| Experiment Ⅰ Establishment of reversible mechanical small bowel obstruction modelObjective:To establish a novel rat model of reversible complete small bowel obstruction.Methods:Forty-eight healthy Wistar rats were randomly divided into sham operation group(Sham),obstruction 1d group(S1),obstruction 2d group(S2),obstruction 3d group(S3),relief group after 2d obstruction(J2),relief group after 3d obstruction(J3)The rat model was developed by using a flexible polyvinyl chloride ring surgically placed surrounding the terminal ileum and removing the ring through reoperation.General condition of rats,macroscopical and microscopical characteristics were observed to assess the injury of the intestinal mucosa.Results:Compared with Sham group,S1 group showed symptoms of incomplete intestinal obstruction and visible intestinal mucosal damage;S2 group showed symptoms of complete intestinal obstruction and more serious damage of intestinal mucosal;The symptoms and the intestinal mucosa damage of S3 group were further aggravated.After relief of obstruction,the symptoms of intestinal obstruction in the J2 group gradually disappeared,the macroscopical and microscopical characteristics returned to normal level at the 7th day;the symptoms of the J3 group disappeared slowly,there were significant damage of the intestinal mucosal at the 7th day.Conclusion:This study suggests that the flexible polyvinyl chloride ring can be used to successfully establish a rat model of reversible complete small bowel obstruction,which represents an ideal model for small bowel obstruction research and postoperative recovery from the disease.Experiment Ⅱ Alterations of gut microbiota in experimental small bowel obstructionObjective:To investigate structural and quantitative alterations of gut microbiota in an experimental model of small bowel obstruction,and to analyze the correlation between the target flora and indicators of intestinal mucosal barrierMethods:Thirty-two healthy rats were randomly divided into sham operation group(Sham),obstruction 1d group(S1),obstruction 2d group(S2),and obstruction 3d group(S3).The alterations of gut microbiota were studied after intestinal obstruction.Intraluminal fecal samples proximal to the obstruction were collected and analyzed by 16s rDNA high-throughput sequencing technology and quantitative PCR(qPCR)for target bacterial groups.Furthermore,intestinal claudin-1 mRNA expression was examined by real-time polymerase chain reaction analysis,and serum sIgA levels were determined by enzyme-linked immunosorbent assay.Results:1.The composition of intestinal flora showed that the number of OTU after obstruction was significantly higher than that of sham operation group(P<0.01).At the phylum level,the 16S rDNA sequences showed a marked decrease in the relative abundance of Firmicutes and increased abundance of Proteobacteria,Verrucomicrobia and Bacteroidetes within 24 hours(P<0.01).The abundance of Firmicutes increased slightly and Proteobacteria,Verrucomicrobia decreased gradually with Bacteroidetes continuously increased between 24 and 72 hours.At the genus level,the relative abundance of Romboutsia,Lactobacillus and Turicibacter decreased significantly within 24h(P<0.01),Escherichia-Shigella,Akkermansia and Bacteroides increased significantly(P<0.01).2.The qPCR analysis showed the absolute quantity of total bacteria increased significantly within 24 hours but did not change distinctly from 24 to 72 hours.3.Claudin-1 and sIgA were positively correlated with Romboutsia and negatively correlated with Escherichia coli,Akkermansia,Bacteroides.Conclusion:1.Small bowel obstruction led to significant bacterial overgrowth and profound alterations in gut microbiota composition and diversity;2.The predominant flora differs at different stages of the disease,with Escherichia coli and Akkermansia within 24 hours,Bacteroides between 24 and 72 hours;3.Intestinal mucosal barrier damage during obstruction was related to Escherichia coli,Akkermansia,Bacteroides,Romboutsia could play a protective role in the disease.Experiment Ⅲ The effect of Fufang Dachengqi Decoction on intestinal flora during the recovery of intestinal obstructionObjective:Reversible complete small intestinal obstruction model was used to observe the changes of intestinal flora after the removal of obstruction and the effect of Fufang Dachengqi Decoction.Methods:Thirty-two healthy rats were randomly divided into sham operation group(Sham),obstruction 2d group(S2),natural recovery group(R3),and Chinese medicine treatment group(T3).The structural and quantitative alterations of gut microbiota were studied.Furthermore,intestinal claudin-1 mRNA expression and serum sIgA levels were determined and compared.Results:1.After the obstruction was removed,the relative abundance of Firmicutes of R3 and T3 groups increased significantly(P<0.01),and the abundance of Verrucomicrobia decreased significantly(P<0.01).2.Compared with the R3 group,the number of OTUs and bacterial community composition of the T3 group were closer to the Sham group.3.Compared with the R3 group,the number of Romboutsia and Turicibacter of T3 increased significantly(P<0.01),while the number of Bacteroides decreased significantly(P<0.01).4.sIgA in the Chinese medicine treatment group rose faster than that in the natural recovery group,and it was close to the level of the Sham group after 3 days of medication.Conclusion:Fufang Dachengqi Decoction can effectively promote the stability and integration of intestinal microbiota and the rapid recovery of the body after the removal of obstruction.The biological mechanisms include:1.Reduce the diversity of intestinal flora and inhibit the growth and reproduction of bacteria.2.Promote the growth of probiotics such as Romboutsia,Turicibacter,and reduce the number of pathogenic bacteria such as Akkermansia,Escherichia-Shigella,Bacteroides.3.Promote the synthesis and secretion of sIgA in the body and thus improve the immune function of intestinal mucosal barrier. |
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