title: Peptostreptococcus anaerobius Induces Intracellular Cholesterol Biosynthesis in Colon Cells to Induce Proliferation and Causes Dysplasia in Mice.
abstract: Stool samples from patients with colorectal cancer (CRC) have a higher abundance of Peptostreptococcus anaerobius than stool from individuals without CRC, based on metagenome sequencing. We investigated whether P anaerobius contributes to colon tumor formation in mice and its possible mechanisms of carcinogenesis.
  
  
  
  We performed quantitative polymerase chain reaction analyses to measure P anaerobius in 112 stool samples and 255 colon biopsies from patients with CRC or advanced adenoma and from healthy individuals (controls) undergoing colonoscopy examination at hospitals in Hong Kong and Beijing. C57BL/6 mice were given broad-spectrum antibiotics, followed by a single dose of azoxymethane, to induce colon tumor formation. Three days later, mice were given P anaerobius or Esherichia coli MG1655 (control bacteria), via gavage, for 6 weeks. Some mice were also given the nicotinamide adenine dinucleotide phosphate oxidase inhibitor apocynin. Intestine tissues were collected and analyzed histologically. The colon epithelial cell line NCM460 and colon cancer cell lines HT-29 and Caco-2 were exposed to P anaerobius or control bacteria; cells were analyzed by immunoblot, proliferation, and bacterial attachment analyses and compared in gene expression profiling studies. Gene expression was knocked down in these cell lines with small interfering RNAs.
  
  
  
  P anaerobius was significantly enriched in stool samples from patients with CRC and in biopsies from patients with colorectal adenoma or CRC compared with controls. Mice depleted of bacteria and exposed to azoxymethane and P anaerobius had a higher incidence of intestinal dysplasia (63%) compared with mice not given the bacteria (8.3%; P < .01). P anaerobius mainly colonized the colon compared with the rest of the intestine. Colon cells exposed to P anaerobius had significantly higher levels of proliferation than control cells. We found genes that regulate cholesterol biosynthesis, Toll-like receptor (TLR) signaling, and AMP-activated protein kinase signaling to be significantly up-regulated in cells exposed to P anaerobius. Total cholesterol levels were significantly increased in colon cell lines exposed to P anaerobius via activation of sterol regulatory element-binding protein 2. P anaerobius interacted with TLR2 and TLR4 to increase intracellular levels of reactive oxidative species, which promoted cholesterol synthesis and cell proliferation. Depletion of reactive oxidative species by knockdown of TLR2 or TLR4, or incubation of cells with an antioxidant, prevented P anaerobius from inducing cholesterol biosynthesis and proliferation.
  
  
  
  Levels of P anaerobius are increased in human colon tumor tissues and adenomas compared with non-tumor tissues; this bacteria increases colon dysplasia in a mouse model of CRC. P anaerobius interacts with TLR2 and TLR4 on colon cells to increase levels of reactive oxidative species, which promotes cholesterol synthesis and cell proliferation.
authors.affiliation: Institute of Digestive Disease and Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong-Shenzhen Research Institute, Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Ho
authors.initials: H
authors.lastname: Tsoi
authors.affiliation: Institute of Digestive Disease and Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong-Shenzhen Research Institute, Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Eagle S H
authors.initials: ESH
authors.lastname: Chu
authors.affiliation: Institute of Digestive Disease and Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong-Shenzhen Research Institute, Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Xiang
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Department of Gastroenterology, Beijing Military General Hospital, Beijing, China.
authors.forename: Jianqiu
authors.initials: J
authors.lastname: Sheng
authors.affiliation: Institute of Digestive Disease and Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong-Shenzhen Research Institute, Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Geicho
authors.initials: G
authors.lastname: Nakatsu
authors.affiliation: Institute of Digestive Disease and Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong-Shenzhen Research Institute, Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Siew C
authors.initials: SC
authors.lastname: Ng
authors.affiliation: Department of Anatomical and Cellular Pathology, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Anthony W H
authors.initials: AWH
authors.lastname: Chan
authors.affiliation: Institute of Digestive Disease and Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong-Shenzhen Research Institute, Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Francis K L
authors.initials: FKL
authors.lastname: Chan
authors.affiliation: Institute of Digestive Disease and Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong-Shenzhen Research Institute, Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Joseph J Y
authors.initials: JJY
authors.lastname: Sung
authors.affiliation: Institute of Digestive Disease and Department of Medicine and Therapeutics, State Key Laboratory of Digestive Disease, Li Ka Shing Institute of Health Sciences, Chinese University of Hong Kong-Shenzhen Research Institute, Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China. Electronic address: junyu@cuhk.edu.hk.
authors.forename: Jun
authors.initials: J
authors.lastname: Yu
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism; D000098:Acetophenones / Q000494:pharmacology; D000236:Adenoma / Q000378:metabolism* / Q000382:microbiology; D000368:Aged; D000818:Animals; D001397:Azoxymethane; D001706:Biopsy; D053898:Biosynthetic Pathways / Q000235:genetics; D018938:Caco-2 Cells; D016022:Case-Control Studies; D049109:Cell Proliferation; D002784:Cholesterol / Q000096:biosynthesis*; D003106:Colon / Q000382:microbiology* / Q000473:pathology; D003110:Colonic Neoplasms / Q000139:chemically induced / Q000378:metabolism* / Q000382:microbiology*; D004269:DNA, Bacterial / Q000032:analysis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005243:Feces / Q000382:microbiology; D015870:Gene Expression; D016908:Gram-Positive Bacterial Infections / Q000150:complications / Q000378:metabolism*; D019073:HT29 Cells; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008875:Middle Aged; D010462:Peptostreptococcus / Q000302:isolation & purification; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D051782:Sterol Regulatory Element Binding Protein 2 / Q000378:metabolism; D051195:Toll-Like Receptor 2 / Q000235:genetics / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation
chemical_list: D000098:Acetophenones; D004269:DNA, Bacterial; D004791:Enzyme Inhibitors; D017382:Reactive Oxygen Species; D051782:Sterol Regulatory Element Binding Protein 2; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D002784:Cholesterol; C056165:acetovanillone; D055372:AMP-Activated Protein Kinases; D001397:Azoxymethane
keywords: Colon Cancer; Microbiota; Pathogen; Signal Transduction