Sunny H Wong1, Liuyang Zhao1, Xiang Zhang1, Geicho Nakatsu1, Juqiang Han2, Weiqi Xu1, Xue Xiao1, Thomas N Y Kwong1, Ho Tsoi1, William K K Wu3, Benhua Zeng4, Francis K L Chan1, Joseph J Y Sung1, Hong Wei5, Jun Yu6. 1. Institute of Digestive Disease, Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences and CUHK-Shenzhen Research Institute, The Chinese University of Hong Kong, Hong Kong, Hong Kong. 2. Institute of Digestive Disease, Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences and CUHK-Shenzhen Research Institute, The Chinese University of Hong Kong, Hong Kong, Hong Kong; Institute of Liver Disease, Beijing Military General Hospital, Beijing, China. 3. Department of Anaesthesia and Intensive Care, The Chinese University of Hong Kong, Hong Kong, Hong Kong. 4. Department of Laboratory Animal Science, College of Basic Medical Sciences, Third Military Medical University, Chongqing, China. 5. Department of Laboratory Animal Science, College of Basic Medical Sciences, Third Military Medical University, Chongqing, China. Electronic address: weihong63528@163.com. 6. Institute of Digestive Disease, Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences and CUHK-Shenzhen Research Institute, The Chinese University of Hong Kong, Hong Kong, Hong Kong. Electronic address: junyu@cuhk.edu.hk.
Abstract
BACKGROUND & AIMS: Altered gut microbiota is implicated in development of colorectal cancer (CRC). Some intestinal bacteria have been reported to potentiate intestinal carcinogenesis by producing genotoxins, altering the immune response and intestinal microenvironment, and activating oncogenic signaling pathways. We investigated whether stool from patients with CRC could directly induce colorectal carcinogenesis in mice. METHODS: We obtained stored stool samples from participants in a metagenome study performed in Hong Kong. Conventional (male C57BL/6) mice were given azoxymethane to induce colon neoplasia after receiving a course of antibiotics in drinking water. Mice were gavaged twice weekly with stool from 5 patients with CRC or 5 healthy individuals (controls) for 5 weeks. Germ-free C57BL/6 mice were gavaged once with stool from 5 patients with CRC or 5 controls. We collected intestinal tissues from mice and performed histology, immunohistochemistry, expression microarray, quantitative polymerase chain reaction, immunoblot, and flow cytometry analyses. We performed 16S ribosomal RNA gene sequencing analysis of feces from mice. RESULTS: Significantly higher proportions of conventional mice fed with stool from individuals with CRC than control stool developed high-grade dysplasia (P < .05) and macroscopic polyps (P < .01). We observed a higher proportion of proliferating (Ki-67-positive) cells in colons of germ-free mice fed with stool from patients with CRC vs those fed with stool from controls (P < .05). Feces from germ-free and conventional mice fed with stool from patients with CRC vs controls contained different microbial compositions, with lower richness in mice fed with stool from patients with CRC. Intestines collected from conventional and germ-free mice fed with stool from patients with CRC had increased expression of cytokines that modulate inflammation, including C-X-C motif chemokine receptor 1, C-X-C motif chemokine receptor 2, interleukin 17A (IL17A), IL22, and IL23A. Intestines from conventional and germ-free mice fed with stool from patients with CRC contained higher proportions of T-helper 1 (Th1) cells (2.25% vs 0.44%) and Th17 cells (2.08% vs 0.31%) (P < .05 for each) than mice fed with stool from controls. Real-time polymerase chain reaction arrays revealed up-regulation of genes involved in cell proliferation, stemness, apoptosis, angiogenesis, invasiveness, and metastasis in mice fed with stool from patients with CRC. CONCLUSIONS: We fed stool samples from patients with CRC and heathy individuals to germ-free mice and conventional mice with azoxymethane. We found stool from patients with CRC to increase the numbers of polyps, levels of intestinal dysplasia and proliferation, markers of inflammation, and proportions of Th1 and Th17 cells in colon, compared with stool from individuals without CRC. This study provides evidence that the fecal microbiota from patients with CRC can promote tumorigenesis in germ-free mice and mice given a carcinogen.
BACKGROUND & AIMS: Altered gut microbiota is implicated in development of colorectal cancer (CRC). Some intestinal bacteria have been reported to potentiate intestinal carcinogenesis by producing genotoxins, altering the immune response and intestinal microenvironment, and activating oncogenic signaling pathways. We investigated whether stool from patients with CRC could directly induce colorectal carcinogenesis in mice. METHODS: We obtained stored stool samples from participants in a metagenome study performed in Hong Kong. Conventional (male C57BL/6) mice were given azoxymethane to induce colon neoplasia after receiving a course of antibiotics in drinking water. Mice were gavaged twice weekly with stool from 5 patients with CRC or 5 healthy individuals (controls) for 5 weeks. Germ-free C57BL/6 mice were gavaged once with stool from 5 patients with CRC or 5 controls. We collected intestinal tissues from mice and performed histology, immunohistochemistry, expression microarray, quantitative polymerase chain reaction, immunoblot, and flow cytometry analyses. We performed 16S ribosomal RNA gene sequencing analysis of feces from mice. RESULTS: Significantly higher proportions of conventional mice fed with stool from individuals with CRC than control stool developed high-grade dysplasia (P < .05) and macroscopic polyps (P < .01). We observed a higher proportion of proliferating (Ki-67-positive) cells in colons of germ-free mice fed with stool from patients with CRC vs those fed with stool from controls (P < .05). Feces from germ-free and conventional mice fed with stool from patients with CRC vs controls contained different microbial compositions, with lower richness in mice fed with stool from patients with CRC. Intestines collected from conventional and germ-free mice fed with stool from patients with CRC had increased expression of cytokines that modulate inflammation, including C-X-C motif chemokine receptor 1, C-X-C motif chemokine receptor 2, interleukin 17A (IL17A), IL22, and IL23A. Intestines from conventional and germ-free mice fed with stool from patients with CRC contained higher proportions of T-helper 1 (Th1) cells (2.25% vs 0.44%) and Th17 cells (2.08% vs 0.31%) (P < .05 for each) than mice fed with stool from controls. Real-time polymerase chain reaction arrays revealed up-regulation of genes involved in cell proliferation, stemness, apoptosis, angiogenesis, invasiveness, and metastasis in mice fed with stool from patients with CRC. CONCLUSIONS: We fed stool samples from patients with CRC and heathy individuals to germ-free mice and conventional mice with azoxymethane. We found stool from patients with CRC to increase the numbers of polyps, levels of intestinal dysplasia and proliferation, markers of inflammation, and proportions of Th1 and Th17 cells in colon, compared with stool from individuals without CRC. This study provides evidence that the fecal microbiota from patients with CRC can promote tumorigenesis in germ-free mice and mice given a carcinogen.
Authors: Sarah Tomkovich; Christine M Dejea; Kathryn Winglee; Julia L Drewes; Liam Chung; Franck Housseau; Jillian L Pope; Josee Gauthier; Xiaolun Sun; Marcus Mühlbauer; Xiuli Liu; Payam Fathi; Robert A Anders; Sepideh Besharati; Ernesto Perez-Chanona; Ye Yang; Hua Ding; Xinqun Wu; Shaoguang Wu; James R White; Raad Z Gharaibeh; Anthony A Fodor; Hao Wang; Drew M Pardoll; Christian Jobin; Cynthia L Sears Journal: J Clin Invest Date: 2019-03-11 Impact factor: 14.808
Authors: Wenhan Zhu; Maria G Winter; Luisella Spiga; Elizabeth R Hughes; Rachael Chanin; Aditi Mulgaonkar; Jenelle Pennington; Michelle Maas; Cassie L Behrendt; Jiwoong Kim; Xiankai Sun; Daniel P Beiting; Lora V Hooper; Sebastian E Winter Journal: Cell Host Microbe Date: 2020-02-18 Impact factor: 21.023