论文部分内容阅读
目的:观察小鼠肠道菌群的变化,探讨粪菌结合益生菌移植干预急性严重肝损伤的结局。方法:选雄性BALB/c小鼠40只,随机分为空白对照组10只,模型组10只,普通粪菌移植组10只,粪菌+益生菌移植组10只,除空白对照组外,其余各组给予D-氨基半乳糖(3.0 g/kg)腹腔注射制备急性严重肝损伤模型,普通粪菌移植组和粪菌+益生菌移植组在造模同时分别给予普通粪菌液和益生菌+粪菌液灌肠(1次/d),48 h后取血清用于检测丙氨酸转氨酶、天冬氨酸转氨酶、总胆红素,取肝组织用于病理学检测。取结肠内容物用于提取DNA进行16S V3-V4区高通量测序,用生物信息学分析技术对测序结果进行可操作分类单元聚类分析,α多样性分析,β多样性分析,线性判别分析找到不同分组小鼠的结肠内容物特征性差异细菌。临床生物化学指标组间差异比较采用n t检验,16S V3-V4区测序结果组间差异采用Wilcoxon检验。n 结果:模型组小鼠血清肝脏生物化学指标高于其他3组,差异有统计学意义(n P < 0.05),模型组肝脏HE染色结果显示肝组织镜下呈严重炎性改变;普通粪菌移植组和粪菌+益生菌移植组较模型组明显减轻,炎症减轻。16S rRNA高通量测序分析结果显示,空白对照组小鼠群菌结构与其他3组的Shannon差异无统计学意义,Observed Species差异有统计学意义,菌群构成差异大,粪菌移植增加小鼠肠道内物种数量。β-多样性分析结果显示,空白对照组与其他3组的组间差异大于疾病组之间的组间差异,粪菌+益生菌移植组改变疾病小鼠肠道菌群结构的差异细菌多为产丁酸盐细菌。n 结论:粪菌+益生菌增强粪菌移植治疗效果,改善肝脏炎症,增加肠道内产丁酸盐细菌数量。“,”Objective:To observe the changes of gut flora in mice, and explore the outcome of fecal microbiota transplantation combined with probiotics in the intervention of severe acute liver injury.Methods:Forty male BALB/c mice were selected and randomly divided into blank control group (10 mice), model group (10 mice), ordinary fecal microbiota transplantation group (10 mice), and fecal microbiota + probiotics transplantation group (10 mice). An intraperitoneal injection of d-galactosamine (D-GalN 3.0g/kg) was given to every group except the blank control group to induce severe acute liver injury model. Simultaneously, ordinary fecal microbiota transplantation group and fecal microbiota + probiotics transplantation group and modeling group were given enema solutions (once a day). After 48 hours, fetched serum was taken to detect alanine transaminase, aspartate transaminase and total bilirubin, and liver tissue was taken for pathological detection. The colonic content was used to extract DNA for 16S V3-V4 high-throughput sequencing. The results of sequencing were analyzed by using bioinformatics analysis; including OTU cluster analysis, α diversity analysis, β diversity analysis, and linear discriminant analysis effect size (Lefse) to find the bacteria with different colonic content characteristics in different groups of mice. Differences in clinical biochemical indicators between groups were compared using t-test, and the differences between 16S V3-V4 region sequencing results were compared using Wilcoxon test.Results:Model group mice serum biochemical parameters were higher than the other three groups, and the difference was statistically significant (n P < 0.05). HE staining of liver sections showed severe inflammatory changes under the microscope in the model group. Ordinary fecal microbiota transplantation group and fecal microbiota + probiotic microbiota transplantation group had low levels of inflammation than the model group. The analysis results of 16S rRNA high-throughput sequencing showed that there was no statistically significant difference in Shannon’s index between the blank control and the other three groups. Observed Species difference was statistically significant, and the gut flora composition varied greatly. Species number in the mice gut flora was increased with fecal microbiota transplantation. The results of β - diversity analysis showed that the difference between the blank control group and the other three groups was greater than that between the disease groups. The difference in the structure of the gut flora of the diseased mice in the fecal microbiota + probiotic transplantation group was mostly butyrate-producing bacteria.n Conclusion:Fecal microbiota + probiotics enhance the therapeutic effect of fecal microbiota transplantation, improve liver inflammation, and increase the number of butyrate-producing bacteria in the gut.