Objectives: To explore the effect of acupuncture and moxibustion on intestinal flora in the rats with diarrhea-predominant irritable bowel syndrome (IBS-D) based on 16S rDNA technique.
Methods: Ten rats were randomized from 58 SPF-grade male SD rats to be the blank group. The remained 48 rats were prepared to be IBS-D models by the modified method of acetic acid enema combined with binding tail-clip stress. Forty successfully-modeled rats were randomly divided into a model group, an acupuncture group, a moxibustion group and a western medication group, with 10 rats in each one. In the acupuncture group, the needle was inserted at bilateral "Zusanli" (ST 36) and remained for 15 min in each rat. In the moxibustion group, the suspending moxibustion was delivered at bilateral "Zusanli" (ST 36) for 15 min. The rats in the western medication group were given pinaverium bromide suspension (10 mL/kg) by intragastric administration. The above interventions were performed once daily for consecutive 14 days. The body mass and the score of fecal trait were compared before and after modeling, as well as after intervention in each group. Fecal water content, diarrhea index and colon transit time (CTT) were measured after modeling and intervention in the rats of each group separately. After intervention, the colonic morphology of rats in each group was observed, and using 16S rDNA technique, the intestinal flora was detected.
Results: After modeling, compared with the blank group, the body mass and CTT were reduced (P<0.01); fecal trait scores, fecal water contents and diarrhea index increased (P<0.01) in the other 4 groups. After intervention, the body mass and CTT of the rats decreased (P<0.01), and fecal trait score, fecal water content and diarrhea index increased (P<0.01) in the model group compared with those in the blank group. In the acupuncture group, the moxibustion group and the western medication group, when compared with the model group, the body mass and CTT were elevated (P<0.01), while fecal trait scores, fecal water contents and diarrhea index declined (P<0.01). Compared with the western medication group, fecal water content decreased in the acupuncture group and the moxibustion group (P<0.05), while CTT increased in the acupuncture group (P<0.01), the body mass increased and fecal trait score was dropped in the moxibustion group (P<0.05). The colonic mucosa structure was clear and complete, and there was no obvious inflammatory cell infiltration in the blank group. The mild interstitial edema of intestinal mucosa was presented with the infiltration of few inflammatory cells in the model group. There was the infiltration of few inflammatory cells in the mucosa of the acupuncture group, the moxibustion group and the western medication group. Compared with the blank group, the indexes of Richness, Chao1, ACE and Shannon decreased in the model group (P<0.05). Indexes of Richness, Chao1 and ACE increased in the acupuncture group and the moxibustion group (P<0.05), and the Richness index in the western medication group increased (P<0.05) when compared with those in the model group. The relative abundance of Bacteroidetes, Proteobacteria and Prevotella increased (P<0.05), and that of Firmicutes and Muribaculaceae decreased (P<0.05) in the model group compared with those in the blank group. When compared with the model group, the relative abundance of Bacteroidetes, Proteobacteria and Prevotella was reduced (P<0.05), while that of Firmicutes and Muribaculaceae increased (P<0.05) in the acupuncture group, the moxibustion group and the western medication group; and that of Actinobacteria and Bifidobacterium increased in the acupuncture group and the moxibustion group (P<0.05). Compared with the blank group, the relative abundance of lipopolysaccharide (LPS) biosynthesis was elevated (P<0.05), and that of folate biosynthesis, lipoic acid metabolism, zeatin biosynthesis, ubiquinone and other terpenoid quinone biosynthesis decreased (P<0.05) in the model group. The relative abundance of LPS biosynthesis was dropped (P<0.05), and that of folate biosynthesis, lipoic acid metabolism, zeatin biosynthesis, ubiquinone and other terpenoid quinone biosynthesis increased (P<0.05) in the acupuncture group, the moxibustion group and the western medication group compared with those of the model group.
Conclusions: Either acupuncture or moxibustion can relieve the symptoms of IBS-D and protect intestinal mucosa, which may be associated with regulating the structure of intestinal flora and promoting nutrient metabolism and biosynthesis.
目的: 基于16S rDNA技术探讨针刺、艾灸对腹泻型肠易激综合征（IBS-D）模型大鼠肠道菌群的影响。方法: 从58只SPF级雄性SD大鼠中随机选取10只作为空白组。剩余48只大鼠采用乙酸灌肠联合束缚夹尾应激的改良方法制备IBS-D模型，将造模成功的40只大鼠随机分为模型组、针刺组、艾灸组和西药组，每组10只。针刺组大鼠予针刺双侧“足三里”，每次15 min；艾灸组大鼠予悬灸双侧“足三里”，每次15 min；西药组大鼠予匹维溴铵悬浊液（10 mL/kg）灌胃。均每天1次，连续干预14 d。比较各组大鼠造模前后及干预后的体质量及粪便性状评分。观察各组大鼠造模后和干预后的粪便含水量、腹泻指数、结肠运转时间。干预后，观察各组大鼠结肠组织形态，采用16S rDNA技术检测各组大鼠肠道菌群。结果: 造模后，与空白组比较，其他4组大鼠体质量、结肠运转时间降低（P<0.01），粪便性状评分、粪便含水量、腹泻指数升高（P<0.01）。干预后，与空白组比较，模型组大鼠体质量、结肠运转时间降低（P<0.01），粪便性状评分、粪便含水量、腹泻指数升高（P<0.01）；与模型组比较，针刺组、艾灸组、西药组大鼠体质量、结肠运转时间升高（P<0.01），粪便性状评分、粪便含水量、腹泻指数降低（P<0.01）；与西药组比较，针刺组、艾灸组粪便含水量降低（P<0.05），针刺组结肠运转时间升高（P<0.01），艾灸组大鼠体质量升高、粪便性状评分降低（P<0.05）。空白组大鼠结肠黏膜结构清晰、完整，无明显炎性细胞浸润；模型组大鼠肠黏膜轻度间质水肿，有少量炎性细胞浸润；针刺组、艾灸组、西药组黏膜可见少量炎性细胞浸润。与空白组比较，模型组大鼠Richness指数、Chao1指数、ACE指数及Shannon 指数均降低（P<0.05）。与模型组比较，针刺组、艾灸组大鼠Richness指数、Chao1指数、ACE指数均升高（P<0.05），西药组大鼠Richness指数升高（P<0.05）。与空白组比较，模型组大鼠拟杆菌门（Bacteroidetes）、变形菌门（Proteobacteria）、普氏菌属（Prevotella）相对丰度升高（P<0.05），厚壁菌门（Firmicutes）、拟杆菌门S24-7菌属（Muribaculaceae）相对丰度降低（P<0.05）。与模型组比较，针刺组、艾灸组、西药组大鼠拟杆菌门、变形菌门、普氏菌属相对丰度降低（P<0.05），厚壁菌门、拟杆菌门S24-7菌属相对丰度升高（P<0.05）；针刺组、艾灸组大鼠放线菌门（Actinobacteria）、双歧杆菌属（Bifidobacterium）相对丰度升高（P<0.05）。与空白组比较，模型组大鼠脂多糖生物合成相对丰度升高（P<0.05），叶酸合成、硫辛酸代谢、玉米素生物合成、辅酶Q及其他萜醌合成相对丰度降低（P<0.05）；与模型组比较，针刺组、艾灸组、西药组大鼠脂多糖生物合成相对丰度降低（P<0.05），叶酸合成、硫辛酸代谢、玉米素生物合成、辅酶Q及其他萜醌合成相对丰度升高（P<0.05）。结论: 针刺、艾灸均可以改善IBS-D模型大鼠腹泻症状，保护肠道黏膜，可能与调节肠道菌群结构、促进营养代谢及生物合成有关。.
Keywords: acupuncture; diarrhea-predominant irritable bowel syndrome; gene function prediction; intestinal flora; moxibustion.