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游丰锋,杨光勇,涂小华,何亚敏,喻良锦,李雨彤,李灿,何光志.保和丸对溃疡性结肠炎模型小鼠肠道益生菌A.muciniphila菌变化及肠黏膜屏障的影响[J].浙江中西医结合杂志,2021,31(4):
保和丸对溃疡性结肠炎模型小鼠肠道益生菌A.muciniphila菌变化及肠黏膜屏障的影响
Effect of Baohe Pill on intestinal probiotics A. muciniphila and intestinal mucosal barrier in mice with ulcerative colitis
投稿时间:2020-11-10  修订日期:2021-03-16
DOI:
中文关键词:  小鼠  保和丸  溃疡性结肠炎  ERIC-PCR  半定量PCR  嗜黏蛋白益生菌
英文关键词:mice  Baohe Pill  ulcerative colitis  ERIC-PCR  semi quantitative PCR  mucinophilic probiotics
基金项目:贵州省科技创新人才团队(黔科合平台人才[2020]5010)
作者单位E-mail
游丰锋 贵州中医药大学 975093911@qq.com 
杨光勇 贵州中医药大学 768305874@qq.com 
涂小华 基础医学院  
何亚敏 贵州中医药大学第一临床医学院  
喻良锦 贵州中医药大学第一临床医学院  
李雨彤 贵州中医药大学第二临床医学院  
李灿 贵州中医药大学第二临床医学院  
何光志 基础医学院  
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中文摘要:
      目的 观察保和丸对溃疡性结肠炎(UC)模型小鼠肠道嗜黏蛋白阿克曼氏菌(A.muciniphila)菌变化及肠黏膜屏障的影响。方法 40只SPF级6周龄KM小鼠按随机数字表法分为空白对照组、模型组、美沙拉嗪组和保和丸组,每组10只。除空白对照组外,其余各组参照文献方法,采用3%葡聚糖硫酸钠(DSS)诱导制备UC小鼠模型,空白对照组、模型组小鼠给予等体积蒸馏水灌胃,美沙拉嗪组和保和丸组小鼠分别按0.1 mL/10g鼠重给药,每次给予0.2 mL美沙拉嗪溶液(0.065 g/kg,每天3次)和保和丸溶液(0.78 g/kg,每天2次)灌胃,连续给药7天。计算小鼠病活动指数评分(DAI),采用肠杆菌基因间重复共有序列(ERIC)-聚合酶链式反应(PCR)检测粪便菌群多样性,半定量PCR检测各组A.muciniphila细菌变化,病理切片HE染色观察各组小鼠结肠组织病理。结果 药物干预前,各实验组小鼠DAI评分显著高于空白对照组(P均<0.01);药物干预后,美沙拉嗪组和保和丸组小鼠DAI评分均显著低于本组干预前和对照组[干预3天:(0.61±0.39)分比(1.33±0.47)分、(1.19±0.41)分,(0.69±0.42)分比(1.31±0.40)分、(1.19±0.41)分,P<0.01或P<0.05;干预7天:(0.36±0.24)分比(1.33±0.47)分、(0.84±0.31)分,(0.38±0.26)分比(1.31±0.40)分、(0.84±0.31)分,P<0.01或P<0.05],美沙拉嗪组和保和丸组药物干预后DAI评分比较,差异无统计学意义(P>0.05);ERIC-PCR结果显示,与空白对照组比较,模型组条带数量无明显减少,但条带位置变化较明显,保和丸组和美沙拉嗪组带数量明显减少,肠道菌群的多样性降低;半定量PCR结果显示,与空白对照组比较,模型组条带稍明显减少,美沙拉嗪组亮度稍暗,保和丸组亮度明显升高,即A.muciniphila丰度升高,基因全长329 bp;组织病理结果显示,与空白对照组比较,模型组小鼠结肠组织隐窝和腺体正常结构被破坏,发现大量炎性细胞浸润到黏膜层,与模型组比较,美沙拉嗪组和保和丸组黏膜损伤程度小,腺体结构破坏较轻,炎性细胞的数量和浸润深度都显著减少。结论 保和丸能调节DSS诱导的UC模型小鼠肠道菌群,提高A.muciniphila菌群丰度,可改善UC模型小鼠炎症。
英文摘要:
      Objective:To observe the effect of Baohe Pill on intestinal A. muciniphila and intestinal mucosal barrier in mice with ulcerative colitis (UC). Methods:Forty SPF grade 6-week-old KM mice were randomly divided into blank control group, model group, mesalazine group and Baohe Pill group, with 10 mice in each group. In addition to the blank control group, the other groups were induced by 3% dextran sodium sulfate (DSS) to prepare UC mice model. The blank control group and model group were given equal volume of distilled water by gavage. The mesalazine group and Baohe Pill group were given 0.1 ml/10g mouse weight, 0.2 ml mesalazine solution (0.065 g/kg, 3 times a day) and Baohe Pill solution (0.78 g/kg, twice a day) each time for 7 days. The disease activity index (DAI) score of mice was calculated. The fecal flora diversity was detected by Enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR). The changes of A. muciniphila in each group were detected by semi quantitative PCR. The colonic pathology of each group was observed by HE staining. Result: Before drug intervention, DAI scores of mice in each experimental group were significantly higher than those in the blank control group (P<0.01); after drug intervention, DAI scores of mice in mesalazine group and Baohe Pill group were significantly lower than those before intervention and control group [intervention for 3 days: (0.61±0.39) vs. (1.33±0.47), (1.19±0.41), (0.69±0.42) vs. (1.31±0.40), (1.19±0.41), P<0.01] The DAI scores of mesalazine group and Baohe Pill group after drug intervention had no statistical significance (P>0.05); ERIC-PCR results showed that compared with the blank control group, the number of bands in the model group was significantly lower, but the change of band position was obvious. The number of bands in Baohe Pill group and mesalazine group decreased significantly, and the diversity of intestinal flora decreased; The results of semi quantitative PCR showed that compared with the blank control group, the number of bands in the model group was slightly decreased, the brightness in the mesalazine group was slightly dim, and the brightness in the Baohe Pill group was significantly increased, that is, the abundance of A. muciniphila was increased, and the full length of the gene was 329 bp; The Histopathologic study showed that compared with the blank control group, the normal structure of crypt and gland of colon tissue in the model group was destroyed, and a large number of inflammatory cells were found to infiltrate into the mucosal layer. Compared with the model group, the degree of mucosal damage in the mesalazine group and Baohe Pill group was less, the destruction of gland structure was lighter, and the number and infiltration depth of inflammatory cells were significantly reduced. Conclusion: Baohe Pill can regulate the intestinal flora of UC model mice induced by DSS, increase the abundance of A. muciniphila flora, and improve the inflammation of UC model mice.
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