健脾合剂对脾虚腹泻型肠易激综合征小鼠肠道菌群的干预研究

Intervention Study of Jianpi Mixture on Intestinal Flora of Mice with Spleen Deficiency and Diarrhea-Type Irritable Bowel Syndrome

目的通过观察健脾合剂对脾虚腹泻型肠易激综合征(IBS-D)模型小鼠的肠道菌群的干预作用,研究健脾合剂对脾虚型IBS-D肠道微生物的影响。方法将清洁级幼龄ICR雄性小鼠48只随机分为四组(n=12),并建立脾虚型IBS-D小鼠模型(3n=36),造模评价成功后,空白组和模型组分别予双蒸水(10ml·kg^(-1))、西药组予复合乳酸菌(0.26 g·kg-1)、中药组予健脾合剂(16 ml·kg^(-1))灌胃,治疗周期均为14天。治疗周期结束后,每组随机抽取5只小鼠,分别标记为B1、B2、B3、B4组,随后通过16SrDNA高通量基因测序法分析治疗前后小鼠肠道菌群的变化。结果本研究测序深度充分,样品均具有良好的丰富度与多样性。与B1组比较,B2组小鼠的肠道菌群多样性明显降低(P <0.05);与B2组比较,B3、B4组小鼠的肠道菌群多样性明显提高(P <0.05);与B1组比较,B3、B4组小鼠肠道菌群多样性无明显差异(P> 0.05),说明健脾合剂和复合乳酸菌的干预均能调节脾虚型IBS-D小鼠肠道菌群达到或接近健康小鼠的肠道菌群稳态;且B3与B4组间的小鼠肠道菌群多样性无明显差异(P> 0.05),推测健脾合剂有益生元样作用;健脾合剂干预后,在门水平,Tenericutes、Actinobacteria、Saccharibacteria丰度上调(P <0.05),Proteobacteria丰度下调(P <0.05);在属水平,Alistipes、Ruminiclostridium、Candidatus_Saccharimonas、Rikenella、Intestinimonas、Lachnoclostridium、Oscillibacter、Coprococcus、Faecalibaculum、Enterorhabdus、Peptococcus、Romboutsia、Papillibacter丰度上调(P <0.05), Lactobacillus、 Bacteroides、 Parasutterella、 Escherichia、 Erysipelatoclostridium、 Parabacteroides、Candidatus_Stoquefichus丰度下调(P <0.05)。结论健脾合剂干预可以增加脾虚型IBS-D小鼠肠道菌群多样性,并调节到与健康小鼠相似的平衡状态。

Objective The rats with spleen deficiency and diarrhea-type irritable bowel syndrome(IBS-D)were treated with Jianpi Mixture to observe the intervention effect of Jianpi Mixture on the intestinal flora of mice.Method 48 young ICR male mice were randomly divided into four groups(n=12),and establish a mouse model of spleen deficiency type IBS-D(3n=36).After successful model evaluation,the blank group and the model group were given double steam(10 ml·kg-1)gavage,the western medicine group was given compound lactic acid bacteria 0.26 g·kg-1gavage,and the Chinese medicine group was given Jianpi mixture(16 ml·kg-1)intragastrically gavage.The treatment cycle was 14 days.At the end of the treatment period,5 mice were randomly selected from each group,and marked as B1,B2,B3,B4 group,and then the changes of intestinal microbiota in the mice before and after treatment were analyzed by 16S rDNA high-throughput gene sequencing.Results In the study,the sequencing depth was sufficient,and the samples all had good species richness and species diversity.Compared with group B1,the intestinal flora of mice in group B2 decreased(P<0.05).Compared with the group B2,the intestinal flora of mice in group B3 and group B4 increased(P<0.05);compared with group B1,there was no significant difference in the intestinal flora of mice in group B3 and group B4(P>0.05),indicating that the intervention of Jianpi Mixture and Compound Lactobacillus could regulate the intestinal flora of spleen-deficient IBS-D mice to reach or close to the intestinal tract of healthy mice.The stability of the intestinalf lora was not significantly different between group B3 and group B4(P>0.05),surmising the prebiotic effect of Jianpi Mixture.After the intervention of Jianpi Mixture,at the phylum level,Actinobacteria,Tenericutes,saccharibacteria were up-regulated(P<0.05),and the abundance of Proteobacteria was down-regulated(P<0.05).At the genu level,Alistipes,Ruminiclostridium,Candidatus_Saccharimonas,Rikenella,Intestinimonas,Lachnoclostridium,Oscillibacter,Coprococcus,Faecalibaculum,Enterorhabdus,Peptococcus,Romboutsia,Papillibacter were up-regulated(P<0.05),Lactobacillus,Bacteroides,Parasutterella,Escherichia,Erysipelatoclostridium,Parabacteroides,Candidatus_Stoquefichus were down-regulated(P<0.05).Conclusion Intervention with Jianpi mixture can increase the intestinal flora species diversity of spleen-deficient IBS-D mice and regulate to a similar equilibrium state to healthy mice.