| Disruption of intestinal epithelial barrier structure,the first line of defense against invasion of pathogenic microorganisms and foodborne hazard factors,is closely related to the occurrence and development of many metabolic diseases.Lactobacillus plantarum,one of the most essential probiotics in human intestinal tract,has been shown to successfully regulate the intestinal barrier with obvious strain specificity.Yet,studies on the interaction between L.plantarum and intestinal barrier mainly focus on the strain itself,the material foundations of L.plantarum on protection of intestinal barrier are not well-understood.Besides,most of research mainly focus on commercial strains,especially the analyzation of single strain composition.Therefore,the purpose of this study was to conduct comprehensive in vitro and in vivo evaluation based on the species of plant lactobacillus.Strains with significant difference on the regulation of intestinal epithelial barrier were selected for the genome comparison,separation of related materials,and verification of functionalities.The material foundations of strain functional specificity were analyzed for exploration of common laws of excellent strains on regulating intestinal epithelial barrier.Firstly,a model of Caco-2 cell monolayer destructed by tumor necrosis factor-? was established.The effects of 55 L.plantarum strains on the intestinal epithelial barrier were evaluated.The results showed that these L.plantarum strains have different regulatory effects on the decrease of transepithelial electrical resistance(TEER).The highest recovery rate was94.63% and the lowest was-81.13%.Six strains with significant differences in their ability to regulate TEER were selected to investigate their effects on the other two main indexes of intestinal barrier.These results showed that the three strains(QS6-1,QHLJZD20L2 and VJLHD16L1)which significantly upregulated TEER value could all increase the content of tight junction protein and the epithelial cell viability.The three strains with down-regulated TEER value(VCQBB3-125-L8,QSCPS1L3 and JS-NJ-PK-4-G-1)had no regulatory effect on the above indexes.Subsequently,the regulatory effects of these strains on DSS-induced injury of the intestinal barrier in mice were evaluated.It was found that 3 strains,which were effective in cell experiments,could significantly reduce weight loss and colon length shortening,decrease intestinal permeability and inhibit the down-regulation of the expression levels of intestinal tight junction proteins.However,3 strains which has a negative regulation of the cell monolayer barrier in cell experiments,had no significant protective effect on DSS-induced gut injury in mice.The results of both in vitro and in vivo models showed that there were differences in the regulation effect of L.plantarum on intestinal barrier.Based on the above experimental results,to explore the inducement of functional specificity and reveal the commonalities among the superior strains,a comparative genomic analysis was conducted on 6 L.plantarum strains with different intestinal barrier regulation functions.The results showed that no general rule was observed in the classification of COG,the number of corresponding genes and the distribution of carbohydrate active enzyme gene family in these strains.Besides,in the phylogenetic tree,these strains with different regulatory roles were divided into the same branch separately,indicating that these strains have strain specificity in genetic background.In addition,the 3 strains of L.plantarum with positive regulation shared a specific gene encoding the PTS fructose transporter subunit IIC,which has been reported to be related to the synthesis of capsular polysaccharides(CPS).Finally,through the comparison of live and dead bacteria,it was confirmed that the surface material components of bacteria played a major role in keeping intestinal barrier.Based on this result,following experiments were conducted on the relevant substance components including surface protein and CPS for functionality verification.These results indicated that only CPS produced by 3 strains with positive regulatory effects could significantly up-regulate the tight junction protein content and enhance the epithelial cell viability.In addition,the above regulatory effects were not observed in the surface proteins of all strains.Therefore,the capsular polysaccharide was identified as the main material basis.Then the differences in structure and content of CPS were analyzed among strains with different regulatory effects.The results showed that there was no difference in the structure(monosaccharide composition and functional group structure)of the CPS between the strains with the strongest and the strains with the least regulatory ability.However,there were significant differences in the content of CPS.These results showed that the strains with negative regulation effect generally had a low content of CPS.Then,after unifying the same concentration of CPS in all strains,it was found that the CPS in all strains showed a positive regulation effect on tight junction protein and intestinal epithelial cell activity.Therefore,the content of CPS is the key factor affecting the intestinal barrier regulation ability of the strain.This study proved that L.plantarum has strain specificity in regulating the intestinal epithelial barrier,and the main material basis for the functional specificity of the strains is capsular polysaccharide,and it was found that the strains with positive regulation ability have a common feature,that is,they can produce high concentration of capsular polysaccharide.In this study,the interaction between Lactobacillus plantarum and the host intestinal tract was studied from the population level of Lactobacillus plantarum,and the common characteristics of the superior strains in regulating intestinal barrier function were analyzed,which provided a new screening target for the rapid screening of L.plantarum with gut barrier protection ability in the future. |
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