Basic Study
Copyright ©The Author(s) 2017. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Nov 28, 2017; 23(44): 7830-7839
Published online Nov 28, 2017. doi: 10.3748/wjg.v23.i44.7830
Composition and immuno-stimulatory properties of extracellular DNA from mouse gut flora
Ce Qi, Ya Li, Ren-Qiang Yu, Sheng-Li Zhou, Xing-Guo Wang, Guo-Wei Le, Qing-Zhe Jin, Hang Xiao, Jin Sun
Ce Qi, Jin Sun, The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, Jiangsu Province, China
Ce Qi, Ya Li, Xing-Guo Wang, Guo-wei Le, Jin Sun, School of Food Science and Technology, Jiangnan University, Wuxi 214122, Jiangsu Province, China
Ren-Qiang Yu, Wuxi Maternal and Child Health Hospital, Wuxi 212422, Jiangsu Province, China
Sheng-Li Zhou, Quality of Research and Development Department, COFCO Fortune Food Sales & Distribution Co., Ltd. Tianjin 300452, China
Hang Xiao, Department of Food Science, University of Massachusetts, Amherst, MA 01003, United States
Author contributions: Sun J and Li Y designed the research; Qi C, Li Y, Wang XG, Le GW, and Zhou SL performed the research; Sun J, Li Y, Yu RQ, and Xiao H analyzed the data and wrote the article; Xiao H and Yu RQ revised the paper; all authors have read and approved the final version to be published.
Supported by China Postdoctoral Science Foundation, No. 172774; Fund of Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, No. KLCCB-KF201603; and National Natural Science Foundation of China, No. 31201805.
Institutional animal care and use committee statement: All procedures involving animals were reviewed and approved by the Institutional Animal Care and Use Committee of Jiangnan University (IACUC protocol number: No. 8/2014/JU).
Conflict-of-interest statement: No potential conflicts of interest.
Data sharing statement: No additional data are available.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Jin Sun, PhD, Associate Professor, School of Food Science and Technology, Jiangnan University, Wuxi 214122, Jiangsu Province, China. sunj@jiangnan.edu.cn
Telephone: +86-510-85917780
Received: September 22, 2017
Peer-review started: September 23, 2017
First decision: October 11, 2017
Revised: October 14, 2017
Accepted: October 27, 2017
Article in press: October 27, 2017
Published online: November 28, 2017
Processing time: 65 Days and 12.8 Hours
ARTICLE HIGHLIGHTS
Research background

Many studies strongly suggest that signals, including bacterial DNA, from colonizing microbes greatly alter host local immune system in the gut. Bacterial cells do not contact with enterocytes in normal physiological status. They might release DNA into the mucus layer to influence host innate immune cell through specific receptors, like Toll-like receptor 9. Evidence supporting this hypothesis is needed.

Research motivation

This research investigated the existence of extracellular bacterial DNA (eDNA) in the mouse gut mucus layer, their resource, and immune modulatory function. There were differences in DNA’s immuno-stimulatory properties among different bacteria as reported by other researchers. Therefore, host immune response would be modulated by targeted change of DNA releasing bacteria in the mucus through specific medicine or food components.

Research objectives

This study aimed to confirm the existence of bacterial eDNA in the mouse gut mucus layer, and to identify bacterial genera that release them. Immuno-stimulatory properties of eDNA were also studied in vitro. This provided basic knowledge about bacteria and host interaction through bacterial DNA and related signal pathways. This will also promote nutritional strategy development to modulate local immune response through changing DNA releasing microbiota.

Research methods

Bacterial eDNA in the mucus layer and crypts was visualized by TOTO-1 staining. Small intestinal mucosal microbiota and eDNA were analyzed using T-RFLP and Illumina MiSeq amplicon sequencing. Immuno-stimulatory effects of microbiota and eDNA were determined after incubation with mouse RAW264.7 macrophages.

Research results

TOTO-1 iodide staining confirmed existence of eDNA in the mucus layer. The composition of the eDNA was significantly different from that of the intracellular DNA (iDNA). The eDNA sequences came mainly from Gram-negative bacteria of Bacteroidales S24-7. The eDNA induced significantly lower TNF-α/IL-10 and IL-6/IL-10 ratios in LPS stimulated RAW264.7 cells than iDNA. This is the first report related to bacteria genus responsible for DNA release in the gut mucus layer.

Research conclusions

Our results indicated that eDNA was located in the intestinal mucus layer. The eDNA was degraded bacterial genomic DNA mainly released by Gram-negative bacteria especially Bacteroidales S24-7. They showed decreased pro-inflammatory activity compared with total gut flora genomic DNA.

Research perspectives

Further studies are needed to clarify the specific bacterial species/strains that release eDNA, and its relationship with the gut immune response, especially the production of antimicrobial peptides in Paneth cells of the small intestinal crypt.