Screening And Functional Evaluation Of Lactobacillus For Alleviating Chronic Alcoholic Liver Injury

Alcoholic liver injury is a common clinical liver disease,with high incidence,great harm and complicated pathogenesis.Intestinal barrier dysfunction and its induced endotoxemia are considered to be the key factors in the occurrence and development of alcoholic liver injury.Probiotics have been proven to have various probiotic functions such as improving intestinal barrier function,regulating intestinal flora,and improving body immunity.At present,studies have shown that probiotics can prevent intestinal barrier dysfunction and liver damage caused by alcohol.However,these studies mainly concentrate on foreign commercial strains.And there is no screening model for probiotics against alcohol-induced intestinal barrier dysfunction.Therefore,Lactobacillus were used as the research object to explore the intervention measures for alcoholic liver injury from the perspective of improving intestinal barrier function.An in vitro cell barrier model was established for preliminary screening to obtain strains with different protective effects on the cell epithelial barrier,and then the results of cell model were verified by animal model.Finally,the effects of the selected strains and their compound preparations on chronic alcoholic liver injury were studied.Firstly,a model of Caco-2 cell monolayer exposed to alcohol was established.Lactobacillus rhamnosus GG was used as a positive control strain,and the effects of 86 strains of Lactobacillus on the intestinal epithelial cell barrier were evaluated.The results showed that alcohol caused the impaired integrity of Caco-2 cell epithelial barrier.The 86 experimental strains had different effects on the recovery of the alcohol-exposed Caco-2 cell monolayer barrier,with recovery ranging from 58.40% to 112.60%.And there were no inter-specific differences between them.In order to verify the results of the cell experiments,strains of Lactobacillus casei with good effect(RS8-5,111.67% recovery)and poor effect(V-CQRC7-161-M2,58.40% recovery),as well as strains of Lactobacillus helveticus with good effect(D-7-S146,112.60% recovery)and poor effect(8M-10,67.91% recovery)were selected out.And then the effects of these strains on chronic alcoholic liver injury were evaluated.A mouse model of chronic alcoholic liver injury was established,and these strains were used as interventions.The results showed that chronic alcohol exposure resulted in damage to the intestinal barrier and disturbance of intestinal flora,as well as liver injury,fat accumulation,liver oxidative stress and inflammation in mice.L.casei RS8-5 and L.helveticus D-7-S146,which were effective in cell experiments,could effectively inhibit the down-regulation of the expression levels of intestinal tight junction proteins and endotoxemia in mice caused by chronic alcohol exposure,improved the balance of intestinal flora,and significantly reduced liver fat accumulation,serum aminotransferase and lipid levels,relievedliver oxidative stress and inflammation in mice exposed to chronic alcohol.However,L.casei V-CQRC7-161-M2 and L.helveticus 8M-10,which did not protect the cell monolayer barrier in cell experiments,had no significant protective effect on alcoholic liver injury in mice.These results are consistent with the results of cell experiments.To further verify the reliability of the cell model,the correlation between the results of cell experiments and animal experiments was analyzed.The Pearson correlation coefficient was-0.97.The results showed that the screening results of cell model had a strong correlation with the results of animal experiments,suggesting that Caco-2 cell monolayer model can be used as an in vitro preliminary screening model for probiotics against alcoholic liver injury.Finally,five combinations of probiotic compound preparations composed of L.casei RS8-5,L.helveticus D-7-S146,Pueraria,Semen Hoveniae,hawthorn,Lycium barbarum,mulberry,fructooligosaccharide and galactooligosaccharides were used to evaluate their effects on chronic alcohol liver injury in mice.The results showed that the five probiotic compound preparations could effectively inhibit the intestinal barrier damage and endotoxemia in mice caused by chronic alcohol exposure,and alleviated liver injury,liver oxidative stress and liver inflammation in mice.Among them,the combination of formula 1 +mixed bacteria composed of L.casei RS8-5,L.helveticus D-7-S146,Pueraria,Semen Hoveniae,hawthorn,fructooligosaccharide,had the most significant effect.The results of this study provided a possible in vitro model for screening probiotics with the potential to alleviate alcoholic liver injury,and further screening to obtain functional strains that can effectively alleviate alcoholic liver injury,providing a potential intervention for dietary prevention and treatment of alcoholic liver injury.