as control group and significant difference compared tothe other two groups(P<0.05).Conclusion:The way adding bifidobacteria could prevent the incidenceof NEC.It would presumed that bifidobacteria could increase theexpression of TLR2、9and decrease the expression of TLR4precede thehistological injury of NEC.