高剂量单宁酸对小鼠肠道屏障和肠道菌群的影响

doi:10.13560/j.cnki.biotech.bull.1985.2021-0860

摘要/Abstract

摘要：

以正常饮食小鼠和高脂饮食诱导肥胖模型小鼠为对象,研究高剂量单宁酸对小鼠肠道屏障及肠道菌群的影响。通过H&E染色、RT-qPCR、16S rRNA测序等方法进行检测分析。研究发现高剂量单宁酸（400 mg/kg BW）灌胃可使小鼠的体重和进食量降低,增加小鼠体内各肠段内容物的含量,其中结肠内容物的含量显著增加。并且高剂量单宁酸干预后可造成肠道功能损伤及肠道屏障破坏,如杯状细胞数量和隐窝长度减少,及肠道紧密连接蛋白（ZO-1,Occludin及Claudin）表达降低。此外,口服高剂量单宁酸会导致小鼠结肠肠道菌群的多样性发生变化,并增加了SCFA产生菌Alistipes,Ruminococcus和 Blautia以及肥胖负相关菌群Alistipes和Oscillibacter的含量。结果表明,高剂量单宁酸对肠黏膜屏障的破坏影响了小鼠对食物的消化吸收,这可能是高剂量单宁酸干预后小鼠体重快速下降的主要潜在原因。另一方面,高剂量单宁酸所造成的肠道菌群变化也会对小鼠的体重产生影响。

关键词: 单宁酸, 结肠内容物, 肠道屏障, 肠道菌群

Abstract:

The objective of this study is to analyze the effects of high-dose tannic acid on the intestinal barrier and gut microbiota in normal-diet mice and high-fat diet-induced obese model mice. H&E staining,RT-qPCR,16S rRNA sequencing and other methods were used for detection and analysis. It was found that the intervention of high-dose tannic acid（400 mg/kg）reduced the body weights and food intakes of the mice,and increased the contents of various intestinal segments in the mice,among which the colon content significantly increased. In addition,high-dose tannic acid damaged the intestinal function and intestinal barrier,such as reduced the number of goblet cell and the length of crypt,and decreased intestinal tight junction protein（ZO-1,Occludin,and Claudin）expression. Moreover,oral administration of high-dose tannic acid changed the diversity of gut microbiota in the colon,and increased the abundance of SCFAs-producing bacteria Alistipes,Ruminococcus and Blautia,as well as obesity negatively related bacteria Alistipes and Oscillibacter. In conclusion,the results showed that the damage of intestinal mucosal barrier caused by high-dose tannic acid affected the digestion and absorption of food in mice,which might be the main potential reason for the rapid weight loss in the mice after the intervention of high-dose tannic acid. On the other hand,the changes of gut microbiota caused by high-dose tannic acid also had an effect on the body weights of the mice.

Key words: tannic acid, colon content, intestinal barrier, gut microbiota

何亚伦, 曾丽荣, 刘雄, 张铃, 王琼. 高剂量单宁酸对小鼠肠道屏障和肠道菌群的影响[J]. 生物技术通报, 2022, 38(4): 278-287.

HE Ya-lun, ZENG Li-rong, LIU Xiong, ZHANG Ling, WANG Qiong. Effects of High-dose Tannic Acid on the Intestinal Barrier Function and Gut Microbiota in Mice[J]. Biotechnology Bulletin, 2022, 38(4): 278-287.

图/表 8

表1 RT-qPCR引物序列

Table 1 Primers’ sequences for RT-qPCR

Gene	Forward primer（5'-3'）	Reverse primer（5'-3'）
ZO 1	GCTTTAGCGAACAGAAGGAGC	TTCATTTTTCCGAGACTTCACCA
Occludin	CACACTTGCTTGGGACAGAG	TAGCCATAGCCTCCATAGCC
GAPDH	TGGCCTTCCGTGTTCCTAC	GAGTTGCTGTTGAAGTCGCA

图1 高剂量单宁酸对小鼠体重的影响 A：4组小鼠在实验期间的体重变化情况;B：实验结束后小鼠的体重增加量;C：实验期间各组小鼠的周均饮食摄入量;D：实验结束后各组小鼠的脂肪组织重量。误差线表示均值±标准误,*（0.01<P<0.05）,**（0.001<P<0.01）,***（P<0.001）,下同

Fig. 1 Effects of high-dose tannic acid on the body weights of the mice A：Changes in the body weights of the four groups of mice during the experiment. B：Weight gains of mice after the end of the experiment. C：The weekly dietary intake of each group during the experiment. D：The weight of adipose tissue in each group after the end of the experiment. The error lines in the figure all are the mean ± standard error,and *（0.01<P<0.05）,**（0.001<P<0.01）,and ***（P<0.001）,the same below

图2 高剂量单宁酸对小鼠体脂的影响 A：脂肪组织H&E染色切片;B：脂滴面积

Fig. 2 Effects of high-dose tannic acid on the body fats of the mice A：H&E staining sections of adipose tissue. B：Fat droplet area

图3 高剂量单宁酸对肠道结构的影响 A：小鼠肠道组织重量;B：小鼠肠道组织内容物重量;C：结肠组织H&E染色;D：杯状细胞数量;E：隐窝长度;F：组织损伤评分

Fig. 3 Effects of high-dose tannic acid on the intestinal structure A：Weights of intestinal tissues of the mice. B：Weights of intestinal tissue contents of the mice. C：H&E staining of colon tissue. D：Number of goblet cells. E：Crypt length. F：Tissue damage score

图4 高剂量单宁酸对肠道屏障的影响 A：肠道Muc2 mRNA含量的变化;B：肠道屏障功能相关基因mRNA含量的变化

Fig. 4 Effect of high dose tannic acid on the intestinal barrier A：Changes in intestinal Muc2 mRNA content. B：Changes in the mRNA levels of genes related to intestinal barrier function

图5 高剂量单宁酸对肥胖小鼠肠道微生物多样性的影响 A：Alpha多样性指数分析;B：Beta多样性分析;C：Venn图;D：门水平热图

Fig. 5 Effects of high dose tannic acid on the intestinal microbial diversity of the obese mice A：Alpha diversity index analysis. B：Beta diversity analysis. C：Venn diagram. D：Heatmap at the phylum level

图6 在门水平上的物种组成成分及组间差异分析 A：门水平群落Bar图;B、C：组间差异分析-多组比较

Fig. 6 Analysis of species composition and inter-group differences at the phylum level A：Community bar plot at the phylum level. B and C：Intergroup difference analysis - multi-group comparison

图7 在属水平上的物种组成成分及组间差异分析 A：属水平群落Bar图;B、C：组间差异分析-多组比较

Fig. 7 Analysis of species composition and inter-group differences at the genus level A：Community bar plot at the genus level. B and C：Intergroup difference analysis - multi-group comparison

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