结肠直肠癌患者肠道黏膜区域的菌群分布

Changes in the structure of intestinal mucosal flora in colorectal cancer patients

目的比较健康人群与结肠直肠癌(CRC)患者粪便菌群及肿瘤/瘤旁黏膜菌群的差异性,尝试寻找CRC新的防治途径。方法收集了13例CRC患者粪便、6例患者肿瘤/瘤旁黏膜和20名健康对照人群粪便,利用16SrDNA测序技术与生物信息学方法分别对粪便和黏膜的菌群组成差异进行了分析。在流式细胞仪检测粪便中的细菌总数后,采用几种较为常见的细菌培养基对粪便及黏膜中微生物进行分离鉴定,并利用乳酸脱氢酶检测试剂盒测试了其中7株细菌对3株肠癌细胞凋亡的影响。结果健康人粪便和CRC患者瘤旁黏膜中的菌群α-多样性显著高于CRC患者粪便和肿瘤黏膜中菌群α-多样性(P<0.05)。从特异性微生物来看,与健康人群相比,CRC患者粪便中的特有菌为Lactobacillaceae,而其肿瘤黏膜上特有菌是Escherichia、Enterococcus、Fusobacterium等。通过体外实验发现,分离自CRC患者肿瘤黏膜处的Bacteroidesfragilis与Fusobacterium nucleatum分别对肠道癌细胞的增殖具有显著抑制活性(P<0.0001)与促进作用(P<0.001)。结论CRC患者粪便、肿瘤黏膜及瘤旁黏膜菌群与健康人群粪便菌群差异较大,且CRC患者粪便菌群不能代表肿瘤黏膜特异性菌群。体外实验结果表明,抑制肿瘤黏膜中F.nucleatum定植与促进具有癌细胞毒性的B.fragilis定植可能是治疗CRC潜在的有效方法。

Objective To investigate the changes in bacterial flora in fecal samples,at the tumor loci and in adjacent mucosa in patients with colorectal cancer(CRC).Methods We collected fecal samples from 13 patients with CRC and 20 healthy individuals and tumor and adjacent mucosa samples from 6 CRC patients.The differences in bacterial composition between the fecal and mucosa samples were analyzed with 16S rDNA sequencing and bioinformatics methods.We also detected the total number of bacteria in the feces using flow cytometry,isolated and identified the microorganisms in the fecal and mucosa samples using common bacterial culture media.We further tested the effects of 7 isolated bacterial strains on apoptosis of 3 CRC cell lines using lactate dehydrogenase detection kit.Results The bacterialα-diversity in the feces of healthy individuals and in adjacent mucosa of CRC patients was significantly higher than that in the feces and tumor mucosa in CRC patients(P<0.05).Lactobacillaceae is a specific bacteria in the feces,while Escherichia,Enterococcus,and Fusobacterium are specific bacteria in tumor mucosa of CRC patients as compared with healthy individuals.Cell experiment with3 CRC cell lines showed that Bacteroides fragilis isolated from the tumor mucosa of CRC patients produced significant inhibitory effects on cell proliferation(P<0.0001),while the isolated strain Fusobacterium nucleatum obviously promoted the proliferation of the cell lines(P<0.001).Conclusion The bacterial flora in the feces,tumor mucosa and adjacent mucosa of CRC patients is significantly different from that in the feces of healthy individuals,and the fecal flora of CRC patients can not represent the specific flora of the tumor mucosa.Inhibition of F.nucleatum colonization in the tumor mucosa and promoting B.fragilis colonization may prove beneficial for CRC treatment.