目的 基于微生物组学探讨肠复方抑制裸鼠肠癌原位移植瘤的作用机制。方法 复制荧光标记HCT-116 肠癌细胞原位移植瘤裸鼠模型。术后第10天，采用小动物活体成像系统检测小鼠腹腔荧光信号。将模型复制 成功的小鼠随机分为模型组、肠复方组（36 g·kg-1）、抗生素组、抗生素联合肠复方组（简称联合组），另取健康 裸鼠作为空白对照组，每组 6只。灌胃给药（20 mL·kg-1），每日 1次，连续干预 4周。实验期间每 2 d称小鼠体 质量并记录；采用小动物活体成像系统评估小鼠体内肿瘤生长情况；HE染色法观察肠癌组织病理变化；免疫 组化法检测肠癌组织 Ki67表达水平；采集小鼠粪便，进行肠道菌群 16S rDNA 扩增子测序及生物信息学分析。 结果 （1）与空白对照组比较，模型组小鼠的体质量显著降低（P＜0.001）。与模型组比较，肠复方组小鼠体质 量显著升高（P＜0.01），肠癌细胞总荧光值明显降低（P＜0.05），肠癌组织中 Ki67 表达显著下调（P＜0.001）； 抗生素组小鼠体质量明显降低（P＜0.05），肠癌细胞总荧光值有升高趋势，但差异无统计学意义（P＞0.05）， 肠癌组织中Ki67表达显著上调（P＜0.01）。与抗生素组比较，联合组小鼠体质量显著升高（P＜0.001），肠癌细 胞总荧光值明显降低（P＜0.05），肠癌组织中 Ki67 表达显著下调（P＜0.001）。（2）与空白对照组比较，模型组 的 chao1 指数明显下降（P＜0.05），变形菌门占比明显增加（P＜0.05），屎肠球菌、克雷伯杆菌占比明显增加 （P＜0.05），肠道菌群的双组分系统丰度明显上升（P＜0.05）。与模型组比较，肠复方组的chao1、simpson指数 均无明显变化（P＞0.05），疣微菌门占比增加（P＞0.05），Akk菌占比增加（P＞0.05），LEfSe分析显示疣微菌门、 Akk菌为优势菌种，肠道菌群的双组分系统及肽酶丰度明显下降（P＜0.05）；抗生素组的simpson指数明显下降 （P＜0.05），chao1 指数无明显变化（P＞0.05），变形菌门占比显著增加（P＜0.01），克雷伯杆菌占比显著增加 （P＜0.01），拟杆菌占比明显减少（P＜0.05），肠道菌群的双组分系统、转运蛋白和ABC转运蛋白、转录因子丰 度显著上升（P＜0.001），DNA修复和蛋白质重组、嘌呤代谢和核糖体、肽酶丰度显著下降（P＜0.001）。与抗生 素组比较，联合组的chao1、simpson指数均无明显变化（P＞0.05），厚壁菌门占比显著增加（P＜0.01），屎肠球 菌占比显著减少（P＜0.01），肠道菌群的嘌呤代谢丰度明显上升（P＜0.05）。结论 HCT-116肠癌细胞原位移植 瘤裸鼠中存在肠道菌群紊乱，抗生素会加剧肠道菌群紊乱。肠复方可抑制肠癌生长，其作用可能与调节肠道菌 群结构有关，Akk菌可能是肠复方的主要靶标之一。

Objective To investigate the mechanism of the inhibitory effect of intestinal compound on orthotopic transplantation of intestinal cancer in nude mice through the analysis of microbiomics. Methods The orthotopic transplantation nude mouse model of fluorescently labeled HCT-116 intestinal cancer cells was established. On the 10 th day of post-operation，small animal imaging system was used to detect fluorescence signals in the abdominal cavity of mice. The successfully modeled mice were randomly divided into model group，intestinal compound group（36 g·kg-1）， antibiotic group，antibiotic-combined intestinal compound group，and blank control group，with 6 mice in each group. The corresponding drug was administered at 20 mL·kg-1 /day orally for 4 consecutive weeks. The body weight of mice in each group were observed at every two day intervals. Tumor growth in mice was evaluated by using small animal imaging system. HE staining was used for detection of histopathological changes in various groups of intestinal cancer， and immunohistochemistry was applied for detection of Ki67 expression. Feces from mice in each group were collected for high throughput 16S rDNA amplicon sequencing and bioinformatic analysis. Results （1）Compared with the blank group，the body weight of mice in the model group was significantly decreased（P＜0.001）. Compared with the model group，the body weight of mice in the intestinal compound group was elevated（P＜0.01），the total fluorescence value and Ki67 expression of intestinal cancer tissue in the intestinal compound group obviously decreased（P＜0.05，P＜ 0.001）. The body weight of the antibiotic group was reduced（P＜0.05），and there was a trend of increase in the total fluorescence value of intestinal cancer cells， but the difference was not statistically significant （P＜0.05）. Ki67 expression of intestinal cancer tissue significantly up-regulated（P＜0.01）. Compared with the antibiotic group， the body weight of the mice in the combination group was elevated（P＜0.001），the total fluorescence value of intestinal cancer cells significantly decreased（P＜0.05）， and Ki67 expression of intestinal cancer tissue significantly downregulated（P＜0.001）.（2）Compared with the blank group，Chao1 index in the model group was significantly decreased （P＜0.05）. The proportion of Proteobacteria， Enterococcus faecalis and Klebsiella were remarkably increased（P＜ 0.05）. The abundance of the two-component system in the gut microbiota significantly increased（P＜0.05）. Compared with the model group， Chao1 and Simpson indices showed no obvious change （P＞0.05）. The proportion of Verrucomicrons and AKK bacteria increased（P＞0.05）. LEfSe analysis showed that Verrucobacteria and Akk bacteria were dominant bacterial species， the abundance of the two-component system in the gut microbiota and peptidase significantly decreased（P＜0.05）. Simpson indice in the antibiotic group significantly decreased（P＜0.05）. There was no significant differences in Chao1 index（P＞0.05）. The proportion of Proteobacteria and Klebsiella were remarkably increased（P＜0.01），while the proportion of Bacteroidetes was significantly decreased（P＜0.05）. The abundance of the two-component system in the gut microbiota，transporters，ABC transporters and transcription factors significantly increased（P＜0.001），but the abundance of DNA repair and protein recombination，purine metabolism，ribosome and peptidase significantly decreased（P＜0.001）. Compared with the antibiotic group，Chao1 and Simpson indices in the combination group showed no obvious change（P＞0.05）. The proportion of Firmicutes increased significantly（P＜ 0.01）， while the proportion of Enterococcus faecalis decreased significantly （P＜0.01）. The abundance of purine metabolism in the gut microbiota increased significantly（P＜0.05）. Conclusion There are intestinal flora disorder in nude mice with orthotopic transplantation of HCT-116 intestinal cancer cells， and antibiotics can exacerbate the intestinal flora disorder. Intestinal compound can inhibit the growth of intestinal cancer，its mechanism may be related to the regulation of the structure of intestinal flora. AKK bacteria may be the main target for intestinal compound.

R285.5

湖南省中医药管理局重点项目（2021035）；湖南省“十四五”第二批中医药学科带头人项目