三味清热类中药水提物对肉仔鸡肠道菌群和黏膜屏障功能的影响

doi:10.16431/j.cnki.1671-7236.2020.07.009

摘要/Abstract

摘要： 为了研究单味中药对肠道菌群和肠黏膜损伤的调节和修复作用，选用金银花、连翘和紫花地丁三味清热类中药对大肠杆菌所致肉仔鸡肠道菌群失调和黏膜屏障功能障碍进行治疗，为单味中药在提高肠道细菌的有益菌群和保护肠黏膜屏障提供可靠的理论依据。选取100只1日龄雄性肉仔鸡，分成5组，每组20只，空白组正常饲喂，模型组和中药组于15~17 d腹腔注射大肠杆菌O₇₈（1.0×10⁶ CFU/mL），1 mL/只。中药组在第18天开始饮用中药，连续7 d，各中药组动物给药量为生药量1 g/kg。在给药后第7和14天，每组取6只鸡心脏釆血，采用ELISA法按照试剂盒说明进行D-乳酸（D-LA）、二胺氧化酶（DAO）和内毒素（ET）的检测。采集空肠肠段和盲肠。采用实时荧光定量PCR检测空肠紧密连接蛋白（（闭合小环蛋白-1（ZO-1）、闭锁蛋白（Occludin）和闭合蛋白-1（Claudin-1））的mRNA的相对表达量。盲肠送检测公司检测中药对盲肠菌群多样性的影响。试验结果表明，金银花、紫花地丁和连翘均可以降低给药后第7和14天肉仔鸡血清（DAO活性）、D-LA和ET含量。在给药后第7天，金银花组ZO-1和Occludin的mRNA相对表达量显著升高（P<0.05）；连翘组Claudin-1和Occludin的mRNA相对表达量显著升高（P<0.05）；在给药后第14天，金银花组ZO-1的mRNA相对表达量显著升高（P<0.05），紫花地丁组ZO-1和Claudin-1的mRNA相对表达量显著升高（P<0.05），连翘组ZO-1、Claudin-1的mRNA相对表达量显著升高（P<0.05）。金银花、紫花地丁和连翘均可提高给药后第7天厚壁菌门占比（P<0.05），金银花、紫花地丁可提高给药后第14天拟杆菌门占比（P<0.05），金银花、紫花地丁抑制给药后第7和14天变形菌门的增殖，紫花地丁和连翘抑制给药后第7天放线菌门的生长（P<0.05），连翘、金银花可以显著提高给药后7和14 d乳酸杆菌占比，紫花地丁可以显著提高给药后7 d乳酸杆菌占比（P<0.05），紫花地丁显著提高给药后第14天未分类毛螺旋菌种相关菌属的占比。综上，金银花、紫花地丁和连翘均可以降低肉仔鸡血清DAO活性、D-LA和ET含量，减少毒素入侵。金银花可以提高ZO-1、Occludin的mRNA相对表达量，连翘可提高ZO-1、Claudin-1和Occludin的mRNA相对表达量，保护肠道黏膜；紫花地丁可提高ZO-1、Claudin-1的mRNA相对表达量。金银花、紫花地丁和连翘均可使肠道OTU数目增多，在不同程度上促进有益菌群繁殖，抑制致病菌群生长，具有调节肠道菌群结构的作用。

关键词: 高通量测序; 清热类中药; 肠道菌群; 肠黏膜屏障

Abstract: In order to study the regulation and repair of intestinal flora and intestinal mucosal damage by single-flavor Chinese medicine,the intestinal flora of broiler chickens caused by Escherichia coli was selected from Chinese herbal medicines of Honeysuckle,Forsythia and Herba violet.It provided a reliable theoretical basis for improving the beneficial flora of intestinal bacteria and protecting intestinal mucosal barrier.100 male broilers of 1 day old were selected and divided into five groups,20 in each group.The blank group was fed normally.The model group and the Chinese medicine group were intraperitoneally injected with E.coli O₇₈ (1.0×10⁶ CFU/mL) at 15-17 days,1 mL each.The Chinese medicine group began to drink traditional Chinese medicine on the 18th day for 7 consecutive days.The dose of each Chinese medicine was 1 g/kg.On the 7 and 14 d after administration,6 chickens in each group were treated with blood stasis.D-lactic acid (D-LA),diamine oxidase (DAO) and endotoxin (ET) were performed according to the Kit instructions by ELISA.Jejunum and cecum were collected.The relative expression levels of jejunal tight junction proteins (ZO-1,Occludin,and Claudin-1) were detected by Real-time fluorescent quantitative PCR.Cecal delivery and detection company was sequenced to detect the influence of traditional Chinese medicine on the diversity of cecal flora.The results showed that Honeysuckle,Forsythia and Herba violet could reduce the levels of serum DAO,D-LA and ET on the 7th and 14th day after administration.On the 7th day after administration,the relative mRNA expression of ZO-1 and Occludin in Honeysuckle group was significantly increased (P<0.05).The relative mRNA expression of Claudin-1 and Occludin in the Forsythia group was significantly increased (P<0.05).On the 14th day after administration,the relative mRNA expression levels of ZO-1 in Honeysuckle group were significantly increased (P<0.05),while the relative mRNA expression levels of ZO-1 and Claudin-1 in Forsythia group and Herba violae were significantly increased (P<0.05).Honeysuckle,Forsythia and Herba violet promoted the proportion of Firmicutes on day 7 after administration (P<0.05),while Honeysuckle,Herba violet promoted the proportion of Bacteroidetes on day 14 after administration (P<0.05).Honeysuckle,Herba violet inhibited the proportion of Proteobacteria at 7 and 14 d after administration,and Herba violet and Forsythia inhibited the growth of Actinobacteria at 7 d after administration (P<0.05).Forsythia,Honeysuckle can significantly increase the proportion of Lactobacillus 7 and 14 d after administration,and Herba violet can significantly increase the proportion of Lactobacillus 7 d after administration (P<0.05).The proportion of the uncultured_bacterium_f_Lachnospiraceae on 14 d after administration was significantly increased by Herba violet.Honeysuckle,Herba violet and Forsythia can reduce the content of serum DAO,D-LA and ET in broilers.Honeysuckle could improve the relative mRNA expression of ZO-1,Occludin and Forsythia could contribute to the relative mRNA expression of ZO-1,Claudin-1 and Occludin.The relative mRNA expression levels of ZO-1 and Claudin-1 were increased by Herba violae.Honeysuckle,Herba violet and Forsythia could increase the number of intestinal OTU,promote the reproduction of beneficial bacteria,inhibit the growth of pathogenic bacteria,and regulate the structure of intestinal flora.

Key words: high-throughput sequencing; heat-clearing traditional Chinese medicine; intestinal flora; testinal mucosal barrier

中图分类号:

S859.3

梁小瑞, 王丹阳, 乔彦杰, 张保军, 孙国杰, 邵永斌, 谷新利, 罗燕. 三味清热类中药水提物对肉仔鸡肠道菌群和黏膜屏障功能的影响[J]. 中国畜牧兽医, 2020, 47(7): 2043-2054.

LIANG Xiaorui, WANG Danyang, QIAO Yanjie, ZHANG Baojun, SUN Guojie, SHAO Yongbin, GU Xinli, LUO Yan. Effects of Water Extract of Three Qingre Chinese Medicine on Intestinal Flora and Mucosal Barrier Function of Broilers[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(7): 2043-2054.

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