摘要
目的基于攻补兼施法探讨在大剂量生白术的配伍下不同枳实剂量配伍的作用及枳术丸(ZZW)对慢传输型便秘(STC)小鼠肠道菌群的影响。方法将65只SPF级别昆明(KM)小鼠随机分为空白对照组10只与模型组50只,空白对照组正常喂养,模型组使用番泻叶0.8 g/kg灌胃7 d,制成脾虚模型后禁食禁水,隔天喂食生大米,饮水0.5 h,连续8 d建立脾虚便秘模型。15 d后将模型组随机分为5组,每组10只,即模型组、西药组、ZZW低剂量组、ZZW中剂量组、ZZW高剂量组。ZZW低、中、高剂量分别为9.7 g/kg、11.6 g/kg、13.7 g/kg,西药组给予莫沙必利2.5 mg/kg。空白对照组和模型组灌胃同等量蒸馏水。治疗1周后收集肠道内容物,行16S rRNA测序,取结肠组织HE染色行病理形态学观察。结果ZZW高剂量组能够显著提高肠道推进率(P<0.05)。结肠切片肉眼观察发现小鼠结肠组织黏膜光滑,未见充血、糜烂及器质性损伤。HE病理学检查结果显示,模型组存在轻微水肿,腺体不齐现象,其余各组小鼠结肠黏膜无变薄、固有层炎症浸润,结肠腺体形态正常。16S rRNA测序结果显示,STC小鼠肠道菌群物种丰富度和均匀度显著下降(P<0.05),菌群多样性升高(P<0.01)。门水平上,与空白对照组比较,模型组拟杆菌门相对丰度升高、厚壁菌门相对丰度降低,但差异无统计学意义(P>0.05)。与模型组比较,ZZW低剂量组较模型组拟杆菌门相对丰度显著降低(P<0.01),厚壁菌门、变形菌门相对丰度升高(P<0.05),ZZW高剂量组较模型组拟杆菌门相对丰度降低(P<0.05),厚壁菌门相对丰度升高(P<0.05)。属水平上,模型组较空白对照组Prevotellamassilia、葡萄球菌属、拟杆菌属、丁酸弧菌属相对丰度升高(P<0.05),螺杆菌属、另枝菌属相对丰度显著降低(P<0.01),经治疗后ZZW低剂量组Prevotellamassilia、葡萄球菌属、拟杆菌属相对丰度降低(P<0.05),ZZW中剂量组丁酸弧菌属相对丰度降低(P<0.05);ZZW高剂量组Prevotellamassilia、葡萄球菌属相对丰度显著降低(P<0.01)。此外,与模型组比较,ZZW低剂量组狄氏副拟杆菌属相对丰度降低(P<0.05)。结论攻补兼施代表方ZZW能明显缓解STC小鼠便秘症状,以ZZW高剂量组效果最佳,但治疗效果与枳实剂量不存在量效关系,其治疗机制可能与ZZW水煎液调整菌群丰度、多样性及菌群物种数目,恢复正常肠道生态环境有关。
Objective To observe the effects of different doses of Aurantii Fructus Immaturus(AFI)in combination with large dose of Atractylodis Macrocephalae Rhizoma and the effects of Zhizhuwan(ZZW)on intestinal flora of mice with slow transit constipation based on the method of combination of attacking and replenishing.Methods Sixty-five SPF KM mice were randomly divided into control group(n=10)and model group(n=50).The control group was fed normally.The model group was given 0.8 g/kg senna leaf by gavage for 7 days to make spleen deficiency model,followed by food and water fasting.After 15 days,the model group was randomly divided into five groups(n=10),model group,western medicine group,and low-dose,medium-dose and high-dose AFI groups.The western medicine group was given 2.5 mg/kg mosapride,while the control group and model group were given the same amount of distilled water.After 1 week of treatment,the intestinal contents were collected for 16S rRNA gene sequencing,and colon tissues were HE stained for pathomorphological observation.Results The intestinal propulsion rate significantly increased in the high-dose AFI group(P<0.05).Macroscopic observation of colon section showed that the mucosa of colon tissue was smooth without hyperemia,erosion or organic damage.HE pathological examination showed slight edema and glandular asymmetry in the model group.There was no thinning of colonic mucosa and lamina propria inflammatory infiltration in other groups,and the morphology of colonic glands was normal.The results of 16S rRNA sequencing showed that the species richness and diversity of intestinal microflora of STC mice significantly decreased.At the phylum level,compared with the control group,the abundance of Bacteroidetes in the model group increased and that of Firmicutes decreased,but without statistical significance(P>0.05).Compared with the model group,the abundance of Bacteroidetes in low-dose AFI group significantly decreased(P<0.01),while those of Firmicutes and Proteobacteria significantly increased(P<0.05);the abundance of Bacteroidetes and Firmicutes in high-dose ZZW group significantly decreased,while that of Firmicutes significantly increased(P<0.05).At the genus level,the abundances of Prevotellamassilia,Staphylococcus,Bacteroides and Butyricimonas in model group increased significantly(P<0.05),while that of Alistipes decreased significantly(P<0.01),compared with the control group.After treatment,the abundances of Prevotellamassilia,Staphylococcus and Bacteroides significantly reduced in low dose ZZW group(P<0.05),that of Butyricimonas in medium dose ZZW group significantly decreased(P<0.05),and those of Prevotellamassilia and Staphylococcus in high dose ZZW group significantly decreased(P<0.01).In addition,compared with the model group,the abundance of Parabacteroides in low-dose ZZW group decreased(P<0.05).Conclusion The attacking-replenishing based ZZW can significantly alleviate the symptoms of constipation of STC mice,with the best effect seen in high dose ZZW group,though there was no dose-effect relationship between the efficacy and the dose of AFI.The mechanism may be related to the restoration of normal intestinal ecological environment by ZZW through adjusting the abundance,diversity and amount of flora.
作者
张婷
刘富林
谭周进
夏旭婷
滕广飞
施敏
廖陈敏
ZHANG Ting;LIU Fu-lin;TAN Zhou-jin;XIA Xu-ting;TENG Guang-fei;SHI Min;LIAO Chen-min(College of Traditional Chinese Medicine,Hu’nan University of Traditional Chinese Medicine,Changsha,Hu’nan 410208,China)
出处
《中国微生态学杂志》
CAS
CSCD
2022年第11期1275-1282,共8页
Chinese Journal of Microecology
基金
湖南省教育厅重点项目(21A0232)
湖南省自然科学基金(2022JJ30437)。
