基于16S rDNA测序分析胆囊结石及胆囊切除对结直肠癌患者肠道菌群的影响

Effects of cholecystolithiasis and cholecystectomy on intestinal flora of colorectal cancer patients based on 16S rDNA sequencing

目的探讨胆囊结石伴胆囊炎和胆囊切除对结直肠癌患者肠道菌群的影响。方法选取2020年6月至2021年3月期间就诊于甘肃省人民医院肛肠科的结直肠癌患者168例,最终纳入符合条件的29例结直肠癌患者作为研究对象,其中结直肠癌合并胆囊结石伴胆囊炎患者10例(胆囊结石伴胆囊炎+结直肠癌组),胆囊切除术后结直肠癌患者10例(胆囊切除+结直肠癌组),胆囊正常结直肠癌患者9例(正常胆囊+结直肠癌组)。收集并比较3组患者的临床资料;采集纳入研究患者的新鲜大便样本,采用16S rDNA高通量测序技术测定并分析获得样本肠道菌群的组成和分布情况。结果胆囊结石伴胆囊炎+结直肠癌组的白细胞介素-6较正常胆囊+结直肠癌组和胆囊切除+结直肠癌组升高(P<0.05)。3组患者的大便菌群在门水平上:①3组样本中拟杆菌(Bacteroidetes)门、厚壁菌(Firmicutes)门、变形杆菌(Proteobactenia)门、梭杆菌(Fusobacteria)门和疣微菌(Verrucomicrobia)门的相对丰度都较高,几乎占据了肠道细菌总量的95%以上;②梭杆菌门的相对丰度在胆囊结石伴胆囊炎+结直肠癌组较正常胆囊+结直肠癌组增高(P<0.05);③疣微菌门的相对丰度在正常胆囊+结直肠癌组较胆囊结石伴胆囊炎+结直肠癌组和胆囊切除+结直肠癌组增高(P<0.05);④互养菌(Synergistetes)门的相对丰度在胆囊切除+结直肠癌组较胆囊结石伴胆囊炎+结直肠癌组和正常胆囊+结直肠癌组增高(P<0.05)。在属水平上:①拟杆菌属和罗斯菌(Roseburia)属的相对丰度在胆囊结石伴胆囊炎+结直肠癌组较胆囊切除+结直肠癌组和正常胆囊+结直肠癌组均降低(P<0.05);②志贺杆菌(Shigella)属的相对丰度在胆囊切除+结直肠癌组较胆囊结石伴胆囊炎+结直肠癌组增高(P<0.05);③毛螺旋菌(Lachnospira)属的相对丰度在胆囊结石伴胆囊炎+结直肠癌组较正常胆囊+结直肠癌组降低(P<0.05);④普氏菌(Prevotella)属和梭杆菌属的相对丰度在胆囊结石伴胆囊炎+结直肠癌组较胆囊切除+结直肠癌组和正常胆囊+结直肠癌组均增高(P<0.05);⑤梭菌(Clostridium)属和阿克曼菌(Akkermansia)属的相对丰度在胆囊结石伴胆囊炎+结直肠癌组和胆囊切除+结直肠癌组较正常胆囊+结直肠癌组均降低(P<0.05);⑥肠球菌(Enterococcus)属的相对丰度在正常胆囊+结直肠癌组较胆囊切除+结直肠癌组增高(P<0.05)。结论①长期患胆囊结石伴胆囊炎会引起拟杆菌属、罗斯菌属、毛螺旋菌属等菌群丰度显著降低;②胆囊切除术后会引起梭菌属、肠球菌属、疣微菌门、互养菌门等菌群相对丰度改变;③梭杆菌属的相对丰度在结直肠癌合并胆囊结石伴胆囊炎患者显著增高,促进炎症细胞因子释放引起肠道炎症,有利于条件致病菌的生长,可能影响结直肠癌的发生发展。

Objective To investigate the effect of cholecystolithiasis with cholecystitis and cholecystectomy on intestinal flora in patients with colorectal cancer.Methods A total of 168 patients with colorectal cancer who admitted to the Department of Anorectal Surgery in Gansu Provincial Hospital from June 2020 to March 2021 were selected,and 29 patients with colorectal cancer who met the criteria were selected as the research objects,including 10 colorectal cancer patients with gallstones and cholecystitis(cholecystolithiasis with cholecystitis+colorectal cancer group),10 colorectal cancer patients after cholecystectomy(cholecystectomy+colorectal cancer group),and 9 colorectal cancer patients with normal gallbladder(normal gallbladder+colorectal cancer group).Clinical data of the patients in three groups were collected and compared.The fresh fecal samples of the patients included in the study were collected,and the 16S rDNA high-throughput sequencing method was used to determine and analyze the composition and distribution of the intestinal flora in the obtained samples.Results The interleukin-6 level in the cholecystolithiasis with cholecystitis+colorectal cancer group was statistically higher than that in the normal gallbladder+colorectal cancer group and the cholecystectomy+colorectal cancer group(P<0.05).At the phylum level of the fecal flora in three groups patients:①In the samples of three groups,the relative abundances of Bacteroidetes,Firmicutes,Proteobacteria,Fusobacteria and Verrucomicrobia phylums were all high,accounting for almost more than 95% of the total intestinal bacteria.②The relative abundance of Fusobacteria phylum in the cholecystolithiasis with cholecystitis+colorectal cancer group was statistically higher than that in the normal gallbladder+colorectal cancer group(P<0.05).③The relative abundance of Verrucomicrobia phylum in the normal gallbladder+colorectal cancer group was statistically higher than that in the cholecystolithiasis with cholecystitis+colorectal cancer group and the cholecystectomy+colorectal cancer group(P<0.05).④The relative abundance of Synergistetes phylum in the cholecystectomy+colorectal cancer group was statistically higher than that in the cholecystolithiasis with cholecystitis+colorectal cancer group and the normal gallbladder+colorectal cancer group(P<0.05).At the genus level:①The relative abundances of Bacteroidetes and Roseburia genus were lower in the gallstone with cholecystitis+colorectal cancer group than those in the cholecystectomy+colorectal cancer group and the normal gallbladder+colorectal cancer group(P<0.05).②The relative abundance of Shigella genus in the cholecystectomy+colorectal cancer group was higher than that in the cholecystolithiasis with cholecystitis+colorectal cancer group(P<0.05).③The relative abundance of the Lachnospira genus in the cholecystolithiasis with cholecystitis+colorectal cancer group was lower than that in the normal gallbladder+colorectal cancer group(P<0.05).④The relative abundances of Prevotella and Fusobacteria genus were higher in the cholecystolithiasis with cholecystitis+colorectal cancer group than that in the cholecystectomy+colorectal cancer group and the normal gallbladder+colorectal cancer group(P<0.05).⑤The relative abundances of Clostridium and Akkermansia genus were lower in the cholecystolithiasis with cholecystitis+colorectal cancer group and the cholecystectomy+colorectal cancer group than that in the normal gallbladder+colorectal cancer group(P<0.05).⑥The relative abundance of Enterococcus genus was higher in the normal gallbladder+colorectal cancer group than that in the cholecystectomy+colorectal cancer group(P<0.05).Conclusions①Long-term occurrence of cholecystolithiasis with cholecystitis can cause obvious decrease in the abundances of Bacteroides,Roseburia,Lachnospira,etc.②Cholecystectomy can cause changes in the relative abundances of Clostridium,Enterococcus,Verrucomicrobia,Synergistetes,etc.③The relative abundance of Fusobacterium is obviously increased in colorectal cancer patients with gallstones and cholecystitis,then promotes the release of inflammatory cytokines and causes intestinal inflammation,which is conducive to the growth of opportunistic pathogens,thus may affect the occurrence and development of colorectal cancer.