Study On The Correlation Between Gut Microbiota And IL-22 And LPS In NAFLD Rats

Objective:To explore the changes of gut microbiota in NAFLD model rats induced by high-fat diet,and to further explore the correlation between intestinal flora and IL-22,LPS.Method:1.Twenty male SD rats were allowed to acclimate for 1 week,and then randomly divided into two groups : the control group and the treatment group.The control group was given normal diet,the treatmentl group was given high-fat diet.The changes of body weight of rats were observed.Four weeks later,the rats were killed,and the changes of liver pathology were observed.2.The levels of serum interleukin-22(IL-22)and lipopolysaccharide(LPS)were detected by ELISA,and the serum ALT,AST,GGT,TC,TG were detected by automatic biochemical analyzer.3.The gene sequencing and bioinformatics analysis of the 16 SrRNA V3-V4 region of intestinal mucosal bacteria were carried out by high-throughput sequencing method: a piece of rat intestinal mucosa was taken,and the total DNA of rat intestinal mucosa was extracted.The target region was amplified and purified by PCR method and the library was constructed.The Illumina high-throughput sequencing platform was used for sequencing,and the results of sequencing were analyzed by bioinformatics,such as diversity analysis,the difference analysis of gut microbiota and functional analysis etc.4.To analyze the correlation between gut microbiota and metabolic indexes of non-alcoholic fatty liver: to find out the different species between the control group and treatment group,and to analyze with ALT,AST,GGT,TC,TG,IL-22 and LPS by Spearman correlation analysis.5.SPSS22.0 statistical analysis software was used for data analysis.Mean ±standard deviation was used for measurement data in accordance with normal distribution.T-testwas used for inter-group comparison.Spearman correlation analysis was used for correlation analysis.P < 0.05 is considerded statistically significant.Result:1.In this study,4 rats were selected in control group,9 rats were selected in treatment group.Four weeks later,the body weight of rats in the contorl group was significantly lower than that in the t group(P<0.05).2.Pathological changes of liver: obvious hepatocyte steatosis and inflammatory infiltration of hepatic lobules appeared in the liver of rats in the high-fat diet group.3.The changes of serum metabolic indexes: the serum levels of ALT,AST,GGT,TC and TG in the treatment group were significantly higher than those in the control group(P < 0.05),and the serum levels of LPS and IL-22 in the treatment group were significantly higher than those in the control group(P<0.05).4.Analysis of gut microbiota: In the phylum level,the diversity of gut microbiota in the high-fat diet group increased compared with the control group,and at the phylum level,the abundance of Proteobacteria?Bacteroides and Verrucomicrobia in the high-fat diet group was higher than that in the normal diet group.the abundance of Firmcutes was lower than that in the normal diet group(P<0.05).5.Correlation analysis: In the phylum level,Proteobacteria was positively correlated with serum ALT,AST and GGT(P<0.05),and was positively correlated with LPS(P<0.05),Proteobacteria was negatively correlated with serum IL-22(P <0.05).Bacteroidetes were positively correlated with serum ALT,TC and TG(P<0.05),and positively correlated with serum LPS(P<0.05).Verrucomicrobia was positively correlated with serum AST,GGT,TC and TG(P< 0.05),negatively correlated with serum IL-22(P<0.05),and positively correlated with serum LPS(P<0.05).Firmicutes were negatively correlated with serum LPS(P< 0.05).Conclusion:1.Compared with the normal diet group,the gut microbiota of high-diet rats changed,suggesting that the gut flora may be involved in the occurrence of NAFLD.2.In the phylum level,Proteobacteria in intestinal flora was positively correlated with LPS,and negatively correlated with serumIL-22.Bacteroidetes were positivelycorrelated with serum LPS.Verrucomicrobia was negatively correlated with serum IL-22,and positively correlated with serum LPS.Firmicutes were negatively correlated with LPS.3.IL-22 may be a protective factor in the occurrence of NAFLD,LPS may be a pathogenic factor in the occurrence of NAFLD.